Research facilities
Laboratories, space and equipment:
Immunology
Over the years, the Department of Medical microbiology has built capacity for research and
training in immunology. The department has the first and only fully fledged immunology
training laboratory in the University. The laboratory collaborates with the Makerere University
Walter Reed, Medical Research Council and Join Clinical Research Centre Laboratories. The
laboratory can perform and teach all T-cell and immunoglobulin based immunological
techniques.
Equipment available in the immunology lab: Water baths, centrifuges, pipette aides; ELISA
reader; ELISA washer; ELISA spot; Refrigerators and freezers; Liquid nitrogen tanks.
Clinical Microbiology
The Department started as a clinical microbiology department for the University and has been on
the fore front of promoting clinical microbiology for the University and country. Below is a
summary of the recent developments in the department:
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Clinical Microbiology Laboratory: The Department of Medical Microbiology has the
oldest teaching clinical microbiology laboratory in the country that has embarked on an
international (ISO) accreditation process. This laboratory has taught well qualified and
successful 10 students in Master of Medicine in Microbiology as well as 3 doctoral
students.
Equipment available in the Clinical Microbiology lab: Microscopes; Centrifuges; Freezers;
Refrigerators; Incubators; Fluorescent Microscope; Autoclave; weighing scale.
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BSL-3 Mycobacteriology laboratory: The Department has just completed construction of
a state of the art BSL-3 laboratory that can enable it handle highly infectious agents. It is
the only one in the University. This will enable the Department to practically teach on
wide range infectious agents that no other University in the country can.
Equipment available in the BSL-3 laboratory: A high containment area, well equipped P3
facility; MGIT machine; 2 Level two bio-safety cabinets CO2 incubators; inverted microscopes;
refrigeration facilities.
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Mycology laboratory: The Department of Medical Microbiology has the only clinical
mycology laboratory in the country. It has been instrumental in detecting existing and
new fungal infections in the era of HIV/AIDS. This will be a powerful resource to
disseminate knowledge on diagnosis of fungal diseases.
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Virology: Currently the Department collaborates with Uganda Virus Research Institute to
teach practical virology. This collaboration will continue as we aim to build our own
capacity.
Molecular biology laboratory
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The Molecular Biology Laboratory is the main research and training laboratory in the
department. Research scope is wide, spanning from basic molecular studies to apply and
operational research in infectious diseases that cause most of the disease burden in
Uganda. Other studies focus on TB Molecular Epidemiology, Drug resistance studies and
Molecular Diagnostics services offered to the teaching hospitals and other research
laboratories in Uganda.
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Equipment available in the molecular Diagnostic laboratory: Two Thermocyclers; Real
time PCR machine; Gel documentation systems (UV illuminators and Stratalinker,
networked gel Bioimaging system); Basic equipment for manipulation of DNA/RNA
(Incubators, spectrophotometers, electrophoretic apparatus (vertical and horizontal), etc),
air and water incubators, hybridization oven, Gene quants, Vacuum blotters, pH meter;
Ice making machine, autoclave, water distiller, Refrigerators and freezers, equipped dark
room.
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Molecular Diagnostic laboratory: The Department has acquired funds to construct a
clinical diagnostic molecular laboratory. It will be the first one in the region. This will
tremendously increase the capacity of the Department to teach and introduce molecular
techniques in the diagnosis of ailments in the University and country at large.
Lecture Space of the department of Medical Microbiology
The Department has nine spacious office laboratories, one of which is being used as a computer
room for post-graduate students. There are also two study rooms reserved for MSc and PhD
students. The department has a big teaching laboratory, accommodating up to 400 students. The
College of Health Sciences has provided funds for extensive renovation of this teaching
laboratory: the learning environment has never been better for student in the College of Health
Sciences.
Computer laboratory
The Department has a computer laboratory (6 x 8 meters) for post-graduate students, equipped
with 5 desktop computers, each fully connected to internet.
Personnel
The department of Medical Microbiology has a total number of 15 academic staff members. Two
are full Professors, two associate Professors, two senior lecturers, four full lecturers, three
assistant lecturers and two teaching assistants. Five of the academic staff members are doing
doctoral studies and they are in the advanced stage of their studies. 90% of the support staff
members have first degrees and 50% have master’s degrees.
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Clinical microbiologists: The Department has 7 well trained clinical microbiologists and
5 technologists.
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Immunologists: The department is working with five dedicated immunologists and many
visiting immunologists with keen interest to develop the proposed program.
Library:The College of Health Sciences has a well equipped library containing current editions of text
books of immunology and clinical microbiology which are available to students. The library
also has access to the leading journals and electronic books in immunology and clinical
microbiology. Graduate students are also provided passwords to access medical literature
electronically using the WHO/HINARI initiative (World Health Organization / Health
InterNetwork Access to Research Initiative). In addition, the department also has a fully
equipped book bank.
Graduate studies
The department has two running graduate programmes:
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MMED Microbiology
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Doctoral program: The department has gone ahead and started a doctoral program in
immunology with 5 fully sponsored PhD immunology students. The doctoral program
will also serve as an avenue for the masters graduates. Although it is new, the doctoral
program has been very favourably reviewed by Welcome Trust, Karolinska institute and
Case Western Reserve Universities that are well recognized immunology teaching
institutions. With capacity to handle a doctoral program, it should be feasible to start a
master’s program and later an undergraduate one.
Researchers wishing to utilize the departmental research facilities are welcome.
Download the departmental Research Policy (pdf)
Download application for utilizing the departmental research facilities (pdf)
Research Programmes
The department of Medical Microbiology strongly believes that microbiological research will
play a leading role in solving Uganda’s medical ailments. In order to address the need for
continued research and training in infectious diseases, the department has rapidly transformed
into a vibrant research and training center within the College of Health Sciences. Research in our
department is integrated, spanning from basic research to applied research and operation/clinical
research. We believe in this integration since basic research provides a firm foundation for
applied and operational research. Undergraduate and graduate (masters and PhD) students from a
wide range of the university’s programs are actively involved in our research programs. Upon
finishing, the graduates pursue careers in research or health care or the academia.
Research in the department can be an individual grant support to a departmental faculty,
or a collaborative study between the faculty and any other scientist or study group.
The department’s research focus is mainly in the following areas, mirrored by the
laboratories where various research activities are localized:
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Molecular biology of infectious diseases: based at the Molecular Biology and
diagnostics laboratories
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Immunological research: based at the Immunology laboratory
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Strategic basic research on the biology of pathogens aiming at drug/vaccine
discovery and development of diagnostic tests and molecular epidemiological
tools for screening of infectious diseases
Molecular epidemiology of infectious diseases
Molecular mechanisms of drug resistance
Routine Molecular diagnostics
Immunology of infectious diseases
Pregnancy immunology
Mycobacteriology: based at the BSL3 Mycobacteriology laboratory
TB Vaccine study clinical trials
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Clinical/Operational research: based at the clinical Microbiology and mycology
laboratories
1) Current Research projects in the Molecular Biology Laboratory
Cell to Cell signaling in Mycobacteria- this project is funded by NIH grant #1R01A175637-01.
This project has is supporting three graduate students, one doing a PhD in Molecular
Microbiology and the other two are doing Masters degree in Molecular Biology. All the the
students are in their advanced years of study. PI: Dr Moses L. Joloba
Molecular Biology of Mycobacterium tuberculosis: Molecular Characterization of MTB
complex in Kampala- this project is funded by Sida-Sarec. The project supported a PhD student
through Makerere University-Karolinska Institute collaboration, and a Masters student who
studied the Molecular Epidemiology of Mycobacterium bovis in Kampala. These students
accomplished their studies successfully and their work was published in referral journals. PI: Dr
Moses L. Joloba
Molecular Biology of Mycobacterium tuberculosis: Evaluation of Rapid Methods for
Diagnosis of Multi-Drug Resistant TB in Kampala” This is another Sida-Sarec supported
project. The project is funding one PhD student and one Msc student, now in their final years of
study. A manuscript from this work was published by BMC Infectious diseases, and it is one of
the most highly accessed papers this year. PI: Dr Moses L. Joloba
Molecular Biology of Mycobacterium tuberculosis: Phenotypic and Microevolution of MTC of
Uganda Genotype: this study belongs to the Mycobacteria tuberculosis Strain working groupof
the Tuberculosis Research Union (TBRU).This project is aiming at investigating the role of
different genotypes of MTC in transmission, treatment response, clinical and radiological
presentation of MTC. A PhD student and an international student are supported by this project.
PI: Prof. Henry Boom; CO-PI: Dr Moses L. Joloba
Molecular Biology of Streptococcus pneumoniae: Transformation Efficiencies of Drug
susceptible and Drug Resistant Pneumococci- This study was completed last year and the data
revealed that there were no differences in transformation efficiencies among drug resistant and
drug susceptible serotypes of Streptococcus pneumoniae. The study funded a Tanzanian Masters
student. A manuscript is ready for submission for publication.
PI: Dr Moses L. Joloba
Prevalence of infection with multiple strains of Mycobacterium tuberculosis among patients
with pulmonary tuberculosis in Kampala, Uganda: This study was done with Howard Hughes
funding, aiming at evaluating the presence of multiple strain infections in a high TB burden
country using the Mycobacterial Interspersed Repetitive Units-Variable Number Tandem
Repeats (MIRU-VNTR) molecular typing technique in our laboratory.
