Coinfections identified from metagenomic analysis of cervical lymph nodes from tularemia patients.

BACKGROUND: Underlying coinfections may complicate infectious disease states but commonly go unnoticed because an a priori clinical suspicion is usually required so they can be detected via targeted diagnostic tools. Shotgun metagenomics is a broad diagnostic tool that can be useful for identifying multiple microbes simultaneously especially if coupled with lymph node aspirates, a clinical matrix known to house disparate pathogens. The objective of this study was to analyze the utility of this unconventional diagnostic approach (shotgun metagenomics) using clinical samples from human tularemia cases as a test model. Tularemia, caused by the bacterium Francisella tularensis, is an emerging infectious disease in Turkey. This disease commonly manifests as swelling of the lymph nodes nearest to the entry of infection. Because swollen cervical nodes are observed from many different types of human infections we used these clinical sample types to analyze the utility of shotgun metagenomics. METHODS: We conducted an unbiased molecular survey using shotgun metagenomics sequencing of DNA extracts from fine-needle aspirates of neck lymph nodes from eight tularemia patients who displayed protracted symptoms. The resulting metagenomics data were searched for microbial sequences (bacterial and viral). RESULTS: F. tularensis sequences were detected in all samples. In addition, we detected DNA of other known pathogens in three patients. Both Hepatitis B virus (HBV) and Human Parvovirus B-19 were detected in one individual and Human Parvovirus B-19 alone was detected in two other individuals. Subsequent PCR coupled with Sanger sequencing verified the metagenomics results. The HBV status was independently confirmed via serological diagnostics, despite evading notice during the initial assessment. CONCLUSION: Our data highlight that shotgun metagenomics of fine-needle lymph node aspirates is a promising clinical diagnostic strategy to identify coinfections. Given the feasibility of the diagnostic approach demonstrated here, further steps to promote integration of this type of diagnostic capability into mainstream clinical practice are warranted.

Contributions of GeneXpert® to TB diagnosis in Myanmar.

BACKGROUND: Xpert® MTB/RIF, a rapid, molecular TB diagnostic assay, can detect Mycobacterium tuberculosis and rifampin resistance directly from clinical sputum samples in <2 h with high sensitivity and specificity. The added diagnostic value of Xpert over smear microscopy at a national level in Myanmar has not been previously reported.METHODS: We evaluated 339,358 Xpert and demographic records captured from January 2015 to December 2018 as part of the Myanmar National TB Program Data Utilization and Connectivity Project to examine the additional diagnostic yield of Xpert relative to smear for the detection of M. tuberculosis for TB diagnosis in Myanmar, with a focus on people living with HIV (PLHIV) and sample type.RESULTS: Use of Xpert increased TB case detection by 40% compared to smear microscopy results. Among PLHIV, use of Xpert increased TB case detection by almost 100% compared to smear microscopy results.CONCLUSION: Xpert testing identified more patients with TB than smear microscopy alone, particularly in cohorts with significant proportions of PLHIV. The use of Xpert as a screening tool in countries with a high burden of TB could lead to significantly increased diagnosis of TB at a regional and national level.

The effect of sodium thiosulfate on the recovery of Mycobacterium chimaera from heater-cooler unit water samples.

BACKGROUND: Heater-cooler units (HCUs) have been implicated in the recent global outbreak of invasive Mycobacterium chimaera infection among patients following cardiothoracic surgery. Because infected patients tend to remain asymptomatic for extended periods, detection of M. chimaera from HCUs in real time is essential to halting the ongoing M. chimaera HCU-associated outbreak. Sample collection protocols to evaluate the presence of M. chimaera offer conflicting recommendations regarding the addition of sodium thiosulfate (NaT) during the collection process. AIM: To study the effect of NaT on M. chimaera recovery and culture contamination. METHODS: Seventy-six paired HCU water samples (with and without NaT) were collected, processed and cultured simultaneously into Lowenstein-Jensen slants, Middlebrook 7H10 agar plates, and mycobacterial growth indicator tubes (MGITs), and incubated at 37°C. A subset of 31 paired samples was additionally cultured on MGITs and incubated at 30°C. FINDINGS: Of 76 samples incubated at 37°C in each of the three media, with and without NaT, M. chimaera was identified in at least one aliquot of 21 samples. CONCLUSION: The presence of NaT did not significantly increase the probability of recovering M. chimaera in a multi-variable conditional logistic model and culture contamination rates were similar between aliquots with and without NaT. In the subset of samples cultured on MGITs at both 30°C and 37°C, the presence of NaT again was not associated with M. chimaera recovery, but was significantly associated with reduced culture contamination.

Evaluation of a centrifugal blood cell processor for washing platelet concentrates.

A semiautomated saline wash procedure using a blood cell processor was evaluated as a technique for removing plasma from platelet concentrates. In vitro studies demonstrated 92 to 99.6 percent (mean, 96%) removal of total plasma protein (n = 30) with 84 to 97 percent (mean, 90.8%) platelet recovery (n = 28) in post-wash units. Post-wash pH values changed by +0.2 to -0.86 (mean, -0.47) (n = 30); the level of recovery from hypotonic shock was 69 to 97 percent (mean, 86%) (n = 11) of pre-wash units; weighted morphology scores decreased from a mean of 248 to 223 (n = 9). Aggregation response to arachidonic acid, collagen, and adenosine diphosphate plus epinephrine showed essentially no change following the wash procedure, and electron microscopy demonstrated slight morphologic alteration. Autologous platelets labeled with indium-111 demonstrated 43 +/- 20 percent recovery (n = 11) for washed units, compared to 41 +/- 10 percent for control unwashed units (n = 5); mean survivals were 140 +/- 41 hours (n = 11) for washed platelets and 185 +/- 28 hours for unwashed units (n = 5). Thirteen alloimmunized patients receiving 55 washed platelet concentrates demonstrated a mean 1- to 4-hour corrected count increment of 3.99 X 10(3) per microliter, compared to 3.02 X 10(3) per microliter for 77 unwashed platelet units given to the same patients. This study documents that platelet concentrates maintain viability and efficacy following a semiautomated saline wash method using the Cobe 2991 Blood Cell Processor, a technique that may be helpful for patients who require plasma-depleted platelet transfusions.