Sub-acute and sub-chronic toxicity assessment of the antimicrobial peptide Dermaseptin B2 on biochemical, haematological and histopathological parameters in BALB/c mice and Albino Wistar rats.

Background: Dermaseptins (Drs) are peptides found in the skin secretions of a variety of Hylid frogs, particularly those belonging to the Agalychnis and Phyllomedusa families. Dermaseptin B2 (Drs B2), an amphipathic, α-helical polypeptide was reported as the most active of the Dermaseptin B family. We have previously shown that Drs B2 has strong anti-proliferative activities against RD cells in vitro and thus required further evaluations for future medical applications. Aim: The aim the study was to evaluate the 14-day sub-acute and 90-day sub-chronic toxicities Drs B2 in vivo. Materials and Methods: BALB/c mice were treated with increasing concentrations of 5-25 mg/kg of Drs B2. Rats were treated with 2, 4 and 10-fold concentrations of the calculated LD50 of Drs B2 following OECD recommendations. At the end of the experimentation periods, the animals were sacrificed and dissected to collect blood and selected organs for analysis of any effects caused by Drs B2 treatment on the biochemical, haematological, and histological parameters. Results: The 14-day sub-acute toxicity tests did not cause significant alteration in the biochemical, hematological and histological parameters. The 90-day sub-chronic toxicity study showed lower ALT and AST than control at doses 1.9 mg/kg and 4.6 mg/kg, respectively. Their haematology results also showed higher platelet count than the controls but the differences were not statistically significant. Histological analysis showed increased megakaryocytes in the spleen for both the mice and the rats. Conclusion: The results of this study indicate that short term treatment of Drs B2 could be safe to the animals, however, long-term treatment can have mild effects on the liver parameters and cause an inflammatory response in the spleen.

Whole-genome sequence analysis reveals the circulation of multiple SARS-CoV-2 variants of concern in Nairobi and neighboring counties, Kenya between March and July 2021.

The emergence and rapid spread of SARS-CoV-2 variants of concern (VOC) have been linked to new waves of COVID-19 epidemics occurring in different regions of the world. The VOC have acquired adaptive mutations that have enhanced virus transmissibility, increased virulence, and reduced response to neutralizing antibodies. Kenya has experienced six waves of COVID-19 epidemics. In this study, we analyzed 64 genome sequences of SARS-CoV-2 strains that circulated in Nairobi and neighboring counties, Kenya between March 2021 and July 2021. Viral RNA was extracted from RT-PCR confirmed COVID-19 cases, followed by sequencing using the ARTIC network protocol and Oxford Nanopore Technologies. Analysis of the sequence data was performed using different bioinformatics methods. Our analyses revealed that during the study period, three SARS-CoV-2 variants of concern (VOC) circulated in Nairobi and nearby counties in Kenya. The Alpha (B.1.1.7) lineage predominated (62.7%), followed by Delta (B.1.617.2, 35.8%) and Beta (B.1.351, 1.5%). Notably, the Alpha (B.1.1.7) VOC were most frequent from March 2021 to May 2021, while the Delta (B.1.617.2) dominated beginning June 2021 through July 2021. Sequence comparisons revealed that all the Kenyan viruses were genetically similar to those that circulated in other regions. Although the majority of Kenyan viruses clustered together in their respective phylogenetic lineages/clades, a significant number were interspersed among foreign strains. Between March and July 2021, our study's findings indicate the prevalence of multiple lineages of SAR-CoV-2 VOC in Nairobi and nearby counties in Kenya. The data suggest that the recent increase in SARS-CoV-2 infection, particularly in Nairobi and Kenya as a whole, is attributable to the introduction and community transmission of SARS-CoV-2 VOC among the populace. In conclusion, the findings provide a snapshot of the SARS-CoV-2 variants that circulated in Kenya during the study period.

Spatial and Spatio-Temporal Distribution of Human Respiratory Syncytial Virus, Human Parainfluenza Virus, and Human Adenoviruses Cases in Kenya 2007-2013.

Background: Human Respiratory Syncytial Virus (HRSV), Human Parainfluenza Virus (HPIV), and Human Adenovirus (HAdV) epidemics differ in geographical location, time, and virus type. Regions prone to infections can be identified using geographic information systems (GIS) and available methods for detecting spatial and time clusters. We sought to find statistically significant spatial and time clusters of HRSV, HPIV, and HAdV cases in different parts of Kenya. Methods: To analyse retrospective data, we used a geographical information system (GIS) and the spatial scan statistic. The information was gathered from surveillance sites and aggregated at the county level in order to identify purely spatial and Spatio-temporal clusters. To detect the presence of spatial autocorrelation, the local Moran's I test was used. To detect the spatial clusters of HRSV, HPIV, and HAdV cases, we performed the purely spatial scan statistic. Furthermore, space-time clusters were identified using space-time scan statistics. Both spatial and space-time analyses were based on the discrete Poisson model with a pre-specified statistical significance levelof p<0.05. Results: The findings showed that HRSV, HPIV, and HAdV cases had significant autocorrelation within the study areas. Furthermore, in the Western region of the country, the three respiratory viruses had local clusters with significant positive autocorrelation (p<0.05). Statistically, the Western region had significant spatial clusters of HRSV, HPIV, and HAdV occurrence. Furthermore, the space-time analysis revealed that the HPIV primary cluster persisted in the Western region from 2007 to 2013. However, primary clusters of HRSV and HAdV were observed in the Coastal region in 2009-11 and 2008-09, respectively. Conclusion: Human respiratory syncytial virus (HRSV), human parainfluenza virus (HPIV), and human adenovirus (HAdV) hotspots (clusters) occurred in Kenya's Western and Coastal regions from 2007 to 2013. The Western region appeared to be more prone to the occurrence of allthree respiratory viruses throughout the study period. Strategic mitigation should focus on these locations to prevent future clusters of HRSV, HPIV, and HAdV infections that could lead to epidemics.

