Comparative identification of protein profiles and major allergens of saliva, salivary gland and whole body extracts of mosquito species in Thailand.

Allergic reactions to mosquito bites, such as generalized urticaria or severe local reactions are common problems worldwide. The diverse sources of allergen prepared from different mosquito body parts usage are a major obstacle to obtaining safe and effective tests and immunotherapy for mosquito bite allergy. Thus, the reactions are often not recognized and allergen immunotherapy is seldom used for severe reaction to mosquito bites. In a search for appropriate allergen sources, the protein profiles of saliva, salivary glands and whole body extracts were comparatively analyzed from 4 common mosquito species of Thailand and/or South East Asia; viz. Culex quinquefasciatus, Aedes aegypti, Aedes albopictus and a zoophilic strain, Anopheles minimus. The major allergens in the extracts which elicited specific IgE responses in the pooled sera of subjects allergic to mosquito bites were identified. It was concluded that mosquito saliva was the best source of allergens. Additionally, both species-specific and species-shared allergens of the 4 mosquito species were identified. The major saliva allergens having MWs of 36, 32 and 22 kDa were identified. The identificstion of major allergens should facilitate the production of specific recombinant allergens and contribute to improvement in the diagnosis and specific immunotherapy of Thai mosquito bite allergy patients.

Adherence to antimalarial drug therapy among vivax malaria patients in northern Thailand.

Vivax malaria is a significant cause of morbidity due to malaria in northern Thailand, accounting for approximately 50% of all malaria cases. The objective of this study was to determine the behavioural factors associated with adherence to the standard 14-day course of chloroquine and primaquine, prescribed from malaria clinics, among patients with vivax malaria. A retrospective study was conducted among 206 patients living in Muang and Mae Sa Riang districts of Mae Hon Son province in northern Thailand. Data on adherence and potential behavioural factors relating to adherence were collected using a structured interviewer-administered questionnaire and supplemented with qualitative data from focus-group interviews. The results indicated that 76.21% of the 206 patients with vivax malaria did not complete the medication course. The adherence of the patients was associated with knowledge scores of malaria (adjusted odds ratio [AOR]=2.2, 95% confidence interval [CI] 1.1-4.5) and accessing drug prescription scores (AOR=5.6, 95% CI 2.13-15.3). Therefore, further effort is needed to educate patients with vivax malaria on knowledge of malaria and its treatment with simple health messages and encourage them to adhere to their treatment.

Control of mosquito vectors of tropical infectious diseases: (2) pyrethroid susceptibility of Aedes aegypti ( l) collected from different sites in Thailand.

Four strains (SS, BS, A and B) of Aedes aegypti collected from different sites in Bangkok and at different times were examined for their pyrethroid susceptibility. Mosquito coils containing dl, d-T80-allethrin, d, d-T-prallethrin and methoxymethyl-tetrafluorobenzyl tetramethyl-cyclopropanecarboxylate (K-3050) with or without a synergist were tested by the 25 m3 semi-field test method. One strain (SS) was the most susceptible with KT50 values of about < 30 minutes for all mosquito coils, while the other three strains (BS, A and B) were found to be around 10 to 20 times more tolerant to pyrethroids than the SS strain. A similar tendency for the pyrethroid susceptibility of the four strains was obtained with tests by topical application method. In field efficacy tests, mosquito coils with d, d-T-prallethrin 0.20% plus N-(2-ethylhexyl)bicycle-[2,2,1]- hept-5- ene-2,3-dicarboxyimide as a synergist exhibited a repellent effect of about 85%, while those with K-3050 0.10% plus the synergist exhibited a greater repellent effect of about 90%. In contrast, the repellent effect of commercial dl, d-T80-allethrin 0.20% coils was as low as about 50%. The d, d-T-prallethrin and K-3050 coils with the synergist were confirmed to be highly effective in repelling Ae. aegypti.

Control of mosquito vectors of tropical infectious diseases: (1) bioefficacy of mosquito coils containing several pyrethroids and a synergist.