This study was pioneered by an international Medical student from the University of Pittsburg,
USA, with initial support from AITRIP and the Forgarty scholarship program. The study also
supports a local Msc student. PI: Kate Dickmanns, CO-PI: Dr Moses L. Joloba
Tuberculosis Drug resistance survey in Kampala: This study is funded by the European Union,
and it has supported two Master’s students: one in Public Health and another in Molecular
biology. PI: Dr Moses L. Joloba
Characterization of Extended spectrum B-lactamases elaborated in Uganda: this is a doctoral
study funded by Makerere University School of Graduate studies and belongs to Dr. F.C.
Najjuka, a lecturer in the department. The study has so far supported up to seven undergraduate
students. PI: Dr F.C. Najjuka
A multi-centre comparative trial of efficacy and safety of sodium stibogluconate (SSG) versus
paromomycin (PM) versus combination of SSG and PM as the first line treatment for visceral
leishmaniasis in Ethiopia, Kenya, Sudan and Uganda. PI: Dr. E. Ssentongo & Prof J.Olobo
Malaria Vaccine Studies in Uganda: Site Preparation, Infrastructure Development, And
Capacity Building For Clinical Trials. PI: Prof. J. Olobo
The efficacy of artemether- lumefantrine therapy and assessment of possible molecular markers
of resistance in Uganda. PI: Dr. Hakim Sendagire
Comparison of the development of thymidine analogue mutations with CD4 monitoring alone
versus CD4 monitoring plus viral load monitoring in naïve HIV-1 individuals on first-line
antiretroviral therapy in Africa. PI: Dr. Hakim Sendagire
The phylogeography of Kaposi’s sarcoma-associated herpesvirus 8 in Uganda. CO-PI: Dr
Henry Kajumbula
Services in the new Molecular Diagnostic Laboratory
Increased emphasis on diagnosis of human diseases by molecular genetic analysis has occurred
in the recent years. Clinicians have become increasingly aware of the tremendous power of
molecular-based tests for the diagnosis of human diseases. Molecular diagnostic tests, originally
developed in a research setting, have been commercially packaged into a myriad of formats that
are sufficiently quick and simple for effective use in clinical diagnosis of human diseases. The
use of these diagnostic assays is becoming increasingly common in many clinical settings in
Uganda, and is likely to extend to major referral hospitals in the near future. Thus, as the
molecular basis of more diseases is elucidated, these technologies will continue to be methods
utilized in clinical settings in the near future. This surge of new molecular-based tests has created
the need for individuals trained in the theory and practice of performing or developing these
tests. Funded by Sida-Sarec, this laboratory is constructed to meet this need. Upon completion,
this laboratory will be fully equipped with facilities to detect all human pathogens and genetic
diseases.
In 2006, the molecular biology laboratory introduced PCR for identification of MTC
infections as a routine for the first time in Uganda. Since then, approx 5000 samples
(culture and sputum) from the Joint Clinical Research Center have been speedily and
accurately worked on. In addition, A number research programmes also bring samples to
the laboratory for identification of MTC. Occasionally, clinicians and health workers bring
suspect specimen to rule out MTC infection from their patients. So we deal with a diverse
source of specimen and samples. Rigorous steps and utmost care are taken to ensure
quality of results disseminated.
Ugandan clinicians, health workers and researchers: We look forward to processing your
samples using robust and reliable molecular techniques
Download PCR protocol for identification of M. tuberculosis
Download IS6110-RFLP Protocol for fingerprinting of M. tuberculosis
Completed projects in the Molecular Biology laboratory
Evaluation of various methods for rapid detection of multi-drug resistant tuberculosis: This
was a WHO/TDR grant support to Moses Joloba. In this study four new methods (one molecular
and other 3 culture based) were compared with the conventional indirect Bactec method for rapid
detection of rifampicin resistance. The methods were compared for turnaround time, cost and
technical ease. Year of completion: 2007. Manuscript published in the open access journal, BMC
Infectoius diseases where it is highly accessed. PI: Dr. Moses Joloba
Role of Cell-Cell signaling homologues in Mycobacteria Signaling: This was an RO3 NIH
grant support to Moses Joloba. A genetic approach was used to identify and characterize
mycobacteria genes that are required for production or sensing extracelluar signals. In addition,
biochemical methods were used to purify signals. The grant was upgraded to RO1 (see cell to
cell signaling in mycobacteria). PI: Dr. Moses Joloba
2) Current Research Projects in the Immunology Laboratory
This is the first and only fully fledged teaching immunology teaching laboratory in the
department and Makerere University. The laboratory collaborates with the Makerere University
Walter Reed, Medical Research Council, Uganda Virus Research Institute and the Joint Clinical
Research Centre Laboratories. The laboratory is fully equipped performs and teaches all T-cell
and immunoglobulin based immunological techniques. The department started a doctoral
program in immunology with 5 fully sponsored PhD immunology students, based in this
laboratory. The doctoral program also serves as an avenue for the training of masters graduates.
Although it is new, the doctoral program has been favorably reviewed by Welcome Trust,
Karolinska institute and Case Western Reserve Universities that are well recognized
immunology teaching institutions.
Collaborative Research studies
The University of Washington PIP (Partners in prevention study) Study: This completed study
was about Randomized Placebo Controlled Trial of HSV-2 Suppression to prevent HIV
transmission among HIV discordant couples. The serology and immunological screening on
patient samples were done in the immunology laboratory.
The University of Washington COS (Couples Observational Study): This is a study among HIV
discordant couples. This study explores the role of HSV-2 suppression in prevention of HIV
transmission in discordant couples. Unlike previous studies, this study includes all participants
irrespective of their level of immune suppression or being on ARVs
University of Washington PrEP Study: This study is about Parallel comparison of Tenofovir
and Emtricitabine/Tenofovir Pre- Exposure Prophylaxis to prevent HIV-1 Acquisition with in
HIV-1 Discordant couples.
The immunological tests done in the laboratory:
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HIV-ELISA
HSV-2 ELISA
Hepatitis B ELISA
RPR + TPHA
Agglutination tests
Western blotting/Immunoblotting
3) Current Research Projects in the BSL3 Mycobacteriology Laboratory
The AERAS Mycobacteriology BSL2 laboratory
Aeras TB vaccine study supported construction of the BSL2 Laboratory in Makerere University.
This laboratory is the first of its kind in the University and it is used for liquid and solid culture
of mycobacteria, Molecular tests as well as Drug susceptibility testing.
Epidemiological Studies towards Phase III TB Vaccine Trials in Uganda
Many TB cases and deaths occur in resource-limited settings in Africa where access to health
services is often limited. Efforts are currently underway for developing and testing of more
effective TB vaccines. Testing of these vaccines in areas where they are most needed has
advantages in spite of the limited capacity to conduct such trials in these settings. Capacity
building activities in these settings including TB epidemiological studies are therefore necessary.
In Uganda, TB vaccine activities to build capacity for future phase III TB vaccine trials are being
trained. Two prospective cohorts of 2500 BCG-vaccinated infants and 7000 adolescents aged 1218 years will be under TB surveillance for two years. The studies are conducted in the
Iganga/Mayuge Demographic Surveillance Site (DSS) located in a rural/peri-urban setting in
Eastern Uganda.
The Biosafety level III Mycobacterialogy Laboratory
The Department of Medical Microbiology College of Health Sciences Makerere University
provided space and other technical support, and AERAS global TB Vaccine foundation funded
the construction and staffing of Bio-safety Level 3 laboratory for the TB vaccine study. The
laboratory has a Director, a Manager, a supervisor, a field Manager, two technologists, one data
Clark, two support staffs and two drivers.
4) Research and services in the Clinical Microbiology laboratory
This is the clinical diagnostic branch of the department of Medical Microbiology. The clinical
laboratory offers affordable and quality diagnostic services to a plethora of specimens from
patients suspected to have infectious diseases. We offer bacterial, parasitic, fungal, and viral
diagnosis detection in the specimens. The laboratory is strategically located centrally in
Kampala, surrounded by a number of hospitals and clinics within the city. The clinical laboratory
has increasingly become the nations referral centre for the public and private hospitals/clinics
within the Kampala area, and countrywide.
The Clinical Laboratory receives specimens from a number of sources. These include Mulago
Hospital, Infectious Diseases Institute, Paediatric Infectious Diseases Institute, Makerere
University-Johns Hopkins University Clinic, Private Clinics and Private not-for-profit health
facilities in Kampala City and upcountry, as well as from walk in individual patients.
Specimens of any type are processed here. Modern techniques raging from biochemical,
serological, culture to molecular biology techniques are employed to identify disease causing
Microorganisms. This work is executed by dedicated personnel composed of Specialist Clinical
Microbiologists, technologists and support staff.
During 2008, the laboratory processed about 7000 specimens.
