Browsing by Author "Okee, Moses"

Now showing 1 - 5 of 5
  • Thumbnail Image
    Item
    Application of antibodies to recombinant heat shock protein 70 in immunohistochemical diagnosis of mycobacterium avium subspecies paratuberculosis in tissues of naturally infected cattle
    (Irish veterinary journal, 2017) Okuni, Julius Boniface; Kateete, David Patrick; Okee, Moses; Nanteza, Anna; Joloba, Moses; Ojok, Lonzy
    Detection of Mycobacterium avium subspecies paratuberculosis (MAP) infection is key to the control of Johne’s disease. Immunohistochemistry is one of the methods of detection of MAP infection in tissues. However, unavailability of commercial antibodies that can detect the organism is a limiting factor for the use of immunohistochemistry. This study was aimed at developing an immunohistochemistry method to diagnose MAP in infected tissues using antibodies against MAP recombinant heat shock protein 70kd.MAP Heat shock protein 70 gene was amplified and cloned into an expression vector, Champion pET-SUMO, then expressed in E coli, purified and used to produce polyclonal rabbit antibodies against the Heat shock protein. Immunohistochemistry was performed in 35 MAP infected tissues with anti-HSP70 polyclonal antibodies. All 35 MAP infected tissues were positive for MAP within macrophages, epithelioid cells and giant cells either in clumps or singly as individual bacilli. No positive staining was seen in the three uninfected normal tissues and in MAP infected tissues where primary antibodies were substituted with PBS or pre-immune serum from the same rabbit.Anti-HSP70 produced in this study offers an opportunity for improved diagnosis, screening of MAP in animal tissues and in studies on the pathogenesis of MAP
  • Thumbnail Image
    Item
    Pertussis Prevalence and Its Determinants among Children with Persistent Cough in Urban Uganda
    (PLoS ONE, 2015) Kayina, Vincent; Kyobe, Samuel; Katabazi, Fred A.; Kigozi, Edgar; Okee, Moses; Odongkara, Beatrice; Babikako, Harriet M.; Whalen, Christopher C.; Joloba, Moses L.; Musoke, Philippa M.; Mupere, Ezekiel
    We determined prevalence of pertussis infection and its associated host and environmental factors to generate information that would guide strategies for disease control. Methods In a cross-sectional study, 449 children aged 3 months to 12 years with persistent cough lasting 14 days were enrolled and evaluated for pertussis using DNA polymerase chain reaction (PCR) and ELISA serology tests. Results Pertussis prevalence was 67 (15% (95% Confidence Interval (CI): 12–18)) and 81 (20% (95% CI: 16–24)) by PCR and ELISA, respectively among 449 participating children. The prevalence was highest in children with >59 months of age despite high vaccination coverage of 94% in this age group. Study demographic and clinical characteristics were similar between pertussis and non-pertussis cases. Of the 449 children, 133 (30%) had a coughing household member and 316 (70%) did not. Among 133 children that had a coughing household member, sex of child, sharing bed with a coughing household member and having a coughing individual in the neighborhood were factors associated with pertussis. Children that had shared a bed with a coughing household individual had seven-fold likelihood of having pertussis compared to children that did not (odds ratio (OR) 7.16 (95% CI: 1.24– 41.44)). Among the 316 children that did not have a coughing household member, age <23 months, having or contact with a coughing individual in neighborhood, a residence with one room, and having a caretaker with >40 years of age were the factors associated with pertussis. Age <23months was three times more likely to be associated with pertussis compared to age 24–59 months (OR 2.97 (95% CI: 1.07–8.28)). Conclusion Findings suggest high prevalence of pertussis among children with persistent cough at a health facility and it was marked in children >59 months of age, suggesting the possibility of waning immunity. The factors associated with pertussis varied by presence or absence of a coughing household member.