Effects of cytochrome P450 2B6 and constitutive androstane receptor genetic variation on Efavirenz plasma concentrations among HIV patients in Kenya.

The effects of genetic variation of cytochrome P450 2B6 (CYP2B6) and constitutive androstane receptor (CAR) on efavirenz (EFV) plasma concentration was evaluated among 312 HIV patients in Nairobi Kenya. The EFV plasma concentration at steady-state were determined using ultra-high-performance liquid chromatography with a tandem quadruple mass spectrometer (LC-MS/MS). Thirteen CYP2B6 (329G>T, 341T>C, 444 G>T/C, 15582C>T, 516G>T, 548T>G, 637T>C, 785A>G, 18492C>T, 835G>C, 1459C>T and 21563C>T) and one CAR (540C>T) single nucleotide polymorphisms (SNPs) were genotyped using real-time polymerase chain reaction. HIV drug resistance mutations were detected using an in-house genotypic assay. The EFV concentration of patients ranged from 4 ng/mL to 332697 ng/mL (median 2739.5 ng/mL, IQR 1878-4891.5 ng/mL). Overall, 22% patients had EFV concentrations beyond therapeutic range of 1000-4000 ng/mL (4.5%% < 1000 ng/mL and 31.7% > 4000 ng/mL). Five SNPs (15582C>T, 516G>T, 785A>G, 983T>C and 21563C>T) were associated with higher EFV plasma concentration while 18492C>T with lower EFV plasma concentration (p<0.05). Strong linkage disequilibrium (LD) was observed for 15582C>T, 516G>T, 785A>G, 18492C>T, 983T>C, 21563C>T, 1459C>T and CAR 540C>T. Sixteen haplotypes were observed and CTGCTTCC, CTGCTTCT, TTGCTTCT and CGACCCCT were associated with high EFV plasma concentration. In multivariate analysis, factors significantly associated with EFV plasma concentration included; the presence of skin rash (ß = 1379, 95% confidence interval (CI) = 3216.9-3416.3; p < 0.039), T allele of CYP2B6 516G>T (ß = 1868.9, 95% CI 3216.9-3416.3; p < 0.018), the C allele of CYP2B6 983T>C (ß = 2638.3, 95% CI = 1348-3929; p < 0.0001), T allele of CYP2B6 21563C>T (ß = 1737, 95% CI = 972.2-2681.9; p < 0.0001) and the presence of 5 to 7 numbers of SNPs per patient (ß = 570, 95% CI = 362-778; p < 0.0001) and HIV viral load ≤1000 cells/mL (ß = -4199.3, 95% CI = -7914.9 --483.6; p = 0.027). About 36.2% of the patients had EFV plasma concentrations beyond therapeutic window, posing high risk of treatment failure or toxicity. The SNPs of CYP2B6 516G>T, CYP2B6 983T>C, 21563C>T, presence of higher numbers of SNPs per patient and haplotypes CTGCTTCC, CTGCTTCT, TTGCTTCT and CGACCCCT could efficiently serves as genetic markers for EFV plasma concentration and could guide personalization of EFV based ART treatment in Kenya.

Spatial-temporal distribution and sequence diversity of group a human respiratory syncytial viruses in Kenya preceding the emergence of ON1 genotype.

BACKGROUND: Human respiratory syncytial virus (HRSV) is a major cause of severe viral acute respiratory illness and contributes significantly to severe pneumonia cases in Africa. Little is known about its spatial-temporal distribution as defined by its genetic diversity. METHODS: A retrospective study conducted utilizing archived nasopharyngeal specimens from patients attending outpatient clinics in hospitals located in five demographically and climatically distinct regions of Kenya; Coast, Western, Highlands, Eastern and Nairobi. The viral total RNA was extracted and tested using multiplex real time RT-PCR (reverse transcriptase polymerase chain reaction). A segment of the G-gene was amplified using one-step RT-PCR and sequenced by Sanger di-deoxy method. Bayesian analysis of phylogeny was utilized and subsequently median joining methods for haplotype network reconstruction. RESULTS: Three genotypes of HRSVA were detected; GA5 (14.0%), GA2 (33.1%), and NA1 (52.9%). HRSVA prevalence varied by location from 33% to 13.2% in the Highlands and the Eastern regions respectively. The mean nucleotide diversity (Pi[π]) varied by genotype: highest of 0.018 for GA5 and lowest of 0.005 for NA1. A total of 58 haplotypes were identified (GA5 10; GA2 20; NA1 28). These haplotypes were introduced into the population locally by single haplotypes and additional subsidiary seeds amongst the GA2 and the NA1 haplotypes. CONCLUSIONS: HRSVA was found across all the regions throughout the study period and comprised three genotypes; GA5, GA2, and NA1 genotypes. The genotypes were disproportionately distributed across the regions with GA5 gradually increasing toward the Western zones and decreasing toward the Eastern zones of the country.