The major clients for the clinical microbiology laboratory include:
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The Infectious Disease Clinic of The Infectious Disease Institute (IDI)- idi.mak.ac.ug
The Baylor Uganda, Paediatric HIV Clinic
The Makerere University- Johns Hopkins Collaboration
A number of Graduate students (Masters and PhD) doing Research Projects for their
dissertations
Collaborative Research
A joint grant application ‘The Environmental Transmission of the AIDS associated Pathogen
Cryptococcus neoformans in Africa’ has been submitted in collaboration with a group from the
University of Minnesota for funding.
Through this grant, a number of equipment will be bought for the lab.
Accreditation:
The Clinical Microbiology laboratory has embarked on the process of accreditation.
Specimens Tests Performed
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Biopsy Tissues e.g. Pleura, Bone marrow etc
Blood
Corneal scrapping
CSF (Cerebrospinal fluid)
Fluid aspirates (Pleural, Peritoneal, Joint, Pericardial, Cysts, Hydrocele)
Genital Specimens e.g urethral swab, High vaginal swab
Hair, Skin, & Nails for Fungal diagnosis
Nasopharyngeal and oro-pharyngeal swabs
Pus
Semen
Skin snips for Microfilaria
Sputum, Induced sputum, Bronchoalveolar Lavage, Tracheal aspirates
Stool for bacterial pathogens
Stool for parasites
Swabs
Urine
5) The Mycology laboratory
The Department of Medical Microbiology has the only clinical mycology laboratory in the
country. It is instrumental in detecting existing and new fungal infections in the era of
HIV/AIDS. The lab is a powerful resource to disseminate knowledge on diagnosis of fungal
diseases in Uganda.
Undergraduate Research and Industrial Training
The laboratories of the department of medical microbiology provide a vibrant research and
training environment for both undergraduate and graduate students. It is these laboratories
(Clinical Microbiology, Molecular Biology, Molecular Diagnostics, Immunology,
Mycobacteriology and Mycology laboratories) that support students’ research and/or training.
We serve health professionals, research scientists and students’ interests spanning from high
school vacationists, undergraduates, Masters, and PhD candidates. Whether you are a student,
health professional or high caliber Researcher (local or International), the department of Medical
Microbiology warmly welcomes you to visit her research facilities for a friendly discussion.
The molecular biology laboratory and the clinical microbiology labs continue to be hubs for
undergraduate industrial and research training in clinical microbiology and Molecular
techniques. The laboratories have so far trained approx 400 undergraduate students from
undergraduate programmes of Makerere University, Kyambogo University and Kampala
International University. Likewise, the molecular biology laboratory has been a hub for
undergraduate research training. The laboratory has trained approx 50 undergraduate students for
their first degree undergraduate research projects.
Below is a summary of representative students and their research topics.
2004:
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B-lactamase production among Escherichia coli isolate of community origin. By
Katabazi Ashaba Fred, BBLT, Mak.
Supervisor: Florence Najjuka, MBChB, MMED, MSC.
2005:
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Quantitative sputum bacillary response to short course chemotherapy in new and
re-treated adults with pulmonary tuberculosis at JCRC. By Lukyamuzi George,
BBLT, Mak.
Supervisor: Moses L. Joloba, PhD.
2006:
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Comparison of various concentrations of carbolfuchsin in Zeihl-Neelsen for
detection of acid-fast bacilli in sputum samples in Kampala district. By Ezati
Nicholas, BBLT, Mak.
Supervisor: Moses L. Joloba, PhD.
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Time to detection of Mycobacterium tuberculosis in Sputum Cultured on
Lowenstein-Jensen Media: A case study of the National Tuberculosis and Leprosy
Programme Unit, Ministry of Health. By Natukwatsa K. Johnson, BBLT, Mak.
Supervisor: Moses L. Joloba, PhD.
2008:
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Evaluation of PCR for detection of Mycobacterium tuberculosis complex in sputum
smear negative samples using chelex-100 as DNA extraction method. By Nakanjako
Ritah, BBLT, Mak.
Supervisor: Benon B. Asiimwe, PhD & Moses L. Joloba, PhD.
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Effect of cell density on the transformation efficiency of mycobacterium smegamtis
with palsimd DNA. By Mboowa Gerald, BBLT, Mak.
Supervisor: David P. Kateete, BVM, MSC & Moses L. Joloba, PhD
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Improvement of template DNA extraction and determination of optimum
concentration of DNA for PCR detection detection of mycobacterium tuberculosis in
a Ugandan laboratory setting. By Ssenyonjo Andrew, BBLT, Mak.
Supervisor: David P. Kateete, BVM, MSC. & Moses L. Joloba, PhD.
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Direct PCR detection of Mycobacterium tuberculosis complex in cerebralspinal
fluid (CSF) of patients with suspected meningeal tuberculosis in Mulago hospital,
Kampala. By Okeng Alfred, BBLT, Mak.
Supervisor: Bwanga Freddie, MbChB, MMED.
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Detection of the intercellular adhesion D (icaD) gene and biofilm production in a
collection of Staphylococcus epidermidis isolates. By Okee Moses, BBLT, Mak.
Supervisor: David P. Kateete, BVM, MSC & Moses L. Joloba, PhD.
2009
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Comparison of the specificity and sensitivity of sheep and human plasma in
detecting coagulase production by Staphylococcus aureus clinical isolates. By
Ndung’u Kimani Cyrus, BBLT, Mak.
Supervisors: Florence Najjuka, MBChB, MMED, MSC, & David P. Kateete, BVM, MSC.
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Steven Tukwasibwe, BBLT, Mak.
Supervisors: David P. Kateete, BVM, MSC., Moses L. Joloba, PhD & Matovu Enoch, PhD
Graduate Research
Masters student’s research
The laboratories of the department of medical microbiology provide a vibrant research and
training environment for both undergraduate and graduate students. It is these laboratories
(Clinical Microbiology, Molecular Biology, Molecular Diagnostics, Immunology,
Mycobacteriology and Mycology laboratories) that support students’ research and/or training.
We serve health professionals, research scientists and students’ interests spanning from high
school vacationists, undergraduates, Masters, and PhD candidates. Whether you are a student,
health professional or high caliber Researcher (local or International), the department of Medical
Microbiology warmly welcomes you to visit her research facilities for a friendly discussion.
Below is a list of Master’s students who successfully completed their graduate research in
the department of Medical Microbiology. Except where noted, students’ research was
totally funded by the department.
2006:
1) Factors associated with multi-drug resistant tuberculosis among retreatment patients
attending mulago tuberculosis clinic. By Alice Asiimwe Rwego, MbChB, MSC. (Clinical
Epidemiology)-Mak.
Supervisors: Moses L. Joloba, PhD, & Alphonse Okwera, MMED
THESIS ABSTRACT
Tuberculosis (TB) drug resistant tuberculosis (MDR-TB) in particular is emerging as an
increasing important cause of morbidity and death. Drug-resistant tuberculosis is a significant
threat to tuberculosis control because only a few effective drugs are available against m.
tuberculosis. Even the best available treatment is often unsuccessful. MDR-TB among retreatment patients attending the Mulago TB clinic has been found to be 13.3% and is increasing.
The factors associated with MDR-TB are not well known in Uganda.
Objective: to determine the factors associated with multi- drug resistant tuberculosis among retreatment patients attending Mulago TB clinic with the aim of informing strategy on preventing
further spread and development of MDR-TB.
Methods: A case control design, the cases being patients with MDR-TB and the controls being
patients with susceptible TB. A study setting was Mulago TB clinic in Kampala with participants
enrolled from the welcome trust study.
Study participants: all TB patients with MDR-TB who fulfilled the eligibility criteria were
recruited as cases and all TB patients with susceptible TB who fulfilled the eligibility criteria
were recruited as controls.
Sampling: cases and controls were recruited and enrolled consecutively until the number was
realized. Data collection, management and analysis: Data was collected using a semi-structured
questionnaire, entered into Epi-info version 6.04 and exported to SPSS for analysis.
Results: 58.7% of the cases were male with median age of 33 years compared to 66.4% of the
controls with a median age of 36years. Adherence level to anti-TB drugs was low among cases
(55.6%) and controls (56.3%). Having worked in the hospital was the only socio-demographic
factor that was negatively associated with multi-drug resistant tuberculosis (or=0.14; 95%CI
0.03-0.76). A participant who had worked in the hospital was almost 7 times less likely to have
MDR-TB than one who had never. None of the clinical factors (adherence to treatment, HIV
status and admission to hospital) was found to be significantly associated with MDR-TB.
Participants with MDR-TB were 18 times more likely to have heard about MDR-TB than those
who did not (or=17.85, 95% CI 6.45-49.30). Receiving drugs from private clinics/pharmacies
and duration after diagnosis of TB before start of treatment were not significantly associated with
MDR-TB
Conclusion: adherence to TB drugs was low among both cases and controls. Participants who
had MDR-TB had a greater chance of hearing about MDR-TB while working in hospital seemed
to be protective from having MDR-TB.
2) Reliability of multiplex PCR method in detection of methicilin resistance and pantonvalentine leukocidin genes in Staphylococcus aureus. By Freddie Bwanga, MbChB, MMED
(Microbiology)-Mak.
