Genotypic Distribution and the Epidemiology of Multidrug Resistant Tuberculosis in Upper Northern Thailand.

The epidemiology and genotypes of multidrug-resistant tuberculosis (MDR-TB), a global public health threat, remain limited. The genotypic distribution and factors associated with MDR-TB in upper northern Thailand between 2015 and 2019 were investigated. The DNA sequencing of rpoB, katG, and inhA promoter of 51 multidrug-resistant Mycobacterium tuberculosis isolates revealed nine patterns of the rpoB gene mutation distributed in seven provinces. The S531L mutation was the most common mutation in all provinces. The rpoB mutation in Chiang Rai, Chiang Mai, and Lampang was highly diverse compared to other areas. Here, the mutation profiles that have yet to be reported in northern Thailand (H526P, Q513P, and H526C) were detected in Chiang Rai province. The S315T katG mutation was the most common genotype associated with INH resistance, especially in Chiang Mai and Lampang. Further analysis of data from 110 TB patients (42 MDR-TB and 68 drug-susceptible TB) revealed that <60 years of age was a significant factor associated with MDR-TB (OR = 0.316, 95% CI 0.128−0.784, p = 0.011) and ≥60 years of age was a significant factor associated with the S315T katG-mutation (OR = 8.867, 95% CI 0.981−80.177, p = 0.047). This study highlighted the necessity for continuous surveillance and risk factor monitoring for effective control of MDR-TB.

Hb Athens-Georgia (beta 40(C6) Arg > Lys, HBB:c.122G > A) with a single α-globin gene (Hb H disease) in a Thai family: molecular, hematological, and diagnostic aspects.

Interaction of structural hemoglobin (Hb) variants with α- or ß-globin defects are occasional in Southeast Asia. Herein we provide the first description of Hb Athens-Georgia (Hb A-Ga) in association with deletional Hb H disease, a novel combination previously undescribed in the population. Hematological, Hb and DNA analysis, and ß-globin haplotype analyses were performed in seven participants from one ethnic Thai family. Hemoglobin analysis by capillary electrophoresis revealed an abnormal Hb fraction in the proband, his father and grandmother (I-2). DNA sequencing revealed that the G > A substitution at codon 40 of the ß-globin gene was identical to the Hb A-Ga (HBB:c.122G > A). Interestingly, α-thal-1 (SEA deletion) and α-thal-2 (-α3.7 deletion) were identified in the proband resulting in Hb H disease, while α-thal-1 was identified in the father, and no α-thal was observed in I-2. Hematological analysis indicated that the proband (ßA-Ga/ßA, -SEA/-α3.7) had moderate anemia and was markedly hypochromic with microcytic red blood cells (RBCs). The father (ßA-Ga/ßA, -SEA/αα) presented mild microcytic anemia, while normal hematology was observed in the I-2 who was heterozygous for Hb Athens-Georgia (ßA-Ga/ßA, αα/αα). The relative level of Hb A-Ga was distinctly reduced according to the degree of α-globin defects. The developed allele-specific PCR method can successfully be used for confirmation of Hb A-Ga. The Thai Hb A-Ga allele associated with a ß-haplotype [+ - - - - - +]. These findings were in accordance with the previous conclusion that this variant is a non-pathological ß-Hb variant.

Feasibility of using dried blood spots for HIV viral load testing among HIV-infected individuals in Thailand using QIAGEN QIAsymphony-artus HIV-1 platform.

