Follow-Up of HIV Positive-Sputum Smear Negative Presumptive Tuberculosis Patients in The EAPHLNP Study Site in Kenya

Tuberculosis (TB) is a global health problem that causes ill-health among millions of people each year. The most common method for diagnosing TB worldwide was sputum smear microscopy. However the advent of HIV/AIDS has complicated the diagnosis and  management of TB with associated emergence of multidrug resistant TB (MDR-TB) and extensively drug resistant TB (XDR-TB). Studies that evaluate methods for diagnosing TB and follow-up of Presumptive TB patients to verify the presence or absence of active TB are crucial for prompt diagnosis and treatment. Objective: The study aimed to assess adherence to attend follow up clinical and sputum reexamination clinics for symptomatic HIV positive presumptive TB patients initially assumed to be TB negative. To find out at what stage HIV positive smear negative presumptive TB patients would become positive for TB with which diagnostic method within a 6 month follow up. Methodology: This was cohort study conducted in nine East African Public Health Laboratory Networking Project study sites in Kenya.  Eligible new and previously treated presumptive TB patients i.e. a patient presenting with symptoms and signs suggestive of TB who tested positive for HIV and had sputum smear negative on initial sputum smear examination were enrolled in to the study. Study participants were expected to be followed up for repeat sputum smear examination at 2 weeks, 2, 4, and 6 months. Results: Out of the 1323 presumptive TB patients enrolled between February 2013 and February 2014, only 201 (15.2%) were eligible for follow up and 164 (81.6%) did not return at 2 weeks for a follow up. Out of the 37 (18.4%) who returned for a follow up at 2 weeks, none was either sputum smear positive or Gene-Xpert positive. Initial sputum samples from all the 201 participants were subjected to culture. Culture results were available for only 156 patients of whom 15/156, (9.6%) were culture positive. Follow up adherence at 2 weeks was significantly associated with history of previous treatment (p-value = 0.010) but not associated with gender (p-value =0.268). Conclusion: The high level of non-adherence to follow up among HIV positive presumptive TB patients is a cause for concern to the  national TB program. We recommend active follow up of this group who has initial sputum smear negative results using the new  diagnostic tools that are available and can diagnosis TB early. Future studies should evaluate strategies that should be put in place to ensure enhanced follow up of the presumptive TB patients as part of the protocol. Key Words: Presumptive tuberculosis, HIV positive smear negative, Follow up&nbsp

Inter / Intra Examiner Calibration of Clinical Signs and Symptoms in Comparison with Laboratory Results in the Diagnosis of Pulmonary Tuberculosis in Patients from EAPHLN Project Study Sites in Kenya

Introduction: The clinical diagnosis of pulmonary tuberculosis (PTB) is based on occurrence of four cardinal signs and symptoms which include current cough, night sweats, weight loss, or low grade fever. However few studies have determined the validity and reliability of these diagnoses by intra and inter-examiner calibration of clinicians for appropriateness of detection of tuberculosis (TB) in resource constrained settings. Objective: The study's aim was to determine the sensitivity of concordance and reliability (Kappa values) of inter-examiner and intra-examiner findings of clinicians in the EAPHLN project. Methodology: The study was a cross-sectional study in nine sites. It included 155 patients for intraexaminer and 57 patients for inter-examiner calibrations selected from eligible people with symptoms or signs suggestive of TB during the implementation of the East African Public Health Laboratory Network Project (EAPHLNP) in Kenya. TB clinical symptoms and signs were recorded in a structured medical form included the following: productive cough, weight loss, night sweats, low grade fever (classical cardinal signs and symptoms). Using quality assurance sampling for a total population of ten thousand people with symptoms or signs suggestive of TB from the sites with a minimum defective sample acceptable of 0 and a probability of defect accepted of 1% andan alpha of 5%, the sample size of repeatable samples is 262 for total patients for the sites per year. Intra-examiner calibration involved examination of the same patient independently by the same clinician within one day interval. Inter-examiner calibration involved examination of the same patient by two clinicians independently the same day. Calibration of the clinical tools used during examination of patients was done. TB laboratory diagnosis was first done by sputum  smear microscopy Ziehl–Neelsen stain.(ZN), secondly by optimized sputum smear microscopy with a Light Emitting Diode microscope ( LED) or fluorescent microscopy(FM), and thirdly by Gene-expert technique (Gene Xpert or Gx). The results from the clinicians and reference laboratory findings for these patients were entered in a computer, verified and analyzed in SPSS for reliability statistics. These unweighted Cohen Kappa scores were interpreted as follows: poor0.01–0.20, moderate 0.21–0.40, fair 0.41–0.60, good 0.61–0.80, or excellent 0.81–1.0 based on the agreement between the intraexaminer and inter-examiner findings Results: A significant difference was found between concordant diagnosis of a least 4 signs and symptoms of TB compared to fewer by the same examiner on the same patient in all TB test/HIV status categories except the ZN positive /HIV positives and GeneXpert negative /HIV positives and HIV negative categories. The highest sensitivity rate was 81.8 %( 95%CI=52.3-94.9) in the Gx+ve/HIV+ve category. The significance difference in senstivity results of TB/HIV test vs at least presence of the 4 signs and symptoms. However, This did not occur in ZN+ve/HIV+ve, FM+ve/HIV+ve , Gx+ve/HIV+ve Gx+ve/HIV-ve categories. Kappa values for cough and fever were consistently significantly higher than zero kappa. Conclusion: Excellent kappa can be achieved in low resource settings by clinician using all four cardinal signs and symptoms of TB with laboratory results. There is possibility of using the clinical diagnosis using the four signs and symptoms where laboratory diagnosis is not present but specificity is low. Good clinical practice would improve the specificity

