Performance of methylcellulose and Avicel overlays in plaque and focus assays of Chikungunya virus

Background: Chikungunya virus is a re-emerging pathogen that is responsible for Chikungunya fever periodic outbreaks along the Kenyan coast and in other African countries.  Epidemiological data from the World Health Organization show that in 2014-2015, there was a major outbreak of Chikungunya fever in the Americas and Pacific Islands.  Surveillance and correct diagnosis are therefore key in controlling the spread and management of the disease. Plaque and focus assays are key techniques in viral characterization or quantification, and both assays typically require overlay with gelling polymers to limit the spread of viruses in cell culture.  There are anecdotal reports that Avicel may be superior to methylcellulose in assay of Influenza virus. However, it is unclear whether this would apply to other viruses. Objective: The objective of this study was to determine the performance of methylcellulose and Avicel overlays in plaque and focus assays of Chikungunya virus. Methods: Confluent Vero cells were seeded in 6- or 96-well plates for plaque and focus assays respectively. Cells were inoculated with serially diluted Chikungunya virus, and incubated to allow adherence of the virus to the cells. The inoculum was removed; replaced with Avicel or methylcellulose overlay at various concentrations and stained with crystal violet or immunostained.  Statistical significance was computed using the Holm-Sidak test. Results: The size of plaques formed by Chikungunya virus was dependent on the concentration of both Avicel and methylcellulose gels used as overlays, with Avicel overlays giving consistently larger plaques than methylcellulose.  Chikungunya virus formed plaques nearly 2.5 times larger in diameter (2 vs 0.8 mm) with 1.2 % Avicel than with 1.25 % methylcellulose after 60 hr growth.  Plaques formed with Avicel were better defined and easier to count after 48 hr growth period compared to a 60 hr period. However, methycellulose overlays provided smaller, more distinct and better defined foci in focus assays. Conclusion: Both methylcellulose and Avicel are good overlay media for viral assays. Avicel is marginally better for plaque assays while methylcellulose provides more distinct and easier to count foci in focus assays. Key words: Chikungunya virus, plaque assay, focus assay, methylcellulose, Avice

Profiles of patients on warfarin anticoagulation therapy in a leading tertiary referral hospital in Kenya; findings and implications for Kenya

Background: Patients’ profiles affect the outcome with warfarin; however, this data, and its implications, is scarce in resource-poor countries without access to pharmacogenetics or regular INR testing. Objectives: To characterize the profiles of patients on long-term warfarin therapy and subsequently use these to guide future anticoagulation management. Methods: Cross-sectional study among 180 adult patients receiving warfarin therapy in at a leading referral hospital in Kenya. Sociodemographic characteristics were obtained through face-to-face interviews. Details of warfarin therapy, concomitant medication and comorbidities were retrieved from medical records. Associations between patients’ profiles and the clinical indications of anticoagulation were computed at p ≤ 0.05. Results: Warfarin maintenance dose was 6.17 (±2.75) mg per day. Venous thromboembolism (56.6%) amongst obese patients (p = 0.0019) and cardioembolic events (48.3%) among males (p = 0.0316) aged ≤50 years (p = 0.0436) whose body mass indices were ≤ 25 (p < 0.0001) were the most common indications. Two-fifths and 45.0% of the patients had at least one other disease and concomitant medications. Conclusions: Long term warfarin therapy among Kenyans is mainly for overweight or lean middle-aged individuals suffering from venous or cardioembolic diseases. Studies should correlate patients’ profiles with warfarin response to guide future management

Determinants of Discontinuation of Contraceptive Methods among Women at Kenyatta National Hospital, Kenya

Background: Contraceptive use prevalence in Kenya is at 58% according to the 2014 Kenya Demographic Health Survey. Several factors lead to discontinuation and switch of contraceptives. Discontinuation rate as per the 2008/2009 KDHS for one year was 35.8%. Objectives: To find out the determinants of discontinuation and switching of hormonal and IUD contraceptive methods. Methodology: The study was a cross-sectional hospital based one, where data was collected by use of an interviewer administered questionnaire at Kenyatta National Hospital, where 400 women were interviewed. The data was then analyzed by use of the statistical software, SPSS Version 20. Results: The study population mainly comprised married women with a mean age of 31.45±6.40 years. Most women chose a contraceptive method which they perceived had the least adverse effects. Discontinuation rate by the time of study in the study population was 60.8%, with adverse effects as the most cited reason for discontinuation. The method of contraception and presence of co-morbidity were the factors associated with discontinuation of a contraceptive. Survival analysis showed that depot injection and Jadelle® (Levonorgestrel releasing implant 75mg) had the highest median months of use and Impanon® (Etonogestrel implant 68mg) the least. Conclusion: The reasons for discontinuation of contraceptive use include primarily concerns for adverse effects, use inconvenience, desire for pregnancy, contraceptive failure, and doctor’s advice. Key words: Contraceptive discontinuation, adverse effects, duration of us

Evaluation of malaria rapid diagnostic tests among children in a malaria endemic region in coastal Kenya

