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DNA barcoding and mini-barcoding as a powerful tool for feather mite studies
Abstract
Feather mites (Astigmata: Analgoidea, Pterolichoidea) are among the most abundantand commonly occurring bird ectosymbionts. Basic questions on the ecology and evolution of feather mites remain unanswered because feather mite species identification is often only possible for adult males and it is laborious even for specialised taxonomists, thus precluding large-scale identifications. Here, we tested DNA barcoding as a useful molecular tool to identify feather mites from passerine birds. 361 specimens of 72 species of feather mites from 68 species of European passerine birds from Russia and Spain were barcoded. The accuracy of barcoding and mini-barcoding was tested. Moreover, threshold choice (a controversial issue in barcoding studies) was also explored in a new way, by calculating through simulations the effect of sampling effort (in species number and species composition) on threshold calculations. We found one 200 bp mini-barcode region that showed the same accuracy as the full-length barcode (602 bp) and was surrounded by conserved regions potentially useful for group-specific degenerate primers. Species identification accuracy was perfect (100%) but decreased when singletons or species of the Proctophyllodes pinnatus group were included. In fact, barcoding confirmed previous taxonomic issues within the Proctophyllodes pinnatus group. Following an integrative taxonomy approach, we compared our barcode study with previous taxonomic knowledge on feather mites, discovering three new putative cryptic species and validating three previous morphologically different (but still undescribed) new species.
... Most commonly, the cytochrome c oxidase subunit 1 (cox1) mitochondrial gene has been used. Cox1 is an effective barcode for feather mite species delineation (Doña et al. 2015a) and can be useful for high-throughput biodiversity surveys ). However, it potentially has limited functionality to detect cryptic variation between closely related taxa or species undergoing incipient speciation, including symbionts like avian malaria parasites (Apicomplexa: Haemosporida; Galen et al. 2018) and mutualistic attine ants (Hymenoptera: Formicidae; Beigel et al. 2021). ...
... A previous study of feather mites suggests a conservative and reliable 3.4% (K2P) threshold value for barcoding genes (Doña et al. 2015a). We used the best partitioned multilocus tree from IQ-Tree as input into the online version of bPTP (Zhang et al. 2013). ...
... ischyros is 10.1% to 10.7% (Table 2). These genetic distance values are approximately three times greater than the suggested species threshold for feather mites of 3.42% (Doña et al. 2015a). Additional collection and sequencing of A. ischyros from more host individuals may help to further understand the breadth of this species' genetic diversity. ...
A fundamental aspect of symbiotic relationships is host specificity, ranging from extreme specialists associated with only a single host species to generalists associated with many different species. Although symbionts with limited dispersal capabilities are expected to be host specialists, some are able to associate with multiple hosts. Understanding the micro- and macroevolutionary causes of variations in host specificity is often hindered by sampling biases and the limited power of traditional evolutionary markers. Here, we studied feather mites to address the barriers associated with estimates of host specificity for dispersal-limited symbionts. We sampled feather mites (Proctophyllodidae) from a nearly comprehensive set of North American breeding warblers (Parulidae) to study mite phylogenetic relationships and host-symbiont codiversification. We used pooled-sequencing (Pool-Seq) and short-read Illumina technology to interpret results derived from a traditional barcoding gene (cytochrome c oxidase subunit 1) versus 11 protein-coding mitochondrial genes using concatenated and multispecies coalescent approaches. Despite the statistically significant congruence between mite and host phylogenies, mite-host specificity varies widely, and host switching is common regardless of the genetic marker resolution (i.e., barcode versus multilocus). However, the multilocus approach was more effective than the single barcode in detecting the presence of a heterogeneous Pool-Seq sample. These results suggest that presumed symbiont dispersal capabilities are not always strong indicators of host specificity or of historical host-symbiont coevolutionary events. Comprehensive sampling at fine phylogenetic scales may help to better elucidate the microevolutionary filters that impact macroevolutionary processes regulating symbioses, particularly for dispersal-limited symbionts.
... In ubiquitous hosts spread by humans, such as rock pigeons, Columba livia Gmelin, 1789 and domestic chickens, Gallus gallus domesticus (L., 1758), mite assemblages apparently change in new localities, indicating that some mite species have been acquired in recent events of interaction with native and phylogenetically similar host species (Gaud 1992). As for feather mites on naturally widespread host species, i.e. generalist hosts with high adaptive plasticity, a molecular identification of mites is often required to investigate their patterns of distribution, either for a proper identification of different mite haplotypes across different localities, as for the identification of potential cryptic species (Doña et al. 2015). Notwithstanding, assessing the geographic distribution of symbionts is a background needed to measure their dispersal potential and their specificity level. ...
... The smallest genetic distance between samples from different territories was found in the P. thraupis clade, where mites from AMF differed in at least 10% with Atlantic Forest mites (SAF and NAF). This COX-1 genetic variation was expressively above the threshold of 3.4% found for species delimitation of feather mites in European passerines (Doña et al. 2015). Yet, in these European passerines, no genetic structure among mites collected from different geographic territories and different host species was found, and instead, putative cryptic species for feather mites on close related hosts were recognized (Doña et al. 2015). ...
... This COX-1 genetic variation was expressively above the threshold of 3.4% found for species delimitation of feather mites in European passerines (Doña et al. 2015). Yet, in these European passerines, no genetic structure among mites collected from different geographic territories and different host species was found, and instead, putative cryptic species for feather mites on close related hosts were recognized (Doña et al. 2015). In North America, a minimum COX-1 distance of 6.6% among proctophyllodid species was identified on parulid warblers, with mites phylogenetically related according to their host's nesting ecology (Matthews et al. 2018). ...
Feather mites are the most common ectosymbionts on birds. These obligatory symbionts are mainly transmitted during their host’s parental care, which creates high host specificity. Due to this intimate relationship, it is thought that their geographic distribution is restricted by their host distribution, or that a host species harbors the same mite composition across its whole range. However, our knowledge regarding the geographic distribution of feather mites remains scarce, with only a few studies indicating disconnections between mite and host distributions, especially in widespread hosts. Here, we investigate the feather mites distribution on four tanager species, three widespread – Thraupis sayaca (L.), T. palmarum (Wied), and Stilpnia cayana (L.) from Northern and Southern Brazil; and the Amazonian T. episcopus (L.). Feather mites were identified using the molecular barcode marker COX-1 using K2P genetic distances. We found a strong genetic structure between Northern and Southern populations of tanagers of more than 10%, even among conspecific hosts. Therefore, the mite distribution on Brazilian tanagers is predominantly shaped by geography rather than by host species. These features in turn reflect historical horizontal transmissions among the hosts, suggesting a high potential for frequent host switches in these symbionts.
... Feather mites are host-specific symbionts with limited dispersal capabilities, but also with some clade-limited host-switching [25][26][27][35][36][37][38][39] . Previous studies have documented multi-host species [35][36][37]40 and cases of morphologically-cryptic but genetically different mite species [35][36][37]40 . ...
... Feather mites are host-specific symbionts with limited dispersal capabilities, but also with some clade-limited host-switching [25][26][27][35][36][37][38][39] . Previous studies have documented multi-host species [35][36][37]40 and cases of morphologically-cryptic but genetically different mite species [35][36][37]40 . Also, genetic structure has been observed (1) among populations of mites at the level of infrapopulations 41 , (2) among feather mite populations inhabiting different bird host species [35][36][37]40 and (3) among different populations of the same passerine bird species 41 . ...
... Feather mites are host-specific symbionts with limited dispersal capabilities, but also with some clade-limited host-switching [25][26][27][35][36][37][38][39] . Previous studies have documented multi-host species [35][36][37]40 and cases of morphologically-cryptic but genetically different mite species [35][36][37]40 . Also, genetic structure has been observed (1) among populations of mites at the level of infrapopulations 41 , (2) among feather mite populations inhabiting different bird host species [35][36][37]40 and (3) among different populations of the same passerine bird species 41 . ...
Some symbiont species are highly host-specific, inhabiting only one or a very few host species, and typically have limited dispersal abilities. When they do occur on multiple host species, populations of such symbionts are expected to become genetically structured across these different host species, and this may eventually lead to new symbiont species over evolutionary timescales. However, a low number of dispersal events of symbionts between host species across time might be enough to prevent population structure and species divergence. Overall, processes of evolutionary divergence and the species status of most putative multi-host symbiont systems are yet to be investigated. Here, we used DNA metabarcoding data of 6,023 feather mites (a total of 2,225 OTU representative sequences) from 147 infracommunities (i.e., the assemblage consisting of all mites of different species collected from the same bird host individual) to investigate patterns of population genetic structure and species status of three different putative multi-host feather mite species Proctophyllodes macedo Vitzthum, 1922, Proctophyllodes motacillae Gaud, 1953, and Trouessartia jedliczkai (Zimmerman, 1894), each of which inhabits a variable number of different closely related wagtail host species (genus Motacilla). We show that mite populations from different host species represent a single species. This pattern was found in all the mite species, suggesting that each of these species is a multi-host species in which dispersal of mites among host species prevents species divergence. Also, we found evidence of limited evolutionary divergence manifested by a low but significant level of population genetic structure among symbiont populations inhabiting different host species. Our study agrees with previous studies showing a higher than expected colonization opportunities in host-specific symbionts. Indeed, our results support that these dispersal events would allow the persistence of multi-host species even in symbionts with limited dispersal capabilities, though additional factors such as the geographical structure of some bird populations may also play a role.
... On the other hand, molecular differences may occur not only at the species level but also between conspecific populations (Rosenberger et al., 2013). Besides, species in some taxa are only known by a single development stage, for instance, adults in oribatid and feather mites (Doña et al., 2015;Lehmitz and Decker, 2017) and larvae in chigger mites (Kudryashova, 1998). Another common problem is the status of morphological forms occurring in sympatric areas (Dabert et al., 2011;Stålstedt et al., 2013). ...
... The COI gene fragment has been successfully used for species identification in feather mites (Astigmata: Analgoidea and Pterolichoidea) (Dabert et al., 2008;Doña et al., 2015). In a study including extensive material (72 species of feather mites associated with 68 species of European passerine birds), this marker revealed neither high intraspecific variation nor correlation with the geographic origin of samples (Doña et al., 2015). ...
... The COI gene fragment has been successfully used for species identification in feather mites (Astigmata: Analgoidea and Pterolichoidea) (Dabert et al., 2008;Doña et al., 2015). In a study including extensive material (72 species of feather mites associated with 68 species of European passerine birds), this marker revealed neither high intraspecific variation nor correlation with the geographic origin of samples (Doña et al., 2015). Moreover, these feather mite species could be delimited using a small part of the COI fragment (the so-called minibarcode) that is shorter than the standard COI barcode: 200 and 602 bp, respectively (Doña et al., 2015). ...
This study reviews the available molecular studies on chigger mites (Acari, Trombiculidae). The vast majority of publications focus on sample identifications. Nucleotide sequences of chigger mites deposited in databases have been obtained through research on biodiversity, population structure, mitochondrial gene arrangement, and mite phylogeny. The COI sequences of mites deposited in GenBank and BOLDSystems were analyzed to estimate the usefulness of barcoding in chigger mite systematics. Our results show the division of the sequences into two groups: one (Leptotrombidium species) lacks two amino acids while the other has them. The variable site matches one of the loops of the COI protein. The current level of knowledge of the nucleotide sequences in chigger mites may lead to misidentifications of samples if only molecular tools are used. Further studies on other species and other genes, especially nuclear ones, are needed.
... DNA-sequencing libraries were prepared by amplifying a region of the mitochondrial COI gene (Doña, Diaz-Real et al., 2015;Doña, Moreno-García, Criscione, Serrano, & Jovani, 2015;, and by adding the Illumina-specific sequencing primers, indices and adaptors in a two-step PCR. Finally, libraries were pooled together and analysed in a total of eight MiSeq 300PE runs (miseq reagent kit v3). ...
... We also checked whether representative sequences contained stop codons. Overall, this sequencing-bioinformatic approach allowed us to get more than 300-400 bp of COI sequenced (after quality-trimming), a sequence length over the 200 bp minibarcode known to give similar results in species identification than the total length barcode (Doña, Diaz-Real et al., 2015). ...
... Samples containing representative sequences unclassified at the species level or containing unexpected mites were further analysed by S.M. based on morphological characters of the exoskeletons, thanks to the fact that our DNA extraction protocol preserves this material (Doña, Diaz-Real et al., 2015); that is, the same mite individuals were used for molecular and later on morphological analyses. ...
The high relevance of host‐switching for the diversification of highly host‐specific symbionts (i.e., those commonly inhabiting a single host species) demands a better understanding of host‐switching dynamics at an ecological scale. Here we used DNA metabarcoding to study feather mites on passerine birds in Spain, sequencing mtDNA (COI) for 25,540 individual mites (representing 64 species) from 1,130 birds (representing 71 species). Surprisingly, 1,228 (4.8%) mites from 84 (7.4%) birds were found on host species that were not the expected to be a host according to a recent bird‐feather mite associations catalog. Unexpected associations were widespread across studied mite (40.6%) and bird (43.7%) species and showed smaller average infrapopulation sizes than typical associations. Unexpected mite species colonized hosts being distantly related to the set of their usual hosts, but with similar body size. The network of bird‐mite associations was modular (i.e., some groups of bird and mite species tended to be more associated with each other than with the others), with 75.9% of the unexpected associations appearing within the module of the typical hosts of the mite species. Lastly, 68.4% of mite species found on unexpected hosts showed signatures of genetic differentiation, and we found evidence for reproduction or the potential for it in many of the unexpected associations. Results show host colonization as a common phenomenon even for these putatively highly host‐specific symbionts. Thus, host‐switching by feather mites, rather than a rare phenomenon, appears as a relatively frequent phenomenon shaped by ecological filters such as host morphology and is revealed as a fundamental component for a dynamic coevolutionary and codiversification scenario
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... On the other hand, molecular differences may occur not only at the species level but also between conspecific populations (Rosenberger et al., 2013). Besides, species in some taxa are only known by a single development stage, for instance, adults in oribatid and feather mites (Doña et al., 2015;Lehmitz and Decker, 2017) and larvae in chigger mites (Kudryashova, 1998). Another common problem is the status of morphological forms occurring in sympatric areas (Dabert et al., 2011;Stålstedt et al., 2013). ...
