Quantifying Viable M. tuberculosis Safely Obviating the Need for High Containment Facilities.

Mycobacterium tuberculosis is an infectious pathogen that requires biosafety level-3 laboratory for handling. The risk of transmission is high to laboratory staff, and to manage the organism safely, it is necessary to construct high containment laboratory facilities at great expense. This limits the application of tuberculosis diagnostics to areas where there is insufficient capital to invest in laboratory infrastructure. In this method, we describe a process of inactivating sputum samples by either heat or guanidine thiocyanate (GTC) that renders them safe without affecting the quantification of viable bacteria. This method eliminates the need for level 3 containment laboratory for the tuberculosis molecular bacterial load assay (TB-MBLA) and is applicable in low- and middle-income countries.

Improved Diagnosis and Treatment Monitoring of Tuberculosis Using Stool and the Tuberculosis Bacterial Load Assay (TB-MBLA).

The diagnosis and monitoring of tuberculosis treatment is difficult as many patients are unable to produce sputum. This means that many patients are treated on the basis of clinical findings and consequently some will be exposed to anti-tuberculosis drugs unnecessarily. Moreover, for those appropriately on treatment and unable to produce a sputum sample, it will be impossible to monitor the response to treatment. We have shown that stool is a potential alternative sample type for diagnosis of tuberculosis. Currently, available protocols like the Xpert MTB/RIF use DNA as a target to detect Mycobacterium tuberculosis in stool but DNA survives long after the organism is dead so it is not certain whether a positive test is from an old or a partially treated infection. The TB MBLA only detects live organisms and thus, can be used to follow the response to treatment. In this chapter, we describe a protocol for TB-MBLA, an RNA-based assay, and apply it to quantify TB bacteria in stool.

Comparison of microscopic and xpert MTB diagnoses of presumptive mycobacteria tuberculosis infection: retrospective analysis of routine diagnosis at Cape Coast Teaching Hospital.

INTRODUCTION: Tuberculosis is a global health problem that causes 1. 4 million deaths every year. It has been estimated that sputum smear-negative diagnosis but culture-positive pulmonary TB diagnosis contribute to 12.6% of pulmonary TB transmission. TB diagnosis by smear microscopy smear has a minimum detection limit (LOD) of 5,000 to 10,000 bacilli per milliliter (CFU/ml) of sputum result in missed cases and false positives. However, GeneXpert technology, with a LOD of 131-250 CFU/ml in sputum samples and its implementation is believe to facilitate early detection TB and drug-resistant TB case. Since 2013, Ghana health Service (GHS) introduce GeneXpert MTB/RIF diagnostic in all regional hospitals in Ghana, however no assessment of performance between microscopy and GeneXpert TB diagnosis cross the health facilities has been reported. The study compared the results of routine diagnoses of TB by microscopy and Xpert MTB from 2016 to 2020 at the Cape Coast Teaching Hospital (CCTH). METHODS: The study compared routine microscopic and GeneXpert TB diagnosis results at the Cape Coast Teaching Hospital (CCTH) from 2016 to 2020 retrospectively. Briefly, sputum specimens were collected into 20 mL sterile screw-capped containers for each case of suspected TB infection and processed within 24 h. The samples were decontaminated using the NALC-NaOH method with the final NaOH concentration of 1%. The supernatants were discarded after the centrifuge and the remaining pellets dissolved in 1-1.5 ml of phosphate buffer saline (PBS) and used for diagnosis. A fixed smears were Ziehl-Neelsen acid-fast stain and observed under microscope and the remainings were used for GeneXpert MTB/RIF diagnosis. The data were analyze using GraphPad Prism. RESULTS: 50.11% (48.48-51.38%) were females with an odd ratio (95% CI) of 1.004 (0.944-1.069) more likely to report to the TB clinic for suspected TB diagnosis. The smear-positive cases for the first sputum were 6.6% (5.98-7.25%), and the second sputum was 6.07% (5.45-6.73%). The Xpert MTB-RIF diagnosis detected 2.93% (10/341) (1.42-5.33%) in the first and 5.44% (16/294) (3.14-8.69%) in the second smear-negative TB samples. The prevalence of Xpert MTB-RIF across smear positive showed that males had 56.87% (178/313) and 56.15% (137/244) and females had 43.13% (135/313) and 43.85% (107/244) for the first and second sputum. Also, false negative smears were 0.18% (10/5607) for smear 1 and 0.31% (16/5126) for smear 2. CONCLUSION: In conclusion, the study highlights the higher sensitivity of the GeneXpert assay compared to traditional smear microscopy for detecting MTB. The GeneXpert assay identified 10 and 16 positive MTB from smear 1 and smear 2 samples which were microscopic negative.