In settings where plasma preparation and sample centralization are not feasible or inconvenient, dried blood spots (DBS) could be used as an alternative specimen to plasma to assess antiretroviral treatment response among HIV-infected individuals. This study was aimed to (1) validate the recent QIAsymphony-artus assay for DBS HIV viral load (VL) and (2) assess the feasibility of measuring HIV VL on DBS using this assay in Thailand. Ethylenediaminetetraacetic acid-blood samples from 99 HIV-infected individuals were used to prepare paired DBS and plasma. Also, DBS samples were shipped to three distant hospitals in the northern region. After short-term storage, DBS were returned by regular post to the AMS laboratory and were re-tested for HIV VL using the same platform. HIV VL results were compared using Pearson's correlation and Bland-Altman analysis. DBS HIV VL fairly correlated to plasma HIV VL (R = 0.62) with a mean difference of 0.02 log10 IU/mL (SD = 1.06). A high correlation (R = 0.79) was observed between HIV VL in DBS before and after shipping (mean difference = 0.14 log10 IU/mL, SD = 0.74), indicating good stability of HIV RNA in DBS. DBS can be used as an alternative specimen for HIV VL monitoring in Thailand. However, measurement of HIV VL with the QIAGEN QIAsymphony-artus assay should be improved, especially the DBS pre-extraction process.

Dietary supplements for dysmenorrhoea.