... The COI gene fragment has been successfully used for species identification in feather mites (Astigmata: Analgoidea and Pterolichoidea) (Dabert et al., 2008;Doña et al., 2015). In a study including extensive material (72 species of feather mites associated with 68 species of European passerine birds), this marker revealed neither high intraspecific variation nor correlation with the geographic origin of samples (Doña et al., 2015). ...
... The COI gene fragment has been successfully used for species identification in feather mites (Astigmata: Analgoidea and Pterolichoidea) (Dabert et al., 2008;Doña et al., 2015). In a study including extensive material (72 species of feather mites associated with 68 species of European passerine birds), this marker revealed neither high intraspecific variation nor correlation with the geographic origin of samples (Doña et al., 2015). Moreover, these feather mite species could be delimited using a small part of the COI fragment (the so-called minibarcode) that is shorter than the standard COI barcode: 200 and 602 bp, respectively (Doña et al., 2015). ...
... Accurate identification requires advanced taxonomic skills and microscopic preparations of many if not all of the specimens from a sample, making large-scale studies of many host individuals very time-consuming. DNA barcoding of single specimens using a region of the COI gene have proven successful for species identification and delimitation (Doña et al. 2015a), and is becoming common in species descriptions thus increasing the coverage of DNA barcode libraries (e.g., Dabert et al. 2008;Mironov et al. 2012Mironov et al. , 2015. In addition, DNA metabarcoding using the now-obsolete Roche 454 sequencing technology allowed for the quantification of mites from some species on bulk samples (Diaz-Real et al. 2015). ...
... The number of primer mismatches (i.e., the number of bases that did not match between the primer and the target annealing site) was also taken into account to investigate whether this affected the accuracy of the abundance estimates (see Table 2). This was done using previously published mitogenomes or sequences from the species under study (Doña et al. 2015a(Doña et al. , 2017b. ...
... script of QIIME was used for taxonomic assignment of each representative sequence. Assignment was done with the RDP classifier (Wang et al. 2007) and a minimum confidence score of 96.6% against a reference database (Doña et al. 2015a). The reference database (Appendices 1, 2, Supporting Material) contained one sequence from each of the feather mite species considered in Doña et al. (2015a, b). ...
Feather mites are among the most common and diverse ectosymbionts of birds, yet basic questions such as the nature of their relationship remain largely unanswered. One reason for feather mites being understudied is that their morphological identification is often virtually impossible when using female or young individuals. Even for adult male specimens this task is tedious and requires advanced taxonomic expertise, thus hampering large-scale studies. In addition, molecular-based methods are challenging because the low DNA amounts usually obtained from these tiny mites do not reach the levels required for high-throughput sequencing. This work aims to overcome these issues by using a DNA metabarcoding approach to accurately identify and quantify the feather mite species present in a sample. DNA metabarcoding is a widely used molecular technique that takes advantage of high-throughput sequencing methodologies to assign the taxonomic identity to all the organisms present in a complex sample (i.e., a sample made up of multiple specimens that are hard or impossible to individualise). We present a high-throughput method for feather mite identification using a fragment of the COI gene as marker and Illumina Miseq technology. We tested this method by performing two experiments plus a field test over a total of 11,861 individual mites (5360 of which were also morphologically identified). In the first experiment, we tested the probability of detecting a single feather mite in a heterogeneous pool of non-conspecific individuals. In the second experiment, we made 2 × 2 combinations of species and studied the relationship between the proportion of individuals of a given species in a sample and the proportion of sequences retrieved to test whether DNA metabarcoding can reliably quantify the relative abundance of mites in a sample. Here we also tested the efficacy of degenerate primers (i.e., a mixture of similar primers that differ in one or several bases that are designed to increase the chance of annealing) and investigated the relationship between the number of mismatches and PCR success. Finally, we applied our DNA metabarcoding pipeline to a total of 6501 unidentified and unsorted feather mite individuals sampled from 380 European passerine birds belonging to 10 bird species (field test). Our results show that this proposed pipeline is suitable for correct identification and quantitative estimation of the relative abundance of feather mite species in complex samples, especially when dealing with a moderate number (> 30) of individuals per sample.
... Indeed, the morphological identification of feather mites is difficult. Immature stages of many taxa and females of closely-related species are often indistinguishable, while male morphological characterization is a laborious task even for experienced taxonomists (Doña et al., 2015b). Molecular studies hold much promise for documenting and understanding diversity in this group, but are still relatively scarce (but see Dabert et al., 2001Dabert et al., , 2015Klimov and OConnor, 2008;Knowles and Klimov, 2011;Štefka et al., 2011;Doña et al., 2015b). ...
... Immature stages of many taxa and females of closely-related species are often indistinguishable, while male morphological characterization is a laborious task even for experienced taxonomists (Doña et al., 2015b). Molecular studies hold much promise for documenting and understanding diversity in this group, but are still relatively scarce (but see Dabert et al., 2001Dabert et al., , 2015Klimov and OConnor, 2008;Knowles and Klimov, 2011;Štefka et al., 2011;Doña et al., 2015b). ...
... One of the difficulties in measuring diversity and in studying host-parasite associations is the presence of cryptic species. Cryptic speciation appears to be common in ectosymbionts, such as lice (Malenke et al., 2009) and mites (Miller et al., 2013;Doña et al., 2015b). The strong host-associated adaptive pressures on these ectosymbionts can promote convergence in morphology, rendering traditional morphological methods insufficient for species identification (Doña et al., 2015b). ...
Feather mites are useful models for studying speciation due to their high diversity and strong degree of host specialization. However, studies to date have focused on the evolution of higher-level mite taxa while much hidden diversity likely occurs at the level of host genera and species. In this study, we examined the diversity and evolution of feather mites infesting six sympatric seabird species from six genera, breeding in the Cape Verde archipelago. We report 32 feather mite morphospecies categorized into ten genera and three families, of which nine correspond to new, undescribed species. Molecular data corroborated morphological species descriptions, except for two morphologically-cryptic, but genetically distinct mite lineages related to Zachvatkinia oceanodromae and Laminalloptes simplex. Using these communities, we then applied a co-structure approach to test the contribution of ectosymbiont and host factors in driving feather mite evolution. Most seabird species hosted specific and unique feather mite species, even under sympatric conditions, and in general, feather mite species exhibited strong host-driven genetic structure. However, patterns of genetic differentiation were variable. That is, some mite species are more generalist than others and mite lineages/haplotypes can be shared by related seabird species. Interestingly, host-specific mites (eg., Zachvatkinia spp.) tend to display much higher intra-specific diversity compared to more generalist mites (eg., Microspalax and Plicatalloptes spp.). We discuss ectosymbiont and host life-history traits that might generate these patterns, such as host dispersal and breeding behavior and/or mite spatial and trophic specialization. Our findings highlight both the vast and largely unrecognized diversity of avian feather mites on seabirds, and the intrinsic complexity of the ecological processes underlying the evolution of these ectosymbionts.
... Reporting cryptic species is relevant for a better description of biodiversity and an aid for taxonomy, and it has been previously documented in other mites, such as feather mites (Dabert et al. 2001;Doña et al. 2015), Tenuipalpidae like Brevipalpus spp. (Navia et al. 2013), ticks like Rhipicephalus microplus (Low et al. 2015) or in rhinonyssid mites (de Rojas et al. 2002). ...
... However, the integration of molecular and morphological (and other) evidence can also inform us about underlying evolutionary processes at play (Struck et al. 2018). In this sense, the two cryptic Tinaminyssus species studied here showed strong differentiation for the two molecular markers studied, displaying genetic distances similar (and even larger) to those from clearly morphologically distinct Tinaminyssus species (de Rojas et al. 2002), feather mites (Doña et al. 2015(Doña et al. , 2017, tetranychid mites (Navajas and Boursot 2003), Demodex species (de Rojas et al. 2012;Zhao et al. 2013Zhao et al. , 2014 or ticks (Hornok et al. 2015). This suggests that the cryptic species found here may be an example of evolutionary stasis, likely because the morphological similarity between close bird host species does not impose strong selection pressures for morphological differentiation in these mite species. ...
... Finally, our results also show the potential usefulness of COI as a barcode for rhinonyssids, as it has been found for related mite species such as tetranychid mites (Ros and Breeuwer 2007), feather mites (Doña et al. 2015) or quill mites (Glowska et al. 2016). Given the difficulty of identifying rhinonyssid species based on morphological traits (see above), DNA barcoding opens a promising future for advancing on the taxonomy of this neglected group of mites. ...
The study of cryptic species allows to describe and to understand biodiversity, and the evolutionary processes shaping it. Mites of the family Rhinonyssidae are permanent parasites of the nasal cavities of birds, currently including about 500 described species and 12 genera. Here, we tested the hypothesis that mites from five populations of the genus Tinaminyssus—three isolated from European turtle doves (Streptopelia turtur), and two from Eurasian collared doves (Streptopelia decaocto; Aves: Columbiformes)—are, in fact, two cryptic species inhabiting different hosts. First, we performed a morphometrical study on 16 traits. Then, we used the ITS1-5.8S rDNA-ITS2 nuclear region (ITS region), and a fragment of the mitochondrial cytochrome c-oxidase 1 (COI) to carry out phylogenetic and species delimitation analyses on Tinaminyssus species. Morphological analyses revealed a lack of biometric differentiation among Tinaminyssus populations from the two host species. However, molecular analyses indicated a high degree of genetic differentiation between populations of Tinaminyssus sp. from S. turtur and S. decaocto. Overall, results show that they can be considered as different cryptic species, suggesting a case of evolutionary stasis, likely because of the anatomical similarity between closely-related bird host species.
... (iii) specific distribution on particular feathers and microsites on feathers (Fern andez-Gonz alez, P erez-Rodr ıguez, de la Hera, Proctor, & P erez-Tris, 2015;Jovani & Serrano, 2001Stefan et al., 2015); and (iv) mainly vertical mode of transmission (Doña, Potti, et al., 2017;Jovani, Tella, Sol, & Ventura, 2001;Mironov & Malyshev, 2002). However, as with many other symbionts, they are challenging to study, and this has strongly hampered our comprehension of this system (Doña, Diaz-Real, et al., 2015;Proctor, 2003;Proctor & Owens, 2000). ...
... A negative control that contained no sample was included in every extraction round to check for contamination during the experiments. This procedure preserves exoskeletons for morphological identifications (see Doña, Diaz-Real, et al., 2015). However, in contrast to more aggressive isolation methods, DNA from Gram-positive bacteria, undigested diatoms and intact fungal spores may not have been amplified. ...
... Kerr, Lijtmaer, Barreira, Hebert, & Tubaro, 2009). In addition, we amplified the COI gene of feather mites (bcdF05/bcdR04, Dabert, Ehrnsberger, & Dabert, 2008) to molecularly confirm the mite DOÑA ET AL. species identity (Doña, Diaz-Real, et al., 2015). Only bacterial and fungal regions were amplified from the external samples. ...
Parasites and other symbionts are crucial components of ecosystems, regulating host populations and supporting food webs. However, most symbiont systems, especially those involving commensals and mutualists, are relatively poorly understood. In this study, we have investigated the nature of the symbiotic relationship between birds and their most abundant and diverse ectosymbionts: the vane‐dwelling feather mites. For this purpose, we studied the diet of feather mites using two complementary methods. First, we used light microscopy to examine the gut contents of 1,300 individual feather mites representing 100 mite genera (18 families) from 190 bird species belonging to 72 families and 19 orders. Second, we used high‐throughput sequencing (HTS) and DNA metabarcoding to determine gut contents from 1,833 individual mites of 18 species inhabiting 18 bird species. Results showed fungi and potentially bacteria as the main food resources for feather mites (apart from potential bird uropygial gland oil). Diatoms and plant matter appeared as rare food resources for feather mites. Importantly, we did not find any evidence of feather mites feeding upon bird resources (e.g., blood, skin) other than potentially uropygial gland oil. In addition, we found a high prevalence of both keratinophilic and pathogenic fungal taxa in the feather mite species examined. Altogether, our results shed light on the long‐standing question of the nature of the relationship between birds and their vane‐dwelling feather mites, supporting previous evidence for a commensalistic–mutualistic role of feather mites, which are revealed as likely fungivore–microbivore–detritivore symbionts of bird feathers.
... Martin et al., 2010;Skoracka & Dabert, 2010;Miller et al., 2013;Pešić et al., 2017Pešić et al., , 2019, including feather mites (e.g. Dabert et al., 2008Dabert et al., , 2015Doña et al., 2015). In this study, we used DNA sequence data to test the hypotheses that Analges species should be grouped into two subgenera, Analges and Analgopsis, or the two species groups A. chelopus and A. passerinus. ...
... Vilas et al., 2005;Smith et al., 2006Smith et al., , 2007Skoracka et al., 2015;Liu et al., 2019), including feather mites (e.g. Whiteman et al., 2006;Dabert et al., 2015;Doña et al., 2015;Stefan et al., 2018). Our results of molecular species delimitation not only supported all Analges species defined a priori based on morphological characteristics but also highlighted seven groups that probably represent new species (Fig. 2). ...
Mites of the genus Analges (Acariformes: Analgidae) inhabit the down feathers of passeriform birds. The evolutionary history of Analges and the co-phylogentic relationships between these mites and their hosts are unknown. Our phylogenetic analysis supported the monophyly of the genus, but it did not support previous taxonomic hypotheses subdividing the genus into the subgenera Analges and Analgopsis or arranging some species into the A. chelopus and A. passerinus species groups. Molecular data reveal seven new species inhabiting Eurasian passerines and support the existence of several multi-host species. According to molecular dating, the origin of the Analges (c. 41 Mya) coincided with the Eocene diversification of Passerida into Sylvioidea and Muscicapoidea–Passeroidea. The initial diversification of Analges took place on the Muscicapoidea clade, while remaining passerine superfamilies appear to have been colonized because of host-switching. Co-speciation appears to be relatively common among Analges species and their hosts, but the most striking pattern in the co-phylogenetic scenario involves numerous complete host-switches, spreads and several failures to speciate. The mechanism of long-term gene-flow among different populations of multi-host Analges species is enigmatic and difficult to resolve. Probably, in some cases mites could be transferred between birds via feathers used as nest material.