  • Thumbnail Image
    Item
    Prevalence and Antimicrobial Susceptibility Patterns of Bacteria from Milkmen and Cows with Clinical Mastitis in and around Kampala, Uganda
    (PloS one, 2013) Kateete, David Patrick; Kabugo, Usuf; Baluku, Hannington; Nyakarahuka, Luke; Kyobe, Samuel; Okee, Moses; Najjuka, Christine Florence; Joloba, Moses Lutaakome
    Identification of pathogens associated with bovine mastitis is helpful in treatment and management decisions. However, such data from sub-Saharan Africa is scarce. Here we describe the distribution and antimicrobial susceptibility patterns of bacteria from cows with clinical mastitis in Kampala, Uganda. Due to high concern of zoonotic infections, isolates from milkmen are also described.Ninety seven milk samples from cows with clinical mastitis and 31 nasal swabs from milkmen were collected (one sample per cow/human). Fifty eight (60%) Gram-positive isolates namely Staphylococci (21), Enterococci (16), Streptococci (13), Lactococci (5), Micrococci (2) and Arcanobacteria (1) were detected in cows; only one grew Staphylococcus aureus. Furthermore, 24 (25%) coliforms namely Escherichia coli (12), Klebsiella oxytoca (5), Proteus vulgaris (2), Serratia (2), Citrobacter (1), Cedecea (1) and Leclercia (1) were identified. From humans, 24 Gram-positive bacteria grew, of which 11 were Staphylococci (35%) including four Staphylococcus aureus. Upon susceptibility testing, methicillin-resistant coagulase-negative staphylococci (CoNS) were prevalent; 57%, 12/21 in cows and 64%, 7/11 in humans. However, methicillin-resistant Staphylococcus aureus was not detected. Furthermore, methicillin and vancomycin resistant CoNS were detected in cows (Staphylococcus hominis, Staphylococcus lugdunensis) and humans (Staphylococcus scuiri). Also, vancomycin and daptomycin resistant Enterococci (Enterococcus faecalis and Enterococcus faecium, respectively) were detected in cows. Coliforms were less resistant with three pan-susceptible isolates. However, multidrug resistant Klebsiella, Proteus, Serratia, Cedecea, and Citrobacter were detected. Lastly, similar species grew from human and bovine samples but on genotyping, the isolates were found to be different. Interestingly, human and bovine Staphylococcus aureus were genetically similar (spa-CC435, spa-type t645 corresponding to ST121) but with different susceptibility patterns.CoNS, Enterococci, Streptococci, and Escherichia coli are the predominant pathogens associated with clinical bovine-mastitis in Kampala, Uganda. Multidrug resistant bacteria are also prevalent. While similar species occurred in humans and cows, transmission was not detected.
  • Thumbnail Image
    Item
    Retrospective in silico mutation profiling of SARS-CoV-2 structural proteins circulating in Uganda by July 2021: Towards refinement of COVID-19 disease vaccines, diagnostics, and therapeutics
    (Plos one, 2022) Odongo, Steven; Okella, Hedmon; Ndekezi, Christian; Okee, Moses; Namayanja, Monica; Mujuni, Brian; Sterckx, Yann G. J.; Kizito, Dennison; Mwiine, Frank Nobert; Lutwama, Julius Julian; Ibingira, Charles
    The SARS-CoV-2 virus, the agent of COVID-19, caused unprecedented loss of lives and economic decline worldwide. Although the introduction of public health measures, vaccines, diagnostics, and therapeutics disrupted the spread of the SARS-CoV-2, the emergence of variants poses substantial threat. This study traced SARS-CoV-2 variants circulating in Uganda by July 2021 to inform the necessity for refinement of the intervention medical products. A comprehensive in silico analysis of the SARS-CoV-2 genomes detected in clinical samples collected from COVID-19 patients in Uganda revealed occurrence of structural protein variants with potential of escaping detection, resisting antibody therapy, or increased infectivity. The genome sequence dataset was retrieved from the GISAID database and the open reading frame encoding the spike, envelope, membrane, or nucleocapsid proteins was translated. The obtained protein sequences were aligned and inspected for existence of variants. The variant positions on each of the four alignment sets were mapped on predicted epitopes as well as the 3D structures. Additionally, sequences within each of the sets were clustered by family. A phylogenetic tree was constructed to assess relationship between the encountered spike protein sequences and Wuhan-Hu-1 wild-type, or the Alpha, Beta, Delta and Gamma variants of concern. Strikingly, the frequency of each of the spike protein point mutations F157L/Del, D614G and P681H/R was over 50%. The furin and the transmembrane serine protease 2 cleavage sites were unaffected by mutation. Whereas the Delta dominated the spike sequences (16.5%, 91/550), Gamma was not detected. The envelope protein was the most conserved with 96.3% (525/545) sequences being wild-type followed by membrane at 68.4% (397/580). Although the nucleocapsid protein sequences varied, the variant residue positions were less concentrated at the RNA binding domains. The dominant nucleocapsid sequence variant was S202N (34.5%, 205/595). These findings offer baseline information required for refining the existing COVID-19 vaccines, diagnostics, and therapeutics.
