A prospective field study was carried out to investigate any preferential differences of gravid female Aedes mosquitoes in ovipositing their eggs in man-made containers placed in different environmental conditions. The findings of this study show that gravid female Aedes mosquitoes preferred to breed in containers found in the outdoor garden than those placed on the patio and or inside the house. The findings also show that if the breeding habitats in the garden were removed, they would favorably use the breeding habitats found on the patio or inside the house as alternatives. An incidental interesting finding in this study shows that ultra-low volume fogging of insecticides using the vehicle-mounted equipment carried out outside the house may promote the gravid female Aedes mosquitoes to enter the house to breed.
A study was made of the oviposit behavior of gravid female Aedes mosquitos in man-made habitats under field conditions. The study showed that the gravid female Aedes mosquitos preferred containers with relatively easy access but not too open to external environmental influence. The dark surface of the containers served as the initial and long-range attractant to the breeding sites. Volatile chemicals generated by the decaying vegetation in the container may serve as a close-range attractant. Finally, the water quality and the quantity of 'food' derived from decaying vegetative matter in the water determined the amount of eggs deposited in each container. The study confirmed previous findings that each gravid female Aedes mosquito had the tendency to lay her eggs in more than one container. However, the results of the study suggests that under favorable conditions, each gravid female Aedes mosquito could be encouraged to lay all her eggs in a single breeding site.
Dengue and dengue haemorrhagic fever are common and serious arboviral diseases endemic in a number of countries situated in both the tropical and subtropical belts.
Since a safe and effective mass vaccination program against dengue fever is not presently available, a good way to prevent and control dengue outbreaks depends mainly on controlling the mosquito vectors. Aedes aegypti mosquito populations can be monitored and reduced by using ovitraps baited with organic infusions. A series of laboratory experiments were conducted which demonstrated that the bacteria in bamboo leaf infusion produce volatile attractants and contact chemical stimulants attractive to the female mosquitoes. The results showed that the female mosquitoes laid most of their eggs (59.9 ± 8.1 vs 2.9 ± 2.8 eggs, P<0.001) in bamboo leaf infusions when compared to distilled water. When the fresh infusion was filtered with a 0.45 μm filter membrane, the female mosquitoes laid significantly more eggs (64.1 ± 6.6 vs 4.9 ± 2.6 eggs, P<0.001) in unfiltered infusion. However when a 0.8 μm filter membrane was used, the female laid significantly more eggs (62.0 ± 4.3 vs 10.1 ± 7.8 eggs, P<0.001) in filtrate compared to a solution containing the residue. We also found that a mixture of bacteria isolated from bamboo leaf infusion serve as potent oviposition stimulants for gravid Aedes mosquitoes. Aedes aegypti laid significantly more eggs (63.3 ± 6.5 vs 3.1 ± 2.4 eggs, P<0.001) in bacteria suspension compared to sterile R2A medium. Our results suggest microbial activity has a role in the production of odorants that mediate the oviposition response of gravid mosquitoes.
Artificial feeding of mosquitoes with blood meal is an important technique in the studies of mosquito feeding. Owing to the difficulty in obtaining suitable artificial membranes for mosquito feeding from other sources, several easily obtainable membranes in Malaysia were tested for their suitability as a replacement. Skin of chicken, fish, and salted sausage were obtained and tested against cattle skin membrane as a control. The results showed that cattle skin is still the most favorable membrane to be used, with full engorgement rate of around 57% using fresh human blood. However, processed chicken skin was shown having potential for further testing since with feeding using human blood kept overnight at 4 degrees C, an engorgement rate of 50% was obtained.
