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Confidence Limits on Phylogenies: An Approach Using the Bootstrap

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Abstract

The recently-developed statistical method known as the "bootstrap" can be used to place confidence intervals on phylogenies. It involves resampling points from one's own data, with replacement, to create a series of bootstrap samples of the same size as the original data. Each of these is analyzed, and the variation among the resulting estimates taken to indicate the size of the error involved in making estimates from the original data, In the case of phylogenies, it is argued that the proper method of resampling is to keep all of the original species while sampling characters with replacement, under the assumption that the characters have been independently drawn by the systematist and have evolved independently. Majority-rule consensus trees can be used to construct a phylogeny showing all of the inferred monophyletic groups that occurred in a majority of the bootstrap samples. If a group shows up 95% of the time or more, the evidence for it is taken to be statistically significant. Existing computer programs can be used to analyze different bootstrap samples by using weights on the characters, the weight of a character being how many times it was drawn in bootstrap sampling. When all characters are perfectly compatible, as envisioned by Hennig, bootstrap sampling becomes unnecessary; the bootstrap method would show significant evidence for a group if it is defined by three or more characters.

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... The evolutionary history was inferred using the neighbor-joining method [40]. The bootstrap consensus tree inferred from 500 replicates [41] and the percentage of replicate trees in which the associated taxa clustered together in the bootstrap test (500 replicates) are shown next to the branches [41]. The evolutionary distances were computed using Tamura-Kumar method [42]. ...
... The evolutionary history was inferred using the neighbor-joining method [40]. The bootstrap consensus tree inferred from 500 replicates [41] and the percentage of replicate trees in which the associated taxa clustered together in the bootstrap test (500 replicates) are shown next to the branches [41]. The evolutionary distances were computed using Tamura-Kumar method [42]. ...
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... Dash (-) indicates sequence data for that gene are unavailable for that specimen. Capital letters distinguish different specimens of the same species and are repeated in Figures (Nguyen et al., 2015), using the models suggested for each unlinked partition, the -spp option that allowed each partition to have an individual evolutionary rate, and 1,000 non-parametric bootstrap replicates (MLB; Felsenstein, 1985) on "Hydra," the Smithsonian Institution High Performance Cluster (SI/HPC). Bayesian Inference analyses were performed using MrBayes v3.2.6 on the CIPRES Science Gateway (Miller et al., 2010). ...
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... Phylogenetic trees were constructed using the NJ, ML, and MP methods using the MEGA V.11.0 program [40]. The robustness of the topology of the phylogenetic trees was evaluated through bootstrap analysis with 1000 re-samplings [41], using the 16S rRNA gene of Nocardia aobensis strain IFM 0372T as an outgroup to root the tree [42]. ...
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... The phylogenetic distance matrices were estimated by the Kimura two-parameter model (Kimura, 1980). The topology of the phylogenetic tree was evaluated by using the bootstrap resampling method by Felsenstein (1985) with 1,000 replicates. ...
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... The most parsimonious trees returned by the analysis were summarized in one consensus tree using the strict consensus method. Bootstrap support (BS) was performed using 1,000 pseudoreplicates to assess the support of the clades (Felsenstein 1985). Bayesian phylogenetic analyses were also performed using MrBayes 3.1.23 ...
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... Sequence alignments and phylogenetic analysis were conducted using Molecular Evolutionary Genetics Analysis (MEGA 11) software, with the ClustalW algorithm employed for alignment (Thompson et al. 1994) (Tamura et al. 2021). A phylogenetic tree was constructed using the neighbor-joining (NJ) method with 1000 bootstrap replicates (Felsenstein 1985). Evolutionary distances were computed using the Maximum Composite Likelihood method (Tamura et al. 2004). ...
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... Although a point estimate of a single best tree can be useful, uncertainty quantification is another critical aspect of biological hypothesis testing, yet generally prohibitive in very large modern datasets. Existing solutions include bootstrapping (Felsenstein, 1985;Efron et al., 1996) and Bayesian posterior supports (Rannala and Yang, 1996). For all inferential approaches, bootstrapping is possible and has highly efficient implementations (Hoang et al., 2018). ...
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As whole genomes become widely available, maximum likelihood and Bayesian phylogenetic methods are demonstrating their limits in meeting the escalating computational demands. Conversely, distance-based phylogenetic methods are efficient, but are rarely favoured due to their inferior performance. Here, we extend distance-based phylogenetics using an entropy-based likelihood of the evolution among pairs of taxa, allowing for fast Bayesian inference in genome-scale datasets. We provide evidence of a close link between the inference criteria used in distance methods and Felsenstein's likelihood, such that the methods are expected to have comparable performance in practice. Using the entropic likelihood, we perform Bayesian inference on three phylogenetic benchmark datasets and find that estimates closely correspond with previous inferences. We also apply this rapid inference approach to a 60-million-site alignment from 363 avian taxa, covering most avian families. The method has outstanding performance and reveals substantial uncertainty in the avian diversification events immediately after the K-Pg transition event. The entropic likelihood allows for efficient Bayesian phylogenetic inference, accommodating the analysis demands of the genomic era.
... As a branch support measure, Bremer support was calculated (Bremer 1994), and as a measure of branch stability, a bootstrap resampling analysis (Felsenstein 1985) was conducted, performing 10,000 pseudoreplicates. Bremer support was calculated after searching for suboptimal trees. ...
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... The ML analyses were conducted in IQ-TREE 2 (Minh et al. 2020). We performed four different runs, in order to test the tree's topologies: one with IQ-TREE's default settings, in which the programme determined the best-fit model for our alignment (TIM2+F+I+R4 under BIC), with 100,000 bootstrap replicates for the SH-like approximate likelihood ratio branch test (SH-aLRT; Guindon et al. (2010)) and with 100,000 ultrafast bootstrap replicates (UFBoot; Hoang et al. (2018)) to assess the branch support; a second run with the three partition schemes identified by Partition-Finder and with the same number of SH-aLRT and UFBoot replicates (100,000); a third run with the default settings, but with the -bnni option (optimised, hill-climbing nearest neighbour interchange, NNI, search) in order to reduce the risk of overestimating the branch supports of the UFBoot, with 10,000 replicates for both SH-aLRT and UFBoot; and finally, a fourth run with the default settings, but the branch support evaluated with 500 standard non-parametric bootstrap (Boot; Felsenstein (1985)) searches. The phylograms were visualised and edited with FigTree (Rambaut 2014) and the uncorrected genetic p-distances were calculated for 16S with MEGA6 (Tamura et al. 2013). ...
