Immunoglobulin G1 subclass responses can be used to detect specific allergy to the house dust mites Dermatophagoides farinae and Dermatophagoides pteronyssinus in atopic dogs.

BACKGROUND: In dogs with atopic dermatitis, intradermal testing (IDT) or allergen specific IgE serological testing are routinely employed to identify causative allergens. These allergens can then be used for allergen-specific immunotherapy and allergy management. The clinical relevance of this testing is affected by the source of allergen, and other biomarkers that are more related to specific allergens still need to be identified. The aim of this study was to investigate levels of specific IgE, total IgG, and IgG1 and IgG2 subclasses against the local house dust mites (HDM) Dermatophagoides farinae (DF) and D. pteronyssinus (DP) as biomarkers by using in-house ELISAs in healthy (n = 33) and atopic dogs (AD) (n = 44) that were either positive or negative by IDT to HDM. RESULTS: Being over 3 years of age was a risk factor for AD (Odds Ratio (OD) = 4.10, 95% Confidence interval (CI) 1.57-10.75, p = 0.0049), but there was no relation to IDT outcomes (OR = 0.9091, 95% CI 0.22-3.74, p = 1.00). High levels of all antibody isotypes (IgE, IgG, IgG1 and IgG2) against HDM were found in aged healthy dogs (> 3 years old). In AD, HDM-IgE and IgG1 levels were higher in dogs that were IDT positive to HDM than in IDT negative animals. Levels of IgE and IgG1 could be used to distinguish the specific allergens, whereas total IgG and IgG2 levels were not different between IDT-positive and IDT-negative AD. By the receiver operating characteristic curve at a false-positive rate = 0.10, both IgE and IgG1 showed better sensitivity than IgG and IgG2. Similar to IgE, serum IgG1 concentration was also relevant to IDT outcomes. CONCLUSIONS: Our in-house ELISAs coated with local HDM were useful for evaluating antibody levels, and we propose use of the HDM-specific IgG1 subclass as a biomarker to detect HDM specific allergens in AD, potentially together with an IgE based platform.

Genetic variation of NADH dehydrogenase subunit 1 (nad1) mitochondrial gene sequence in adult Necator americanus hookworms recovered from a female patient in Thailand.

Two female and one male adult hookworms were recovered from a female patient in Thailand. Based on gross and microscopic morphology, the three hookworms are members of Necator americanus. Phylogenetic reconstruction based on partial NADH dehydrogenase subunit 1 (nad1) mitochondrial gene sequences shows that these hookworms belong to the same genetic lineage as N. americanus adult worm from Zhejiang, China. The male and female hookworms were genetically distinct, belonging to two different nad1-haplotypes. This is the first report targeting the nad1 gene on the identification and genetic characterization of the human hookworms originated from infected patient. The nad1 gene marker is useful for species and higher taxa differentiation of hookworms.

Cytochrome c oxidase subunit I haplotype reveals high genetic diversity of Angiostrongylus malaysiensis (Nematoda: Angiostrongylidae).

The rat lungworm Angiostrongylus malaysiensis is a metastrongyloid nematode parasite. It has been reported in Malaysia, Thailand, Laos, Myanmar, Indonesia and Japan. In this study, A. malaysiensis adult worms recovered from the lungs of wild rats in different geographical regions/provinces in Thailand were used to determine their haplotype by means of the mitochondrial partial cytochrome c oxidase subunit I (COI) gene sequence. The results revealed high COI haplotype diversity of A. malaysiensis from Thailand. The geographical isolates of A. malaysiensis from Thailand and other countries formed a monophyletic clade distinct from the closely related A. cantonensis. In the present study, five new haplotypes were identified in addition to the four haplotypes reported in the literature. Phylogenetic analysis revealed that four of these five new haplotypes - one from Mae Hong Song (northern region), two from Tak (western region) and one from Phang Nga (southern region) - formed a distinct clade with those from Phatthalung (southern region) and Malaysia. The haplotype from Malaysia was identical to that of Phatthalung (haplotype AM1). In general, the COI sequences did not differentiate unambiguously the various geographical isolates of A. malaysiensis. This study has confirmed the presence of high COI genetic diversity in various geographical isolates of A. malaysiensis. The COI gene sequence will be suitable for studying genetic diversity, population structure and phylogeography.