Background: In Kenya, malaria case management is based on clinical suspicion and detection of parasite in blood by parasitological or confirmatory diagnosis. Confirmatory diagnosis can be achieved with either microscopy or Rapid diagnostic tests (RDTs). RDTs are relatively new technologies, and their performance in actual conditions of use needs to be evaluated to provide information for appropriate use and to support decision making in procurement. Objectives: To evaluate performance and operational characteristics of three malaria RDTs: CareStart™, First Response®, and SD Bioline™ in the field for diagnosis of infection by Plasmodium falciparum monospecies as well as mixed infections with P. ovale and P. malariae. Methodology: A prospective study with blind comparisons to a gold standard was carried out at Pingilikani dispensary in Kilifi County, Kenya. Blood samples were obtained from 500 febrile children. Three RDTs: CareStart™, First Response® and SD Bioline™ were evaluated against microscopy of Giemsa stained blood films for detection of Plasmodium falciparum and non-falciparum malarial parasites. RDTs specific for P. falciparum only (HRP2 RDTs) and non-falciparum malarial parasites (HRP2/pLDH (Pf/pan) RDTs) were evaluated. Results: Plasmodium sp were detected by microscopy in 242 (48.40%) study participants. Plasmodium falciparum species were the most prevalent (93.3%) in comparison with other Plasmodium species: P. ovale and P. malariae whose prevalence were 2.89% and 3.71% respectively. Compared to microscopy the sensitivities of CareStart™, SD Bioline™, and First Response® RDTs for Plasmodium falciparum using Pf (mono species) kits were: 95.04% (95% CI: 92.34 - 97.73), 95.04% (95% CI: 92.34 - 97.73) and 94.21% (95% CI: 91.3 - 97.11) respectively while the specificities were 78.12% (95% CI: 72.98 - 83.25), 81.10% (95% CI: 76.23 - 85.96) and 78.74%  (95% CI: 73.65 - 83.82) respectively. Sensitivities of CareStart™, SD Bioline™ and First Response® RDTs for Plasmodium falciparum using Pf/Pan kits were: 99.02% (95% CI: 98.92 - 99.15), 99.04% (95% CI: 98.92 – 99.15) and 97.56% (95% CI: 97.78 – 97.99), respectively while the specificities were 78.46% (95% CI: 77.61 - 79.30), 78.46% (95% CI: 75.78 - 81.13) and 80.28% (95% CI: 76.73 - 83.82) respectively. CareStart™, SD Bioline™, and First Response® RDTs for non-falciparum sp using Pf/Pan kits both had 100% sensitivity and specificity. Conclusion: Data from this study demonstrate that CareStart™, SD Bioline™ and First Response® RDTs have good operational characteristics and are reliable alternatives to microscopy for diagnosing malaria in children. Key words: malaria, rapid diagnostic tests, microscopy, Plasmodiu

Prevalence and determinants of antibiotic related adverse drug reactions in Kenya : spontaneously reported cases at the pharmcy and poisons board database

Background: Antibiotics are useful in the treatment and prevention of many infections. Despite this, they may cause adverse drug reactions (ADRs) which could further increase the morbidity, mortality and treatment costs. The national pharmacovigilance system of the Pharmacy and Poisons board in Kenya has a database in which all actual and suspected adverse drug reactions nationally from hospitals are reported and analysed. Objective: To describe the prevalence, characteristics, severity and outcome of antibiotic related adverse reactions between January 2010 and December 2015. Methods: This was a retrospective cross sectional study in which all case reports of the reported antibiotic related ADRs were reviewed. Information on the patient demographics, types of antibiotics and concomitant drugs used, adverse reaction reported, the severity and outcome was abstracted and analysed using STATA version 13. Bivariate analysis and logistic regression were conducted to determine the risk factors associated with severity and outcomes of antibiotic related ADRs. Ethical approval was obtained from the Kenyatta National Hospital/ University of Nairobi Research and Ethics Committee and the Pharmacy and Poisons board. Results: A total of 550 case reports were analysed. The majority of patients were female (62.3%, n=330), median age of 34 (IQR 22.0-45.0) years. The most commonly affected system was the integumentary system (60.9%, n=388), with skin rash as the most commonly reported ADR (39.7%,n=253). Cotrimoxazole contributed the majority of the adverse reactions (55.3%, n=304). Most of the reactions were mild to moderate (82.6%, n=440), leading to drug withdrawal for 79.1% (n=435). Sulphonamides and anti-TB drugs produced the most severe reactions (n=15.8%, n=84), while fatal reactions were only 1.5% (n=8). Older age was significantly associated with severity of the reported ADRs (p=0.003) while HIV status (p=0.011) and severity of the ADR (p=<0.001) were associated with poor outcomes. Causality assessment attributed 15.6% (n=86) of the ADRs to the suspected antibiotic while 56.5% (n=311) were probable. Conclusion: There is a high burden of antibiotic related ADRS, most of which are skin related. HIV status and severity of the ADRs are associated with poor outcomes, and the elderly experience more severe ADRs

