Exploring the Genomic Diversity and Antimicrobial Susceptibility of Bifidobacterium pseudocatenulatum in a Vietnamese Population.

Bifidobacterium pseudocatenulatum is a member of the human gut microbiota, and specific variants of B. pseudocatenulatum have been associated with health benefits such as improving gut integrity and reducing inflammatory responses. Here, we aimed to assess the genomic diversity and predicted metabolic profiles of B. pseudocatenulatum cells found colonizing the gut of healthy Vietnamese adults and children. We found that the population of B. pseudocatenulatum from each individual was distinct and highly diverse, with intraclonal variation attributed largely to a gain or loss of carbohydrate-utilizing enzymes. The B. pseudocatenulatum genomes were enriched with glycosyl hydrolases predicted to target plant-based nondigestible carbohydrates (GH13, GH43) but not host-derived glycans. Notably, the exopolysaccharide biosynthesis region from organisms isolated from healthy children showed extensive genetic diversity and was subject to a high degree of genetic modification. Antimicrobial susceptibility profiling revealed that the Vietnamese B. pseudocatenulatum cells were uniformly susceptible to beta-lactams but exhibited variable resistance to azithromycin, tetracycline, ciprofloxacin, and metronidazole. The genomic presence of ermX and tet variants conferred resistance against azithromycin and tetracycline, respectively; ciprofloxacin resistance was associated with a mutation(s) in the quinolone resistance-determining region (GyrA, S115, and/or D119). Our work provides the first detailed genomic and antimicrobial resistance characterization of B. pseudocatenulatum found in the Vietnamese population, which can be exploited for the rational design of probiotics. IMPORTANCE Bifidobacterium pseudocatenulatum is a beneficial member of the human gut microbiota. The organism can modulate inflammation and has probiotic potential, but its characteristics are largely strain dependent and associated with distinct genomic and biochemical features. Population-specific beneficial microbes represent a promising avenue for the development of potential probiotics, as they may exhibit a more suitable profile in the target population. This study investigates the underexplored diversity of B. pseudocatenulatum in Vietnam and provides more understanding of its genomic diversity, metabolic potential, and antimicrobial susceptibility. Such data from indigenous populations are essential for selecting probiotic candidates that can be accelerated into further preclinical and clinical investigations.

The Gut Microbiome of Healthy Vietnamese Adults and Children Is a Major Reservoir for Resistance Genes Against Critical Antimicrobials.

Antimicrobials are a key group of therapeutic agents. Given the animal/human population density and high antimicrobial consumption rate in Southeast Asia, the region is a focal area for monitoring antimicrobial resistance (AMR). Hypothesizing that the gastrointestinal tract of healthy individuals in Vietnam is a major source of AMR genes that may be transferred to pathogens, we performed shotgun metagenomic sequencing on fecal samples from 42 healthy Vietnamese people (21 children and 21 adults). We compared their microbiome profiles by age group and determined the composition of AMR genes. An analysis of the taxonomic profiles in the gut microbiome showed a clear differentiation by age, with young children (age <2 years) exhibiting a unique structure in comparison to adults and older children. We identified a total of 132 unique AMR genes, with macrolide, lincosamide, and streptogramin class resistance genes (ermB and lnuC) and tetracycline resistance genes being almost ubiquitous across the study population. Notably, samples from younger children were significantly associated with a greater number of AMR genes than other age groups, including key signature genes associated with AMR pathogens (eg, blaCTX-M, mphA). Our data suggest that the gut microbiome of those living in Vietnam, particularly young children, is a substantial reservoir of AMR genes, which can be transferred to circulating enteric pathogens. Our data support the generation of longitudinal cohort studies of those living in urban and rural areas of developing countries to understand the behavior of these AMR reservoirs and their role in generating multidrug-resistant and extensively drug-resistant pathogens.

The transfer and decay of maternal antibody against Shigella sonnei in a longitudinal cohort of Vietnamese infants.

