Nejvíce citovaný článek - PubMed ID 20227841
Distribution of Mycobacterium avium subsp. avium and M. a. hominissuis in artificially infected pigs studied by culture and IS901 and IS1245 quantitative real time PCR
A total of 152 aerosol and spider web samples were collected: 96 spider's webs in karst areas in 4 European countries (Czech Republic, France, Italy, and Slovakia), specifically from the surface environment (n = 44), photic zones of caves (n = 26), and inside (aphotic zones) of caves (n = 26), 56 Particulate Matter (PM) samples from the Sloupsko-Sosuvsky Cave System (speleotherapy facility; n = 21) and from aerosol collected from the nearby city of Brno (n = 35) in the Czech Republic. Nontuberculous mycobacteria (NTM) were isolated from 13 (13.5%) spider's webs: 5 isolates of saprophytic NTM (Mycobacterium gordonae, M. kumamotonense, M. terrae, and M. terrae complex) and 6 isolates of potentially pathogenic NTM (M. avium ssp. hominissuis, M. fortuitum, M. intracellulare, M. peregrinum and M. triplex). NTM were not isolated from PM collected from cave with the speleotherapy facility although mycobacterial DNA was detected in 8 (14.3%) samples. Temperature (8.2 °C, range 8.0-8.4 °C) and relative humidity (94.7%, range 93.6-96.6%) of air in this cave were relatively constant. The average PM2.5 and PM10 mass concentration was 5.49 µg m-3 and 11.1 µg m-3. Analysed anions (i.e., F-, Cl-, NO2-, SO42-, PO43- and NO3-) originating largely from the burning of wood and coal for residential heating in nearby villages in the surrounding area. The air in the caves with speleotherapy facilities should be monitored with respect to NTM, PM and anions to ensure a safe environment.
A total of 281 guano samples were collected from caves (N = 181) in eight European countries (Bulgaria, Czech Republic, France, Hungary, Italy, Romania, Slovakia and Slovenia) and attics in the Czech R. (N = 100). The correlation of detection of mycobacteria between Ziehl-Neelsen (ZN) microscopy and culture examination and qPCR was strong. ZN microscopy was positive in guano from caves (58.6%) more than double than positivity in guano from attics (21.0%; p < 0.01). From 89 mycobacterial isolates (73 isolates from cave guano and 16 isolates from attics' guano), 68 (76.4%) isolates of 19 sp., ssp. and complex were identified as members of three Groups (M. fortuitum, M.chelonae, and M. mucogenicum) and four complexes (M. avium, M. terrae, M.vaccae, and M.smegmatis). A total of 20 isolates (22.5%) belonged to risk group 1 (environmental saprophytes), 48 isolates (53.9%) belonged to risk group 2 (potential pathogens), and none of the isolates belonged to risk group 3 (obligatory pathogens). When comparing bat guano collected from caves and attics, differences (p < 0.01; Mann-Whitney test) were observed for the electrical conductivity, total carbon, total organic, and total inorganic carbon. No difference (p > 0.05; Mann-Whitney test) was found for pH and oxidation-reduction potential parameters.
For epidemiology studies, a decontamination method using a solution containing 4.0% NaOH and 0.5% tetradecyltrimethylammonium bromide (TDAB) represents a relatively simple and universal procedure for processing heavily microbially contaminated matrices together with increase of mycobacteria yield and elimination of gross contamination. A contamination rate only averaging 7.3% (2.4% in Cluster S; 6.9% in Cluster R and 12.6% in Cluster E) was found in 787 examined environmental samples. Mycobacteria were cultured from 28.5% of 274 soil and water sediments samples (Cluster S), 60.2% of 251 samples of raw and processed peat and other horticultural substrates (Cluster R), and 29.4% of 262 faecal samples along with other samples of animal origin (Cluster E). A total of 38 species of slow and rapidly growing mycobacteria were isolated. M. avium ssp. hominissuis, M. fortuitum and M. malmoense were the species most often isolated. The parameters for the quantitative detection of mycobacteria by PCR can be significantly refined by treating the sample suspension before DNA isolation with PMA (propidium monoazide) solution. This effectively eliminates DNA residue from both dead mycobacterial cells and potentially interfering DNA segments present from other microbial flora. In terms of human exposure risk assessment, the potential exposure to live non-tuberculous mycobacteria can be more accurately determined.