Genetic differentiation of Anisakis species (Nematoda: Anisakidae) in marine fish Priacanthus tayenus from Gulf of Thailand.

Members of the genus Anisakis are parasitic nematodes of the family Anisakidae. They are potential zoonotic parasites, causing anisakiasis in humans who consume raw or undercooked seafood (fish or squid) infected with the larvae of this nematode. In the present study, anisakid nematodes collected from the marine fish Priacanthus tayenus (Purplespotted big-eye) caught from the Gulf of Thailand were examined morphologically and characterized genetically by DNA sequence analysis. Sequence data from the mitochondrial cytochrome c oxidase subunit II (mtDNA cox2) gene were used to identify these nematodes to species level and to evaluate the phylogenetic relationship among various taxa. All the 15 third-stage larvae of Anisakis nematodes investigated in this study belonged to the same genetic lineage as the A. typica species complex (named here as A. typica sp. T - T for Thailand). Eight mtDNA cox2 haplotypes were revealed in the 15 isolates of this Anisakis from Thailand. The mtDNA cox2 haplotypes of A. typica sp. T from Thailand were genetically distinct from those of the A. typica sensu stricto. Taxonomic description of this A. typica sp. T as a distinct species however awaits the availability of adult specimens.

Alteration of -656(G/T) and -607(C/A) polymorphisms in interleukin-18 (IL-18) gene in house dust mite-sensitive allergic rhinitis patients in Thailand.

Allergic rhinitis (AR) is an IgE-mediated inflammation of the nasal membranes, which is naturally triggered by aeroallergens. House dust mites (HDM) are the most common inhalant allergens. Interleukin-18 (IL-18) has been established as an essential cytokine that can activate the generation of IgE. This randomized controlled study aimed to identify the possible relationship of the genetic variations in the IL-18 gene with AR in mite-sensitive Thai patients. Study subjects consisted of 150 AR patients and 50 normal participants. Genomic DNA of 30 randomized AR patients and 30 randomized controls were screened by sequencing for the selection of candidate single nucleotide polymorphisms (SNPs), and further analyzed by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) assay for all subjects. The following five SNPs were detected in the IL-18 gene: -656 G/T, -607 C/A, and -137 G/C in promoter 1 and -920 C/T and -373 C/G in promoter 2. The results showed that -656 G/T and -607 C/A SNPs were significantly correlated with IgE levels specific to Dermatophagoides pteronyssinus (Der p) allergen (P = 0.045 and P = 0.045, respectively), and significant differences were observed in the genotype distribution of AR patients when compared with controls [P = 0.044 and P = 0.044, respectively; odds ratios (ORs): 1.941 (95%CI, 1.014-3.715) and 1.941 (95%CI, 1.014-3.715), respectively]. Our findings indicate that the IL-18 alleles, -656T (rs1946519) and -607A (rs1946518), might be associated with the higher production of Der p allergen-specific IgE in mite-sensitive AR patients.

Exome sequencing in Thai patients with familial obesity.

Obesity is a major worldwide health issue, with increasing prevalence in adults and children from developed and developing countries. Obesity causes several chronic diseases, including cardiovascular and respiratory diseases, osteoarthritis, hypertension, stroke, type II diabetes, obstructive sleep apnea, and several types of cancer. Previous genome-wide association studies have identified several genes associated with obesity, including LEP, LEPR, POMC, PCSK1, FTO, MC3R, MC4R, GNPDA2, TMEM18, QPCTL/GIPR, BDNF, ETV5, MAP2K5/SKOR1, SEC16B, SIM1, and TNKS/MSRA. However, most of these variants are found in the intronic or intergenic regions, making it difficult to elucidate the underlying mechanisms. Therefore, in this study, we performed a whole exome sequencing of the protein-coding regions in the total genome (exome) of two obese and one normal subject belonging to the same Thai family to identify the genes responsible for obesity. We identified 709 functional variants that were differentially expressed between obese and normal subjects; of these, 65 were predicted to be deleterious to protein structure or function. The minor allele frequency of 14 of these genes (ALOX5AP, COL9A2, DEFB126, GDPD4, HCRTR1, MLL3, OPLAH, OR4C45, PRIM2, RXFP2, TIGD6, TRPM8, USP49, and ZNF596) was low, indicating causal variants that could be associated with complex traits or diseases. Genotyping revealed HCRTR1, COL9A2, and TRPM8 to be associated with the regulation of feeding behavior and energy expenditure. These genes constituted a network of pathways, including lipid metabolism, signaling transduction, immune, membrane transport, and gene regulation pathways, and seemed to play important roles in obesity.