The bioefficacy of mosquito coils containing several pyrethroids were tested in a 25 m3 room against Culex pipiens quinquefasciatus, Aedes aegypti and Anopheles dirus. The test results were compared with tests against Culex pipiens pallens in Japan. Based on the KT50 values (the 50% knockdown time) of mosquito coils containing dl, d-T80-allethrin, d, d-T-prallethrin and methoxymethyl-tetrafluorobenzyl tetramethyl-cyclopropanecarboxylate (K-3050) at doses of 0.05-0.5% (w/w) with or without a synergist, the pyrethroid susceptibility of the four mosquito species was as follows: Cx. p. quinquefasciatus was several times more tolerant to pyrethroids than Cx. p. pallens, Ae. aegypti was a further several times more tolerant than Cx. p. quinquefasciatus, and An. dirus was more susceptible than Cx. p. pallens (KT50 value: about half of Cx. p. pallens). The order of their susceptibilities is common for pyrethroids. Mosquito coils containing d, d-T-prallethrin and K-3050 at doses of 0.05-0.2% (w/w) and N-(2-ethylhexyl)bicycle-[2,2,1]-hept-5-ene-2,3-dicarboxyimide as a synergist at a ratio of 2 times the active ingredient were highly effective against Ae. aegypti, the most important mosquito vector for dengue fever.

An empowerment program to enhance women's ability to prevent and control malaria in the community, Chiang Mai Province, Thailand.

Paulo Freire's theory was modified to empower a women's group in Chiang Mai Province, Thailand, to prevent and control malaria. This study conducted an intervention in Mueang Na Wan Village, Mueang Na Sub-district, Chiang Dao District, Chiang Mai Province, where 45 women were systematically recruited into the study cohort. Navail Village was selected as a control village because it resembled the intervention village. The empowerment program emphasized enhancement of malaria preventive levels, using insecticide-treated bed nets, self-esteem, and self confidence expectation to prevent and control malaria. Intensive training was conducted and activities performed among the women's group, with 10 participatory meetings in all. Data collection was conducted for the pre-test in month 1, and post-intervention in months 3, 6, 9, and 12. The qualitative methods used were focus-group discussions, non-participant observations, and in-depth interviews with housewives, their husbands, and youths at risk for malaria. The results showed that, post-intervention, there were significantly increased levels for malaria preventive behaviors, behaviors of using insecticide-treated nets, self-esteem, and self confidence expectations, in the intervention village compared with the control village. Insecticide-treated net usage and insecticide-treated net usage behaviors increased in the intervention village more than before and more than that in the control village. The women's group in the intervention village created the following plans, which were crucial to malaria prevention: (1) a family protection plan, (2) providing malaria education to community members, (3) a mosquito-control campaign, (4) scaling-up insecticide-impregnated bed nets, and (5) malaria control among foreign laborers. Finally, the empowered women's group performed sustainable activities. Between malaria-prevention activities, they conducted a joint program to raise income for their families.

Laboratory colonization of Mansonia mosquitoes with an emphasis on Ma. annulata and Ma. bonneae.

The present study records the first successful colonization of Mansonia annulata and describes colony maintenance with modification of rearing medium and host plants. Three species of Mansonia mosquitoes (Ma. uniformis, Ma. indiana and Ma annulifera) were successfully reared in ambient environments with adult emergence rates > 50%, while Ma. bonneae and Ma. dives yielded emergence rates > 30%. Colonization of Ma. annulata was modified and improved so that they were successfully raised to adult with emergence rates of 23%. Tube sedge, Lepironia articulata, was utilized as a host plant and peat swamp water was used as a rearing medium. Yeast and small lizard droppings were added daily to the larval medium to maintain microorganisms and pH in the infusion. However, identifying suitable culture medium remains an obstacle to establishing colonies of Ma. annulata, as the culture medium is difficult to mimic in the laboratory. Further study, focusing particularly on larval attachment substrates and rearing medium, is needed to develop a standardized and practical rearing technique for Mansonia mosquitoes.

Insecticide susceptibility of mosquitoes invading tsunami-affected areas of Thailand.