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We report the rediscovery, a century after its last record, of two specimens of Pristimantis ruidus from approximately 6 km northeast of its type locality, in a montane forest from the western Andes of southern Ecuador. This species, described by John D. Lynch in 1979, based on specimens collected in 1922 by George H. Tate, was previously known only from the type series, all specimens originating from a single locality (Molleturo, Provincia Azuay, Ecuador, 2317 m elev.). The rediscovery occurred in November 2022 in the Reserva de Conservación Quitahuaycu, Molleturo Parish, Azuay Province. The two specimens, both females, are characterised by rough skin with numerous rounded low warts and tubercles on their dorsum, a W-shaped occipital fold, tympanum and tympanic ring not externally visible and by the lack of cranial crests, characters that correspond with the description of the holotype. We evaluate the phylogenetic relationships, based on two mitochondrial genes (12S and 16S rRNA) and one nuclear gene (RAG-1). These analyses reveal Pristimantis ruidus as a member of the subgenus Huicundomantis, being most closely related to P. philipi, a species with a distribution restricted to the paramo of the same mountain region (Macizo del Cajas). Pristimantis ruidus co-exists with Pristimantis jimenezi, three additional species of Pristimantis and one Noblella with uncertain taxonomic status. This rediscovery underscores the urgency of implementing effective conservation and monitoring measures for the few remnants of the original ecosystems from western Ecuador, which face ongoing anthropogenic threats and harbour a diversity that remains insufficiently studied.
... These substitution models were used to analyse phylogenetic relationships with PHYML v.3.0. Topology support was examined by bootstrapping the original dataset 1000 times (heuristic option) [34,35]. Bayesian phylogenetic analyses were conducted using MrBayes 3.2.7 [33]. ...
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The family Rhinonyssidae includes endoparasitic, blood-feeding mites that are parasitic on birds and that remain largely unstudied despite their potential role as vectors or reservoirs for various pathogens, like other Dermanyssoidea. Traditionally, the taxonomy of the group has been based on morphometric characteristics, which makes identification very difficult in many groups of closely related species. On the other hand, studies on the phylogenetic relationships within this group of mites have been neglected until the early years of the present century. In this study, twelve species belonging to five different species complexes were identified, and domains D1-D3 28S rRNA of each one were sequenced, for the first time, to investigate the sequence variation and its taxonomic implications for phylogenetic inference. Our data indicate that this molecular marker can effectively differentiate between species within the “motacillae”, “sairae”, “pari”, and “hirsti” complexes of the genus Ptilonyssus and the “melloi” and “columbae” complexes of the genus Tinaminyssus. Furthermore, the phylogenetic tree that can be derived from the domain D1-D3 28S rRNA sequences presented in this study is congruent with the current taxonomy of the Rhinonyssidae. This research calls for a reassessment of the taxonomic status of some group of species.
... Phylogenetic analysis based on the concatenated sequences of the ITS and D1/D2 domains of the LSU rRNA gene was performed in mega X using the neighbour-joining (NJ) method with Kimura's two-parameter distance correction (Fig. 1). Confidence levels of the clades were derived from 10 000 bootstrap replicates [22]. Wickerhamomyces anomalus CBS 5759 T (KY110078/NR_111210) was used as the outgroup for this analysis. ...
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Hanseniaspora , a genus of yeasts in which many species reproduce sexually, has attracted the attention of researchers because of its prevalence in diverse ecological niches. Building on our extensive collection efforts since 2020, three previously unknown yeast strains from wild Drosophila species trapped in ripe bananas in Okinawa, Japan, were isolated. Using a multifaceted approach, including physiological assessments and sequence analysis of the D1/D2 domain of the 26S LSU rRNA gene and the internal transcribed spacer (ITS) region, it was revealed that these strains are novel members of the genus Hanseniaspora . The three strains, JCM 36741 T , JCM 36742 and JCM 36748, had identical sequences in their respective D1/D2 and ITS regions, justifying their classification as a single species. Moreover, the new species exhibited a remarkable degree of sequence divergence from its closest relatives, differing by 7 nucleotide substitutions (1.28%) in the D1/D2 domain, 29 nucleotide substitutions and 4 gaps (4.08%) in the ITS regions. These substantial sequence differences highlight the distinctiveness of this novel species in the genus Hanseniaspora . Further analysis revealed physiological characteristics that distinguished the new species from its closest relative, Hanseniaspora hatyaiensis (nom. inval.). These findings culminated in the proposed name Hanseniaspora drosophilae sp. nov., which recognizes the unique ecological niche within the Drosophila microbiota. By uncovering this novel species, this study not only adds to the growing body of knowledge on yeast diversity but also sheds light on the intricate ecological relationships that shape microbial communities. The implications of this discovery extend beyond taxonomic boundaries, inviting further exploration of the evolutionary dynamics and ecological significance of yeast–fly interactions. We propose accommodating H. drosophilae sp. nov. in the genus Hanseniaspora with JCM 36741 T as the holotype. The MycoBank accession number is MB 853822.
... The tree is drawn to scale, with branch lengths in the same units as those of the evolutionary distances used to infer the phylogenetic tree. The evolutionary distances were computed using the Maximum Composite Likelihood method [18]. This analysis involved 15 nucleotide sequences of tested M. phaseolina and 9 six nucleotide sequences of M. phaseolina of sesame from different parts of country, 4 nucelotide sequences of M. phaseolina causing disease in other host plant and Alternaria sesami from portugal used as an out source group for constructing phylogenetic tree. ...
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... The sequences underwent alignment by the ClustalW algorithm with default settings and were also manually trimmed. Afterward, we used the Models function to find a suitable substitution model for the Neighbor-Joining Tree method [6,53]. The same procedure and method were used to construct the phylogenetic tree in Figure 1 [54], in which the accession numbers are also displayed. ...
