Amplification of 1-amino-cyclopropane-1-carboxylic (ACC) deaminase from plant growth promoting rhizobacteria in Striga-infested soil

Experiments were conducted in pots to determine the growth effect of different rhizobacteria on maize under Striga hermonthica infestation. Three bacteria were selected based on their plant growth promoting effects. Whole bacterial cells of the rhizobacteria were used to amplify 1-amino-cyclopropane-1-carboxylic acid (ACC) deaminase gene by polymerase chain reaction (PCR). Each bacterial inoculation increased agronomic characteristics of maize although not always to a statistically significant extent. The extent of growth enhancement differs between the isolates. Enterobacter sakazakii 8MR5 had the ability to stimulate plant growth, however in the PCR study, ACC deaminase was not amplified from this isolate, indicating that not all plant growth-promoting rhizobacteria contain the enzyme ACC deaminase. In contrast, an ACC deaminase specific product was amplified from Pseudomonas sp. 4MKS8 and Klebsiella oxytoca 10MKR7.  This is the first report of ACC deaminase in K. oxytoca. Key words: 1-amino-cyclopropane-1-carboxylic acid, ACC deaminase, PCR, rhizobacteria, Striga hermonthica. (African Journal of Biotechnology: 2003 2(6): 157-160

Serotype Diversity of Respiratory Human Adenoviruses amongst Pediatric Patients from Western Kenya, 2010-2012

Background: Respiratory illnesses are common among pediatric patients in Kenya, and many are attributed to viral causes. However, there is limited knowledge of the diversity of viral etiologies associated with these illnesses. Objective: To characterize respiratory adenoviruses isolates using serological and molecular approaches. Methods: A total of 1,879 samples were collected from symptomatic pediatric patients seeking medical care at New Nyanza Provincial General Hospital during the period of June 2010 to June 2012 and screened for adenoviruses as well as other respiratory viruses. Sixteen respiratory human adenoviruses (HAdVs) were isolated in Hep2 cell culture and characterized them using Immunofluorescence Assay, viral DNA amplification, sequencing and phylogenomics. Results: Phylogenetic characterization of the HAdVs using the hyper variable region 7 of the hexon gene identified HAdV B and C as the major species associated with respiratory infections during the study period. Amongst these, a single B-type and four C-type serotypes were identified.  The serotype distribution consisted of 31% HAdV B7, 25% HAdV C1, 25% HAdV C2, 6% HAdV C5, and 13% HAdV C6. Positive selection was observed in the nucleotide sequences from HAdV B7 and HAdV C5 signaling evolution of these two serotypes. Conclusion: These finding may be useful to policy makers regarding appropriate strain selection for vaccination in Kenya. Keywords: Respiratory Human adenovirus, Kenya, Pediatric, Serotype, Hexon, HVR-

The prevalence of TEM and SHV genes among Extended-Spectrum Beta-Lactamase-producing Klebsiella pneumoniae and Escherichia coli

Background: Antimicrobial resistance to cephalosporin, penicillin and aztreonam is mediated by Extended-Spectrum Beta-Lactamases (ESBL) via hydrolysis of antibiotics. The most common bacteria associated with ESBL among the Enterobacteriaceae are Escherichia coli and Klebsiella pneumonia. Pathogenic Escherichia coli is associated with diarrhoea affecting mostly elderly, children under five years and the immunocompromised. There are a number of antibiotic regimens for treatment among them cephalosporins. There is reported increase in microbial resistance to cephalosporin use and the resistance is mediated by either TEM or SHV genes. Objective: To investigate the prevalence of ESBL-producing E. coli and Klebsiella pneumonia from patients presenting with diarrhea in Machakos District Hospital, Kenya. Methods: Bacterial isolates were identified to species level by biochemical methods and tested for sensitivity to twelve different antibiotics including cephalosporins, aminoglycosides and quinolones. Those resistant to cephalosporins with a zone diameter of ≤20 mm were tested phenotypically for Extended Spectrum Beta Lactamase (ESBL) phantom development and confirmed by MicroScan. Resistant strains to cephalosporin were further tested for presence and frequency of TEM and SHV genes. Results: Out of the 200 K. pneumonia and 100 E.coli tested, 18 (6%) were positive for ESBL production phenotypically. These 18 (100 %) isolates demonstrated phantom phenomena phenotypically. Eight (4%) and 2 (1%) of the 200 K. pneumonia isolates had TEM and SHV resistant genes, respectively. There were 5 (5%) TEM and 3 (3%) SHV detected from 100 E. coli isolates. The 18 phenotypically detected and E-test-positive strains (10 Klebsiella spp. and 8 E. coli) were retested with VITEK (GNS-532 card), and 17 of these strains (94.4%) were subsequently found to be ESBL positive. One strain (5.6%) tested ESBL negative by VITEK. The cefotaxime ESBL strip detected the presence of ESBL activity in these 18 phenotypically-positive strains. Discussion: The detection of ESBL-producing E. coli and Klebsiella isolates from Machakos District Hospital was 6%. The findings point out the need for continuous surveillance to determine prevalence of ESBL-producing Enterobacteria strains for better management of diarrheal illness. Key words: Extended spectrum Beta Lactamases; Cephalosporin resistance genes; Enterobacteriacea