... Martin et al., 2010;Skoracka & Dabert, 2010;Miller et al., 2013;Pešić et al., 2017Pešić et al., , 2019, including feather mites (e.g. Dabert et al., 2008Dabert et al., , 2015Doña et al., 2015). In this study, we used DNA sequence data to test the hypotheses that Analges species should be grouped into two subgenera, Analges and Analgopsis, or the two species groups A. chelopus and A. passerinus. ...
... Vilas et al., 2005;Smith et al., 2006Smith et al., , 2007Skoracka et al., 2015;Liu et al., 2019), including feather mites (e.g. Whiteman et al., 2006;Dabert et al., 2015;Doña et al., 2015;Stefan et al., 2018). Our results of molecular species delimitation not only supported all Analges species defined a priori based on morphological characteristics but also highlighted seven groups that probably represent new species (Fig. 2). ...
Mites of the genus Analges (Acariformes: Analgidae) inhabit the down feathers of passeriform birds. The evolutionary history of Analges and the co-phylogentic relationships between these mites and their hosts are unknown. Our phylogenetic analysis supported the monophyly of the genus, but it did not support previous taxonomic hypotheses subdividing the genus into the subgenera Analges and Analgopsis or arranging some species into the A. chelopus and A. passerinus species groups. Molecular data reveal seven new species inhabiting Eurasian passerines and support the existence of several multi-host species. According to molecular dating, the origin of the Analges (c. 41 Mya) coincided with the Eocene diversification of Passerida into Sylvioidea and Muscicapoidea–Passeroidea. The initial diversification of Analges took place on the Muscicapoidea clade, while remaining passerine superfamilies appear to have been colonized because of host-switching. Co-speciation appears to be relatively common among Analges species and their hosts, but the most striking pattern in the co-phylogenetic scenario involves numerous complete host-switches, spreads and several failures to speciate. The mechanism of long-term gene-flow among different populations of multi-host Analges species is enigmatic and difficult to resolve. Probably, in some cases mites could be transferred between birds via feathers used as nest material.
... DNA barcoding is considered as an advance technique which is commonly being used taxonomic identifications (Nagoshi et al., 2011;Doña et al., 2015;Xu et al., 2015). It uses short standardized COI (cytochrome C Oxidase subunit I) gene region of mitochondrial DNA for identification of species (Herbert et al., 2003). ...
... Infact identification of the known adult samples is also time taking, as the spider species need quite detailed examination and sometimes even dissection of sexual organs for firm identification (Locket and Millidge, 1951). Under such circumstances, DNA barcoding is likewise a significant and trustworthy method Barrett and Herbert, 2005;Blagoev et al., 2013;Raso et al., 2014;Doña et al., 2015;Xu et al., 2015). For the first time DNA Barcoding was discovered by Hebert et al. (2003), now this method is used in biosecurity (Armstrong and Ball, 2005), consumer protection (Lowenstein et al., 2010), ecology (Jurado-Rivera et al., 2009), conservation (Francis et al., 2010), biodiversity assessment (Janzen et al., 2009) and taxonomy (Benziger et al., 2011). ...
Spiders are natural predators of many insect pests and preferred as they are carnivores. In the present study we identified the cursorial spiders of different families using DNA barcoding as it is very helpful in identifying species where morphological identifications can be difficult e.g., delimitation of juvenile stages. Standard barcode region of CO1 gene of 64 samples was amplified. The sequences of 658 base pairs were recovered from 62 samples, representing 7 families, 20 genera and 27 species. Araneidae was the most dominant family followed by Salticidae, Oxyopidae, Clubionidae, Tetragnathida, Thomisidae, Mitergidae and Lycosidae. The interspecific value of divergence was more than the intraspecific value of divergence for all seven families which described a clear barcode gap. No overlap was recorded in intraspecific and interspecific divergence value. Furthermore, distance to NN was higher than the maximum intraspecific value for all species. A barcode reference library of the cursorial spiders of Punjab University, Lahore and Soon Sakeser Valley Punjab was also established. It is concluded that CO1 has potentially enough information for fast and accurate identification of spiders. Although, morphological studies alone are working satisfactory for the identification of spiders, still its efficiency increased when combined with DNA barcoding.
... It is unclear what active component of G. gecko is used in related functional foods and Chinese patent medicines [20,21]. Although DNA barcoding technology based on COI region has the ability to distinguish G. gecko from related species and to identify other animal species successfully [11,22,23], this technology cannot be applied to samples with degraded DNA because it is impossible to amplify the COI region (> 600 bp) of highly processed materials [24][25][26][27]. ...
... The results were consistent with a previously reported study that identified G. gecko from adulterants using COI regions [11,22]. DNA minibarcoding and specific-PCR identification are becoming more popular methods for effective species identification [24,25,31], especially for specimens with degraded DNA, such as archival specimens, processed foods, functional foods, and Chinese patent medicines [26,27,[32][33][34][35]. For a more sensitive identification of G. gecko in Fig. 3 The sensitivity of amplification with universal and specific primers for detection of Gekko gecko. ...
Background:
The dry body of the Tokay Gecko (Gekko gecko) is the source of a valuable traditional Chinese medicine, it is therefore listed as a Class II protected animal species in China. Due to increasing market demand and a declining supply of the species, a considerable number of adulterants have emerged in the market. Thus, it is necessary to establish an accurate and rapid method of identification for distinguishing G. gecko from its adulterants and for separating it from highly processed products.
Methods:
A total of 274 COI sequences were analyzed by using MEGA 5.0 software. Several specific primers were designed to amplify mini-barcode regions and identify G. gecko from its counterfeits and products.
Results:
274 COI sequences of G. gecko and 15 adulterants species were analyzed. G. gecko could be distinguished from its adulterants through BLAST analysis, intra- and inter-specific distance analyses, and an NJ tree based on COI sequences. Two pairs of specific primers designed for this study, COISF2/COISR2 and COISF3/COISR3, amplified 200- and 133-bp fragments of the COI region, respectively, both of which were suitable for the identification of G. gecko and its adulterants. Furthermore, COISF3/COISR3 detected G. gecko in 15 batches of products.
Conclusion:
Therefore, the specific DNA mini-barcoding method developed here may be a powerful tool for the identification of G. gecko and counterfeits, and may also be used to distinguish G. gecko from its highly processed by-products.
... It is unclear what active component of G. gecko is used in related functional foods and Chinese patent medicines [20,21]. Although DNA barcoding technology based on COI region has the ability to distinguish G. gecko from related species and to identify other animal species successfully [11,22,23], this technology cannot be applied to samples with degraded DNA because it is impossible to amplify the COI region (>600 bp) of highly processed materials [24][25][26][27]. ...
... The results were consistent with a previously reported study that identi ed G. gecko from adulterants using COI regions [11,22]. DNA mini-barcoding and speci c-PCR identi cation are becoming more popular methods for effective species identi cation [24,25,31], especially for specimens with degraded DNA, such as archival specimens, processed foods, functional foods, and Chinese patent medicines [26,27,[32][33][34][35]. For a more sensitive identi cation of G. gecko in 2001, it was distinguished from 14 adulterants through allele-speci c diagnostic PCR which ampli ed 260-bp fragments of the 12S rRNA gene [36]. ...
Background: The dry body of the Tokay Gecko (Gekko gecko) is the source of a valuable traditional Chinese medicine, it is therefore listed as a Class II protected animal species in China. Due to increasing market demand and a declining supply of the species, a considerable number of adulterants have emerged in the market. Thus, it is necessary to establish an accurate and rapid method of identification for distinguishing G. gecko from its adulterants and for separating it from highly processed products.
Methods: A total of 274 COI sequences were analyzed by using MEGA 5.0 software. Several specific primers were designed to amplify mini-barcode regions and identify G. gecko from its counterfeits and products.
Results: 274 COI sequences of 16 G. gecko species and adulterants were analyzed. G. gecko could be distinguished from its adulterants through BLAST analysis, intra- and inter-specific distance analyses, and an NJ tree based on COI sequences. Two pairs of specific primers designed for this study, COISF2/COISR2 and COISF3/COISR3, amplified 200- and 133-bp fragments of the COI region, respectively, both of which were suitable for the identification of G. gecko and its adulterants. Furthermore, COISF3/COISR3 detected G. gecko in 15 batches of products.
Conclusion: Therefore, the specific DNA mini-barcoding method developed here may be a powerful tool for the identification of G. gecko and counterfeits, and may also be used to distinguish G. gecko from its highly processed by-products.
... DNA barcoding is considered as an advance technique which is commonly being used taxonomic identifications (Nagoshi et al., 2011;Doña et al., 2015;Xu et al., 2015). It uses short standardized COI (cytochrome C Oxidase subunit I) gene region of mitochondrial DNA for identification of species (Herbert et al., 2003). ...
... Infact identification of the known adult samples is also time taking, as the spider species need quite detailed examination and sometimes even dissection of sexual organs for firm identification (Locket and Millidge, 1951). Under such circumstances, DNA barcoding is likewise a significant and trustworthy method Barrett and Herbert, 2005;Blagoev et al., 2013;Raso et al., 2014;Doña et al., 2015;Xu et al., 2015). For the first time DNA Barcoding was discovered by Hebert et al. (2003), now this method is used in biosecurity (Armstrong and Ball, 2005), consumer protection (Lowenstein et al., 2010), ecology (Jurado-Rivera et al., 2009), conservation (Francis et al., 2010), biodiversity assessment (Janzen et al., 2009) and taxonomy (Benziger et al., 2011). ...
... The single best window identified through this procedure was then subjected to BM and BCM analyses in Species Identifier similar to those described above for the full barcode sequence set. Following Doña et al. (2015), we used DNASP Bagley et al. 5 (Librado & Rozas, 2009) to evaluate the suitability of regions flanking the best window for development of fish-specific primers to amplify the mini-barcode, based on DNA polymorphism of sliding windows across the gene. ...
... Bagley et al. Doña et al., 2015). Our sliding window results identified a 200 bp mini-barcode region of the cox1 gene for Cerrado freshwater fishes that performed equally well at specimen identification as compared with the entire DNA barcode (Table 2) and seems well positioned for the development of degenerate primers for PCR (Figure 4). ...
The extraordinary species diversity of the Neotropical freshwater fish fauna is world renown. Yet, despite rich species diversity, taxonomic and genetic resources for its Cerrado ichthyofauna remain poorly developed. We provide a reference library of 149 DNA barcodes for 39 species/lineages of Cerrado headwater stream fishes from the Brazilian Distrito Federal and nearby areas and test the utility of distance-based criteria, tree-based criteria and minibarcodes for specimen identification. Mean Kimura 2-parameter genetic distances within species to orders ranged 1·8-12·1%. However, mean intraspecific v. congeneric-interspecific distances (0·9-1·3%) overlapped extensively and distance-based barcoding failed to achieve correct identifications due to c. 4-12·1% error rates and 19·5% ambiguous identifications related to the presence of singletons. Overlap was reduced and best-match success rates improved drastically to 83·5% when Characidium barcodes representing potential misidentifications or undescribed species were removed. Tree-based monophyly criteria generally performed similarly to distance methods, correctly differentiating up to c. 85% of species/lineages despite neighbour-joining and Bayesian tree errors (random lineage-branching events, long-branch attraction). Five clusters (Ancistrus aguaboensis, Characidium spp., Eigenmannia trilineata, Hasemania hanseni and Hypostomus sp. 2) exhibited deep intraspecific divergences or para-/polyphyly and multiple Barcode Index Number assignments indicative of putative candidate species needing taxonomic re-examination. Sliding-window analyses also indicated that a 200 bp minibarcode region performed just as well at specimen identification as the entire barcode gene. Future DNA barcoding studies of Distrito Federal-Cerrado freshwater fishes will benefit from increased sampling coverage, as well as consideration of minibarcode targets for degraded samples and next-generation sequencing.
... Based on our curated Pyroglyphidae + Psoroptidae dataset, a 'conservative' distance of > 9.52% K2P distance was able to distinguish species that have clear morphological differences (Table 1). This value is very close to the average smallest interspecific distances (9%) reported for feather mites [34]. Below the 9.52% 'conservative' distance there was a "gray" species delimitation zone, where OTUs could not be unambiguously assigned to species based on morphology. ...
... Below the 9.52% 'conservative' distance there was a "gray" species delimitation zone, where OTUs could not be unambiguously assigned to species based on morphology. It is notable that our 'conservative' cox1 threshold is much higher than values used in literature (4% [11], 3.14% [34], 3% [6,12],~2% [7,12,13], or lower [6]). Applying even the highest of these threshold values to our dataset will split species having large, strongly structured and presumably panmictic populations. ...
We report two host-specific lineages of scab mites of the genus Caparinia, parasitizing European and African hedgehogs. Based on morphology, these mite lineages are closely related sister groups. The morphological differences, however, are subtle and do not provide clear-cut evidence for the existence of separate species. CO1 divergence between these lineages was 7.4–7.8%, well above the CO1 barcoding gaps or thresholds commonly used to separate species, whereas divergence of five nuclear genes was very low, 0.06–0.53%, suggesting that these lineages could belong to a single species with gene flow between them. Thus, there is a conflict between the mitochondrial (CO1) gene and nuclear genes (i.e mito-nuclear discordance). We attribute this conflict to the ‘gray zone’ where species delimitation is ambiguous due to substantial gene flow. We also report another ‘gray zone’ species, Psoroptes ovis (a species of veterinary importance), whose within-species CO1 distances reached 6.0%. We provide a detailed morphological description and figures of C. ictonyctis stat. res. from the African hedgehog, using light and SEM microscopy and give morphometric data for this species and its sister species, Caparinia tripilis from Europe. For all known species of Caparinia, we document their host associations and give a key to species of the world based on results of our morphological and molecular analyses and a nearly exhaustive study of museum specimens.