BACKGROUND: Dysmenorrhoea refers to painful menstrual cramps and is a common gynaecological complaint. Conventional treatments include non-steroidal anti-inflammatory drugs (NSAIDs) and oral contraceptive pills (OCPs), which both reduce myometrial activity (contractions of the uterus). A suggested alternative approach is dietary supplements. We used the term 'dietary supplement' to include herbs or other botanical, vitamins, minerals, enzymes, and amino acids. We excluded traditional Chinese medicines. OBJECTIVES: To determine the efficacy and safety of dietary supplements for treating dysmenorrhoea. SEARCH METHODS: We searched sources including the Cochrane Gynaecology and Fertility Group Specialised Register, the Cochrane Central Register of Controlled Trials (CENTRAL), MEDLINE, EMBASE, AMED, PsycINFO (all from inception to 23 March 2015), trial registries, and the reference lists of relevant articles. SELECTION CRITERIA: We included randomised controlled trials (RCTs) of dietary supplements for moderate or severe primary or secondary dysmenorrhoea. We excluded studies of women with an intrauterine device. Eligible comparators were other dietary supplements, placebo, no treatment, or conventional analgesia. DATA COLLECTION AND ANALYSIS: Two review authors independently performed study selection, performed data extraction and assessed the risk of bias in the included trials. The primary outcomes were pain intensity and adverse effects. We used a fixed-effect model to calculate odds ratios (ORs) for dichotomous data, and mean differences (MDs) or standardised mean differences (SMDs) for continuous data, with 95% confidence intervals (CIs). We presented data that were unsuitable for analysis either descriptively or in additional tables. We assessed the quality of the evidence using Grading of Recommendations Assessment, Development and Evaluation (GRADE) methods. MAIN RESULTS: We included 27 RCTs (3101 women). Most included studies were conducted amongst cohorts of students with primary dysmenorrhoea in their late teens or early twenties. Twenty-two studies were conducted in Iran and the rest were performed in other middle-income countries. Only one study addressed secondary dysmenorrhoea. Interventions included 12 different herbal medicines (German chamomile (Matricaria chamomilla, M recutita, Chamomilla recutita), cinnamon (Cinnamomum zeylanicum, C. verum), Damask rose (Rosa damascena), dill (Anethum graveolens), fennel (Foeniculum vulgare), fenugreek (Trigonella foenum-graecum), ginger (Zingiber officinale), guava (Psidium guajava), rhubarb (Rheum emodi), uzara (Xysmalobium undulatum), valerian (Valeriana officinalis), and zataria (Zataria multiflora)) and five non-herbal supplements (fish oil, melatonin, vitamins B1 and E, and zinc sulphate) in a variety of formulations and doses. Comparators included other supplements, placebo, no treatment, and NSAIDs.We judged all the evidence to be of low or very low quality. The main limitations were imprecision due to very small sample sizes, failure to report study methods, and inconsistency. For most comparisons there was only one included study, and very few studies reported adverse effects. Effectiveness of supplements for primary dysmenorrhoea We have presented pain scores (all on a visual analogue scale (VAS) 0 to 10 point scale) or rates of pain relief, or both, at the first post-treatment follow-up. Supplements versus placebo or no treatmentThere was no evidence of effectiveness for vitamin E (MD 0.00 points, 95% CI -0.34 to 0.34; two RCTs, 135 women).There was no consistent evidence of effectiveness for dill (MD -1.15 points, 95% CI -2.22 to -0.08, one RCT, 46 women), guava (MD 0.59, 95% CI -0.13 to 1.31; one RCT, 151 women); one RCT, 73 women), or fennel (MD -0.34 points, 95% CI -0.74 to 0.06; one RCT, 43 women).There was very limited evidence of effectiveness for fenugreek (MD -1.71 points, 95% CI -2.35 to -1.07; one RCT, 101 women), fish oil (MD 1.11 points, 95% CI 0.45 to 1.77; one RCT, 120 women), fish oil plus vitamin B1 (MD -1.21 points, 95% CI -1.79 to -0.63; one RCT, 120 women), ginger (MD -1.55 points, 95% CI -2.43 to -0.68; three RCTs, 266 women; OR 5.44, 95% CI 1.80 to 16.46; one RCT, 69 women), valerian (MD -0.76 points, 95% CI -1.44 to -0.08; one RCT, 100 women), vitamin B1 alone (MD -2.70 points, 95% CI -3.32 to -2.08; one RCT, 120 women), zataria (OR 6.66, 95% CI 2.66 to 16.72; one RCT, 99 women), and zinc sulphate (MD -0.95 points, 95% CI -1.54 to -0.36; one RCT, 99 women).Data on chamomile and cinnamon versus placebo were unsuitable for analysis. Supplements versus NSAIDSThere was no evidence of any difference between NSAIDs and dill (MD 0.13 points, 95% CI -1.01 to 1.27; one RCT, 47 women), fennel (MD -0.70 points, 95% CI -1.81 to 0.41; one RCT, 59 women), guava (MD 1.19, 95% CI 0.42 to 1.96; one RCT, 155 women), rhubarb (MD -0.20 points, 95% CI -0.44 to 0.04; one RCT, 45 women), or valerian (MD points 0.62 , 95% CI 0.03 to 1.21; one RCT, 99 women),There was no consistent evidence of a difference between Damask rose and NSAIDs (MD -0.15 points, 95% CI -0.55 to 0.25; one RCT, 92 women).There was very limited evidence that chamomile was more effective than NSAIDs (MD -1.42 points, 95% CI -1.69 to -1.15; one RCT, 160 women). Supplements versus other supplementsThere was no evidence of a difference in effectiveness between ginger and zinc sulphate (MD 0.02 points, 95% CI -0.58 to 0.62; one RCT, 101 women). Vitamin B1 may be more effective than fish oil (MD -1.59 points, 95% CI -2.25 to -0.93; one RCT, 120 women). Effectiveness of supplements for secondary dysmenorrhoea There was no strong evidence of benefit for melatonin compared to placebo for dysmenorrhoea secondary to endometriosis (data were unsuitable for analysis). Safety of supplements Only four of the 27 included studies reported adverse effects in both treatment groups. There was no evidence of a difference between the groups but data were too scanty to reach any conclusions about safety. AUTHORS' CONCLUSIONS: There is no high quality evidence to support the effectiveness of any dietary supplement for dysmenorrhoea, and evidence of safety is lacking. However for several supplements there was some low quality evidence of effectiveness and more research is justified.

Novel targeting of the lepB gene using PCR with confronting two-pair primers for simultaneous detection of Mycobacterium tuberculosis complex and Mycobacterium bovis.

