Biomedical subjects
Publications and source records attributed to I Vythilingam.
The susceptibility of Culex sitiens to Japanese Encephalitis (JE) virus was examined in the laboratory. Cx. sitiens became infected with JE virus on day 8 and subsequently it is able to transmit the virus when it takes a blood meal. Both parts of the experiment were carried out using artificial membrane feeding technique.
Malaria vector surveys were carried out in 8 provinces in Lao PDR in 1999. The surveys were conducted in 4 provinces - Savannakhet, Champasak, Luang Perbang and Sayaboury in May and in another 4 provinces - Bolikhamsay, Sarvan, Sekong and Vientiane in December 1999. Bare leg collection were carried out indoors and outdoors from 6 pm to 5 am. All anopheline mosquitos were identified, dissected and the gut, gland and ovaries were examined. A total of 438 Anopheles mosquitos belonging to 19 species were obtained. Of these only 3 species were found to be infected with oocysts - An. maculatus, An. dirus and An. minimus. All these species were found biting both indoors and outdoors. An. aconitus was the predominant species obtained in the December collection but its vectorial status remains unknown.
Two formulations of lambda-cyhalothrin (EC-Emulsion concentrate and MC-Microencapsulated) were impregnated into bednets made of polyethylene and polyester. The nets were treated at a dosage of 15 mg/m2. For bioassay of insecticidal efficacy, female Anopheles maculatus and Aedes aegypti were exposed to the nets for two minutes and mortality was scored 24 hours later. The nets were also tested after repeated washings with water and with soap and water. Microencapsulated (2.5CS) formulation was more effective than emulsion concentrate (2.5EC) formulation on both net materials--polyethylene and polyester. Repeated washing with water and soap reduces the efficacy of all bednet treatment combinations. Microencapsulated formulation on polyethylene gave best results; it could sustain up to five washes with water and two with soap and water.
Dried Anopheles farauti mosquitos caught in Solomon Islands in 1990 were examined for malaria sporozoites by ELISA and nested polymerase chain reaction (PCR). Only heads and thoraces were used. Plasmodium genus-specific nested PCR amplifications were carried out on all samples. Of the 402 pools of mosquitos that were processed, 30 were positive for malaria. Nest 1 products of positive samples were subjected to further PCR amplifications with species-specific primers for P. falciparum and P. vivax. Twenty pools were positive for P. vivax by PCR while only 7 were positive by ELISA. For P. falciparum 2 pools were positive by both ELISA and PCR, and one of these was a pool which was positive for P. vivax by PCR and ELISA. Thus the sensitivity of PCR for P. vivax was 100% while the specificity was 96.7%. For P. falciparum the sensitivity and specificity were 100%. The PCR technique is highly sensitive and can be used on dried mosquitos which makes it a valuable tool for determining sporozoite rates of mosquitos, even in remote areas.
Accurate identification of filarial parasites in mosquitoes poses a major problem for the coordination of filariasis control programs. Traditional methods are tedious, and some are not specific enough to give satisfactory results. Amplification of specific gene sequences by primer-directed polymerase chain reaction (PCR) has been increasingly utilized as a diagnostic tool. However, current protocols for the extraction of parasite DNA from mosquito samples are tedious and could lead to failure of PCR amplification. We demonstrate that the use of Chelex is an efficient method for DNA extraction from mosquitoes and the parasite and that PCR amplification with primers specific for Brugia malayi yields a band of the expected size. The PCR products were transferred to a nylon membrane with Southern blotting, and a B. malayi-specific digoxigenin-labeled probe confirmed the sequence similarity of the PCR-amplified fragment and increased the sensitivity of the PCR assay. Use of this probe enabled us to detect PCR-amplified product from B. malayi even when a product was not visible on an ethidium bromide-stained agarose gel. This increased sensitivity allowed us to detect the parasite in the heads of mosquitoes.