HAP-FAST: a feasibility study incorporating qualitative, mechanistic and costing sub-studies alongside a randomised pilot trial comparing chest x-ray to low-dose CT scan and empirical antibiotics to antibiotics guided by the BIOFIRE® FILM ARRAY® pneumonia plus panel in adults with suspected non-ventilator-associated hospital-cquired pneumonia.

INTRODUCTION: Non-ventilator-associated hospital-acquired pneumonia (nv-HAP) is the most common healthcare-associated infection (HCAI), is associated with high mortality and morbidity and places a major burden on healthcare systems. Diagnosis currently relies on chest x-rays to confirm pneumonia and sputum cultures to determine the microbiological cause. This approach leads to over-diagnosis of pneumonia, rarely identifies a causative pathogen and perpetuates unnecessary and imprecise antibiotic use. The HAP-FAST study aims to evaluate the feasibility of a randomised trial to evaluate the clinical impact of low-dose, non-contrast-enhanced thoracic CT scans and rapid molecular sputum analysis using the BIOFIRE® FILMARRAY® pneumonia plus panel (FAPP) for patients suspected with nv-HAP. METHODS AND ANALYSIS: The HAP-FAST feasibility study consists of a pilot randomised trial, a qualitative study, a costing analysis and exploratory analyses of clinical samples to investigate the immune-pathophysiology of HAP. Participants are identified and recruited from four acute hospitals in the Northwest of the UK. Using a Research Without Prior Consent model, the pilot trial will recruit 220 adult participants, with or without mental capacity, and with suspected HAP. HAP-FAST is a non-blinded, sequential, multiple assignment, randomised trial with two possible stages of randomisation: first, chest x-ray (CXR) or CT; second, if treated as nv-HAP, FAPP or standard microbiological processing alone (no FAPP). Pathogen-specific antibiotic guidance will be provided for FAPP results. Randomisation uses a web-based platform and followed up for 90 days. The feasibility of a future trial will be determined by assessing trial processes, outcome measures and patient and staff experiences. ETHICS AND DISSEMINATION: This study has undergone combined review by the UK NHS Research Ethics Committee and Health Research Authority. Results will be disseminated via peer-reviewed journals, via the funders' website and through a range of media to engage the public. TRIAL REGISTRATION NUMBER: NCT05483309.

EasyNAT MTC Assay: a Rapid Method to Detect Mycobacterium Tuberculosis in Respiratory Specimens.

BACKGROUND: This study aimed to effectively evaluate the diagnostic performance of the EasyNAT Mycobacterium tuberculosis complex (MTC) assay for tuberculosis (TB) detection from sputum. METHODS: The retrospectively analyzed data was collected from September 1, 2021, to November 1, 2023, in our hospital. RESULTS: Forty EasyNAT-positive sputum specimens were simultaneously detected using the GeneXpert MTB/ rifampicin (RIF) assay. The concordance rate between the EasyNAT and GeneXpert MTB/RIF assays was 100%. CONCLUSIONS: Because of the complexity of detecting RIF resistance data information, the rapid EasyNAT system used in conjunction with GeneXpert might be a better choice for the detection of TB in hospitals.

Rapid detection of Mycobacterium tuberculosis based on cyp141 via real-time fluorescence loop-mediated isothermal amplification (cyp141-RealAmp).

