This was across sectional study conducted in the renal and general outpatients’ units of Mulago National Referral Hospital, Kampala, Uganda. The Hospital receives referred patients from all districts of the country with official capacity of 1,790 beds, although it often houses over 3,000 patients. The hospital has 2,057 health care professionals (Africa Health Workforce Observatory, 2009). The study population comprised of both female and male patients attending renal and general outpatient units in Mulago hospital during the study period. The study participants were enrolled purposively until the required sample size of 254 was achieved. And they were of age categories of young, youth, adults and elderly. All processes involved in this study were executed in accordance with the guidelines and regulations stated in the ethical approval document (reference number -MHREC 1347).
Data collection tools and methods
The questionnaire was administered to all the 254 participants in the renal and general out patient’s units while waiting to be examined by the clinicians. This was purposely to obtain (address, gender and age), socioeconomic data (occupation and education level) and information on (contact with rodents, flooding of household and surroundings and water source) (details refer to appendix II). Occupation was grouped into two categories namely Professionals (those with a minimum of certificates academic qualification) and Non-professionals (those without a minimum of a certificate academic qualification).
The research also used patient’s information of body temperature, physical examination and history of previous treatment clerked by the clinician and filled special laboratory request form (details refer to appendix I). This focused on answering the associated factors with the prevalence of leptospirosis.
Blood samples were collected from participants and allowed to stand for between 30 and 60 minutes to clot, then centrifuged for 5 minutes at a speed of 3000 rpm centrifugal force to separate the serum. The Serum was placed in aliquots in the cryovials, packed in the cool box and transported to Central Diagnostic Laboratory found in College of Veterinary and Biosecurity, Makerere University for analysis. During testing at the central Diagnostic Laboratory, the standard microscopic agglutination test (MAT) with a panel of 14 Leptospira serovars belonging to 10 serogroups. These included: Autumnalis represented by (L. interrogans serovar Autumnalis) Ballum (L. borgpetersenii serovar Kenya), Canicola (L. interrogans serovarCanicola), Tarassovi (L.borgapetersenii serovar Tarassovi) Hebdomadis (L.borgapetersenii serovar Nona),Pomona (L. interrogansserovar Pomona), shermani (L.santarosai serovar Shermani), Djasiman (L. interrogans serovar Djasiman), Pyrogenes (L.borgapetersenii serovar Nigeria), Sejroe (L.borgapetersenii serovar Sejroe) Icterohaemorrhagiae (L.kirschneri serovar Sokoline), Icterohaemorrhagiae (L.interrogans Serovar Icterohaemorrhagiae, Copenhageni).
Plate plan was drawn for each run and samples were diluted in the ratio of 1:50 by adding 100ul of test serum in 4.9mls of saline for step one screening. Then two-fold dilutions of each sample up to 1:1600 for step two titration. And 100ul of controls were added to each well as indicated on the Titration plate plan. Finally, 100ul of the test serovars(s) to each of the appropriate wells. The plate setup shaking was for 30 seconds at 50 rpm on a mikrotitre plate shaker (Orbit P4, Labnet International Inc. NJ-USA). The preparation was incubated at 29oC for 2.5 hours and the examined using a stereo microscope at high light intensity for agglutination. And only sera with a titre of ≥ 100 was interpreted as positive.
Data analysis
Data from the questionnaire was first entered in the research laboratory registers and then later double entered into the computer using EPI DATA package 2.1, and then subsequently exported to a statistical software programme R version 3.5.2 (2018-12-20) for analysis. The frequencies and proportions were determined. P values of below 0.05 were considered significant and a confidence interval of 95% was used. Multivariate logistic regression models were done to show the association between independent variables and the dependent variable (leptospirosis). Odds ratios were calculated showing predictive risk factors. Data were summarized in tables and graphs.
Ethics consideration
This study was approved by Mulago National Referral Hospital Research and Ethics Committee (approval number -MHREC 1347). In addition, written informed consent was sought from all the study participants and those that did not consent were allowed to access health services and were excluded from the study. At every subsequent stage of handling samples, every effort was made to ensure that the patient’s identity is protected from third parties: “the use of study numbers instead of the actual patients’ names was adhered to. In circumstances were the participants involved minors (below 18 years old), written informed consent was sought from the parent or guardian that escorted them to the health facility.