Adrian Muwonge

Isolation of non-tuberculous mycobacteria from pastoral ecosystems of Uganda: public health significance

BackgroundThe importance of non-tuberculous mycobacteria (NTM) infections in humans and animals in sub-Saharan Africa at the human-environment-livestock-wildlife interface has recently received increased attention. NTM are environmental opportunistic pathogens of humans and animals. Recent studies in pastoral ecosystems of Uganda detected NTM in humans with cervical lymphadenitis and cattle with lesions compatible with bovine tuberculosis. However, little is known about the source of these mycobacteria in Uganda. The aim of this study was to isolate and identify NTM in the environment of pastoral communities in Uganda, as well as assess the potential risk factors and the public health significance of NTM in these ecosystems.MethodA total of 310 samples (soil, water and faecal from cattle and pigs) were examined for mycobacteria. Isolates were identified by the INNO-Lipa test and by 16S rDNA sequencing. Additionally, a questionnaire survey involving 231 pastoralists was conducted during sample collection. Data were analysed using descriptive statistics followed by a multivariable logistic regression analysis.ResultsForty-eight isolates of NTM were detected; 25.3% of soil samples, 11.8% of water and 9.1% from animal faecal samples contained mycobacteria. Soils around water sources were the most contaminated with NTM (29.8%). Of these samples, M. fortuitum-peregrinum complex, M. avium complex, M. gordonae, and M. nonchromogenicum were the most frequently detected mycobacteria. Drinking untreated compared to treated water (OR = 33), use of valley dam versus stream water for drinking and other domestic use (OR = 20), sharing of water sources with wild primates compared to antelopes (OR = 4.6), sharing of water sources with domestic animals (OR = 5.3), and close contact with cattle or other domestic animals (OR = 13.8) were the most plausible risk factors for humans to come in contact with NTM in the environment.ConclusionsThe study detected a wide range of potentially pathogenic NTM from the environment around the pastoral communities in Uganda. Drinking untreated water and living in close contact with cattle or other domestic animals may be risk factors associated with the possibility of humans and animals acquiring NTM infections from these ecosystems.

Q fever abortions in ruminants and associated on-farm risk factors in northern Cyprus

Background Q fever is caused by the obligate intracellular bacterium, Coxiella burnetii [1,2]. This disease is regarded as endemic worldwide, with the exception of New Zealand [3-6]. Cattle, sheep and goats are considered to be the primary source of transmission for humans [7,8]. Humans are infected mainly by inhalation of contaminated aerosols or by the ingestion of infected milk and/or fresh dairy products. In animals, Q fever is mainly subclinical but has especially been associated with reproductive disorders such as late abortions, stillbirths, weak off springs, metritis and infertility in ruminants [8-11]. Abortions during Q fever epizootics have been described in goats and sheep, but rarely documented in dairy cows [7,12]. Domestic pets, such as cats, dogs and wild-domestic birds such as rock doves (Columba livia) and geese (Anser anser) are known to be an additional source of infection [8,12-14]. Previous studies have reported occurrences of C. burnetii in migratory wild birds, rodents and ticks in southern Cyprus [15-17]. More than 40 species of ticks are naturally infected with C. burnetii. However, besides the aerosol route, the significance of ticks in transmitting the disease in ruminants and humans has previously been documented [8,9]. On the other hand, recent studies showed that ticks seem to play a major role in the circulation of C. burnetii in cycles of nature especially in wild life cycles. Ticks are also believed to probably play another crucial role in the transmission of the agent from infected wild vertebrates to domestic animals [5,18,19]. In humans, Q fever is mostly asymptomatic, the acute disease form is mainly limited flu-like illness, pneumonia or hepatitis while the chronic disease manifests with chronic fatigue syndrome or endocarditis [4,5,20]. On the reproductive health point of view, C. burnetii infections are known to cause abortions, stillbirth and premature deliveries in pregnant women. In the past, a series of Q fever outbreaks in both human and animal populations resulting in abortions on the island of Cyprus have been reported [10,21]. Studies done on the islands as far back as the 1970’s showed that Q fever has been an ongoing public health problem. Recently, the prevalence of IgG antibodies against C. burnetii phase II antigens was estimated to be at 52.7% for humans, 48.2% for goats, 18.9% for sheep, and 24% for cows. In this context, control of C. burnetii infection in ruminants is a vital component of public health [19]. There is no known record of humans contracting Q fever in northern Cyprus, which might be linked with lack of routine screenings and/or insufficient diagnostic units for C. burnetii [Northern Cyprus Ministry of Health, 2008]. The diagnostic enigma is that C. burnetii is difficult to culture, and detection in Cyprus was first done by complement fixation of antibodies [22,23]. Lately detection and diagnosis of C. burnetii, has been more effectively done by PCR based techniques, targeting the isocitrate dehydrogenase, superoxide dismutase gene and a transposon-like repetitive region [15,24-28]. The current technological advancement in these techniques has made them the most useful diagnostic tools for detection of C. burnetii in bovine aborted foetuses and ovine genital swabs [29-31]. Since the division of Cyprus in 1974, there has not been any research work on this disease in the northern region. Therefore, the aim of this study was to determine the occurrence of Q fever abortion using a PCR based method on DNA isolated from aborted foetal abomasal contents and placental tissues from ruminants in northern Cyprus. In addition, to determine the on-farm risk factors associated with the disease.