Background: The rapid detection of Mycobacterium tuberculosis (MTB) is essential for controlling tuberculosis. Methods We designed a portable thermocycler-based real-time fluorescence loop-mediated isothermal amplification assay (cyp141-RealAmp) using six oligonucleotide primers derived from cyp141 to detect MTB. A combined number of 213 sputum samples (169 obtained from clinically diagnosed cases of pulmonary TB and 44 from a control group without tuberculosis) underwent Acid-fast bacillus (AFB) smear, culture, Xpert MTB/RIF assays, and cyp141-RealAmp assay. Results: By targeting MTB cyp141, this technique could detect as low as 10 copies/reaction within 30 min, and it was successfully rejected by other mycobacteria and other bacterial species tested. Of the 169 patients, there was no statistical difference between the detection rate of cyp141-RealAmp (92.90%, 95% CI: 89.03-96.07) and that of Xpert MTB/RIF (94.67%, 95% CI: 91.28-98.06) (P > 0.05), but both were statistically higher than that of culture (65.68%, 95% CI: 58.52-72.84) (P< 0.05) and AFB (57.40%, 95% CI: 49.94-64.86) (P< 0.05). Both cyp141-RealAmp and Xpert MTB/RIF had a specificity of 100%. Furthermore, a high concordance between cyp141-RealAmp and Xpert MTB/RIF was found (Kappa = 0.89). Conclusion: The cyp141-RealAmp assay was shown to be effective, responsive, and accurate in this study. This method offers a prospective strategy for the speedy and precise detection of MTB.

Comparison of efficacies of different treatments for nontuberculous mycobacterial pulmonary disease in Anhui Province, China.

INTRODUCTION: Although nontuberculous mycobacterial (NTM) infection is a common cause of pulmonary disease worldwide, few studies have focused on epidemiological and therapeutic factors related to NTM cases in Anhui Province, China. This retrospective study aimed to identify aetiological and clinical factors, and treatment outcomes of patients with NTM pulmonary disease (NTMPD) in Anhui. METHODOLOGY: Retrospective clinical data obtained from medical records of NTMPD patients seeking care at Anhui Chest Hospital from July 2019 to June 2022 were analyzed. Treatment outcomes were compared between two patient groups: one receiving a standardised NTM treatment regimen and the other receiving precision treatment regimens. RESULTS: Genotypic analysis of 672 clinical NTMPD-associated isolates revealed that most were Mycobacterium intracellulare, while drug-susceptibility test results demonstrated diverse antibiotic resistance profiles for these isolates. Cough was the most common symptom for 101 NTMPD patients. After patients of both groups received treatment, symptoms improved, sputum culture conversion was observed for some patients, imaging findings stabilised; however, no statistically significant intergroup differences in treatment outcomes were found. CONCLUSIONS: In this study, M. intracellulare was the predominant NTM species identified in isolates obtained from NTMPD patients. Drug resistance profiles of our patient isolates were complex, highlighting the need for administration of timely, more effective, standardised treatments for patients with NTMPD in Anhui Province, China.

Are omicron infections more frequently associated with bacterial co-infections?

Introduction: Clinical observations suggest that Omicron infections may present with different radiographic findings and be more frequently associated with bacterial co-infections, but there is a paucity of published data. This study aimed to compare the clinical and radiographic findings of patients hospitalized with Omicron versus alpha-delta infections. Materials and Methods: Between January 1, 2021 and June 30, 2021 (alpha and delta period) and between January 1, 2022 and July 31, 2022 (Omicron period), respectively 149 and 163 COVID-19 PCR-positive patients who were followed up in the COVID-19 ward and intensive care unit of a tertiary care center were included in the study. Clinical (presence of fever and purulent sputum), laboratory and radiologic findings of the two groups were compared. Sputum culture results and antibiotic use were also evaluated. Result: In the alpha/delta group, ground glass opacities were seen in 75.2% (112) of the patients, consolidation in 2.7% (4), and both findings together in 6.0% (9). In the Omicron group, ground glass was seen in 40.5% (66), consolidation in 5.5% (9), and both ground glass and consolidation together in 8.7% (13) (p< 0.001). Procalcitonin levels were 0.25 µg/L or higher in 29.6% and 43.9% of the patients in the alpha/delta and Omicron groups, respectively. Mean PCT values were 0.36 µg/L and 1.93 µg/L, respectively (p> 0.05). CRP levels were similar in both groups. Mean LDH level in the Omicron group was 278 U/L and was significantly lower than the alpha/delta group (381 U/L) (p< 0.001). The proportion of patients requiring intensive care during hospitalization was higher in the alpha/delta group (36.2% vs 26.4%) (p= 0.06). Conclusions: Lower LDH levels, less need for intensive care and less frequent development of ARDS indicate that Omicron causes milder disease, while a higher rate of consolidation and higher procalcitonin levels suggest a higher frequency of bacterial co-infections.

