Characterization of Vibrio parahaemolyticus isolated from clinical specimens and oysters in Thailand.

INTRODUCTION: Vibrio parahaemolyticus is a common pathogen that can cause seafood-borne gastroenteritis in humans. We determined the prevalence and characteristics of V. parahaemolyticus isolated from clinical specimens and oysters in Thailand. METHODOLOGY: Isolates of V. parahaemolyticus from clinical specimens (n = 77) and oysters (n = 224) were identified by biochemical testing, polymerase chain reaction (PCR) assays, and serotyping. The toxin genes, antimicrobial resistance, and ß-lactamase production were determined. RESULTS: A total of 301 isolates were confirmed as V. parahaemolyticus by PCR using specific primers for the toxR gene. The majority of clinical isolates carried the tdh+/trh- genotype (82.1%), and one of each isolate was tdh-/trh+ and tdh+/trh+ genotypes. One isolate from oyster contained the tdh gene and another had the trh gene. Twenty-six serotypes were characterized among these isolates, and O3:K6 was the most common (37.7%), followed by OUT:KUT, and O4:K9. In 2010, most clinical and oyster isolates were susceptible to antibiotics, with the exception of ampicillin. In 2012, clinical isolates were not susceptible to cephalothin (52.4%), streptomycin (95.2%), amikacin (66.6%), kanamycin (61.9%), and erythromycin (95.2%), significantly more frequently than in 2010. More than 95% of isolates that were not susceptible to ampicillin produced ß-lactamase enzymes. CONCLUSIONS: We found toxin genes in two oyster isolates, and the clinical isolates that were initially determined to be resistant to several antibiotics. Toxin genes and antimicrobial susceptibility profiles of V. parahaemolyticus from seafood and environment should be continually monitored to determine the spread of toxin and antimicrobial resistance genes.

Spoligotype-based population structure and isoniazid-resistance gene mutation of Mycobacterium tuberculosis isolates from Thailand.

OBJECTIVES: The present study aims to investigate the population structure of Thai Mycobacterium tuberculosis (MTB) isolates and anti-tuberculosis (TB) drug resistance and to determine the most frequent genetic mutations conferring isoniazid (INH) resistance. METHODS: Genomic DNA from 287 MTB clinical isolates were extracted and used for spoligotyping, amplification and sequencing analysis of the region of different (RD) 105, and of the INH resistance (IR) associated genes, inhA, katG and oxyR-ahpC genes. RESULTS: Eighty-one clinical isolates were resistant to at least one first-line drug; 53 of these were resistant to INH. All strains were classified into three lineages based on their spoligotypes: East Asia (EA)/Beijing, Indo-Oceanic (IO), and Euro-American (EuA). EA and IO lineages revealed a strong association with anti-TB drug resistance (P = 0.005 and 0.013, respectively). A total of 33 mutations were found among IR isolates, which for 28 (84.8%), 3 (9.1%), and 2 (6.1%) occurred in katG, inhA, and oxyR-ahpC genes, respectively. Moreover, the most common mutations found were 54.7% of IR presented Ser315Thr at katG (54.7%) and C-15T at inhA (15.1%) presented. This result suggests the involvement of other genetic markers or other mechanisms of resistance. CONCLUSION: This study provides information about strains diversity, drug resistance profiles, and their possible association. EA and IO lineages were predominant in Thailand, and they were highly associated with anti-TB drug resistance. Testing two mutations including katGSer315Thr and inhA-15C→T could detect 68% of the IR strains.

Nasopharyngeal SARS-CoV-2 Viral Load Response among COVID-19 Patients Receiving Favipiravir.