Mutation patterns of resistance genes for macrolides, aminoglycosides, and rifampicin in non-tuberculous mycobacteria isolates from Kenya.

BACKGROUND: Non-tuberculous mycobacteria (NTM) treatment constitutes a macrolide-based antibiotic regimen in combination with aminoglycosides for Rapid-Growing Mycobacteria (RGM), and rifampicin for Slow-Growing Mycobacteria (SGM). Mutations in the anti-NTM drug target regions promote NTM evolution to mutant strains that are insusceptible to NTM drugs leading to treatment failure. We, therefore, described the mutation patterns of anti-NTM drug target genes including rrl, rrs, and rpoB in NTM isolates from Kenya.  Methods: We carried out a cross-sectional study that included 122 NTM obtained from the sputum of symptomatic tuberculosis-negative patients in Kenya. All 122 NTM underwent targeted sequencing of the rrl gene. The 54 RGM were also sequenced for rrs, and the 68 SGM were sequenced for rpoB genes using ABI 3730XL analyzer. The obtained sequences were aligned to their wild-type reference sequences for each gene using Geneious then mutations were identified. Pearson chi-square at a 95% confidence interval tested the association of NTM to mutation patterns for each gene. RESULTS: NTM harboring mutations associated with resistance to at least one of the antibiotics used in the macrolide-based therapy were 23% (28/122). Of these NTM, 10.4% (12/122) had mutations in the rrl gene with 58.3% (7/12) comprising RGM and 41.7% (5/12) being SGM. Mutation at position 2058 (A2058G, A2058C, A2058T) of the rrl gene was seen for 83.3% (10/12) of NTM, while 16.6% (2/12) harbored a A2059G mutation. Of the 54 RGM included for rrs characterization, 11.1% (6/54) exhibited mutations  at position 1408(A1408G), while 14.7% (10/68) of the SGM had mutations in the rpoB gene at positions S531W, S531L, S531Y, F506L, E509H with M.gastri having multiple mutations at positions D516V, H526D and, S531F. CONCLUSION: We demonstrated a significant level of mutations associated with drug resistance for macrolides, aminoglycosides, and rifampicin in NTM isolated from symptomatic TB negative patients in Kenya.

Association between social psychological status and efavirenz and nevirapine plasma concentration among HIV patients in Kenya.

HIV-related stigma, lack of disclosure and social support are still hindrances to HIV testing, care, and prevention. We assessed the association of these social-psychological statuses with nevirapine (NVP) and efavirenz (EFV) plasma concentrations among HIV patients in Kenya. Blood samples were obtained from 254 and 312 consenting HIV patients on NVP- and EFV-based first-line antiretroviral therapy (ART), respectively, and a detailed structured questionnaire was administered. The ARV plasma concentration was measured by liquid chromatography-tandem mass spectrometry (LC-MS/MS). There were 68.1% and 65.4% of the patients on NVP and EFV, respectively, who did not feel guilty for being HIV positive. The disclosure rates were approximately 96.1% and 94.6% of patients on NVP and EFV, respectively. Approximately 85% and 78.2% of patients on NVP and EFV, respectively, received social support as much as needed. There were 54.3% and 14.2% compared to 31.7% and 4.5% patients on NVP and EFV, respectively, with supratherapeutic and suboptimal plasma concentrations. Multivariate quantile regression analysis showed that feeling guilty for being HIV positive was associated with increased 954 ng/mL NVP plasma concentrations (95% CI 192.7 to 2156.6; p = 0.014) but not associated with EFV plasma concentrations (adjusted ß = 347.7, 95% CI = - 153.4 to 848.7; p = 0.173). Feeling worthless for being HIV positive was associated with increased NVP plasma concentrations (adjusted ß = 852, 95% CI = 64.3 to 1639.7; p = 0.034) and not with EFV plasma concentrations (adjusted ß = - 143.3, 95% CI = - 759.2 to 472.5; p = 0.647). Being certain of telling the primary sexual partner about HIV-positive status was associated with increased EFV plasma concentrations (adjusted ß 363, 95% CI, 97.9 to 628.1; p = 0.007) but not with NVP plasma concentrations (adjusted ß = 341.5, 95% CI = - 1357 to 2040; p = 0.692). Disclosing HIV status to neighbors was associated with increased NVP plasma concentrations (adjusted ß = 1731, 95% CI = 376 to 3086; p = 0.012) but not with EFV plasma concentrations (adjusted ß = - 251, 95% CI = - 1714.1 to 1212.1; p = 0.736). Obtaining transportation to the hospital whenever needed was associated with a reduction in NVP plasma concentrations (adjusted ß = - 1143.3, 95% CI = - 1914.3 to - 372.4; p = 0.004) but not with EFV plasma concentrations (adjusted ß = - 6.6, 95% CI = - 377.8 to 364.7; p = 0.972). HIV stigma, lack disclosure and inadequate social support are still experienced by HIV-infected patients in Kenya. A significant proportion of patients receiving the NVP-based regimen had supra- and subtherapeutic plasma concentrations compared to EFV. Social-psychological factors negatively impact adherence and are associated with increased NVP plasma concentration compared to EFV.

