[Show abstract][Hide abstract] ABSTRACT: Nine and 11 isolines of Anopheles argyropus and Anopheles pursati, respectively, were established from individual females collected from cow-baited traps, and the characteristics of metaphase chromosomes were investigated in their F1-progenies. As determined by the different amounts of extra heterochromatin on sex chromosomes, 2 types of X (X1, X2) and Y (Y1,Y2), and 2 types of X (X1, X2) and 3 types of Y (Y1, Y2, Y3) chromosomes were obtained from An. argyropus and An. pursati, respectively. These types of sex
chromosomes comprised 2 [Forms A (X1, Y1) and B (X1, X2, Y2)] and 3 [Forms A (X1, X2, Y1), B (X1, X2, Y2) and C (X2, Y3)] karyotypic forms of An. argyropus and An. pursati, respectively. All karyotypic forms acquired from An. pursati are new one that were discovered in this study, of which Forms A, B and C were found generally in Chiang Mai Province, while only 1 isoline of Form B was obtained in Ratchaburi Province. Form A was recovered from An. argyropus only in Ubon Ratchathani Province, whereas Form B from that species was found commonly in both Ubon Rathchathani and Nakhon Si Thammarat Provinces. Crossing
experiments among the 2 and 3 isolines representing 2 and 3 karyotypic forms of An. argyropus and An. pursati, respectively, indicated genetic compatibility in yielding viable progenies and synaptic salivary gland polytene chromosomes through F2-generations. The conspecific natures of these karyotypic forms in both species were further supported by very low intraspecific sequence variations (average genetic distance: An. argyropus = 0.003-0.007, An. pursati = 0-0.005) of ribosomal DNA (ITS2) and mitochondrial DNA (COI and COII).
[Show abstract][Hide abstract] ABSTRACT: Twenty-nine isolines of Anopheles crawfordi were established from wild-caught females collected from cow-baited traps in Thailand and Cambodia. Three types of X (X1, X2, X3) and four types of Y (Y1, Y2, Y3, and Y4) chromosomes were identified, according to differing amounts of extra heterochromatin. These sex chromosomes represent four metaphase karyotypes, i.e., Forms A (X1, X2, X3, Y1), B (X1, X2, X3, Y2), C (X2, Y3) and D (X2, Y4). Forms C and D are novel metaphase karyotypes confined to Thailand, whereas forms A and B appear to be common in both Thailand and Cambodia. Cross-mating experiments between the four karyotypic forms indicated genetic compatibility in yielding viable progenies and synaptic salivary gland polytene chromosomes. The results suggest that the forms are conspecific and A. crawfordi comprises four cytological races, which is further supported by very low intraspecific variation (mean genetic distance = 0.000–0.018) of the nucleotide sequences in ribosomal DNA (ITS2) and mitochondrial DNA sequences (COI, COII).
[Show abstract][Hide abstract] ABSTRACT: In this study, we focused on the molecular detection of Coxiella-like bacteria using a PCR technique to identify Coxiella 16S rRNA sequences in Haemaphysalis tick samples (105 adults, 8 nymph pools and 19 larval pools). Seven Haemaphysalis species obtained from 5 locations in Thailand were evaluated in this work. Coxiella endosymbionts could be detected in samples representing all 3 growth stages examined. The results also revealed that only 4 of 7 tick species were positive for Coxiella-like endosymbiont: Haemaphysalis hystricis, Haemaphysalis lagrangei, Haemaphysalis obesa, and Haemaphysalis shimoga. Haemaphysalis shimoga demonstrated the highest percentage of Coxiella-like positive samples (58.33% with n = 24), while Haemaphysalis hystricis had the lowest percentage; only 1 female tick was positive for Coxiella-like bacteria (n = 6). Interestingly, the results indicated that female Haemaphysalis ticks tended to harbour Coxiella symbionts more frequently than male ticks (59.32% of females and 21.27% of males of all species studied). Phylogenetic analyses based on 16S rRNA sequences illustrated that Coxiella-like spp. from the same tick species always grouped in same clade, regardless of the location from which they were isolated. Moreover, a phylogenetic tree also showed that Coxiella-like endosymbionts from other genera (for example, the tick genus Rhipicephalus) formed a separate group compared to Coxiella-like symbionts in the genus Haemaphysalis. This suggests that a high amount of DNA sequence variation is present in Coxiella-like bacteria harboured by ticks.