We retrospectively studied nasopharyngeal severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) viral load in coronavirus disease 2019 (COVID-19) patients who were hospitalized between January 13 and April 1, 2020. Quantitative real-time reverse transcription-polymerase chain reaction (RT-PCR) was conducted using primers and probes targeting the ORF1ab and N genes. All patients were classified in the following groups: Group 1: received favipiravir + chloroquine or hydroxychloroquine + lopinavir/ritonavir or darunavir/ritonavir for 5-10 days, Group 2: received chloroquine or hydroxychloroquine + lopinavir/ritonavir or darunavir/ritonavir for 5-10 days, and Group 3: no antiviral medication. Among the 115 patients, 38 (33%), 54 (47%), and 23 (20%) were in Groups 1, 2, and 3, respectively. The median (IQR) baseline viral loads on day 0 of Groups 1, 2, and 3 were 7.2 (6.0-8.1), 6.9 (5.8-7.8), and 6.9 (5.8-7.6) log10 copies/mL, respectively. The reductions of mean viral loads on day 3 from baseline were 2.41, 1.38, and 2.19 log10 copies/mL in the corresponding groups (P < 0.05). There were no differences in the reduction of mean viral loads from baseline among the three groups on days 5 and 10 (P > 0.05). Multiple logistic regression analysis showed that receiving favipiravir was associated with nasopharyngeal viral load reduction at three days (P = 0.001). Significant nasopharyngeal SARS-CoV-2 viral load reduction was achieved in COVID-19 patients who received a favipiravir-containing regimen.

Characteristics of HIV-infected tuberculosis patients in Thailand.

To improve understanding about the epidemiology and clinical features of HIV-associated tuberculosis (TB) infection we conducted a prospective, multi-center observational study of HIV-infected TB patients in Thailand. We enrolled HIV-infected patients diagnosed with TB at public health facilities from three provinces and the national infectious diseases referral hospital in Thailand. Patients underwent standardized interviews, evaluations, and laboratory testing at the beginning of TB treatment. We analyzed demographic and clinical characteristics of patients and stratified our findings by level of immune-suppression and whether antiretroviral therapy (ART) was used before TB diagnosis. Of 769 patients analyzed, pulmonary TB was diagnosed in 461 (60%). The median CD4+ T-lymphocyte (CD4) count was 63 cells/microl [interquartile range (IQR), 23-163.5] and the median HIV RNA viral load was 308,000 copies/ml (IQR, 51,900-759,000) at the time of TB diagnosis. Methamphetamine use was reported by 304 patients (40%), marijuana by 267 patients (35%), and injection drug use by 199 patients (26%). Three hundred three patients (40%) reported having been previously incarcerated. Among sexually active patients, 142 (42%) reported never using condoms at all. Patients with CD4 counts <200 cells/microl were significantly more likely than patients with CD4 counts > or =200 cells/microl to have extra-pulmonary TB, fever, fatigue, muscle weakness, no hemoptysis, tachycardia, low body mass index, jaundice, or no pleural effusion. Of the 94 patients that received ART before TB diagnosis, the median time from ART initiation to TB diagnosis was 105 days (IQR, 31-468). HIV-infected patients who developed TB after ART initiation were more likely than other HIV-infected TB patients to have extra-pulmonary TB, a normal chest radiograph, low HIV RNA viral load, or a history of previous TB treatment.

Mutations in Streptomycin Resistance Genes and Their Relationship to Streptomycin Resistance and Lineage of Mycobacterium tuberculosis Thai Isolates.

