Identification of the Autochaperone Domain in the Type Va Secretion System (T5aSS):Prevalent Feature of Autotransporters with a β-Helical Passenger

Autotransporters (ATs) belong to a family of modular proteins secreted by the Type V, subtype a, secretion system (T5aSS) and considered as an important source of virulence factors in lipopolysaccharidic diderm bacteria (archetypical Gram-negative bacteria). While exported by the Sec pathway, the ATs are further secreted across the outer membrane via their own C-terminal translocator forming a β-barrel, through which the rest of the protein, namely the passenger, can pass. In several ATs, an autochaperone domain (AC) present at the C-terminal region of the passenger and upstream of the translocator was demonstrated as strictly required for proper secretion and folding. However, considering it was functionally characterised and identified only in a handful of ATs, wariness recently fells on the commonality and conservation of this structural element in the T5aSS. To circumvent the issue of sequence divergence and taking advantage of the resolved three-dimensional structure of some ACs, identification of this domain was performed following structural alignment among all AT passengers experimentally resolved by crystallography before searching in a dataset of 1523 ATs. While demonstrating that the AC is indeed a conserved structure found in numerous ATs, phylogenetic analysis further revealed a distribution into deeply rooted branches, from which emerge 20 main clusters. Sequence analysis revealed that an AC could be identified in the large majority of SAATs (self-associating ATs) but not in any LEATs (lipase/esterase ATs) nor in some PATs (protease autotransporters) and PHATs (phosphatase/hydrolase ATs). Structural analysis indicated that an AC was present in passengers exhibiting single-stranded right-handed parallel β-helix, whatever the type of β-solenoid, but not with α-helical globular fold. From this investigation, the AC of type 1 appears as a prevalent and conserved structural element exclusively associated to β-helical AT passenger and should promote further studies about the protein secretion and folding via the T5aSS, especially toward α-helical AT passengers

The evolving SARS-CoV-2 epidemic in Africa: Insights from rapidly expanding genomic surveillance

INTRODUCTION Investment in Africa over the past year with regard to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) sequencing has led to a massive increase in the number of sequences, which, to date, exceeds 100,000 sequences generated to track the pandemic on the continent. These sequences have profoundly affected how public health officials in Africa have navigated the COVID-19 pandemic. RATIONALE We demonstrate how the first 100,000 SARS-CoV-2 sequences from Africa have helped monitor the epidemic on the continent, how genomic surveillance expanded over the course of the pandemic, and how we adapted our sequencing methods to deal with an evolving virus. Finally, we also examine how viral lineages have spread across the continent in a phylogeographic framework to gain insights into the underlying temporal and spatial transmission dynamics for several variants of concern (VOCs). RESULTS Our results indicate that the number of countries in Africa that can sequence the virus within their own borders is growing and that this is coupled with a shorter turnaround time from the time of sampling to sequence submission. Ongoing evolution necessitated the continual updating of primer sets, and, as a result, eight primer sets were designed in tandem with viral evolution and used to ensure effective sequencing of the virus. The pandemic unfolded through multiple waves of infection that were each driven by distinct genetic lineages, with B.1-like ancestral strains associated with the first pandemic wave of infections in 2020. Successive waves on the continent were fueled by different VOCs, with Alpha and Beta cocirculating in distinct spatial patterns during the second wave and Delta and Omicron affecting the whole continent during the third and fourth waves, respectively. Phylogeographic reconstruction points toward distinct differences in viral importation and exportation patterns associated with the Alpha, Beta, Delta, and Omicron variants and subvariants, when considering both Africa versus the rest of the world and viral dissemination within the continent. Our epidemiological and phylogenetic inferences therefore underscore the heterogeneous nature of the pandemic on the continent and highlight key insights and challenges, for instance, recognizing the limitations of low testing proportions. We also highlight the early warning capacity that genomic surveillance in Africa has had for the rest of the world with the detection of new lineages and variants, the most recent being the characterization of various Omicron subvariants. CONCLUSION Sustained investment for diagnostics and genomic surveillance in Africa is needed as the virus continues to evolve. This is important not only to help combat SARS-CoV-2 on the continent but also because it can be used as a platform to help address the many emerging and reemerging infectious disease threats in Africa. In particular, capacity building for local sequencing within countries or within the continent should be prioritized because this is generally associated with shorter turnaround times, providing the most benefit to local public health authorities tasked with pandemic response and mitigation and allowing for the fastest reaction to localized outbreaks. These investments are crucial for pandemic preparedness and response and will serve the health of the continent well into the 21st century

Multidrug resistance and extended-spectrum β-lactamases genes among Escherichia coli from patients with urinary tract infections in Northwestern Libya

Introduction: Multidrug resistance (MDR) and emergence of extended-spectrum β-lactamases (ESBLs) that mediate resistance to β-lactam drugs among Escherichia coli and other uropathogens have been reported worldwide. However, there is little information on the detection of ESBLs genes in E. coli from patients with urinary tract infections (UTIs) in the Arab countries using polymerase chain reaction (PCR), and in Libya such information is lacking. Methods: All patients attending Zawiya Teaching Hospital in Zawiya city between November 2012 and June 2013 suspected of having UTIs and from whom midstream urine samples were taken as part of the clinical workup were included in this prospective study. Samples were examined for uropathogens by standard bacteriological procedures. VITEK-2 automated microbiology system was used to identify the isolated uropathogens and determine the susceptibility of E. coli and Klebsiella spp. isolates to antimicrobials. In addition, phenotypically ESBLs-positive E. coli isolates were tested for ESBLs genes by PCR. Results: The present study enrolled 1,790 patients with UTIs. Uropathogens were found in 371 (20.7%) urine specimens examined. Mixed pathogens were detected in two specimens with 373 total pathogens isolated. E. coli and Klebsiella spp. were the predominant uropathogens at 55.8% (208/373) and 18.5% (69/373), respectively. Other pathogens were detected in 25.7% (96/373) of urine samples. Of the E. coli and Klebsiella spp. tested, 69.2 and 100% were resistant to ampicillin, 6.7 and 33.3% to ceftriaxone, and 23.1 and 17.4% to ciprofloxacin, respectively. MDR (resistance to ≥3 antimicrobial groups) was found in 69 (33.2%) of E. coli and in 29 (42%) of Klebsiella spp. isolates. ESBLs were detected phenotypically in 14 (6.7%) of E. coli and in 15 (21.7%) of Klebsiella spp. isolates. Thirteen out of the 14 phenotypically ESBL-positive E. coli were positive for ESBL genes by PCR. blaTEM gene was detected in seven isolates, blaOXA gene in 10 isolates and blaCTX-M gene in six isolates. blaSHV gene was not detected in the present study. Conclusion: The isolation of MDR ESBL-producing uropathogens undoubtedly will limit the choices clinicians have to treat their patients with UTIs. Therefore, there is an urgent need for surveillance studies on antimicrobial resistance and prevalence of ESBLs among uropathogens to guide the clinical treatment of UTIs in Libya in the future