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Simulium dermatitis is an IgE-mediated skin reaction in animals and humans caused by the bites of black flies. Although Simulium nigrogilvum has been incriminated as the main human-biting black fly species in Thailand, information on its salivary allergens is lacking. Salivary gland extract of S. nigrogilvum females was subjected to sodium dodecylsulfate-polyacrylamide gel electrophoresis, and the separated components were applied onto nitrocellulose membranes for immunoblotting, which was performed by probing the protein blots with sera from 17 individuals who were allergic to the bites of S. nigrogilvum. IgE-reactive protein bands were characterized further by liquid chromatography-mass spectrometry (LC-MS/MS) analysis. Nine protein bands (79, 42, 32, 25, 24, 22, 15, 13, and 11 kDa) were recognized in the serum of the subjects. Four of the nine protein bands (32, 24, 15, and 11 kDa) showed IgE reactivity in all (100%) of the tested sera, and they were identified as salivary secreted antigen 5-related protein, salivary serine protease, erythema protein, and hypothetical secreted protein, respectively. Three other proteins, salivary serine protease (25 kDa), salivary D7 secreted protein (22 kDa), and hypothetical protein (13 kDa), reacted with > 50% of the sera. The relevance of the identified protein bands as allergens needs to be confirmed by using pure recombinant proteins, either in the in vivo skin prick test or in vitro detection of the specific IgE in the serum samples of allergic subjects. This will be useful for the rational design of component-resolved diagnosis and allergen immunotherapy for the allergy mediated by the bites of black flies.

Timely identification and treatment of malaria transmission "hot spots" is essential to achieve malaria elimination. Here we investigate the relevance of using an Anopheles salivary biomarker to estimate Plasmodium falciparum malaria exposure risk along the Thailand-Myanmar border to guide malaria control. Between May 2013 and December 2014, > 9,000 blood samples collected in a cluster randomized control trial were screened with serological assays to measure the antibody responses to Anopheles salivary antigen (gSG6-P1) and P. falciparum malaria antigens (circumsporozoite protein, merozoite surface protein 119 [MSP-1 19 ]). Plasmodium falciparum infections were monitored through passive and active case detection. Seroprevalence to gSG6-P1, MSP-1 19 , and CSP were 71.8% (95% Confidence interval [CI]: 70.9, 72.7), 68.6% (95% CI: 67.7, 69.5), and 8.6% (95% CI: 8.0, 9.2), respectively. Multivariate analysis showed that individuals with the highest Ab response to gSG6-P1 had six times the odds of being positive to CSP antigens ( P < 0.001) and two times the odds of P. falciparum infection compared with low gSG6-P1 responders ( P = 0.004). Spatial scan statistics revealed the presence of clusters of gSG6-P1 that partially overlapped P. falciparum infections. The gSG6-P1 salivary biomarker represents a good proxy for estimating P. falciparum malaria risk and could serve to implement hot spot-targeted vector control interventions to achieve malaria elimination.

Objective: To determine the clinical features of mosquito allergy in children and the ability of commercially available mosquito allergy tests to detect children with mosquito allergy in Thailand., Methods: Patients with mosquito allergy aged 1 month to 18 years were recruited. Demographic data, history of mosquito allergy (onset of the reaction, reaction type) and clinical features were recorded. A skin prick test using a commercially available whole body allergen extract from Culex pipiens was performed, and serum was tested for specific IgE antibodies to Aedes communis whole body extract., Results: A total of 50 patients with mosquito allergy were enrolled. The median age of enrolled children was 6.2 years with an average age of onset of 2 years [interquartile range (IQR) 1-6]. Half of the children were female. The most common skin lesion from mosquito allergy was erythematous papules (n = 45, 76.3%). The majority of children (58%) were in stage 3 (immediate and delayed type of reactions). One child (2%) was in the desensitization stage after 4.6 years of symptoms. The causative mosquito species could be identified only in 26 (52%) children: 16 (32%) children were positive for Aedes communis, 17 (34%) children were positive for Culex pipiens and 7 (14%) children were positive for both Aedes communis and Culex pipiens. Having positive IgE antibodies against Aedes communis was significantly more common in boys (n = 13, 48.1%) than girls (n = 3, 13%) (p < 0.01)., Conclusions: Immediate and delayed skin reaction is the most common manifestation in mosquito allergy children. Commercially available tests for mosquito allergy can detect only 30-50% of children with mosquito allergy.

Dengue viruses (DENV; family Flaviviridae, genus Flavivirus) are transmitted by Aedes aegypti mosquitoes and can cause dengue fever (DF), a relatively benign disease, or more severe dengue haemorrhagic fever (DHF). Arthropod saliva contains proteins delivered into the bite wound that can modulate the host haemostatic and immune responses to facilitate the intake of a blood meal. The potential effects on DENV infection of previous exposure to Ae. aegypti salivary proteins have not been investigated. We collected Ae. aegypti saliva, concentrated the proteins and fractionated them by nondenaturing polyacrylamide gel electrophoresis (PAGE). By the use of immunoblots, we analysed reactivity with the mosquito salivary proteins (MSP) of sera from 96 Thai children diagnosed with secondary DENV infections leading either to DF or DHF, or with no DENV infection, and found that different proportions of each patient group had serum antibodies reactive to specific Ae. aegypti salivary proteins. Our results suggest that prior exposure to MSP might play a role in the outcome of DENV infection in humans., (© 2011 Blackwell Publishing Ltd.)