Nepenthes pitcher plants are colonized by a variety of specialized arthropods. As Aedes mosquitoes are container breeders, Nepenthes pitchers are a potential candidate oviposition site for vector species, such as Aedes aegypti (L.) and Aedes albopictus (Skuse). However, Aedes spp. are not commonly encountered in Nepenthes pitchers, and the environment inside the pitchers of some species is lethal to them. One exception is Nepenthes ampullaria Jack, whose pitchers are known to be colonized by Ae. albopictus on very rare occasions. Given that Ae. albopictus larvae can survive in N. ampullaria pitcher fluids, we sought to determine why pitcher colonization is rare, testing the hypothesis that gravid Aedes mosquitoes are deterred from ovipositing into container habitats that have similar characteristics to N. ampullaria pitchers. Using plastic ovitraps of different sizes, colors, and with different types of fluids (based on the characteristics of N. ampullaria pitchers), we compared oviposition rates by Aedes mosquitoes in urban and rural areas within the geographical range of N. ampullaria near Kuala Lumpur, Malaysia. Ovitraps that were black and large (>250-ml capacity) accumulated significantly more eggs than ovitraps that were smaller, or green in color. In terms of size and color, small, green ovitraps are analogous to N. ampullaria pitchers, indicating that these pitchers are not particularly attractive to gravid Ae. albopictus. Although Aedes spp. are capable of colonizing N. ampullaria pitchers, the pitchers are relatively unattractive to gravid females and do not represent a significant habitat for larvae of dengue vectors at present.
Zika virus (ZIKV) has emerged since 2013 as a significant global human health threat following outbreaks in the Pacific Islands and rapid spread throughout South and Central America. Severe congenital and neurological sequelae have been linked to ZIKV infections. Assessing the ability of common mosquito species to transmit ZIKV and characterizing variation in mosquito transmission of different ZIKV strains is important for estimating regional outbreak potential and for prioritizing local mosquito control strategies for Aedes and Culex species. In this study, we evaluated the laboratory vector competence of Aedes aegypti, Culex quinquefasciatus, and Culex tarsalis that originated in areas of California where ZIKV cases in travelers since 2015 were frequent. We compared infection, dissemination, and transmission rates by measuring ZIKV RNA levels in cohorts of mosquitoes that ingested blood meals from type I interferon-deficient mice infected with either a Puerto Rican ZIKV strain from 2015 (PR15), a Brazilian ZIKV strain from 2015 (BR15), or an ancestral Asian-lineage Malaysian ZIKV strain from 1966 (MA66). With PR15, Cx. quinquefasciatus was refractory to infection (0%, N = 42) and Cx. tarsalis was infected at 4% (N = 46). No ZIKV RNA was detected in saliva from either Culex species 14 or 21 days post feeding (dpf). In contrast, Ae. aegypti developed infection rates of 85% (PR15; N = 46), 90% (BR15; N = 20), and 81% (MA66; N = 85) 14 or 15 dpf. Although MA66-infected Ae. aegypti showed higher levels of ZIKV RNA in mosquito bodies and legs, transmission rates were not significantly different across virus strains (P = 0.13, Fisher's exact test). To confirm infectivity and measure the transmitted ZIKV dose, we enumerated infectious ZIKV in Ae. aegypti saliva using Vero cell plaque assays. The expectorated plaque forming units PFU varied by viral strain: MA66-infected expectorated 13±4 PFU (mean±SE, N = 13) compared to 29±6 PFU for PR15-infected (N = 13) and 35±8 PFU for BR15-infected (N = 6; ANOVA, df = 2, F = 3.8, P = 0.035). These laboratory vector competence results support an emerging consensus that Cx. tarsalis and Cx. quinquefasciatus are not vectors of ZIKV. These results also indicate that Ae. aegypti from California are efficient laboratory vectors of ancestral and contemporary Asian lineage ZIKV.
Colonies of three strains of Aëdes aegypti resistant to DDT were obtained from Trinidad, Haiti and Malaya and reared beside a normal colony. From their relative resistance to a series of compounds analogous to DDT, characteristic resistance "spectra" were obtained. The two colonies from the West Indies showed a similar type of resistance, rather different from the (smaller) resistance of the Malay strain. No resistance to methoxychlor or dieldrin was found.
Aedes albopictus (Skuse) is an Asian mosquito that recently has colonized North America via used tire transport. Temperate Ae. albopictus populations overwinter as diapausing eggs, but tropical populations cannot diapause. Eggs of tropical (SABAH) and temperate (INDY) Ae. albopictus were obtained in diapause-inducing conditions and placed inside a scrap tire to monitor overwintering survival in northern Indiana during the winters of 1989-1990 and 1990-1991. Diapause eggs of Ae. triseriatus (Say), a native North American mosquito, were included for comparison. Tropical Ae. albopictus from Malaysia did not survive either winter. Temperate Ae. albopictus from Indianapolis, IN, did not survive the winter of 1989-1990, but 78% survived the winter of 1990-1991. In contrast, 92 and 96% of Ae. triseriatus survived the winters of 1989-1990 and 1990-1991, respectively. Neither mean temperature nor absolute minimum temperature (a winter's lowest temperature) accurately predicted Ae. albopictus overwintering survivorship in the field. The possible effect of snow and other insulating materials on the overwintering survivorship of Ae. albopictus eggs is discussed.