... Based on our curated Pyroglyphidae + Psoroptidae dataset, a 'conservative' distance of > 9.52% K2P distance was able to distinguish species that have clear morphological differences (Table 1). This value is very close to the average smallest interspecific distances (9%) reported for feather mites [34]. Below the 9.52% 'conservative' distance there was a "gray" species delimitation zone, where OTUs could not be unambiguously assigned to species based on morphology. ...
... Below the 9.52% 'conservative' distance there was a "gray" species delimitation zone, where OTUs could not be unambiguously assigned to species based on morphology. It is notable that our 'conservative' cox1 threshold is much higher than values used in literature (4% [11], 3.14% [34], 3% [6,12],~2% [7,12,13], or lower [6]). Applying even the highest of these threshold values to our dataset will split species having large, strongly structured and presumably panmictic populations. ...
Background: The cox1-barcoding approach is currently extensively used for high-throughput species delimitation and discovery. However, this method has several limitations, particularly when organisms have large effective population sizes. Paradoxically, most common, abundant, and widely distributed species may be misclassified by
this technique.
Results: We conducted species delimitation analyses for two host-specific lineages of scab mites of the genus Caparinia, having small population sizes. Cox1 divergence between these lineages was high (7.4–7.8%) while that of nuclear genes was low (0.06–0.53%). This system was contrasted with the medically important American house dust mite, Dermatophagoides farinae, a globally distributed species with very large population size. This species has two distinct, sympatric cox1 lineages with 4.2% divergence. We tested several species delimitation algorithms PTP, GMYC, ABGD, BPP, STACEY and PHRAPL, which inferred different species boundaries for these entities. Notably, STACEY recovered the Caparinia lineages as two species and D. farinae as a single species. BPP agreed with these
results when the prior on ancestral effective population sizes was set to expected values, although delimitation of Caparinia was still equivocal. No other cox1 species delimitation algorithms inferred D. farinae as a single species, despite the fact that the nuclear CPW2 gene shows some evidence for introgression between the cox1 groups. This indicates that the cox1-barcoding approach may result in excessive species splitting.
Conclusions: Our research highlights the importance of using nuclear genes and demographic characteristics to infer species boundaries rather than relying on a single-gene barcoding approach, particularly for putative species having large effective population sizes.
... Despite being of poor utility in discerning deeper phylogenetic relationships, DNA barcodes have nevertheless been shown to be extremely useful for rapid and accurate species delimitation in invertebrates (Wang et al. 2018), especially in cases where species show low levels of morphological divergence (e.g. Hebert et al. 2004;Schlick-Steiner et al. 2006;Fisher & Smith 2008;Liu et al. 2013;Doña et al. 2015). Rhopalomastix species are small (ca. ...
... Resolving the phylogeny of Rhopalomastix is beyond the scope of this study, but we advocate an integrated approach based on morphology and more comprehensive DNA sequencing for future systematic work on this genus, given the complexities of morphological variation between and within populations. That said, we do not discount the value of short fragment DNA barcodes-these barcodes still serve as useful tools to aid initial species delimitation (Doña et al. 2015;Wang et al. 2018), but should not constitute the single basis for species inference. Genetic information should always be analysed in conjunction with morphology and/or other forms of biological evidence. ...
The true diversity of the Asian ant genus Rhopalomastix Forel is poorly understood. We use an integrated approach to
review the known species and subspecies of Rhopalomastix in Southeast Asia. Based on morphology and supporting DNA
evidence, we recognize six species. We raise two subspecies of R. rothneyi Forel to species rank (R. johorensis Wheeler
stat. n, R. javana Wheeler stat. n.), synonymize R. janeti Donisthorpe (syn. nov.) with R. johorensis, and describe four
new species from Singapore: R. glabricephala sp. n., R. murphyi sp. n., R. striata sp. n., and R. tenebra sp. n. All six
species found in Southeast Asia are distinct from each other based on morphology; morphological delimitation of these
species is further supported by and congruent with mOTUs generated from objective clustering of short fragment COI
barcodes using the best close match criteria. Different castes and sexes of most species are described, including redescriptions
of the queen of R. javana and male of R. johorensis. A key to the Southeast Asian species based on the worker caste
is also provided. Variation among sympatric and also geographically distant populations, and the possibilities of cryptic
species, are discussed.
... Despite being of poor utility in discerning deeper phylogenetic relationships, DNA barcodes have nevertheless been shown to be extremely useful for rapid and accurate species delimitation in invertebrates (Wang et al. 2018), especially in cases where species show low levels of morphological divergence (e.g. Hebert et al. 2004;Schlick-Steiner et al. 2006;Fisher & Smith 2008;Liu et al. 2013;Doña et al. 2015). Rhopalomastix species are small (ca. ...
... Resolving the phylogeny of Rhopalomastix is beyond the scope of this study, but we advocate an integrated approach based on morphology and more comprehensive DNA sequencing for future systematic work on this genus, given the complexities of morphological variation between and within populations. That said, we do not discount the value of short fragment DNA barcodes-these barcodes still serve as useful tools to aid initial species delimitation (Doña et al. 2015;Wang et al. 2018), but should not constitute the single basis for species inference. Genetic information should always be analysed in conjunction with morphology and/or other forms of biological evidence. ...
The true diversity of the Asian ant genus Rhopalomastix Forel is poorly understood. We use an integrated approach to review the known species and subspecies of Rhopalomastix in Southeast Asia. Based on morphology and supporting DNA
evidence, we recognize six species. We raise two subspecies of R. rothneyi Forel to species rank (R. johorensis Wheeler stat. n, R. javana Wheeler stat. n.), synonymize R. janeti Donisthorpe (syn. nov.) with R. johorensis, and describe four new species from Singapore: R. glabricephala sp. n., R. murphyi sp. n., R. striata sp. n., and R. tenebra sp. n. All six species found in Southeast Asia are distinct from each other based on morphology; morphological delimitation of these species is further supported by and congruent with mOTUs generated from objective clustering of short fragment COI barcodes using the best close match criteria. Different castes and sexes of most species are described, including redescriptions of the queen of R. javana and male of R. johorensis. A key to the Southeast Asian species based on the worker caste is also provided. Variation among sympatric and also geographically distant populations, and the possibilities of cryptic species, are discussed.
... One of the most widely employed markers is the mitochondrial Cytochrome Oxidase subunit I (COI), which has been used to investigate intra-and interspecific variation in many arthropods including various mites, such as, feather mites (Bochkov et al. 2014;Doña et al. 2015;Klimov et al. 2017), phytophagous mites (Navajas et al. 1996;Skoracka and Dabert 2010;Khaing et al. 2015), predatory mites (Li et al. 2012), and stored product mites (Webster et al. 2004;Yang et al. 2011;Khaing et al. 2014;Sun et al. 2014;Erban et al. 2016;Yang and Li 2016;Que et al. 2016). This locus possesses a high level of variability, enabling distinction between two or more groups of individuals that originally were described by morphological characters either as a single or as distinct species and helps to establish the species status of these individuals (Hebert et al. 2003(Hebert et al. , 2004aPentinsaari et al. 2016). ...
... The within-species distance of T. putrescentiae (4.3 or 5.8%, full dataset vs. the Folmer fragment only), is relatively high and broadly overlaps with between-species distances reported in the literature, e.g., 4% (Dowton et al. 2014), 3.1% (Doña et al. 2015), 3% (Hebert et al. 2004b;Smith et al. 2005), 2% (Rossini et al. 2016;Smith et al. 2005), or lower (Hebert et al. 2004a). However, species having compatible or higher within-species genetic distances are known as well: 10.1% in the human follicle mite, Demodex folliculorum (Demodecidae) (conseravtively recalculated from Palopoli et al. 2015), 5.7-6.8% in the common blue butterfly, Polyommatus icarus (Lycaenidae) (Wiemers and Fiedler 2007), and about 6% in the sea snail, Echinolittorina vidua (Littorinidae) (Williams and Reid 2004). ...
Mites of the genus Tyrophagus (Acari: Acaridae) are among the most widespread and common mites, inhabiting diverse natural and anthropogenic habitats. Some species are pests of agricultural products and stored food and/or live in house dust, causing allergies to humans. We sequenced 1.2 kb of the mitochondrial COI gene for 38 individuals belonging to seven species of Tyrophagus, including T. curvipenis, T. putrescentiae, T. fanetzhangorum, T. longior, T. perniciosus, and T. cf. similis. Molecular phylogenetic analyses (1) recovered two major clades corresponding to the presence or absence of eyespots, and (2) separated all included morphological species. Tyrophagus curvipenis and T. putrescentiae had the lowest between-species genetic distances (range, mean ± SD): 14.20–16.30, 15.17 ± 0.40 (K2P). The highest within-species variation was found in T. putrescentiae 0.00–4.33, 1.78 ± 1.44 (K2P). In this species, we recovered two distinct groups; however, no geographical or ecological dissimilarities were observed between them. Based on our analyses, we document important morphological differences between T. curvipenis and T. putrescentiae. For the first time, we record the occurrence of T. curvipenis in the New World and suggest that it may be an emerging pest as it is currently spreading in agricultural produce.
... However, the main drawback is the relative difficulty in designing universal primers compared with that for normal barcode regions (ITS2, psbA-trnH, etc.) when working with degraded DNA. The amplification and sequencing of mini-barcodes often require the design of specific primers, which also reduces the universality of this method [105]. Second, the reduced size of mini-barcodes with less genetic information and variation sites inevitably causes a decrease in species resolution [106]. ...
DNA barcoding has been widely used for herb identification in recent decades, enabling safety and innovation in the field of herbal medicine. In this article, we summarize recent progress in DNA barcoding for herbal medicine to provide ideas for the further development and application of this technology. Most importantly, the standard DNA barcode has been extended in two ways. First, while conventional DNA barcodes have been widely promoted for their versatility in the identification of fresh or well-preserved samples, super-barcodes based on plastid genomes have rapidly developed and have shown advantages in species identification at low taxonomic levels. Second, mini-barcodes are attractive because they perform better in cases of degraded DNA from herbal materials. In addition, some molecular techniques, such as high-throughput sequencing and isothermal amplification, are combined with DNA barcodes for species identification, which has expanded the applications of herb identification based on DNA barcoding and brought about the post-DNA-barcoding era. Furthermore, standard and high-species coverage DNA barcode reference libraries have been constructed to provide reference sequences for species identification, which increases the accuracy and credibility of species discrimination based on DNA barcodes. In summary, DNA barcoding should play a key role in the quality control of traditional herbal medicine and in the international herb trade.
... The COI intraspecific diversity gene in animals was found to be significantly lower than the interspecific diversity. The COI gene barcoding was effective for classifying and identifying vertebrates and invertebrates, so it has been widely used in various fisheries' biological groups [16][17][18][19]. This is what makes COI genes very important in the identification and classification of biological groups. ...
Objective: This study describes the newest deoxyribonucleic acid (DNA) barcoding Asian redtail
catfish (Hemibagrus nemurus) in the Progo River, Magelang, Central Java, Indonesia.
Materials and Methods: Ten fish were caught in the Progo River, Magelang, Central Java,
Indonesia. The polymerase chain reaction was the molecular diagnosis to detect the sequences
of DNA of Cytochrome Oxidase I compared to National Center for Biotechnology Information data
(GenBank).
Results: The results showed that the percent identity was not 100% with H. nemurus data from
other locations (GenBank), including Indonesia. The closest percent identity was H. nemurus from
Java Island (Accession ID: MK312566.1) with 97.6% similarity. The genetic mutation that hap�pened might be due to environmental change (pollution) in the Progo River recently.
Conclusions: This study showed a genetic mutation in H. nemurus from Progo River may be
caused by environmental change. Low pollution exposure levels may not be detrimental (lethal)
to fish. However, it can affect fish fertility, which leads to population degradation (gene variation).
Attention must be increased for fish survival in the future.
... In contrast, phylogenetic relationships within the genus remain almost unexplored. The relationships between small numbers of species-mainly from a few species groups of European passerines-were recently analyzed based on molecular data (Doña et al. 2015(Doña et al. , 2017Constantinescu et al. 2018b). ...
The paper provides major references to Trouessartia species described since Santana’s (1976) world revision of this genus, unified diagnoses of 11 currently recognized species groups (africana, appendiculata, capensis, crucifera, estrildae, minutipes, picumni, rosterii, stelgidopteryx, tenuipilata and viduae), and a world checklist of currently known species. The updated checklist includes 147 valid species and is supplemented with data on type hosts, distribution and references to most useful descriptions.
... The COI intraspecific diversity gene in animals was found to be significantly lower than the interspecific diversity. The COI gene barcoding was effective for classifying and identifying vertebrates and invertebrates, so it has been widely used in various fisheries' biological groups [16][17][18][19]. This is what makes COI genes very important in the identification and classification of biological groups. ...
Objective:
This study describes the newest deoxyribonucleic acid (DNA) barcoding Asian redtail catfish (Hemibagrus nemurus) in the Progo River, Magelang, Central Java, Indonesia.
Materials and methods:
Ten fish were caught in the Progo River, Magelang, Central Java, Indonesia. The polymerase chain reaction was the molecular diagnosis to detect the sequences of DNA of Cytochrome Oxidase I compared to National Center for Biotechnology Information data (GenBank).
Results:
The results showed that the percent identity was not 100% with H. nemurus data from other locations (GenBank), including Indonesia. The closest percent identity was H. nemurus from Java Island (Accession ID: MK312566.1) with 97.6% similarity. The genetic mutation that happened might be due to environmental change (pollution) in the Progo River recently.
Conclusions:
This study showed a genetic mutation in H. nemurus from Progo River may be caused by environmental change. Low pollution exposure levels may not be detrimental (lethal) to fish. However, it can affect fish fertility, which leads to population degradation (gene variation). Attention must be increased for fish survival in the future.
... The very high identification efficiencies (98.1 and 97.2% for Dinaric Chthoniidae and Neobisiidae, respectively), as estimated from BCM analyses, suggest that DNA barcoding is an effective approach to identify many morphospecies based on COI divergences. The high success rate of this technique us comparable to other studies involving arachnid groups globally, including eriophyoid mites (99%; 67 ), feather mites (100%; 68 ) and spiders (98%; 69 ). ...