BACKGROUND: Shigella sonnei is an emergent and major diarrheal pathogen for which there is currently no vaccine. We aimed to quantify duration of maternal antibody against S. sonnei and investigate transplacental IgG transfer in a birth cohort in southern Vietnam. METHODS AND RESULTS: Over 500-paired maternal/infant plasma samples were evaluated for presence of anti-S. sonnei-O IgG and IgM. Longitudinal plasma samples allowed for the estimation of the median half-life of maternal anti-S. sonnei-O IgG, which was 43 days (95% confidence interval: 41-45 days). Additionally, half of infants lacked a detectable titer by 19 weeks of age. Lower cord titers were associated with greater increases in S. sonnei IgG over the first year of life, and the incidence of S. sonnei seroconversion was estimated to be 4/100 infant years. Maternal IgG titer, the ratio of antibody transfer, the season of birth and gestational age were significantly associated with cord titer. CONCLUSIONS: Maternal anti-S. sonnei-O IgG is efficiently transferred across the placenta and anti-S. sonnei-O maternal IgG declines rapidly after birth and is undetectable after 5 months in the majority of children. Preterm neonates and children born to mothers with low IgG titers have lower cord titers and therefore may be at greater risk of seroconversion in infancy.

Dexamethasone and long-term outcome of tuberculous meningitis in Vietnamese adults and adolescents.

BACKGROUND: Dexamethasone has been shown to reduce mortality in patients with tuberculous meningitis but the long-term outcome of the disease is unknown. METHODS: Vietnamese adults and adolescents with tuberculous meningitis recruited to a randomised, double-blind, placebo-controlled trial of adjunctive dexamethasone were followed-up at five years, to determine the effect of dexamethasone on long-term survival and neurological disability. RESULTS: 545 patients were randomised to receive either dexamethasone (274 patients) or placebo (271 patients). 50 patients (9.2%) were lost to follow-up at five years. In all patients two-year survival, probabilities tended to be higher in the dexamethasone arm (0.63 versus 0.55; p = 0.07) but five-year survival rates were similar (0.54 versus 0.51, p = 0.51) in both groups. In patients with grade 1 TBM, but not with grade 2 or grade 3 TBM, the benefit of dexamethasone treatment tended to persist over time (five-year survival probabilities 0.69 versus 0.55, p = 0.07) but there was no conclusive evidence of treatment effect heterogeneity by TBM grade (p = 0.36). The dexamethasone group had a similar proportion of severely disabled patients among survivors at five years as the placebo group (17/128, 13.2% vs. 17/116, 14.7%) and there was no significant association between dexamethasone treatment and disability status at five years (p = 0.32). CONCLUSIONS: Adjunctive dexamethasone appears to improve the probability of survival in patients with TBM, until at least two years of follow-up. We could not demonstrate a five-year survival benefit of dexamethasone treatment which may be confined to patients with grade 1 TBM. TRIAL REGISTRATION: ClinicalTrials.gov NCT01317654.

A retrospective analysis of the haemodynamic and metabolic effects of fluid resuscitation in Vietnamese adults with severe falciparum malaria.