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Acetic acid bacteria are single-celled organisms well-known for their ability to convert ethanol into acetic acid. Still, recent research suggests they may harbor another attractive characteristic—the production of proteins with remarkable similarities to sirtuins. Sirtuins have been linked to lifespan extension in various organisms, raising intriguing questions about the potential connection between acetic acid bacteria and the biology of aging. This article delves into the characterization of sirtuin homologs in acetic acid bacteria. Up to three types of sirtuin homologs have been identified in 21% of acetic acid bacteria genomes deposited in NCBI. All three types were present only in the genera Acetobacter and Novacetimonas, which are known to survive in the harshest environmental conditions (high concentrations of acetic acid and ethanol). Interestingly, two types of these sirtuin homologs (SirAAB-L and SirAAB-S) constitute a separate group (SirAAB), distinctive from all other presently known sirtuins. The results obtained in silico thus encourage further studies into the function of these types of sirtuins and their interplay with metabolic pathways in these industrially important bacteria.
... Phylogenetic trees were generated using the neighbor-joining DNA distance method in MEGA 7 [38]. Tree topologies were assessed using bootstrap analysis with 1000 resampled datasets [39]. The partial 16S rRNA gene sequences of the selected eight Bacillus strains have been deposited in NCBI GenBank with assigned accession numbers. ...
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Oral bacterial pathogens, including Bacillus species, form biofilms that enhance antibiotic resistance, promote bacterial adherence, and maintain structural integrity. The ability of bacteria to form biofilms is directly linked to several oral diseases, including gingivitis, dental caries, periodontitis, periapical periodontitis, and peri-implantitis. These biofilms act as a predisposing factor for such infections. Nanoparticles, known for their strong antibacterial properties, can target specific biofilm-forming microorganisms without disturbing the normal microflora of the oral cavity. This study focuses on the biofilm-forming ability and clindamycin (CM) resistance of Bacillus species found in the oral cavity. It aims to evaluate the antibacterial and antibiofilm properties of zinc oxide nanoparticles (ZnO-NPs) against oral Bacillus species and assess the effectiveness of combining CM with ZnO-NPs in reducing antibiotic resistance. The antibacterial susceptibility of Bacillus isolates was tested using ZnO-NPs and CM, demonstrating synergistic effects that reduced the minimum inhibitory concentrations by up to 8-fold. The fractional inhibitory concentration (FIC) index indicated a significant synergistic effect in most strains, with FIC values ranging from 0.375 to 0.5. It was found that the majority of Bacillus strains exhibited significant biofilm-forming capabilities, which were reduced when treated with the ZnO-NPs and CM combination. The study also evaluated the cytotoxicity of ZnO-NPs on cancer cells (CAL27) and normal fibroblasts (HFB4). CAL27 cells showed stronger cytotoxicity, with an IC50 of 52.15 µg/mL, compared to HFB4 cells, which had an IC50 of 36.3 µg/mL. Genetic analysis revealed the presence of biofilm-associated genes such as sipW and tasA, along with antibiotic resistance genes (ermC), which correlated with the observed biofilm phenotypes. Overall, this study demonstrates the potential of combining ZnO-NPs with CM to overcome antibiotic resistance and biofilm formation in the oral bacterial pathogens, Bacillus species. These findings suggest new approaches for developing more effective dental treatments targeting oral biofilm-associated infections and antibiotic resistance.
... Neighbour-joining [7], maximum-likelihood [8] and maximumparsimony [9] algorithms were applied to reconstruct the phylogenetic trees using Molecular Evolutionary Genetics Analysis (mega) software package version X [10]. A bootstrap method with 1000 repetitions was applied to evaluate the topology stability of the phylogenetic tree [11], and Kimura's two-parameter model was applied to calculate the evolutionary distance [12]. The result of EzBioCloud analysis based on the almost complete 16S rRNA gene sequence (1496 bp) indicated that strain MR15.9 T belonged to the genus Tersicoccus, and the closest relatives were T. solisilvae CGMCC 1.15480 T (98.9% sequence similarity), followed by T. phoenicis 1P05MA T (98.6% sequence similarity). ...
Article
A Gram-stain-positive, aerobic and non-motile actinobacterium, designated strain MR15.9 T , was isolated from sediment collected from a mangrove ecosystem in Semarang city, Indonesia. Strain MR15.9 T grew at 4–37 °C (optimum 30 °C), pH 6.0–11.0 (optimum 7.0–8.0), with 0–10% (w/v) NaCl (optimum 0–3%). The genome of strain MR15.9 T was 3.67 Mbp with 71.7 mol% G+C content. Phylogenetic analysis based on 16S rRNA gene sequence and genome sequence directed that strain MR15.9 T formed a well-supported clade with Tersicoccus solisilvae CGMCC 1.15480 T and Tersicoccus phoenicis 1P05MA T and shared the highest similarity to T. solisilvae CGMCC 1.15480 T (98.9% sequence similarity) and T. phoenicis 1P05MA T (98.6% sequence similarity). However, the comparative genome analysis between strain MR15.9 T and T. solisilvae CGMCC 1.15480 T gave average nt identity value of 85.2% and digital DNA–DNA hybridization value of 29.0%, which were below the statistical threshold for the delineation of the species. Chemotaxonomic data showed that the major fatty acids were anteiso-C 15:0 , anteiso-C 17:0 and iso-C 16:0 . The polar lipids consisted of diphosphatidylglycerol, phosphatidylglycerol, phosphatidylinositol, two unidentified glycolipids and two unidentified phospholipids. MK-8(H 2 ) and MK-9(H 2 ) were noted as the predominant respiratory quinones in this strain. The results of polyphasic characterization indicated that strain MR15.9 T represents a novel species of the genus Tersicoccus . The name Tersicoccus mangrovi sp. nov. is proposed, with the type strain MR15.9 T (=MCCC 1K08875 T =KCTC 59105 T ).
... Maxtrees were set to 5,000, branches of zero length were collapsed and all parsimonious trees were saved. Clade robustness was assessed using bootstrap (BT) analysis with 1,000 pseudo-replicates (Felsenstein 1985). Descriptive tree statistics -tree length (TL), composite consistency index (CI), composite retention index (RI), composite rescaled consistency index (RC) and composite homoplasy index (HI) were calculated for each maximum parsimonious tree generated. ...
