Prevalence of blaCTX-M, blaTEM, and blaSHV Genes among Extended-spectrum β-lactamases-producing Clinical Isolates of Enterobacteriaceae in Different Regions of Sudan

Background: This study aimed to characterize blaCTX-M, blaTEM, and blaSHV genes among extended-spectrum beta-lactamases (ESBLs)-producing Enterobacteriaceae species in different regions of Sudan. Methods: In this cross-sectional study, different clinical samples (n = 985) were collected randomly from symptomatic patients from four geographical regions of Sudan and cultured on chromogenic media. Following bacterial identification, phenotypic screening of ESBLs was done according to CLSI guidelines using cefotaxime (30 μg), ceftazidime (30 μg), and cefepime (30 μg) discs with and without clavulanic acid. The DNA was extracted by guanidine hydrochloride protocol, and then conventional PCR was used to detect blaCTX-M, blaTEM, and blaSHV genes. The presence of genes’ subtypes was characterized by DNA Sanger sequencing for selected samples.  Results: Enterobacteriaceae represented 31% (305/985) of all isolates, 42 (128/305) of which were ESBLs producer, confirmed by phenotypic confirmatory test (75% [96/128] of them were positive for blaCTX-M genes, 61% [78/128] for blaTEM genes, and 38% [48/128] for blaSHV genes). Fourteen isolates (11%) were negative for all genes. Forty-eight percent (63/75) of Escherichia coli isolates were positive for blaCTX-M, while in Klebsiella pneumoniae, the dominant gene was blaTEM (82%) and had a low amount of blaSHV (59%). There was a significant association (P-value = 0.001 for all except for chloramphenicol, P = 0.014, and amikacin, P = 0.017) between resistance to third-generation cephalosporins and ciprofloxacin, nalidixic acid, meropenem, chloramphenicol, and amikacin.  Forty-two percent (40/96) of CTX-M-positive isolates were in Gizera State, 33% (32.96) in Sinnar, 24% (23/96) in Khartoum, and 1% (1/96) in White Nile.  Conclusion: We conclude that blaCTX-M genes are the most dominant genes in ESBLs-producing isolates and are more prevalent in big cities than in rural areas. Keywords: phenotypic, blaCTX-M, blaTEM, and blaSHV ESBLs genes, Enterobacteriaceae, Suda

Neutrophil elastase promotes Leishmania donovani infection via interferon-β

Visceral leishmaniasis is a deadly illness caused by Leishmania donovani that provokes liver and spleen inflammation and tissue destruction. In cutaneous leishmaniasis, the protein of L. major named inhibitor of serine peptidases (ISP2) inactivates neutrophil elastase (NE) present at the macrophage surface, resulting in blockade of TLR4 activation, prevention of TNF and IFN production and parasite survival. We report poor intracellular growth of L. donovani in macrophages from knock-out mice for NE (ela-/-), TLR4 or TLR2. NE and TLR4 co-localized with the parasite in the parasitophorous vacuole. Parasite load in the liver and spleen of ela-/- mice were reduced and accompanied by increased nitric oxide and decreased TGF production. Expression of ISP2 was not detected in L. donovani and a transgenic line constitutively expressing ISP2, displayed poor intracellular growth in macrophages and decreased burden in mice. Infected ela-/- macrophages displayed significantly lower IFN mRNA than background mice macrophages and the intracellular growth of was fully restored by exogenous IFN. We propose that L. donovani utilizes the host NE-TLR machinery to induce IFN necessary for parasite survival/growth during early infection. Low or absent expression of parasite ISP2 in L. donovani is necessary to preserve the activation of the NE-TLR pathway

Design, Development and Evaluation of rK28-Based Point-of-Care Tests for Improving Rapid Diagnosis of Visceral Leishmaniasis

