Given that there is limited available information on the insensitive acetylcholinesterase in insect species in Malaysia, the present study aims to detect the presence of G119S mutation in the acetylcholinesterase gene of Culex quinquefasciatus from 14 residential areas across 13 states and a federal territory in Malaysia.
There has been no comprehensive study on biochemical characterization of insecticide resistance mechanisms in field populations of Malaysian Culex quinquefasciatus. To fill this void in the literature, a nationwide investigation was performed to quantify the enzyme activities, thereby attempting to characterize the potential resistance mechanisms in Cx. quinquefasciatus in residential areas in Malaysia.
The resistance status of riceland Culex vishnui against four major groups of insecticides (i.e., organochlorines, carbamates, organophosphates and pyrethroids) was investigated. Biochemical assays (ESTα, ESTβ, MFO and GST) were also conducted to detect the resistance levels. Throughout a 12-month study period, multiple insecticide resistance was observed in both larvae and adult Cx vishnui. Culex vishnui larvae exhibited low resistance against malathion, temephos and permethrin with resistance ratio (RR) values < 5. In adult bioassay, Cx. vishnui were highly resistant against all tested adulticides with 24h post-treatment mortality < 70%. Correlations between permethrin and malathion resistance, as well as between deltamethrin and cyfluthrin resistance were found in Cx. vishnui. The results indicated that mixed function oxidases activity of Cx. vishnui was the highest compared to ESTα, ESTβ and GST. Spearman rank-order analysis showed that ESTα, ESTβ and GST were involved in multiple resistances in Cx. vishnui. The findings of this study established a baseline of insecticide susceptibility and revealed the effects of agricultural insecticide pressure on the vectors of Japanese encephalitis in Malaysia.
This study reported the ant species that were recovered from monkey carcasses in three different ecological habitats in Malaysia. The study was conducted from 9 May - 10 October 2007, 6 May - 6 August 2008 and 26 May - 14 July 2009 in forested area (Gombak, Selangor), coastal area (Tanjong Sepat, Selangor) and highland area (Bukit Cincin, Pahang), respectively. Monkey carcass was used as a model for human decomposition in this study. A total of 4 replicates were used in each of the study sites. Ants were observed to prey on eggs, larvae, pupae and newly emerged flies. This study found that ant species could be found at all stages of decomposition, indicating that ants were not a significant indicator for faunal succession. However, different species of ants were obtained from monkey carcasses placed in different ecological habitats. Cardiocondyla sp. was only found on carcasses placed in the coastal area; while Pheidole longipes, Hypoponera sp. and Pachycondyla sp. were solely found on carcasses placed in the highland area. On the other hand, Pheidologeton diversus and Paratrechina longicornis were found in several ecological habitats. These data suggests that specific ant species can act as geographic indicators for different ecological habitats in forensic entomology cases in Malaysia.
The present study explored the intraspecific genetic diversity, dispersal patterns and phylogeographic relationships of Culex quinquefasciatus Say (Diptera: Culicidae) in Malaysia using reference data available in GenBank in order to reveal this species' phylogenetic relationships. A statistical parsimony network of 70 taxa aligned as 624 characters of the cytochrome c oxidase subunit I (COI) gene and 685 characters of the cytochrome c oxidase subunit II (COII) gene revealed three haplotypes (A1-A3) and four haplotypes (B1-B4), respectively. The concatenated sequences of both COI and COII genes with a total of 1309 characters revealed seven haplotypes (AB1-AB7). Analysis using tcs indicated that haplotype AB1 was the common ancestor and the most widespread haplotype in Malaysia. The genetic distance based on concatenated sequences of both COI and COII genes ranged from 0.00076 to 0.00229. Sequence alignment of Cx. quinquefasciatus from Malaysia and other countries revealed four haplotypes (AA1-AA4) by the COI gene and nine haplotypes (BB1-BB9) by the COII gene. Phylogenetic analyses demonstrated that Malaysian Cx. quinquefasciatus share the same genetic lineage as East African and Asian Cx. quinquefasciatus. This study has inferred the genetic lineages, dispersal patterns and hypothetical ancestral genotypes of Cx. quinquefasciatus.