Regional differences of Mycobacterium tuberculosis complex infection and multidrug resistance epidemic in Luoyang.

BACKGROUND: Tuberculosis (TB) remains a global public health event of great concern, however epidemic data on TB covering entire areas during the special period of the COVID-19 epidemic have rarely been reported. We compared the dissemination and multidrug-resistance patterns of Mycobacterium tuberculosis complex (MTBC) in the main urban area of Luoyang City, China (including six municipal jurisdictions) and nine county and township areas under its jurisdiction, aimed to establish the epidemiology of TB in this region and to provide reference for precision anti-TB in places with similar settings. METHODS: From 2020 to 2022, sputum samples were collected from 18,504 patients with confirmed, suspected and unexcluded TB in 10 designated TB medical institutions. Insertion sequence 6110 was amplified by PCR (rpoB gene detection if necessary) to confirm the presence of MTBC. PCR-positive specimens were analyzed by multicolor melting curve analysis to detect multidrug resistance. RESULTS: Among the 18,504 specimens, 2675 (14.5%) were MTBC positive. The positive rate was higher in the main urban area than in the county and township areas (29.8% vs. 10.9%, p < 0.001). Male, re-treated and smear-positive groups were high-burden carriers of MTBC. Individuals aged > 60 years were the largest group infected with MTBC in the main urban area, compared with individuals aged < 61 years in the county and township areas. The detection of multidrug-resistant TB (MDR-TB) was higher in the main urban area than in the county and township areas (13.9% vs. 7.8%, p < 0.001). In all areas, MDR-TB groups were dominated by males, patients with a history of TB treatment, and patients aged < 61 years. Stratified analysis of MDR-TB epidemiology showed that MDR4 (INH þ RIF þ EMB þ SM) was predominant in the main urban area, while MDR3 (INH þ RIF þ SM) was predominant in the county and township areas. MDR-TB detection rate and epidemiology differed among the county and township areas. CONCLUSIONS: For local TB control, it is necessary to plan more appropriate and accurate prevention and control strategies according to the regional distribution of MTBC infection.

Artificial intelligence-based radiographic extent analysis to predict tuberculosis treatment outcomes: a multicenter cohort study.

Predicting outcomes in pulmonary tuberculosis is challenging despite effective treatments. This study aimed to identify factors influencing treatment success and culture conversion, focusing on artificial intelligence (AI)-based chest X-ray analysis and Xpert MTB/RIF assay cycle threshold (Ct) values. In this retrospective study across six South Korean referral centers (January 1 to December 31, 2019), we included adults with rifampicin-susceptible pulmonary tuberculosis confirmed by Xpert assay from sputum samples. We analyzed patient characteristics, AI-based tuberculosis extent scores from chest X-rays, and Xpert Ct values. Of 230 patients, 206 (89.6%) achieved treatment success. The median age was 61 years, predominantly male (76.1%). AI-based radiographic tuberculosis extent scores (median 7.5) significantly correlated with treatment success (odds ratio [OR] 0.938, 95% confidence interval [CI] 0.895-0.983) and culture conversion at 8 weeks (liquid medium: OR 0.911, 95% CI 0.853-0.973; solid medium: OR 0.910, 95% CI 0.850-0.973). Sputum smear positivity was 49.6%, with a median Ct of 26.2. However, Ct values did not significantly correlate with major treatment outcomes. AI-based radiographic scoring at diagnosis is a significant predictor of treatment success and culture conversion in pulmonary tuberculosis, underscoring its potential in personalized patient management.

Nanopore sequencing for smear-negative pulmonary tuberculosis-a multicentre prospective study in China.