In order to control the mosquitoes invading tsunami-affected areas of Thailand, the insecticide susceptibility status of field larvae and mosquitoes (Anopheles sundaicus and Culex sitiens) was tested under laboratory conditions. Larval bioassay tests were conducted using the WHO standard method. Three larvicides: temephos, malathion, and plant extract (ethanolic extract of the Southeast Asian long pepper. Piper retrofractum Vahl), were used in the experiments. The results revealed that Cx. sitiens was more susceptible to temephos than malathion and the plant extract, with LC50 ranges of 0.0008-0.0014 mg/l, 0.0046-0.0078 mg/l, and 5.3180-10.1030 mg/l, respectively. Cx. quinquefasciatus showed greater tolerance to every tested larvicide than Cr sitiens. Adult bioassay tests using a WHO test kit and diagnostic doses of 5% malathion, 0.75% permethrin, 0.05% deltamethrin, and 4% DDT were also conducted. The results revealed that Cx. sitiens and An. sundaicus were susceptible to all tested insecticides. The LT50 of 5% malathion ranged between 25.7-26.0 minutes for Cx. sitiens, and 44.7 minutes for An. sundaicus. In addition, Cx. quinquefasciatus showed susceptibility to malathion, with LT10 of 19.7 minutes. However, it showed resistance to both pyrethroid insecticides, with LT50 of 33.1 minutes for 0.75% permethrin, and 19.6 minutes. for 0.05% deltamethrin; it showed low percentage mortality at 24 hour post-exposure, of 48 and 32%, respectively. In conclusion, every tested larvicide could be used for controlling Cx. sitiens larvae, even in brackish water, pyrethroid insecticides for adult Cx. sitiens and An. sundaicus, and malathion for all three species.

Determination of lead toxicity in Culex quinquefasciatus mosquitoes in the laboratory.

Laboratory investigations were carried out to study the effects of lead toxicity and lead uptake on Culex quinquefasciatus mosquitoes. Three different concentrations of lead nitrate were used in laboratory tests (0.05, 0.1, and 0.2 mg/l). An atomic absorption spectrometer (AAS) was used to the determine lead concentrations. The results showed that lead significantly reduced hatching, egg-production, and emergence rates, compared with the unexposed group (p < 0.05). The ratio of female to male offspring was 3.64:1, which was observed in the second generation, after the parents were exposed to 0.2 mg/l lead. No effects were observed on oviposition preference, larval weight, or larval deformation. The LC50 of lead against Cx. quinquefasciatus larvae within 24 hours was 0.18 mg/l. There was a significant increase in lead uptake related to increased lead exposure in mosquito larvae (p < 0.05). The bioconcentration factor (BCF) showed that the lead concentration in the larvae was 62 times greater than in the water. The lead concentration from parents to offspring reduced in the first and second generations (p < 0.05). There was no significant difference between female and male mosquitoes in lead concentration (p > 0.05).

First isolation of Japanese encephalitis from Culex quinquefasciatus in Thailand.

Isolation of Japanese encephalitis (JE) virus using C6/36 cell and immunofluorescence virus antigen detection techniques was attempted from female mosquitoes collected with CDC gravid traps in Samut Songkhram Province in the central region and in Phuket Province in southern Thailand, in 2003. One thousand and eighty female mosquitoes including 6 species of the Culicidae family (Culex quinquefasciatus, Cx. gelidus, Cx. tritaeniorhynchus, Cx. whitmorei, Cx. vishnui complex, Cx. s.g. culiciomyia) (pooled by specific specimen), were processed for virus isolation. Two pools of Cx. quinquefasciatus yielded a JE virus isolation. This represents the first report of JE virus isolation from Cx. quinquefasciatus in Thailand.

Malaria and enterobiasis among Karen Long-neck tribe in Mae Hong Son Province.

In Thailand, Mae Hong Son Province is highly endemic for malaria. Knowing this, the local Health Department has introduced a program to educate local residents about the risk factors, in particular the dangers and symptoms of malaria. This study was conducted to evaluate these efforts, by determining the number of malaria infections in a segment of the population, and also by testing for enterobiasis among a group of its children. Two villages in Mae Hong Son Province were chosen for this purpose with a combined population of about 300. Of these, 195 were screened for malaria. Two subjects were diagnosed positive for malaria by microscopy. One of these two villages was chosen to screen for Enterobius vermicularis infection in children as well. Out of 69 stool samples, five (7%) showed infection with E. vermicularis: three with a low number of eggs (1-50), and two with a high number of eggs (>100). Compared with infection rates in similar studies, the results of this study indicate that the Health Department's efforts are meeting with relative success. The low prevalence of infection indicates that the villagers are using the information they have received to help combat infection.

Parasitic infections among Karen in Kanchanaburi Province, western Thailand.

