The potential mechanism of the progression from latent to active tuberculosis based on the intestinal microbiota alterations.

INTRODUCTION: Tuberculosis (TB) poses a serious challenge to global health systems. The altered intestinal microbiota is associated with the pathogenesis of TB, but the exact links remain unclear. METHODS: 16 S rDNA sequencing was performed to comprehensively detect the changes in the intestinal microbiota of feces from active TB (ATB), latent TB infection (LTBI) and healthy controls (HC). RESULTS: The rarefaction curves demonstrated the sequencing results' validity. The alpha diversity was lowest in ATB, while highest in HC. Boxplot of beta diversity showed significant differences in every two groups. LDA Effect Size (LEfSe) Analysis revealed differences in probiotic bacteria like Romboutsia, Bifidobacterium and Lactobacillus in LTBI, and pro-inflammatory bacteria like R. gnavus, Streptococcus and Erysipelatoclostridium in ATB, corresponding to the cluster analysis. PICRUST2 analysis revealed the pentose phosphate pathway was active in ATB and LTBI (more active in ATB). The differences between the groups are statistically significant at the P<0.05 level. CONCLUSION: Our study indicated that from LTBI to ATB, some intestinal microbiota inhibit the synthesis of interferon (INF)-γ and interleukin (IL)-17, promoting the survival and spread of Mycobacterium tuberculosis (M. tb). In addition, the metabolites secreted by intestinal microbiota and dysbiosis in intestine also have an effect on the development of LTBI to ATB.

Untargeted Metabolomics of Feces Reveals Diagnostic and Prognostic Biomarkers for Active Tuberculosis and Latent Tuberculosis Infection: Potential Application for Precise and Non-Invasive Identification.

Purpose: Distinguishing latent tuberculosis infection (LTBI) from active tuberculosis (ATB) is important to control the prevalence of tuberculosis; however, there is currently no effective method. The aim of this study was to discover specific metabolites through fecal untargeted metabolomics to discriminate ATB, individuals with LTBI, and healthy controls (HC) and to probe the metabolic perturbation associated with the progression of tuberculosis. Patients and Methods: Liquid chromatography-tandem mass spectrometry (LC-MS/MS) was performed to comprehensively detect compounds in fecal samples from HC, LTBI, and ATB patients. Differential metabolites between the two groups were screened, and their underlying biological functions were explored. Candidate metabolites were selected and enrolled in LASSO regression analysis to construct diagnostic signatures for discriminating between HC, LTBI, and ATB. A receiver operating characteristic (ROC) curve was applied to evaluate diagnostic value. A nomogram was constructed to predict the risk of progression of LTBI. Results: A total of 35 metabolites were found to exist differentially in HC, LTBI, and ATB, and eight biomarkers were selected. Three diagnostic signatures based on the eight biomarkers were constructed to distinguish between HC, LTBI, and ATB, demonstrating excellent discrimination performance in ROC analysis. A nomogram was successfully constructed to evaluate the risk of progression of LTBI to ATB. Moreover, 3,4-dimethylbenzoic acid has been shown to distinguish ATB patients with different responses to etiological tests. Conclusion: This study constructed diagnostic signatures based on fecal metabolic biomarkers that effectively discriminated HC, LTBI, and ATB, and established a predictive model to evaluate the risk of progression of LTBI to ATB. The results provide scientific evidence for establishing an accurate, sensitive, and noninvasive differential diagnosis scheme for tuberculosis.

Effects of a false-positive result in newborn congenital hypothyroidism screening on parents in Guangxi, China.

Background: As more than 500,000 neonates participate in newborn congenital hypothyroidism (CH) screening in Guangxi Zhuang Autonomous Region each year, the overall number of false-positive (FP) cases has increased. We aim to assess the parental stress in parents of neonates with FP CH results in Guangxi, find out the influence factors related to demographics, and provide the basis for personalized health education. Methods: The parents of neonates with FP CH results were invited to participate in the FP group, and the parents of neonates with all negative results were invited to participate in the control group. The parents completed a questionnaire on demographics, knowledge of CH, and the parental stress index (PSI) in the hospital for the first time. The follow-up visits for PSI were conducted 3, 6, and 12 months afterward through telephone and online. Results: A total of 258 and 1,040 parents participated in the FP and control groups, respectively. The parents in the FP group had better knowledge of CH and higher PSI scores than the parents in the control group. The result of logistic regression showed that the major influence factors related to the knowledge of CH were FP experience and source of knowledge. The parents in the FP group who were well-informed during the recall phone call had lower PSI scores than the other parents. The parents in the FP group showed decreasing PSI scores gradually in follow-up visits. Conclusion: The results suggested that FP screening results may affect parental stress and parent-child relationship. FP results increased the stress on the parents and increased their knowledge of CH passively.

