METHODS: In this study, 135 mitochondrial cytochrome c oxidase subunit I (COI) sequences were established for 45 species in the genus Simulium in Vietnam, encompassing three subgenera (Gomphostilbia, Nevermannia, and Simulium), with 64 paratypes of 27 species and 16 topotypes of six species. Of these COI sequences, 71, representing 27 species, are reported for the first time.
RESULTS: Combined with GenBank sequences of specimens from Malaysia, Myanmar, Thailand, and Vietnam, a total of 234 DNA barcodes of 53 nominal species resulted in a 71% success rate for species identification. Species from the non-monophyletic Simulium asakoae, S. feuerborni, S. multistriatum, S. striatum, S. tuberosum, and S. variegatum species groups were associated with ambiguous or incorrect identifications. Pairwise distances, phylogenetics, and species delimitation analyses revealed a high level of cryptic diversity, with discovery of 15 cryptic taxa. The current study also revealed the limited utility of a fast-evolving nuclear gene, big zinc finger (BZF), in discriminating closely related, morphologically similar nominal species of the S. asakoae species group.
CONCLUSION: This study represents the first comprehensive molecular genetic analysis of the black fly fauna in Vietnam to our knowledge, providing a foundation for future research. DNA barcoding exhibits varying levels of differentiating efficiency across species groups but is valuable in the discovery of cryptic diversity.
METHODS: Genomic DNA of Indonesian black fly samples were extracted and sequenced, producing 86 COI sequences in total. Two hundred four COI sequences, including 118 GenBank sequences, were analysed. Maximum likelihood (ML) and Bayesian inference (BI) trees were constructed and species delimitation analyses, including ASAP, GMYC and single PTP, were performed to determine whether the species of Indonesian black flies could be delineated. Intra- and interspecific genetic distances were also calculated and the efficacy of COI sequences for species identification was tested.
RESULTS: The DNA barcodes successfully distinguished most morphologically distinct species (> 80% of sampled taxa). Nonetheless, high maximum intraspecific distances (3.32-13.94%) in 11 species suggested cryptic diversity. Notably, populations of the common taxa Simulium (Gomphostilbia) cheongi, S. (Gomphostilbia) sheilae, S. (Nevermannia) feuerborni and S. (Simulium) tani in the islands of Indonesia were genetically distinct from those on the Southeast Asian mainland (Malaysia and Thailand). Integrated morphological, cytogenetic and nuclear DNA studies are warranted to clarify the taxonomic status of these more complex taxa.
CONCLUSIONS: The findings showed that COI barcoding is a promising taxonomic tool for Indonesian black flies. The DNA barcodes will aid in correct identification and genetic study of Indonesian black flies, which will be helpful in the control and management of potential vector species.
METHODS: H. contortus specimens (n = 57) were isolated from wild blue sheep (Pseudois nayaur) inhabiting Helan Mountains (HM), China and additional H. contortus specimens (n = 20) were isolated from domestic sheep that were grazed near the natural habitat of the blue sheep. Complete ITS2 (second internal transcribed spacer) sequences and partial sequences of the nad4 (nicotinamide dehydrogenase subunit 4 gene) gene were amplified to determine the sequence variations and population genetic diversities between these two populations. Also, 142 nad4 haplotype sequences of H. contortus from seven other geographical regions of China were retrieved from database to further examine the H. contortus population structure.
RESULTS: Sequence analysis revealed 10 genotypes (ITS2) and 73 haplotypes (nad4) among the 77 specimens, with nucleotide diversities of 0.007 and 0.021, respectively, similar to previous studies in other countries, such as Pakistan, Malaysia and Yemen. Phylogenetic analyses (BI, MP, NJ) of nad4 sequences showed that there were no noticeable boundaries among H. contortus populations from different geographical origin and population genetic analyses revealed that most of the variation (94.21%) occurred within H. contortus populations. All phylogenetic analyses indicated that there was little genetic differentiation but a high degree of gene flow among the H. contortus populations among wild blue sheep and domestic ruminants in China.
CONCLUSIONS: The current work is the first genetic characterization of H. contortus isolated from wild blue sheep in the Helan Mountains region. The results revealed a low genetic differentiation and high degree of gene flow between the H. contortus populations from sympatric wild blue sheep and domestic sheep, indicating regular cross-infection between the sympatrically reared ruminants.
METHODS: In the present study, we included a larger sample size of P. knowlesi (83 samples) covering eight states of Malaysia to determine the genetic polymorphism, natural selection and haplotype groups of the gene fragment coding PkMSP-142. The region flanking PkMSP-142 was amplified by PCR and directly sequenced. Genetic diversity, haplotype diversity, population genetic differentiation and natural selection were determined in order to study the polymorphic characteristic of PkMSP-142.