Effects of Sputum Quality on Xpert® MTB/RIF Results in The Detection of Mycobacterium Tuberculosis from Persons Presumed To Have TB in EAPHLN Project Operational Research Study Sites in Kenya

Background: The common problem in tuberculosis (TB) management is mis-diagnosis or underdiagnosis of cases leading to high  morbidity and mortality. In order to reverse this, new diagnostic tools for detection of Mycobacterium tuberculosis (MTB) the causative agent of TB disease have been developed. However, in the evaluation process of these tools many studies have not considered attributes of sputum quality in their testing algorithm. Objectives: This study aimed at evaluating the effect of sputum quality in detection of MTB when using Xpert® MTB/RIF (GeneXpert) among patients presumed to have pulmonary TB. Methodology: Between February 2013 and August 2014 a total of 3585 Spot and morning sputum specimens were collected from 1918 persons presumed to have pulmonary TB enrolled in nine East Africa Public Health Laboratory Networking (EAPHLN) Project study sites in Kenya. The mean age was 40 (+17SD) years ranging between 18 and 95 years. Some of these specimens (512) were rejected and 3073 were analyzed. The specimens were appropriately packaged and transported to KEMRI Mycobacteriology research laboratory where they were macroscopically characterized into muco-purulent; mucoid, salivary or blood stained. The sputum specimens having reddish color was labeled as blood-stained sputum. Each specimen was processed for GeneXpert testing and culture. Results: Upon macroscopic characterization, out of the 3073 specimens received, 46.1% were mucoid, 44% salivary, 7.5% muco-purulent,  while 2.4% were blood stained. Bivariate analysis revealed that there was a significant association between sputum quality and gender  (p<0.001), age (p=0.022), specimen type (p<0.001), and HIV status (p=0.003). Performance of GeneXpert on the different specimen  categories, muco-purulent (85.7%; CI 95%, 67.4-100%) and mucoid (85.3%, 95%CI: 77.393.3%) specimens had higher sensitivity when  compared to salivary specimens (76.7%, 95%CI: 64.1-89.3). However when stratified by HIV status, GeneXpert detected more MTB on salivary specimens produced by HIV positive (85.7%, 95%CI: 67.4-100%) patients than those from HIV negative patients (71.4%, 95%CI: 52.1- 90.7%). Conclusion: By macroscopic characterization, any sputum specimen type from HIV Positive or Negative persons presumed to have  tuberculosis can be used in diagnosis of tuberculosis regardless of sputum quality classification. However, the sensitivity of GeneXpert was higher in morning sputum specimens that were muco-purulent and mucoid with high MTB yield than in spot sputum specimens which were salivary with low MTB yield. Also, GeneXpert sensitivity was higher, though not significant, in salivary specimens from HIV positive individuals than those of HIV negative individuals. Sputum specimen quality assessment should be considered as an integral part of routine laboratory diagnosis of TB especially in HIV negative individuals. Keywords: Specimen quality, Sputum, Tuberculosis, Xpert® MTB/RIF, GeneXper

Impact of Research Methodology and Scientific Writing Training in Transforming Clinical and Laboratory Personnel to Research Scientists at the East Africa Public Health Laboratory Networking Project Study Sites in Kenya