Profiling of RNA Viruses in Biting Midges (Ceratopogonidae) and Related Diptera from Kenya Using Metagenomics and Metabarcoding Analysis.

Vector-borne diseases (VBDs) cause enormous health burden worldwide, as they account for more than 17% of all infectious diseases and over 700,000 deaths each year. A significant number of these VBDs are caused by RNA virus pathogens. Here, we used metagenomics and metabarcoding analysis to characterize RNA viruses and their insect hosts among biting midges from Kenya. We identified a total of 15 phylogenetically distinct insect-specific viruses. These viruses fall into six families, with one virus falling in the recently proposed negevirus taxon. The six virus families include Partitiviridae, Iflaviridae, Tombusviridae, Solemoviridae, Totiviridae, and Chuviridae. In addition, we identified many insect species that were possibly associated with the identified viruses. Ceratopogonidae was the most common family of midges identified. Others included Chironomidae and Cecidomyiidae. Our findings reveal a diverse RNA virome among Kenyan midges that includes previously unknown viruses. Further, metabarcoding analysis based on COI (cytochrome c oxidase subunit 1 mitochondrial gene) barcodes reveal a diverse array of midge species among the insects used in the study. Successful application of metagenomics and metabarcoding methods to characterize RNA viruses and their insect hosts in this study highlights a possible simultaneous application of these two methods as cost-effective approaches to virus surveillance and host characterization. IMPORTANCE The majority of the viruses that currently cause diseases in humans and animals are RNA viruses, and more specifically arthropod-transmitted viruses. They cause diseases such as dengue, West Nile infection, bluetongue disease, Schmallenberg disease, and yellow fever, among others. Several sequencing investigations have shown us that a diverse array of RNA viruses among insect vectors remain unknown. Some of these could be ancient lineages that could aid in comprehensive studies on RNA virus evolution. Such studies may provide us with insights into the evolution of the currently pathogenic viruses. Here, we applied metagenomics to field-collected midges and we managed to characterize several RNA viruses, where we recovered complete and nearly complete genomes of these viruses. We also characterized the insect host species that are associated with these viruses. These results add to the currently known diversity of RNA viruses among biting midges as well as their associated insect hosts.

Diagnostic performance of a colorimetric RT -LAMP for the identification of SARS-CoV-2: A multicenter prospective clinical evaluation in sub-Saharan Africa.

BACKGROUND: Management and control of the COVID-19 pandemic caused by the severe acute respiratory syndrome coronavirus SARS-CoV-2 is critically dependent on quick and reliable identification of the virus in clinical specimens. Detection of viral RNA by a colorimetric reverse transcription loop-mediated isothermal amplification (RT-LAMP) is a simple, reliable and cost-effective assay, deployable in resource-limited settings (RLS). Our objective was to evaluate the intrinsic and extrinsic performances of RT-LAMP in RLS. METHODS: This is a multicenter prospective observational study of diagnostic accuracy, conducted from October 2020 to February 2021 in four African Countries: Cameroon, Ethiopia, Kenya and Nigeria; and in Italy. We enroled 1657 individuals who were either COVID-19 suspect cases, or asymptomatic and presented for screening. RNA extracted from pharyngeal swabs was tested in parallel by a colorimetric RT-LAMP and by a standard real time polymerase chain reaction (RT-PCR). FINDINGS: The sensitivity and specificity of index RT LAMP compared to standard RT-PCR on 1657 prospective specimens from infected individuals was determined. For a subset of 1292 specimens, which underwent exactly the same procedures in different countries, we obtained very high specificity (98%) and positive predictive value (PPV = 99%), while the sensitivity was 87%, with a negative predictive value NPV = 70%, Stratification of RT-PCR data showed superior sensitivity achieved with an RT-PCR cycle threshold (Ct) below 35 (97%), which decreased to 60% above 35. INTERPRETATION: In this field trial, RT-LAMP appears to be a reliable assay, comparable to RT-PCR, particularly with medium-high viral loads (Ct < 35). Hence, RT-LAMP can be deployed in RLS for timely management and prevention of COVID-19, without compromising the quality of output.