PURPOSE: In this prospective study, the diagnosis accuracy of nanopore sequencing-based Mycobacterium tuberculosis (MTB) detection was determined through examining bronchoalveolar lavage fluid (BALF) samples from pulmonary tuberculosis (PTB) -suspected patients. Compared the diagnostic performance of nanopore sequencing, mycobacterial growth indicator tube (MGIT) culture and Xpert MTB/rifampin resistance (MTB/RIF) assays. METHODS: Specimens collected from suspected PTB cases across China from September 2021 to April 2022 were tested then assay diagnostic accuracy rates were compared. RESULTS: Among the 111 suspected PTB cases that were ultimately diagnosed as PTB, the diagnostic rate of nanopore sequencing was statistically significant different from other assays (P < 0.05). Fleiss' kappa values of 0.219 and 0.303 indicated fair consistency levels between MTB detection results obtained using nanopore sequencing versus other assays, respectively. Respective PTB diagnostic sensitivity rates of MGIT culture, Xpert MTB/RIF and nanopore sequencing of 36.11%, 40.28% and 83.33% indicated superior sensitivity of nanopore sequencing. Analysis of area under the curve (AUC), Youden's index and accuracy values and the negative predictive value (NPV) indicated superior MTB detection performance for nanopore sequencing (with Xpert MTB/RIF ranking second), while the PTB diagnostic accuracy rate of nanopore sequencing exceeded corresponding rates of the other methods. CONCLUSIONS: In comparison with MGIT culture and Xpert MTB/RIF assays, BALF's nanopore sequencing provided superior MTB detection sensitivity and thus is suitable for testing of sputum-scarce suspected PTB cases. However, negative results obtained using these assays should be confirmed based on additional evidence before ruling out a PTB diagnosis.

Microbial community organization designates distinct pulmonary exacerbation types and predicts treatment outcome in cystic fibrosis.

Polymicrobial infection of the airways is a hallmark of obstructive lung diseases such as cystic fibrosis (CF), non-CF bronchiectasis, and chronic obstructive pulmonary disease. Pulmonary exacerbations (PEx) in these conditions are associated with accelerated lung function decline and higher mortality rates. Understanding PEx ecology is challenged by high inter-patient variability in airway microbial community profiles. We analyze bacterial communities in 880 CF sputum samples collected during an observational prospective cohort study and develop microbiome descriptors to model community reorganization prior to and during 18 PEx. We identify two microbial dysbiosis regimes with opposing ecology and dynamics. Pathogen-governed PEx show hierarchical community reorganization and reduced diversity, whereas anaerobic bloom PEx display stochasticity and increased diversity. A simulation of antimicrobial treatment predicts better efficacy for hierarchically organized communities. This link between PEx, microbiome organization, and treatment success advances the development of personalized clinical management in CF and, potentially, other obstructive lung diseases.

Current approaches for diagnosis of subclinical pulmonary tuberculosis, clinical implications and future perspectives: a scoping review.

INTRODUCTION: Subclinical tuberculosis (TB) is the presence of TB disease among people who are either asymptomatic or have minimal symptoms. AREAS COVERED: Currently, there are no accurate diagnostic tools and clear treatment approaches for subclinical TB. In this study, a comprehensive literature search was conducted across major databases. This review aimed to uncover the latest advancements in diagnostic approaches, explore their clinical implications, and outline potential future perspectives. While innovative technologies are in development to enable sputum-free TB tests, there remains a critical need for precise diagnostic tools tailored to the unique characteristics of subclinical TB. Given the complexity of subclinical TB, a multidisciplinary approach involving clinicians, microbiologists, epidemiologists, and public health experts is essential. Further research is needed to establish standardized diagnostic criteria and treatment guidelines specifically tailored for subclinical TB, acknowledging the unique challenges posed by this elusive stage of the disease. EXPERT OPINION: Efforts are needed for the detection, diagnosis, and treatment of subclinical TB. In this review, we describe the importance of subclinical TB, both from a clinical and public health perspective and highlight the diagnostic and treatment gaps of this stage.

Nocardia implantans sp. nov. isolated from a patient with pulmonary infection.