Introduction: The East Africa Public Health Laboratory Networking (EAPHLN) Project which was being implemented in Kenya, Uganda, Tanzania, Burundi and Rwanda had several components which included Operational Research (OR). The OR component had two strategic objectives one was to provide oversight and guidance in carrying out operational research activities under the regional project secondly, was to facilitate local and regional capacity to carry out operational research and evaluation of medical diagnostics. Research Methodology and Scientific writing training sharpens the fundamental skills upon which the conceptual framework of scientific expertise is built. In order to facilitate the local and regional capacity to carry out operational research, there is need to build human capital at the study sites.Methodology:  A qualitative study was carried out using a semi-structured questionnaire to determine the research training needs among clinical and laboratory personnel at the World Bank-funded EAPHLNP facilities in Kenya. Based on the findings, a short training in Research Methodology and Scientific Writing training was developed in April 2012 to support the implementation of operational research activities at the health facilities in Kenya and the region. A team of research experts from KEMRI and an external facilitator with vast knowledge on curriculum development was constituted to develop the training. The training content was delivered through lectures, self directed learning, group discussions, individual and group assignments. The participants were instructed to develop concept proposals and were assigned mentors to guide them through the process of document development and approval.Findings: The research training needs assessment study revealed that there was limited knowledge in research undertaking and scientific writing. The curriculum was developed and piloted in April 2012. A total of nineteen participants from Kenyan sites were trained in Mombasa for two weeks. Seven concept proposals were developed. The proposals are at different stages of scientific review process. After undergoing this training, two of the participants were motivated to pursue further studies and enrolled for higher degrees (Masters and Doctoral).The training curriculum was reviewed in October 2012 by the KEMRI OR team and packaged into three manuals namely: - Facilitators, Participant and Exercise. The curriculum was adopted by the East, Central and Southern African Health Community (ECSAHC) to train participants from the region. Recommendations:  Strengthen capacity building in operational research at the centres of excellence in order to adequately address public health related issues. Capacity build, other facilities country wide and regionally by rolling out the training and develop structured mentorship programs

Emerging Antimicrobial Resistance Patterns of Enteric Pathogens Isolated from Children under 5 years in EAPHLNP Satellite Sites in Kenya

Introduction: The emergence of resistance to antimicrobial agents in bacterial pathogens is a worldwideproblem that has been associated with inappropriate use in human and veterinary medicine.Epidemiological studies from several African countries by the year 2001 established that, diarrhoeawas the most common illness reported by the United States military service members deployed toAfrica for strategic training and contingency operations. Out of 15,000 US military personnel whoparticipated, more than 500 service members were affected by acute diarrhoea [7]. Objective: To determine the susceptibility of common circulating enteric bacterial pathogens to antimicrobials. Methodology: Between 12th February 2013 and 30th July 2014, a total of 420 children under 5 years of age with diarrhea were analyzed for bacterial enteric pathogens of which E. coli isolates were characterized by Polymerase Chain Reaction for the presence of virulence genes. Results:  Patients from whom bacterial enteric pathogens were isolated and identified from the 5 satellite sites were= 145, Wajir = 21, Malindi= 42, Kitale = 34, Machakos = 18 and Busia = 30 County Referral Hospitals. Antibiotic susceptibility testing was done on all isolates: pathogenic E. coli = 55, Salmonella =23 and Shigella =72 using disk-diffussion methods containing Ampicillin, Cefotaxime, Tetracycline, Erythromycin Gentamicin, Chloramphenicol, Trimethoprim / Sulphamethoxazole, Ciprofloxacin, Furasolidine and Nalidixic acid. E. coli, Shigella and Salmonella isolates showed up to 100% level of resistance to ampicillin, trimethoprin / sulphamethoxazole and erythromycin.Furthermore, pathogenic E. coli revealed tetracycline resistance ranging from 67% to 76% in all sites. Emerging resistance to ciprofloxacin ranged from 14.3% in Wajir to 50.0% in Machakos and gentamycin resistance ranged from 20% in Kitale to 100% in Wajir. Salmonella isolates showed levels of resistance ranging from 25% to 100% in Busia and 14% to 100% in Wajir for all the antimicrobials tested.Conclusion: Our findings on diarrhea due to enteric bacteria show that a high percentage is caused by antimicrobial-resistant strains, thus illustrating the effect of long-standing unregulated antimicrobial use. Most enteric pathogens easily share genes for antimicrobial  resistance. There was emerging resistance to newly prescribed antibiotics. This may have policy implications on the use of antibiotics in Keny