Nested within the Mediterranean biodiversity hotspot, the Dinaric Karst of the western Balkans is one of the world's most heterogeneous subterranean ecosystems and renowned for its highly diverse and mostly endemic fauna. The evolutionary processes leading to both endemism and diversity remain insufficiently understood, and large-scale analyses on taxa that are abundant in both subterranean and surface habitats remain infrequent. Here, we provide the first comprehensive molecular study on Croatian pseudoscorpions, a lineage of arachnids that is common and diverse in both habitats. Phylogenetic reconstructions using 499 COI sequences derived from 128 morphospecies collected across the Dinaric Karst show that: (i) occurrence in karstic microhabitats boosters speciation and endemism in the most diverse genera Chthonius C.L. Koch, 1843 (37 morphospecies) and Neobisium Chamberlin, 1930 (34 morphospecies), (ii) evidence for ongoing diversification is found in many species and species complexes through low optimal thresholds (OTs) and species delineation analyses, and (iii) landscape features, such as mountain ranges, correlate with patterns of genetic diversity in the diverse genus Neobisium. We present two synonymies: Protoneobisium Ćurčić, 1988 = Neobisium, syn. nov., and Archaeoroncus Ćurčić and Rađa, 2012 = Roncus L. Koch, 1873, syn. nov. Overall, our study suggests that karstic microhabitats promote diversification in soil-and cave-dwelling arthropods at all taxonomic levels, but also provide important refugia for invertebrates in past and present periods of environmental change.
... DNA barcoding is also effective in revealing cryptic species, but most past studies have targeted vertebrates (April et al., 2011;Kerr et al., 2007), insects (Dincă et al., 2021;Schmid-Egger et al., 2019), or spiders (Blagoev et al., 2016. Only a few studies have used this approach to examine mites (Donã et al., 2015;de Rojas et al., 2018;Young et al., 2012Young et al., , 2019. The Barcode of Life Data Systems database (BOLD; http:// www.bolds ystems.org/) ...
Eriophyoid mites (Acari: Eriophyoidea) are among the smallest of terrestrial arthropods and the most species‐rich group of herbivorous mites with a high host specificity. However, knowledge of their species diversity has been impeded by the difficulty of their morphological differentiation. This study assembles a DNA barcode reference library that includes 1,850 mitochondrial COI sequences which provides coverage for 45% of the 930 species of eriophyoid mites known from China, and for 37 North American species. Sequence analysis showed a clear barcode gap in nearly all species, reflecting the fact that intraspecific divergences averaged 0.97% versus a mean of 18.51% for interspecific divergences (minimum nearest‐neighbor distances) in taxa belonging to three families. Based on these results, we used DNA barcoding to explore the species diversity of eriophyoid mites as well as their host interactions. The 1,850 sequences were assigned to 531 Barcode Index Numbers (BINs). Analyses examining the correspondence between these BINs and species identifications based on morphology revealed that members of 45 species were assigned to two or more BINs, resulting in 1.16 times more BINs than morphospecies. Richness projections suggest that over 2,345 BINs occurred at the sampled locations. Host plant analysis showed that 89% of these mites (BINs) attack only one or two congeneric host species, but the others have several hosts. Furthermore, host‐mite network analyses demonstrate that eriophyoid mites are high host‐specific, and modularity is high in plant‐mite networks. By creating a highly effective identification system for eriophyoid mites in BOLD, DNA barcoding will advance our understanding of the diversity of eriophyoid mites and their host interactions.
... Species delimitation through an integrative approach to taxonomy constitutes a solution in studies on Trombiculidae. Attention should be paid to the potential presence of cryptic species confirmed in other systematic groups of mites (de Rojas et al. 2002;Navia et al. 2013;Doña et al. 2015;Low et al. 2015). Use of non-morphological criteria in species discrimination of chiggers has been rare up to now. ...
The study aims to ascertain the diversity of trombiculid species associated with Chiroptera in Poland, and for the first time in the case of research on Central European Trombiculidae, we use both DNA and morphology in an integrative taxonomic approach to determine species identities of trombiculids. The research was carried out from 2015 to 2019. In total, 2725 larvae were collected from 300 specimens of bats belonging to 11 species. Deutonymphs were obtained through laboratory rearing of larvae; few larvae and deutonymphs were collected also from bats' daily roosts. The presence of trombiculid larvae on hosts was observed between July and April of the following year, with the highest numbers recorded in autumn, during bat swarming. Male bats were infested more often than females (16.4 vs. 6.6%). The highest infestation rate was recorded for Barbastella barbastellus, Myotis nattereri and Plecotus auritus, and the highest prevalence of chiggers (> 30%) for Myotis bechsteinii and P. auritus. The larvae found on bats occupied the areas with free access to the host’s skin: auricles, tragus, and snout. Morphological identification of specimens to the species level was hindered by the mosaic distribution of diagnostic traits. Morphological analyses indicated the presence of Leptotrombidium russicum and Leptotrombidium spp. in the examined material, whereas molecular analyses additionally suggested three other potential species assigned to the same genus based on the assessed scope of intrageneric variation (ASAP method). We argue that the identification of the parasitic larvae (chiggers) using morphological characters does not address the question of actual species boundaries, which, in turn, affects the inferences about host specificity and host range.
... He has made a breakthrough in taxonomy, wherein mitochondrial DNA, rather than nuclear DNA, formed another most attractive and remarkable protocol in the molecular taxonomy of species by using a novel gene called Cytochrome c Oxidase I (COI). Globally, after Hebert's publication in 2004, several studies have been made and documented the species diversity based on COI, for example, Eurasian birds (Zink et al., 2008), Korean birds (Yoo et al., 2006), Kingfishers (Moyle et al., 2007), Palearctic birds (Kerr et al., 2009a), Scandinavian birds (Johnsen et al., 2010) (Radulovici et al., 2010), Continental patterns of avian diversification (Lijtmaer et al., 2011) House sparrow (Schrey et al., 2011), Avian evolutionary studies (Aliabadian et al., 2013), feather mite studies (Doña et al., 2015), Neotropical birds (Chaves et al., 2015) and Migratory birds (Bilgin et al., 2016). ...
Cytochrome c oxidase I (COI) is one of the mitochondrial genes, an excellent marker used for the DNA barcoding of various organisms. The COI was isolated from twelve different species of waterbirds using the Gene Elute DNA miniprep Kit. The relationship among the waterbirds was assessed by making a phylogenetic tree with the software MEGA-X. The phylogenetic tree's dendrogram showed two main branches in which seven species of water birds aligned one group with four subgroups and the remaining five species aligned with two subgroups based on their similar COI sequences. The Little egret showed 96% similarity with the Cattle egret, Purple heron and Oriental Darter produced 94% similarity with the Grey heron, pond heron shared 90% similarity with the Black-crowned night heron, Asian openbill shared 89% with Pheasant-tailed Jacana and the Common coot 94% similarity with White-breasted waterhen. The pond heron and the Black-crowned night heron showed zero % pairwise distance, but the Cattle egret, Little egret, Oriental Darter, Purple heron and Grey heron showed <0.29%. However, the other seven species of water birds showed >12% of the pairwise distance. Twenty-one conserved haplotypes have been shown in their COI sequences based on the multiple sequences alignment. DNA barcoding identifies the species with their genetic property rather than based on their ecology and behvaviour.
... Species identification based on morphology is not only time-consuming but often inconclusive for closely related species, and even not feasible in the absence of key morphological characters [33]. Conversely, DNA barcode is a quick and reliable molecular method for species identification and can successfully overcome those limitations [34][35][36][37]. In this study, barcodes based on the mitochondrial CO1 gene successfully identified R. maculosus specimens in pupal and larval stages and assigned specimen sequences in GenBank to their appropriate taxon. ...
The oak flea weevil, Rhynchaenus maculosus Yang et Zhang 1991, is a newly emerging pest that severely damages oak (genus Quercus) in China. The first R. maculosus outbreak occurred in 2020 and caused spectacular damage to all oak forests in Jilin province, northeast China. The lack of key morphological characters complicates the identification of this native pest, especially in larva and pupa stages. This is problematic because quick and accurate species identification is crucial for early monitoring and intervention during outbreaks. Here, we provided the first detailed morphological description of R. maculosus at four life stages. Additionally, we used DNA barcodes from larva and pupa specimens collected from three remote locations for molecular identification. The average pairwise divergence of all sequences in this study was 0.51%, well below the 2% to 3% (K-2-parameter) threshold set for one species. All sample sequences matched the R. maculosus morphospecies (KX657706.1 and KX657707.1), with 99.23% to 100% (sequence identity, E value: 0.00) matching success. The tree based on barcodes placed the specimens into the Rhynchaenus group, and the phylogenetic relationship between 62 sequences (30 samples and 32 from GeneBank) had high congruence with the morphospecies taxa. The traditional DNA barcodes were successfully transformed into quick response codes with larger coding capacity for information storage. The results showed that DNA barcoding is reliable for R. maculosus identification. The integration of molecular and morphology-based methods contributes to accurate species identification of this newly emerging oak pest.
... Therefore, since 2003, a molecular approach has been used to identify species within taxa. The use of DNA barcodes, short sequences of mitochondrial genes that are present in a wide number of species, assists traditional taxonomy (Hebert et al. 2003a(Hebert et al. , 2003b and is a useful tool for resolving taxonomic issues (Hebert and Gregory 2005;Okassa et al. 2009;Doña et al. 2015;Mendoza et al. 2016;Hernández-Triana et al. 2019). In addition, the molecular barcode provides the possibility to identify immature stages, which cannot be distinguished in many groups solely on the basis of morphological observations, because the DNA sequences of an individual are identical at all stages of development (Thomas et al. 2005;Ahrens et al. 2007;Okassa et al. 2012;Bakhoum 2018). ...
The bean Phaseolus vulgaris is widely consumed by the Congolese population, because it is rich in protein and has a good nutritional quality. Beetles of the subfamily Bruchinae are pests of bean seeds in the field and storage in the Republic of Congo (RC). Their attacks cause a reduction in seed weight, leading to a decrease in the nutritional and commercial value of the seeds. The purpose of our study was to identify the post-harvest diversity of bean bruchids in two bean producing regions in the Congo (Niari and Bouenza) using molecular taxonomy (12S rRNA marker). The bean weevil, Acanthoscelides obtectus (Say), and cowpea weevil, Callosobruchus maculatus (Fabricius), were identified. Acanthoscelides obtectus, recognised as the main pest of beans, was found in all ten localities sampled, whereas C. maculatus, recognised as the main pest of pigeon peas (Cajanus cajan) and chickpeas (Cicer arietinum), was found in two of the ten. Better knowledge of the diversity of bruchids present in the RC will make it possible to propose effective control methods with little impact on environmental and human health.
... The use of any dataset carries some intrinsic biases, such as issues with species identification and heterogeneity of sampling efforts . We believe our dataset is robust to avoid misidentifications, due to the expected low number of cryptic species in feather mites (Doña et al., 2015(Doña et al., , 2016. On the other hand, the predominance of migratory birds would generate bias because of the seasonal habit of these species. ...
Aim
The simultaneous influence of abiotic and biotic factors as main drivers of global species distributions remains poorly understood, especially in host‐dependent groups. In this study, we diverge from traditional macroecological approaches by considering both biotic (avian species diversity) and abiotic (climatic) factors in determining the global distribution pattern of feather mite species richness, one of the most abundant and diverse bird ectosymbionts.
Location
Global.
Methods
We used a global dataset of feather mite–bird interactions published in 2016, complemented with an up‐to‐date literature survey. We created statistical models designed to explain the effect of abiotic (i.e., temperature, precipitation and energy‐related variables) and biotic factors (bird species richness) on the species richness of feather mites. We used these models to predict global distribution patterns of mites and estimate each explanatory variable's relative importance in temperate and tropical regions.
Results
According to our models, bird species richness accounts for ~63% of the global distribution pattern of mites, which is ten times more relevant than climatic variables. Among abiotic drivers, precipitation intensity and seasonality were the most important variables, accounting for 10% of mite species richness. This figure is lower in tropical regions, where biotic factors are seven times more important than in temperate regions.
Main conclusions
We demonstrate that global mite diversity was primarily determined by biotic and, to a lesser extent, abiotic factors. The relative importance of the predictive variables, however, varied between tropical and temperate regions. The strong association between bird species richness and feather mite species diversity at a global scale raises concerns about the potential for future co‐extinctions.
... This is particularly true for some morphologically cryptic insect pests. In these circumstances, our study, in accordance with previous studies [20,[82][83][84][85][86], indicated that DNA barcoding offers a fast and reliable means of identifying organisms. However, incorporating DNA barcodes with morphological and ecological characters will aid the rapid and authentic classification of Nigerian butterflies. ...
Accurate identification of species is a prerequisite for successful biodiversity management and further genetic studies. Species identification techniques often require both morphological diagnostics and molecular tools, such as DNA barcoding, for correct identification. In particular, the use of the subunit I of the mitochondrial cytochrome c oxidase (COI) gene for DNA barcoding has proven useful in species identification for insects. However, to date, no studies have been carried out on the DNA barcoding of Nigerian butterflies. We evaluated the utility of DNA barcoding applied for the first time to 735 butterfly specimens from southern Nigeria. In total, 699 DNA barcodes, resulting in a record of 116 species belonging to 57 genera, were generated. Our study sample comprised 807 DNA barcodes based on sequences generated from our current study and 108 others retrieved from BOLD. Different molecular analyses, including genetic distance-based evaluation (Neighbor-Joining, Maximum Likelihood and Bayesian trees) and species delimitation tests (TaxonDNA, Automated Barcode Gap Discovery, General Mixed Yule-Coalescent, and Bayesian Poisson Tree Processes) were performed to accurately identify and delineate species. The genetic distance-based analyses resulted in 163 well-separated clusters consisting of 147 described and 16 unidentified species. Our findings indicate that about 90.20% of the butterfly species were explicitly discriminated using DNA barcodes. Also, our field collections reported the first country records of ten butterfly species—Acraea serena, Amauris cf. dannfelti, Aterica galena extensa, Axiocerces tjoane rubescens, Charaxes galleyanus, Papilio lormieri lormieri, Pentila alba, Precis actia, Precis tugela, and Tagiades flesus. Further, DNA barcodes revealed a high mitochondrial intraspecific divergence of more than 3% in Bicyclus vulgaris vulgaris and Colotis evagore. Furthermore, our result revealed an overall high haplotype (gene) diversity (0.9764), suggesting that DNA barcoding can provide information at a population level for Nigerian butterflies. The present study confirms the efficiency of DNA barcoding for identifying butterflies from Nigeria. To gain a better understanding of regional variation in DNA barcodes of this biogeographically complex area, future work should expand the DNA barcode reference library to include all butterfly species from Nigeria as well as surrounding countries. Also, further studies, involving relevant genetic and eco-morphological datasets, are required to understand processes governing mitochondrial intraspecific divergences reported in some species complexes.