RESULTS: A high level of genetic diversity (Hd = 0.970 ± 0.007; л = 0.01079 ± 0.00033) was observed among the 83 P. knowlesi samples, confirming the extensive genetic polymorphism exhibited among the P. knowlesi population found in Malaysia. A total of 18 distinct haplotypes with 17 amino acid changes were identified, whereby 15 were new haplotypes. High population differentiation values were observed within samples from Peninsular Malaysia and Malaysian Borneo. The 42 kDa fragments of P. knowlesi from Malaysian Borneo were found to be acting on balancing selection whilst purifying selection was suggested to act on isolates from Peninsular Malaysia. The separation of PkMSP-142 haplotypes into two main groups based on geographical separation has further supported the existence of two distinct P. knowlesi lineages.
CONCLUSIONS: A high level of genetic diversity was observed among PkMSP-142 in Malaysia, whereby most of the polymorphisms were found within the 33 kDa region. Taken together, these data will be useful in order to understand the nature of P. knowlesi population in Malaysia as well as the design and development of a MSP-142 based knowlesi malaria vaccine.
METHODS: The helminth community structures of two urban rat populations in Kuala Lumpur, Malaysia were investigated. The rats were from two contrasting sites in the city caught over a period of 21 months in 2000-2002.
RESULTS: Eleven species of helminth parasites comprising seven nematodes (Heterakis spumosum, Mastophorus muris, Nippostrongylus brasiliensis, Syphacia muris, Pterygodermatites tani/whartoni, Gongylonema neoplasticum, Angiostrongylus malaysiensis), three cestodes (Hymenolepis (Rodentolepis) nana, H. diminuta and Taenia taeniaeformis) and one acanthocephalan (Moniliformis moniliformis) were recovered from 346 Rattus rattus and 104 R. norvegicus from two urban sites, Bangsar and Chow Kit, during 2000-2002. Rattus rattus harboured over 60% of all helminths compared with R. norvegicus, although both host species played a dominant role in the different sites with, for example R. norvegicus at Bangsar and R. rattus at Chow Kit accounting for most of the nematodes. Overall 80% of rats carried at least one species of helminth, with the highest prevalences being shown by H. diminuta (35%), H. spumosum (29.8%) and H. nana (28.4%). Nevertheless, there were marked differences in prevalence rates between sites and hosts. The influence of extrinsic (year, season and site) and intrinsic (species, sex and age) factors affecting infracommunity structure (abundance and prevalence of infection) and measures of component community structure were analyzed.
CONCLUSIONS: Since at least two species of rat borne helminths in Kuala Lumpur have the potential to infect humans, and these showed high prevalences in the rats, the assessment and regular monitoring of infections carried by wild rodents have important roles to play in public health.
RESULTS: An RNAi construct targeting the RNA recognition motif of the Aedes aegypti transformer-2 (tra-2) gene does not trigger female-to-male sex conversion as commonly observed among dipterous insects. Instead, homozygous insects show greater mortality among m-chromosome-bearing sperm and mm zygotes, yielding up to 100% males in the subsequent generations. The performance of transgenic males was not significantly different to wild-type males in narrow-cage competitive mating experiments.
CONCLUSION: Our data provide preliminary evidence that the knockdown of Ae. aegypti tra-2 gene expression causes segregation distortion acting at the level of gametic function, which is reinforced by sex-specific zygotic lethality. This finding could promote the development of new synthetic sex distorter systems for the production of genetic sexing mosquito strains.
METHODS: A retrospective study was performed on diagnostic results of Fasciola spp., Toxocara spp., Strongyloides stercoralis and Taenia solium IgG ELISA tests from Medic Medical Center Laboratory, Ho Chi Minh City in 2012. The data were first stratified before statistical analyses were performed. Seroprevalence of fascioliasis, toxocariasis, strongyloidiasis and cysticercosis was determined and the age and gender risk factors were evaluated.
RESULTS: Seroprevalence of fascioliasis, toxocariasis, strongyloidiasis and cysticercosis was 5.9 % (590/10,084; 95 % CI: 5.44-6.36), 45.2 % (34,995/77,356; 95 % CI: 44.85-45.55), 7.4 % (3,174/42,920; 95 % CI: 7.15-7.65) and 4.9 % (713/14,601; 95 % CI: 4.55-5.25), respectively. Co-exposure to multiple parasites was detected in 890 males (45.7 %; 95 % CI: 43.49-47.91) and 1,059 females (54.3 %; 95 % CI: 52.09-56.51). Social structure and differences in behavioural factors caused the gender factor to have a significant effect on the prevalence of all the diseases, while the seropositivity for fascioliasis and strongyloidiasis were age group-related.
CONCLUSIONS: The seroprevalence of fascioliasis, toxocariasis, strongyloidiasis and cysticercosis in the blood samples diagnosed in Medic Medical Center Laboratory, Ho Chi Minh City, in year 2012 were comparatively high. The Vietnamese customs and cultures, dietary habits and agricultural practices exposed them to high risk of contracting NTDs. Despite the possibility of false positive results due to antigenic cross-reactions, detection of IgG antibodies remains as a reliable method in sero-epidemiological study as it is non-invasive and demonstrates previous exposure of individuals to the parasites. Besides the implementation of strategies to control these diseases, epidemiological analysis and surveillance of diseases should also be continually strengthened to monitor the effectiveness of regimens and interventions.