BACKGROUND: Optimising the fluid resuscitation of patients with severe malaria is a simple and potentially cost-effective intervention. Current WHO guidelines recommend central venous pressure (CVP) guided, crystalloid based, resuscitation in adults. METHODS: Prospectively collected haemodynamic data from intervention trials in Vietnamese adults with severe malaria were analysed retrospectively to assess the responses to fluid resuscitation. RESULTS: 43 patients were studied of whom 24 received a fluid load. The fluid load resulted in an increase in cardiac index (mean increase: 0.75 L/min/m(2) (95% Confidence interval (CI): 0.41 to 1.1)), but no significant change in acid-base status post resuscitation (mean increase base deficit 0.6 mmol/L (95% CI: -0.1 to 1.3). The CVP and PAoP (pulmonary artery occlusion pressure) were highly inter-correlated (r(s) = 0.7, p<0.0001), but neither were correlated with acid-base status (arterial pH, serum bicarbonate, base deficit) or respiratory status (PaO(2)/FiO(2) ratio). There was no correlation between the oxygen delivery (DO(2)) and base deficit at the 63 time-points where they were assessed simultaneously (r(s) = -0.09, p = 0.46). CONCLUSIONS: In adults with severe falciparum malaria there was no observed improvement in patient outcomes or acid-base status with fluid loading. Neither CVP nor PAoP correlated with markers of end-organ perfusion or respiratory status, suggesting these measures are poor predictors of their fluid resuscitation needs.

Risk factors of Streptococcus suis infection in Vietnam. A case-control study.

BACKGROUND: Streptococcus suis infection, an emerging zoonosis, is an increasing public health problem across South East Asia and the most common cause of acute bacterial meningitis in adults in Vietnam. Little is known of the risk factors underlying the disease. METHODS AND FINDINGS: A case-control study with appropriate hospital and matched community controls for each patient was conducted between May 2006 and June 2009. Potential risk factors were assessed using a standardized questionnaire and investigation of throat and rectal S. suis carriage in cases, controls and their pigs, using real-time PCR and culture of swab samples. We recruited 101 cases of S. suis meningitis, 303 hospital controls and 300 community controls. By multivariate analysis, risk factors identified for S. suis infection as compared to either control group included eating "high risk" dishes, including such dishes as undercooked pig blood and pig intestine (OR(1) = 2.22; 95%CI = [1.15-4.28] and OR(2) = 4.44; 95%CI = [2.15-9.15]), occupations related to pigs (OR(1) = 3.84; 95%CI = [1.32-11.11] and OR(2) = 5.52; 95%CI = [1.49-20.39]), and exposures to pigs or pork in the presence of skin injuries (OR(1) = 7.48; 95%CI = [1.97-28.44] and OR(2) = 15.96; 95%CI = [2.97-85.72]). S. suis specific DNA was detected in rectal and throat swabs of 6 patients and was cultured from 2 rectal samples, but was not detected in such samples of 1522 healthy individuals or patients without S. suis infection. CONCLUSIONS: This case control study, the largest prospective epidemiological assessment of this disease, has identified the most important risk factors associated with S. suis bacterial meningitis to be eating 'high risk' dishes popular in parts of Asia, occupational exposure to pigs and pig products, and preparation of pork in the presence of skin lesions. These risk factors can be addressed in public health campaigns aimed at preventing S. suis infection.

Dexamethasone in Vietnamese adolescents and adults with bacterial meningitis.

BACKGROUND: It is uncertain whether all adults with bacterial meningitis benefit from treatment with adjunctive dexamethasone. METHODS: We conducted a randomized, double-blind, placebo-controlled trial of dexamethasone in 435 patients over the age of 14 years who had suspected bacterial meningitis. The goal was to determine whether dexamethasone reduced the risk of death at 1 month and the risk of death or disability at 6 months. RESULTS: A total of 217 patients were assigned to the dexamethasone group, and 218 to the placebo group. Bacterial meningitis was confirmed in 300 patients (69.0%), probable meningitis was diagnosed in 123 patients (28.3%), and an alternative diagnosis was made in 12 patients (2.8%). An intention-to-treat analysis of all the patients showed that dexamethasone was not associated with a significant reduction in the risk of death at 1 month (relative risk, 0.79; 95% confidence interval [CI], 0.45 to 1.39) or the risk of death or disability at 6 months (odds ratio, 0.74; 95% CI, 0.47 to 1.17). In patients with confirmed bacterial meningitis, however, there was a significant reduction in the risk of death at 1 month (relative risk, 0.43; 95% CI, 0.20 to 0.94) and in the risk of death or disability at 6 months (odds ratio, 0.56; 95% CI, 0.32 to 0.98). These effects were not found in patients with probable bacterial meningitis. Results of multivariate analysis indicated that dexamethasone treatment for patients with probable bacterial meningitis was significantly associated with an increased risk of death at 1 month, an observation that may be explained by cases of tuberculous meningitis in the treatment group. CONCLUSIONS: Dexamethasone does not improve the outcome in all adolescents and adults with suspected bacterial meningitis; a beneficial effect appears to be confined to patients with microbiologically proven disease, including those who have received prior treatment with antibiotics. (Current Controlled Trials number, ISRCTN42986828 [controlled-trials.com] .).