Visceral Leishmaniasis caused by Leishmania donovani is endemic in several parts of South Asia, East Africa, South and Central America. It is a vector-borne disease transmitted by bites of infected sand flies and often fatal in the absence of chemotherapy. Timely diagnosis is an essential first step in providing proper patient care and in controlling transmission. VL diagnosis in East Africa and Latin America are currently based on microscopic confirmation of parasites in tissue aspirates. The Kalazar Detect rapid test is widely used as a confirmatory test in India with very high accuracy, but sensitivity issues have severely limited its usefulness in the African sub-continent. Direct Agglutination Test is another confirmatory test used widely in East Africa and offers high sensitivity but is not field-friendly. We report on the design of a novel synthetic fusion protein capable of sequestering antibodies against three different Leishmania donovani antigens and the development of point-of-care tests for improving VL diagnosis. We believe the ease of use of these rapid tests and their high accuracy in detecting VL cases could make them useful as a first-line test, thereby eliminating the need for painful biopsies and ensuring better patient care

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

Prevalence of blaCTX-M, blaTEM, and blaSHV Genes among Extended-spectrum -lactamases-producing Clinical Isolates of Enterobacteriaceae in Different Regions of Sudan

Background: This study aimed to characterize blaCTX-M, blaTEM, and blaSHV genes among extended-spectrum beta-lactamases (ESBLs)-producing Enterobacteriaceae species in different regions of Sudan. Methods: In this cross-sectional study, different clinical samples (n = 985) were collected randomly from symptomatic patients from four geographical regions of Sudan and cultured on chromogenic media. Following bacterial identification, phenotypic screening of ESBLs was done according to CLSI guidelines using cefotaxime (30 μg), ceftazidime (30 μg), and cefepime (30 μg) discs with and without clavulanic acid. The DNA was extracted by guanidine hydrochloride protocol, and then conventional PCR was used to detect blaCTX-M, blaTEM, and blaSHV genes. The presence of genes’ subtypes was characterized by DNA Sanger sequencing for selected samples. Results: Enterobacteriaceae represented 31% (305/985) of all isolates, 42 (128/305) of which were ESBLs producer, confirmed by  phenotypic confirmatory test (75% [96/128] of them were positive for blaCTX-M genes, 61% [78/128] for blaTEM genes, and 38% [48/128] for blaSHV genes). Fourteen isolates (11%) were negative for all genes. Forty-eight percent (63/75) of Escherichia coli isolates were positive for blaCTX-M, while in Klebsiella pneumoniae, the dominant gene was blaTEM (82%) and had a low amount of blaSHV (59%). There was a significant association (P-value = 0.001 for all except for chloramphenicol, P = 0.014, and amikacin, P = 0.017) between resistance to third-generation cephalosporins and ciprofloxacin, nalidixic acid, meropenem, chloramphenicol, and amikacin. Forty-two percent (40/96) of CTX-M-positive isolates were in Gizera State, 33% (32.96) in Sinnar, 24% (23/96) in Khartoum, and 1% (1/96) in White Nile. Conclusion: We conclude that blaCTX-M genes are the most dominant genes in ESBLsproducing isolates and are more prevalent in big cities than in rural areas. Keywords: phenotypic, blaCTX-M, blaTEM, and blaSHV ESBLs genes, Enterobacteriaceae, Suda

Cytokine concentrations of stimulated PBMCs of eumycetoma patients and healthy controls.

<p>(A) IL-10 concentrations (pg/ml) measured after stimulating PBMCs from eumycetoma patients and healthy endemic controls with <i>M. mycetomatis</i> antigen. Statistically significantly different concentrations were found between eumycetoma patients and healthy controls (<i>p</i> = 0.004) as determined by the Mann–Whitney <i>U</i> test. (B) IFN-γ concentrations (pg/ml) measured after stimulating PBMCs from eumycetoma patients and healthy endemic controls with <i>M. mycetomatis</i> antigen. Statistically significantly different concentrations were found between eumycetoma patients and healthy controls (<i>p</i> = 0.007) as determined by the Mann–Whitney <i>U</i> test. (C) TNF-α concentrations (pg/ml) measured after stimulating PBMCs from eumycetoma patients and healthy endemic controls with <i>M. mycetomatis</i> antigen. No statistically significantly different concentrations were found between eumycetoma patients and healthy controls (<i>p</i> = 0.85) as determined by the Mann–Whitney <i>U</i> test. (D) TGF-β concentrations (pg/ml) measured after stimulating PBMCs from eumycetoma patients and healthy endemic controls with <i>M. mycetomatis</i> antigen. No statistically significantly different concentrations were found between eumycetoma patients and healthy controls (<i>p</i> = 0.89) as determined by the Mann–Whitney <i>U</i> test.</p