... The intraspecific diversity of the COI gene in animals had lower resolving power than interspecific diversity as a DNA barcode. The COI gene is used extensively for DNA barcoding in other biological groups, but less so for fish (Doña et al., 2015). ...
Fish is a fundamentally healthy food, loaded with essential nutrients, high protein content, vitamin D, and omega-three fatty acid. Mislabeling is a common problem in the fish industry that causes an imbalance in prices and fluctuation in the market. DNA barcoding is a potential technique for authentication of mislabeled and misidentified fish species. In this study, 11 freshwater and 6 marine fish species were used for DNA barcoding and further authentication using the mitochondrial cytochrome b (Cyt b) gene. Cyt b was amplified using PCR, producing an average read length of 1,141 bp. The obtained sequences were compared to the National Center for Biotechnology Information database (NCBI) using the Basic Local Alignment Search Tool (BLAST). The average AT content (55.20%) was higher than the average GC content (44.78%) in marine and freshwater fish species. The mean genetic Kimura 2-parameter distances for species, genus, families, and orders were 0.311, 0.308, 0.023, and 0.337, respectively. Phylogenetic tree analysis revealed that most of the freshwater fish species clustered together due to the fact that they were in the same order or family, while the marine fish species clustered distantly. Single nucleotide polymorphism (SNP) analysis of all species in the study revealed distinct features regarding unique sites. All fish species could be identified based on their unique SNP profiles. Based on SNP data, DNA sequence based QR codes were developed for accurate identification of fish species. This is the first study to develop DNA-based QR barcodes for proper authentication of species during the chain of custody using simple technology.
... DNA barcoding uses a fragment of the mitochondrial gene cytochrome c oxidase subunit I (CO1) as a unique species diagnosis/identification tool in the animal kingdom (Hebert et al., 2003), with analogous single to several locus protocols applied for vascular plants, ferns, mosses, algae and fungi (Saunders, 2005;Kress & Erickson, 2007;Nitta, 2008;Chase & Fay, 2009;Liu et al., 2010), protists (Scicluna, Tawari & Clark, 2006), and prokaryotes (Barraclough et al., 2009). Due to relative ease and inexpensive sequencing, DNA barcoding is a popular tool in species identification and taxonomic applications (e.g., Doña et al., 2015;Xu et al., 2015; see also Collins & Cruickshank, 2013), and the method is no longer fundamentally controversial at the species level (Pentinsaari, Hebert & Mutanen, 2014;Lopardo & Uhl, 2014;Anslan & Tedersoo, 2015;Wang et al., 2015). ...
The use of unique DNA sequences as a method for taxonomic identification is no longer fundamentally controversial, even though debate continues on the best markers, methods, and technology to use. Although both existing databanks such as GenBank and BOLD, as well as reference taxonomies, are imperfect, in best case scenarios “barcodes” (whether single or multiple, organelle or nuclear, loci) clearly are an increasingly fast and inexpensive method of identification, especially as compared to manual identification of unknowns by increasingly rare expert taxonomists. Because most species on Earth are undescribed, a complete reference database at the species level is impractical in the near term. The question therefore arises whether unidentified species can, using DNA barcodes, be accurately assigned to more inclusive groups such as genera and families—taxonomic ranks of putatively monophyletic groups for which the global inventory is more complete and stable. We used a carefully chosen test library of CO1 sequences from 49 families, 313 genera, and 816 species of spiders to assess the accuracy of genus and family-level identifications. We used BLAST queries of each sequence against the entire library and got the top ten hits resulting in 8160 hits. The percent sequence identity was reported from these hits (PIdent, range 75-100%). Accurate identification (PIdent above which errors totaled less than 5%) occurred for genera at PIdent values > 95 and families at PIdent values ≥ 91, suggesting these as heuristic thresholds for generic and familial identifications in spiders. Accuracy of identification increases with numbers of species/genus and genera/family in the library; above five genera per family and fifteen species per genus all identifications were correct. We propose that using percent sequence identity between conventional barcode sequences may be a feasible and reasonably accurate method to identify animals to family/genus. However, the quality of the underlying database impacts accuracy of results; many outliers in our dataset could be attributed to taxonomic and/or sequencing errors in BOLD and GenBank. It seems that an accurate and complete reference library of families and genera of life could provide accurate higher level taxonomic identifications cheaply and accessibly, within years rather than decades.
... The COI is a mitochondrial gene, whose partial length of 655 bp is analyzed in fish based on its universal primers. 655 bp COI gene fragment, is used as a barcode and extremely effective for classification and identification of fish (Doña et al., 2015;Staffen et al., 2017). DNA barcoding technology comes with various merits. ...
... Synonymies have also been cited in the scientific literature, which adds to the difficulty of using only traditional taxonomy for identification (Moraes et al. 1986(Moraes et al. , 2004Chant & McMurtry 1994. Use of the DNA barcode using short sequences of mitochondrial genes that are present in a wide number of species assists traditional taxonomy (Hebert et al. 2003) and presents a useful tool that has permitted the resolution of a number of questions of this taxonomic order (Hebert & Gregory 2005;Doña et al. 2015;Mendoza et al. 2016;M Hernández-Triana et al. 2019). Within the mites, the molecular barcode has allowed the confirmation of the specific status of closely related species using 12S rRNA (Okassa et al. 2009(Okassa et al. , 2010Kanouh et al. 2010;Tixier et al. 2014;Navia et al. 2014) and ITSS (Tixier et al. 2006;Kanouh et al. 2010;Navia et al. 2014). ...
The purpose of this study was to identify the taxonomic status of several individuals belonging to the genus Euseius, collected from three host plants (cassava, okra, and chilli) in the Republic of the Congo, using morphometric and molecular analysis. For this, hierarchical ascending classification on principal component analysis was performed using morphological measurements from 21 and 22 individuals of E. fustis and Euseius sp., respectively, with the original description of Euseius neodossei considered. For molecular analysis, two DNA fragments were amplified and sequenced: the mitochondrial 12S rRNA and the nuclear ribosomal region ITSS. This integrative approach found that (1) morphological differences existed between the Euseius sp. and both E. neodossei and E. fustis, and (2) both mitochondrial DNA fragments showed a clear delineation between Euseius sp. and E. fustis. The results obtained highlight the value of using integrative taxonomy to improve the identification of species in under-sampled areas and the need to increase the number of DNA sequences deposited in GenBank database from this region. This is the first time that DNA sequences of mites from the Congo (Brazzaville) have been deposited in this database.
... The COI gene is a commonly applied marker for identification studies in various animals, such as in Antartic ocean's animals (Grant et al 2010); marine crustacea (Radulovici et al 2009), marine metazoa (Bucklin et al 2011). The COI gene sequences was also effective for classifying and identifying vertebrates and invertebrates, and has been widely used in various biological groups (Weigt et al 2012;Dona et al 2015;Nzelu et al 2015;Staffen et al 2017). Similar results were obtained in a barcoding study on fish species (Ward et al 2005(Ward et al , 2008a(Ward et al , 2008bHubert et al 2008;Steinke et al 2009;Lakra et al 2011;Liu et al 2013). ...
This study determined the genetic characteristics of ricefish from Lake Poso, Sulawesi Tengah-Indonesia based on cytochrome c oxidase subunit I (COI) gene marker. We collected and analyzed 2 tissue samples from Lake Poso, about 54 sequences from GenBank, and 100 sequences from Bold taxonID of ricefishes. The two tissue samples were extracted to obtain whole genome then amplified and sequenced to obtained COI nucleotides. The nucleotide frequencies were A = 23.8%, T/U = 29.8%, C = 26.8%, and G = 19.6%. The AT content (53.85%) in the COI region of ricefish was higher than that of GC (46.2%). We identified 217 codons from two sequences of our samples. Ninety-six percent of nucleotide changes were observed at the 3rd codon position of COI sequence, but the amino acid compositions translated by COI sequences of two samples stayed the same. We identified also 170 cut positions from two sequences of our samples. The phylogenetic tree estimated by Neighbour-joining method, Blastn and Boldsystem results, and nucleotide composition indicated that the ricefish is Oryzias nebulosus. Individuals from operational taxonomic units designated as same ricefish species, supporting morphological evidence for each of these being similar species. It is suggested that the COI barcoding can be used to identify ricefish species of Oryzias.
... In 1993, only 49 Acari DNA sequences were reported in the GenBank database, whereas at the beginning of 2006 there were almost 93,000 sequences (Dabert 2006). DNA amplification and DNA sequencing techniques have greatly been improved after this time (Dabert 2006;Desloire et al. 2006;Auger et al. 2013;Dona et al. 2015). In some biological programs using phytoseiid mites, only molecular assays can reliably identify the species status, because of the release of mixed species of predators in the field for biological control and only small amounts of initial material are necessary for DNA sequences (Dos Santos & Tixier 2018). ...
Obtaining clean DNA from mites to be used in molecular studies has always been a challenging task. In this study, five protocols (CTAB-based handmade kit, salting out protocol, STE protocol, modified CTAB-based protocol and wizard Genomic DNA purification kit) were evaluated to extract DNA from Typhlodromus (Anthoseius) bagdasarjani. Quantitative and qualitative analyses of the DNA concentrations and a comparison between the ratio of absorption of wavelength from 260 to 280 nm in a completely randomized split plot design for five extraction protocols and three levels of 1, 5, 10 mites were performed. PCR protocol was used to examine the amplification of the ITS1 fragment from the specimens. The results showed that wizard kit and CTAB modified protocol were the bests. However, economically, modified CTAB-based protocol is the best choice for extensive experiments, since this protocol allows extracting DNA from an individual mite and provides the highest concentration DNA (143.35±0.85ng) and the best OD (1.80±0.03) for molecular studies.
... The barcode region has been useful for delineating species in various groups of parasitiform and acariform mites (e.g., Roy et al. 2009, Knee et al. 2012a, Glowska et al. 2014, Doña et al. 2015, Ondrejicka et al. 2016, Fisher et al. 2017. In Canada, approximately 60-70% of 230 morphologically recognized species of oribatids across 45 families are currently supported by barcode clusters (M Young and L Lumley unpubl. ...
Summaries of taxonomic knowledge are provided for all acarine groups in Canada, accompanied by references to relevant publications, changes in classification at the family level since 1979, and notes on biology relevant to estimating their diversity. Nearly 3000 described species from 269 families are recorded in the country, representing a 56% increase from the 1917 species reported by Lindquist et al. (1979). An additional 42 families are known from Canada only from material identified to family- or genus-level. Of the total 311 families known in Canada, 69 are newly recorded since 1979, excluding apparent new records due solely to classification changes. This substantial progress is most evident in Oribatida and Hydrachnidia, for which many regional checklists and family-level revisions have been published. Except for recent taxonomic leaps in a few other groups, particularly of symbiotic mites (Astigmata: feather mites; Mesostigmata: Rhinonyssidae), knowledge remains limited for most other taxa, for which most species records are unpublished and may require verification. Taxonomic revisions are greatly needed for a large majority of families in Canada. Based in part on species recorded in adjacent areas of the USA and on hosts known to be present here, we conservatively estimate that nearly 10,000 species of mites occur in Canada, but the actual number could be 15,000 or more. This means that at least 70% of Canada’s mite fauna is yet unrecorded. Much work also remains to match existing molecular data with species names, as less than 10% of the ~7500 Barcode Index Numbers for Canadian mites in the Barcode of Life Database are associated with named species. Understudied hosts and terrestrial and aquatic habitats require investigation across Canada to uncover new species and to clarify geographic and ecological distributions of known species.
... Identifications of eggs and instars of pest species are difficult on the basis of morphological characters (Ball & Armstrong, 2006). In these circumstances, DNA barcoding provide quick and authentic means for species identification (Hebert et al. 2004;Ball and Armstrong 2006;Blagoev et al. 2013;Raso et al. 2014;Dona et al. 2015;Xu et al. 2015). The authenticity of DNA barcoding relies on barcode gap. ...
Accurate identification of agricultural pests is key requirement for the successful integrated pest management (IPM) program. However, due to limitations of conventional morphological methods, other molecular method like DNA barcoding is used. The current study was designed to evaluate the accuracy of morphological identification of insect pests using DNA barcoding. Morphologically, a total of 247 insect pests, representing 10 families, 18 genera, 22 species were identified. However, molecular identifications confirmed the presence of 11 families, 16 genera, and 20 species of agricultural pests. A total of 59 specimens were processed for DNA barcoding but genomic sequences of mt COI gene up to 600 bp were revived from 48 samples. Specimens that were misidentified through morphological studies were placed to their appropriate taxon, using this molecular approach. Results revealed the existence of clear barcode gap for different pest species. Moreover, the values of distance with the nearest neighbour recorded were higher than the maximum intra-sequence divergences for all species. It is concluded that DNA barcoding is a reliable technique for identification of agricultural pests, especially for immature stages when morphometric studies are ambiguous and will be helpful in the development of more effective pest management options for regulating pest species.
