Diversity Of Mosquito Species Ovipositing In Different Zones of Light Intensity Within Limestone Caves In Thailand.

Climate change and human activity affect the geographical and annual distribution and population abundance of mosquitoes. As natural habitats are reduced, it is hypothesized that mosquitoes may seek refuge in more stable environments such as cave habitats. Therefore, we explored the species diversity of mosquitoes exploiting cave habitats in Thailand. Ten species belonging to 4 genera were collected, of which none were considered to be true cave-dwelling species (Troglobiont). The known cavernicolous species, Aedes cavaticus, was observed to oviposit outside of the cave and therefore should be categorized as a subtroglophilic species. Other species were also oviposited inside the cave but should be regarded as trogloxenic species. There was no clear association between environmental factors and mosquito abundance inside the 4 limestone caves, except for Ae. cavaticus, which was positively correlated with rainfall. This study indicates that different biotic or abiotic factors may be involved in mosquito oviposition site selection inside caves.

Phylogenetic Relationships Among Malaria Vectors and Closely Related Species in Thailand Using Multilocus DNA Sequences.

The evolutionary and taxonomic status is important for understanding speciation events and phylogenetic relationships between closely related vector and nonvector species. This information is useful for targeting important disease vector species groups for the development of novel genetic-based vector and pathogen control methods. In this study, different phylogenetic analyses were performed to reconstruct phylogenetic trees for the primary malaria vectors in Thailand based on sequence information of 4 DNA fragments from the nuclear and mitochondrial regions. The primary Anopheles species in the subgenus Cellia involved in malaria transmission in Thailand separate clearly into 3 distinct clades: the Leucosphyrus group, Minimus subgroup, and Maculatus group. The phylogenetic trees based on different reconstructed algorithms and different gene regions provided congruent phylogenetic status of the mosquito species studied. The phylogenetic relationships of malaria vector species examined followed similar patterns based on morphological characters. An estimate of the divergence time among the Anopheles species infers that they were present during the Eocene and Miocene periods (>41 million years ago). Congruent phylogenetic analysis of malaria vectors is presented with different algorithms and gene regions. The nuclear TOLL6 fragment appears useful for molecular phylogenetic, species DNA barcode, and Anopheles population genetic analyses.

Molecular Identification of Four Members of the Anopheles dirus Complex Using the Mitochondrial Cytochrome C Oxidase Subunit I Gene.

Precise mosquito species identification is an essential step for proper management and control of malaria vectors. Misidentification of members in the Anopheles dirus complex, some which are primary malaria vectors in Thailand and mainland Southeast Asia, remains problematic because of indistinguishable or overlapping morphological characters between sibling species. Moreover, there is a need for alternative methods, since the existing molecular techniques in the literature are not entirely satisfactory in differentiating all members in the An. dirus complex. The nucleotide polymorphisms in the mitochondrial cytochrome c oxidase subunit I (COI) sequences were developed to identify the 4 species within the An. dirus complex using an allele-specific (AS) multiplex polymerase chain reaction (PCR). The identified primers amplified and clearly differentiated the 4 members of the complex found in Thailand, Anopheles dirus, An. cracens, An. scanloni, and An. baimaii with PCR products 428/104, 236, 625, and 428 bp, respectively. These results demonstrate that an AS-PCR based on the COI region can accurately identify 4 members of An. dirus complex and would be useful as an alternative PCR-based method for accurate species identification.

Active Compounds Against Anopheles minimus Carboxypeptidase B for Malaria Transmission-Blocking Strategy.