Zoonotic tuberculosis in human beings caused by Mycobacterium bovis—a call for action

Mycobacterium tuberculosis is recognised as the primary cause of human tuberculosis worldwide. However, substantial evidence suggests that the burden of Mycobacterium bovis, the cause of bovine tuberculosis, might be underestimated in human beings as the cause of zoonotic tuberculosis. In 2013, results from a systematic review and meta-analysis of global zoonotic tuberculosis showed that the same challenges and concerns expressed 15 years ago remain valid. These challenges faced by people with zoonotic tuberculosis might not be proportional to the scientific attention and resources allocated in recent years to other diseases. The burden of zoonotic tuberculosis in people needs important reassessment, especially in areas where bovine tuberculosis is endemic and where people live in conditions that favour direct contact with infected animals or animal products. As countries move towards detecting the 3 million tuberculosis cases estimated to be missed annually, and in view of WHOs end TB strategy endorsed by the health authorities of WHO Member States in 2014 to achieve a world free of tuberculosis by 2035, we call on all tuberculosis stakeholders to act to accurately diagnose and treat tuberculosis caused by M bovis in human beings.

African swine fever among slaughter pigs in Mubende district, Uganda

Owing to frequent reports of suspected outbreaks and the presence of reservoir hosts and vectors (warthogs, bushpigs and O. moubata ticks), African swine fever (ASF) is believed to be an endemic disease in Uganda. There have, however, been very few studies carried out to confirm its existence in Uganda. This study was carried out to describe the prevalence of ASF based on pathologic lesions and analysis of serum samples from slaughtered pigs during a suspected outbreak in the Mubende district of Uganda. The study was based on visits to 22 slaughterhouses where individual pigs were randomly selected for a detailed ante-mortem and post-mortem inspections. Sera were also collected for laboratory analysis. A total of 997 pigs (53.7% male and 46.3% female) were examined for lesions suggestive of ASF and sero-positivity of sera for ASF antibodies. The sera were tested using enzyme-linked immunosorbent assay (ELISA) and positive samples were further confirmed with an immunoblot assay. The results showed that 3.8% (38/997) of the pigs examined had clinical signs and post-mortem lesions suggestive of ASF. Two of 997 (0.2%) sera analysed were positive for ASF antibodies. Of the sub-counties investigated, Bagezza (12%) and Kiyuni (11%) had the highest prevalence of lesions suggestive of ASF based on ante- and post-mortem examination results, while Mubende town council (1.7%) had the lowest. This study found a low number of pigs (3.8%) with lesions suggestive of ASF at slaughter and an even lower number of pigs (0.2%) that were seropositive at slaughter, however a significantly higher number of pigs were slaughtered during the outbreak as a strategy for farmers to avoid losses associated with mortality.

Prevalence and associated risk factors of mycobacterial infections in slaughter pigs from Mubende district in Uganda

To date, the public health relevance of mycobacterial infections in pigs is not well investigated despite high risk of infection. Recently, there has been a documented increase in opportunistic infections and risk of acquiring opportunistic mycobacterial infections in HIV/AIDS patients in Mubende district; unfortunately, there has been no published information on the epidemiology of mycobacterial infections in this area. This study was carried out between September 2008 and February 2009. Investigations were done to assess the prevalence and associated risk factors of mycobacterial infections in slaughtered pigs in Mubende district of Uganda. A total of 997 pigs (53.7% male and 46.3% female) from 31 different slaughterhouses were examined for the presence of lesions compatible with TB and mycobacterial infections. Pathologic tissue specimens were collected for culturing and isolation of mycobacteria. A cross-sectional technique was used based on convenient visits to slaughterhouses but random selection of individual slaughtered pigs for a detailed post-mortem inspection on a daily basis. The results reflected a 9.3% and 3.1% (95% CI) prevalence of Mycobacterium species based on necropsy examinations and culture isolation, respectively. The highest prevalence of mycobacterial infection was recorded in Buwekula County (the mixed agro-zone) whilst the lowest was in Kassanda County (pastoral zone). A multivariable logistical regression analysis identified age (P ≤ 0.001) and sex (P ≤ 0.05) as risk factors for mycobacterial infections in pigs. Post-estimation statistics of the regression model evaluation and validation fit it well into the data (HL, χ2 = 5.9; P = 0.69 for necropsy, HL χ2 = 2.9; P = 0.94 for culturing). This study documented a high prevalence of mycobacterial infections in slaughter pigs in Mubende district. The fact that pigs and human often share common housing and environment poses a high risk of zoonotic transmission. This then warrants further molecular investigation to identify the specific Mycobacterium species and their public health importance in this area.