Clinical evaluation of a loop-mediated isothermal amplification (LAMP) assay for rapid detection of Neisseria meningitidis in cerebrospinal fluid.

BACKGROUND: Neisseria meningitidis (Nm) is a leading causative agent of bacterial meningitis in humans. Traditionally, meningococcal meningitis has been diagnosed by bacterial culture. However, isolation of bacteria from patients' cerebrospinal fluid (CSF) is time consuming and sometimes yields negative results. Recently, polymerase chain reaction (PCR)-based diagnostic methods of detecting Nm have been considered the gold standard because of their superior sensitivity and specificity compared with culture. In this study, we developed a loop-mediated isothermal amplification (LAMP) method and evaluated its ability to detect Nm in cerebrospinal fluid (CSF). METHODOLOGY/PRINCIPAL FINDINGS: We developed a meningococcal LAMP assay (Nm LAMP) that targets the ctrA gene. The primer specificity was validated using 16 strains of N. meningitidis (serogroup A, B, C, D, 29-E, W-135, X, Y, and Z) and 19 non-N. meningitidis species. Within 60 min, the Nm LAMP detected down to ten copies per reaction with sensitivity 1000-fold more than that of conventional PCR. The LAMP assays were evaluated using a set of 1574 randomly selected CSF specimens from children with suspected meningitis collected between 1998 and 2002 in Vietnam, China, and Korea. The LAMP method was shown to be more sensitive than PCR methods for CSF samples (31 CSF samples were positive by LAMP vs. 25 by PCR). The detection rate of the LAMP method was substantially higher than that of the PCR method. In a comparative analysis of the PCR and LAMP assays, the clinical sensitivity, specificity, positive predictive value, and negative predictive value of the LAMP assay were 100%, 99.6%, 80.6%, and 100%, respectively. CONCLUSIONS/SIGNIFICANCE: Compared to PCR, LAMP detected Nm with higher analytical and clinical sensitivity. This sensitive and specific LAMP method offers significant advantages for screening patients on a population basis and for diagnosis in clinical settings.

An expanded age range for meningococcal meningitis: molecular diagnostic evidence from population-based surveillance in Asia.

BACKGROUND: To understand epidemiologic patterns of meningococcal disease in Asia, we performed a retrospective molecular analysis of cerebrospinal fluid (CSF) specimens collected in prospective surveillance among children aged < 5 years of age in China, South Korea, and Vietnam. METHODS: A total of 295 isolates and 2,302 CSFs were tested by a meningococcal species- and serogroup-specific polymerase chain reaction (PCR) assay targeting the Neisseria meningitidis (Nm) ctrA gene. Multi-locus sequence typing (MLST) was performed in Nm gene amplification analysis and incidence rates for meningococcal meningitis were estimated. RESULTS: Among 295 isolates tested, 10 specimens from Vietnam were confirmed as serogroup B and all were Sequence Type (ST) 1576 by MLST. Among the 2,032 CSF specimen tested, 284 (14%) were confirmed by PCR (ctrA gene), including 67 (23.6%) from China, 92 (32.4%) from Korea, and 125 (44.0%) from Vietnam. Neonates and infants aged < 6 months of age accounted for more than 50% of Nm-PCR positive CSF. Two CSF specimens from Vietnam were identified as serogroup B using MLST. In addition, 44 specimens underwent sequencing to confirm meningococcal serogroup; of these, 21 (48%) were serogroup C, 12 (27%) were serogroup X, 9 (20%) were serogroup Y and 2 (5%) were serogroup B. The incidence rates of meningococcal meningitis among children < 5 years of age was highest in Vietnam (7.4/100,000 [95% CI, 3.6-15.3] followed by Korea (6.8/100,000 [95% CI, 3.5-13.5] and China (2.1/100,000) [95% CI, 0.7-6.2]). CONCLUSIONS: These results suggest that there is a previously undetected, yet substantial burden of meningococcal meningitis among infants and young children. Standardized, sensitive and specific molecular diagnostic assays with Nm serogrouping capacity are needed throughout Asia to understand the true burden of N. meningitidis disease.