Molecular characterization of fluoroquinolone resistance genes in isolates obtained from patients with diarrhea in Machakos District Hospital, Kenya

Background: Diarrhea caused by Enterobacteriaceae such as Shigella species and Escherichia coli (E. coli) is endemic throughout the world, and is one of the most important causes of global childhood mortality and morbidity. There is a range of antibiotics that can be used for treatment among them quinolones. However, there is emerging increase in microbial resistance to quinolones use, with E. coli and Shigellae among the species of bacteria commonly associated with quinolone resistance. Objective: To investigate the prevalence of quinolone resistance genes in Shigellae and E. coli from patients presenting with diarrhea in Machakos District Hospital. Methods: Bacteria isolates were identified to species level by biochemical methods and serology and thereafter tested for 12 different antibiotics including quinolones, cephalosporins and aminoglycosides. Those resistant to quinolones with a zone diameter of ≤20 mm were tested for the presence of quinolone resistance genes using PCR. The gyrA resistance genes were further analyzed by sequencing to determine mutations within the quinolone resistance regions. Results: There were different E. coli pathotypes and Shigellae spp.  They resisted more than four antibiotics: Ciprofloxacin (4%), (Chloramphenical (28%), Cotrimoxazole (78%), Co-amoxilav (70%) Erythromycin (98%) Cefotoxime (18%) and Tetracycline (56%). Mutations responsible for fluoroquinolone resistance in the gyrA, gyrB, parC, and parE genes of E. coli and Shigella spp were: gyrA (17/30, 36%) gyrB (7/30, 23.3%) topoisomerase (parC 3/30, 10%) parE (3/30, 10%). Discussion: There is an increase in fluoroquinolone resistance in Shigellae and E.coli which points to a major challenge in current treatment strategies. In addition, detection of high resistance found to commonly used antibiotics should serve as a warning call for close surveillance and understanding of the epidemiology of the resistance. Key words: Quinolone antibiotics, resistance, Shigella, Escherichia col

Molecular Characterization of Human Enteroviruses Detected in Children Under Five Years Old in Kenya 2009 - 2015

   Introduction:  Human enterovirus (HEVs) infection is common, with an extensive array of clinical displays ranging from asymptomatic to life-threatening. Presentation include nonspecific febrile illness often accompanied by muscle pain, sore throat, abdominal discomfort, rash, headache, encephalitis, aseptic meningitis and acute flaccid paralysis [2]. Objectives: The study objective was to investigate the natural selection and genetic variability of HEVs and to identify HEV serotypes in circulation among children below 5 years old with diarrhea in an informal settlement(Kibera) in Kenya. Methodology: Specimens (n=628) from a prospective cohort study assessing the incidence and etiology of diarrhea from 2009-2015 were analyzed. Enteric Taqman array cards (TAC) were used for initial screening where two hundred and nine (78%) tested positive for HEVs. Of these specimens, 72 (42%) had a cycle threshold (Ct) ≤30 and were tested by conventional PCR targeting the 3’ regions of the viral protein 1 (VP1) gene. A total of 48 (67%) underwent sequencing; 11 (23%) of which yielded nucleotide sequences. Phylogenetic analyses clustered the Kenyan serotypes to HEVs groups C, B and A. Evaluation of the VP1 amino acid sequences revealed numerous amino acid substitutions in relation to reference strains, which were confirmed to be due to natural selection by negative or positive selection. Conclusion: The Heterogeneous nature of stool samples is known to influence disparities in viral nucleic acid yields. TAC detected 209 of which 171 (82%) were confirmed positive for HEVs by real-time reverse transcription polymerase chain reaction (RRT-PCR), targeting the 5’ NTR regions. Therefore, the results may not be a representative of all circulating HEVs in the study area. Since this was a retrospective study of previously collected samples, it is possible that some HEVs strains may have failed to amplify