  • Thumbnail Image
    Item
    Rhomboids of Mycobacteria: Characterization Using an aarA Mutant of Providencia stuartii and Gene Deletion in Mycobacterium smegmatis
    (Plos One, 2012) Kateete, David Patrick; Katabazi, Fred Ashaba; Okeng, Alfred; Okee, Moses; Musinguzi, Conrad; Asiimwe, Benon Byamugisha; Kyobe, Samuel; Asiimwe, Jeniffer; Boom, W. Henry; Joloba, Moses Lutaakome
    Rhomboids are ubiquitous proteins with unknown roles in mycobacteria. However, bioinformatics suggested putative roles in DNA replication pathways and metabolite transport. Here, mycobacterial rhomboid-encoding genes were characterized; first, using the Providencia stuartii null-rhomboid mutant and then deleted from Mycobacterium smegmatis for additional insight in mycobacteria.Using in silico analysis we identified in M. tuberculosis genome the genes encoding two putative rhomboid proteins; Rv0110 (referred to as “rhomboid protease 1”) and Rv1337 (“rhomboid protease 2”). Genes encoding orthologs of these proteins are widely represented in all mycobacterial species. When transformed into P. stuartii null-rhomboid mutant (ΔaarA), genes encoding mycobacterial orthologs of “rhomboid protease 2” fully restored AarA activity (AarA is the rhomboid protein of P. stuartii). However, most genes encoding mycobacterial “rhomboid protease 1” orthologs did not. Furthermore, upon gene deletion in M. smegmatis, the ΔMSMEG_4904 single mutant (which lost the gene encoding MSMEG_4904, orthologous to Rv1337, “rhomboid protease 2”) formed the least biofilms and was also more susceptible to ciprofloxacin and novobiocin, antimicrobials that inhibit DNA gyrase. However, the ΔMSMEG_5036 single mutant (which lost the gene encoding MSMEG_5036, orthologous to Rv0110, “rhomboid protease 1”) was not as susceptible. Surprisingly, the double rhomboid mutant ΔMSMEG_4904–ΔMSMEG_5036 (which lost genes encoding both homologs) was also not as susceptible suggesting compensatory effects following deletion of both rhomboid-encoding genes. Indeed, transforming the double mutant with a plasmid encoding MSMEG_5036 produced phenotypes of the ΔMSMEG_4904 single mutant (i.e. susceptibility to ciprofloxacin and novobiocin).Mycobacterial rhomboid-encoding genes exhibit differences in complementing aarA whereby it's only genes encoding “rhomboid protease 2” orthologs that fully restore AarA activity. Additionally, gene deletion data suggests inhibition of DNA gyrase by MSMEG_4904; however, the ameliorated effect in the double mutant suggests occurrence of compensatory mechanisms following deletion of genes encoding both rhomboids.