Extensive Comparative Genomic Analysis of Enterococcus faecalis and Enterococcus faecium Reveals a Direct Association between the Absence of CRISPR-Cas Systems, the Presence of Anti-Endonuclease (ardA) and the Acquisition of Vancomycin Resistance in E. faecium.

Here, we performed a comparative genomic analysis of all available genomes of E. faecalis (n = 1591) and E. faecium (n = 1981) and investigated the association between the presence or absence of CRISPR-Cas systems, endonuclease/anti-endonuclease systems and the acquisition of antimicrobial resistance, especially vancomycin resistance genes. Most of the analysed Enterococci were isolated from humans and less than 14% of them were from foods and animals. We analysed and detected CRISPR-Cas systems in 75.36% of E. faecalis genomes and only 4.89% of E. faecium genomes with a significant difference (p-value < 10-5). We found a negative correlation between the number of CRISPR-Cas systems and genome size (r = -0.397, p-value < 10-5) and a positive correlation between the genome %GC content and the number of CRISPR-Cas systems (r = 0.215, p-value < 10-5). Our findings showed that the presence of the anti-endonuclease ardA gene may explain the decrease in the number of CRISPR-Cas systems in E. faecium, known to deactivate the endonucleases' protective activities and enable the E. faecium genome to be versatile in acquiring mobile genetic elements, including carriers of antimicrobial resistance genes, especially vanB. Most importantly, we observed that there was a direct association between the absence of CRISPR-Cas, the presence of the anti-CRISPR ardA gene and the acquisition of vancomycin resistance genes.

HCK and ABAA: A Newly Designed Pipeline to Improve Fungi Metabarcoding Analysis.

INTRODUCTION: The fungi ITS sequence length dissimilarity, non-specific amplicons, including chimaera formed during Polymerase Chain Reaction (PCR), added to sequencing errors, create bias during similarity clustering and abundance estimation in the downstream analysis. To overcome these challenges, we present a novel approach, Hierarchical Clustering with Kraken (HCK), to classify ITS1 amplicons and Abundance-Base Alternative Approach (ABAA) pipeline to detect and filter non-specific amplicons in fungi metabarcoding sequencing datasets. MATERIALS AND METHODS: We compared the performances of both pipelines against QIIME, KRAKEN, and DADA2 using publicly available fungi ITS mock community datasets and using BLASTn as a reference. We calculated the Precision, Recall, F-score using the True-Positive, False-positive, and False-negative estimation. Alpha diversity (Chao1 and Shannon metrics) was also used to evaluate the diversity estimation of our method. RESULTS: The analysis shows that ABAA reduced the number of false-positive with all metabarcoding methods tested, and HCK increases precision and recall. HCK, coupled with ABAA, improves the F-score and bring alpha diversity metric value close to that of the BLASTn alpha diversity values when compared to QIIME, KRAKEN, and DADA2. CONCLUSION: The developed HCK-ABAA approach allows better identification of the fungi community structures while avoiding use of a reference database for non-specific amplicons filtration. It results in a more robust and stable methodology over time. The software can be downloaded on the following link: https://bitbucket.org/GottySG36/hck/src/master/.

A tuberculosis nationwide prevalence survey in Gambia, 2012.

OBJECTIVE: To estimate the population prevalence of active pulmonary tuberculosis in Gambia. METHODS: Between December 2011 and January 2013, people aged ≥ 15 years participating in a nationwide, multistage cluster survey were screened for active pulmonary tuberculosis with chest radiography and for tuberculosis symptoms. For diagnostic confirmation, sputum samples were collected from those whose screening were positive and subjected to fluorescence microscopy and liquid tuberculosis cultures. Multiple imputation and inverse probability weighting were used to estimate tuberculosis prevalence. FINDINGS: Of 100 678 people enumerated, 55 832 were eligible to participate and 43 100 (77.2%) of those participated. A majority of participants (42 942; 99.6%) were successfully screened for symptoms and by chest X-ray. Only 5948 (13.8%) were eligible for sputum examination, yielding 43 bacteriologically confirmed, 28 definite smear-positive and six probable smear-positive tuberculosis cases. Chest X-ray identified more tuberculosis cases (58/69) than did symptoms alone (43/71). The estimated prevalence of smear-positive and bacteriologically confirmed pulmonary tuberculosis were 90 (95% confidence interval, CI: 53-127) and 212 (95% CI: 152-272) per 100 000 population, respectively. Tuberculosis prevalence was higher in males (333; 95% CI: 233-433) and in the 35-54 year age group (355; 95% CI: 219-490). CONCLUSION: The burden of tuberculosis remains high in Gambia but lower than earlier estimates of 490 per 100 000 population in 2010. Less than half of all cases would have been identified based on smear microscopy results alone. Successful control efforts will require interventions targeting men, increased access to radiography and more accurate, rapid diagnostic tests.