Characterization of some Brucella species from Zimbabwe by biochemical profiling and AMOS-PCR

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Biofilm formation by Mycobacterium avium isolates originating from humans, swine and birds

<p>Abstract</p> <p>Background</p> <p><it>Mycobacterium avium </it>includes the subspecies <it>avium</it>, <it>silvaticum</it>, <it>paratuberculosis </it>and <it>hominissuis</it>, and <it>M. avium </it>subspecies has been isolated from various environments all over the world including from biofilms in water distribution systems. The aim of this study was to examine isolates of <it>M. avium </it>subsp. <it>avium </it>and <it>M. avium </it>subsp. <it>hominissuis </it>of different origin for biofilm formation and to look for correlations between biofilm formation and RFLP-types, and to standardise the method to test for biofilm formation. In order to determine the best screening method, a panel of 14 isolates of <it>M. avium </it>subsp. <it>avium </it>and <it>M. avium </it>subsp. <it>hominissuis</it>, were tested for their ability to form biofilm in microtiter plates under different conditions. Subsequently, 83 additional isolates from humans, swine and birds were tested for biofilm formation. The isolates were tested for the presence of selected genes involved in the synthesis of glycopeptidolipids (GPLs) in the cell wall of <it>M. avium</it>, which is believed to be important for biofilm formation. Colony morphology and <it>hsp65 </it>sequvar were also determined.</p> <p>Results</p> <p>Nine isolates from swine produced biofilm. There was a significant higher frequency of porcine isolates forming biofilm compared to human isolates. All isolates were previously characterised by IS<it>1311</it>- and IS<it>1245</it>-RFLP typing. The ability to form biofilm did not correlate with the RFLP-type, <it>hsp65 </it>sequevar, colony morphology or the presence of gene sequences related to GPL synthesis.</p> <p>Conclusion</p> <p>The observed differences in biofilm forming abilities between porcine and human isolates raises questions regarding the importance of biofilm formation for infectious potential. The optimised method worked well for screening of multiple isolates.</p

New probes used for IS1245 and IS1311 restriction fragment length polymorphism of Mycobacterium avium subsp. avium and Mycobacterium avium subsp. hominissuis isolates of human and animal origin in Norway

BACKGROUND: Mycobacterium avium is an environmental mycobacterium that can be divided into the subspecies avium, hominissuis, paratuberculosis and silvaticum. Some M. avium subspecies are opportunistic pathogens for animals and humans. They are ubiquitous in nature and can be isolated from natural sources of water, soil, plants and bedding material. Isolates of M. avium originating from humans (n = 37), pigs (n = 51) and wild birds (n = 10) in Norway were examined by IS1245 and IS1311 RFLP using new and specific probes and for the presence of IS901 and ISMpa1 by PCR. Analysis and generation of a dendrogram were performed with the software BioNumerics. RESULTS: IS1311 RFLP provided clear results that were easy to interpret, while IS1245 RFLP generated more complex patterns with a higher discriminatory power. The combination of the two methods gave additional discrimination between isolates. All avian isolates except one were M. avium subsp. avium with two copies of IS1311 and one copy of IS1245, while the isolates of human and porcine origin belonged to M. avium subsp.hominissuis. The isolates from human patients were distributed randomly among the clusters of porcine isolates. There were few identical isolates. However, one isolate from a human patient was identical to a porcine isolate. Regional differences were detected among the porcine isolates, while there was no clustering of human isolates according to type of clinical symptoms or geographical location of the patient's home addresses. CONCLUSION: The results demonstrate that a wide range of M. avium subsp.hominissuis are present in pigs and humans in Norway, and that some of these isolates are very similar. It remains to be determined whether humans are infected from pigs or if they are infected from common environmental sources

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

<p>Abstract</p> <p>Background</p> <p>The 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.</p> <p>Methods</p> <p>A 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.</p> <p>Results</p> <p>Mycobacteria 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 <it>Mycobacterium (M) avium</it>, of these 78% and 22% were <it>M. avium</it> sub sp. <it>Hominissuis</it> and <it>avium</it> respectively. Other mycobacterial species included <it>M. senuense</it> (16%)<it>, M. terrae</it> (7%) and <it>M. asiaticum</it> (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.</p> <p>Conclusions</p> <p>This study demonstrated a high prevalence of NTM infections among slaughter pigs in Mubende district of Uganda. <it>M. avium</it> 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.</p