Patterns of Prescribing Practices in Makueni County Referral Hospital, Kenya

Background: Prescribing is said to be irrational if it does not conform to good standards of treatment. Irrational prescribing leads to increased cost of drug therapy, increased risk for adverse drug reactions and emergence of drug resistance. Objective: The study objective was to determine the quality and patterns of prescribing in Makueni County Referral Hospital, Kenya, using World Health Organization prescribing indicators. Methodology: The design was a descriptive retrospective cross-sectional study. Data was abstracted from 824 patient encounters selected through quasi-random sampling. Data was collected from the sampled prescriptions using a pre-tested data collection form, entered into and analyzed using Stata version 10.0 software. Results: The mean number of drugs per patient encounter was 2.7. Only 45.5% of the total drugs were prescribed using generic names. Antibiotics and injections were prescribed in 74% and 13.2% of the prescriptions surveyed respectively. Discussion: On average, inpatients received a higher number of drugs per encounter compared to outpatients, probably because they usually have more severe disease than outpatients which may require management with more drugs. Conclusion: The results showed a trend towards irrational prescribing, particularly polypharmacy, underuse of generic names and over-prescription of antibiotics. Relevant educational, managerial and regulatory interventions are recommended to remedy the problems. Keywords: Irrational prescribing, prescribing indicators, polypharmac

Molecular Surveillance of Adamantane Resistance among Human Influenza A Viruses Isolated in Four Epidemic Seasons in Kenya

Background: Adamantanes impede influenza A virus replication and are important in the treatment and prophylaxis of disease caused by these viruses. Genotypic characterization of influenza A viruses for mutations associated with resistance to adamantanes has not been fully investigated in Kenya. Objective: To characterize susceptibility of influenza A virus subtypes that circulated in Kenya from 2008-2011 to adamantanes. Methods: Archived influenza A virus strains obtained from 2008 to 2011 were propagated in MDCK cells prior to sequencing of the matrix and hemagglutinin gene segments, followed by bioinformatics analyses. Results: Ninety two virus strains consisting of 21 A/H3N2, 18 A/H1N1 and 53 A/H1N1pdm09 were analyzed.  All A/H3N2 and A/H1N1pdm09 viruses displayed resistance to adamantanes due to the S31N/S31D amino acid substitution. All A/H1N1pdm09 virus strains belonged to the N-lineage characterized by S203T amino acid substitution in the HA1. All A/H1N1 viruses were sensitive to adamantane and were characterized by K140E amino acid substitution in the HA1. Conclusion: All Kenyan influenza A/H3N2 and A/H1N1pdm09 virus strains were resistant to adamantanes while seasonal A/H1N1 strains were sensitive to these drugs. During the study period, Amantadine and Rimantadine were inappropriate for prophylaxis and treatment of influenza disease caused by A/H3N2 and A/H1N1pdm09 virus subtypes in Kenya. Key words: Kenya, influenza A/H3N2, A/H1N1pdm09, A/H1N1, adamantane

Microbial Quality of Unregulated Herbal Medicinal Products in Kenya

Background: The use of herbal medicines is global, with the majority of the world’s population depending on traditional medicines, particularly herbal remedies for their primary healthcare needs. In Kenya, it is estimated that conventional healthcare system provides for approximately 30 % of the population, while nearly 70 % rely on herbal remedies. Herbal medicines, however, present safety concerns as they carry a relatively high risk of contamination by pathogenic microbes, organic and inorganic pollutants including toxic metals and non-metals, organic, mycotoxins, endotoxins, and agrochemical residues. Objective: This study was designed to assess the microbial quality of regulated and unregulated herbal medicinal products in diverse Kenyan markets, such as the supermarkets, roadside vendors, retail pharmacies and herbal clinics, for levels of microbial contaminants. Materials and Methods: Thirty samples of registered and unregistered herbal medicinal products were collected by purposive sampling from five Kenyan provinces.  Microbial load analysis was performed in accordance to pharmacopoeial methods (BP and USP). Microorganisms were further isolated and characterized using differential and selective media and by biochemical analyses. Results: All registered products had microbial load below 100 cfu/ml, and complied with BP and USP requirements. However, none of the unregistered samples complied with pharmacopoeial limits for either or both bacterial and fungal load limits. Most of the unregistered samples had microbial loads ranging from 3.00×106 to 1.56 × 1010 cfu/ml, thus exceeding by far BP or USP standards. The microbial isolates belonged to fifteen (15) different bacterial genera and seven (7) fungal genera.  Escherichia coli was the most frequently isolated bacteria from 75 % of the unregistered product samples while Klebsiella pneumoniae, Enterobacter aerogenes, and Staphylococcus aureus followed in 70 %, 60 % and 45 % of the samples, respectively. Salmonella spp was isolated in 40 % of the samples while Shigella spp was found in 20 % of the samples. Conclusion: Unregulated herbal medicinal products that are available in diverse Kenyan markets show poor microbial quality and exhibit contamination by pathogenic microorganisms.  There is need to extend regulatory control by the drug authorities to herbal medicinal products to enhance microbial quality and safety. Keywords: Microbial contaminants, herbal medicines, microbial loa