Molecular Epidemiology, Drug Susceptibility and Economic Aspects of Tuberculosis in Mubende District, Uganda

Background Tuberculosis (TB) remains a global public health problem whose effects have major impact in developing countries like Uganda. This study aimed at investigating genotypic characteristics and drug resistance profiles of Mycobacterium tuberculosis isolated from suspected TB patients. Furthermore, risk factors and economic burdens that could affect the current control strategies were studied. Methods TB suspected patients were examined in a cross-sectional study at the Mubende regional referral hospital between February and July 2011. A questionnaire was administered to each patient to obtain information associated with TB prevalence. Isolates of M. tuberculosis recovered during sampling were examined for drug resistance to first line anti-TB drugs using the BACTEC-MGIT960TMsystem. All isolates were further characterized using deletion analysis, spoligotyping and MIRU-VNTR analysis. Data were analyzed using different software; MIRU-VNTR plus, SITVITWEB, BioNumerics and multivariable regression models. Results M. tuberculosis was isolated from 74 out of 344 patients, 48 of these were co-infected with HIV. Results from the questionnaire showed that previously treated TB, co-infection with HIV, cigarette smoking, and overcrowding were risk factors associated with TB, while high medical related transport bills were identified as an economic burden. Out of the 67 isolates that gave interpretable results, 23 different spoligopatterns were detected, nine of which were novel patterns. T2 with the sub types Uganda-I and Uganda-II was the most predominant lineage detected. Antibiotic resistance was detected in 19% and multidrug resistance was detected in 3% of the isolates. Conclusion The study detected M. tuberculosis from 21% of examined TB patients, 62% of whom were also HIV positive. There is a heterogeneous pool of genotypes that circulate in this area, with the T2 lineage being the most predominant. High medical related transport bills and drug resistance could undermine the usefulness of the current TB strategic interventions.

Molecular investigation of multiple strain infections in patients with tuberculosis in Mubende district, Uganda

Multiple strain tuberculosis (TB) infections are now an acceptable facet of tuberculosis epidemiology. Identification of patients infected with more than one strain gives an insight in disease dynamics at individual and population level. This study therefore aimed at identifying multiple strain infections among TB infected patients. Furthermore, to determine factors associated with multiple strain infections in Mubende district of Uganda. A total of 72 Mycobacterium tuberculosis isolates from patients at Mubende regional referral hospital were characterized using 15 loci MIRU-VNTR, Spoligotyping and deletion analysis. Genotypic and epidemiological data were analyzed using MIRU-VNTR plus, Bionumerics software version 6.1 and an exact logistic regression model respectively. Eight (11.1%) of the 72 patients had mixed TB infections. Five were exclusively pulmonary mixed infections while three had both pulmonary and extra-pulmonary infections (Compartmentalized TB infections). Unlike previous studies that have linked this phenomenon to Beijing strains, multiple strains in this study belonged to T2-Uganda, X2 and T1 lineages. Two of the pulmonary mixed infections were resistant to rifampicin or isoniazid. All except one were HIV positive, newly diagnosed cases and urban residents of Mubende district. The study revealed that one in nine urban dwelling, HIV/TB co-infected patient were infected with more than one M. tuberculosis strains. The molecular findings give indications of a vital component of the disease dynamics that is most likely under looked at clinical level.

Non-tuberculous mycobacteria isolated from slaughter pigs in Mubende district, Uganda