... In 2003, Hebert et al. [10] proposed DNA barcoding technology, in which the mitochondrial cytochrome c oxidase subunit I (COI) gene sequence was used as a barcode for species identification with the expectation of barcoding all species for the purpose of species identification and classification. It was found that the intraspecific diversity of the COI gene in animals was significantly lower than the interspecific diversity, using the COI gene as a barcode was effective for classifying and identifying vertebrates and invertebrates, and the COI gene has been widely used in various biological groups [11][12][13][14]. Compared with the traditional morphological classification methods, the advantages of the DNA barcoding technology are mainly as follows: 1) Some species have extremely similar external morphological characteristics; therefore, it is difficult to distinguish them from each other merely by morphological characteristics. ...
DNA barcoding based on a fragment of the cytochrome c oxidase subunit I (COI) gene in the mitochondrial genome is widely applied in species identification and biodiversity studies. The aim of this study was to establish a comprehensive barcoding reference database of fishes in the Taiwan Strait and evaluate the applicability of using the COI gene for the identification of fish at the species level. A total of 284 mitochondrial COI barcode sequences were obtained from 85 genera, 38 families and 12 orders of fishes. The mean length of the sequences was 655 base pairs. The average Kimura two parameter (K2P) distances within species, genera, families, orders and classes were 0.21%, 6.50%, 23.70% and 25.60%, respectively. The mean interspecific distance was 31-fold higher than the mean intraspecific distance. The K2P neighbor-joining trees based on the sequence generally clustered species in accordance with their taxonomic classifications. High efficiency of species identification was demonstrated in the present study by DNA barcoding, and we conclude that COI sequencing can be used to identify fish species.
... NGS barcoding data can also be used for studying species richness and community patterns over time and space because each barcode comes with precise information on collection locality and date. demonstrated similar discriminatory powers of "minibarcodes" (e.g., Doña et al., 2015;Hajibabaei et al., 2006;Meusnier et al., 2008), but they were comparatively small scale while large scale in silico studies implied that minibarcodes may have lower power for discriminating dense species samples (Hajibabaei et al., 2006). We note, however, that our study is a limited test; results may differ when more intraspecific variability across the species' ranges is sampled (Bergsten et al., 2012). ...
Molecular markers are increasingly used for species identification and new taxa description. However, rules to determine frontiers between populations and species are not clear depending on taxa considered. For mites, few studies deal with molecular diagnoses, making rules for associated decision difficult. The present study focuses on a species of the predatory mite family Phytoseiidae (Phytoseius finitimus), considered for biological control of mites and small insect pests in fruit orchards and vineyards in the Mediterranean basin. This paper aims to elucidate the causes of great molecular variations and questions the occurrence of cryptic species. Molecular (12S rRNA, CytB mtDNA, ITSS) and morphological analyses were performed on four populations collected in Corsica and Italy in crops (vine and kiwi) and in an uncultivated environment (Viburnum lantana). Different methods for identifying species have been used (tree approaches, distances and ABGD algorithms). A reference database of distances within and between Phytoseiidae species has been elaborated to inform the present question and to assist with further diagnosis within Acari. Mitochondrial DNA analyses show that specimens from V. lantana were well separated from the three other populations with high genetic distances, suggesting the existence of a cryptic species. Molecular ITSS analyses coupled with morphological features show however that the four populations seem to belong to the same species. The great mitochondrial polymorphism is discussed in regards to: (i) genetic distances reported for Phytoseiidae species and (ii) potential biological differences between populations (cultivated versus uncultivated areas). This study clearly emphasizes the necessity of integrative taxonomy approaches for diagnosis decisions. Furthermore, based on the polymorphism herein detected, maximal intraspecific distances are proposed (9, 23 and 2.8 % for 12S rRNA, CytB mtDNA and ITSS) for diagnosis decisions within Phytoseiidae. Further statistical analyses are however clearly required to determine statistical error for general and reliable decision making.
Positive correlations between the shapes of male and female sexual structures can be interpreted as cooperative or as combative. In the feather mite genus Trouessartia Canestrini, 1899, the spermaducts of females range from entirely internal to extending externally for varying lengths, while male primary genitalia range from gracile to massive. Males also possess a pair of adanal suckers used to hold onto the dorsal surface of the female during copulation. In the area of this attachment, females exhibit ornamentation and have strongly developed dorsal setae (setae h1), which we hypothesized serve to weaken the male’s hold during copulation. In male and female Trouessartia from 51 bird species, we compared female external spermaduct length and male genitalic ‘massiveness’ and explored whether patterns of female dorsal ornamentation and/or h1 seta size correlate with male adanal sucker size. Our results indicate that females with longer external spermaducts are associated with males with relatively massive genitalia. However, we found no significant relationship between male adanal sucker size and female ornamentation or h1 seta size. Further information regarding how the genitalia interact during sperm transfer is necessary to interpret correlations in genitalia size and strong intersexual difference in dorsal ornamentation and seta size in Trouessartia.
Morphological identification of gastropods can be difficult considering the larva of species and high similarity among the same genera. DNA barcoding has been widely used in species identification and biodiversity research. The purpose of this study was to explore the feasibility of using the cytochrome c oxidase subunit I (COI) gene for the identification of gastropod species and to construct a reliable DNA barcoding reference database of gastropods in Hainan island, China. A total of 306 mitochondrial COI barcode sequences were obtained from 120 species, 35 families and 7 orders of gastropods. The average length of the sequence was 640 base pairs. The average genetic distances based on Kimura two parameter (K2P) within species, genera, families, orders and classes were 0.9 %, 14.7 %, 18.9 %, 24.5 % and 28.6 %, respectively. Most of the gastropod species could be identified using COI sequences. Our results confirmed that the identification method combining morphology and DNA barcode greatly improved the efficiency of species identification. In this study, we found three new record species in China, namely Semiricinula tissoti (Petit de la Saussaye, 1852), Engina alveolata (Kiener, 1836) and Wallaconchis ater (Lesson, 1831). Overall, this study revealed that the identification of gastropods by DNA barcoding is efficient, and COI sequencing technology can be used for the identification of gastropod species and thereby can be used to manage fisheries and assess biodiversity.
Whether ribosomal genes can be used as DNA barcodes for molecular identification of Demodex (Acariformes: Demodicidae) is unclear. To examine this, Demodex folliculorum, D. brevis, D. canis, and D. caprae were collected for DNA extraction, rDNA fragments amplification, sequencing, and analysis. The V2 and V4 regions of SSU rDNA; D5, D6, and D8 regions of LSU rDNA; and ITS region were obtained from the four morphospecies. BLAST analysis showed that the obtained sequences matched those of Demodex or Aplonobia (Acariformes: Tetranychidae) in Raphignathae. Phylogenetic trees derived from V2, V4, D5, D6, and D8 regions, but not from ITS region, showed that the four species of Demodex clustered independently. Sequence divergence analysis further demonstrated that D5, D6, and D8 regions had obvious barcoding gap between intraspecific and interspecific divergences, with the gap of D5 (16.91%) larger than that of D6 (11.82%) and D8 (4.66%). The V2 and V4 regions did not have a barcoding gap, as the intraspecific and interspecific divergences partially overlapped. For the ITS region, intraspecific and interspecific divergences completely overlapped. These results suggest that the D5, D6, and D8 regions of LSU rDNA, especially D5, are suitable DNA barcodes for Demodex.
Mites of the family Pachylaelapidae Berlese, 1913 are a group of predatory mites with free-living species, feeding on arthropods and soil-inhabiting nematodes. Identification of these mites is difficult, because of morphological similarities in a significant number of species and also the fact that new taxons descriptors have referred to some morphological features that have not sufficient distinction criteria to differentiate them from close taxons. Therefore, the use of new identification methods can be helpful in this context. In the present study, along with identification, based on morphological characteristics and size (morphometric identification), we used a molecular method, based on the sequencing of the internal transcribed spacer (ITS) gene to identify two species Olopachys compositus Koroleva, 1976 and Olopachys caucasicus Koroleva, 1976. After DNA extraction, ITS region were amplified and sequenced, using the appropriate primers and afterward the sequences were recorded in GeneBank. Alignment of sequences was performed and the genetic differences between species was calculated as 2.3 percent. As a result of these investigations, in addition to the separation of the two species O. compositus and O. caucasicus based on morphological differences, the molecular method also well confirmed the difference between these two species.
Obligate symbionts may be genetically structured among host individuals and among phenotypically distinct host populations. Such processes may in turn determine within-host genetic diversity of symbionts, which is relevant for understanding symbiont population dynamics. We analysed the population genetic structure of two species of feather mites ( Proctophyllodes sylviae and Trouessartia bifurcata ) in migratory and resident blackcaps Sylvia atricapilla that winter sympatrically. Resident and migratory hosts may provide mites with habitats of different qualities, what might promote specialization of mite populations. We found high genetic diversity of within-host populations for both mite species, but no sign of genetic structure of mites between migratory and resident hosts. Our results suggest that, although dispersal mechanisms between hosts during the non-breeding season are unclear, mite populations are not limited by transmission bottlenecks that would reduce genetic diversity among individuals that share a host. Additionally, there is no evidence that host phenotypic divergence (associated with the evolution of migration and residency) has promoted the evolution of host-specialist mite populations. Unrestricted dispersal among host types may allow symbiotic organisms to avoid inbreeding and to persist in the face of habitat heterogeneity in phenotypically diverse host populations.
In check-list of feather mites recorded from 90 species of the passerines living in the North-West of Russia is given. It includes 118 mite species belonging to 21 genera, 9 families, 2 superfamilies. The data for each mite species contain a list of passerine hosts recorded in the North-West of Russia, regions of recording, type host, main host-parasite associations and geographical distribution in general lines. A taxonomic composition of the feather mite fauna discovered and some its peculiarities are discussed. The brief review of recent papers on feather mites associated with the passerines in Europe and the European part of Russia is given.
Although much biological research depends upon species diagnoses, taxonomic expertise is collapsing. We are convinced that the sole prospect for a sustainable identification capability lies in the construction of systems that employ DNA sequences as taxon 'barcodes'. We establish that the mitochondrial gene cytochrome c oxidase I (COI) can serve as the core of a global bioidentification system for animals. First, we demonstrate that COI profiles, derived from the low-density sampling of higher taxonomic categories, ordinarily assign newly analysed taxa to the appropriate phylum or order. Second, we demonstrate that species-level assignments can be obtained by creating comprehensive COI profiles. A model COI profile, based upon the analysis of a single individual from each of 200 closely allied species of lepidopterans, was 100% successful in correctly identifying subsequent specimens. When fully developed, a COI identification system will provide a reliable, cost-effective and accessible solution to the current problem of species identification. Its assembly will also generate important new insights into the diversification of life and the rules of molecular evolution.
A new feather mites species, Proctophyllodes valchukae sp. n., is described from the Long-tailed Tit, Aegithalos caudatus (Linnaeus, 1758) (Passeriformes: Aegithalidae), captured in the Primoriye (Russian Far East). The new species belongs to the tricetratus species group and is most closely related to P. stachyris Atyeo et Braasch, 1966. For the first time for feather mites the standard morphological description is supplemented by sequence data of the mitochondrial cytochrome c oxidase subunit I gene fragment (COI) and nuclear D2 region of 28S rDNA
Here we present the first evidence of female dimorphism in ectoparasitic quill mites of the family Syringophilidae (Actinotrichida: Prostigmata: Cheyletoidea). Stibarokris phoeniconaias Skoracki et OConnor, 2010 and Ciconichenophilus phoeniconaias Skoracki et OConnor, 2010 so far have been treated as two distinct species cohabiting inside the quills of feathers of the lesser flamingo Phoeniconaias minor (Geoffroy Saint-Hilaire) and the American flamingo Phoenicopterus ruber Linnaeus. Although females of these species differ morphologically by the extent of body sclerotisation, presence/absence of lateral hypostomal teeth, and shape of dorsal setae, their important common features are the lack of leg setae vs II, and both stylophore and peritremes shape. Here, we apply the DNA barcode markers to test whether the differences between S. phoeniconaias and C. phoeniconaias have a genetic basis, indicating that they really are distinct taxa, or whether they just represent two morphs of a single species. All analysed sequences (616 bp for COI and 1159 bp for 28S rDNA) obtained for specimens representing females of both studied taxa as well as male, tritonymph, protonymph and larva of S. phoeniconaias were identical, which indicates that S. phoeniconaias and C. phoeniconaias are conspecific. The formal taxonomic consequence of our results is denial of the genus status of Ciconichenophilus Skoracki et OConnor, 2010 and species status of C. phoeniconaias, and recommendation that they should be treated as junior synonyms of Stibarokris Kethley, 1970 and S. phoeniconaias, respectively.
Gradients of environmental stress may affect biotic interactions in unpredictable ways responding to climate variation, depending on the abiotic stress tolerance of interacting partners. Here, we study the effect of local climate on the intensity of feather mites in six mountain passerines along a 1400 m elevational gradient characterized by shifting temperature and rainfall. Although obligatory symbionts of warm-blooded organisms are assumed to live in mild and homeothermic environments, those inhabiting external, non-blood-irrigated body portions of the host organism, such as feather mites, are expected to endure exposure to the direct influence of a fluctuating climate. As expected, feather mite intensity declined with elevation in all bird species, a pattern that was also found in cold-adapted passerines that have typical alpine habits. The elevation cline was mainly explained by a positive effect of the average temperature upon mite intensity in five of the six species studied. Precipitation explained less variance in mite intensity than average temperature, and showed a negative correlation in half of the studied species. We found no climate-driven migration of mites along the wings of birds, no replacement of mite species along elevation gradients and no association with available food resources for mites (estimated by the size of the uropygial gland). This study suggests that ectosymbionts of warm-blooded animals may be highly sensitive to climatic variation and become less abundant under stressful environmental conditions, providing empirical evidence of the decline of specialized biotic interactions among animal species at high elevations.
As a discipline, phylogenetics is becoming transformed by a flood of molecular data. These data allow broad questions to be
asked about the history of life, but also present difficult statistical and computational problems. Bayesian inference of
phylogeny brings a new perspective to a number of outstanding issues in evolutionary biology, including the analysis of large
phylogenetic trees and complex evolutionary models and the detection of the footprint of natural selection in DNA sequences.