Mosquito bite allergic reactions are due to sensitization to mosquito salivary proteins. The purpose of our study was to assess the clinical features of Thai patients with mosquito allergy, and to define the natural desensitization course. Seventy patients with mosquito allergy were enrolled. Most patients were female. Half of the patients had a personal history of atopy. The average age of onset of mosquito reaction was 5.7 years old (range 2-58). Four patients were in the desensitization phase in which clinical symptoms disappeared within 9.5 years (median). The common cutaneous lesions were erythematous papules (68.6%) and immediate wheals (67.1%). The most common area of involvement was the leg. Forty of 50 patients (80%) had positive skin prick testing to mosquito allergen (Culex quinquefasciatus) and 31 of 42 patients (73.8%) had positive immunoglobulin E antibodies against allergens in C. quinquefasciatus saliva proteins., (© 2010 Japanese Dermatological Association.)

Among cockroaches (CR) that live in people's homes, two species, i.e., German CR (Blattella germanica) and American CR (Periplaneta americana) predominate in temperate and tropical areas, respectively. CR is an important source of inhalant indoor allergens that sensitize atopic subjects to (localized) type I hypersensitivity or atopy including allergic rhinitis and atopic asthma. In Thailand the predominant CR species is P. americana. CR allergens are found throughout CR infested houses; the number found in kitchens correlates with the degree of CR infestation while sensitization and reactivation of the allergic morbidity are likely to occur in the living room and bedroom. Levels of the CR allergens in homes of CR allergic Thais, measured by using locally made quantification test kits, revealed that the highest levels occur in dust samples collected from the wooden houses of urban slums and in the cool and dry season. CR allergens are proteins that may be derived from any anatomical part of the insect at any developmental stage. The allergens may be also from CR secretions, excretions, body washes or frass. The proteins may be the insect structural proteins, enzymes or hormones. They may exist as dimers/multimers and/or in different isoforms. Exposure to CR allergens in infancy leads to allergic morbidity later in life. Clinical symptoms of CR allergy are usually more severe and prolonged than those caused by other indoor allergens. The mechanisms of acute and chronic airway inflammation and airway hyper-responsiveness (AHR) have been addressed including specific IgE- and non-IgE-mediated mechanisms, i.e., role of protease-activated receptor-2 (PAR2). Participation of various allergen activated-CD4+ T cells of different sublineages, i.e., Th2, Th17, Th22, Th9, Th25, Tregs/Th3 as well as invariant NKT cells, in asthma pathogenesis have been mentioned. The diagnosis of CR allergy and the allergy intervention by CR population control are also discussed.

Monitoring the levels of cockroach (CR) allergen in the environment has medical relevance as a clear dose response relationship between CR allergen exposure, sensitization and hospitalization has been reported. In this study, a cross-sectional survey of the levels of a major American cockroach (Periplaneta americana) allergen, i.e. Per a 9 (arginine kinase) in dust samples collected in various seasons throughout the year 2007 from 76 houses of CR allergic Thai patients in the Bangkok metropolitan area were determined. A monoclonal antibody-polyclonal antibody (MAb-PAb) based-sandwich ELISA was used. The MAb was specific to Per a 9 and the PAb was raised in a rabbit against the crude extract of P. americana. The detection limit of the assay was 122 pg of the allergen or 0.024 microg per gram of fine dust powder. The concentrations of Per a 9 were found to be highest during the winter months and lowest in summer. The levels of this CR allergen had a direct correlation with disease exacerbation; i.e. the majority of the CR allergic patients had their most severe clinical manifestations during winter. Moreover, the CR allergen levels were found to be higher in wood based-houses than in concrete houses.

In this study, native tropomyosin (Per a 7) of American cockroach (CR), Periplaneta americana, caught in Thailand was purified. Also, gene sequence encoding full length tropomyosin of the CR was PCR amplified by using degenerate primers designed from gene sequences coding for P. americana tropomyosin of the database (Per a 7.0101 and Per a 7.0102; accession no.Y14854 and AF106961, respectively). Amino acid sequence deduced from the nucleotide sequence encoding P. americana tropomyosin of this study (GenBank accession no. FJ976895) had 98.59% identity with the sequences of Per a 7.0101 and Per a 7.0102 and was 97.18% identical to the Bla g 7 sequence of German cockroach, Blatella germanica (accession no. AF260897). The native and recombinant tropomyosins (approximately 34 kDa) were used as antigens in sandwich ELISA for detecting specific IgE in serum samples of 14 consented allergic patients who were positive by skin test to crude CR extract in comparison to 5 individuals who were skin test negative. It was found that 8 (57%) and 6 (43%) of the CR allergic patients gave positive IgE binding results to the native and the recombinant proteins, respectively, while none of the non-allergic counterparts was positive. Results of immunoblotting conformed to the ELISA results. Tropomyosin extracted from the P. americana caught in Thailand has potential as standard P. americana allergen in clinical monitoring of the allergic Thai patients.