Association between rs155971 in the PCSK1 gene and the lipid profile of obese Thai children: a family-based study.

Genetic variants of the POMC and PCSK1 genes cause severe obesity among patients in the early stages of childhood. This family-based study analyzed the links between single nucleotide polymorphisms (SNPs) in either the POMC or PCSK1 genes and obesity, as well as obesity-related traits among obese Thai children and their families. The variants rs1042571 and rs6713532 in the POMC gene in a sample of 83 obese children and their family members were investigated using polymerase chain reaction (PCR)-restriction fragment length polymorphism. In addition, the SNPs rs6232, rs155971, rs3762986, rs3811942, and rs371897784 of PCSK1 were analyzed in all samples using PCR and gene sequencing methods. Participants with the homozygous variant genotype in rs155971 had significantly elevated cholesterol and low-density lipoprotein cholesterol (LDL-C) levels (P = 0.011, OR = 1.025, 95%CI = 1.006-1.045; and P = 0.006, OR = 1.030, 95%CI = 1.009-1.053, respectively) after adjustment for age, gender, and body mass index (BMI). In addition, patients with the heterozygous variant genotype in rs371897784 of PCSK1 had a 1.249- fold higher risk (95%CI = 1.081-1.444, P = 0.027) of increased waist circumference than patients with the normal genotype, after adjustment for age, gender, and BMI. However, this analysis did not find any correlation between obesity and SNPs in PCSK1 and POMC. Therefore, these common variants in PCSK1 and POMC were not the major cause of obesity in the Thai subjects sampled. However, variants in PCSK1 did affect cholesterol level, LDL-C level, and waist circumference.

Genetic variant screening of MC3R and MC4R genes in early-onset obese children and their relatives among a Thai population: family-based study.

MC3R (melanocortin-3 receptor) and MC4R (melanocortin-4 receptor) play important roles in energy homeostasis. Severe early-onset obesity, known as monogenic obesity when it is the consequence of a mutation in a single-gene product, may result when energy homeostasis is disrupted. The purpose of our study was to screen for variations of the MC3R and MC4R genes and observe the mode of inheritance of variations in affected families. We used polymerase chain reaction and direct sequencing to analyze the 11 early-onset obese children (probands) with their 71 family members, together with DNA from 100 healthy subjects used as controls. No novel mutations were found in the MC3R gene. Two previously described polymorphisms, rs3746619 and rs3827103, were detected in the MC3R gene. It was not associated with any obesity-related phenotypes. Three heterozygous variations of the MC4R gene were detected in 3 of 11 probands. The rs34114122 and rs61741819 variations have previously been reported, but rs182455344 was novel. Moreover, each MC4R variant was also found in a number of family members, indicating that this molecular analysis of a family-based study showed an autosomal dominant pattern. Our study indicated that MC4R variations in early-onset obese Thai children were found, and transmission of these variations in each family is in the dominant pattern. These variants could possibly contribute to a genetic influence of early-onset obesity in Thais. There is no evidence of any association between MC3R variations and obesity.

Staphylococcus aureus clinical isolates: antibiotic susceptibility, molecular characteristics, and ability to form biofilm.

Periodic monitoring of Staphylococcus aureus characteristics in a locality is imperative as their drug-resistant variants cause treatment problem. In this study, antibiograms, prevalence of toxin genes (sea-see, seg-ser, seu, tsst-1, eta, etb, and etd), PFGE types, accessory gene regulator (agr) groups, and ability to form biofilm of 92 S. aureus Thailand clinical isolates were investigated. They were classified into 10 drug groups: groups 1-7 (56 isolates) were methicillin resistant (MRSA) and 8-10 (36 isolates) were methicillin sensitive (MSSA). One isolate did not have any toxin gene, 4 isolates carried one toxin gene (seq), and 87 isolates had two or more toxin genes. No isolate had see, etb, or tsst-1; six isolates had eta or etd. Combined seg-sei-sem-sen-seo of the highly prevalent egc locus was 26.1%. The seb, sec, sel, seu, and eta associated significantly with MSSA; sek was more in MRSA. The sek-seq association was 52.17% while combined sed-sej was not found. Twenty-three PFGE types were revealed, no association of toxin genes with PFGE types. All four agr groups were present; agr group 1 was predominant (58.70%) but agr group 2 strains carried more toxin genes and were more frequent toxin producers. Biofilm formation was found in 72.83% of the isolates but there was no association with antibiograms. This study provides insight information on molecular and phenotypic markers of Thailand S. aureus clinical isolates which should be useful for future active surveillance that aimed to control a spread of existing antimicrobial resistant bacteria and early recognition of a newly emerged variant.