Supervisors: Moses L. Joloba, PhD, & Deogratious H. Kaddu-Mulindwa, PhD
THESIS ABSTRACT
There has been a recent increase in community-acquired methicillin resistant Staphylococcus
aureus infections associated with strains possessing lukS and lukF genes that code for a highly
virulent bi-component cytotoxin, the Panton-Valentine Leukocidin (PVL). Thus detection of
PVL genes in a methicillin resistant S.aureus isolate is a genetic marker of community acquired
infection and could be used to characterize the source of MRSA infection. Methicillin resistance
in S. aureus is predominantly associated with production of penicillin binding protein 2a, which
is encoded by the mecA gene. Currently the mecA and the lukS-lukF genes can be detected in S.
aureus using PCR runs and it is regarded as the reference method. To detect all the three genes,
two separate PCR runs are done. However it is possible to detect the presence of the three genes
in one PCR run which could be cost effective. The aim of this study was to assess the reliability
of the multiplex PCR test compared to the reference individual tests in simultaneous detection of
the mecA and lukS-lukF genes in S.aureus. Individual and multiplex PCR tests for the mecA and
lukS-lukF genes were performed on 220frozen stocks of S. aureus isolated from various clinical
specimens, between 200 and 2006. The sensitivity and specificity of the multiplex test in
detecting the mecA gene were 85.1% and 94.8% respectively. For the PVL genes, the
corresponding values were 55.8% and 98.3% respectively. This study has shown that multiplex
PCR is a sensitive and specific method of detecting the mecA gene but has low sensitivity in
detecting the PVL-encoding genes lukS-lukF in S. aureus. The values obtained in this study were
lower than those in two studies in Canada and Cleveland where the sensitivity and specificity
were 100% for mecA and 98% and 100% for PVL. The multiplex method should be further
evaluated before it is recommended for routine co-detection of the mecA and lukS-lukF genes
3) The distribution of mycobacterium tuberculosis complex species among PTB patients
presenting at the national TB treatment center at Mulago hospital, Kampala, Uganda, by
PCR method. By Candin Godfrey Mawa, MbChB, MMED (Microbiology)-Mak.
Supervisors: Moses L. Joloba, PhD, & William Worodria, MMED.
THESIS ABSTRACT
Tuberculosis (TB), caused by the bacterium mycobacterium tuberculosis complex (MTC) is one
of the deadliest pathogens in human history. Tuberculosis control today relies on case
identification, treatment (case holding) until cure and in certain instances prophylaxis with drugs
such as in case of recent infection without clinical or radiological manifestation in non- BCG
vaccinated children less than 5 years of age, living in close contact with patients and presenting
with a strong positive reaction to tuberculin. The gold standard of TB diagnosis and speciation is
based on the demonstration of AAF bacilli in clinical specimens, culture and setting biochemical
tests. However classification based on physical and biochemical properties are tedious,
interruption subjective and therefore can give ambiguous results. Molecular typing techniques
particularly based on amplification or lack of specific DNA sequences has been found to give
superior results. Despite high prevalence of TB in Uganda species identification of MTC is still a
remote practice, particularly speciation based on molecular techniques, although there have been
attempts to classify these organisms mainly on basis of biochemical tests. Strain typing allows
the tracing of epidemiologically related cases including virulent or MDR strains, identification of
nosocomial infections, differentiations of new cases from relapses and identification of
laboratory contaminants.
This study describes molecular typing of 300 M. tuberculosis complex isolates cultured from
sputum of patients described in the literature based on genomic deletion sequences namely: RD9,
TbD1, RD4, RD1, RD12. The distribution of the molecular type pattern suggests that the
population of M. tuberculosis complex strains isolated from PTB patients attending nation TB
treatment centre at Mulago was predominantly m. tuberculosis constituting 89.3% contrary to the
previous study a decade ago which showed that M. africanum was the predominant species in
Kampala constituting 67%. This time M. africanum constituted only 2.7% of the isolates,22
(7.3%)were identified as M. tuberculosis ancient strain, and 2 (0.7%) were identified as M.
canettii. the study concludes that M.tuberclosis is the predominant MTC species causing TB
among patients aged twelve years and above presenting at the national TB treatment centre at
Mulago hospital, Kampala, Uganda. M. africanum constitutes a small percentage of only 2.7%
and M. canettii 0.7%. From the above findings, the study safely recommends that there is urgent
need to carry out similar speciation studies up- country. Such studies should in addition search
for possible accentuating factors for TB infection such as HIV. The study further recommends
that molecular methods of diagnosis and speciation be imported for routine use in all
microbiology laboratories in this country, as this would promote good TB management and
control.
4) Campylobacter spp isolates and their sensitivity pattern from children with acute
diarrhohea at Mulago hosipital complex Kampala, Uganda. By Mshana Eliatosha, MD,
MMED (Microbiology)- Mak.
Supervisors: Moses L. Joloba, PhD, Deogratious H. Kaddu-Mulindwa, PhD & Kakooza
Mwesigwa Angelina, MMED.
THESIS ABSTRACT
Campylobacter species are a frequent cause of entries and less often of extra intestinal infection
in humans. Infection is usually transmitted through contaminated water and food with animal’s
excreta especially from poultry. Infection rate in broilers in Uganda is 87% but there is no data
on these organisms in human infection in Uganda. In other developing countries infection rate
has been found to be between 5-20%. This study aimed at finding the prevalence of
campylobacter spp among children with acute diarrhea attending Mulago hospital, Kampala,
Uganda.
Main objective: The aim of the study was to determine the proportional of campylobacter spp
infection and the antibacterial sensitivity of these organisms among children with acute diarrhea
at Mulago hospital, Kampala, Uganda.
Materials and methods: A cross sectional study was conducted on 226 children with acute
diarrhea attending Mulago hospital from June to October 2005. Serial sampling method was used
to obtain the sample size. Stool specimens were obtained, examined macroscopically and
microscopically for white blood cells (WBC) and gram stain; and culture was done in microaerophilic environment using blood free campylobacter media (containing cefoperazone,
charcoal, and deoxycholate). Identification was done using gram stain, catalase, oxidase and
susceptibility to nalidixic acid, cephlothin and sodium hippurate hydrolysis. Disc susceptibility
tests for erythromycin (15µg), ampicillin (10µg) and ciprofloxine (5 µg) were done.
Results: Study population was made up of 226 children with acute diarrhea, the majority were
from Kampala district 138 (61.1%), the mean age was 16 months, and 42.5% were infants. A
total of 68 (30.5%) had used antibiotics before stool culture. While blood cells were seen in
56.2% of stool specimen, 78% of the study population didn’t keep any animal at home.
Campylobacter spp were isolated in 21 (9.8%) from 226 stool specimens cultured:
Campylobacter jejuni 17 (80.9%), campylobacter coli 1 (4.8%) campylobacter lari 2 (9.5%) and
campylobacter jeji/coli (4.8%). There was association between presence of white blood cells and
culture results (p=0.001), also there was association between use of antibiotics and low culture
rate of campylobacter spp (p=0.031). There was no association between keeping animals at home
and isolation rate of campylobacter spp (p=0.617) also being an infant did not predispose to
infection with campylobacter spp (p=0.176). All campylobacter isolates were sensitive to
erythromycin using disc susceptibility test, 20% were resistant to ampicilin and only 1 (5%) was
resistant to ciprofloxin. This isolate was identified as campylobacter lari. The sensitivity and
specificity of gram stain in diagnosing campylobacter infection was 76% and 99.5%,
respectively.
Conclusion: Campylobacter infection is prevalent in Ugandan’s in other developing countries.
There is strong association between the presence of white blood cells in stool and positive
culture of campylobacter. Use of antibiotics affects the culture of campylobacter spp and the
gram stain is specific for diagnosing campylobacter infection where facilities are limited
.2007:
1) Aetiology and antimicrobial susceptibility pattern of pyoderma in children presenting to
Mulago hospital. By Joyce N. Balagadde, MBChB, MMED., Mak.
Supervisors: Philippa Musoke, PhD., Fred Kambugu, MMED & Moses L. Joloba, PhD
THESIS ABSTRACT
Introduction: P. yoderma is well-recognizes cause of morbidity in children in the underdeveloped tropical environment. It is curable if diagnosed early and appropriately treated,
otherwise can potentially result in life-threatening complication. Treatment is empiric and relies
on knowledge of the likely etiologic bacteria and their local antibacterial susceptibility pattern.
Objective: To determine the etiology and antibacterial susceptibility pattern of pyoderma in
children presenting to skin clinic and assessment center of Mulago hospital.
Methods: Children aged 2 month to 12 years who fulfilled the eligibility criteria were enrolled.
Clinical diagnostic was followed by specimen collection for bacteria culture and sensitivity
testing HIV testing was done.
Results: We enrolled 398 children. The prevalence of HIV infection was 3.8%. Primary and
secondary pyoderma was diagnosed in 34% and 56% of children respectively.
Ecthyma, folliculitis and furunculosis were the most common primary pyoderma while infected
eczema, tinea capitis, popular urticaria and scabies were the most common secondary pyoderma.
S .aureus and P. pyogenes were recovered from 70% and 43% of children respectively. Gram
negative bacteria were recovered from 8%. Sensitivity of S. aureus to vancomyicin and oxacillin
was 99.6% and 98% respectively. Resistance of S. aureus to penicillin was 97%. Resistance of S.
aureus and S. pyogenes to erythromyicin was 36% and 42% respectively.