A 2-yr study of Japanese encephalitis (JE) virus in Sepang District, Selangor, Malaysia, was carried out to identify the mosquito vectors and to determine their seasonal abundance, parity, and infection rates. In total, 81,889 mosquitoes belonging to 9 genera and > 50 species were identified from CDC trap collections augmented with dry ice during 1992 and 1993. Culex tritaeniorhynchus Giles and Culex gelidus Giles were the most abundant species, and both increased in numbers with increases in rainfall. Overall, 45 JE virus isolations were made from 7 species-Cx. tritaeniorhynchus (24), Cx. gelidus (12), Culex fuscocephala Theobald (2), Aedes butleri Theobald (4), Culex quinquefasciatus Say (1), Aedes lineatopennis Ludlow (1), and Aedes (Cancraedes) sp. (1). Based on elevated abundance and JE infection rates, Cx. tritaeniorhynchus appears to be the most important vector of JE virus in Sepang.
Two hundred forty nucleotides from the pre-membrane gene region of 12 Japanese encephalitis virus (JEV) strains isolated from three different regions of Malaysia from 1993 to 1994 were sequenced and compared with each other and with the JEV strains from different geographic areas in Asia. These 12 Malaysian isolates were classified into two genotypes. The four JEV strains isolated from Sarawak in 1994 and the four JEV strains isolated from Sepang, Selangor in 1993 were classified into one genotype that included earlier isolated strains from Malaysia (JE-827 from Sarawak in 1968 and WTP/70/22 from Kuala Lumpur in 1970). The four JEV strains from Ipoh, Perak in 1994 were classified into another genotype that included JEV strains isolated from northern Thailand and Cambodia. In an earlier report, 10 JEV strains from Sabak Bernam, Selangor in 1992 were classified into the largest genotype that included strains isolated in temperate regions such as Japan, China, and Taiwan. The data indicate that at least three genotypes of JEV have been circulating in Malaysia.
A study was carried out to determine the distribution of cockroaches in two different housing areas with central sewerage or individual septic tanks in an urban area in Kuala Lumpur, Malaysia. Six species of cockroaches were present and of these Periplaneta americana and Periplaneta brunnea were found in greater abundance. Seventeen species of bacteria were isolated and of these Escherichia coli and Klebsiella p. pneumoniae were isolated in greatest numbers. Control measures carried out using lambda cyhalothrin showed that there was no significant difference between treated and control sites.
Studies were carried out to observe the species composition of mosquitos and to determine the vectors responsible for the transmission of filariasis in Grik, Perak, Malaysia. A total of 2,155 mosquitos belonging to 7 genera and 30 species were collected. Anopheles donaldi comprised 24.1% of the collection. Twelve out of 519 An. donaldi were infected with L3 larvae of Brugia malayi. The peak biting time was around 23.00-24.00 hours. The infective bites per month ranged from 0 to 6.3.
Detection and isolation of Japanese encephalitis (JE) virus were attempted from female mosquitoes collected in Kampong Pasir Panjang, Sabak Bernam, Selangor, from May to November 1992. A total of 7,400 mosquitoes consisting of 12 species in 148 pools were processed and inoculated into Aedes albopictus clone C6/36 cell cultures. Of these, 26 pools showed the presence of viral antigens in the infected C6/36 cells by specific immunoperoxidase staining using an anti-JE virus polyclonal antibody. Presence of JE virus genome was confirmed in the infected culture fluid for 16 pools by using reverse transcriptase-polymerase chain reaction and JE virus-specific primers. Of these, 3 pools were from Culex tritaeniorhynchus, 4 from Culex vishnui, 3 from Culex bitaeniorhynchus, 2 from Culex sitiens, one from Aedes species, and 3 from Culex species. Isolation of JE virus from Cx. sitiens, Cx. bitaeniorhynchus, and Aedes sp. (Aedes butleri and Ae. albopictus) is reported for the first time in Malaysia.
The effect of permethrin impregnated bednets on Anopheles maculatus Theobald was studied in four villages in Pos Betau, Pahang, Malaysia from August 1990 to July 1992. Collections of mosquitos were carried out indoors and outdoors from 1900 to 0700 hours. All mosquitos were dissected for sporozoites and parity. In May 1991 two villages received bednets impregnated with permethrin at 0.5 g/m2 and two villages received placebo bednets. There was a significant difference in the sporozoite and parous rates between the treated and control villages after the distribution of bednets (p < 0.05). There was no significant difference in the bites/man/night of An. maculatus between the pre and post treatment periods in the control villages. However there was a significant difference in bites/man/night between pre and post treatment in the treated villages (p < 0.001).