Performance of Ziehl-Neelsen Microscopy, Light Emitting Diode – FM and Xpert MTB/RIF in the Diagnosis of Tuberculosis in People with Presumptive TB from EAPHLNP study sites in Kenya

In Kenya, sputum smear microscopy, especially Ziehl-Neelsen (ZN) method has been the cornerstone for tuberculosis (TB) diagnosis at  most public health facilities. Recently, Led Emitted Diode (LED) fluorescent microscopy (FM) and Xpert MTB/ RIF (GeneXpert), have been introduced in selected health facilities for diagnosis of TB and Drug Resistant TB. This study was undertaken to determine and compare the performance (sensitivity, specificity, positive and negative predictive test values) of these two new TB diagnostics with ZN microscopy as a benchmark, LED/FM and GeneXpert using either LJ or MGIT culture, whichever was available or if one was positive while the other negative, as a gold standard. Methodology: A cross-sectional study was conducted between February 2013 and August 2014 in nine selected public health, facilities in Kenya. People with presumptive TB aged 18 years and above, both new and re-treatment cases attending the facilities with symptoms suggestive of TB (including cough of two or more weeks) were eligible for the study and consecutively recruited. Two sputum specimens (spot and early morning) were collected over two consecutive days. A total of 3073 sputum samples were collected from 1891 people with presumptive TB. The specimens from the study sites were appropriately packaged and shipped to the TB research laboratory in KEMRI, Nairobi, whereby samples were received and processed for ZN, LED, GeneXpert, LJ and MGIT culture in accordance with standard  procedures. Culture was used as a gold standard. The study was approved by the Ethical Review Committee of KEMRI. Results: A total of 639 specimens from 390 patients with culture results were included in the analysis. GeneXpert showed significantly  higher sensitivity (83.7% (95%CI: 76.6-90.8)) than ZN  6 African Journal of Health Sciences Volume 32, Issue No. 6, November - December, 2019 (65.4% (95% CI: 56.3-74.5)) and FM (68.3% (95% CI: 59.4-77.2)) microscopy methods in the diagnosis of TB. On the contrary, specificity of GeneXpert (87.9% (95% CI: 85.1-90.7)) was significantly lower than that of ZN (93.5% (95% CI: 91.4-95.6)) and FM (93.3% (95% CI: 91.295.4)) microscopy. GeneXpert sensitivity in smear positive culture positive was (95.6% (95% CI: 90.7-100.0)) and (97.2% (95% CI: 93.4-100.0)) for ZN and FM, respectively, it was significantly lower in smear negative culture positive specimens with (61.1% (95% CI: 45.2-77.0)) and (54.5% (95%CI: 37.5-71.5)) for ZN and FM, respectively. Sensitivity rate was significantly higher in specimens from non-previously treated with presumptive TB (71.1% (95%CI: 61.4-80.9)) for ZN, (73.5% (95% CI: 64.0-83.0)) for FM and (89.2% (95% CI: 82.5-95.9)) for GeneXpert than those from retreatment cases (42.9% (95% CI: 21.7-64.1)), (47.6% (95% CI: 26.2-69.0)) and (61.9% (95%CI: 41.1-82.7)), respectively. Overall, HIV status did not affect the performance of GeneXpert. However, Sensitivity of GeneXpert (84.4% (95% CI: 71.8-97.0)) was significantly higher in HIV positive than that of ZN (53.1% (95% CI: 35.8-70.4)) and FM (56.3% (95% CI: 39.1-73.5)) microscopy. There were no significant differences in sensitivity of ZN (70.8% (95% CI: 60.3-81.3)) and FM (73.6% (95% CI: 63.4-83.8)) in HIV negative specimens compared to sensitivity of ZN (53.1% (95% CI: 35.8-70.4)) and FM (56.3% (95% CI: 39.1-73.5)) in HIV positive specimens. A small proportion (6.2%) of specimens with ZN and culture negative results was positive by GeneXpert. Conclusions: Performance of GeneXpert was higher than both ZN and FM microscopy for diagnosis of TB in Kenya and is comparable with performance indicated in a few previous studies in Africa. Despite the low sensitivity in smear negative culture positive specimens, GeneXpert has potential to increase diagnostic yield in smear and culture negative specimens, especially from HIV positive people with presumptive TB. Further studies are required to ascertain its specificity and application in specific patient population. This will be possible when patients' clinical details are linked with respective laboratory data as a result of combination of tests to improve diagnostic yield. Key words: ZN, LED-FM; GeneXpert: Performance; Tuberculosis Diagnosis