Conclusions: S. aureus and S. pyogenes were the predominant etiologic agents of pyoderma.
The prevalence of methicillin resistant S. aureus was very low (2%). Resistance of S. aureus and
S. pyogenes of erythromycin was high.
2) Prevalence of methicillin resistant Staphylococcus aureus among isolates from surgical
site infections in Mulago hospital, Kampala, Uganda. By Ojulong Julius, MbChB, MMED
(Microbiology)- Mak.
Supervisors: Moses L. Joloba, PhD, & Deogratious H. Kaddu-Mulindwa, PhD
THESIS ABSTRACT
Background: Methicillin resistant Staphylococcus aureus( MRSA) is a worldwide health
problem. MRSA isolates are resistant to penicillins and all other B-lactam antibiotics.
Nosocomial MRSA is also resistant to variety of other antibodies classes. MRSA infections are
associated with a high morbidity and mortality particularly in developing countries where more
expensive drugs like vancomycin are affordable. The objective of this study was to determine the
prevalence of MRSA among S. aureus isolates from surgical sites infections in Mulago hospital,
Kampala, Uganda.
Methods: One hundred and eight pus swabs were collected from patients with surgical site
infections. Swabs were introduced for culture at microbiology laboratory faculty of medicine,
Makerere University. S. aurues was identified biochemically. All S. aurues isolates were
subjected to oxacllin agar screen and then tested with a polymerase chain reaction (PCR) assay
for detection of the mecA gene which codes for oxacillin resistance.
Results: Out of the 188 specimen cultures, 54(28.7%) grew S. aurues. Seventeen (31.5) of the 54
isolates were confirmed as MRSA by PCR.
Conclusion: This study shows a high pr
evalence of MRSA in surgical site infections in Mulago hospital.
3) Clinical profile and antimicrobial susceptibility of pnuemococcal bacteria among febrile
patients admitted to the emergency medical ward at Mulago hospital. By Grace
Namayanja, MbChB, MMED., Mak.
Supervisors: Alice Namale, MMED., Moses L. Joloba, PhD, Robert A. Salata, MD & Harriet
Mayanja Kizza, MD, MSC.
THESIS ABSTRACT
Introduction: Streptococcus penumoniae is major cause of morbidity and mortality worldwide
more especially in the immuno-compromised individuals. An estimated 50-60% of in-patients on
the medical wards of Mulago hospital are immuno-compromised due to HIV infection.
Increasing resistance of S. pneumonia bacteraemia to available antimicrobial agents may worsen
clinical outcome in resources constrained settings, there are limited data on prevalence, clinical
profile, and antimicrobial susceptibility of S. pneumonia among hospitalized patients in Uganda.
Objectives: To determine the prevalence, clinical profile and antimicrobial susceptibility
patterns of S. pneumonia bacteraemia among febrile patients admitted to the emergency medical
ward at Mulago hospital.
Methods: Descriptive cross sectional study with follow up of patients with confirmed S.
pneumonia bacteraemia on blood culture. Febrile patients with an oral temperature of greater
than or equal to 37.8°C were sampled consecutively until the sample size was achieved. Using a
standardized questionnaire, data on socio-demographics clinical features and outcome were
collected. Blood was drawn for complete blood count, serum chemistry, bacterial culture and
sensitivity. Data was analyzed using SPSS version 12.0.
Study setting: Emergency medical ward, Mulago hospital Kampala, Uganda.
Study participants: A total number of 386 febrile patients aged 13-81 years, who were admitted
from November 2006 to March 2007 in the emergency medical ward were enrolled.
Results: The prevalence of S. pneumonia bacteraemia was 9.8% (38/386). Of these, 68%
(26/38)were HIV infected. The mean oral temperature was 38.6°C and mean duration of fever
was 3 weeks. Cough was reported by 78.9 %( 30/38) and headache by 376.8 %( 14/38) with a
mean duration of 3 weeks. Cigarette smoking was reported by 15.8%.
Multilobar consolidation on chest x-ray was noted in 58% (11/19). The mean neutrophil
percentage was 77.4= 12.6% with a neutrophilia of greater than or equal to 75% present in 68%
(26/38). Impaired renal function with creatinine of greater than or equal to1.3mg/id was found in
68% (26/38). Cough (p=0.014), chest signs (p=0.048), meningeal signs (p=0.001), neutrophil
percentage (p=0.004), multilobar consolidation (p=0.001) were significantly associated with S.
pneumonia bacteraemia, but cigarette smoking (p=0775) was not. All S. pneumonia bacteraemia
isolates were resistant to contromoxazole, but all were susceptible to cenftriaxone and
ethromycin while only 21.1% were susceptible to penicillin. On follow up of the patients, the
mean hospital stay was 8.6 days; 34.2% developed septicaemia, 28.9% pneumonia while 13.0%
developed meningitis. Complete recovery was noted in 78.9% (30/38), and mortality in 7.9%
(3/386), Staphylococcus aureus 1.6 %( 6/386), Pseudomonas areruginosa 5, E. coli 5, Klebisella
pneumonia 3, Haemophilus influenza 2, A cinetobacter 2 and Norcadia 1.
Conclusion: S. pneumoniae bacteraemia is common among febrile patients admitted on the
medical emergence ward at Mulago hospital. Presentation is characterized by fever, cough, and
headache. The isolates are resistant to contrimoxazole and penicillin which are the commonly
available antibiotics. Mortality is more likely in patients with leucopenia, anemia, HIV infection,
meningitis and dehydration.
4) Accuracy of sputum polymerase chain reaction in the diagnosis of tuberculosis among
sputum smear negative adult PTB suspects in Mulago hospital. By Lydia Nakiyingi,
MbChB, MMED (Internal Medicine)-Mak.
Supervisors: Roy Mugerwa, Harriet Mayanja & Moses Joloba, PhD.
THESIS ABSTRACT
Introduction and rationale: Accurate and early diagnosis of TB is crucial for effective patient
management and TB control. The sensitivity of the available diagnosis method, sputum smear
microscopy is very low; ranging from 30to 70% and it is even lower in TB/HIV co-infected
patients. This has resulted into an increased number of SSN PTB suspects, yet sputum culture for
confirmation of TB is not readily available and not routinely done. Therefore a need for rapid
and accurate tests for the diagnosis of SSN TB, particularly those using molecular techniques
like sputum PCR. Sputum PCR is now available in Uganda in research settings and no study has
been done to evaluate its accuracy in the diagnosis of TB among SSN adult PTB suspects.
Objective: To evaluate the accuracy of in house sputum PCR as compared to sputum culture in
the diagnosis of TB among AFB sputum smear negative adult PTB suspects in Mulago hospital
and to also describe the clinical characteristics associated with positive sputum
PCR.
Methods: This cross sectional study was conducted from September 2007 to February 2008 on
adult patients admitted on the emergency medical wards of Mulago hospital complex. A pretested and standardized questionnaire was administed to consenting PTB suspects who were then
asked to provide 2 early morning sputum samples and a proportion of each sample was subjected
to smear microscopy using ZN staining. SSN PTB suspects meeting the inclusion criteria were
recruited consecutively into the study until a sample size of 205 patients was attained. The
remaining portions were each subjected to sputum culture using LJ media and a proportion of the
second sputum was subjected to sputum PCR after processing
Data was collected using a coded questionnaire, entered using EPI-INFO 6.04 and analyzed
using STATA version 10.0. using LJ sputum culture as the “gold standard”, we analyzed for the
diagnostic accuracy of sputum PCR by computing Sensitivity, specificity, Positive and negative
values as well as diagnostic likelihood ratios. Bivariate analytical methods were conducted to
describe the factors associated with positive sputum PCR.
Results: of the 320 consenting PTB suspects screened, 115 patients were AFB smear positive
and were started on anti-tuberculosis treatment, 205 were AFB smear negative and the inclusion
criteria. Compared to LJ culture, the sensitivity and specificity of the in-house sputum PCR were
75% and 35% respectively and the positive and negative predictive values were 39% and 72.4%
respectively.
Body temperature below 37.5° (or 0.423(0.22-0.82), p- value 0.0009) was significantly
associated with positive sputum PCR among AFB smear negative PTB suspects.
Conclusion: the diagnostic accuracy of the available in-house sputum PCR is low. Body
temperature below 37.5°C is associated with positive PCR in AFB smear negative PTB suspects
Recommendations: the available in- house sputum PCR cannot be adopted as a rapid and
accurate alternative to LJ culture detection of TB in AFB smear negative PTB suspects. The
diagnostic accuracy of this test, therefore, has improved in order for it to be worthwhile and
beneficial. the in-house PCR should be compared to a more sensitive “gold standard” like
mycobacteria growth indicator tube (MIGIT) system and a study should be done to compare
other possible in-house PCR methods in our setting to the “gold standard”.
Evaluation of PCR for direct detection of Mycobacterium tuberculosis complex from
sputum samples. By Alarakol Simon Peter.
Supervisors: Moses L. Joloba, PhD & Nakavuma Jessica, PhD.
THESIS ABSTRACT
BACKGROUND: Tuberculosis (TB) is a public health problem causing up to 3 million deaths
worldwide. In Uganda TB is the third top killer disease especially in the HIV patients.
