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Find video protocols related to scientific articles indexed in Pubmed.
Genome Sequence of Yersinia similis Y228T, a Member of the Yersinia pseudotuberculosis Complex.
Genome Announc
PUBLISHED: 03-29-2014
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We report here on the genome sequence of Yersinia similis 228(T) isolated in Germany. The genome has a size of 4.9 Mb and a G+C content of 47% and is predicted to contain 4,135 coding sequences. Annotation of the 60,687-bp extrachromosomal element predicted 67 coding sequences and a G+C content of 47.8%.
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Animal brucellosis in Egypt.
J Infect Dev Ctries
PUBLISHED: 02-18-2014
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Brucellosis is a highly contagious zoonosis that affects the public health and economic performance of endemic as well as non-endemic countries. In developing nations, brucellosis is often a very common but neglected disease. The purpose of this review is to provide insight about brucellosis in animal populations in Egypt and help to understand the situation from 1986 to 2013. A total of 67 national and international scientific publications on serological investigations, isolation, and biotyping studies from 1986 to 2013 were reviewed to verify the current status of brucellosis in animal populations in Egypt. Serological investigations within the national surveillance program give indirect proof for the presence of brucellosis in cattle, buffaloes, sheep, goats, and camels in Egypt. Serologic testing for brucellosis is a well-established procedure in Egypt, but most of the corresponding studies do not follow the scientific standards. B. melitensis biovar (bv) 3, B. abortus bv 1, and B. suis bv 1 have been isolated from farm animals and Nile catfish. Brucellosis is prevalent nationwide in many farm animal species. There is an obvious discrepancy between official seroprevalence data and data from scientific publications. The need for a nationwide survey to genotype circulating Brucellae is obvious. The epidemiologic situation of brucellosis in Egypt is unresolved and needs clarification.
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Detection of Brucella melitensis in bovine milk and milk products from apparently healthy animals in Egypt by real-time PCR.
J Infect Dev Ctries
PUBLISHED: 02-14-2014
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Brucellosis in Egypt is an endemic disease among animals and humans. In endemic developing countries, dairy products produced from untreated milk are a potential threat to public health. The aim of this study was to detect brucellae in milk and milk products produced from apparently healthy animals to estimate the prevalence of contamination.
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Clostridium difficile genotypes in piglet populations in Germany.
J. Clin. Microbiol.
PUBLISHED: 09-11-2013
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Clostridium difficile was isolated from 147 of 201 (73%) rectal swabs of piglets from 15 farms of Lower Saxony and North Rhine-Westphalia. In 14 farms, 14 to 100% (mean, 78%) of the animals tested were culture positive. The rate of isolation was 68% postpartum, increased to 94% in animals 2 to 14 days of age, and declined to 0% for animals 49 days of age and older. There was no link between isolation and antibiotic treatment or diarrhea of piglets. Strains were assigned to 10 PCR ribotypes, and up to 4 PCR ribotypes were found to be present at the same time on a farm. The closely related PCR ribotypes 078 (55%) and 126 (20%) were most frequently recovered and were present in 13 of the 14 positive farms. The comparison of multilocus VNTR (variable number of tandem repeats) analysis (MLVA) data from this study and previously published data on human, porcine, and bovine PCR ribotype 078 isolates from 5 European countries revealed genetic differences between strains of different geographic origin and confirmed the relatedness of human and porcine C. difficile isolates. This study demonstrated that the human-pathogenic PCR ribotypes 078 and 126 are predominant in piglets in Germany. The results suggest that presence of C. difficile is correlated with animal age but not with antibiotic treatment or clinical disease. MLVA indicated that strains of the same geographical origin are often genetically related and corroborated the hypothesis of a close epidemiological connection between human and porcine C. difficile isolates.
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Interlaboratory comparison of intact-cell matrix-assisted laser desorption ionization-time of flight mass spectrometry results for identification and differentiation of Brucella spp.
J. Clin. Microbiol.
PUBLISHED: 07-12-2013
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Classical microbiological diagnosis of human brucellosis is time-consuming, hazardous, and subject to variable interpretation. Intact-cell matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) was evaluated for the routine identification of Brucella spp. Analysis of mass peak patterns allowed accurate identification to the genus level. However, statistical models based on peak intensities were needed for definite species differentiation. Interlaboratory comparison confirmed the reproducibility of the results.
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Isolation and identification of bovine Brucella isolates from Pakistan by biochemical tests and PCR.