Early detection of biohazardous bacteria that can be misused as biological weapons is one of the most important measures to prevent the spread and outbreak of biological warfare. For this reason, many instrument platforms need to be introduced into operation in the field of biological warfare detection. Therefore the purpose of this study is to establish a new detection panel for biothreat bacteria (Bacillus anthracis, Yersinia pestis, Francisella tularensis, and Brucella spp.) and confirm it by collaborative validation by using a multiplex oligonucleotide ligation followed by polymerase chain reaction and hybridization to microspheres by MagPix detection platform (MOL-PCR). Appropriate specific sequences in bacterial DNA were selected and tested to assemble the detection panel, and MOLigo probes (short specific oligonucleotides) were designed to show no cross-reactivity when tested between bacteria and to decrease the background signal measurement on the MagPix platform. During testing, sensitivity was assessed for all target bacteria using serially diluted DNA and was determined to be at least 0.5 ng/µL. For use as a diagnostic kit and easier handling, the storage stability of ligation premixes (MOLigo probe mixes) was tested. This highly multiplex method can be used for rapid screening to prevent outbreaks arising from the use of bacterial strains for bioterrorism, because time of analysis take under 4 h.
- Klíčová slova
- MOL-PCR, bioterrorism, biothreat bacteria, detection panel, magnetic bead,
- Publikační typ
- časopisecké články MeSH
The monitoring of wastewater treatment plants is important for their proper functioning as well as for re-use of water and also to avoid possible circulation of human or animal pathogens in our environment. The samples in this study originated from a full-scale wastewater treatment plant where the structure of the bacterial community was monitored using 454-pyrosequencing. The composition differed in different parts of the plant. In the effluent, bacteria belonging to phyla Proteobacteria, Actinobacteria, TM7 and Bacteroidetes were most frequently detected. The presence of Mycobacterium sp., Mycobacterium avium, Norovirus, Hepatitis A and E viruses was examined using quantitative real-time PCR. Mycobacterium sp. was detected in the effluent in quantities of up to 10(4) cells/ml. Mycobacterium avium subsp. paratuberculosis and subsp. hominissuis were detected in amounts of up to 10(3) cells/ml, and Norovirus group 1 and 2 were also detected. Our findings show the importance of monitoring and controlling the occurrence of specific pathogens in effluent, mainly because of the negative impact on human health when the water is reused.
The environment is a reservoir of nontuberculous mycobacteria and is considered a source of infection for animals and humans. Mycobacteria can persist in different types of environments for a relatively long time. We have studied their possible internalization into plant tissue through intact, as well as damaged, root systems of different types of plants grown in vitro and under field conditions. The substrate into which plants were seeded was previously contaminated with different strains of Mycobacterium avium (10(8) to 10(10) cells/g of soil) and feces from animals with paratuberculosis. We detected M. avium subsp. avium, hominissuis, and paratuberculosis in the stems and leaves of the plants by both culture and real-time quantitative PCR. The presence of mycobacteria in the plant tissues was confirmed by microscopy. The concentration of mycobacteria found inside plant tissue was several orders of magnitude lower (up to 10(4) cells/g of tissue) than the initial concentration of mycobacteria present in the culture medium or substrate. These findings led us to the hypothesis that plants may play a role in the spread and transmission of mycobacteria to other organisms in the environment.
- MeSH
- bakteriologické techniky MeSH
- endocytóza * MeSH
- kvantitativní polymerázová řetězová reakce MeSH
- listy rostlin mikrobiologie MeSH
- mikroskopie MeSH
- Mycobacterium genetika růst a vývoj fyziologie MeSH
- rostliny mikrobiologie MeSH
- stonky rostlin mikrobiologie MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Non-tuberculous mycobacteria are increasingly described as infectious agents in immunocompromised patients. A 17-year-old male patient suffering from secondary non-Hodgkin's lymphoma and treated with chemotherapeutic agents was admitted to hospital due to pleuropneumonia. Mycobacterium neoaurum was cultured repeatedly from his sputum and, Mycobacterium avium subsp. avium (M. a. avium) was detected by IS901 qPCR from detached fragments of his intestinal mucosa. We attempted to determine the possible sources of infection by analysing environmental samples from the closed oncology unit and conventional unit in the hospital, and from the patient's home residence and places which he frequented. The environment of the patient harboured mycobacteria (41 isolates in total); however, M. neoaurum was not recovered. M. a. avium was detected by qPCR in the environmental samples from a small flock of hens kept by his neighbour. Although it was not confirmed by DNA fingerprinting methods, the M. a. avium infection could have been acquired through the eating of incompletely cooked eggs.
- MeSH
- antituberkulotika terapeutické užití MeSH
- antitumorózní látky terapeutické užití MeSH
- atypické mykobakteriální infekce * MeSH
- ciprofloxacin terapeutické užití MeSH
- hostitel s imunodeficiencí * MeSH
- lidé MeSH
- mikrobiologie životního prostředí MeSH
- mladiství MeSH
- Mycobacterium avium izolace a purifikace MeSH
- nehodgkinský lymfom farmakoterapie MeSH
- netuberkulózní mykobakterie izolace a purifikace MeSH
- sputum mikrobiologie MeSH
- střevní sliznice mikrobiologie MeSH
- tuberkulóza * MeSH
- Check Tag
- lidé MeSH
- mladiství MeSH
- mužské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- kazuistiky MeSH
- práce podpořená grantem MeSH
- Názvy látek
- antituberkulotika MeSH
- antitumorózní látky MeSH
- ciprofloxacin MeSH
Mycobacterium marinum, the cause of chronic systemic infections in fish, occasionally causes granulomatous skin and soft tissue lesions in humans. Cutaneous mycobacterial infection in two patients owing to unusual circumstances is presented in this report. The first patient was infected through improper hygienic behavior, while infection in the second patient was previously misdiagnosed as rheumatoid arthritis and treated with methylprednisolone for a period of three months, which resulted in a rare systemic spread of M. marinum into the bones of the hand, testis, and epididymis. Simultaneously, screening for possible sources of M. marinum infection in patients' aquaria revealed positive fish harboring VNTR profiles identical to those obtained for clinical isolates from patients.