Prevalence of human respiratory syncytial virus, parainfluenza and adenoviruses in East Africa Community partner states of Kenya, Tanzania, and Uganda: A systematic review and meta-analysis (2007-2020).

BACKGROUND: Viruses are responsible for a large proportion of acute respiratory tract infections (ARTIs). Human influenza, parainfluenza, respiratory-syncytial-virus, and adenoviruses are among the leading cause of ARTIs. Epidemiological evidence of those respiratory viruses is limited in the East Africa Community (EAC) region. This review sought to identify the prevalence of respiratory syncytial virus, parainfluenza, and adenoviruses among cases of ARTI in the EAC from 2007 to 2020. METHODS: A literature search was conducted in Medline, Global Index Medicus, and the grey literature from public health institutions and programs in the EAC. Two independent reviewers performed data extraction. We used a random effects model to pool the prevalence estimate across studies. We assessed heterogeneity with the I2 statistic, and Cochran's Q test, and further we did subgroup analysis. This review was registered with PROSPERO under registration number CRD42018110186. RESULTS: A total of 12 studies met the eligibility criteria for the studies documented from 2007 to 2020. The overall pooled prevalence of adenoviruses was 13% (95% confidence interval [CI]: 6-21, N = 28829), respiratory syncytial virus 11% (95% CI: 7-15, N = 22627), and parainfluenza was 9% (95% CI: 7-11, N = 28363). Pooled prevalence of reported ARTIs, all ages, and locality varied in the included studies. Studies among participants with severe acute respiratory disease had a higher pooled prevalence of all the three viruses. Considerable heterogeneity was noted overall and in subgroup analysis. CONCLUSION: Our findings indicate that human adenoviruses, respiratory syncytial virus and parainfluenza virus are prevalent in Kenya, Tanzania, and Uganda. These three respiratory viruses contribute substantially to ARTIs in the EAC, particularly among those with severe disease and those aged five and above.

Morbidity burden, seasonality and factors associated with the human respiratory syncytial virus, human parainfluenza virus, and human adenovirus infections in Kenya.

Background: Human respiratory syncytial viruses (HRSV), human parainfluenza viruses (HPIV), and human adenoviruses (HAdVs) cause a substantial morbidity burden globally. Objective: We sought to estimate morbidity burden, assess seasonality, and determine factors associated with these respiratory viruses in Kenya. Methods: The data were obtained from Kenyan sites included in the Köppen-Geiger climate classification system. We defined the proportion of morbidity burden by descriptive analysis and visualized time-series data for January 2007-December 2013. Logistic regression was used to identify factors associated with infection outcomes. Results: The morbidity burden for HRSV was 3.1%, HPIV 5.3% and HAdVs 3.3%. Infants were more likely to be infected than other age groups. HRSV exhibited seasonality with high occurrence in January-March (odds ratio[OR] = 2.73) and April-June (OR = 3.01). Hot land surface temperature (≥40 °C) was associated with HRSV infections (OR = 2.75), as was warmer air temperature (19-22.9 °C) (OR = 1.68), compared with land surface temperature (<30) and cooler air temperature (<19 °C) respectively. Moderate rainfall (150-200 mm) areas had greater odds of HRSV infection (OR = 1.32) than low rainfall (<150 mm). Conclusion: HRSV, HPIV and HAdVs contributed to morbidity burden, and infants were significantly affected. HRSV had a clear seasonal pattern and were associated with climate parameters, unlike HPIV and HAdVs.

Plasmid mediated penicillin and tetracycline resistance among Neisseria gonorrhoeae isolates from Kenya.

BACKGROUND: Treatment of gonorrhea is complicated by the development of antimicrobial resistance in Neisseria gonorrhoeae (GC) to the antibiotics recommended for treatment. Knowledge on types of plasmids and the antibiotic resistance genes they harbor is useful in monitoring the emergence and spread of bacterial antibiotic resistance. In Kenya, studies on gonococcal antimicrobial resistance are few and data on plasmid mediated drug resistance is limited. The present study characterizes plasmid mediated resistance in N. gonorrhoeae isolates recovered from Kenya between 2013 and 2018. METHODS: DNA was extracted from 36 sub-cultured GC isolates exhibiting varying drug resistance profiles. Whole genome sequencing was done on Illumina MiSeq platform and reads assembled de-novo using CLC Genomics Workbench. Genome annotation was performed using Rapid Annotation Subsystem Technology. Comparisons in identified antimicrobial resistance determinants were done using Bioedit sequence alignment editor. RESULTS: Twenty-four (66.7%) isolates had both ß-lactamase (TEM) and TetM encoding plasmids. 8.3% of the isolates lacked both TEM and TetM plasmids and had intermediate to susceptible penicillin and tetracycline MICs. Twenty-six (72%) isolates harbored TEM encoding plasmids. 25 of the TEM plasmids were of African type while one was an Asian type. Of the 36 isolates, 31 (86.1%) had TetM encoding plasmids, 30 of which harbored American TetM, whereas 1 carried a Dutch TetM. All analyzed isolates had non-mosaic penA alleles. All the isolates expressing TetM were tetracycline resistant (MIC> 1 mg/L) and had increased doxycycline MICs (up to 96 mg/L). All the isolates had S10 ribosomal protein V57M amino acid substitution associated with tetracycline resistance. No relation was observed between PenB and MtrR alterations and penicillin and tetracycline MICs. CONCLUSION: High-level gonococcal penicillin and tetracycline resistance in the sampled Kenyan regions was found to be mediated by plasmid borne blaTEM and tetM genes. While the African TEM plasmid, TEM1 and American TetM are the dominant genotypes, Asian TEM plasmid, a new TEM239 and Dutch TetM have emerged in the regions.