A total of 29 female black flies were captured by a hand net as they swarmed around humans in Tam Dao National Park, Vinh Phuc Province, Vietnam. They included one species of the subgenus Gomphostilbia (Simulium (Gomphostilbia) asakoae Takaoka & Davies) and five species of the subgenus Simulium, of which one species is described as Simulium (Simulium) vietnamense sp. nov. and the other four species (S. (S.) chungi Takaoka & Huang, S. (S.) grossifilum Takaoka & Davies, S. (S.) maenoi Takaoka & Choochote, and S. (S.) rufibasis Brunetti) are newly recorded from Vietnam.
Four new species of black flies are described, and three others are reported as newly recorded, based on adults reared from pupae, pupae and larvae collected in and near Tam Dao National Park, Vinh Phuc Province, Vietnam. New species include Simulium (Gomphostilbia) hongthaii sp. nov., S. (G.) tamdaoense sp. nov. (both species placed in the asakoae species-group), S. (Simulium) taythienense sp. nov. and S. (S.) xuandai sp. nov. (the two latter species placed in the striatum species-group). Newly recorded species are S. (G.) brinchangense Takaoka, Sofian-Azirun & Hashim, S. (Nevermannia) aureohirtum Brunetti and S. (S.) brevipar Takaoka & Davies. These discoveries increase the number of species of black flies known in Vietnam from 21 to 28.
The population genetic structure of Simulium tani was inferred from mitochondria-encoded sequences of cytochrome c oxidase subunits I (COI) and II (COII) along an elevational gradient in Cameron Highlands, Malaysia. A statistical parsimony network of 71 individuals revealed 71 haplotypes in the COI gene and 43 haplotypes in the COII gene; the concatenated sequences of the COI and COII genes revealed 71 haplotypes. High levels of genetic diversity but low levels of genetic differentiation were observed among populations of S. tani at five elevations. The degree of genetic diversity, however, was not in accordance with an altitudinal gradient, and a Mantel test indicated that elevation did not have a limiting effect on gene flow. No ancestral haplotype of S. tani was found among the populations. Pupae with unique structural characters at the highest elevation showed a tendency to form their own haplotype cluster, as revealed by the COII gene. Tajima's D, Fu's Fs, and mismatch distribution tests revealed population expansion of S. tani in Cameron Highlands. A strong correlation was found between nucleotide diversity and the levels of dissolved oxygen in the streams where S. tani was collected.
Three new species of black flies, Simulium (Nevermannia) langbiangense, S. (N.) phami, and S. (N.) bachmaense, are described on the basis of females, males, pupae and larvae collected in Vietnam. All three species are assigned to the Simulium feuerborni species-group of Simulium (Nevermannia). Simulium (N.) langbiangense sp. nov. is characterized by the female sensory vesicle with a large opening, and a short common basal stalk of the six pupal gill filaments, S. (N.) phami sp. nov. is most striking in having the pupal gill with five filaments, a character not reported in species of the S. feuerborni species-group, and S. (N.) bachmaense sp. nov. is characterized by the female genital fork with a triangular lobe-like projection pointed posteromedially on each arm, and the small larval postgenal cleft. This represents the first record of the S. feuerborni species-group from Vietnam.
Susceptibility status of Aedes aegypti (L.) and Aedes albopictus Skuse larvae obtained from 12 states in Malaysia were evaluated against five insect growth regulators (IGRs), namely, pyriproxyfen, methoprene, diflubenzuron, cyromazine, and novaluron under laboratory conditions. Field populations of Ae. aegypti exhibited moderate resistance toward methoprene and low resistance toward pyriproxyfen, with resistance ratios of 12.7 and 1.4, respectively, but susceptibility to diflubenzuron, cyromazine, and novaluron. On the other hand, field populations of Ae. albopictus exhibited low resistance against diflubenzuron and novaluron, with resistance ratio of 2.1 and 1.0, respectively, but susceptibility to other tested IGRs. Our study concluded that the tested IGRs provide promising results and can be used to control field population of Ae. aegypti and Ae. albopictus, especially cyromazine. The use of IGR should be considered as an alternative when larvae develop resistance to conventional insecticides.