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In the present study, four new species, Geastrum artocarpicola, G. fibulatum, G. sinense and G. trachelium collected from China, are proposed based on a combination of morphological characteristics and molecular evidence. Geastrum artocarpicola is characterized by shallowly saccate to deep saccate exoperidium, bubble-shaped to flask shaped basidia, spherical basidiospores. G. fibulatum is characterized by shallowly saccate to deep saccate exoperidium, spherical basidiospores, generative hyphae with clamp connections in the mycelium layer. G. sinense has arched exoperidium, long stipe expanded basidiomata, and spherical basidiospores. G. trachelium has deep saccate exoperidium, flask-shaped basidia, and spherical basidiospores. Sequences of the internal transcribed spacers (ITS), large subunit (nrLSU), the largest subunit of ribosomal polymerase II (RPB1), and subunit 6 of ATP synthase (ATP6) of the nuclear ribosomal DNA (rDNA) markers of the studied samples were generated, and the phylogenetic analyses were performed with maximum likelihood, maximum parsimony and Bayesian inference methods. The results showed that our collection clustered within Geastrum but distinctly from the others. Full morphological descriptions, illustrations, and phylogenetic analyses results for the four new species are provided. In addition, G. sanglinense is treated as a synonym of beijingense.
... A phylogenetic rescontruction based on Maximum Likelihood method was also performed in MegaX software, with substitution models and other parameters determined by jModeltest software as described previously. Branch supports were obtained by bootstrap method (Felsenstein 1985) with 100 replicates. ...
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The species-rich genus Inga (Leguminosae) presents ca. 300 species widespread throughout the Neotropics and is recognized by its recent and rapid diversification. Forty-eight species of Inga are endemic of the Brazilian Atlantic Forest. Among them, Inga subnuda, with two subspecies: subsp. subnuda, which occurs from the state of Paraiba to Rio de Janeiro; and subsp. luschnathiana, which occurs from Espirito Santo to the state of Santa Catarina. Both subspecies occur in sympatry in the southeastern region and share floral and leaf characters, which hampers the morphological delimitation. The co-occurrence of Inga vera subsp. affinis. with both I. subnuda subspecies results in intermediate morphologies between I. vera subsp. affinis and I. subnuda subsp. luschnathiana, making the distinction between the latter species even harder. We sampled 94 individuals from 8 natural populations and evaluated morphological characters previously described as distinctive among the subspecies of I. subnuda in addition to others not measured yet. We used 84 plastid (trnD-trnT spacer) and 58 nuclear (ITS 1 and 2) sequences to characterize the phylogenetic relationships between the taxa. The results obtained point out I. subnuda subsp. subnuda as a more structured taxon in relation to the other subspecies, whilst I. subnuda subsp. luschnathiana and I. vera subsp. affinis constituted a cohesive group. The apportionment of haplotypes differed between the markers used, thus evincing a retention of ancestral polymorphism between the subspecies, given their recent diversification. This paper explores different lines of evidence, thus contributing to the delimitation of these species. Demographic and biogeographic scenarios are also discussed. A new status of for the taxon currently circumscribed as Inga subnuda subsp. luschnathiana is suggested.
... Evolutionary analyses were conducted in MEGA X [60]. The bootstrap consensus tree was inferred from 500 replicates [65]. Branches corresponding to partitions reproduced in less than 50% bootstrap replicates are collapsed. ...
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Background Malaria parasites establish new permeation pathways (NPPs) at the red blood cell membrane to facilitate the transport of essential nutrients from the blood plasma into the infected host cell. The NPPs are critical to parasite survival and, therefore, in the pursuit of novel therapeutics are an attractive drug target. The NPPs of the human parasite, P. falciparum, have been linked to the RhopH complex, with the monoallelic paralogues clag3.1 and clag3.2 encoding the protein RhopH1/CLAG3 that likely forms the NPP channel-forming component. Yet curiously, the combined knockout of both clag3 genes does not completely eliminate NPP function. The essentiality of the clag3 genes is, however, complicated by three additional clag paralogs ( clag2, clag8 and clag9 ) in P. falciparum that could also be contributing to NPP formation. Methods Here, the rodent malaria species, P. berghei , was utilised to investigate clag essentiality since it contains only two clag genes, clagX and clag9. Allelic replacement of the regions encompassing the functional components of P. berghei clagX with either P. berghei clag9 or P. falciparum clag3.1 examined the relationship between the two P. berghei clag genes as well as functional orthology across the two species. An inducible P. berghei clagX knockout was created to examine the essentiality of the clag3 ortholog to both survival and NPP functionality. Results It was revealed P. berghei CLAGX and CLAG9 , which belong to two distinct phylogenetic clades, have separate non-complementary functions, and that clagX is the functional orthologue of P. falciparum clag3. The inducible clagX knockout in conjunction with a guanidinium chloride induced-haemolysis assay to assess NPP function provided the first evidence of CLAG essentiality to Plasmodium survival and NPP function in an in vivo model of infection. Conclusions This work provides valuable insight regarding the essentiality of the RhopH1 clag genes to the NPPs functionality and validates the continued investigation of the RhopH complex as a therapeutic target to treat malaria infections.
... The genetic relationships between the 20 bread wheat genotypes were also estimated by using the Ward's hierarchical clustering method based on a dissimilarity matrix using DARWIN software v.6.0.010 (http://darwin.cirad.fr), with bootstrapping of 1000 replicates to determine the support for each node [37]. A clustering algorithm on the Bayesian model STRUCTURE software 2.3.4 was used to obtain an explicit picture of genetic composition [38]. ...
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Bread wheat; Genetic diversity; Heterozygosity, SSR; Algeria Bread wheat (Triticum aestivum L.) genotypes was the most important cereal crop in Algeria. Twenty Algerian and introduce genotypes used in this study with a set of 18 wheat microsatellites molecular markers (Simple Sequence Repeat-SSR). This study is to use SSR marker to examine their utility in detecting DNA polymorphism and assessing genetic diversity. A total of 62 alleles were detected at 13 loci using 13 microsatellites primer pairs. The total numbers of alleles per locus (Na) ranged from 4 to 7 with a average of 4,7 alleles. The mean values of the number of effective alleles (Ne), and Shannon's information index (I) were estimated at 3,425 and 1,31 respectively. The observed heterozygosity (Ho) values scored 1.00 at all loci, and the Polymorphic information content (PIC) values ranged from 0,60 at the locus WMC19 to 1,00 for the primer WMC25 averaging at 0.72. The mean observed heterozygosity (Ho) was slightly higher than the mean expected heterozygosity (He), determining a negative fixation index (F) at all loci, (average F=-0.45). Both groups of local bread wheat populations (Algerian) and those introduced from CYMMIT have almost the same number of alleles, (Na) 55,0 and 54,0 respectively. While the mean observed heterozygosity (Ho) was similar in the two groups, (He) was higher in Algerian genotypes, resulting in a negative fixation index (F) for both these two groups. The phylogenetic tree divided the 20 genotypes into two clusters. AMOVA revealed diversity within populations at 99% and between populations at 1%. The current populations were found to be highly structured. STRUCTURE analysis showed three groups at K =3. Kara, et.al, 2024 AMA, Agricultural Mechanization in Asia, Africa and Latin America 19204 The first Group has the largest number of genotypes. The results proved the microsatellite markers SSR utility in detecting polymorphism due to the discrimination of various genotypes and estimating genetic diversity. This work is licensed under a Creative Commons Attribution Non-Commercial 4.0 International License.