Association of HTTLPR and 5-HT2A T102C polymorphisms with smoking characteristics and anthropometric profiles of Thai males.

Nicotine increases serotonin release in the brain. Gene polymorphisms in the serotonergic system have been suggested to be associated with smoking behavior. We investigated a possible association between two polymorphisms in the serotonergic system - HTTLPR of a serotonin transporter gene and 5-HT(2A) at position T102C - with biochemical and anthropometric parameters, and with cigarette smoking in an investigation of 200 smokers and 111 non-smokers. The two polymorphisms, HTTLPR and 5-HT(2A) at position T102C, were genotyped by PCR-RFLP. They were not significantly associated with smoking status in these Thai males. Among the smokers, thiocyanate concentrations and quantity of cigarettes smoked (cigarette pack-years) were significantly higher for individuals with LL/LS genotypes than SS genotypes of 5-HTTLPR (all P < 0.05), whereas "age at starting smoking" and "duration of smoking" were not significantly different between these two genotypes. Moreover, anthropometric variables, comprising triceps skinfold thickness, arm circumference, waist circumference, hip circumference, and waist-to-hip ratio, were significantly higher for the CC/TC genotypes of 5-HT(2A) than the TT genotype (all P < 0.05), except for body mass index. HTTLPR and 5-HT(2A) T102C polymorphisms were not significantly associated with smoking status among Thai males; however, the HTTLPR polymorphism among smokers appears to be an indicator of increased smoking intensity consisting of cigarette pack-years and thiocyanate concentrations. The 5-HT(2A) T102C polymorphism plays a role in the anthropometric profiles, triceps skinfold thickness, arm circumference, waist circumference, hip circumference, and waist-to-hip ratio, but not smoking status in Thai subjects.

Association of T869C gene polymorphism of transforming growth factor-ß1 with low protein levels and anthropometric indices in osteopenia/osteoporosis postmenopausal Thai women.

Osteoporosis is the most common metabolic bone disease; it is an important health problem among postmenopausal women. We evaluated the association of three polymorphisms, T869C, C-509T and G915C, of the TGF-ß1 gene with bone mineral density (BMD) serum TGF-ß1 levels in 278 postmenopausal female osteopenia/osteoporosis subjects and 95 postmenopausal female control subjects. Serum TGF-ß1 levels were significantly lower in osteopenia/osteoporosis subjects than in control subjects. Serum TGF-ß1 levels of the CT+CC (T869C) genotype group were significantly lower in osteopenia/osteoporosis subjects than in control subjects (11.3 vs 15.8 ng/mL). There was a significant difference in the CT+CC (T869C) genotype frequencies between the osteopenia/osteoporosis and control subjects (74.18 vs 60.22%; OR = 1.90, 95%CI = 1.16-3.12). In the age group of more than 50 years, subjects with the TC+CC genotype of T869C polymorphism had significantly increased risk of osteopenic/ osteoporotic bones at L1 (OR = 2.36, 95%CI = 1.37-4.07), L2 (OR = 1.71, 95%CI = 1.01-2.90), L3 (OR = 2.21, 95%CI = 1.23-3.98), L4 (OR = 1.74, 95%CI = 1.00-3.03) and the femoral neck (OR = 1.80, 95%CI = 1.04-3.12). The CT+CC genotype of the T869C polymorphism of the TGF-ß1 gene was found to be associated with lower serum TGF-ß1 in osteopenia/osteoporosis subjects and increased risk of osteopenic and osteoporotic fracture at L1-4, femoral neck and total hip in postmenopausal Thai women. Logistic regression analysis showed that T869C polymorphism is a significant risk factor for osteopenia/ osteoporosis. We concluded that T869C polymorphism of the TGF-ß1 gene has an impact on decreased serum TGF-ß1 levels and influences susceptibility to osteopenia/osteoporosis in Thai women.