BACKGROUND: Streptomycin (SM) is recommended by the World Health Organization (WHO) as a part of standard regimens for retreating multidrug-resistant tuberculosis (MDR-TB) cases. The incidence of MDR-TB in retreatment cases was 19% in Thailand. To date, information on SM resistance (SMR) gene mutations correlated to the SMR of Mycobacterium tuberculosis Thai isolates is limited. In this study, the mutations in rpsL, rrs, gidB, and whiB7 were investigated and their association to SMR and the lineage of M. tuberculosis were explored. METHODS: The lineages of 287 M. tuberculosis collected from 2007 to 2011 were identified by spoligotyping. Drug susceptibility profiles were evaluated by the absolute concentration method. Mutations in SMR genes of 46 SM-resistant and 55 SM-susceptible isolates were examined by DNA sequencing. RESULTS: Three rpsL (Lys43Arg, Lys88Arg, and Lys88Thr) and two gidB (Trp45Ter and Gly69Asp) mutations were present exclusively in the SM resistant M. tuberculosis. Lys43Arg rpsL was the most predominant SMR mutations (69.6%) and prevailed among Beijing isolates (p<0.001). No SMR-related mutation in was found rrs. The combination of rpsL and gidB mutations provided 76.1% sensitivity for detecting SMR in M. tuberculosis Thai isolates. whiB7 was not responsible for SMR in SM resistant isolates lacking rpsL and rrs mutations. The significance of the three gidB mutations, 276A>C, 615A>G, and 330G>T, as lineage signatures for Beijing and EAI were underscored. This study identified 423G>A gidB as a novel sub-lineage marker for EAI6-BGD1. CONCLUSION: Our study suggested that the majority of SMR in M. tuberculosis Thai isolates were responsible by rpsL and gidB polymorphisms constantly providing the novel lineage specific makers.

Antifungal susceptibilities of Cryptococcus neoformans cerebrospinal fluid isolates and clinical outcomes of cryptococcal meningitis in HIV-infected patients with/without fluconazole prophylaxis.

OBJECTIVES: To compare the MICs of FLUconazole (FLU) and amphotericin B against isolates of Cryptococcus neoformans (C. neoformans) obtained from the CerebroSpinal Fluid (CSF); and clinical outcomes of HIV-infected patients diagnosed with cryptococcal meningitis. MATERIAL AND METHOD: There were two groups including those who did not receive FLU (group A) and those who did receive either FLU 400 mg/week for primary prophylaxis cryptococosis or 200 mg/day for secondary prophylaxis cryptococosis (group B). CSF isolates of C. neoformans from group A and group B between January 2003 and October 2004 were retrospectively studied. The MICs were determined by using the standard NCCLS broth microdilution methods (M27-A). The MICs of FLU and amphotericin B, and clinical outcomes after 10 weeks of cryptococcal meningitis treatment were determined. RESULTS: There were 98 isolates; 80 in group A and 18 in group B. The patients in group B had a higher proportion of previous opportunistic infections (p = 0.008). The other baseline characteristics between the two groups were not different. The median (range) MIC of FLU was 8.0 (0.5-32) microg/ml in group A, and 6.0 (0.5-32) microg/ml in group B (p = 0.926). The median (range) MIC of amphotericin B was 0.25 (0.03-1.0) microg/ml in group A, and 0.25 (0.12-1.0) microg/ml in group B (p = 0.384). Sixty patients from group A and 14 from group B received standard treatment and continued to follow-up. After the 10-week treatment, 39/60 (65%) patients in group A and 7/14 (50%) in group B had complete recovery (p = 0.364; RR = 0.538, 95%CI = 0.166-1.742). The overall mortality rate was 14/60 (23.3%) in group A and 7/14 (50.0%) in group B (p = 0.096; RR = 3.286, 95%CI = 0.983-10.979). CONCLUSION: The MICs of FLU and amphotericin B against CSF isolates of C. neoformans and clinical outcomes between HIV-infected patients who receive or did not receive FLU prophylaxis are not different.

Time to initiate antiretroviral therapy between 4 weeks and 12 weeks of tuberculosis treatment in HIV-infected patients: results from the TIME study.