The mosquito, Aedes albopictus, has recently become established in a number of cities throughout the United States. An initial survey of allozyme and genotypic frequencies in U.S. populations (Black et al., 1988) revealed an extensive amount of local differentiation of populations and suggested that much genetic drift may have accompanied colonization. A study of gene flow was initiated in native habitats of Ae. albopictus in Malaysia to determine if the result observed in the U.S. was a consequence of colonization or simply followed the natural breeding structure of the species. Allelic and genotypic frequencies were monitored at ten enzymatic loci in 11 populations from peninsular Malaysia and Borneo. Multiple populations were sampled within the districts of Kuala Lumpur and Kuala Trengganu. Peninsular Malaysian and Borneo populations were strongly genetically differentiated. Allele frequencies were significantly different among and within districts in both regions. Variance in allele frequencies among all collections was partitioned into the variance among regions, districts within regions and collections within districts. Almost all of the variance within regions was attributable to local differentiation suggesting that genetic drift is an important component of the natural breeding structure of this species. This indicates that the large amounts of local differentiation found in U.S. populations was not a consequence of recent colonization.
Urbanization has caused an increase in favorable habitats for Aedes aegypti (Diptera: Culicidae), given their ability to reproduce in small and often non-degradable artificial water-containers. While much work has been done on Ae. aegypti biology and ecology in urban landscapes, the role of shading on immature stages as an independent factor from temperature, and any possible interactions between these factors, remains unexamined. We assessed how temperature and shading affected egg hatch-rate, larval/pupal mortality, and larval development to adult stage under different factorial temperature (28; 31; 34; 37; 40° C) and shade (0%, 3,100 lux; 40%, 1,860 lux; 75%, 775 lux; 100%, 0 lux) regimes. Hatch-rate was significantly lower at 37° C (57 %), and no eggs hatched at 40° C. There was no significant effect caused by shading on hatchability. Larval and pupal mortality at 37° C was significantly higher (35%) compared to lower temperature groups, while the effects of shading were emergent at low temperatures. Developmental times from hatching to adult emergence were significantly reduced with increasing temperatures and with greater light exposures. The eco-physiological response of Ae. aegypti larvae to temperature and light regimes suggest a photosensitivity previously unstudied in this species.
The mosquito Aedes albopictus is indigenous to Southeast Asian and is a vector for arbovirus diseases. Studies examining the population genetics structure of A. albopictus have been conducted worldwide; however, there are no documented reports on the population genetic structure of A. albopictus in Malaysia, particularly in Penang. We examined the population genetics of A. albopictus based on a 445-base pair segment of the mitochondrial DNA cytochrome oxidase 1 gene among 77 individuals from 9 localities representing 4 regions (Seberang Perai Utara, Seberang Perai Tengah, Northeast, and Southwest) of Penang. A total of 37 haplotypes were detected, including 28 unique haplotypes. The other 9 haplotypes were shared among various populations. These shared haplotypes reflect the weak population genetic structure of A. albopictus. The phylogenetic tree showed a low bootstrap value with no genetic structure, which was supported by minimum spanning network analysis. Analysis of mismatch distribution showed poor fit of equilibrium distribution. The genetic distance showed low genetic variation, while pairwise FST values showed no significant difference between all regions in Penang except for some localities. High haplotype diversity and low nucleotide diversity was observed for cytochrome oxidase 1 mtDNA. We conclude that there is no population genetic structure of A. albopictus mosquitoes in the Penang area.