[Show abstract][Hide abstract] ABSTRACT: Some reptile ticks are potential vectors of pathogens such as spotted fever group (SFG) rickettsiae. Here, we report for the first time in detail the molecular evidence, DNA sequences and phylogenetic studies, for the presence of Rickettsia spp. in Amblyomma ticks (Amblyomma helvolum and Amblyomma varanense) from snakes in Thailand. A total of 24 tick samples was collected from 4 snake species and identified. A phylogenetic analysis inferred from the partial sequences of the gltA gene indicated that the Rickettsia spp. from 2 Amblyomma helvolum and 1 Amblyomma varanense belong to the same group as the SFG rickettsiae, which are closely related to Rickettsia raoultii strains. In contrast, there was 1 Rickettsia sp. from Amblyomma helvolum grouped into the same clade with other SFG rickettsiae (Rickettsia tamurae, Rickettsia monacensis, and a Rickettsia endosymbiont of Amblyomma dubitatum from Brazil). However, another Rickettsia sp. from Amblyomma varanense was closely related to Rickettsia bellii and Rickettsia sp. strain RDa420 from Thailand. In addition, from phylogenetic results based on the 16S rRNA gene and a concatenated tree of the 3 genes (gltA, ompA, and ompB), we found what may be a novel SFG rickettsia species closely related to Rickettsia raoultii (from both Amblyomma varanense and Amblyomma helvolum). In conclusion, our findings are the first report on the presence of novel SFG rickettsiae in 2 snake tick species, Amblyomma varanense and Amblyomma helvolum in Thailand and in south-eastern Asia.
[Show abstract][Hide abstract] ABSTRACT: Southeast Asia harbours abundant biodiversity, hypothesized to have been generated by Pliocene and Pleistocene climatic and environmental change. Vicariance between the island of Borneo, the remaining Indonesian archipelago and mainland Southeast Asia caused by elevated sea levels during interglacial periods has been proposed to lead to diversification in the littoral zone mosquito Anopheles (Cellia) sundaicus (Rodenwaldt) sensu lato. To test this biogeographical hypothesis we inferred the population history and assessed gene flow of An. sundaicus s.l. sampled from 18 populations across its pan-Asian species range, using sequences from mitochondrial cytochrome c oxidase subunit 1 (CO1); the internal transcribed spacer 2 (ITS2); and the mannose phosphate isomerase (Mpi) gene. A hypothesis of ecological speciation for An. sundaicus involving divergent adaptation to brackish and freshwater larval habitats was also previously proposed, based on a deficiency of heterozygotes for Mpi allozyme alleles in sympatry. This hypothesis was not supported by Mpi sequence data, exhibiting no differentiation between brackish and freshwater larval habitats. Mpi and CO1 supported the presence of up to eight genetically distinct population groupings. Counter to the hypothesis of three allopatric species, divergence was often no greater between Borneo, Sumatra/Java and the Southeast Asian mainland than it was between genetic groupings within these landmasses. An Isolation-with-Migration (IM) model indicates recurrent gene flow between the current major landmasses. Such gene flow would have been possible during glacial periods when the current landmasses merged, presenting opportunities for dispersal along expanding and contracting coastlines. Consequently, Pleistocene climatic variation has proved a homogenising, rather than diversifying, force for An. sundaicus diversity.This article is protected by copyright. All rights reserved.
[Show abstract][Hide abstract] ABSTRACT: Ticks are important vectors of several bacterial pathogens, including Francisella. In this study, a total of 24 adult ticks (Amblyomma varanense and Amblyomma helvolum) collected from 4 species of snake from 3 provinces of Thailand was screened for the presence of Francisella-like endosymbionts (FLEs) by PCR. FLEs were detected in 46% (11/24) of all ticks examined. A phylogenetic analysis of the 16S rDNA sequence indicated that the 11 distinct genotypes of FLEs amplified from A. varanense and A. helvolum (from Khon Kaen and Pichit provinces, respectively) are in the same group, along with other FLEs amplified from the other tick genera. Interestingly, these FLEs are closely related to, but distinct (different clade) from, the FLEs isolated from different tick species previously reported. This work represents the first report of Francisella spp. in snake ticks from Thailand.