... ML inference was performed using the PhyML program (Guindon and Gascuel 2003), available on the public web server Phylemon2 (http://phylemon.bioinfo.cipf.es). Statistical support for the resulting phylogenies was assessed by bootstrapping with 1,000 replicates (Felsenstein 1985). All trees were edited with FigTree software. ...
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Populations of invasive species can have a negative impact on the natural environment and socioeconomic effects of artificial systems. The relationship of Corbicula fluminea with humans, together with the its great adaptive-reproductive capacity, make it one of the most expansive species of invasive bivalves worldwide. Here we report on the dispersion of C. fluminea, and based on genetic analysis we describe the presence and potentially negative effect of this species for the first time in an anthropic environment, an aquaculture facility in Ecuador.
... Using Darwin software, version 6.0.21 [37] (http://darwin.cirad.fr, accessed on 26 April 2019), the genotypes of centenary olive trees, together with those of the commercial olive varieties and the Spanish, Italian, and Greek olive germplasm, were hierarchically classified by applying the neighbor-joining (NJ) method based on a dissimilarity matrix, with bootstrapping of 1000 replicates to determine the support for each node [38]. ...
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The centennial olive trees of Tunisia serve as enduring symbols of resilience, having withstood the test of time while witnessing the effects of climate change, rising temperatures, water scarcity, and the emergence of new diseases. Presently, there is a notable lack of research on the genomic analysis of ancient trees. This study investigates the genetic diversity of twenty-eight ancient olive specimens collected from archeological sites in nine governorates from the north to the south of Tunisia. Using nine highly polymorphic microsatellite markers, these ancient olive trees were compared with twenty-five local Tunisian cultivars and sixty olive varieties from other Mediterranean countries (Greece, Italy, and Spain). The ancient olive trees were revealed to have a high genetic diversity, with 67 alleles and a Shannon index of 1.68. The key findings identify the ancient trees M25, M1, M28, and M24 as synonyms for local olive cultivars, while “M10” is noted as a first-generation migrant from Tunisian olives. Cluster analysis methods, including structure, neighbor-joining (NJ), and principal coordinates (PCoA), show that these ancient trees share a common genetic background and ancestry with varieties from Tunisia, Italy, Spain, and Greece. The conservation and evaluation of these genotypes will increase the genetic diversity available for breeding programs and strengthen the resilience of agriculture, which is currently facing unprecedented pressure worldwide.
... The Neighbor-Joining approach in MEGA 6.0 was used to infer the evolutionary history (Saitou & Nei, 1987). The evolutionary history of the taxa under study was assumed to be represented by the bootstrap consensus tree that was estimated from 500 replicates (Felsenstein, 1985). The Jukes-Cantor method (Jukes & Cantor, 1969) was utilised to calculate the evolutionary distances. ...
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Using accepted methods, the physicochemical characteristics and microalgae composition of the water and sediment of Otamiri river were assessed. The results showed that the electrical conductivity (EC), turbidity, and biological oxygen demand (BOD) levels were higher than those allowed by the World Health Organisation (WHO). As expected, the dissolved oxygen (DO) level of 8.33mgL-1 was below the WHO standard of 10mgL-1. Out of all the heavy metals discovered in the river water, iron had the highest quantity (0.542mgL-1). Similarly, concentrations of arsenic, mercury, and lead were found to exceed the WHO drinking water quality criteria. Nevertheless, there was no nickel, copper, or cadmium in the river water. The lowest and greatest quantities of arsenic (0.004 mg kg-1) and iron (1.789 mg kg-1) were found in the sediment, respectively. Benzethonium chloride was the most prevalent cationic surfactant in both the river water (3.9634mgL-1) and the sediment (1.5631mgkg-1). Diethyl heptadecyl imidazolinium chloride and benzalkonium chloride exhibited the lowest concentrations in water and sediment, measuring 0.5408mgL-1 and 0.9758mgkg-1 , respectively. The amounts of cationic surfactants detected in the river water were all higher than the permissible limit of 0.5mgL-1 established by the Environmental Protection Agency (EPA) in 1979. The most concentrated anionic surfactant in sediment (5.6551mgkg-1) and water (2.0938mgL-1) was discovered to be sodium decylsulfate. Sodium tetradecylsulfate had the lowest amount in the water (0.0521mgL-1), however sodium octadecylsulfate was absent from the sediment. Both sodium decylsulfate and sodium octadecylsulfate levels were higher than those advised by the EPA. The concentrations of certain physicochemical parameters in soil samples and river water point to possible human-caused pollution of the aquatic environment. Aquatic creature disturbance, alterations in water chemistry, and bioaccumulation in the food chain are just a few of the ecological effects that these contaminants may have. The microalgae isolates were identified as Micractinium pusillum, Chlorella sorokiniana, and Dictyosphaeriun ehrenbagianum by BLASTn's partial 18S rRNA gene sequence similarity analysis.
... The CLUSTAL X program was used to align the sequences. Using the neighbor-joining approach (Saitou & Nei 1987), phylogenetic trees were built using evolutionary distance data derived from Kimura's two-parameter model, and 1,000 bootstrap analyses were carried out (Felsenstein 1985). ...