The purpose of this study was to determine the rate of intestinal parasitic and malarial infections during a period of low infection among the residents of remote Karen villages in Thailand. Fifty-five males and 64 females, aged 6 months to 70 years, were examined for malaria by thick blood smears using the Giemsa staining technique. Of the 119 subjects, 4 (3.36%) showed positive for malaria with vivax gametocytes. Results suggested that mass screening was not an effective way for diagnosing malaria. Stool samples were examined under a light microscope. The overall intestinal parasitic infection rates were 38.24% in 34 males, and 36.11% in 36 females. These were hookworm (17.14%), Ascaris lumbricoides (7.14%), Trichuris trichiura (1.43%), Strongyloides stercoralis (7.14%), Taenia spp (1.43%), Entamoeba histolytica (1.43%), Entamaeba coli (10.00%) and Giatdia lamblia (1.43%). The highest (55.55%) and lowest (16.66%) rates of infection were observed in age groups 0-5 and over 45, years respectively. In addition, A. lumbricoides and T. trichiura were found more frequently in children, while hookworms was found similarly in every age group. Results showed that the Karen living along the western border of Thailand possessed high rates of intestinal parasitic infections. Strict monitoring and control programs for these parasites should be implemented.

Comparative studies on the biology and filarial susceptibility of selected blood-feeding and autogenous Aedes togoi sub-colonies

Blood-feeding and autogenous sub-colonies were selected from a laboratory, stock colony of Aedes togoi, which was originally collected from Koh Nom Sao, Chanthaburi province, Southeast Thailand. Comparative biology and filarial susceptibility between the two sub-colonies (blood-feeding: F11, F13; autogeny: F38, F40) were investigated to evaluate their viability and vectorial capacity. The results of comparison on biology revealed intraspecific differences, i.e., the average egg deposition/gravid female (F11/F38; F13/F40), embryonation rate (F13/F40), hatchability rate (F11/F38; F13/F40), egg width (F11/F38), wing length of females (F13/F40), and wing length and width of males (F11/F38) in the blood-feeding sub-colony were significantly greater than that in the autogenous sub-colony; and egg length (F11/F38) and width (F13/F40), and mean longevity of adult females (F11/F38) and males (F13/F40) in the blood-feeding sub-colony were significantly less than that in the autogenous sub-colony. The results of comparison on filarial susceptibility demonstrated that both sub-colonies yielded similar susceptibilities to Brugia malayi [blood-feeding/autogeny = 56.7 percent (F11)/53.3 percent(F38), 60 percent(F13)/83.3 percent(F40)] and Dirofilaria immitis [blood-feeding/autogeny = 85.7 percent(F11)/75 percent(F38), 45 percent(F13)/29.4 percent(F40)], suggesting autogenous Ae. togoi sub-colony was an efficient laboratory vector in study of filariasis

Intraspecific hybridization of two karyotypic forms of Anopheles vagus (Diptera: Culicidae) and the related egg surface topography.

Hybridization tests of the two karyotypic forms (Form A and B) of laboratory-raised, isolines of Anopheles vagus, were conducted by induced copulation. The results of reciprocal- and back-crosses indicated that they were genetically compatible, providing viable progeny. Comparative egg morphometry and morphology, aided by scanning electron microscopy (SEM), revealed that the eggs of the two karyotypic forms were morphometrically and morphologically similar.

Intraspecific hybridization of Anopheles minimus (Diptera: Culicidae) species A and C in Thailand.

Hybridization tests of laboratory-raised, isolines of Anopheles minimus, species A and C were conducted by induced copulation. The three isolines were established based on three morphological variants of wild-caught, fully engorged females and two distinct types of metaphase chromosomes. They were An. minimus species A: V form (X1,Y1), M form (X2,Y1); species C: P form (X3,Y2). The results of reciprocal and back crosses indicated that the two morphologically variant forms of species A were genetically compatible, providing viable progeny and completely synaptic salivary gland polytene chromosomes, whereas they were genetically incompatible with species C and/or the P form. Hybrid progeny was only obtained from both forms of species A females x species C males, but asynaptic salivary gland polytene chromosomes on 3L and partial development of ovarian follicles in females were seen. Back crosses of F1 hybrid males with parental species A females provided viable progeny, while back crosses of F1 hybrid females with parental species C males provided progeny of low viability and adult males with abnormal spermatozoa, suggesting the partial reproductive isolation of An. minimus species A and C.