Routine diagnosis of TB in the laboratory uses ZN microscopy which has low sensitivity.
Objective: this study evaluated PCR for direct detection of MTB complex from sputum samples.
Method: A total of 180 sputum samples were collected from 60 suspected TB patients.
Thirty ZN negative and 30ZN positive were included in the study. Three specimens were
collected from each patient. Specimens were digested in N- Acetyl L- Cysteine (NALC)-4
NaOH. DNA extracted from sputum samples was used for PCR amplification of 1S6110
sequence which is specific for M.tb complex.
Results: The proportion of samples detected by PCR for ZN negative and ZN positive smears
were 46 (51%) and 73 (81.1%) respectively. Using three samples from each patient, PCR
detected M.tb in all (100%) of ZN positive samples and 80% of ZN negative samples.
Conclusion/recommendation: PCR is a highly sensitive diagnostic tool and therefore can be
used in the detection of mycobacterium tuberculosis complex in ZN negative patients.
2008:
1) Prevalence and factors associated with risk of HIV occupational exposure and PEP
utilization among healthcare workers in Mwanza referral hospitals-tanzania. By Samuel
Sumba James, MbChB, MMED., Mak.
Supervisors: Moses L. Joloba, PhD, Sarah Nabwire Ssali, PhD & B. Gumodoka, MMED.
THESIS ABSTRACT
Introduction: Its is estimated that about 40 million people are living with HIV/AIDS globally,
and of those two thirds are sub Saharan Africa. In Tanzania the prevalence of HIV/AIDS among
adults is 7%. The increasing number of persons being treated for HIV associated illness makes it
likely that more health care workers will encounter patients infected with HIV and therefore they
are at a high risk of occupational exposure, more so in developing countries with high incidence
of blood borne viruses and increased risk of occupational injuries. The use of post exposure
prophylaxis (PEP) for HIV reduces the chance of infection.
Objective: The objective of this study was to study prevalence and factors associated with risk
of HIV occupational exposure and PEP utilization among healthcare workers in Mwanza referral
hospital.
Methods A cross sectional study was conducted between January and march 2008 in Mwanza
referral hospital(North west of Tanzania mainland). For quantitative data, a total of 363 health
care workers were initially selected by sampling proportionate to size of each hospital and by
occupational category in the respective hospitals. Consecutive sampling was done within the
different categories (units). Qualitative data were collected from a total of six key informants,
three from each hospital selected purposively.
Results: The overall prevalence of risk of HIV occupational exposure was 33.9% (95% CI=29.038.8). Risk of HIV occupational exposure was high among healthcare workers in Mwanza
regional hospital (Sekou-Toure), (OR 2.44, 95% CI-1.54-3.85). Those working in the department
of surgery and obstetrics were more likely to experience, (OR 1.92, 95% CI= 1.19-3.13) and (OR
1.45, 95% CI=0.84-2.50) respectively. The rate of utilization was higher among those who
worked for forty hours or more per week (OR 5.44,95% CI 1.04-28.59). Health care workers
who knew the procedures for PEP were more likely to utilization the services, ( OR 5.88, 95% CI
1.64-20.00).Some of the possible key barriers to utilization of PEP services for HIV among other
were lack of knowledge and information on PEP services for HIV among healthcare workers,
stigma and professional discrimination.
Conclusion: T his study demonstrates that healthcare workers in Mwanza referral hospital are at
an increased risk of HIV occupational exposure, and the utilization of PEP services for HIV is
suboptimal.
2) Assessment of smear microscopy in a TB programme setting in Kampala, Uganda:
combining blinded re-checking, culture and polymerase chain reaction. By Sande Obondo
James, MbChB, MMED (Microbiology)- Mak.
Supervisors: Moses L. Joloba, PhD., & William Worodria, MbChB, MMED.
THESIS ABSTRACT
Background: In developing countries TB case detection by quality-assured bacteriology using
Acid- Fast direct smear microscopy is one of the DOTS components of the stop TB strategy in
an era of increasing HIV- related TB. Uganda’s TB case detection rate is low may be due to poor
performance of AFB smear microscopy at peripheral laboratories and failure to use new rapid
diagnostic tools such as PCR. Blinded rechecking of smears, the EQA method of AFB smear
microscopy has not been performed against mycobacterial culture in assessing performance of
AFB direct smear microscopy at peripheral laboratories, and the accuracy of per in TB case
detection is not known in our setting.
Objective: This study aimed at determining the magnitude of missed smear-positive cases at
peripheral laboratories in Kampala by performing blinded rechecking of smears against LJ
culture as the gold- standard, and also determines the accuracy of PCR using Lowenstein Jensen
(LJ) culture as the gold- standard.
Methods: This was a cross-sectional study in four health units in Kampala, from February 2008
to may 2008. ZN smears from 296 spot sputum samples of new TB suspects were prepared and
read by technologists at the peripheral laboratories and the re-read (blinded rechecking)by
technologists at reference laboratory. LJ culture and IS6110 PCR were performed on
NaOH/NALC – processed sputum from which the original smears were prepared. HIV status of
participants was determined.
Results: Sixty-eight percent of the TB suspects were HIV- positive, 23% HIV negative and 90%
unknown status. The magnitiutude of missed smear-positive TB cases at peripherical laboratories
was 19.2% (9 missed smear- positive cases) when compared to culture, and 5.7% (3 missed
smear-positive cases) when compared to blinded rechecking. There was 91.8% observed
agreement between blinded rechecking and culture, 98.7% observed agreement between reading
of smears at peripherical laboratory and re- reading at the reference laboratory (Kaapa=0.930).
PCR showed sensitivity, specificity, positive and negative values of 98.0%, 84.6% and 99.4%
respectively in all suspects. Of the 19 culture cases (94.7%). Of the 111 culture- negative smearnegative HIV- positive TB suspects, negative predictive values of PCR in smear- negative HIVpositive TB suspects were 94.7%, 97.3%,85.7% and 99%, respectively.
Conclusion:
Blinded rechecking is a satisfactory EQA method of smear microscopy in our setting. PCR is
very sentive and highly specific in detecting TB cases in smear-negative HIBV-Positive TB
suspects. The magnitude of missed smear- positive cases was worse at peripheral laboratories
than at reference laboratory, but not statically different.
3) Molecular characterization of Mycobacterium bovis isolates from selected slaughter
houses in Kampala. By Jeniffer Asiimwe, BVM, MSC (Molecular Biology)-Mak.
Supervisors: Moses L. Joloba, PhD & Nakavuma Jessica, PhD.
THESIS ABSTRACT
In order to gain an insight into the genetic diversity and geographical sub-structuring of M. bovis
strains in Uganda,170 samples were collected from the major slaughter houses in Kampala, and
cultured for mycobacteria isolation. A total of 21 mycobacteria isolates (12.4% of the samples
collected) were obtained from the study. PCR based identification revealed that 48% (n=10) 0f
the mycobacteria other than tuberculosis (MOTTS). Spoligotyping revealed that three of the M.
bovis isolates had spoligopatterns that had previously been reported in Uganda while the seven
were new. In this study, one of the M. bovis had a spoligo pattern identical to that observed in a
TB-HIV co-infected individual from a parallel study in Rubaga division, Kampala (unpublished
observations). The M. tuberclosis isolates lack spacer 40 which is characteristic to most of the
isolates previously isolated from humans in Uganda. IS6110 RFLP analysis of nine of the M.
bovis isolates obtained in this study revealed that most of these strains (except two)were high
copy number strains with more than five copies of IS6110. Molecular typing of the M. bovis and
M. tuberculosis isolates revealed a high degree of heterogeneity among the strains and a high
level of strain dissemination in a country where cattle movements are not controlled. Although a
few clusters were identified on analysis of the spoligotypes and RFLP patterns, no geographical
sub-structuring was observed. This study also highlights the fact that in regions with high
prevalence of HIV positivity, a cycle of cattle to human to human and human to cattle could be
easily established especially in a country where many communities are economically dependent
upon cattle and are in frequent close contact with them. It is therefore recommended that there
should be a grater degree of co-operation between veterinary and medical policies that will
provide adequate data for the formulation of sustainable control strategies for TB in Uganda.
Download thesis
4) Analysis of HIV-1 subtypes among blood donars in Uganda using a multi-region
hybridization assay. By Bagaya Ssentalo, BBLT, MSC (Molecular Biology)-Mak.
Supervisors: Moses L. Joloba, PhD., Fred Wabwire-Mangen, PhD., and Miguel A. Arroyo, PhD.
THESIS ABSTRACT
Background: Uganda has been a focus of HIV/AIDS intervention efforts, including vaccine
clinical trials. HIV -1 genetic diversity poses challenges for design of efficacious vaccines.
Data on HIV-1 genetic diversity is crucial for design of an effective vaccine in Uganda. Some
previous studies have reported discrepant results probably due to varied sensitivity of different
sub-typing methods and different study populations. Also no study sampled HIV-1 across the
entire country.
Study objectives: (i) to determine the HIV-1 prevalence and (ii) HIV-1 subtype distribution
among blood donors attending donation centers located in five different regions of Nakasero/
Kampala (central), Mbale (eastern), Fortportal (wersten), Mbarara (sourthen), and Gulu (northern
region).