Molecular characterization of Mycobacterium bovis isolates from Ethiopian cattle

<p>Abstract</p> <p>Background</p> <p>Bovine Tuberculosis (BTB) is a widespread and endemic disease of cattle in Ethiopia. Information relating to genotypic characteristics of <it>Mycobacterium bovis </it>strains affecting the cattle population in Ethiopia is limited. We carried out molecular characterization of <it>M. bovis </it>strains isolated from BTB infected cattle using the spoligotyping technique. The relationship between distribution of spoligotypes and recorded variables was also investigated. A new approach that can numerically reflect the degree of genetic polymorphism in a <it>M. bovis </it>population was also developed. The study was conducted from July 2006 to January 2007 in cattle slaughtered at five representative abattoirs in Ethiopia.</p> <p>Results</p> <p>Forty-five <it>M. bovis </it>isolates were obtained from 406 pathologic tissue specimens collected from 337 carcasses with lesions compatible with BTB. Twelve spoligotypes were identified from 34 distinct strains; with SB1176 as a dominant spoligotype (41.2% of the isolates) followed by SB0133 (14.7%). Comparison of spoligotypes with an <it>M. bovis </it>global database <url>http://www.mbovis.org</url> revealed six new spoligotypes which were subsequently registered in the database with international identification codes of SB1517, SB1518, SB1519, SB1520, SB1521 and SB1522. The majority of strains were obtained from cattle slaughtered at Addis Ababa abattoir. On the basis of the Spoligotype Evolutionary Index, SEI (a numeric expression approach to make standardized comparison of spoligotype evolution), <it>M. bovis </it>isolates from Ethiopia were relatively more heterogeneous (SEI = 3.2) compared to isolates from other countries. This might be attributed to extensive livestock movement linked to trading or seasonal migration, high degree of livestock mingling, and also diversities of the country's agricultural and livestock ecosystems, in addition to lack of disease control measures that led to high infection prevalence. Multiple spoligotype infection was recorded in nine (50%) of infected carcasses and this may indicate the prevailing high degree of super infection.</p> <p>Conclusions</p> <p>This study provided molecular evidence for the widespread distribution of <it>M. bovis </it>in the cattle population in Ethiopia. It also demonstrated a relatively high degree of genetic polymorphism of the isolates. Further molecular investigation of <it>M. bovis </it>strains in humans and other domestic animals is recommended in order to elucidate the zoonotic importance as well as reservoirs and pattern of transmission among various hosts.</p

Isolation and characterization of Mycobacterium bovis strains from indigenous Zambian cattle using Spacer oligonucleotide typing technique

<p>Abstract</p> <p>Background</p> <p>Bovine tuberculosis (BTB), caused by <it>Mycobacterium bovis</it>, has remained a major source of concern to public health officials in Zambia. Previous investigations have used traditional epidemiological methods that are unable to identify the causative agent and from which dynamics of disease dispersion is difficult to discern. The objective of this study was to isolate, characterize and determine the genetic diversity and relatedness of <it>M. bovis </it>from major cattle rearing districts in Zambia by spoligotyping. A total of 695 carcasses were examined and 98 tissues had gross post-mortem lesions compatible with BTB.</p> <p>Results</p> <p>Forty-two out of the ninety-eight suspected tissues examined had culture properties characteristic of mycobacteria from which 31 isolates yielded interpretable spoligotypes. This technique showed good discriminatory power (HGDI = 0.98), revealing 10 different spoligotype patterns. Twenty-seven isolates belonged to one cluster with more than 95% similarity and inside the cluster, one predominant spoligotype was found in 20 (64.5%) of the isolates tested. The highest number of spoligotypes was observed among samples from Namwala district. Spoligotypes from 26 (83.9%) of the isolates belonged to five spoligotypes that have been reported before while the remaining 5 (16.1%) isolates had unique spoligotypes that are being reported for the first time; these have been assigned numbers SB1763 to SB1767. Five of the 6 districts had the predominant spoligotype (SB0120).</p> <p>Conclusion</p> <p>The study has described the dispersion patterns of <it>M. bovis </it>in Zambian cattle for the first time and has identified 5 spoligotype patterns specific to Zambia. The observation of an overlap in the spoligotype pattern SB0120 in 5 of the 6 districts suggests the probability of sharing a common source of infection.</p