Surveillance of mosquitoes and their distribution in association with rainfall, relative humidity, and temperature were conducted in selected suburban and forested areas, namely, Sungai Penchala (Kuala Lumpur) and Taman Alam (Selangor) for 12 months. Armigeres kesseli was the most abundant species in Sungai Penchala while Aedes butleri was the most dominant species in Taman Alam. A positive correlation between mosquito distribution and rainfall was observed in selected mosquito species in Sungai Penchala (Armigeres kesseli, r = 0.75; Armigeres subalbatus, r = 0.62; and Aedes albopictus, r = 0.65) and Taman Alam (Armigeres sp, r = 0.59; Ae. butleri, r = 0.85; and Ae. albopictus, r = 0.62). However, no significant cor- relation was found either between selected mosquito species in both study areas and relative humidity or temperature. Results obtained suggested that vector control programs to be conducted based on temporal distribution of vectors in order to achieve beneficial outcomes with effective costing.
Given the lack of molecular evidence in altered target-site insecticide resistance mechanism in Aedes albopictus (Skuse) worldwide, the present study aims to detect the presence of A302S mutation in the gene encoding the gamma aminobutyric acid receptor resistant to dieldrin (Rdl) in Ae. albopictus for the first time from its native range of South East Asia, namely Malaysia. World Health Organization (WHO) adult susceptibility bioassay indicated a relatively low level of dieldrin resistance (two-fold) in Ae. albopictus from Petaling Jaya, Selangor. However, PCR-RFLP and direct sequencing methods revealed the presence of the A302S mutation with the predomination of heterozygous genotype (40 out of 82 individuals), followed by the resistant genotype with 11 individuals. This study represents the first field evolved instance of A302S mutation in Malaysian insect species.
A multi-locus approach was used to examine the DNA sequences of 10 nominal species of blackfly in the Simulium subgenus Gomphostilbia (Diptera: Simuliidae) in Malaysia. Molecular data were acquired from partial DNA sequences of the mitochondria-encoded cytochrome c oxidase subunit I (COI), 12S rRNA and 16S rRNA genes, and the nuclear-encoded 18S rRNA and 28S rRNA genes. No single gene, nor the concatenated gene set, resolved all species or all relationships. However, all morphologically established species were supported by at least one gene. The multi-locus sequence analysis revealed two distinct evolutionary lineages, conforming to the morphotaxonomically recognized Simulium asakoae and Simulium ceylonicum species groups.
Species of the Simulium (Simulium) melanopus species-group in Sabah are taxonomically revised by examining type specimens of S. (S.) crassimanum S. (S.) laterale, and S. (S.) nigripilosum, all described from females by Edwards in 1933, and newly collected samples from the vicinity of Mt. Kinabalu. The females of these three species are redescribed, and their males and pupae are described for the first time based on adults reared from pupae. Simulium (S.) liewi Takaoka, 2007 and S. (S.) kinabaluense Smart & Clifford, 1969 are synonymized with S. (S.) crassimanum and S. (S.) laterale, respectively. Simulium (S.) cheedhangi Takaoka, Sofian-Azirun & Ya'cob, 2015 is newly recorded from Sabah. Two new related species, S. (S.) lardizabalae and S. (S.) timpohonense, are described from males reared from pupae. Keys to identify eight species of the S. melanopus species-group in Sabah are provided for females, males, pupae and mature larvae.
Surveys of pupae and larvae of black flies (Diptera: Simuliidae) were carried out in Thua Thien Hue Province of central Vietnam, and Lam Dong Province of southern Vietnam in 2014. A total of 26 species belonging to the genus Simulium were collected, consisting of eight known species, one newly recorded species, and 17 new species (of which three species of the subgenus Nevermannia were described in 2014). The remaining 14 new species (nine of the subgenus Gomphostilbia and five of the subgenus Simulium) are described here based on females, males, pupae and mature larvae. The total number of species of black flies in Vietnam is now 46. Keys to identify all 26 species recorded from the two provinces of Vietnam are given for females, males, pupae and mature larvae.