TwoGram-stain-positive and rod-shaped actinomycetes (strains CDC186T and CDC192) were isolated from sputum samples of a patient in Chongqing, PR China, and were investigated to determine their taxonomic status. The results of phylogenetic analysis based on the 16S rRNA gene indicated that CDC186T and CDC192 represented members of the genus Nocardia, and the sequence similarity with Nocardia beijingensis DSM 44636T was the highest, at 99.71 and 99.78 %, respectively. The DNA G+C content of both CDC186T and CDC192 was 69.1 %. Genomic diversity analysis revealed that the average nucleotide identity and in silico DNA‒DNA hybridisation values between the two novel strains and closely related species were significantly below the thresholds of 95-96 and 70 %, respectively, but these values between the two novel strains were 99.96 and 99.90 %, respectively. The phylogenetic relationship based on the dapb1 gene and the single-copy core genes further indicated that the two novel strains were clustered in separate branch adjacent to N. beijingensis DSM 44636T. Growth occurred within the ranges of 20-42 °C, pH 6.0-9.0 and NaCl concentrations of 0.5-4.5 % (w/v). The major fatty acids of CDC186T and CDC192 were C16 : 0 and C18 : 0 10-methyl [tuberculostearic acid (TBSA)]. The predominant respiratory menaquinone was MK-9. The polar lipid profile contained diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylinositol mannoside, one unidentified glycolipid, one unidentified phospholipid and one unidentified phosphoglycolipid. All the genomes of the studied strains were annotated with virulence factor (VF)-associated genes homologous to those of Mycobacterium tuberculosis, and the results of susceptibility testing indicated that CDC186T and CDC192 were resistant to amoxicillin-clavulanic acid and tigecycline. On the basis of chemotaxonomic characteristics and the results of phylogenetic analyses, strains CDC186T and CDC192 represent a novel species within the genus Nocardia, for which the name Nocardia implantans sp. nov. is proposed. The type strain is CDC186T (=GDMCC 4.206T= JCM 34959T).

Impact of diurnal temperature variations on sputum bacterial detection in hospitalized patients with acute COPD exacerbation: a retrospective study from Fuzhou, China.

OBJECTIVE: To investigate the association between meteorological data three days before admission and the status of sputum pathogens culture in hospitalized patients with Acute exacerbation of Chronic obstructive pulmonary disease (AECOPD) and respiratory infections. METHODS: Data from 1,370 AECOPD patients (80.66% males, approximately 80% age > 70) with respiratory infections hospitalized in Fujian Provincial Hospital between December 2013 and December 2019 were collected. This cohort comprised, along with concurrent meteorological data from Fuzhou. Group differences were analyzed to compare the meteorological data three days prior to admission between patients with positive sputum pathogen cultures and those without. Logistic regression models were employed to investigate the association between meteorological parameters and the status of sputum pathogen cultures in patients with AECOPD and respiratory infections. Sensitivity analyses was conducted among the hospitalized patients from 2013 to 2016 and 2017-2019. Stratified analysis was performed to explore the factors affecting the effect of temperature differences and their interactions. RESULTS: 578(42.19%) cases had a positive sputum culture report indicating pathogen growth. 323 cases were found with Gram-negative bacteria, 160 with Gram-positive bacteria, and 114 with fungi. Uni-variate analysis revealed statistical differences in DTD three days prior to admission (DTD-3d) between the positive and negative sputum culture groups (p = 0.019). Multivariate analysis indicated that an increase in the risk of positive sputum pathogen cultures was associated with greater DTD three days before admission (DTD-3d), with OR1.657 (95%CI [ 1.328-1.981]). The risk of positive sputum pathogen cultures was higher in groups with greater DTD-3d. The findings were consistent across different admission periods. Stratified analysis showed that patients without respiratory failure were more affected by DTD-3d, and an interaction effect was observed (p < 0.001). CONCLUSION: In coastal areas, the diurnal temperature difference three days prior to admission affects the sputum pathogen status in AECOPD patients with respiratory infections.

The etiological diagnostic value of metagenomic next-generation sequencing in suspected community-acquired pneumonia.