Evaluation of the MODS culture technique for the diagnosis of tuberculous meningitis.

BACKGROUND: Tuberculous meningitis (TBM) is a devastating condition. The rapid instigation of appropraite chemotherapy is vital to reduce morbidity and mortality. However rapid diagnosis remains elusive; smear microscopy has extremely low sensitivity on cerebrospinal fluid (CSF) in most laboratories and PCR requires expertise with advanced infrastructure and has sensitivity of only around 60% under optimal conditions. Neither technique allows for the microbiological isolation of M. tuberculosis and subsequent drug susceptibility testing. We evaluated the recently developed microscopic observation drug susceptibility (MODS) assay format for speed and accuracy in diagnosing TBM. METHODOLOGY/PRINCIPAL FINDINGS: Two hundred and thirty consecutive CSF samples collected from 156 patients clinically suspected of TBM on presentation at a tertiary referal hospital in Vietnam were enrolled into the study over a five month period and tested by Ziehl-Neelsen (ZN) smear, MODS, Mycobacterial growth Indicator tube (MGIT) and Lowenstein-Jensen (LJ) culture. Sixty-one samples were from patients already on TB therapy for >1day and 19 samples were excluded due to untraceable patient records. One hundred and fifty samples from 137 newly presenting patients remained. Forty-two percent (n = 57/137) of patients were deemed to have TBM by clinical diagnostic and microbiological criteria (excluding MODS). Sensitivity by patient against clinical gold standard for ZN smear, MODS MGIT and LJ were 52.6%, 64.9%, 70.2% and 70.2%, respectively. Specificity of all microbiological techniques was 100%. Positive and negative predictive values for MODS were 100% and 78.7%, respectively for HIV infected patients and 100% and 82.1% for HIV negative patients. The median time to positive was 6 days (interquartile range 5-7), significantly faster than MGIT at 15.5 days (interquartile range 12-24), and LJ at 24 days (interquartile range 18-35 days) (P<0.01). CONCLUSIONS: We have shown MODS to be a sensitive, rapid technique for the diagnosis of TBM with high sensitivity, ease of performance and low cost (0.53 USD/sample).

Serial MRI to determine the effect of dexamethasone on the cerebral pathology of tuberculous meningitis: an observational study.

BACKGROUND: Adjunctive dexamethasone increases survival from tuberculous meningitis, but the underlying mechanism is unclear. We aimed to determine the effect of dexamethasone on cerebral MRI changes and their association with intracerebral inflammatory responses and clinical outcome in adults treated for tuberculous meningitis. METHODS: Cerebral MRI was undertaken, when possible, at diagnosis and after 60 days and 270 days of treatment in adults with tuberculous meningitis admitted to two hospitals in Vietnam. Patients were randomly assigned either dexamethasone (n=24) or placebo (n=19) and received 9 months of treatment with standard first-line antituberculosis drugs. We assessed associations between MRI findings, treatment allocation, and resolution of fever, coma, cerebrospinal fluid inflammation, and neurological outcome. FINDINGS: 83 scans were done for 43 patients: 19 given placebo, 24 given dexamethasone. Basal meningeal enhancement (82%) and hydrocephalus (77%) were the most common presenting findings. Fewer patients had hydrocephalus after 60 days of treatment with dexamethasone than after placebo treatment (p=0.217). Tuberculomas developed in 74% of patients during treatment and in equal proportions in the treatment groups; they were associated with long-term fever, but not relapse or poor clinical outcome. The basal ganglia were the most common site of infarction; the proportion with infarction after 60 days was halved in the dexamethasone group (27%vs 58%, p=0.130). INTERPRETATION: Dexamethasone may affect outcome from tuberculous meningitis by reducing hydrocephalus and preventing infarction. The effect may have been under-estimated because the most severe patients could not be scanned.