The state of Johore suffered a massive flood disaster from 19th December 2006 to 1st January and from 12th January to 19th February 2007. The possible upsurge of dengue was of foremost concern and led to efforts in increasing control activities. Anyone with history of high fever with at least two symptoms of severe headache, pain behind the eyes, muscles and joint paint, rashes and petechiae were notified as dengue. Active and passive case finding was initiated at all 371 evacuation centres as well through health facilities and hospitals through an active surveillance system. Presumptive larval survey was also carried together with control activities by 46 health teams. Data were collected using the format ‘Aktiviti harian kawalan denggi di kawasan pos banjir- Lampiran E‘ and ‘Laporan aktiviti harian kawalan denggi di pusat pemindahan banjir – Lampiran D2’. Dengue serology and blood film for malaria was sent for as well as vector species identification. A total of 594 dengue cases were reported for the period of 19th December 2006 till 19th February 2007, which was an increase in comparison to the 5-year median but less than that reported in year 2006. However only 14 (2.3%) cases were from flood affected areas. During the flood phase, a total of 5,929 inspections were carried out at the evacuation centres with Aedes Index (AI) of 1.86%, while the post flood period showed a lower index. However Breteau Index (BI) and Container Index (CI) were higher. Preventive fogging were carried out at the evacuation centres using adulticides, thermal fogging was carried out at 21,959 premises (40.04% of inspected premises) and 350.6 L adulticides (malathion, fenitrothion and permethrin) were used. Dengue was expected to increase during flood as a result of increase Aedes potential breeding sites. However with intensive and integrated control activities, Johore was able to minimize the impact of flood for vector-borne diseases as seen from the low cases reported in flood related areas. A special guidelines for surveillance and control was developed during this flood as a reference for future occurrences.
World Health Organization (WHO) estimated over 100 million dengue infections to happen annually worldwide involving more than 2.5 billion people. Temephos or abate is a larvicide that has been used in vector control to eradicate mosquito larvae. Though practically low risk, there had been resistance problem reported with continuous use. This study seeks to find an effective and safer alternative to abate by assessing the use of ethanolic extract of Murraya koenigii leaves as larvicidal agent against Aedes aegypti. M.koenigii leaves were macerated for 3 days with absolute ethanol and evaporated using rotary vapor to produce the crude extract. The crude extract was subjected to phytochemical screening using standard qualitative method. For bioassay, the crude
extract underwent a serial dilution to produce 3 concentrations of 100 ppm (C1), 50 ppm (C2) and 10 ppm (C3) with abate and absolute ethanol as negative and positive control respectively. Bioassay for larvicidal effect was conducted in accordance to WHO standard method. Phytochemical screening of ethanolic extract of M. koenigii leaves revealed the presence of alkaloid, steroid and saponin. The bioassay shows that after 24 hours, the mortality rate of C1, C2 and C3 larvae were 100%, 38% and 0% and when further extended to 48 hours, the rate increased to 100% and 46% for C2 and C3 respectively. The LC50 and LC99 post 24 hours were 54.489 ppm and 93.961 ppm respectively whilst at post 48 hours, the LC50 and LC99 were 10.263 ppm and 16.176 ppm respectively. The results show that up to 48 hours duration of exposure, the mortality
rate increase whilst the lethal concentration (LC50 and LC99) decreases. Upon examination on larvae deformities at post 24 and 48 hours, all test concentrations and negative control exhibit normal morphology. Positive control, however, exhibit deformities characterized by twisted and fragmented insides. When statistically analyzed, C1 larvicidal activity was proven comparable with abate at 24 hours while C2 needed 48 hours exposure to be on par. Based on the results, it could be argued that the ethanolic extract of M.koenigii leaves does hold promising value to be further developed as larvicidal.
Adaptation to environmental conditions within the native range of exotic species can condition the invasion success of these species outside their range. The striking success of the Asian tiger mosquito, Aedes albopictus, to invade temperate regions has been attributed to the winter survival of diapause eggs in cold environments. In this study, we evaluate genetic polymorphisms (SNPs) and wing morphometric variation among three biogeographical regions of the native range of A. albopictus. Reconstructed demographic histories of populations show an initial expansion in Southeast Asia and suggest that marine regression during late Pleistocene and climate warming after the last glacial period favored expansion of populations in southern and northern regions, respectively. Searching for genomic signatures of selection, we identified significantly differentiated SNPs among which several are located in or within 20 kb distance from candidate genes for cold adaptation. These genes involve cellular and metabolic processes and several of them have been shown to be differentially expressed under diapausing conditions. The three biogeographical regions also differ for wing size and shape, and wing size increases with latitude supporting Bergmann's rule. Adaptive genetic and morphometric variation observed along the climatic gradient of A. albopictus native range suggests that colonization of northern latitudes promoted adaptation to cold environments prior to its worldwide invasion.