BACKGROUND: The emergence of metagenomic next-generation sequencing (mNGS) may provide a promising tool for early and comprehensive identification of the causative pathogen in community-acquired pneumonia (CAP). In this study, we aim to further evaluate the etiological diagnostic value of mNGS in suspected CAP. METHODS: A total of 555 bronchoalveolar lavage fluid (BALF) samples were collected for pathogen detection by mNGS from 541 patients with suspected CAP. The clinical value was assessed based on infection diagnosis and treatment guidance. The diagnostic performance for pathogen identification by mNGS and sputum culture and for tuberculosis (TB) by mNGS and X-pert MTB/RIF were compared. To evaluate the potential for treatment guidance, we analyzed the treatment regimen of patients with suspected CAP, including imaging changes of lung after empirical antibacterial therapy, intensified regimen, antifungal treatment, and a 1-year follow up for patients with unconfirmed diagnosis and non-improvement imaging after anti-infective treatment and patients with high suspicion of TB or NTM infection who were transferred to the Wuhan Pulmonary Hospital for further diagnosis and even anti-mycobacterium therapy. RESULTS: Of the 516 BALF samples that were analyzed by both mNGS and sputum culture, the positivity rate of mNGS was significantly higher than that of sputum culture (79.1% vs. 11.4%, P = 0.001). A total of 48 samples from patients with confirmed TB were analyzed by both mNGS and X-pert MTB/RIF, and the sensitivity of mNGS for the diagnosis of active TB was significantly lower than that of X-pert MTB/RIF (64.6% vs. 85.4%, P = 0.031). Of the 106 pathogen-negative cases, 48 were ultimately considered non-infectious diseases, with a negative predictive value of 45.3%. Of the 381 pathogen-positive cases, 311 were eventually diagnosed as CAP, with a positive predictive value of 81.6%. A total of 487 patients were included in the evaluation of the therapeutic effect, and 67.1% improved with initial empirical antibiotic treatment. Of the 163 patients in which bacteria were detected, 77.9% improved with antibacterial therapy; of the 85 patients in which fungi were detected, 12.9% achieved remission after antifungal therapy. CONCLUSIONS: Overall, mNGS had unique advantages in the detection of suspected CAP pathogens. However, mNGS was not superior to X-pert MTB/RIF for the diagnosis of TB. In addition, mNGS was not necessary as a routine test for all patients admitted with suspected CAP. Furthermore, when fungi are detected by mNGS, antifungal therapy should be cautious.

Spectrum and antimicrobial resistance in acute exacerbation of chronic obstructive pulmonary disease with pneumonia: a cross-sectional prospective study from Vietnam.

BACKGROUND: Respiratory infections have long been recognized as a primary cause of acute exacerbation of chronic obstructive pulmonary disease (AE-COPD). Additionally, the emergence of antimicrobial resistance has led to an urgent and critical situation in developing countries, including Vietnam. This study aimed to investigate the distribution and antimicrobial resistance of bacteria in patients with AE-COPD using both conventional culture and multiplex real-time PCR. Additionally, associations between clinical characteristics and indicators of pneumonia in these patients were examined. METHODS: This cross-sectional prospective study included 92 AE-COPD patients with pneumonia and 46 without pneumonia. Sputum specimens were cultured and examined for bacterial identification, and antimicrobial susceptibility was determined for each isolate. Multiplex real-time PCR was also performed to detect ten bacteria and seven viruses. RESULTS: The detection rates of pathogens in AE-COPD patients with pneumonia were 92.39%, compared to 86.96% in those without pneumonia. A total of 26 pathogenic species were identified, showing no significant difference in distribution between the two groups. The predominant bacteria included Klebsiella pneumoniae, Haemophilus influenzae, Moraxella catarrhalis, and Streptococcus pneumoniae, followed by Acinetobacter baumannii and Streptococcus mitis. There was a slight difference in antibiotic resistance between bacteria isolated from two groups. The frequency of H. influenzae was notably greater in AE-COPD patients who experienced respiratory failure (21.92%) than in those who did not (9.23%). S. pneumoniae was more common in patients with stage I (44.44%) or IV (36.36%) COPD than in patients with stage II (17.39%) or III (9.72%) disease. ROC curve analysis revealed that C-reactive protein (CRP) levels could distinguish patients with AE-COPD with and without pneumonia (AUC = 0.78). CONCLUSION: Gram-negative bacteria still play a key role in the etiology of AE-COPD patients, regardless of the presence of pneumonia. This study provides updated evidence for the epidemiology of AE-COPD pathogens and the appropriate selection of antimicrobial agents in Vietnam.

Relationship of neutrophil lymphocyte ratio, monocyte lymphocyte ratio and neutrophil monocyte ratio with treatment response in pulmonary tuberculosis patients during intensive phase treatment.