MLVA-16 typing of 295 marine mammal Brucella isolates from different animal and geographic origins identifies 7 major groups within Brucella ceti and Brucella pinnipedialis

<p>Abstract</p> <p>Background</p> <p>Since 1994, <it>Brucella </it>strains have been isolated from a wide range of marine mammals. They are currently recognized as two new <it>Brucella </it>species, <it>B. pinnipedialis </it>for the pinniped isolates and <it>B. ceti </it>for the cetacean isolates in agreement with host preference and specific phenotypic and molecular markers. In order to investigate the genetic relationships within the marine mammal <it>Brucella </it>isolates and with reference to terrestrial mammal <it>Brucella </it>isolates, we applied in this study the Multiple Loci VNTR (Variable Number of Tandem Repeats) Analysis (MLVA) approach. A previously published assay comprising 16 loci (MLVA-16) that has been shown to be highly relevant and efficient for typing and clustering <it>Brucella </it>strains from animal and human origin was used.</p> <p>Results</p> <p>294 marine mammal <it>Brucella </it>strains collected in European waters from 173 animals and a human isolate from New Zealand presumably from marine origin were investigated by MLVA-16. Marine mammal <it>Brucella </it>isolates were shown to be different from the recognized terrestrial mammal <it>Brucella </it>species and biovars and corresponded to 3 major related groups, one specific of the <it>B. ceti </it>strains, one of the <it>B. pinnipedialis </it>strains and the last composed of the human isolate. In the <it>B. ceti </it>group, 3 subclusters were identified, distinguishing a cluster of dolphin, minke whale and porpoise isolates and two clusters mostly composed of dolphin isolates. These results were in accordance with published analyses using other phenotypic or molecular approaches, or different panels of VNTR loci. The <it>B. pinnipedialis </it>group could be similarly subdivided in 3 subclusters, one composed exclusively of isolates from hooded seals (<it>Cystophora cristata</it>) and the two others comprising other seal species isolates.</p> <p>Conclusion</p> <p>The clustering analysis of a large collection of marine mammal <it>Brucella </it>isolates from European waters significantly strengthens the current view of the population structure of these two species, and their relative position with respect to the rest of the <it>Brucella </it>genus. MLVA-16 is confirmed as being a rapid, highly discriminatory and reproducible method to classify <it>Brucella </it>strains including the marine mammal isolates. The <it>Brucella2009 </it>MLVA-16 genotyping database available at <url>http://mlva.u-psud.fr/</url> is providing a detailed coverage of all 9 currently recognized <it>Brucella </it>species.</p

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

Molecular epidemiology, drug susceptibility and economic aspects of tuberculosis in mubende district, Uganda

<div><p>Background</p><p>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 <i>Mycobacterium tuberculosis</i> isolated from suspected TB patients. Furthermore, risk factors and economic burdens that could affect the current control strategies were studied.</p><p>Methods</p><p>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 <i>M. tuberculosis</i> recovered during sampling were examined for drug resistance to first line anti-TB drugs using the BACTEC-MGIT960^TMsystem. All isolates were further characterized using deletion analysis, spoligotyping and MIRU-VNTR analysis. Data were analyzed using different software; MIRU-VNTR <i>plus</i>, SITVITWEB, BioNumerics and multivariable regression models.</p><p>Results</p><p><i>M. tuberculosis</i> 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.</p><p>Conclusion</p><p>The study detected <i>M. tuberculosis</i> 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.</p></div

Isolation of non-tuberculous mycobacteria from pastoral ecosystems of Uganda: Public Health significance

<p>Abstract</p> <p>Background</p> <p>The 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.</p> <p>Method</p> <p>A 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.</p> <p>Results</p> <p>Forty-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, <it>M. fortuitum-peregrinum </it>complex, <it>M. avium </it>complex, <it>M. gordonae</it>, and <it>M. nonchromogenicum </it>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.</p> <p>Conclusions</p> <p>The 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.</p