Tuberculosis (TB), caused by members of the Mycobacterium tuberculosis complex (MTC), is the leading cause of infectious disease-related mortality worldwide. The standard method for TB diagnosis usually requires long periods of mycobacteria cultivation, leading to delayed diagnosis, inefficient treatment and widespread occurrence of the disease. Therefore, a rapid method for the detection and differentiation of MTC from other mycobacteria is essential for disease diagnosis. Here, we describe the potential of using the type I signal peptidase (lepB) gene as a novel target for TB diagnosis, based on confronting two-pair primers PCR (PCRCTPP) that can detect MTC and simultaneously differentiate M. bovis. The limit of detection of the developed technique was equivalent to 12­120 bacilli. PCR-CTPP was highly specific to only MTC and M. bovis, and no cross-reaction was detected in 27 DNA of the non-tuberculous mycobacterial and bacterial strains tested. Thirty-nine blinded clinical isolates and 72 sputum samples were used to validate the PCR-CTPP in comparison with the standard mycobacterial culture method. The sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) of PCR-CTPP were equal to 95, 100, 100 and 95 %, respectively, when tested with clinical isolates. Furthermore, upon testing with the sputum samples, the sensitivity, specificity, PPV and NPV were observed to be 84, 76, 90 and 67 %, respectively. Hence, this highly sensitive novel technique, which is rapid, easy to conduct and cost-effective, is a potential method for TB diagnosis and epidemiological studies, especially in resource-limited countries with a high TB burden.

Clinical indicators for severe prognosis of scrub typhus.

BACKGROUND: The study explored clinical risk characteristics that may be used to forecast scrub typhus severity under routine clinical practices. METHODS: Retrospective data were collected from patients registered at two university-affiliated tertiary care hospitals in the north of Thailand, from 2004 to 2010. Key information was retrieved from in-patient records, out patient cards, laboratory reports and registers. Patients were classified into three severity groups: nonsevere, severe (those with at least one organ involvement), and deceased. Prognostic characteristics for scrub typhus severity were analyzed by a multivariable ordinal continuation ratio regression. RESULTS: A total of 526 patients were classified into nonsevere (n = 357), severe (n = 100), and deceased (n = 69). The significant multivariable prognostic characteristics for scrub typhus severity were increased body temperature (odds ratio [OR] = 0.58, 95% confidence interval [CI] = 0.45-0.74, P < 0.001), increased pulse rate (OR = 1.03, 95% CI = 1.01-1.05, P < 0.001), presence of crepitation (OR = 3.25, 95% CI = 1.52-6.96, P = 0.001), increased percentage of lymphocytes (OR = 0.97, 95% CI = 0.95-0.98, P = 0.001), increased aspartate aminotransferase (every 10 IU/L) (OR = 1.04, 95% CI = 1.02-1.06, P < 0.001), increased serum albumin (OR = 0.47, 95% CI = 0.27-0.80, P = 0.001), increased serum creatinine (OR = 1.83, 95% CI = 1.50-2.24, P < 0.001), and increased levels of positive urine albumin (OR = 1.43, 95% CI = 1.17-1.75, P < 0.001). CONCLUSION: Patients suspicious of scrub typhus with low body temperature, rapid pulse rate, presence of crepitation, low percentage of lymphocyte, low serum albumin, elevated aspartate aminotransferase, elevated serum creatinine, and positive urine albumin should be monitored closely for severity progression.

Semi-nested Taqman real-time quantitative PCR for noninvasive prenatal diagnosis of Bart's hydrops fetalis.