OBJECTIVE: To determine the relationship of Neutrophil Lymphocyte Ratio (NLR), Monocyte Lymphocyte Ratio (MLR), and Neutrophil Monocyte Ratio (NMR) with treatment response in Pulmonary Tuberculosis (PTB) patients during intensive phase treatment (IPT). METHODS: This analytical cross-sectional study was conducted at Ojha Institute of Chest Diseases (OICD), Dow University of Health Sciences, from February to December 2021. 100 patients were enrolled using purposive sampling technique. Both male and female of age 18 and above, rifampicin sensitive newly diagnosed cases of PTB by Acid Fast Bacilli (AFB) microscopy and Gene Xpert MTB/RIF were included. SPSS version 26 was used to analyze data. Numerical data was expressed in median and interquartile range and categorical data was expressed in frequencies and percentages. RESULTS: Out of total 100 patients, 81% (n = 81) showed treatment response with negative AFB Sputum Smear Microscopy (SSM) after 2nd month. Out of 81% (n = 81) of the patients who achieved treatment response, 83.9% (n = 68) also had decreased NLR, 85.2% (n = 69) had decreased MLR and 83.9% (n = 68) had decreased NMR from baseline. However 19% (n = 19) did not achieved treatment response with positive AFB SSM after 2nd month of ATT (Anti tuberculosis treatment), among them 10.52% (n = 2) were INH resistant with no decrease in all the ratios after 2nd month. CONCLUSION: Leukocyte ratios decreased significantly from baseline as PTB was treated in patients who achieved treatment response with negative AFB SSM after two months of ATT and hence these ratios could be used as markers to monitor the treatment response.

Implication of Negative GeneXpert Mycobacterium tuberculosis/Rifampicin Results in Suspected Tuberculosis Patients: A Research Study.

OBJECTIVE: GeneXpert Mycobacterium tuberculosis/rifampicin (MTB/RIF) is a conceptually helpful tool for establishing tuberculosis (TB) disease. Negative results from the GeneXpert test do not exclude the possibility of diagnosing non-tuberculous mycobacteria lung disease (NTMLD) as a chronic pulmonary disease. When a patient is diagnosed on a clinical basis, and there is no bacteriological evidence of TB, it is necessary to consider NTM as one of the causes of disease with TB-like symptoms. The prevalence of non-tuberculous mycobacteria (NTM) disease is rising globally, but its diagnosis is still delayed and often misdiagnosed as multidrug-resistant TB (MDR-TB). This study highlights the implication of negative GeneXpert MTB/RIF results in suspected TB patients who conducted mycobacteria culture and detected the incidence of NTMLD. METHODS: In this experimental study, the performance of GeneXpert MTB/RIF-negative results with those of mycobacteria cultures and lung abnormalities among suspected TB patients in a referral hospital in Indonesia were evaluated. From January to August 2022, 100 sputum samples from suspected chronic pulmonary TB patients with GeneXpert MTB/RIF assay-negative results were cultured in Lowenstein-Jensen medium, and the implication among negative GeneXpert result MTB/RIF assay. RESULTS: 7% were confirmed to have MTB and 1% had NTM by culture assay. Moreover, 34% were diagnosed with clinical TB and treated with anti-TB drugs. CONCLUSION: For patients with negative assay results of GeneXpert MTB/RIF regarding clinically suspected chronic TB infection, further diagnostic tests to determine the causative agents of the lung abnormalities should be carried out.

Fatal Nocardia paucivorans Lower Respiratory Tract Infection in a Tuberculosis Suspect Who Worked as Gold Miner.

Nocardiosis is an opportunistic infection that affects both immunocompromised as well as immunocompetent patients. The main infections occur as soft tissue and lung infections although they might disseminate to various organs. This is a case study aimed to reflect the severity of the disease and the patient's risk factors associated with the infection. A sputum sample was collected from tuberculosis (TB) suspects for culture. Nocardia-like colonies were isolated, purified, and sent to BGI Company (Hongkong, China). Standard forward sequencing of 16S rRNA was done by ABI Genetic Analyzer (Applied Biosystems). Sequence alignment and nucleotide basic local alignment search tool (BLAST) were done using National Center for Bioinformatics (NCBI) Nucleotide BLAST. Biochemical identification to the colonies was done using an automation system (BD Phoenix™) to confirm the identification. Nocardia paucivorans was identified from the TB suspect. Risk factors were identified as extensive contact to dust, absence of primary care units with complete facilities, and old age. Since the infection of the lungs caused by Nocardia might be similar to pulmonary TB, this case report highlights the importance of accurate diagnosis and identification procedures to differentiate between the two.