The enhanced pneumococcal LAMP assay: a clinical tool for the diagnosis of meningitis due to Streptococcus pneumoniae.

BACKGROUND: Streptococcus pneumoniae is a leading cause of invasive bacterial disease in developed and developing countries. We studied the loop-mediated isothermal amplification (LAMP) technique to assess its suitability for detecting S. pneumoniae nucleic acid in cerebrospinal fluid (CSF). METHODOLOGY/PRINCIPAL FINDINGS: We established an improved LAMP assay targeting the lytA gene (Streptococcus pneumoniae [Sp] LAMP). The analytical specificity of the primers was validated by using 32 reference strains (10 Streptococcus and seven non-Streptococcus species) plus 25 clinical alpha-hemolytic streptococcal strains, including four S. pneumoniae strains and 21 other strains (3 S. oralis, 17 S. mitis, and one Streptococcus species) harboring virulence factor-encoding genes (lytA or ply). Within 30 minutes, the assay could detect as few as 10 copies of both purified DNA and spiked CSF specimens with greater sensitivity than conventional polymerase chain reaction (PCR). The linear determination range for this assay is 10 to 1,000,000 microorganisms per reaction mixture using real-time turbidimetry. We evaluated the clinical sensitivity and specificity of the Sp LAMP assay using 106 randomly selected CSF specimens from children with suspected meningitis in Korea, China and Vietnam. For comparison, CSF specimens were also tested against conventional PCR and culture tests. The detection rate of the LAMP method was substantially higher than the rates of PCR and culture tests. In this small sample, relative to the LAMP assay, the clinical sensitivity of PCR and culture tests was 54.5% and 33.3%, respectively, while clinical specificity of the two tests was 100%. CONCLUSIONS/SIGNIFICANCE: Compared to PCR, Sp LAMP detected S. pneumoniae with higher analytical and clinical sensitivity. This specific and sensitive LAMP method offers significant advantages for screening patients on a population basis and for diagnosis in clinical settings.

[Surveillance and research on acute meningitis, encephalitis syndrome in Guangxi, China].

OBJECTIVE: To estimate disease burden and epidemiological characteristics of acute meningitis/encephalitis, and provide the basis for the disease control strategy development. METHODS: A syndrome surveillance system was established in Guigang city with a population of 5 020 000. For the suspected cases, serum and CSF were collected, and bacterial culture, latex agglutination test, real-time PCR and ELISA tests were carried out. All involved cases were identified to 6 categories according to WHO case definition. RESULTS: 1424 suspected cases were evaluated in a surveillance of 30 months, yielding the incidence, mortality and mortality of 11.35/100 000 (1424/12 546 500 person years), 0.43/100 000 (54/12 546 500 person years), 3.79% (54/1424) respectively. A total of 103 and 51 cases were confirmed for JE, bacterial meningitis, with a incidence of 0.82/100 000 (103/12 546 500 person years), 0.41/100 000 (51/12 546 500 person years). 96.10% (99/103) of JE cases and 37.30% (19/51) bacterial meningitis cases occurred in < 10 years old children and < 5 years old children. A clinical misdiagnosis rate of 19.42% (20/103) and 15.69% (8/51) were observed for JE and bacterial meningitis. CONCLUSION: Acute encephalitis, meningitis syndrome can cause a higher burden of disease, of which the main components of viral encephalitis. Most of syndrome is occurred in summer and autumn, mainly reported in children of younger than 10 years old. A quite misdiagnosis would be made among meningitis and encephalitis syndrome cases.

[On-site assessment on the definition and classification of suspected cases in the manual of WHO Japanese Encephalitis Surveillance Standards].