[Show abstract][Hide abstract] ABSTRACT: Thirteen isoline colonies of Anopheles nigerrimus were established from individual wild-caught females collected from cow-baited traps at locations in Thailand and Cambodia. Three types of X (X1 , X2 , X3 ) and 4 types of Y (Y1 , Y2 , Y3 , Y4 ) chromosomes were recovered, according to differing amounts of extra heterochromatin. Four karyotypic forms were designed depending upon apparently distinct figures of X and Y chromosomes, i.e., Form A (X1 , X2 , X3 , Y1 ), B (X2 , X3 , Y2 ), C (X1 , Y3 ), and D (X3 , Y4 ). Forms C and D were new metaphase karyotypes discovered in this study. Form A appeared to be common in both Thailand and Cambodia. Forms B and D were found to be rather specific to southern and northeastern Thailand, respectively, whereas Form C was confined to Cambodia. Hybridization experiments among the eight isoline colonies, which were representative of four karyotypic forms of An. nigerrimus, demonstrated genetic compatibility in giving viable progenies and synaptic salivary gland polytene chromosomes through F2 -generations. These results elucidated the conspecific relationship, comprising four cytological forms within this taxon. Supportive evidence was confirmed further by very low intraspecific sequence variations (average genetic distance = 0.002-0.007) of the nucleotide sequences in ribosomal DNA [second internal transcribed spacer (ITS2)] and mitochondrial DNA [cytochrome c oxidase subunit I (COI) and subunit II (COII)].
[Show abstract][Hide abstract] ABSTRACT: Eight species members of the Thai Hyrcanus Group were identified based on the intact morphology and molecular analysis (COI barcoding, 658bp) of F1-progenies. Five iso-female lines of each species were pooled in order to establish stock colonies. A stenogamous colony of each species was investigated by making 200 and 300 newly emerged adult females and males co-habit in a 30cm cubic cage for one week. After ovipositon, the spermathecae of females were examined for sperms. The results revealed that Anopheles argyropus, Anopheles crawfordi, Anopheles nitidus, Anopheles pursati, Anopheles sinensis, Anopheles nigerrimus, Anopheles paraliae and Anopheles peditaeniatus yielded insemination rates of 0%, 0%, 0%, 31%, 33%, 42%, 50% and 77%, respectively. Continuous selection to establish stenogamous colonies indicated that An. sinensis, An. pursati, An. nigerrimus, An. paraliae and An. peditaeniatus provided insemination rates of 33-34%, 27-31%, 42-58%, 43-57% and 61-86% in 1, 2, 5, 6 and 20 generations of passages, respectively.
[Show abstract][Hide abstract] ABSTRACT: The Hyrcanus Group comprises important vectors of malaria because of Plasmodium vivax Grassi and Feletti and filariasis caused by Brugia malayi Brug in many countries of South, Southeast and East Asian regions. In Thailand, eight species members (Anopheles argyropus Swellengrebel, An. crawfordi Reid, An. nigerrimus Giles, An. nitidus Harrison, Scanlon & Reid, An. paraliae Sandosham, An. peditaeniatus Leicester, An. pursati Laveran
and An. sinensis Wiedemann) of the Hyrcanus Group have been recognized. Due to morphological overlap, adult females of the Hyrcanus Group in Thailand have been
misidentified markedly among the eight species, particularly when using the traumatic scales of wild-caught specimens from epidemiology and control approaches.
Therefore, this study first developed a simple and robustmultiplex PCR assay, based on second internal transcribed spacer sequences of ribosomal DNA, for differentiating the
eight species members of the Thai Hyrcanus Group.
[Show abstract][Hide abstract] ABSTRACT: Sixteen isoline colonies of Anopheles paraliae were established from wild-caught females collected from cow-baited traps at 4 locations in Thailand. They showed 3 types of X (X1, X2, X3) and 5 types of Y (Y1, Y2, Y3, Y4, Y5) chromosomes based on the number and amount of major block(s) of heterochromatin present in the heterochromatic arm, and were designated as Forms A (X3, Y1), B (X1, X2, X3, Y2), C (X3, Y3), D (X1, X2, X3, Y4) and E (X3, Y5). Form A was found in Songkhla Province, Form B was obtained in Ratchaburi, Nakhon Si Thammarat and Songkhla Provinces, Form C was acquired in Chanthaburi Province, Form D was recovered in Ratchaburi and Songkhla Provinces, and Form E was encountered in Ratchaburi Province. Hybridization experiments among the 7 isoline colonies, which represented the 5 karyotypic forms of An. paraliae, revealed genetic compatibility in providing viable progenies and synaptic salivary gland polytene chromosomes through F2-generations, and thus suggest the conspecific nature of these karyotypic forms. These results were supported by the very low intraspecific sequence divergence (mean genetic distance = 0.000-0.002) of the nucleotide sequences in ribosomal DNA (ITS2) and mitochondrial DNA (COI and COII) of the 5 forms.