Article
Yeasts are potential biocontrol agents for Aspergillus flavus, an aflatoxin-producing fungus that is present in various agricultural products, including nutmeg. This study aimed to obtain yeast isolates from nutmeg (seeds, pulps, and leaves), characterise them, and identify their antagonistic effects on A. flavus. The antagonistic activities toward A. flavus were determined by a dual-culture method. Moreover, the possible mechanism responsible for these antagonistic effects was also analysed. The results showed that 51 yeast isolates were successfully isolated from nutmeg. The inhibition percentages of 47.25 ± 1.66% (isolate DP 1341a) and 55.98 ± 1.31% (isolate DP 1342) were statistically significant (p < 0.05). The antagonistic mechanisms of the DP 1341a isolate were associated with the production of volatile organic compounds (32.79 ± 1.01%), a chitinolytic index (2.51 ± 0.55), and hyperparasitism but not toxin activity. Moreover, the DP 1342 isolate produced volatile organic compounds (54.33 ± 3.13%), exhibited toxin activity (2.74 ± 0.22) and exhibited hyperparasitism but did not exhibit chitinase activity. Molecular identification showed that the two yeast isolates (DP 1341a and DP 1342) were identified as Pseudozyma hubeiensiswith sequence similarity > 99%. Therefore, the selected yeast isolates, P. hubeiensis DP 1341a and DP 1342, could be further developed as biological control agents for A. flavus. This finding will also be useful for improving biological control agents as an eco-friendly and economically viable disease management strategy.
... Nucleotide sequences were assembled and edited with MEGAX (Kumar et al. 2018), and the best-fit model of amino acid substitutions for maximum-likelihood (ML) and the general reversible mitochondrial model (Adachi & Hasegawa 1996) was selected to fit the datasets best using the frequency response model. Phylogenetic analysis was conducted by the ML method and bootstrap analysis (Felsenstein 1985) with 1000 iterations in MEGAX. Thor, 1932 in Cyprus. ...
Article
The slug mite Riccardoella oudemansi Thor, 1932, is reported for the first time from Cyprus. Two terrestrial mollusc species: Limacus flavus (Linnaeus, 1758) (Limacidae) and, tentatively, Carpathica cf. cretica (Forcart, 1950) (Oxychilidae) are reported as hosts. The mites were identified on the basis of palps and leg chaetotaxy, and morphological characters including the shape of famulus on tibia I. Results of an initial molecular analysis are presented. Other Riccardoella species may be present in Cyprus.
... [35]. For the ML analysis, the GTRGAMMA model was employed for all datasets [36], and bootstrap values were calculated from 1000 bootstrap replicates through heuristic searches [37]. For the MP analysis, a heuristic search with 1000 random addition sequence replicates and tree-bisection-reconnection (TBR) branch swapping was performed. ...
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The Coelogyne s.l. is one of the emblematic genera of the Asian orchids, with high horticultural and medicinal values. However, the phylogenetic relationships of the genus inferred from previous studies based on a limited number of DNA markers remain ambiguous. In this study, we newly sequenced and assembled the complete plastomes of seven Coelogyne species: C. bulleyia, C. fimbriata, C. flaccida, C. prolifera, C. tricallosa, C. uncata, and an unknown taxa, Coelogyne sp. The plastomes of Coelogyne exhibited a typical quadripartite structure, varying in length between 157,476 bp and 160,096 bp, accompanied by a GC content spanning from 37.3% to 37.5%. A total of 132 genes were annotated for each plastome, including 86 protein-coding genes, eight rRNA genes, and 38 tRNA genes. Among these, 19 genes underwent duplication within the inverted repeat (IR) regions, and 18 genes exhibited the presence of introns. Additionally, we detected 54 to 69 simple sequence repeats (SSRs) and 30 to 49 long repeats. In terms of codon usage frequency, leucine (Leu) emerged with the highest frequency, while cysteine (Cys) exhibited the lowest occurrence. Furthermore, eight hypervariable regions (atpB-rbcL, psbK-psbI, rps8-rpl14, rps16-trnQUUG, psaC-ndhE, ndhF-rpl32, psbB-psbT, and ycf1) were identified. Phylogenetic analyses using complete plastomes and protein-coding genes indicated that Coelogyne s.l. was monophyletic. Moreover, the results robustly supported the division of Coelogyne s.l. into five clades. This study provides a comprehensive analysis of the structural variation and phylogenetic analysis of the Coelogyne s.l. based on plastome data. The findings offer significant insights into the plastid genomic characteristics and the phylogenetic relationships of Coelogyne s.l., contributing to a deeper understanding of its evolutionary history.
... The phylogenetic analysis was carried out using the MEGA (v.11) [20] software with the maximum-likelihood [21], neighbor-joining [22], and maximum-parsimony [23] algorithms. The phylogenetic trees were evaluated by bootstrap analysis with 1000 replications [24]. ...
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A novel Gram-negative, orange-colored, rod-shaped, oxidase and catalase-positive, non-spore-forming bacterium, designated as zp-37T, was isolated from the rhizosphere soil of Phragmites australis (Cav.) Trin. ex Steud in Kashgar County, Xinjiang, China. The phylogenetic analysis, based on the 16S rRNA genes, revealed that strain zp-37T belongs to the genus Halomonas. Growth of strain zp-37T was observed at 10–43 °C, pH 6.0–11.0, and 0–20% NaCl (w/v). The principal fatty acids of strain zp-37T were summed feature 8 (C18:1ω7c and/or C18:1ω6c, 55.67%) and summed feature 3 (C16:1ω7c and/or C16:1ω6c, 20.16%). The polar lipid profile contained diphosphatidylglycerol (DPG), phosphatidylglycerol (PG), phosphatidylethanolamine (PE), unidentified phospholipids (UPL 1–3), unidentified aminophospholipids (UAPL 1–2), and unidentified lipid (UL). Its main respiratory quinone was ubiquinone Q-9 (100%). The genome of strain zp-37T was 3,489,967 bp in size, containing two plasmids with lengths of 18,112 bp and 4364 bp, respectively. The genomic DNA G+C content of strain zp-37T was 59.3%. By the genome annotation, various genes related to the function of saline-alkaline stress tolerance and plant growth promotion were predicted. The average nucleotide identity (ANI) and digital DNA–DNA hybridization (dDDH) values between strain zp-37T and its five closely related strains were 72.64–75.59% and 19.70–20.40%, respectively, which were lower than the threshold for species delineation (ANI: 95–96%, dDDH: 70%). Based on the phylogenetic, phenotypic, and chemotaxonomic analyses and genomic comparisons, strain zp-37T was suggested to represent a novel species within the genus Halomonas, for which the name Halomonas kashgarensis sp. nov. is proposed. The strain type was designated zp-37T (=CGMCC 1.62213T = JCM 37305T).