Methodology: 6,192 samples were collected from anonymous blood donors in five regional
blood banks Uganda. All samples were tested for HIV-1 using MUWRP laboratory/FIDA
approved algorithm. HIV-1 viral load and HIV-1 sub typing was performed on HIV-1 positive
samples using Roche Amplicator HIV-1 monitor Test v1.5 and MHAacd respectively.
Results: HIV-1 prevalence among blood donors was 1.3% but was highest in Kampala at 1.8%
and Gulu at 1.5% and was the lowest in Fortportal and Mbarara at0.9% and 1.0% respectively.
Prevalence increased with increased age and was 3.2% among the 34-39 and 3.4% in the 50+ age
groups. HIV-1 subtypes A accounted for 50% of cases, 25% subtype D, 2% subtype C and
recombinants AD made up 20% and 3% for AC. Subtype A was dominant in 4 out of 5 regional
blood banks while subtype D predominated in fort portal. Subtype distribution was compared
across gender but the HIV-1 prevalence was higher in female (1.6%) blood donors than in males
(1.3%)
Conclusions: this study adds information to the HIV-1 subtype distribution in Uganda and
informs vaccine design and clinical trials. HIV- 1 pure sub type A was the most predominant sub
type among blood donors in Uganda and the proportion of pure sub type C was very low in this
study. Recombinant HIV being responsible for almost a quarter of cases among blood; a low risk
population further depicts the increasing role of recombinants and dynamic nature of the
HIV/AIDS pandemic HIV -1 subtype distribution was comparable with respect to gender. This
study demonstrates development of the capacity to genotype HIV-1 using the real-time PCR
based MHA acid technique for the time in Uganda.
5) Bacterial aetiology and antimicrobial susceptibility of chronic suppurative otitis media
in HIV-Infected children attending the paediatric infectious disease clinic in Mulago
hosipital. By Jimmy Sekitoleko, MbChB, MMED., Mak.
Supervisors: Turitwenka Edward, MSc & Moses L. Joloba, PhD
THESIS ABSTRACT
The HIV/AIDS pandemic is one of the most devastating ever seen. Sub-Saharan Africa is one of
the worst hit, although it is home to 10% of the world’s population, 60% of people with
HIV/AIDS live in it. HIV destroys the body’s immune system leading to development of
multiple pathogenic conditions, chronic suppurative otitis media among them. Approximately
15% of children infected with HIV present with chronic suppurative otitis media. Poorly
managed chronic suppurative otitis media can result into complications, among these are;
hearing loss due to tympanic membrane perforation, septicaemia, mastoiditis, facial nerve palsy,
extra and intracranial infections and some of the are fatal. Many antimicrobial agents are on the
Ugandan market; however their efficacy on bacterial agents of C.S.O.M in HIV infected children
is unknown.
Objective: The aim of the study was to determine the bacterial agents of chronic suppurative
otitis media and their antimicrobial susceptibility in HIV infected children.
Study design: This was a case- series study
Study setting: The study was conducted in the pediatric infectious disease clinic located in
Mulago hospital.
Study population: This involved 41 HIV positive children aged between 0-12 years attending
pediatric infectious diseases clinic who met the inclusion criteria.
Outcome measures: The objective was to determine the types of bacterial agent’s of C.S.O.M in
HIV infected children and their antimicrobial susceptibilities to commonly available
antimicrobial agents.
Results: During the study period between October and December 2007, 41 children were
assessed. Bacterial agents isolated in order of percentage frequently included: Proteus mirabilis
(37%), Pseudomonas aeruginosa (21.7%), Klebsiella pneumonae (10.8%), Escherichia coli and
Staphylococcus aureus each contributed (8.7%), Enterobacta (6.5%), Morganella morgani
(4.4%) and finally Streptococcus pneumonae with (2.2%).
Ciprofloxacin was 78% effective on all isolates, followed by gentamycin with effectivity of 65%,
ceftriaxone with 63%, augmentin with 26% and chloramphenicol with 24%.
Chloramphenicol which is commonly used in form of ear drops was found to be effective on
only 24% of all isolates.
Utility of the study results: The findings were recorded on pre-tested data collection sheets and
analyzed. It is hoped that results of this study will contribute into the knowledge base of bacterial
etiology of chronic suppurative otitis media and antimicrobial susceptibility in HIV infected
children.
2009:
1) Prevalence of toxoplasma gondi infection among adult HIV patients in mualgo hospital,
Kampala, Uganda. By Erima Bernard, MbChB, MMED (Microbiology)- Mak.
Supervisors: D.H. Kaddu-Mulindwa, PhD., Fred M. Kironde, PhD., & Edward Ddumba,
MMED.
THESIS ABSTRACT
Introduction: Toxoplasma gondii is a major cause of neurological morbidity and mortality
among patients with advanced acquired immunodeficiency syndrome (AIDS). There are very
few published studies on human toxoplasmosis in Uganda. The magnitude of the problem among
the HIV/AIDS patients in Uganda is not known. The relationship between circulating T- cells
and the infection with Toxoplasma gondii has not been adequately investigated.
Objective: the goal of this study was to determine the prevalence of Toxoplasama gondii
infection and describe its manifestation using the laboratory tests among adult HIV positive
patients attending Mulago Hosipital.
Design: across sectional and descriptive study.
Methods: Three hundred (300) adult HIV infected patients receiving health care on the medical
wards and outpatients clinics at Mulago hospital were enrolled. Three (3) ml of whole blood was
collected for analysis. The circulating CD4+ T-Cell count was determined using FACS caliber
(Becton Dickinson) flow cytometry system. Anti-T. gondii IgG antibodies were screened by an
agglutination technique using Toxoscreen DA kit (bio-merieux); and the presence of T. gondii
specific DNA was examined for using nested PCR in the patients’ blood.
Results: A hundred thirty one (131) males and a hundred sixty nine (169) females were enrolled
in the study. The participants’ age ranged from 18 years, with mean age of 34 years. The mean
CD4+ T cells count was 175 cells/ µL (0.0 to 1361 cells/ µL).
Seventy percent (70%) of samples had CD4+ T-cell count less than 200 cell/ µL.
The ser0- prevalence of T. gondii infection in this study population was 59.7%. out of the 300
samples, 62.0% had T.gondii specific DNA (T. gondii B1 gene). Therefore, 23.3% had acute
toxoplasmosis, 38.7% had reactivated toxoplasmosis, 20% had latent toxoplasmosis, and only
18% participants in the study were not infected with T. gondii. The proportion of patients with
reactivated toxoplasimosis did not have a lower mean CD4+ T cell count compared to those with
latent toxoplasmosis.
Conclusion: The sero- prevalence of toxoplasmosis among HIV/HIV patients attending Mulago
hosipital is very high with significant proportions having acute, latent, or reactivated
toxoplasmosis. Levels of CD4+ T cell count was not related to presence or type of toxoplasma
infection.
2) Benson Kidenya, MD, MSC (Molecular Biology)-Mak.
Supervisors: Moses L. Joloba, PhD., & Nakavuma Jessica, PhD.
THESIS ABSTRACT
Download thesis
3) Mycobacterium tuberculosis genetic diversity in Mbarara, South Western Uganda. By
Joel Bazira. MbChB, MMED-Mbarara Univ.
Supervisors: Moses L. Joloba, PhD.
2010:
Lydia Nabyonga
Sylvia Wanzala
Katabazi Ashaba Freddie
Olia Alex
Eugene
Muyombya William
Gafirita James
Tusubira Evans
Doctoral Student’s Research theses
The laboratories of the department of medical microbiology provide a vibrant research and
training environment for both undergraduate and graduate students. It is these laboratories
(Clinical Microbiology, Molecular Biology, Molecular Diagnostics, Immunology,
Mycobacteriology and Mycology laboratories) that support students’ research and/or training.
We serve health professionals, research scientists and students’ interests spanning from high
school vacationists, undergraduates, Masters, and PhD candidates. Whether you are a student,
health professional or high caliber Researcher (local or International), the department of Medical
Microbiology warmly welcomes you to visit her research facilities for a friendly discussion.
Below is a list of doctoral students who successfully completed their graduate research in
the department of Medical Microbiology. Except where noted, students’ research was
totally funded by the department.
2008:
1) Molecular characterization of mycobacterium tuberculosis complex in Kampala,
Uganda. By Benon B. Asiimwe
Supervisors: Gunilla Kallenius, PhD., Moses L. Joloba, PhD., & Tuija Koivula, PhD.
THESIS ABSTRACT
Uganda is one of the countries with the highest burden of tuberculosis (TB) in sub-saharan
Africa, ranked 16th among the 22 highest-burden countries in the world. Poor peri-urban areas of
developing countries with inadequate living conditions and a high prevalence of HIV infection
have been implicated most in the increase of TB. Different species, strain families and lineages
of the Mycobacterium tuberculosis Complex (MTC) are now known to have differences in
virulence, clinical presentation as well as transmission potential. This study determined the
predominant species as well as strain lineages that cause TB in Rubaga division, Kampala;
analyzed TB transmission in HIV-seropositive and HIV-seronegative TB patients and the
prevalence of resistance to key anti-tuberculosis drugs. Furthermore, the study characterized
cattle-derived isolates of M. bovis from slaughter-cattle at a peri-urban city abattoir so as to set
up a database of M. bovis strains for comparison with infections in humans in future studies.
