Drug repositioning identifies salvinorin A and deacetylgedunin (DCG) enriched plant extracts as novel inhibitors of Mpro, RBD-ACE2 and TMPRRS2 proteins.

The coronavirus disease 2019 (COVID-19) has spread worldwide with severe health, social, and economic repercussions. Although vaccines have significantly reduced the severity of symptoms and deaths, alternative medications derived from natural products (NPs) are vital to further decrease fatalities, especially in regions with low vaccine uptake. When paired with the latest computational developments, NPs, which have been used to cure illnesses and infections for thousands of years, constitute a renewed resource for drug discovery. In the present report, a combination of computational and in vitro methods reveals the repositioning of NPs and identifies salvinorin A and deacetylgedunin (DCG) as having potential anti-SARS-CoV-2 activities. Salvinorin A was found both in silico and in vitro to inhibit both SARS-CoV-2 spike/host ACE2 protein interactions, consistent with blocking viral cell entry, and well as live virus replication. Plant extracts from Azadirachta indica and Cedrela odorata, which contain high levels of DCG, inhibited viral cell replication by targeting the main protease (Mpro) and/or inhibited viral cell entry by blocking the interaction between spike RBD-ACE2 protein at concentrations lower than salvinorin A. Our findings suggest that salvinorin A represent promising chemical starting points where further optimization may result in effective natural product-derived and potent anti-SARS-CoV-2 inhibitors to supplement vaccine efforts.

Whole genome-based antimicrobial resistance and virulence profiling of Staphylococcus aureus isolates from chronic leg ulcer patients in Kilimanjaro, Tanzania.

BACKGROUND: Chronic leg ulcers are hard to treat and can be a burden, particularly in resource-limited settings where diagnosis is a challenge. Staphylococcus aureus is among the common bacteria isolated from chronic wounds with a great impact on wound healing, particularly in patients with co-morbidities. Antimicrobial resistance genes and virulence factors in Staphylococcus aureus isolates were assessed to support healthcare professionals to make better therapeutic choices, and importantly to curb the development and spread of antibiotic resistance. METHODS: A cross-sectional study involved both inpatients and outpatients with chronic leg ulcers was conducted from August 2022 to April 2023 in 2 health facilities in Kilimanjaro region in Tanzania. Antimicrobial susceptibility testing was done using the disc diffusion method. Further, whole genome sequencing was performed to study the genotypic characteristics of the isolates. RESULTS: A total of 92 participants were recruited in which 9 participants were only positive for 10 Staphylococcus aureus isolates upon culture. Five STs among 9 isolates were identified. Most of them belonged to ST8 (44%), with 1 isolate does not belong to any ST. Additionally, 50% of the isolates were methicillin-resistant Staphylococcus aureus (MRSA). All S. aureus isolates had almost similar virulence factors such as hemolysin, proteases and evasions that promote toxin production, protease production and host immune evasion respectively. Moreover, all mecA positive S. aureus isolates were phenotypically susceptible to cefoxitin. CONCLUSION: Presence of mecA positive S. aureus isolates which are also phenotypically susceptible to cefoxitin implies the possibility of classifying MRSA as MSSA. This may result in the possible emergence of highly cefoxitin - resistant strains in health care and community settings when subsequently exposed to beta-lactam agents. Therefore, combination of whole genome sequencing and conventional methods is important in assessing bacterial resistance and virulence to improve management of patients.

Genetic diversity and genomic analysis of G3P[6] and equine-like G3P[8] in children under-five from Southern Highlands and Eastern Tanzania.

Rotavirus group A genomic characterization is critical for understanding the mechanisms of rotavirus diversity, such as reassortment events and possible interspecies transmission. However, little is known about the genetic diversity and genomic relationship of the rotavirus group A strains circulating in Tanzania. The genetic and genomic relationship of RVA genotypes was investigated in children under the age of five. A total of 169 Fecal samples were collected from under-five with diarrhea in Mbeya, Iringa and Morogoro regions of Tanzania. The RVA were screened in children under five with diarrhea using reverse transcription PCR for VP7 and VP4, and the G and P genotypes were determined using Sanger dideoxynucleotide cycle sequencing. Whole-genome sequencing was performed on selected genotypes. The overall RVA rate was 4.7% (8/169). The G genotypes were G3 (7/8) and G6 (1/8) among the 8 RVA positives, while the P genotypes were P[6] (4/8) and P[8] (2), and the other two were untypeable. G3P[6] and G3P[8] were the identified genotype combinations. The genomic analysis reveals that the circulating G3P[8] and G3P[6] isolates from children under the age of five with diarrhea had a DS-1-like genome configuration (I2-R2-C2-M2-axe-N2-T2-E2-H2). The phylogenic analysis revealed that all 11 segments of G3P[6] were closely related to human G3P[6] identified in neighboring countries such as Uganda, Kenya, and other African countries, implying that G3P[6] strains descended from a common ancestor. Whereas, G3P[8] were closely related to previously identified equine-like G3P[P8] from Kenya, Japan, Thailand, Brazil, and Taiwan, implying that this strain was introduced rather than reassortment events. We discovered amino acid differences at antigenic epitopes and N-linked glycosylation sites between the wild type G3 and P[8] compared to vaccine strains, implying that further research into the impact of these differences on vaccine effectiveness is warranted. The phylogenic analysis of VP7 also identified a bovine-like G6. For the first time in Tanzania, we report the emergence of novel equine-like G3 and bovine-like G6 RVA strains, highlighting the importance of rotavirus genotype monitoring and genomic analysis of representative genotypes.