Prevalence and factors influencing the distribution of influenza viruses in Kenya: Seven-year hospital-based surveillance of influenza-like illness (2007-2013).

BACKGROUND: Influenza viruses remain a global threat with the potential to trigger outbreaks and pandemics. Globally, seasonal influenza viruses' mortality range from 291 243-645 832 annually, of which 17% occurs in Sub-Saharan Africa. We sought to estimate the overall prevalence of influenza infections in Kenya, identifying factors influencing the distribution of these infections, and describe trends in occurrence from 2007 to 2013. METHODS: Surveillance was conducted at eight district hospital sites countrywide. Participants who met the case definition for influenza-like illness were enrolled in the surveillance program. The nasopharyngeal specimens were collected from all participants. We tested all specimens for influenza viruses with quantitative reverse transcriptase real-time polymerase chain reaction (RT-qPCR) assay. Bivariate and multivariate log-binomial regression was performed with a statistically significant level of p<0.005. An administrative map of Kenya was used to locate the geographical distribution of surveillance sites in counties. We visualized the monthly trend of influenza viruses with a graph and chart using exponential smoothing at a damping factor of 0.5 over the study period (2007-2013). RESULTS: A total of 17446 participants enrolled in the program. The overall prevalence of influenza viruses was 19% (n = 3230), of which 76% (n = 2449) were type A, 21% (n = 669) type B and 3% (n = 112) A/ B coinfection. Of those with type A, 59% (n = 1451) were not subtyped. Seasonal influenza A/H3N2 was found in 48% (n = 475), influenza A/H1N1/pdm 2009 in 43% (n = 434), and seasonal influenza A/ H1N1 in 9% (n = 88) participants. Both genders were represented, whereas a large proportion of participants 55% were ≤1year age. Influenza prevalence was high, 2 times more in other age categories compared to ≤1year age. Category of occupation other than children and school attendees had a high prevalence of influenza virus (p< <0.001). The monthly trends of influenza viruses' positivity showed no seasonal pattern. Influenza types A and B co-circulated throughout the annual calendar during seven years of the surveillance. CONCLUSIONS: Influenza viruses circulate year-round and occur among children as well as the adult population in Kenya. Occupational and school-based settings showed a higher prevalence of influenza viruses. There were no regular seasonal patterns for influenza viruses.

Origin and evolution of dengue virus type 2 causing outbreaks in Kenya: Evidence of circulation of two cosmopolitan genotype lineages.

Dengue fever (DF) is an arboviral disease caused by dengue virus serotypes 1-4 (DENV 1-4). Globally, DF incidence and disease burden have increased in the recent past. Initially implicated in a 1982 outbreak, DENV-2 recently reemerged in Kenya causing outbreaks between 2011 and 2014 and more recently 2017-8. The origin and the evolutionary patterns that may explain the epidemiological expansion and increasing impact of DENV-2 in Kenya remain poorly understood. Using whole-genome sequencing, samples collected during the 2011-4 and 2017-8 dengue outbreaks were analyzed. Additional DENV-2 genomes were downloaded and pooled together with the fourteen genomes generated in this study. Bioinformatic methods were used to analyze phylogenetic relationships and evolutionary patterns of DENV-2 causing outbreaks in Kenya. The findings from this study have shown the first evidence of circulation of two different Cosmopolitan genotype lineages of DENV-2; Cosmopolitan-I (C-I) and Cosmopolitan-II (C-II), in Kenya. Our results put the origin location of C-I lineage in India in 2011, and C-II lineage in Burkina Faso between 1979 and 2013. C-I lineage was the most isolated during recent outbreaks, thus showing the contribution of this newly emerged strain to the increased DENV epidemics in the region. Our findings, backed by evidence of recent local epidemics that have been associated with C-I in Kenya and C-II in Burkina Faso, add to the growing evidence of expanding circulation and the impact of multiple strains of DENV in the region as well as globally. Thus, continued surveillance efforts on DENV activity and its evolutionary trends in the region, would contribute toward effective control and the current vaccine development efforts.

Multidisciplinary approach towards training of the next generation of forensic DNA analysts in Africa; a Kenyan perspective.