BACKGROUND: Non-invasive prenatal diagnosis based on detection of fetal cell-free DNA is limited when mother and father are both carriers for the same autosomal recessive mutation. OBJECTIVE: Develop the semi-nested Taqman real-time PCR for quantification of alpha-thalassemia-1 SEA type deletion allele in plasma of alpha-thalassemia-1 SEA carriage pregnancies. MATERIAL AND METHOD: Plasma DNA was extracted from six women who carried fetuses with normal, 11 with heterozygote alpha-thalassemia-1 SEA type deletion and seven with Bart's hydrops fetalis. DNA was amplified using conventional PCR with the primary specific primer set for alpha-thalassemia-1 SEA type deletion. PCR product was then subjected to the semi-nested real-time PCR using the secondary specific primer and Taqman probe set for alpha-thalassemia-1 SEA type deletion. The standard curve was constructed using ten-fold serial dilutions of conventional PCR product of the heterozygote alpha-thalassemia-1 SEA type deletion. RESULTS: Women who carried fetuses with Bart's hydrops fetalis displayed a trend toward higher mean copy number of alpha-thalassemia-1 SEA type deletion allele vs. women who carried fetuses with normal and heterozygote, albeit not reaching statistical significance. CONCLUSION: The maternally inheritedfetal allele present in maternal plasma is difficult to discern the fetal cell-free DNA from a higher background DNA of the mother Thus, further investigation is needed to improve the diagnosis ofBart's hydrops fetalis using this technique.

Analysis of real-time PCR cycle threshold of alpha-thalassemia-1 Southeast Asian type deletion using fetal cell-free DNA in maternal plasma for noninvasive prenatal diagnosis of Bart's hydrops fetalis.

BACKGROUND: Noninvasive prenatal diagnosis based on detection of fetal cell-free DNA is hampered when mother and father are both carriers for the same autosomal recessive mutation. OBJECTIVE: To compare the diagnosis of Bart's hydrops fetalis using conventional Gap-PCR analysis of fetal cells/tissues with the measurement of quantitative difference (deltaCp) between alpha-thalassemia-1 SEA type deletion gene (C(T-mutant)) and wild type alpha-globin gene (C(T-wild type)) in plasma of pregnancies by using the Taqman real-time quantitative PCR. MATERIAL AND METHOD: Plasma DNA samples were collected from three groups of pregnancies whose fetuses have known thalasemia status (7 normal, 11 heterozygote alpha-thalassemia-1 SEA type deletion, and 7 Bart's hydrops fetalis). The alpha-thalassemia-1 SEA type deletion gene and wild type alpha-globin gene were quantified by using Taqman real-time quantitative PCR and then the delta C(T) was analyzed by subtracting the C(T-mutant) from C(T-wild type). RESULTS: Mean deltaC(T) values were not significantly different among the three groups. However, women whose fetuses were diagnosed as Bart's hydrops fetalis had a higher proportion (43%) of plasma DNA samples that had negative deltaC(T) value than women whose fetuses were diagnosed as normal or heterozygote alpha-thalassemia-1 SEA type deletion (0 and 27%, respectively). CONCLUSION: Further investigations are needed to improve the diagnosis of Bart's hydrops fetalis using fetal cell-free DNA.

Thalassemia intermedia associated with complex interaction of Hb Beijing [alpha16(A14)Lys-->Asn] and Hb E [beta26(B8)Glu-->Lys] with a deletional alpha-thalassemia-1 in a Thai family.

A Thai family with a complex thalassemia syndrome caused by alpha- and beta-globin defects is described. The proband was a 14-year-old boy who had chronic hypochromic microcytic anemia. Hemoglobin (Hb) and DNA analyses demonstrated that he carried Hb Beijing [alpha16(A14)Lys-->Asn], Hb E [beta26(B8)Glu-->Lys] and alpha-thalassemia-1 (alpha-thal-1). Interaction of the alphaBeijing with the betaE globin chains in the proband leads to a new Hb variant, namely Hb E Beijing with different characteristics to both Hb E and Hb Beijing. Family studies showed that his father carried Hb Beijing and Hb E, whereas his mother was a simple alpha-thal-1 carrier. The genotype-phenotype relationship observed in this Thai family with complex hemoglobinopathies is presented and a simple DNA assay based on allele specific polymerase chain reaction (ASPCR) for detection of Hb Beijing is described.