Prevalence and genomic characterization of rotavirus group A genotypes in piglets from southern highlands and eastern Tanzania.

Animals have been identified as the potential reservoirs of rotavirus group A (RVA) for human infection. However, very little is known regarding the genotype and genomic profiles of circulating RVA in Tanzanian piglets. The rotavirus genetic diversity and genome analysis was assessed among piglets from Southern highlands and Eastern Tanzania. A total of 241 faecal samples were collected from piglets in the regions of Mbeya, Iringa, and Morogoro. RVA was detected and genotyped using reverse transcription polymerase chain reaction (RT-PCR). Sanger dideoxynucleotide cycle sequencing of the viral protein (VP) 4 and VP7 genes was afterwards performed to confirm the RT-PCR results. Selected genotypes were subjected to whole genome sequencing. The overall prevalence of RVA was 35.26% (85/241) in piglets (30.58% in Mbeya, 43.75% in Iringa and 31.16% in Morogoro). Upon genotyping, the G genotypes were G4 (26), G9 (10), G3 (6), G5 (3) and the remaining 40 were untypeable, while the P genotype, were P[6] (35), P[13] (3) and the remaining 47 were untypeable. The G4P[6] were the predominant genotype followed by G3P[6], G3P[13], G4P[13] and G5P[13] were most common genotypes combinations. On phylogenetic analysis, G4 was grouped to lineage V, sublineages VIIa and VIIc, G9 to lineage I, G5 to lineage II, G3 to lineage IV, P[6] to lineage V and sublineage Ic and the P[13] to lineage IV. We revealed amino acid differences between the circulating G4 and the G4 in the ProSystems RCE vaccine used in pigs. The whole genome reveals genomic constellation of G4-P[6]-I1-R1-C1-M1-A8-N1-T1-E1-H1, G5-P[x]-I5-R1-C1-M1-A8-N1-Tx-E1-H1, G3/G4-P[13]/P[6]-Ix-R1-C1-M1-A8-N1-T1-E1-H1, G3-P[6]-Ix-R1-C1-M1-A8-N1-Tx-E1-H1 and G9-P[x]-Ix-R1-C1-M1-Ax-N1-Tx-E1-H1. The VP7 gene of G9, the VP4 gene of P[6] and NSP4 (E1) gene of some genotypes clustered together and closely related to humans origin or porcine-human reassortant strains with nucleotide similarities ranging from 97.90% to 99.74% from neighboring countries, implying possibility intragenogroup reassortment and interspecies transmission. The higher strain diversity observed within the gene segments highlight the importance of genomic analysis and continuous monitoring of RVA genotypes. Further research is needed to determine the risk factors associated with RVA infection in Tanzanian pigs in order to properly design a control program.

Evidence of chikungunya virus infection among febrile patients seeking healthcare in selected districts of Tanzania.

Introduction: Chikungunya virus (CHIKV) infection is an emerging mosquito-borne disease that has been associated with frequent epidemics in the world. However, there is a dearth of information on its magnitude and associated risk factors in Tanzania. Objective: A study was conducted to determine seroprevalence of CHIKV among febrile patients seeking medical care at health facilities in Karagwe, Sengerema, Kilombero and Kyela districts. Methods: Structured questionnaires were administered and 728 serum samples were collected between May and June, 2015 and tested for the presence of CHIKV-IgM and IgG-specific antibodies using Enzyme-linked immunosorbent assay. Results and discussion: The common clinical characteristics exhibited by outpatients were fever, headache and joint pains (100%, 70%, and 68.3% respectively). Out of 728 outpatients screened for CHIKV, 105 (14%) tested CHIKV IgG positive whilst 11 (1.5%) tested CHIKV IgM positive. Chikungunya seropositivity was significantly higher than previously reported in Tanzania. The most affected age group was 20-29 years. Our results indicate that CHIKV infection is prevalent and contributes to the burden of febrile illnesses in Tanzania. The seroprevalence varies between districts, reflecting variation in mosquito vector transmission dynamics in different parts of the country. Abbreviations: CHIKV: Chikungunya virus; EDTA: Ethylenediaminetetraacetic acid; ELISA: Enzyme-linked immunosorbent assay; IgG: Immunoglobulin G; IgM: Immunoglobulin M; NIMR: National Institute for Medical Research; RU: Relative Units; SACIDS: Southern African Centre for Infectious Disease Surveillance; USA: United States of America.