The uptake of forensic DNA testing technologies in Africa has been slow despite the revolutionary technology being discovered and adopted 3 decades ago. African governments and partners have invested in construction and equipping of forensic laboratories in Africa but the benefits are yet to be realised as the laboratories are still faced with the challenge of shortage of adequately trained personnel. This paper describes an innovative multidisciplinary training approach that was developed and used to train officers from the Directorate of Criminal Investigations Kenya. We report on the structure, implementation and effectiveness of the training. It is expected that with the increased number of trained forensic DNA analysts, there will be an improvement in quality of forensic DNA evidence presented in courts and a reduction in backlog in the forensic biology laboratories in Kenya.

Genetic analysis of HA1 domain of influenza A/H3N2 viruses isolated in Kenya during the 2007-2013 seasons reveal significant divergence from WHO-recommended vaccine strains.

BACKGROUND: Influenza viruses evolve rapidly and cause regular seasonal epidemics in humans challenging effective vaccination. The virus surface HA glycoprotein is the primary target for the host immune response. Here, we investigated the vaccine efficacy and evolution patterns of human influenza A/H3N2 viruses that circulated in Kenyan in the period before and after the 2009 A/H1N1 pandemic, targeting the HA1 domain. MATERIALS AND METHODS: A hundred and fifteen HA sequences of Kenyan virus viruses were analyzed relative to the corresponding WHO vaccine reference strains using bioinformatics approaches. RESULTS: Our analyses revealed varied amino acid substitutions at all the five antigenic sites (A-E) of the HA1 domain, with a majority the changes occurring at sites A and B. The Kenyan A/H3N2 viruses isolated during 2007/2008 seasons belonged to A/Brisbane/10/2007-like viruses lineage, while those circulating in 2009-2012 belonged to the lineage of A/Victoria/361/2011-like viruses. The 2013 viruses clustered in clade 3C.3 of the A/Samara/73/2013-like viruses. The mean evolutionary rate of the A/H3N2 viruses analyzed in the study was at 4.17×10-3 (95% HPD=3.09×10-3-5.31×10-3) nucleotide substitutions per site per year, whereas the TMRCA was estimated at 11.18 (95% HPD=9.00-14.12) years ago from 2013. The prediction of vaccine efficacy revealed modest vaccine efficaciousness during 2008, and 2010 influenza seasons, whilst sub-optimal effectiveness was registered in 2007, 2009, 2012 and 2013. Further, the overall selective pressure acting on the HA1 domain was estimated at 0.56 (ω<1), suggesting that a majority of codon sites in the HA1 epitopes were evolving under purifying selection. CONCLUSIONS: Generally, our results highlight the genetic plasticity of A/H3N2 viruses and reveal considerable disparity in vaccine efficaciousness against the A/H3N2 viruses that circulated in Kenya, specifically during 2007, 2009, 2012, and 2013 influenza seasons. Our findings underscore the importance and need for consistent surveillance and molecular characterization of influenza viruses, to inform decision making and enhance early of detection of strains with epidemic/pandemic potential as well as benefit in guiding decisions regarding the appropriate annual influenza vaccine formulations.

Genetic diversity and evolutionary analysis of human respirovirus type 3 strains isolated in Kenya using complete hemagglutinin-neuraminidase (HN) gene.

Human respirovirus type 3 (HRV3) is a leading etiology of lower respiratory tract infections in young children and ranks only second to the human respiratory syncytial virus (HRSV). Despite the public health importance of HRV3, there is limited information about the genetic characteristics and diversity of these viruses in Kenya. To begin to address this gap, we analyzed 35 complete hemagglutinin-neuraminidase (HN) sequences of HRV3 strains isolated in Kenya between 2010 and 2013. Viral RNA was extracted from the isolates, and the entire HN gene amplified by RT-PCR followed by nucleotide sequencing. Phylogenetic analyses of the sequences revealed that all the Kenyan isolates grouped into genetic Cluster C; sub-clusters C1a, C2, and C3a. The majority (54%) of isolates belonged to sub-cluster C3a, followed by C2 (43%) and C1a (2.9%). Sequence analysis revealed high identities between the Kenyan isolates and the HRV3 prototype strain both at the amino acid (96.5-97.9%) and nucleotide (94.3-95.6%) levels. No amino acid variations affecting the catalytic/active sites of the HN glycoprotein were observed among the Kenyan isolates. Selection pressure analyses showed that the HN glycoprotein was evolving under positive selection. Evolutionary analyses revealed that the mean TMRCA for the HN sequence dataset was 1942 (95% HPD: 1928-1957), while the mean evolutionary rate was 4.65x10-4 nucleotide substitutions/site/year (95% HPD: 2.99x10-4 to 6.35x10-4). Overall, our results demonstrate the co-circulation of strains of cluster C HRV3 variants in Kenya during the study period. This is the first study to describe the genetic and molecular evolutionary aspects of HRV3 in Kenya using the complete HN gene.

Assessing antigenic drift and phylogeny of influenza A (H1N1) pdm09 virus in Kenya using HA1 sub-unit of the hemagglutinin gene.