- MeSH
- atypické mykobakteriální infekce diagnóza mikrobiologie patologie MeSH
- bakteriální nemoci kůže diagnóza mikrobiologie patologie MeSH
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- minisatelitní repetice genetika MeSH
- Mycobacterium marinum genetika izolace a purifikace patogenita MeSH
- ryby MeSH
- zvířata MeSH
- Check Tag
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- mužské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- kazuistiky MeSH
- práce podpořená grantem MeSH
Mycobacteria are widely present in diverse aquatic habitats, where they can survive for months or years while some species can even proliferate. The resistance of different mycobacterial species to disinfection methods like chlorination or ozonation could result in their presence in the final tap water of consumers. In this study, the culture method, Mycobacterium tuberculosis complex conventional duplex PCR for detection of non-tuberculous mycobacteria (NTM) and quantitative real-time PCR (qPCR) to detect three subspecies of M. avium species (M. a. avium, M. a. hominissuis, and M. a. paratuberculosis) were used to trace their possible path of transmission from the watershed through the reservoir and drinking water plant to raw drinking water and finally to households. A total of 124 samples from four drinking water supply systems in the Czech Republic, 52 dam sediments, 34 water treatment plant sludge samples, and 38 tap water household sediments, were analyzed. NTM of 11 different species were isolated by culture from 42 (33.9 %) samples; the most prevalent were M. gordonae (16.7 %), M. triplex (14.3 %), M. lentiflavum (9.5 %), M. a. avium (7.1 %), M. montefiorenase (7.1 %), and M. nonchromogenicum (7.1 %). NTM DNA was detected in 92 (76.7 %) samples. By qPCR analysis a statistically significant decrease (P < 0.01) was observed along the route from the reservoir (dam sediments), through water treatment sludge and finally to household sediments. The concentrations ranged from 10(0) to 10(4) DNA cells/g. It was confirmed that drinking water supply systems (watershed-reservoir-drinking water treatment plant-household) might be a potential transmission route for mycobacteria.
- MeSH
- bakteriologické techniky * MeSH
- netuberkulózní mykobakterie izolace a purifikace MeSH
- pitná voda mikrobiologie MeSH
- polymerázová řetězová reakce * MeSH
- prevalence MeSH
- zásobování vodou MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Geografické názvy
- Česká republika MeSH
- Názvy látek
- pitná voda MeSH
"Mycobacterium avium subsp. hominissuis" often causes cervical lymphadenitis in children; its prompt and accurate identification enables adequate therapy, tracing, and prevention. The aims of this study were to determine the causative agent of lymphadenitis using culture, PCR, and triplex quantitative real-time PCR (qPCR) methods with DNA directly isolated from tissue, as well as to identify possible sources of infection from the environment. We confirmed the diagnoses by detecting M. avium subsp. hominissuis using qPCR with DNA directly isolated from lymph node biopsy specimens of two patients. In order to trace the source of infection from the environment, a method of DNA isolation from soil and other environmental samples, such as dust, cobwebs, and compost, was developed. The triplex qPCR examination revealed the presence of M. avium subsp. hominissuis in a high proportion of the environmental samples (42.8% in the first patient's house and 47.6% in the second patient's house). Both patients were also exposed to M. avium subsp. avium, which was present due to the breeding of infected domestic hens. The high infectious dose of M. avium subsp. hominissuis or the increased susceptibility of humans to M. avium subsp. hominissuis compared to M. avium subsp. avium could be the reason why the children were infected with M. avium subsp. hominissuis.
- MeSH
- bakteriologické techniky metody MeSH
- dítě MeSH
- krk mikrobiologie MeSH
- lidé MeSH
- mikrobiologie životního prostředí * MeSH
- molekulární typizace MeSH
- Mycobacterium avium klasifikace genetika růst a vývoj izolace a purifikace MeSH
- polymerázová řetězová reakce metody MeSH
- polymorfismus délky restrikčních fragmentů MeSH
- předškolní dítě MeSH
- tuberkulóza lymfatických uzlin diagnóza mikrobiologie MeSH
- Check Tag
- dítě MeSH
- lidé MeSH
- mužské pohlaví MeSH
- předškolní dítě MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- kazuistiky MeSH
- práce podpořená grantem MeSH