Influenza surveillance among children with pneumonia admitted to a district hospital in coastal Kenya, 2007-2010

Background: Influenza data gaps in sub-Saharan Africa include incidence, case fatality, seasonal patterns, and associations with prevalent disorders. Methods: Nasopharyngeal samples from children aged <12 years who were admitted to Kilifi District Hospital during 2007–2010 with severe or very severe pneumonia and resided in the local demographic surveillance system were screened for influenza A, B, and C viruses by molecular methods. Outpatient children provided comparative data. Results: Of 2002 admissions, influenza A virus infection was diagnosed in 3.5% (71), influenza B virus infection, in 0.9% (19); and influenza C virus infection, in 0.8% (11 of 1404 tested). Four patients with influenza died. Among outpatients, 13 of 331 (3.9%) with acute respiratory infection and 1 of 196 without acute respiratory infection were influenza positive. The annual incidence of severe or very severe pneumonia, of influenza (any type), and of influenza A, was 1321, 60, and 43 cases per 100 000 <5 years of age, respectively. Peak occurrence was in quarters 3–4 each year, and approximately 50% of cases involved infants: temporal association with bacteremia was absent. Hypoxia was more frequent among pneumonia cases involving influenza (odds ratio, 1.78; 95% confidence interval, 1.04–1.96). Influenza A virus subtypes were seasonal H3N2 (57%), seasonal H1N1 (12%), and 2009 pandemic H1N1 (7%). Conclusions: The burden of influenza was small during 2007–2010 in this pediatric hospital in Kenya. Influenza A virus subtype H3N2 predominated, and 2009 pandemic influenza A virus subtype H1N1 had little impact

falciparum dihydrofolate reductase and

Trends in drug resistance codons in Plasmodiu

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

PGF2α Synthase-Like Proteins are Expressed in Promastigotes of Old World Leishmania Species but not in New World Species

Background: Leishmaniasis is a disease caused by protozoan parasites of the genus Leishmania and spread by the bite of infected sand fly species. The disease is characterized by an increase in prostaglandin production in the host. PGF2α is among the prostaglandins that are synthesized by Leishmania sp. Objectives: To compare the expression profiles of PGF2α synthase-like proteins in Old and New World species of Leishmania so as to provide insight into the role of these proteins. Methodology: To detect gene expression at transcription level, polymerase chain reaction was carried out using L. major PGF2α synthase gene specific primers and cDNA from L. major, L. donovani, L. tropica, L. amazonensis, L. braziliensis, L. mexicana and L. chagasi promastigotes. To detect expression at translation level, total protein from promastigotes of the above parasites was analyzed on a Western blot using T. brucei-specific rabbit anti-PGF2α synthase polyclonal antibodies. Results: At the transcription level, PGF2α synthase gene expression was detected in Old World species L. major, L. donovani and L. tropica, but was absent in the New World  L. amazonensis and L. mexicana. It was expressed at low levels in the New World L. chagasi. Western blot analysis confirmed the presence of PGF2α synthase - like proteins in Old World and not in New World species. Discussion: These findings suggest that New World Leishmania may have evolved new ortholog genes to produce PGF2α. Alternatively, the ancestral PGF2α synthase gene may be present in the New World species but has mutated or been lost due to speciation during evolution. Key words: Prostaglandins; PGF2α synthase; Leishmani