... Convergence was indicated by an average standard deviation of split frequencies of 0.009385, potential scale reduction factors ranging from 1.000 to 1.004 for all parameters, and effective sample sizes exceeding 113 for all parameters. Nodal support within the ML tree was evaluated using bootstrap analysis (BS) with 1000 pseudoreplicates (Felsenstein 1985). ...
Article
The superfamily Pseudocerotoidea Lang encompasses Euryleptidae Stimpson, Pseudocerotidae Lang, and Stylostomidae Dittman. These families include conspicuous, flamboyant marine flatworms with a worldwide distribution. Here, we describe a new euryleptid species belonging to Prostheceraeus Schmarda, 1859, based on specimens collected by dredging at depths of 100–180 m off Misaki, Japan. The new species, Prostheceraeus fragum sp. n., is distinguished by its unique dorsal colouration in that oval red maculae are radially distributed along the translucent cream body periphery. Additionally, based on newly collected topotypic specimens, we transfer the poorly known pseudocerotid species Pseudoceros sagamianus Kato, 1937, to the genus Phrikoceros Newman and Cannon, 1996. These specimens possess (1) a pharynx with shallow, simple folds, (2) a deeply ruffled body margin, and (3) ear-like pseudotentacles with folds. Our molecular phylogeny, based on partial 18S and 28S rDNA sequences, yielded two key findings: (1) Pr. fragum sp. n. nested within the clade containing all other Prostheceraeus species, and (2) Phrikoceros sagamianus comb. n. formed a clade with all other Phrikoceros and Monobiceros species, distinct from the Pseudoceros clade. These results support the generic affiliations established through our morphological examinations. Furthermore, based on the resulting phylogenetic trees, we discuss the systematics of Euryleptidae in relation to Stylostomidae.
... Phylogenetic analysis based on single ITS or D1/D2 sequences was executed utilizing the Neighbor-Joining model in MEGA v.7 with evolutionary distances derived from Kimura's two-parameter model (Kimura 1980;Kumar et al. 2016;Lachance 2022). Bootstrap analyses were performed on 1,000 random re-sampling (Felsenstein 1985). Maximum parsimony (MP), Bayesian Inference (BI), and maximum likelihood (ML) analyses were conducted on the combined ITS and D1/D2 sequences using PAUP v.4.0b10 (Swofford 2003), MRBAYES v.3.2 with 1,000,000 generations (Ronquist et al. 2012), and RAX-ML-HPC 7.2.8 with 1,000 bootstrap replicates (Stamatakis 2006), respectively. ...
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A study on yeast species from the genera Geotrichum and Magnusiomyces in southwest and central China was conducted based on morphological and molecular phylogenetic analyses using the ITS region and the D1/D2 domain of the LSU rRNA gene. The research identified two new yeast species: Geotrichum hubeiense and Magnusiomyces pitmudophilus. The study contributed to understanding arthroconidial yeast diversity in fermentation and natural environments and paved the way for future taxonomic and ecological studies. Descriptions, illustrations, and phylogenetic analysis results of the two new taxa are provided.
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Kodamaea ohmeri is an emerging and opportunistic yeast associated with a high mortality rate in humans. As it is commonly found in the environment, it is possible that environmental conditions and agricultural practices contribute to the adaptation of this yeast and the selection of antifungal resistance. During a multicentric study in Brazil, conducted under a One Health perspective, 14 isolates of K. ohmeri were identified from different sources: three from blood cultures, three from animals (swine and poultry), and eight from animal environments (swine and poultry). Yeasts were isolated using CHROmagar® Candida medium and identified by MALDI-TOF MS and ITS rDNA barcoding. Minimum inhibitory concentration (MIC) was determined using the broth microdilution method for clinical (azoles, echinocandins, pyrimidine analogs, and polyenes), and environmental antifungals (tebuconazole, pyraclostrobin, carbendazim, and mancozeb), and hospital disinfectants (quaternary ammonium compounds). Of note, color variations of K. ohmeri were noted on CHROmagar® depending on the incubation time, which is likely to complicate its identification. Following polyphasic identification and taxonomic confirmation, all isolates demonstrated low MIC values for clinical antifungals, disinfectants, and tebuconazole. However, all isolates were able to grow in the presence of carbendazim, mancozeb, and pyraclostrobin. Together, these findings highlight the risks associated with the use of environmental azoles, such as tebuconazole, as they may impact non-target fungi of medical importance, but other fungicides do not present the same risk. This is the first study to demonstrate that K. ohmeri, an important emerging yeast in human medicine, can be isolated from various sources, including patients. Although the isolates exhibited low MIC values for clinical antifungals, it is crucial to monitor changes in sensitivity patterns over time in emerging microorganisms to prevent the development of multidrug resistance, which may originate in the environment.
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This research targets straw return in Farm 852’s albic soil, China. The soil is nutrient-poor with few microbes and slow straw decomposition. Through fixed-point sampling and bacterial screening, an actinomycete consortium consisting of four strains was assembled, and two of them were identified as new actinomycetes. After 7 days of fermentation, the lignocellulose degradation rates of this consortium outstripped those of single strains, with cellulose degraded at 69.07%, hemicellulose at 64.98%, and lignin at 68.95%. FTIR, XRD, and SEM verified the damage inflicted on the straw structure. Lab simulations found group D (with the consortium) had a higher straw weight loss rate than group C (with commercialized microbial agents) and controls. The compound actinomycetes stepped up the bacterial abundance with the passage of time. In contrast, their effect on fungal abundance was hardly noticeable, but they had markedly ameliorated the soil fertility. These findings prove that the microbial consortium effectively accelerates straw decomposition and boosts soil microbe abundance and fertility in albic soil. It shows great potential for straw return and provides a microbial solution for this field.