[Show abstract][Hide abstract] ABSTRACT: To assess differentiation and relationships between Anopheles lesteri and Anopheles paraliae we established three and five iso-female lines of An. lesteri from Korea and An. paraliae from Thailand, respectively. These isolines were used to investigate the genetic relationships between the two taxa by crossing experiments and by comparing DNA sequences of ribosomal DNA second internal transcribed spacer (ITS2) and mitochondrial DNA cytochrome c oxidase subunit I (COI) and subunit II (COII). Results of reciprocal and F1-hybrid crosses between An. lesteri and An. paraliae indicated that they were compatible genetically producing viable progenies and complete synaptic salivary gland polytene chromosomes without inversion loops in all chromosome arms. The pairwise genetic distances of ITS2, COI and COII between these morphological species were 0.040, 0.007-0.017 and 0.008-0.011, respectively. The specific species status of An. paraliae in Thailand and/or other parts of the continent are discussed.
Memórias do Instituto Oswaldo Cruz 05/2013; 108(3). DOI:10.1590/S0074-02762013000300009 · 1.59 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: An engorged female Amblyomma helvolum Koch tick was removed from an adult Varanus salvator Laurenti lizard during field collection in Thailand. After using polymerase chain reaction to amplify three genes (16S rDNA, gltA, and OmpA), we discovered the presence of a Rickettsia sp. of the Spotted Fever Group. Phylogenetic analysis revealed that this Rickettsia sp. is closely related to Rickettsia raoultii Mediannikov. Therefore, we report herein for the first time the detection of a novel Spotted Fever Group Rickettsia in an Amblyomma helvolum from a Varanus salvator in Thailand.
Journal of Medical Entomology 02/2013; 50(1):217-20. DOI:10.1603/ME12010 · 1.95 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The symbiotic microorganisms of arthropod vectors are highly significant from several points of view, partly due to their possible roles in the transmission of pathogenic causative agents by blood-sucking vectors. Although ticks are well studied because of their significance to human health, novel microbial associations remain to be described. This review summarises several endosymbiotic bacterial species in hard ticks from various parts of the world, including Coxiella-, Francisella-, Rickettsia- and Arsenophonus-like symbionts as well as Candidatus Midichloria mitochondrii and Wolbachia. New methodologies for the isolation and characterization of tick-associated bacteria will, in turn, encourage new strategies of tick control by studying their endosymbionts.
[Show abstract][Hide abstract] ABSTRACT: Background
Filariasis, caused by Brugia malayi, is a public health problem in Thailand. Currently, at least two locations in southern Thailand are reported to be active endemic areas. Two and four Mansonia species are primary and secondary vectors, respectively, of the nocturnally subperiodic race, whereas, Coquillettidia crassipes is a vector of the diurnally subperiodic race. Although several Anopheles species have been incriminated extensively as natural and/or suspected vectors of B. malayi, little is known about vector competence between indigenous Anopheles and this filaria in Thailand. Thus, the susceptibility levels of eight species members in the Thai An. hyrcanus group to nocturnally subperiodic B. malayi are presented herein, and the two main refractory factors that affect them in different degrees of susceptibility have been elucidated.
Aedes togoi (a control vector), An. argyropus, An. crawfordi, An. nigerrimus, An. nitidus, An. paraliae, An. peditaeniatus, An. pursati and An. sinensis were allowed to feed artificially on blood containing B. malayi microfilariae, and dissected 14 days after feeding. To determine factors that take effect at different susceptibility levels, stain-smeared blood meals were taken from the midguts of Ae. togoi, An. peditaeniatus, An. crawfordi, An. paraliae, An. sinensis and An. nitidus immediately after feeding, and their dissected-thoraxes 4 days post blood-feedings were examined consecutively for microfilariae and L1 larvae.