Malaria transmission-blocking compounds have been studied to block the transmission of malaria parasites, especially the drug-resistant Plasmodium. Carboxypeptidase B (CPB) in the midgut of Anopheline mosquitoes has been demonstrated to be essential for the sexual development of Plasmodium in the mosquito. Thus, the CPB is a potential target for blocking compounds. The aim of this research was to screen compounds from the National Cancer Institute (NCI) diversity dataset and U.S. Food and Drug Administration (FDA)-approved drugs that could reduce the Anopheles CPB activity. The cDNA fragment of cpb gene from An. minimus (cpbAmi) was amplified and sequenced. The three-dimensional structure of CPB was predicted from the deduced amino acid sequence. The virtual screening of the compounds from NCI diversity set IV and FDA-approved drugs was performed against CPBAmi. The inhibition activity against CPBAmi of the top-scoring molecules was characterized in vitro. Three compounds-NSC-1014, NSC-332670, and aminopterin with IC50 at 0.99 mM, 1.55 mM, and 0.062 mM, respectively-were found to significantly reduce the CPBAmi activity.

Multilocus nuclear DNA markers reveal population structure and demography of Anopheles minimus.

Utilization of multiple putatively neutral DNA markers for inferring evolutionary history of species population is considered to be the most robust approach. Molecular population genetic studies have been conducted in many species of Anopheles genus, but studies based on single nucleotide polymorphism (SNP) data are still very scarce. Anopheles minimus is one of the principal malaria vectors of Southeast (SE) Asia including the Northeastern (NE) India. Although population genetic studies with mitochondrial genetic variation data have been utilized to infer phylogeography of the SE Asian populations of this species, limited information on the population structure and demography of Indian An. minimus is available. We herewith have developed multilocus nuclear genetic approach with SNP markers located in X chromosome of An. minimus in eight Indian and two SE Asian population samples (121 individual mosquitoes in total) to infer population history and test several hypotheses on the phylogeography of this species. While the Thai population sample of An. minimus presented the highest nucleotide diversity, majority of the Indian samples were also fairly diverse. In general, An. minimus populations were moderately substructured in the distribution range covering SE Asia and NE India, largely falling under three distinct genetic clusters. Moreover, demographic expansion events could be detected in the majority of the presently studied populations of An. minimus. Additional DNA sequencing of the mitochondrial COII region in a subset of the samples (40 individual mosquitoes) corroborated the existing hypothesis of Indian An. minimus falling under the earlier reported mitochondrial lineage B.

Molecular population genetics of the NADPH cytochrome P450 reductase (CPR) gene in Anopheles minimus.

Development of insecticide resistance (IR) in mosquito vectors is a primary huddle to malaria control program. Since IR has genetic basis, and genes constantly evolve with response to environment for adaptation to organisms, it is important to know evolutionary pattern of genes conferring IR in malaria vectors. The mosquito Anopheles minimus is a major malaria vector of the Southeast (SE) Asia and India and is susceptible to all insecticides, and thus of interest to know if natural selection has shaped variations in the gene conferring IR. If not, the DNA fragment of such a gene could be used to infer population structure and demography of this species of malaria vector. We have therefore sequenced a ~569 bp DNA segment of the NADPH cytochrome P450 reductase (CPR) gene (widely known to confer IR) in 123 individuals of An. minimus collected in 10 different locations (eight Indian, one Thai and one Vietnamese). Two Indian population samples were completely mono-morphic in the CPR gene. In general, low genetic diversity was found with no evidence of natural selection in this gene. The data were therefore analyzed to infer population structure and demography of this species. The 10 populations could be genetically differentiated into four different groups; the samples from Thailand and Vietnam contained high nucleotide diversity. All the 10 populations conform to demographic equilibrium model with signature of past population expansion in four populations. The results in general indicate that the An. minimus mosquitoes sampled in the two SE Asian localities contain several genetic characteristics of being parts of the ancestral population.

Population structure and genetic bottleneck in sweet cherry estimated with SSRs and the gametophytic self-incompatibility locus.