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Okra (Abelmoschus esculentus L.) is a significant vegetable crop in India, but its cultivation is severely impacted by viral diseases, particularly okra yellow vein mosaic disease (OYVMD).This study was conducted to evaluate the effectiveness of various pest management modules against whiteflies (Bemisia tabaci), the primary vector of OYVMD, and to assess the incidence and severity of OYVMD in okra. The experiment was carried out at the Central Research Station of Odisha University of Agriculture and Technology during the kharif seasons of 2019 and 2020, using a Randomized Complete Block Design (RCBD) with five replications. Four treatments were evaluated: an Integrated Module, a Biointensive Module, a Chemical Module, and an Untreated Control. The results demonstrated that both the Integrated and Chemical modules significantly reduced the whitefly population and managed it effectively throughout the cropping period, with mean populations of 2.19 whiteflies per leaf. These modules also effectively suppressed the incidence and severity of OYVMD with the highest incidence being recorded in bio-intensive module. Molecular characterization of both yellow vein mosaic virus and whiteflies collected from virus infected plants were also done by conducting PCR followed by gel electrophoresis of isolated DNA samples which revealed that the whiteflies from Bhubaneswar grouped phylogenetically with the whitefly isolates from Indonesia, Thailand and China. In case of viruses, okra enation leaf curl virus from Bhubaneswar clustered with Mungbean yellow mosaic virus (MYMV) from Raichur and Hyderabad.
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Fungal keratitis, which is caused primarily by Neocosmospora and Fusarium species, is a significant global health issue that affects more than a million people annually in tropical and subtropical regions. Neocosmospora solani (formerly Fusarium solani) is a leading cause of corneal infections, along with members of the Fusarium oxysporum species complex (FOSC). This study provides new insights by reporting a series of ocular fusariosis cases caused by FOSC members and presenting molecular evidence linking specific haplotypes within FOSC to human infections. We describe three cases of Fusarium keratitis selected from a comprehensive review of clinicopathological data in our institution's archives. These cases were chosen for their distinctive clinical presentations and the involvement of less common Fusarium species. Two of these patients were diagnosed with keratitis and anterior endophthalmitis, and the third patient had a corneal ulcer previously treated with topical antivirals and antibiotics. All patients were successfully treated with topical amphotericin B. The Fusarium isolates from these patients were subjected to detailed molecular characterization, including DNA sequencing (tef1α, rpb2, CaM, tub2, and LSU), amplified fragment length polymorphism (AFLP) marker analysis, and MALDI-TOF MS analysis. Remarkably, our study reports the first case of human infection by F. veterinarium, alongside cases involving F. contaminatum and F. curvatum. Furthermore, a molecular survey using haplotypic networks based on tef1α sequences identified genotypes associated with human infections and revealed the emergence of F. veterinarium clade VII. Our findings emphasize the need for vigilance regarding emerging species within the FOSC, particularly F. veterinarium. This highlights the need for improved diagnostic tools and targeted research to combat fusarioid-related infections effectively.
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Arsenic-resistant Klebsiella oxytoca strain AT-02 was isolated from the ground water of the Multan region of Pakistan. The strain displayed high arsenite and arsenate resistance as minimal inhibitory concentration (MIC) was 600ppm and 10,000ppm respectively. The high tolerance of the isolated strain towards arsenate can be postulated due to significant increase in biofilm in response to arsenate. The bacterial strain exposed to 1/2 and 3/4 MIC showed a significant 10 and 12 folds increase in expression of the arsenite efflux gene arsB. Sequential and structural comparison of the arsB gene showed the presence of conserved arsenic binding residues. Arsenic remediation by AT-02 biomass was 50% after 0.5 hours of incubation and 66% in 2 hours. the increase in remediation efficiency with the increase in incubation time indicates its biosorption potential. the arsenic sensitive strain NK11 showed only 4–5% arsenic remediation. Fourier transform infrared spectroscopy (FTIR) analysis confirmed interaction of arsenate and arsenite with functional groups (aromatic amino acid residues) on the cell surface leading to characteristic peak shifts. Thus, the isolated AT-02 has the potential to remediate both arsenite and arsenate from contaminated environmental sites.
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A method is described for reconstructing presumed cladistic evolutionary sequences of recent organisms and its implications are discussed. Characters of the organisms to be studied are presented in a data matrix of the type employed in numerical taxonomy with the character states arrayed according to a presumed evolutionary sequence. The reconstruction proceeds on the hypothesis that the minimum number of evolutionary steps yields the correct cladogram. The method has been programmed for computer processing.
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There are essentially three ways in which four species may be related in a phylogenetic tree graph. It is usual to compute for each of these three possibilities the smallest number of mutations that could have brought about the observed distribution of characteristics among the four species. The graph that minimizes this number is then preferred. In fact, the hypothesis that the graph chosen in this way is correct may be accepted with confidence if the minimum is strong in a sense described here. In principle, the theory could be extended to treat sets of more than four species.
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In the past few years there has been a surge in the development of new statistical theories and methods that take advantage of the high speed digital computer. The payoff for such intensive computation methods is freedom from two limiting factors that have dominated statistical theory since its beginning: the assumption that the data conform to a bell-shaped curve and the need to focus on statistical measures whose theoretical properties can be analyzed mathematically. The new methods free the statistician to attack more complicated problems, exploiting a wider array of statistical tools. The bootstrap method is examined and evaluated as an example of this new generation of statistical tools.
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This is an invited expository article for The American Statistician. It reviews the nonparametric estimation of statistical error, mainly the bias and standard error of an estimator, or the error rate of a prediction rule. The presentation is written at a relaxed mathematical level, omitting most proofs, regularity conditions, and technical details.
Article
We discuss the following problem given a random sample X = (X 1, X 2,…, X n) from an unknown probability distribution F, estimate the sampling distribution of some prespecified random variable R(X, F), on the basis of the observed data x. (Standard jackknife theory gives an approximate mean and variance in the case R(X, F) = θ(F^)θ(F)\theta \left( {\hat F} \right) - \theta \left( F \right), θ some parameter of interest.) A general method, called the “bootstrap”, is introduced, and shown to work satisfactorily on a variety of estimation problems. The jackknife is shown to be a linear approximation method for the bootstrap. The exposition proceeds by a series of examples: variance of the sample median, error rates in a linear discriminant analysis, ratio estimation, estimating regression parameters, etc.
Statistical inference of phylogenies Parsimony in systematics: Bio-logical and statistical issues Confidence limits on phylogenies with a molecular clock
  • J Felsenstein
FELSENSTEIN, J. 1983a. Statistical inference of phylogenies. J. Roy. Statist. Soc. A 146:246-272.. 1983b. Parsimony in systematics: Bio-logical and statistical issues. Ann. Rev. Ecol. Syst. 14:313-333. 1985. Confidence limits on phylogenies with a molecular clock. Systematic Zoology 34: 152-161.