Adiponectin/ACP30, a collagen-like plasma protein in relation to anthropometric measurement in Thai overweight and obese subjects.

Adiponectin, anthropometric parameters including weight, height, body mass index (BMI), arm circumference, triceps skinfold, subscapular skinfold, waist, hip circumferences and waist/hip ratio were recorded in 48 male and 166 female overweight and obese Thai volunteers (BMI=25.0 kg/m(2)), and in 26 male and 81 female normal subjects (BMI=18.5-24.9 kg/m(2)). Thai volunteers were investigated. Statistically significantly lower adiponectin concentrations in overweight and obese subjects were found when compared with control subjects of both sexes. Anthropometric parameters, including weight, height, BMI, arm circumference, triceps skinfold, subscapular skinfold, waist, hip circumferences and waist/hip ratio, except arm span, were statistically significantly higher in overweight and obese subjects than in control subjects. The overweight and obese subjects had higher glucose concentrations than the control subjects. The BMI and glucose concentrations were found to be significantly related, under these conditions, to adiponectin.

Occurrence of the infective stage of Angiostrongylus cantonensis in the yellow tree monitor (Varanus bengalensis) in five Provinces of Thailand.

Twenty-two yellow tree monitors (Varanus bengalensis) were trapped and transported from 5 provinces, namely Lampang, Phitsanulok, Kamphaeng Phet, Tak and Prachin Buri to look for the infective larvae stage of Angiostrongylus cantonensis. In 4 provinces all of the yellow tree monitors were infected with the infective stage larvae of A. cantonensis altogether. Twenty-one of the 22 yellow tree monitors (95.5%) in the five provinces were infected. Our results of this study extend our knowledge of natural prevalence of A. cantonensis in yellow tree monitors. The snail eating habit of the yellow tree monitor might possibly be the source of the larvae.

Multi-infection with helminths in adults from northeast Thailand as determined by post-treatment fecal examination of adult worms.

Six hundred and eighty-one residents from 16 provinces in northeast Thailand who had previously been found positive for Opisthorchis viverrini and Taenia spp. eggs were given 40 mg Praziquantel (Bayer, Germany), per kg body weight. The total stool output for one to three days was collected and examined for adult worms. The prevalence of O. viverrini in this group was 92.4%. The intestinal flukes Echinostoma malayanum, E. ilocanum and E. revolutum were found to be high in males, with 8.3%, 8.1% and 0.8%, respectively. Minute intestinal flukes such as Haplorchis taichui, H. pumilio, H. yokogawai, Phaneropsolus bonnei, Plagiorchis harinasutai, Prosthodendrium molenkampi and Stellantchasmus falcatus were found to be 7.8%, 6.2%, 2.9%, 15.0%, 0.7%, 19.4% and 0.3%, respectively.

Effects of sensitization on efficacy of mebendazole in mice infected with adult Angiostrongylus costaricensis.

The relation between immunopotentiation and efficacy of mebendazole in sensitized mice infected with adult Angiostrongylus costaricensis was investigated. When compared with the non-treated control, the sensitized control showed better results in almost all parameters with a few mice being positive for the first-stage larvae in feces and eggs in the intestinal section. The results suggest development of protective immunity in the sensitized mice though it was not completely effective for inhibiting infection, worm growth and their functions. The immunity seemed to be developed by producing specific antibodies against the larvae by sensitization. Among 4 infected groups, the sensitized-treated group had the best results in all parameters especially in worm recovery and worm body length which referred that drug action was enhanced by the sensitization.

Studies on chemotherapy of parasitic helminths: efficacy of artemether on Japanese strain of Schistosoma japonicum in mice.

Effects of artemether were examined on Schistosoma japonicum in mice. When the drug was given at a daily dosage of 200 mg/kg for 4 successive days from 46 days post-infection, a significant reduction in worm recovery was observed. A significant reduction in size of worms from the medicated mice was also seen compared with that from non-medicated controls.

Intestinal perforation due to Macracanthorhynchus hirudinaceus infection in Thailand.

The ninth record of a human infection with the acanthocephalan worm Macracanthorhynchus hirudinaceus in Thailand is reported. The parasite was removed during laparotomy after perforation through the intestine of a 30-year-old woman.