We announce the release of an advanced version of the Molecular Evolutionary Genetics Analysis (MEGA) software, which currently contains facilities for building sequence alignments, inferring phylogenetic histories, and conducting molecular evolutionary analysis. In version 6.0, MEGA now enables the inference of timetrees, as it implements our RelTime method for estimating divergence times for all branching points in a phylogeny. A new Timetree Wizard in MEGA6 facilitates this timetree inference by providing a graphical user interface (GUI) to specify the phylogeny and calibration constraints step-by-step. This version also contains enhanced algorithms to search for the optimal trees under evolutionary criteria and implements a more advanced memory management that can double the size of sequence data sets to which MEGA can be applied. Both GUI and command-line versions of MEGA6 can be downloaded from www.megasoftware.net free of charge.
Because many animal species are undescribed, and because the identification of known species is often difficult, interim taxonomic nomenclature has often been used in biodiversity analysis. By assigning individuals to presumptive species, called operational taxonomic units (OTUs), these systems speed investigations into the patterning of biodiversity and enable studies that would otherwise be impossible. Although OTUs have conventionally been separated through their morphological divergence, DNA-based delineations are not only feasible, but have important advantages. OTU designation can be automated, data can be readily archived, and results can be easily compared among investigations. This study exploits these attributes to develop a persistent, species-level taxonomic registry for the animal kingdom based on the analysis of patterns of nucleotide variation in the barcode region of the cytochrome c oxidase I (COI) gene. It begins by examining the correspondence between groups of specimens identified to a species through prior taxonomic work and those inferred from the analysis of COI sequence variation using one new (RESL) and four established (ABGD, CROP, GMYC, jMOTU) algorithms. It subsequently describes the implementation, and structural attributes of the Barcode Index Number (BIN) system. Aside from a pragmatic role in biodiversity assessments, BINs will aid revisionary taxonomy by flagging possible cases of synonymy, and by collating geographical information, descriptive metadata, and images for specimens that are likely to belong to the same species, even if it is undescribed. More than 274,000 BIN web pages are now available, creating a biodiversity resource that is positioned for rapid growth.
Feather mites (Astigmata: Psoroptidia) are commensals and parasites of almost all recent bird species and are closely associated with individual bird hosts. Although the evolutionary mechanisms in feather mites have been intensively studied on medium and higher taxonomic levels, the initial stage of the speciation process is completely unknown. In order to investigate the extent of the genetic isolation, mite populations belonging to oligoxenous feather mite species Avenzoaria totani (collected from tattlers Tringa glareola and T. totanus) and Proctophyllodes stylifer (collected from tits Parus caeruleus, P. montanus, and P. major) were studied. Sequence analyses were carried out on internal transcribed spacers (ITS1 and ITS2) of the nuclear rDNA. In case of A. totani each of the methods used for phylogeny reconstruction has revealed the division of the species into two distinct groups of haplotypes. Nucleotide sequences of both ITS1 and ITS2 of mites sampled from T. totanus and T. glareola built two distinct monophyla. Sequence variation in ITS1 provides evidence that Proctophyllodes stylifer inhabiting Parus caeruleus is genetically isolated from populations living on P. major and P. montanus, and shows lack of genetic isolation between populations of Pr. stylifer from P. montanus and P. major. Duplication of haplotype lines in ITS2 on the ancestor of recent hosts of Pr. stylifer has probably lead to the origin of two lines of mites evolving separately but in parallel on descendent hosts.
Typical species (re)description of feather mites is based on characteristics of the external morphology. We propose extending this standard by sequence data of the cytochrome oxidase subunit I gene fragment (DNA barcode region chosen by the Consortium for the Barcode of Life). The method of nondestructive DNA isolation, which enables to obtain intact feather mite exoskeleton for subsequent morphological analysis, is shown. The description of new feather mite species Glaucalges tytonis (Analgoidea: Xolalgidae) from the plumage of the barn owl Tyto alba (Scopoli, 1769) (Strigiformes: Tytonidae) is presented as an example of the new procedure that may be implemented as well for feather mites as other groups of Acari.
DNA barcoding is a technique for identifying organisms based on a short, standardized fragment of genomic DNA. The standardized sequence region is called a DNA barcode because it is like a barcode tag for each taxon. Since the proposition of this concept and the launch of a large project named the Barcode of Life, this simple technique has attracted attention from taxonomists, ecologists, conservation biologists, agriculturists, plant-quarantine officers and others, and the number of studies using the DNA barcode has rapidly increased. The extreme diversity of insects and their economical, epidemiological and agricultural importance have made this group a major target of DNA barcoding. However, there is some controversy about the utility of DNA barcoding. In this review, we present an overview of DNA barcoding and its application to entomology. We also introduce current advances and future implications of this promising technique.
Feather mites (suborder Astigmata, superfamilies Analgoidea, Pterolichoidea and Freyaniidae) are among the commonest ec-tosymbionts of birds. Most researchers have assumed they are parasites, having negative effects on hosts. Here we present evidence that suggests that feather mites may not be parasites. We develop a framework for considering conditional outcomes in these interspecific associations, dealing with different kinds of relationships between symbionts. The non-parasitic status of feather mites is supported by a literature review as well as by preliminary data on mites' food. We illustrate symbiotic rela-tionships with a graphical model showing different scenarios in which hosts' cost-benefit relations are determined by the interac-tions among their symbionts. Birds are hosts to numerous symbionts, some of which can be parasitic, i.e decreasing the fitness of their hosts. Failure to detect costs of parasitism might be due to methodological problems or analytical limitations, be-cause detrimental effects may be confounded or masked by other factors (e.g. Clayton et al. 1992), including infections by other parasitic taxa (e.g. Merino and Potti 1995) and changes in host behaviour to minimise the costs of being parasitised (Hart 1997). Alternatively, some symbionts assumed to be parasitic might have negligible effects on hosts. This kind of symbionts could be considered as commensals, facultative parasites or facultative mutualists rather than true parasites depend-ing on host condition, the symbiont under study, and the nature of the usually diverse community of symbi-otic organisms (Bronstein 1994). In fact, commensal-ism, where there are neither costs nor benefits to hosts, and mutualism, where both host and symbiont benefit, are common in nature (Thompson 1994) although among birds and their symbionts next to nothing is known about how common they are.
Feather mites are arthropods that live on or in the feathers on birds, and are among the commonest avian ectosymbionts. However, the nature of the ecological interaction between feather mites and birds remains unclear, some studies reporting negative effects of feather mites on their hosts and others reporting positive or no effects. Here we use a large dataset com-prising 20 189 measurements taken from 83 species of birds collected during 22 yr in 151 localities from seven countries in Europe and North Africa to explore the correlation between feather mite abundance and body condition of their hosts. We predicted that, if wing-dwelling feather mites are parasites, a negative correlation with host body condition should be found, while a mutualistic interaction should yield positive correlation. Although negative relationships between feather mite abundance and host body condition were found in a few species of birds, the sign of the correlation was positive in most bird species (69%). The overall effect size was only slightly positive (r 0.066). The effect of feather mite abundance explained 10% of variance in body condition in most species (87%). Results suggest that feather mites are not parasites of birds, but rather that they hold a commensalistic relationship where feather mites may benefit from feeding on uropygial gland secretions of their hosts and birds do not seem to obtain a great benefit from the presence of feather mites.
DNA barcoding remains a challenge when applied to diet analyses, ancient DNA studies, environmental DNA samples and, more generally, in any cases where DNA samples have not been adequately preserved. Because the size of the commonly used barcoding marker (COI) is over 600 base pairs (bp), amplification fails when the DNA molecule is degraded into smaller fragments. However, relevant information for specimen identification may not be evenly distributed along the barcoding region, and a shorter target can be sufficient for identification purposes. This study proposes a new, widely applicable, method to compare the performance of all potential 'mini-barcodes' for a given molecular marker and to objectively select the shortest and most informative one. Our method is based on a sliding window analysis implemented in the new R package SPIDER (Species IDentity and Evolution in R). This method is applicable to any taxon and any molecular marker. Here, it was tested on earthworm DNA that had been degraded through digestion by carnivorous landsnails. A 100 bp region of 16 S rDNA was selected as the shortest informative fragment (mini-barcode) required for accurate specimen identification. Corresponding primers were designed and used to amplify degraded earthworm (prey) DNA from 46 landsnail (predator) faeces using 454-pyrosequencing. This led to the detection of 18 earthworm species in the diet of the snail. We encourage molecular ecologists to use this method to objectively select the most informative region of the gene they aim to amplify from degraded DNA. The method and tools provided here, can be particularly useful (1) when dealing with degraded DNA for which only small fragments can be amplified, (2) for cases where no consensus has yet been reached on the appropriate barcode gene, or (3) to allow direct analysis of short reads derived from massively parallel sequencing without the need for bioinformatic consolidation.
We propose a general working strategy to deal with incomplete reference libraries in the DNA barcoding identification of species. Considering that (1) queries with a large genetic distance with their best DNA barcode match are more likely to be misidentified and (2) imposing a distance threshold profitably reduces identification errors, we modelled relationships between identification performances and distance thresholds in four DNA barcode libraries of Diptera (n = 4270), Lepidoptera (n = 7577), Hymenoptera (n = 2067) and Tephritidae (n = 602 DNA barcodes). In all cases, more restrictive distance thresholds produced a gradual increase in the proportion of true negatives, a gradual decrease of false positives and more abrupt variations in the proportions of true positives and false negatives. More restrictive distance thresholds improved precision, yet negatively affected accuracy due to the higher proportions of queries discarded (viz. having a distance query-best match above the threshold). Using a simple linear regression we calculated an ad hoc distance threshold for the tephritid library producing an estimated relative identification error <0.05. According to the expectations, when we used this threshold for the identification of 188 independently collected tephritids, less than 5% of queries with a distance query-best match below the threshold were misidentified. Ad hoc thresholds can be calculated for each particular reference library of DNA barcodes and should be used as cut-off mark defining whether we can proceed identifying the query with a known estimated error probability (e.g. 5%) or whether we should discard the query and consider alternative/complementary identification methods.
Since its introduction in 2001, MrBayes has grown in popularity as a software package for Bayesian phylogenetic inference using Markov chain Monte Carlo (MCMC) methods. With this note, we announce the release of version 3.2, a major upgrade to the latest official release presented in 2003. The new version provides convergence diagnostics and allows multiple analyses to be run in parallel with convergence progress monitored on the fly. The introduction of new proposals and automatic optimization of tuning parameters has improved convergence for many problems. The new version also sports significantly faster likelihood calculations through streaming single-instruction-multiple-data extensions (SSE) and support of the BEAGLE library, allowing likelihood calculations to be delegated to graphics processing units (GPUs) on compatible hardware. Speedup factors range from around 2 with SSE code to more than 50 with BEAGLE for codon problems. Checkpointing across all models allows long runs to be completed even when an analysis is prematurely terminated. New models include relaxed clocks, dating, model averaging across time-reversible substitution models, and support for hard, negative, and partial (backbone) tree constraints. Inference of species trees from gene trees is supported by full incorporation of the Bayesian estimation of species trees (BEST) algorithms. Marginal model likelihoods for Bayes factor tests can be estimated accurately across the entire model space using the stepping stone method. The new version provides more output options than previously, including samples of ancestral states, site rates, site d(N)/d(S) rations, branch rates, and node dates. A wide range of statistics on tree parameters can also be output for visualization in FigTree and compatible software.
Background:
Poorly regulated international trade in ornamental fishes poses risks to both biodiversity and economic activity via invasive alien species and exotic pathogens. Border security officials need robust tools to confirm identifications, often requiring hard-to-obtain taxonomic literature and expertise. DNA barcoding offers a potentially attractive tool for quarantine inspection, but has yet to be scrutinised for aquarium fishes. Here, we present a barcoding approach for ornamental cyprinid fishes by: (1) expanding current barcode reference libraries; (2) assessing barcode congruence with morphological identifications under numerous scenarios (e.g. inclusion of GenBank data, presence of singleton species, choice of analytical method); and (3) providing supplementary information to identify difficult species.
Methodology/principal findings:
We sampled 172 ornamental cyprinid fish species from the international trade, and provide data for 91 species currently unrepresented in reference libraries (GenBank/Bold). DNA barcodes were found to be highly congruent with our morphological assignments, achieving success rates of 90-99%, depending on the method used (neighbour-joining monophyly, bootstrap, nearest neighbour, GMYC, percent threshold). Inclusion of data from GenBank (additional 157 spp.) resulted in a more comprehensive library, but at a cost to success rate due to the increased number of singleton species. In addition to DNA barcodes, our study also provides supporting data in the form of specimen images, morphological characters, taxonomic bibliography, preserved vouchers, and nuclear rhodopsin sequences. Using this nuclear rhodopsin data we also uncovered evidence of interspecific hybridisation, and highlighted unrecognised diversity within popular aquarium species, including the endangered Indian barb Puntius denisonii.
Conclusions/significance:
We demonstrate that DNA barcoding provides a highly effective biosecurity tool for rapidly identifying ornamental fishes. In cases where DNA barcodes are unable to offer an identification, we improve on previous studies by consolidating supplementary information from multiple data sources, and empower biosecurity agencies to confidently identify high-risk fishes in the aquarium trade.
Towards lower latitudes the number of recognized species is not only higher, but also phylogeographic subdivision within species is more pronounced. Moreover, new genetically isolated populations are often described in recent phylogenies of Neotropical birds suggesting that the number of species in the region is underestimated. Previous COI barcoding of Argentinean bird species showed more complex patterns of regional divergence in the Neotropical than in the North American avifauna.
Here we analyzed 1,431 samples from 561 different species to extend the Neotropical bird barcode survey to lower latitudes, and detected even higher geographic structure within species than reported previously. About 93% (520) of the species were identified correctly from their DNA barcodes. The remaining 41 species were not monophyletic in their COI sequences because they shared barcode sequences with closely related species (N = 21) or contained very divergent clusters suggestive of putative new species embedded within the gene tree (N = 20). Deep intraspecific divergences overlapping with among-species differences were detected in 48 species, often with samples from large geographic areas and several including multiple subspecies. This strong population genetic structure often coincided with breaks between different ecoregions or areas of endemism.
The taxonomic uncertainty associated with the high incidence of non-monophyletic species and disc