Simulium (Gomphostilbia) merapiense sp. nov. is described based on females, males, pupae, and mature larvae from Yagyakarta, Java, Indonesia. This new species is placed in the Simulium epistum species-group, and is characterized by the pupal gill with eight short filaments all arising at the same level from a short stalk, somewhat enlarged basal fenestra, entirely bare pupal head and thoracic integument, and small and short larval postgenal cleft. These characters rarely are found in the subgenus. Taxonomic notes are given to separate this new species from related species of the S. epistum species-group.
Perspicuous assessments of taxonomic boundaries and discovery of cryptic taxa are of paramount importance in interpreting ecological and evolutionary phenomena among black flies (Simuliidae) and combating associated vector-borne diseases. Simulium tani Takaoka & Davies is the largest and perhaps the most taxonomically challenging species complex of black flies in the Oriental Region. We use a DNA sequence-based method to delineate currently recognized chromosomal and morphological taxa in the S. tani complex on the Southeast Asian mainland and Taiwan, while elucidating their phylogenetic relationships. A molecular approach using multiple genes, coupled with morphological and chromosomal data, supported recognition of cytoform K and morphoform 'b' as valid species; indicated that S. xuandei, cytoform L, and morphoform 'a' contain possible cryptic species; and suggested that cytoform B is in the early stages of reproductive isolation whereas lineage sorting is incomplete in cytoforms A, C, and G.
A new simuliid species, Simulium kalimantanense sp. nov., is described on the basis of females, males, pupae, and mature larvae from East Kalimantan, Indonesia, and is assigned to the Simulium banauense species-group of Simulium (Gomphostilbia). This new species has close similarities to S alienigenum Takaoka from the Philippines, in many characters including the adult antennal color pattern and pupal gill with four long filaments arranged in two pairs each bearing a long stalk, but is distinguished from the latter in the female by the longer sensory vesicle and in the pupa by the gill with an elongate common basal stalk. Simulium kalimantanense sp. nov. is the first member of the S. banauense group in Borneo, and marks the most southerly distribution of the group. Keys to identify 19 Bornean species of the subgenus Gomphostilbia are provided.
We access the molecular diversity of the black fly Simulium nobile De Mejiere, using the universal cytochrome c oxidase subunit I (COI) barcoding gene, across its distributional range in Southeast Asia. Our phylogenetic analyses recovered three well-supported mitochondrial lineages of S. nobile, suggesting the presence of cryptic species. Lineage A is composed of a population from Sabah, East Malaysia (Borneo); lineage B represents the type population from Java, Indonesia; and lineage C includes populations from the mainland of Southeast Asia (Peninsular Malaysia and Thailand). The genetic variation of lineage C on the mainland is greater than that of lineages A and B on the islands of Borneo and Java. Our study highlights the value of a molecular approach in assessing species status of simuliids in geographically distinct regions.
The increasing attention on Vietnam as a biodiversity hotspot prompted an investigation of the potential for cryptic diversity in black flies, a group well known elsewhere for its high frequency of isomorphic species. We analyzed the banding structure of the larval polytene chromosomes in the Simulium tuberosum species group to probe for diversity beyond the morphological level. Among 272 larvae, 88 different chromosomal rearrangements, primarily paracentric inversions, were discovered in addition to 25 already known in the basic sequences of the group in Asia. Chromosomal diversity in Vietnam far exceeds that known for the group in Thailand, with only about 5% of the rearrangements shared between the two countries. Fifteen cytoforms and nine morphoforms were revealed among six nominal species in Vietnam. Chromosomal evidence, combined with available molecular and morphological evidence, conservatively suggests that at least five of the cytoforms are valid species, two of which require formal names. The total chromosomal rearrangements and species (15) now known from the group in Vietnam far exceed those of any other area of comparable size in the world, supporting the country's status as a biodiversity hotspot. Phylogenetic inference based on uniquely shared, derived chromosomal rearrangements supports the clustering of cytoforms into two primary lineages, the Simulium tani complex and the Southeast Asian Simulium tuberosum subgroup. Some of these taxa could be threatened by habitat destruction, given their restricted geographical distributions and the expanding human population of Vietnam.