The susceptibility rates of Ae. togoi, An. peditaeniatus, An. crawfordi, An. nigerrimus, An. argyropus, An. pursati, An. sinensis, An. paraliae and An. nitidus to B. malayi were 70–95%, 70–100%, 80–85%, 50–65%, 60%, 60%, 10%, 5%, and 0%, respectively. These susceptibility rates related clearly to the degrees of normal larval development in thoracic muscles, i.e., Ae. togoi, An. peditaeniatus, An. crawfordi, An. paraliae, An. sinensis and An. nitidus yielded normal L1 larvae of 93.15%, 96.34%, 97.33%, 23.60%, 15.38% and 0%, respectively.
An. peditaeniatus, An. crawfordi, An. nigerrimus, An. argyropus and An. pursati were high potential vectors. An. paraliae and An. sinensis were low potential vectors, while An. nitidus was a refractory vector. Two refractory mechanisms; direct toxicity and/or melanotic encapsulation against filarial larval were involved in the refractoriness of development in the thoracic muscles of the mosquito.
[Show abstract][Hide abstract] ABSTRACT: Mitotic karyotypes of five species of tephritid fruit fly parasitoid (Hymenoptera: Braconidae) belonging to the three genera, Psyttalia, Diachasmimorpha and Fopius, are described. The haploid chromosome numbers (n) of the three genera are distinct, i.e., Psyttalia n = 17, Diachasmimorpha n = 20 and Fopius n = 23. The species within each genus share the same haploid chromosome number but the chromosomes differ in size and shape: 7 metacentric (M), 7 submetacentric (SM) and 3 subtelocentric (ST) chromosomes in P. fletcheri compared with 6 M, 9 SM and 2 ST in P. incisi; and 5 M, 13 SM and 2 ST in D. longicaudata compared with 6 M, 12 SM and 2 ST in D. dacusii. F. arisanus showed 5 M, 13 SM and 5 ST. These karyological data are useful for cytotaxonomy and contribute to a better understanding of the chromosomal evolution of these fruit fly parasitoids.
[Show abstract][Hide abstract] ABSTRACT: Fifty-three isolines of Anopheles peditaeniatus were established from individual wild-caught females collected from cow-baited traps in 17 provinces of Thailand. Three types of X (X1, X2, X3) and 6 types of Y (Y1,Y2, Y3, Y4, Y5, Y6) chromosomes were determined based on different amounts of major block(s) of heterochromatin. These sex chromosomes comprised 6 karyotypic forms designated as Forms A (X3, Y1), B (X1, X2, X3, Y2), C (X3, Y3), D (X1, X2, X3, Y4), E (X1, X2, X3,Y5) and F (X2, X3, Y6). Form F is a new metaphase karyotype discovered in this study and is commonly found in all regions. Form A was found only in Lampang province, whereas Form E is widespread throughout the country. Forms B, C and D were obtained from the northern, northeastern, western and southern regions. Crossing experiments among the 11 isoline colonies representing the 6 karyotypic forms of An. peditaeniatus indicated genetic compatibility yielding viable progenies and complete synapsis of salivary gland polytene chromosomes through to the F2-generations. The results suggested the conspecific nature of these karyotypic forms which were further supported by very low intraspecific variation (genetic distance = 0.000-0.003) of nucleotide sequences in ribosomal DNA (ITS2) and mitochondrial DNA (COI and COII).
[Show abstract][Hide abstract] ABSTRACT: The polytene chromosomes of 512 larvae of the Simulium nobile species group collected from 16 stream sites in northern, central and southern Thailand were examined. Band-by- band comparisons relative to the established standard chromosome map for the subgenus Simulium distinguished the 2 species of this group, S. nobile and S. nodosum. The differences are mainly due to the position of the nucleolar organizer, pseudochromocenter, and fixed and polymorphic inversions. S. nobile is a monomorphic species, while S. nodosum is a polymorphic species. The morphology of these taxa is described based on the gill filaments and the color of the antennae and female genitalia. S. nobile is distributed in southern Thailand at low altitudes, whereas the distribution of S. nodosum is in the north at higher altitudes.
Cytogenetic and Genome Research 07/2011; 134(4):308-18. DOI:10.1159/000329713 · 1.56 Impact Factor