Beijing genotype of Mycobacterium tuberculosis is significantly associated with human immunodeficiency virus infection and multidrug resistance in cases of tuberculous meningitis.

Multidrug-resistant tuberculous meningitis is fatal without rapid diagnosis and use of second-line therapy. It is more common in human immunodeficiency virus (HIV)-positive patients. Beijing genotype strains of Mycobacterium tuberculosis are associated with drug resistance, particularly multidrug resistance, and their prevalence is increasing worldwide. The prevalence of Beijing genotype strains among Mycobacterium tuberculosis isolates from the cerebrospinal fluid of HIV-positive (n = 35) and HIV-negative (n = 187) patients in Ho Chi Minh City was determined. The Beijing genotype was significantly associated with HIV status (odds ratio [OR] = 2.95 [95% confidence interval {CI}, 1.38 to 6.44]; P = 0.016), resistance to any drug (OR = 3.34 [95% CI, 1.87 to 5.95]; P < 0.001) and multidrug resistance (Fisher's exact test; P = 0.001). The association of the Beijing genotype with drug resistance was independent of HIV status. This is the first report of Beijing genotype association with HIV status, which may be an association unique to tuberculous meningitis.

Dexamethasone for the treatment of tuberculous meningitis in adolescents and adults.

BACKGROUND: Tuberculous meningitis kills or disables more than half of those affected with the disease. Previous studies have been too small to determine whether adjunctive treatment with corticosteroids can reduce the risk of disability or death among adults with tuberculous meningitis, and the effect of coinfection with the human immunodeficiency virus (HIV) is unclear. METHODS: We performed a randomized, double-blind, placebo-controlled trial in Vietnam in patients over 14 years of age who had tuberculous meningitis, with or without HIV infection, to determine whether adjunctive treatment with dexamethasone reduced the risk of death or severe disability after nine months of follow-up. We conducted prespecified subgroup analyses and intention-to-treat analyses. RESULTS: A total of 545 patients were randomly assigned to groups that received either dexamethasone (274 patients) or placebo (271 patients). Only 10 patients (1.8 percent) had been lost to follow-up at nine months of treatment. Treatment with dexamethasone was associated with a reduced risk of death (relative risk, 0.69; 95 percent confidence interval, 0.52 to 0.92; P=0.01). It was not associated with a significant reduction in the proportion of severely disabled patients (34 of 187 patients [18.2 percent] among survivors in the dexamethasone group vs. 22 of 159 patients [13.8 percent] in the placebo group, P=0.27) or in the proportion of patients who had either died or were severely disabled after nine months (odds ratio, 0.81; 95 percent confidence interval, 0.58 to 1.13; P=0.22). The treatment effect was consistent across subgroups that were defined by disease-severity grade (stratified relative risk of death, 0.68; 95 percent confidence interval, 0.52 to 0.91; P=0.007) and by HIV status (stratified relative risk of death, 0.78; 95 percent confidence interval, 0.59 to 1.04; P=0.08). Significantly fewer serious adverse events occurred in the dexamethasone group than in the placebo group (26 of 274 patients vs. 45 of 271 patients, P=0.02). CONCLUSIONS: Adjunctive treatment with dexamethasone improves survival in patients over 14 years of age with tuberculous meningitis but probably does not prevent severe disability.