In a previous study, we have reported the detection and isolation of dengue virus in Brunei (Osman, O., Fong, M.Y., Devi, S., 2007. A preliminary study of dengue infection in Brunei. JJID 60 (4), 205-208). DEN-2 was the predominant serotype followed by DEN-1. The full genomic sequences of 3 DEN-2 viruses isolated during the 2005-2006 dengue incident in Brunei were determined. Twenty-five primer sets were designed to amplify contiguous overlapping fragments of approximately 500-600 base pairs spanning the entire sequence of the viral genome. The amplified PCR products were sent for sequencing and their nucleotides and the deduced amino acids were determined. All three DEN-2 virus isolated were clustered in the Cosmopolitan genotype of the DEN-2 classification by Twiddy et al. This work constitutes the first complete genetic characterization of three Brunei DEN-2 virus strains.
The purpose of this study was to examine the extent of dengue infection in Brunei and to determine the predominant serotype circulating in the country. The study generated useful epidemiological data on dengue infection in Brunei. A total of 271 samples from patients suspected of having dengue infections were selected and analyzed. All patients were seen in clinics and hospitals in Brunei. The samples were collected from April 2005 to April 2006 and transported to the WHO Collaborating Centre for Arbovirus Reference and Research, University of Malaya, Malaysia. The following tests were used to achieve the objectives: in-house IgM-capture enzyme-linked immunosorbent assay, virus isolation in mosquito albopictus cell line (C6/36), and viral RNA detection and serotyping by reverse transcriptase-polymerase chain reaction (RT-PCR). The results show that 45 people were positive for dengue-specific IgM (27 males and 18 females), while RT-PCR detected dengue viral RNA in 12 patients, 3 identified as DEN-1 and 9 as DEN-2. Dengue virus was isolated from 6 patients using the C6/36 cell line; 3 were DEN-2 isolates and 3 were DEN-1 isolates. These data show that dengue virus is circulating in Brunei and the predominant infecting serotype for that period was DEN-2 followed by DEN-1. This study is the first to report the detection and isolation of dengue virus from Brunei using RT-PCR and culture in the C6/36 albopictus mosquito cell line.
The genetics of glucosephosphate isomerase (E.C. 5.3.1.9) in two strains (Malaysian and Taiwan) of Aedes togoi is reported. Three electrophoretic phenotypes were presented in both sexes. The zymogram patterns were identical in both strains of A. togoi. The phenotypes were governed by a pair of codominant alleles. The allele frequency of the slow-moving band was 0.63 in the Malaysian strain adn was 0,86 and 0.82 in F161 and F169 generations, respectively, of the Taiwan strain. The sample studied was in good accord with Hardy-Weinberg expectation.
Vector-borne diseases are still rife because of the re-emergence of diseases transmitted by mosquitoes. The objective of this paper is to evaluate the larvicidal efficacy of crude leaf extract of Mangifera indica, Gluta renghas, and Melanochyla fasciculiflora against vector of dengue hemorrhagic fever, Aedes aegypti. These plant species are endemic species and widely distributed in Malaysian forests. Leaves of Ma. indica, G. renghas and M. fascculiflora were collected from Teluk Bahang National Park, Penang Malaysia. Fractions of leaves were segregated, air-dried, powdered and extracted using Soxhlet with methanol. The solvent was removed by using rotary evaporator to obtain the crude extract. Using WHO standard larval bioassay test method, third instar larvae of Aedes aegypti were exposed to concentration ranging from 200- 4500 ppm of methanol extract for all plant species. Larval mortality was observed after 24 hours exposure. The highest susceptibility and toxicity was recorded by Mangifera indica with the lowest concentration at 800 ppm followed by M. fasciculiflora and G. renghas. This indicates that crude plant extract is very effective in killing Ae. aegypti mosquitoes. This finding may lead to new low cost alternative, environmentally friendly method for mosquito control programs. To our knowledge, this is the first report on larvicidal bioefficacy from endemic Malaysian plants.