Influenza A (H1N1) pdm09 virus emerged in North America in 2009 and has been established as a seasonal strain in humans. After an antigenic stasis of about six years, new antigenically distinct variants of the virus emerged globally in 2016 necessitating a change in the vaccine formulation for the first time in 2017. Herein, we analyzed thirty-eight HA sequences of influenza A (H1N1) pdm09 strains isolated in Kenya during 2015-2018 seasons, to evaluate their antigenic and molecular properties based on the HA1 sub-unit. Our analyses revealed that the A (H1N1) pdm09 strains that circulated in Kenya during this period belonged to genetic clade 6B, subclade 6B.1 and 6B.2. The Kenyan 2015 and 2016 isolates differed from the vaccine strain A/California/07/2009 at nine and fourteen antigenic sites in the HA1 respectively. Further, those isolated in 2017 and 2018 correspondingly varied from A/Michigan/45/2015 vaccine strain at three and fifteen antigenic sites. The predicted vaccine efficacy of A/California/07/2009 against Kenyan 2015/2016 was estimated to be 32.4% while A/Michigan/45/2015 showed estimated vaccine efficacies of 39.6% - 41.8% and 32.4% - 42.1% against Kenyan 2017 and 2018 strains, respectively. Hemagglutination-inhibition (HAI) assay using ferret post-infection reference antiserum showed that the titers for the Kenyan 2015/2016 isolates were 2-8-fold lower compared to the vaccine strain. Overall, our results suggest the A (H1N1) pdm09 viruses that circulated in Kenya during 2015/2016 influenza seasons were antigenic variants of the recommended vaccine strains, denoting sub-optimal vaccine efficacy. Additionally, data generated point to a swiftly evolving influenza A (H1N1) pdm09 virus in recent post pandemic era, underscoring the need for sustained surveillance coupled with molecular and antigenic analyses, to inform appropriate and timely influenza vaccine update.

gyrA and parC mutations in fluoroquinolone-resistant Neisseria gonorrhoeae isolates from Kenya.

BACKGROUND: Phenotypic fluoroquinolone resistance was first reported in Western Kenya in 2009 and later in Coastal Kenya and Nairobi. Until recently gonococcal fluoroquinolone resistance mechanisms in Kenya had not been elucidated. The aim of this paper is to analyze mutations in both gyrA and parC responsible for elevated fluoroquinolone Minimum Inhibitory Concentrations (MICs) in Neisseria gonorrhoeae (GC) isolated from heterosexual individuals from different locations in Kenya between 2013 and 2017. METHODS: Antimicrobial Susceptibility Tests were done on 84 GC in an ongoing Sexually Transmitted Infections (STI) surveillance program. Of the 84 isolates, 22 resistant to two or more classes of antimicrobials were chosen for analysis. Antimicrobial susceptibility tests were done using E-test (BioMerieux) and the results were interpreted with reference to European Committee on Antimicrobial Susceptibility Testing (EUCAST) standards. The isolates were sub-cultured, and whole genomes were sequenced using Illumina platform. Reads were assembled de novo using Velvet, and mutations in the GC Quinolone Resistant Determining Regions identified using Bioedit sequence alignment editor. Single Nucleotide Polymorphism based phylogeny was inferred using RaxML. RESULTS: Double GyrA amino acid substitutions; S91F and D95G/D95A were identified in 20 isolates. Of these 20 isolates, 14 had an additional E91G ParC substitution and significantly higher ciprofloxacin MICs (p = 0.0044*). On the contrary, norfloxacin MICs of isolates expressing both GyrA and ParC QRDR amino acid changes were not significantly high (p = 0.82) compared to MICs of isolates expressing GyrA substitutions alone. No single GyrA substitution was found in the analyzed isolates, and no isolate contained a ParC substitution without the simultaneous presence of double GyrA substitutions. Maximum likelihood tree clustered the 22 isolates into 6 distinct clades. CONCLUSION: Simultaneous presence of amino acid substitutions in ParC and GyrA has been reported to increase gonococcal fluoroquinolone resistance from different regions in the world. Our findings indicate that GyrA S91F, D95G/D95A and ParC E91G amino acid substitutions mediate high fluoroquinolone resistance in the analyzed Kenyan GC.

First report of Entamoeba moshkovskii in human stool samples from symptomatic and asymptomatic participants in Kenya.

Entamoeba moshkovskii is a member of the Entamoeba complex and a colonizer of the human gut. We used nested polymerase chain reaction (PCR) to differentiate Entamoeba species in stool samples that had previously been screened by microscopy. Forty-six samples were tested, 23 of which had previously been identified as Entamoeba complex positive by microscopy. Of the 46 specimens tested, we identified nine (19.5%) as E. moshkovskii-positive. In seven of these nine E. moshkovskii-positive samples, either E. dispar or E. histolytica (or both) were also identified, suggesting that co-infections may be common. E. moshkovskii was also detected in both symptomatic and asymptomatic participants. To the best of our knowledge, this is the first report of E. moshkovskii in Kenya.