The American cockroach, Periplaneta americana, is the predominant cockroach (CR) species in Thailand and a major source of indoor allergens second only to the house dust mite. The incidence of CR allergy among allergic Thai patients is increasing but basic information on the allergenic components is scarce. In this study a recombinant troponin-T was produced by using cDNA prepared from RNA of the P. americana as a template and PCR primers designed from the P. americana troponin-T sequence deposited in the GenBank database. The recombinant protein (Mr approximately 50) did not bind to IgE in the sera of 18 skin prick test positive CR allergic patients. Rabbit polyclonal antiserum (PAb) against the recombinant troponin-T was produced and used in preparing an affinity column for the purification of native troponin-T from the crude P. americana extract (Mr approximately 47). IgE-immunoblotting revealed that the native protein bound to IgE in 3 of the 18 (16.7%) patients. Our results imply that native P. americana troponin-T, but not its recombinant counterpart, is a minor allergen among the CR allergic Thais.

During blood feeding, arthropod vectors inject saliva into vertebrate hosts. The saliva is biochemically complex and pharmacologically active, and may play an important role in pathogen transmission. To examine whether mosquito saliva could elicit humoral immune response in humans under natural conditions, we have collected sera from malaria patients, healthy villagers, and people from a non-malarious region in Thailand. Here we have demonstrated that anti-Anopheles salivary protein antibodies occurred predominantly in patients with acute Plasmodium falciparum or P. vivax malaria, whereas people from a non-malarious area had no such antibodies. Besides, antibody levels against mosquito salivary proteins in malaria patients were highly variable, which may be related to the levels of mosquito exposure. Despite variability, patients' sera with high IgG titers consistently detected several proteins in Anopheles dirus salivary gland protein extracts. Immunohistochemical staining of Anopheles salivary glands with human sera showed that the salivary gland-specific IgGs reacted strongly with the median lobe. Comparison using Anopheles and Aedes salivary proteins suggests that the anti-salivary protein antibodies detected in malaria patients were Anopheles-specific, consistent with the major malaria vector status of An. dirus in this area.

Unlabelled: Recently, cockroaches have been established as the second most Important allergen, producing allergic diseases, especially in low socioeconomic populations. In Thailand, about 44-61% of atopic patients were positive to cockroach extract by a skin-prick test. This study examined cockroach allergen levels in relation to cockroach species and allergic diseases in the houses of cockroach-sensitive patients. Sixty households of allergic patients in the Bangkok metropolitan area were surveyed using open- and closed-ended questionnaires. Cockroaches were collected using commercial cockroach traps, while dust samples were obtained from the bedrooms, kitchens and living rooms of the houses using a vacuum cleaner. The cockroaches were counted and their species Identified. The levels of cockroach allergens were determined by specific monoclonal antibodies using a monoclonal antibody-polyclonal antibody based sandwich ELISA kit. Six cockroach species were Identified: Periplaneta americana (American cockroach, 72.15%), Supella longlpalpa (2.75%, found in only one house), Periplaneta brunnea (0.78%), Periplaneta australaslae (0.78%), Neostylopyga rhombifolla (0.78%), Blattella germanica (German cockroach, 0.39%) and nymphs (22.35%). Allergens of the predominant species, P. americana, were detectable in all homes studied, with the highest levels in the kitchen areas. The range of allergen levels in house dust varied from 0.40-162.00 microg per g of dust. The median and mean allergen levels in kitchen dust were 59.16 microg and 62.80 microg per g of dust, respectively, while the median allergen level in bedroom dust was only 15.90 microg per g of dust. The German cockroach allergen (Bla g 2) was undetectable in any of the houses., In Conclusion: P. americana was the most common cockroach and may be the species causing allergic diseases, especially asthma, in Thailand, which differs from the USA and Europe

One hundred and twenty patients, aged 3-15 years, attending the Allergy Clinic, Thammasat University Hospital, with allergic rhinitis, asthma or both conditions were studied. Standardized skin prick tests were performed with 15 common aeroallergens. The subjects were 85 boys and 35 girls with the mean age of 8.1 years (SD = 3.4 years). There were 63 cases (53%), 22 cases (18%) and 35 cases (29%) diagnosed with allergic rhinitis only, asthma only and both conditions, respectively. The prevalence of asthma among the allergic rhinitis patients was 36 percent whereas the prevalence of allergic rhinitis among the asthmatic patients was 61 percent. The sensitization patterns for major aeroallergens among the three groups were similar with the most common sensitized allergens as D. pteronyssinus, followed by D. farinae, American cockroach, Kopok and German cockroach, respectively. Among the atopic patients, a greater number of allergen reactivity was significantly associated with the increased risk of having their coexistence of allergic rhinitis and asthma (p = 0.0046; score test for trend of odds ratio).