RESULTS: In the erythrocyte-binding assay, binding level was determined by scoring the number of rosettes that were formed by erythrocytes surrounding transfected mammalian COS-7 cells which expressed PkDBPαII. The assay result revealed a significant difference in the binding level. The number of rosettes scored for Fya+/b+ was 1.64-fold higher than that of Fya+/b- (155.50 ± 34.32 and 94.75 ± 23.16 rosettes, respectively; t(6) = -2.935, P = 0.026).
CONCLUSIONS: The erythrocyte-binding assay provided a simple approach to quantitatively determine the binding level of PkDBPαII to the erythrocyte Duffy antigen. Using this assay, PkDBPαII was found to display higher binding to Fya+/b+ erythrocytes than to Fya+/b- erythrocytes.
METHODS: Water samples were collected from rivers, water tanks, wells and seawater on Tioman Island over the course of April to October 2015. Water samples were indirectly screened for Sarcocystis species by obtaining sediment from respective water sources. PCR amplification of the 18S rRNA gene region was conducted to identify positive samples. Microscopy was used in an attempt to reappraise PCR results, but no sporocysts were detected in any of the samples.
RESULTS: A total of 157 water samples were obtained and 19 were positive for various Sarcocystis species. Through BLASTn and phylogenetic analysis, these species were found to be S. singaporensis, S. nesbitti, Sarcocystis sp. YLL-2013 and one unidentified Sarcocystis species.
CONCLUSIONS: This is the first positive finding of S. nesbitti in water samples on Tioman Island, which was found in a water tank and in river water samples. This finding supports the hypothesis that water was a potential medium for the transmission of S. nesbitti during the outbreak. This will potentially identify areas in which preventive measures can be taken to prevent future outbreaks.
RESULTS: Yeast two-hybrid (Y2H) experiment was used to identify the binding partners of surface antigens of T. gondii by using SAG2 as bait. Colony PCR was performed and positive clones were sent for sequencing to confirm their identity. The yeast plasmids for true positive clones were rescued by transformation into E. coli TOP 10F' cells. The interplay between bait and prey was confirmed by β-galactosidase assay and co-immunoprecipitation experiment. We detected 20 clones interacting with SAG2 based on a series of the selection procedures. Following the autoactivation and toxicity tests, SAG2 was proven to be a suitable candidate as a bait. Thirteen clones were further examined by small scale Y2H experiment. The results indicated that a strong interaction existed between Homo sapiens zinc finger protein and SAG2, which could activate the expressions of the reporter genes in diploid yeast. Co-immunoprecipitation experiment result indicated the binding between this prey and SAG2 protein was significant (Mann-Whitney U-test: Z = -1.964, P = 0.05).
CONCLUSIONS: Homo sapiens zinc finger protein was found to interact with SAG2. To improve the understanding of this prey protein's function, advanced investigations need to be carried out.
METHODS: Using the Center for Disease Control and Prevention (CDC) bottle assays, the insecticide resistance status of nine different Ae. aegypti strains from Selangor was accessed. Synergism tests and biochemical assays were conducted to further understand the metabolic mechanisms of insecticide resistance. Polymerase chain reaction (PCR) amplification and sequencing of the IIP-IIS6 as well as IIIS4-IIIS6 regions of the sodium channel gene were performed to enable comparisons between susceptible and resistant mosquito strains. Additionally, genomic DNA was used for allele-specific PCR (AS-PCR) genotyping of the gene to detect the presence of F1534C, V1016G and S989P mutations.
RESULTS: Adult female Ae. aegypti from various locations were susceptible to malathion and propoxur. However, they exhibited different levels of resistance against dichlorodiphenyltrichloroethane (DDT) and pyrethroids. The results of synergism tests and biochemical assays indicated that the mixed functions of oxidases and glutathione S-transferases contributed to the DDT and pyrethroid resistance observed in the present study. Besides detecting three single kdr mutations, namely F1534C, V1016G and S989P, co-occurrence of homozygous V1016G/S989P (double allele) and F1534C/V1016G/S989P (triple allele) mutations were also found in Ae. aegypti. As per the results, the three kdr mutations had positive correlations with the expressions of resistance to DDT and pyrethroids.
CONCLUSIONS: In view of the above outcomes, it is important to seek new tools for vector management instead of merely relying on insecticides. If the latter must be used, regular monitoring of insecticide resistance should also be carried out at all dengue epidemic areas. Since the eggs of Ae. aegypti can be easily transferred from one location to another, it is probable that insecticide-resistant Ae. aegypti can be found at non-dengue outbreak sites as well.