Two hundred and forty nucleotides from the pre-M gene region of 10 Japanese encephalitis (JE) virus strains isolated in Malaysia in 1992 were sequenced and compared with the other JE virus strains from different geographic areas in Asia. Our JE virus strains belong to the largest genotypic group that includes strains isolated in temperate regions such as Japan, China, and Taiwan. Our Malaysian JE virus strains differed in 32 nucleotides (13.3%) from WTP/70/22 strain isolated from Malaysia in 1970, which belonged to another distinct genotypic group.
Five pyrethroids namely, lambda-cyhalothrin, permethrin, deltamethrin, cyfluthrin and alpha-cypermethrin were evaluated using adult male and female cockroaches, Periplaneta americana (L). The American cockroaches were exposed for 10 minutes to glass jars treated with different concentrations of the five pyrethroids. The cockroaches were susceptible to all five pyrethroids and the susceptibility based on LC50 and LC50 were observed to be in the following order: lambda-cyhalothrin > permethrin > deltamethrin > cyfluthrin > alpha-cypermethrin The results showed that lambda-cyhalothrin was the most effective and alpha-cypermethrin was the least effective against Periplaneta americana (L).
Mosquito collections were carried out for a period of one year from January to December 1992 in a pig farm in Sungai Pelek, Selangor, Malaysia. A total of 41,022 mosquitos belonging to 52 species and 20 genera were collected. Culex tritaeniorhynchus and Cx. gelidus, the important vectors, comprised 63% of all mosquitos collected. Both these species were collected in large numbers during the wet months of May and December. The other predominant species in that area were Cx. fuscocephala, Cx. quinquefasciatus, Cx. sitiens, Aedes butleri, and Armigeres subalbatus.
Isolation of Japanese encephalitis virus (JEV) from mosquitoes in Sabak Bernam, Selangor, Malaysia, was attempted. An aliquot of homogenate from each pool of mosquitoes, 50 per tube, was inoculated into Aedes albopictus clone C6/36 cells for virus isolation. Each cell culture was tested for the presence of viral antigen by immunoperoxidase staining using an anti-JEV polyclonal antibody. Out of 4 Culex sitiens mosquito pools, 2 pools were positive for JEV by cell culture. Presence of JEV genome in the cell cultures for Cx. sitiens was confirmed by using reverse transcriptase-polymerase chain reaction and JEV-specific primers. This is the first report on the isolation of JEV from Cx. sitiens.
The residual effectiveness of 0.005mg/ml of cyhalothrin applied to cattle was determined against three species of mosquitos: Anopheles maculatus Theobald. Anopheles dirus Peyton and Harrison Mansonia uniformis Theobald. Twenty-four hour post exposure mortality and the degree of successful blood engorgement were determined by exposing mosquitos for 10 minutes to cattle. Three replicated assays were conducted and mortality determined at 1, 2, 7, 14 and 21 days after each treatment. An initial mortality of 92-94% for An. dirus and Ma. uniformis and 79% for An. maculatus was obtained. Percentage mortality declined to 10%, 18% and 31% for An. maculatus, An. dirus and Ma. uniformis respectively on day 7 post application. On day 21 post application, percentage mortality was 2-3% for the three species of mosquitos.
Mosquito collections were carried out from May to June 1992 and from September to December 1992 in an area where a case of Japanese encephalitis was confirmed. A total of 40,072 mosquitoes belonging to 35 species and 8 genera were collected. The dominant species in that locality were Culex vishnui, Culex tritaeniorhynchus, Culex pseudovishnui, Culex gelidus, Aedes butleri, and Mansonia uniformis.
A field trial was carried out to study the effect of lambdacyhalothrin on Anopheles maculatus in trap huts in Jeram Kedah, Negeri Sembilan, Malaysia. Two trap huts were built, of which one was sprayed with lambdacyhalothrin at a dosage of 25 mg ai/m2 and the other served as control. Eight collectors commenced collecting mosquitos from 1900 to 2400 hours, two each indoors and outdoors. Bioassay was also carried out in the treated and control huts to determine susceptibility of adult mosquitos to lambdacyhalothrin. In the treated hut more mosquitos were present during the pre- spraying period. Lambdacyhalothrin gave a mortality of 100% against An. maculatus for 8 months.