BACKGROUND: Domestication and breeding involve the selection of particular phenotypes, limiting the genomic diversity of the population and creating a bottleneck. These effects can be precisely estimated when the location of domestication is established. Few analyses have focused on understanding the genetic consequences of domestication and breeding in fruit trees. In this study, we aimed to analyse genetic structure and changes in the diversity in sweet cherry Prunus avium L. RESULTS: Three subgroups were detected in sweet cherry, with one group of landraces genetically very close to the analysed wild cherry population. A limited number of SSR markers displayed deviations from the frequencies expected under neutrality. After the removal of these markers from the analysis, a very limited bottleneck was detected between wild cherries and sweet cherry landraces, with a much more pronounced bottleneck between sweet cherry landraces and modern sweet cherry varieties. The loss of diversity between wild cherries and sweet cherry landraces at the S-locus was more significant than that for microsatellites. Particularly high levels of differentiation were observed for some S-alleles. CONCLUSIONS: Several domestication events may have happened in sweet cherry or/and intense gene flow from local wild cherry was probably maintained along the evolutionary history of the species. A marked bottleneck due to breeding was detected, with all markers, in the modern sweet cherry gene pool. The microsatellites did not detect the bottleneck due to domestication in the analysed sample. The vegetative propagation specific to some fruit trees may account for the differences in diversity observed at the S-locus. Our study provides insights into domestication events of cherry, however, requires confirmation on a larger sampling scheme for both sweet cherry landraces and wild cherry.

Population genetics of speciation in two closely related wild tomatoes (Solanum section Lycopersicon).

We present a multilocus sequencing study to assess patterns of polymorphism and divergence in the closely related wild tomato species, Solanum peruvianum and S. chilense (Solanum section Lycopersicon, Solanaceae). The data set comprises seven mapped nuclear loci (approximately 9.3 kb of analyzed sequence across loci) and four local population samples per species that cover much of the species' range (between 80 and 88 sequenced alleles across both species). We employ the analytical framework of divergence population genetics (DPG) in evaluating the utility of the "isolation" model of speciation to explain observed patterns of polymorphism and divergence. Whereas the isolation model is not rejected by goodness-of-fit criteria established via coalescent simulations, patterns of intragenic linkage disequilibrium provide evidence for postdivergence gene flow at two of the seven loci. These results suggest that speciation occurred under residual gene flow, implying that natural selection is one of the evolutionary forces driving the divergence of these tomato species. This inference is fully consistent with their recent divergence, conservatively estimated to be

Using multilocus sequence data to assess population structure, natural selection, and linkage disequilibrium in wild tomatoes.

We employed a multilocus approach to examine the effects of population subdivision and natural selection on DNA polymorphism in 2 closely related wild tomato species (Solanum peruvianum and Solanum chilense), using sequence data for 8 nuclear loci from populations across much of the species' range. Both species exhibit substantial levels of nucleotide variation. The species-wide level of silent nucleotide diversity is 18% higher in S. peruvianum (pi(sil) approximately 2.50%) than in S. chilense (pi(sil) approximately 2.12%). One of the loci deviates from neutral expectations, showing a clinal pattern of nucleotide diversity and haplotype structure in S. chilense. This geographic pattern of variation is suggestive of an incomplete (ongoing) selective sweep, but neutral explanations cannot be entirely dismissed. Both wild tomato species exhibit moderate levels of population differentiation (average F(ST) approximately 0.20). Interestingly, the pooled samples (across different demes) exhibit more negative Tajima's D and Fu and Li's D values; this marked excess of low-frequency polymorphism can only be explained by population (or range) expansion and is unlikely to be due to population structure per se. We thus propose that population structure and population/range expansion are among the most important evolutionary forces shaping patterns of nucleotide diversity within and among demes in these wild tomatoes. Patterns of population differentiation may also be impacted by soil seed banks and historical associations mediated by climatic cycles. Intragenic linkage disequilibrium (LD) decays very rapidly with physical distance, suggesting high recombination rates and effective population sizes in both species. The rapid decline of LD seems very promising for future association studies with the purpose of mapping functional variation in wild tomatoes.