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

Identification and characterization of influenza A viruses in selected domestic animals in Kenya, 2010-2012

<div><p>Background</p><p>Influenza A virus subtypes in non-human hosts have not been characterized in Kenya. We carried out influenza surveillance in selected domestic animals and compared the virus isolates with isolates obtained in humans during the same period.</p><p>Methods</p><p>We collected nasal swabs from pigs, dogs and cats; oropharyngeal and cloacal swabs from poultry; and blood samples from all animals between 2010 and 2012. A standardized questionnaire was administered to farmers and traders. Swabs were tested for influenza A by rtRT-PCR, virus isolation and subtyping was done on all positive swabs. All sera were screened for influenza A antibodies by ELISA, and positives were evaluated by hemagglutination inhibition (HI). Full genome sequencing was done on four selected pig virus isolates.</p><p>Results</p><p>Among 3,798 sera tested by ELISA, influenza A seroprevalence was highest in pigs (15.9%; 172/1084), 1.2% (3/258) in ducks, 1.4% (1/72) in cats 0.6% (3/467) in dogs, 0.1% (2/1894) in chicken and 0% in geese and turkeys. HI testing of ELISA-positive pig sera showed that 71.5% had positive titers to A/California/04/2009(H1N1). Among 6,289 swabs tested by rRT-PCR, influenza A prevalence was highest in ducks [1.2%; 5/423] and 0% in cats and turkeys. Eight virus isolates were obtained from pig nasal swabs collected in 2011 and were determined to be A(H1N1)pdm09 on subtyping. On phylogenetic analysis, four hemagglutinin segments from pig isolates clustered together and were closely associated with human influenza viruses that circulated in Kenya in 2011.</p><p>Conclusion</p><p>Influenza A(H1N1)pdm09 isolated in pigs was genetically similar to contemporary human pandemic influenza virus isolates. This suggest that the virus was likely transmitted from humans to pigs, became established and circulated in Kenyan pig populations during the study period. Minimal influenza A prevalence was observed in the other animals studied.</p></div

Distribution of hemagglutination-inhibition (HI) titers and proportion positive of enzyme-linked immunosorbent assay (ELISA)-positive pig sera against three reference antigens for household and slaughterhouse sites, May 2010 –August 2012 (n = 172)^*.

<p>Distribution of hemagglutination-inhibition (HI) titers and proportion positive of enzyme-linked immunosorbent assay (ELISA)-positive pig sera against three reference antigens for household and slaughterhouse sites, May 2010 –August 2012 (n = 172)^{<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0192721#t003fn001" target="_blank">*</a>}.</p

Analysis of HA segment of Kenya swine isolates alongside other contemporary isolates.

<p>Branches highlighted in red referto Kenya swine isolates while those in blue are Kenya human isolates. A/California/04/2009 is used to root the phylogenetic tree.</p

Prevalence of PCR positivity for influenza A in nasal, oropharyngeal and cloacal swabs, by species and site, Kenya, May 2010-August 2012.

<p>Prevalence of PCR positivity for influenza A in nasal, oropharyngeal and cloacal swabs, by species and site, Kenya, May 2010-August 2012.</p

Influenza A seroprevalence by ELISA among pigs sampled at the slaughter house, 2010–2012.

<p>Influenza A seroprevalence by ELISA among pigs sampled at the slaughter house, 2010–2012.</p

Proportion of enrolled households (HH) owning different animal types by study site, Kenya, May 2010-August 2012.

<p>Proportion of enrolled households (HH) owning different animal types by study site, Kenya, May 2010-August 2012.</p

Map of Kenya counties showing the location of the study sites, Kibera in Nairobi, Ndumbui-ini slaughterhouse in Kiambu County and Asembo in Siaya County.

<p>Map of Kenya counties showing the location of the study sites, Kibera in Nairobi, Ndumbui-ini slaughterhouse in Kiambu County and Asembo in Siaya County.</p

Outbreak of Middle East Respiratory Syndrome Coronavirus in Camels and Probable Spillover Infection to Humans in Kenya

The majority of Kenya&rsquo;s &gt; 3 million camels have antibodies against Middle East respiratory syndrome coronavirus (MERS-CoV), although human infection in Africa is rare. We enrolled 243 camels aged 0&ndash;24 months from 33 homesteads in Northern Kenya and followed them between April 2018 to March 2020. We collected and tested camel nasal swabs for MERS-CoV RNA by RT-PCR followed by virus isolation and whole genome sequencing of positive samples. We also documented illnesses (respiratory or other) among the camels. Human camel handlers were also swabbed, screened for respiratory signs, and samples were tested for MERS-CoV by RT-PCR. We recorded 68 illnesses among 58 camels, of which 76.5% (52/68) were respiratory signs and the majority of illnesses (73.5% or 50/68) were recorded in 2019. Overall, 124/4692 (2.6%) camel swabs collected from 83 (34.2%) calves in 15 (45.5%) homesteads between April&ndash;September 2019 screened positive, while 22 calves (26.5%) recorded reinfections (second positive swab following &ge; 2 consecutive negative tests). Sequencing revealed a distinct Clade C2 virus that lacked the signature ORF4b deletions of other Clade C viruses. Three previously reported human PCR positive cases clustered with the camel infections in time and place, strongly suggesting sporadic transmission to humans during intense camel outbreaks in Northern Kenya

Building laboratory capacity to detect and characterize pathogens of public and global health security concern in Kenya

Since 1979, multiple CDC Kenya programs have supported the development of diagnostic expertise and laboratory capacity in Kenya. In 2004, CDC's Global Disease Detection (GDD) program within the Division of Global Health Protection in Kenya (DGHP-Kenya) initiated close collaboration with Kenya Medical Research Institute (KEMRI) and developed a laboratory partnership called the Diagnostic and Laboratory Systems Program (DLSP). DLSP built onto previous efforts by malaria, human immunodeficiency virus (HIV) and tuberculosis (TB) programs and supported the expansion of the diagnostic expertise and capacity in KEMRI and the Ministry of Health. First, DLSP developed laboratory capacity for surveillance of diarrheal, respiratory, zoonotic and febrile illnesses to understand the etiology burden of these common illnesses and support evidenced-based decisions on vaccine introductions and recommendations in Kenya. Second, we have evaluated and implemented new diagnostic technologies such as TaqMan Array Cards (TAC) to detect emerging or reemerging pathogens and have recently added a next generation sequencer (NGS). Third, DLSP provided rapid laboratory diagnostic support for outbreak investigation to Kenya and regional countries. Fourth, DLSP has been assisting the Kenya National Public Health laboratory-National Influenza Center and microbiology reference laboratory to obtain World Health Organization (WHO) certification and ISO15189 accreditation respectively. Fifth, we have supported biosafety and biosecurity curriculum development to help Kenyan laboratories safely and appropriately manage infectious pathogens. These achievements, highlight how in collaboration with existing CDC programs working on HIV, tuberculosis and malaria, the Global Health Security Agenda can have significantly improve public health in Kenya and the region. Moreover, Kenya provides an example as to how laboratory science can help countries detect and control of infectious disease outbreaks and other public health threats more rapidly, thus enhancing global health security