BACKGROUND: Optimal timing for initiation of antiretroviral therapy (ART) among HIV-infected patients with tuberculosis (TB) is not well established. METHODS: HIV/TB-coinfected patients were randomized to initiate tenofovir/lamivudine/efavirenz at 4 weeks (4-week group) or 12 weeks (12-week group) of TB treatment. The primary outcome was 1-year all-cause mortality. RESULTS: Of 156 patients, 79 were in 4-week group and 77 in 12-week group. Overall, median (interquartile range) CD4 was 43 (47-106) cells per cubic millimeter and median (interquartile range) HIV-1 RNA was 5.8 (5.4-6.3) log copies per milliliter. Eleven (7%) mortalities occurred in a total follow-up period of 137 patient-years. Seven percent (6/79, 8.76 per 100 patient-years) mortalities were in 4-week group, and 6% (5/77, 7.25 per 100 person-years) mortalities were in 12-week group [relative risk (RR) = 0.845, 95% confidence interval (CI) = 0.247 to 2.893]. Twenty-eight (35%) patients in 4-week group and 25 (32%) patients in 12-week group were hospitalized (RR = 1.142, 95% CI = 0.588 to 2.217). Grade 2-4 adverse events were 39% (31/79) in 4-week group and 34% (26/77) in 12-week group (RR = 1.267, 95% CI = 0.659 to 2.435). In multivariate analysis, "low albumin" (RR = 2.695, 95% CI = 1.353 to 5.475) and "low baseline CD4 count" (RR = 4.878, 95% CI = 1.019 to 23.256) were the independent predictors of mortality. Immune reconstitution inflammatory syndrome was more frequent in 4-week group with an incidence of 8.86 versus 5.02 per 100 person-months in 12-week group over the first 6 months of ART (P = 0.069). CONCLUSIONS: In middle-income countries where ART is initiated at CD4 count of <350 cells per cubic millimeter, immediate initiation of ART in HIV-infected patients with active TB was not associated with survival advantage when compared to initiation of ART at 12 weeks.

Clinical case definition and manifestations of paradoxical tuberculosis-associated immune reconstitution inflammatory syndrome.

BACKGROUND: The International Network for the Study of HIV-associated IRIS (INSHI) recently published criteria for tuberculosis-associated immune reconstitution inflammatory syndrome (TB-IRIS) diagnosis. The performance of this definition and clinical manifestations of TB-IRIS were studied. METHODS: Antiretroviral therapy-naive HIV/TB Thai patients receiving antituberculous therapy were enrolled during 2006-2007 and prospectively followed through 24 weeks of antiretroviral therapy. Patients were defined as having paradoxical TB-IRIS if they fulfilled the 'study definition' by French 2004 and were confirmed by an external reviewer. All were later compared by the classification according to 'INSHI-2008'. RESULTS: For the 126 patients, median baseline CD4 cell count was 43 cells/microl and HIV-1 RNA was 5.9 log(10) Y copies/ml. Seventy-three (58%) had extrapulmonary/disseminated TB. Twenty-two (18%) and 21 (17%) fulfilled TB-IRIS criteria according to the study definition and INSHI-2008 definition, respectively. Two (2%) were diagnosed by study definition only and one (1%) by INSHI-2008 definition only. Twenty (16%) were concordantly diagnosed by both definitions and 103 (82%) were consistently negative. Eighteen (82%) had worsening of a preexisting site, whereas four (18%) had TB-IRIS in a new location. Lymph node enlargement (73%) and fever (59%) were common in TB-IRIS. Sensitivity and specificity of INSHI-2008 was 91% (95% confidence interval, 72-98%) and 99% (95% confidence interval, 95-99.8%), respectively. Positive predictive value was 95% and negative predictive value was 98%. By multivariate analysis, factors predicting TB-IRIS were extrapulmonary TB (odds ratio, 8.63) and disseminated TB (odds ratio, 4.17). CONCLUSION: There was high concordance between the INSHI-2008 and French 2004 definition for TB-IRIS diagnosis in HIV/TB patients with relatively high rate of paradoxical TB-IRIS. This suggests that lack of HIV-1 RNA and CD4 cell count monitoring does not impede the ability to diagnose TB-IRIS.