OBJECTIVE: To assess the new edition of WHO Japanese Encephalitis (JE) Surveillance Standards (WHO Standards) based on syndrome surveillance data and to provide field evidence regarding the standards. METHODS: Based on syndrome surveillance data, acute encephalitis syndrome (AES) case was categorized, according to the WHO Standards. A cohort study was applied to estimate the AES definition set in the Standard and relative risk was computed to estimate the existence and intensity of statistical correlation between AES and JE cases. Percentage of attributable risk was counted to describe the coverage of AES for JE cases in the studied population. Sensitivity, specificity, Youden index and positive predictive value of AES components were calculated for the purpose of identifying the clinical values under the screening program. RESULTS: 1424 suspected cases were evaluated in the surveillance program and 1396 cases with ELISA result, of which 109 positive cases were detected. According to the "standardized" classification, a total of 706 cases in line with AES case definition, were categorized into 83 cases of JE, 425 cases of AES unknown and 198 cases of AES other agent. In the cohort study, a relative risk of 4.62 (95%CI: 2.80 - 7.63) and the percentage of attributable risk as 78.35% (95%CI: 64.25% - 86.89%) were observed. CONCLUSION: The AES definition for JE was significantly effecting on the screening programs and a strong correlation strength was observed in the study. AES syndrome could cover most of the JE cases. "Convulsions", with appreciative screening value, was recommended to be involved into the new version of the WHO Standards.

Trends and disease burden of enteric fever in Guangxi province, China, 1994-2004.

OBJECTIVE: To determine the burden of enteric fever through trends in morbidity and mortality, bacterial species and antimicrobial resistance in Guangxi, a southern, subtropical, coastal province of China with a disproportionally large burden of enteric fever. METHODS: Data on morbidity and mortality caused by enteric fever between 1994 and 2004 were extracted from the Guangxi Center for Disease Control and Prevention. Laboratory-based surveillance and outbreak investigations were integrated with reports of notifiable infectious diseases to estimate the bacterial species-specific incidence of enteric fever. To adjust for underreporting, survey data were collected from three prefectures that represent the hyper-, moderate- and low-endemic regions of Guangxi province. FINDINGS: In Guangxi province, enteric fever incidence rate varied over the study period, with a peak of 13.5 cases per 100 000 population in 1995 and a low of 6.5 in 2003. The disease occurred most frequently during the summer and autumn months and in the group aged 10-49 years. The incidence of enteric fever varied by region within Guangxi province. During the 11-year period covered by the study, 61 outbreaks of enteric fever were reported, and Salmonella paratyphi A (SPA) became the predominant causative agent in the province. CONCLUSION: Prospective studies may provide a better understanding of the reason for the shifting epidemiology of enteric fever in Guangxi province. Given the emergence of resistance to first- and second-line antimicrobials for the treatment of enteric fever, a bivalent vaccine against both SPA and S. typhi would facilitate for disease control.

[Comparative analysis of three diarrhea-household-surveys in Guangxi].

OBJECTIVE: To study the epidemiological characteristics and the effects of control measurements in Guangxi by comparing the results from diarrhea-household-surveys conducted in the three different periods of time and to develop control strategies. METHODS: Data on the incidence, health care seeking and treatment of diarrhea from three household surveys conducted in 1988, 1995 and 2007 was analyzed using SPSS (version 13.0). RESULTS: The incidence rates of diarrhea over the three periods of time were 0.562, 0.456 and 0.221 per person-year, respectively (P < 0.001). No significant difference was found in the incidence between males and females. The disease mainly attacked young age groups and those with lower educational levels. In recent years, longer durations of disease but less severe were observed in patients with diarrhea. The patients mainly visited first line health services with a health care seeking rate of 28.3%. Antibiotics were used by most of the patients (49.8% - 90.2%), while the rate of using oral rehydration salts (ORS) was only 1.4% - 11.5% but the use of traditional Chinese medicines has increased. Intake of untreated water, contaminated foods and contact with patients were important risk factors on diarrhea. CONCLUSION: The prevalence of diarrhea in Guangxi had declined and the health seeking rate was low in the past two decades. It is necessary to further regulate the treatment, in order to strengthen the health education programs to the general population, in order to improve the accessibility of health services and to increase both the health care seeking rate and effective diagnosis rate.