BackgroundThe importance of infections caused by non-tuberculous mycobacteria (NTM) in animals and humans has gained considerable recognition during the past few years. In the developed world, where pig production is extensively practiced, studies on mycobacterial infections and related control strategies have received increasing attention. The infections are reported to be caused by a wide spectrum of NTM. Unfortunately, these infections have been less recognized in sub-Saharan Africa owing to lack of awareness and systematic studies. In this study we aimed at isolating and identifying species of mycobacteria involved in causing infections in slaughter pigs in Mubende district of Uganda. Furthermore we wanted to identify factors associated with infection prevalence in the study area.MethodsA total of 363 lymph nodes were collected and cultured for the presence of mycobacteria. Isolates were identified by 16S rDNA gene sequencing. A questionnaire survey was administered to identify production related factors associated with infection prevalence. Data were assembled and analysed using descriptive statistics and mixed effects logistic regression analysis.ResultsMycobacteria were detected in 39 % (143/363) of the examined lymph nodes, 63 % (59/93) of lymph nodes with gross lesions typical of mycobacteriosis and 31% (84/270) of lymph nodes with no visible lesions. Nineteen per cent of the isolated mycobacteria were identified as Mycobacterium (M) avium, of these 78% and 22% were M. avium sub sp. Hominissuis and avium respectively. Other mycobacterial species included M. senuense (16%), M. terrae (7%) and M. asiaticum (6%). This study found free range systems (OR = 3.0; P = 0.034) and use of water from valley dams (OR = 2.0; P = 0.049) as factors associated with high prevalence of mycobacteria in slaughter pigs.ConclusionsThis study demonstrated a high prevalence of NTM infections among slaughter pigs in Mubende district of Uganda. M. avium was the most prevalent of all NTM isolated and identified. Free range system of pig management and valley dam water were the most significant factors associated with NTM prevalence in Mubende district. These findings could be of a major public health concern given that it is in a predominantly pork consuming population with 18% HIV/AIDS prevalence. Therefore, stringent post-mortem inspection at the slaughter houses is of paramount importance to reduce human exposure.

Mycobacterium bovis infections in slaughter pigs in Mubende district, Uganda: a public health concern

BackgroundBovine tuberculosis (TB) caused by Mycobacterium bovis is primarily a disease of ruminants, particularly cattle (Bos primigenius) and buffalo (Syncerus caffer), and is endemic in most developing countries. To date, studies done in Uganda have documented the prevalence of M. bovis in cattle, humans and wild life, in addition to non-tuberculous mycobacteria in pigs. Pigs are increasingly becoming an important component of the livestock sector and share the human ecosystem in rural Uganda. It is therefore of public health interest that they are not a source of human infections. As a follow up to previously published findings on mycobacteria in pigs, this study was aimed at investigating the occurrence and molecular characteristics of M. bovis detected in slaughter pigs in Mubende district, Uganda. One hundred fifty mesenteric lymph nodes with lesions suggestive of mycobacterial infections were collected from approximately one thousand slaughtered pigs in Mubende district over a period of five months. The isolation and identification of M. bovis was done using conventional mycobacteriological methods. Mycobacteria belonging to the Mycobacterium tuberculosis complex (MTC) were identified to species level using deletion analysis. Molecular typing was done using Spoligotyping and MIRU-VNTR analysis. Molecular data were analysed and interpreted using MIRU-VNTR plus, SpolDB4.0 and the Mycobacterium bovis spoligo database.ResultsOf the examined animals, one boar and two sows from Madudu Sub County were infected with M. bovis which presented as lesions of a deep yellow colour and a grit-like texture in the mesenteric lymph nodes. This represents 2% (3/150) of the lymph nodes where lesions suggestive of mycobacterial infections were detected. Molecular analysis revealed that the isolates from the infected pigs showed identical MIRU-VNTR profile and spoligotype (SB1469).ConclusionsThis is the first study documenting the occurrence of M. bovis in slaughter pigs in Uganda, revealing that one in fifty slaughter pigs with suspected lesions in mesenteric lymph nodes were infected. Molecular analysis revealed that the isolates were identical, showing a spoligotype previously reported from humans and cattle in the north eastern part of the Uganda cattle corridor. This finding is of public health importance, therefore there is a need for close cooperation between medical and veterinary professionals in designing and implementing control and prevention measures that safeguard the public from this potential source of zoonotic TB in Uganda.

Characterization of Mycobacterium bovis from Humans and Cattle in Namwala District, Zambia

Tuberculosis remains a major public health problem in Zambia. While human to human transmission of Mycobacterium tuberculosis is of major importance in driving the tuberculosis epidemic, the impact of Mycobacterium bovis transmission from infected cattle is largely unknown. This cross-sectional study aimed at molecular characterization of M. bovis in humans and cattle. A total of 100 human sputum samples and 67 bovine tissues were collected and analyzed for the presence of mycobacteria. Of 65 human samples that harbored acid fast bacteria (AFB), 55 isolates were obtained of which 34 were identified as M. tuberculosis and 2 as M. bovis. AFB-positive bovine samples (n = 67) yielded 47 mycobacterial isolates among which 25 were identified as M. bovis and no M. tuberculosis was found. Among the M. bovis isolates, spoligotyping revealed a high homogeneity in genotypes circulating in Namwala district. Human and cattle isolates shared identical MIRU-VNTR genotypes, suggesting that transmission between the two hosts may occur. Therefore, this study has documented zoonotic TB in human patients in Namwala district of Zambia. However, further molecular epidemiological studies in the study area are recommended.