To achieve this, 386 consecutive newly presenting sputum smear positive patients resident in and
attending TB clinics in Rubaga division were enrolled. Infecting species for 344 cultures were
determined by a solely PCR-based typing panel that determined presence or absence of regions
of difference (RDs) in the MTC; strain types and families were determined by spoligotyping, and
dynamics of TB spread in HIV co-infected vis-à-vis HIV-seronegative TB patients by standard
IS6110-RFLP fingerprinting methodology. All but one of the 344 isolates in the study were M.
tuberculosis, the other being M. bovis. Spoligotyping revealed predominance of the T2 family,
which was in turn predominated by a previously described “Ugandan genotype” group of
strains. Further characterization of 139 Uganda genotype strains revealed an internal deletion in
the RD724 locus, a polymorphism that defines one major sub-lineages of M. tuberculosis
commonly seen in the central African human host population. Resistance to isoniazid was found
in 8.1% of 344 strains, while all 15 (4.4%) strains resistant to rifampicin were also multi-drug
resistant. IS6110-RFLP analysis of isolates from 80 HIV-seronegative patients revealed no
difference in the level of diversity of DNA fingerprints observed in the two serogroups (P =
0.615), patients aged <40 years (P = 0.100), and sex (P = 0.715). However, 54% (99/183) of the
patients shared fingerprints (average cluster size of 2.9), suggesting a high transmission rate in
this community. There was no association between any starin types in the sample with either
drug resistance or HIV sero-status of the patients. Eleven M. bovis and six non tuberculous
mycobacteria were isolated from tissue samples of 87 carcasses. Worryingly, six carcasses
showing obvious and multiple sites of infection were not condemned as unfit for human
consumption, creating a potential for spread of M. bovis in the food chain and to humans through
consumption of contaminated meat, a very important health concern in a resource-poor high
disease-burden setting.
The study has shown that M. tuberculosis is the predominant species of the MTC in Kampala,
and the spoligotype-specific and RD724-deleted “Uganda genotype” the predominant strain type.
The TB epidemic in Kampala is localized, mainly caused by the closely knit T2 spoligotype
family of strains, and strain types common in neighboring countries were minimal. Additionally,
strain types were neither associated with drug resistance, nor HIV sero-status. The study further
showed evidence of a high rate of recent transmission of TB in Rubaga with a high average
cluster size, but infection with an isolate with a fingerprint found to be part of a cluster was not
associated with any demographic or clinical characteristic, including HIV sero-status.
Download thesis
2009:
2) Characterization of in vitro anti-malarial sensitivity and evaluation of genetic
polymorphisms in P. falciparum associated with resistance in Uganda. By Nsobya Samuel,
BBLT, MSC (Molecular Biology), PhD thesis submitted.
2010:
Bwanga, Freddie
RAPID TESTS FOR MULTIDRUG RESISTANT TUBERCULOSIS IN LOW INCOME
SETTINGS
Fredagen den 24 september 2010, kl. 09.00.
Swedish Institute for Infectious Diseases Control Solna, Sweden in Gard-Aulan hall.
ISBN: 978-91-7457-034-2
Supervisors:
1. Moses L. Joloba, PhD, Makerere University College of Health Sciences, Kampala,
Uganda
2. Hoffner S, Karolinska Institute, Stockholm, Sweden
Abstract:
Tuberculosis (TB) is at epidemic levels in the resource-limited settings (RLSs) due to
HIV/AIDS, poverty and insufficient TB control programmes. These factors are also contributing
to TB drug resistance. Patients with multidrug drug resistant tuberculosis (MDR-TB) do not
respond to first line drugs. These patients require unique drug regimens, making it necessary to
routinely screen for MDR-TB. Screening for MDR-TB with the Lowenstein-Jensen proportion
method (LJPM), which is common in the RLSs is a very slow process – taking 2-3 months. More
rapid tests suitable for RLSs are urgently needed. In this thesis, a comparison of the technical and
operational performance of several rapid tests for MDR-TB was done, and the most optimal tests
for RLSs are proposed.
In paper I, a meta-analysis of rapid tests for direct detection of MDR-TB was conducted. The
direct nitrate reductase assay (NRA), microscopic observation drug susceptibility (MODS) and
Genotype® MTBDRplus (GT-DRplus) were highly sensitive and specific, and far more rapid
than the conventional indirect drug susceptibility testing (DST).
In paper II, the NRA, MODS, Mycobacterium Growth Indicator Tube (MGIT 960), GT-DRplus,
Alamar blue, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and resazurin
assays were compared head-to-head for indirect detection of MDR-TB at the National
Tuberculosis Reference Laboratory (NTRL) Kampala. The NRA, MGIT 960, GT-DRplus and
MODS were the most sensitive and specific tests, with significantly shorter time to results
compared to the LJPM.
In paper III, the direct NRA and MODS assays were compared at the NTRL on sputum
specimens from consecutive re-treatment TB patients. Interpretable results were obtained in over
90% of the samples with both assays. The median days to results were 10 with the NRA and 7
with MODS. The direct NRA was more sensitive and specific, and was cheaper.
In paper IV, the sensitivity, specificity, time to results (TTR) and reproducibility of the direct
GTDRplus against the MGIT 960 was assessed. Sensitivity and specificity were 100% and 96%
for detection of rifampicin resistance; 81%, and 100% for isoniazid resistance; and 92%, and
96%, for MDR-TB, respectively. The TTR was 1-3 days, and concordance of results between the
Molecular Laboratory at Makerere University and the FIND Diagnostics Laboratory was 98%.
In paper V, we applied spoligotyping to study the clustering rate and predominant genotypic
strains of 99 MDR-TB strains isolated from patients in Kampala. Eighty-three percent of the
strains occurred in clusters, and the T2 lineage was the largest single cluster.
Conclusion. The direct NRA and the GT-DRplus appear to be the most appropriate tests for
MDR-TB in RLSs. The NRA being the cheapest test can be applied where resources are
extremely limited, while the ultra rapid but commercially available GT-DRplus can be used
where resources permit.
List of papers from the dissertation
Direct susceptibility testing for multi drug resistant tuberculosis: a meta-analysis
Bwanga F, Hoffner S, Haile M, Joloba ML.
BMC Infect Dis, 2009; 9: 67
Evaluation of seven tests for the rapid detection of multidrug-resistant tuberculosis in Uganda.
Bwanga F, Joloba ML, Haile M, Hoffner S.
Int J Tuberc Lung Dis, 2010; 14: 890-895
Direct Nitrate Reductase Assay versus Microscopic Observation Drug Susceptibility for rapid
detection of MDR-TB in Uganda.
Bwanga Freddie, Melles Haile, Sven Hoffner, Emmanuel Ochom, Moses L. Joloba.
Manuscript
Rapid screening of MDR-TB using molecular Line Probe Assay is feasible in Uganda.
Albert H, Bwanga F, Mukkada S, Nyesiga B, Ademun JP, Lukyamuzi G, Haile M, Hoffner S,
Joloba M, OBrien R.
BMC Infect Dis, 2010; 10: 41
High clustering of MDR-TB strains in Kampala, Uganda: Predominance of the T2 lineage.
Bwanga Freddie, William George Muyombya, Sven Hoffner, Melles Haile, Benon Asiimwe,
David Kateete, Fred Katabazi, Jennifer Asiimwe, Maria Wijkander, Moses L Joloba.
Manuscript
International Student’s Research
The department of medical microbiology welcomes international students who wish to train in
infectious diseases research. The department has so far partly supported a couple of international
students either to learn clinical microbiological methods, immunology and molecular techniques
or to carry out an investigation of their choice in an area of infectious diseases.
Listed below are some of the international students who have trained with us.
1. Kate Dickman
Kate Dickmans was an international medical student from the University of Pittsburg, USA, who
pioneered the introduction of MIRU-NVTR technique in our laboratory. She was partly
supported by AITRIP and the Forgarty scholarship program. She left a written manuscript of her
research and will soon be published. Kate Dickman is also credited for supporting research for a
local Master of Molecular Biology student. She is currently at the pediatrics department of
Harvard Medical School.
Research title and aims:
Prevalence of infection with multiple strains of Mycobacterium tuberculosis among
patients with pulmonary tuberculosis in Kampala, Uganda: This study was done with
Howard Hughes funding, aiming at evaluating the presence of multiple strain infections in a high
TB burden country using the Mycobacterial Interspersed Repetitive Units-Variable Number
Tandem Repeats (MIRU-VNTR) molecular typing technique in our laboratory.
Supervisors: Moses L. Joloba & Chris Whalen
Duration: 2007-2009(with breaks in between)
Funding: Howard Hughes, AITRIP and the Forgarty scholarship program.
Nationality: USA
2) Bethany Cadwell
Research title: The use of RFLP and Spoligotyping in understanding transmission of
Mycobactreium tuberculosis Complex in a house hold
Supervisors: Moses L. Joloba
Duration: 2009-2010
Funding: American ambassador Rotarians
Nationality: USA
School of admission: Ohio State University School of Medicine