Molecular characterization of enterotoxigenic Escherichia coli toxins and colonization factors in children under five years with acute diarrhea attending Kisii Teaching and Referral Hospital, Kenya.

BACKGROUND: Enterotoxigenic Escherichia coli (ETEC) is one of the leading causes of infectious diarrhea in children. There are no licensed vaccines against ETEC. This study aimed at characterizing Escherichia coli for ETEC enterotoxins and colonization factors from children < 5 years with acute diarrhea and had not taken antibiotics prior to seeking medical attention at the hospital. METHODS: A total of 225 randomly selected archived E. coli strains originally isolated from 225 children with acute diarrhea were cultured. DNA was extracted and screened by multiplex polymerase chain reaction (PCR) for three ETEC toxins. All positives were then screened for 11 colonization factors by PCR. RESULTS: Out of 225 E. coli strains tested, 23 (10.2%) were ETEC. Heat-stable toxin (ST) gene was detected in 16 (69.6%). ETEC isolates with heat-stable toxin of human origin (STh) and heat-stable toxin of porcine origin (STp) distributed as 11 (68.8%) and 5 (31.2%) respectively. Heat-labile toxin gene (LT) was detected in 5 (21.7%) of the ETEC isolates. Both ST and LT toxin genes were detected in 2 (8.7%) of the ETEC isolates. CF genes were detected in 14 (60.9%) ETEC strains with a majority having CS6 6 (42.9%) gene followed by a combination of CFA/I + CS21 gene detected in 3 (21.4%). CS14, CS3, CS7 and a combination of CS5 + CS6, CS2 + CS3 genes were detected equally in 1 (7.1%) ETEC isolate each. CFA/I, CS4, CS5, CS2, CS17/19, CS1/PCFO71 and CS21 genes tested were not detected. We did not detect CF genes in 9 (39.1%) ETEC isolates. More CFs were associated with ETEC strains with ST genes. CONCLUSION: ETEC strains with ST genes were the most common and had the most associated CFs. A majority of ETEC strains had CS6 gene. In 9 (39.1%) of the evaluated ETEC isolates, we did not detect an identifiable CF.

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.

Evaluation of the performance of a multiplex reverse transcription polymerase chain reaction kit as a potential diagnostic and surveillance kit for rotavirus in Kenya.

BACKGROUND: Diarrhea is a serious concern worldwide, especially in developing countries. Rotavirus is implicated in approximately 400,000 infant deaths annually. It is highly contagious elevating the risk of outbreaks especially in enclosed settings such as daycare centers, hospitals, and boarding schools. Reliable testing methods are critical for early detection of infections, better clinical management, pathogen surveillance and evaluation of interventions such as vaccines. Enzyme immunoassays have proved to be reliable and practical in most settings; however, newer multiplex reverse transcription polymerase assays have been introduced in the Kenya market but have not been evaluated locally. METHODS: Stool samples collected from an ongoing Surveillance of Enteric Pathogens Causing diarrheal illness in Kenya (EPS) study were used to compare an established enzyme immunoassay, Premier™ Rotaclone® (Meridian Bioscience, Cincinnati, Ohio, U.S.A.), that can only detect group A rotavirus against a novel multiplex reverse transcription polymerase chain reaction kit, Seeplex® Diarrhea-V ACE Detection (Seegene, Seoul, Republic of Korea), that can detect rotavirus, astrovirus, adenovirus, and norovirus genogroups I and II. Detection frequency, sensitivity, specificity, turnaround time, and cost were compared to determine the suitability of each assay for clinical work in austere settings versus public health work in well-funded institutes in Kenya. RESULTS: The Premier™ Rotaclone® kit had a detection frequency of 11.2%, sensitivity of 77.8%, specificity of 100%, turnaround time of 93 min and an average cost per sample of 13.33 United States dollars (USD). The Seeplex® Diarrhea-V ACE Detection kit had a detection frequency of 16.0%, sensitivity of 100%, specificity of 98.1%, turnaround time of 359 min and an average cost per samples 32.74 United States dollars respectively. The detection frequency sensitivity and specificity of the Seeplex® Diarrhea-V ACE Detection kit mentioned above are for rotavirus only. CONCLUSIONS: The higher sensitivity and multiplex nature of the Seeplex® Diarrhea-V ACE Detection kit make it suitable for surveillance of enteric viruses circulating in Kenya. However, its higher cost, longer turnaround time and complexity favor well-resourced clinical labs and research applications. The Premier™ Rotaclone®, on the other hand, had a higher specificity, shorter turnaround time, and lower cost making it more attractive for clinical work in low complexity labs in austere regions of the country. It is important to continuously evaluate assay platforms' performance, operational cost, turnaround time, and usability in different settings so as to ensure quality results that are useful to the patients and public health practitioners.