Trop Anim Health Prod
PUBLISHED: 07-02-2013
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Brucellosis is endemic in bovines in Pakistan. The Brucella species and biovars involved, however, are unknown. The objectives of the present study were to isolate and characterize brucellae from seropositive milk samples, aborted fetuses, and vaginal swabs of cattle and buffaloes which had recently aborted. The seropositive milk samples, aborted fetuses, and vaginal swabs of cattle and buffaloes were collected from the Potohar Plateau, Pakistan. Isolation of brucellae was done on modified Farrells serum dextrose agar. Isolates were characterized by conventional biotyping methods, while molecular typing was done by genus (B4/B5) and species-specific (Brucella abortus, Brucella melitensis, Brucella ovis, and Brucella suis) polymerase chain reaction (PCR). A total of 30 isolates were recovered from milk (n?=?5), aborted fetuses (n?=?13), and vaginal swabs (n?=?12). Most isolates were from cattle (56.7 %). All of them were identified as B. abortus biovar 1 based on conventional biotyping methods and genus and species-specific PCR. This preliminary study provides the first report on the prevalence of B. abortus biovar 1 in cattle and buffaloes in Pakistan.
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A probabilistic model of absolute auditory thresholds and its possible physiological basis.
Adv. Exp. Med. Biol.
PUBLISHED: 05-30-2013
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Detection thresholds for auditory stimuli, specified in terms of their -amplitude or level, depend on the stimulus temporal envelope and decrease with increasing stimulus duration. The neural mechanisms underlying these fundamental across-species observations are not fully understood. Here, we present a "continuous look" model, according to which the stimulus gives rise to stochastic neural detection events whose probability of occurrence is proportional to the 3rd power of the low-pass filtered, time-varying stimulus amplitude. Threshold is reached when a criterion number of events have occurred (probability summation). No long-term integration is required. We apply the model to an extensive set of thresholds measured in humans for tones of different envelopes and durations and find it to fit well. Subtle differences at long durations may be due to limited attention resources. We confirm the probabilistic nature of the detection events by analyses of simple reaction times and verify the exponent of 3 by validating model predictions for binaural thresholds from monaural thresholds. The exponent originates in the auditory periphery, possibly in the intrinsic Ca(2+) cooperativity of the Ca(2+) sensor involved in exocytosis from inner hair cells. It results in growth of the spike rate of auditory-nerve fibers (ANFs) with the 3rd power of the stimulus amplitude before saturating (Heil et al., J Neurosci 31:15424-15437, 2011), rather than with its square (i.e., with stimulus intensity), as is commonly assumed. Our work therefore suggests a link between detection thresholds and a key biochemical reaction in the receptor cells.
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Presence of Clostridium difficile PCR ribotype clusters related to 033, 078 and 045 in diarrhoeic calves in Germany.
J. Med. Microbiol.
PUBLISHED: 05-02-2013
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This study provides data on the distribution and relationship of C. difficile PCR ribotypes in diarrhoeic calves in Germany. C. difficile was isolated from 176 of 999 (17.6?%) faecal samples or swabs of diarrhoeic calves from 603 farms collected between January 2010 and August 2012 by eight federal laboratories of six states. Strains were assigned to 17 PCR ribotypes. PCR ribotypes 033 (57?%), 078 (17?%) and 045/FLI01 (closest match to 045 in the WEBRIBO database; 9?%) were found the most frequently. Nine per cent of all culture-positive tested animals shed more than one multiple locus variable number tandem repeat analysis (MLVA) or PCR ribotype. Eight PCR ribotypes with related profiles (including 033, 078 and 045/FLI01) representing 92?% of all isolates were grouped into three clusters. Molecular relatedness was supported by the absence of the MLVA locus A6Cd only in clustered strains and identical toxin gene profiles for strains within each cluster. Previously reported mulitilocus sequence typing analysis for PCR ribotypes that were also recovered in this study found identical sequence types and a tcdC deletion (?39 bp) for 033, 045, 078 and 126 (ST-11), confirming this clustering. A different geographical occurrence of PCR ribotypes was shown for cluster 033 (found more frequently in southern Germany) and 045 (found more frequently in northern Germany). This study showed that clusters of C. difficile PCR ribotypes related to 033, 078 and 045 are predominant in diarrhoeic calves in Germany. The high number of strains belonging to PCR ribotype 078 demonstrated that diarrhoeic calves are also potential reservoirs for human pathogenic C. difficile strains.
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Seroprevalence and risk factors associated with brucellosis as a professional hazard in Pakistan.
Foodborne Pathog. Dis.
PUBLISHED: 04-06-2013
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The present study was conducted to determine the seroprevalence and identify risk factors associated with brucellosis in humans at high risk in the Potohar plateau of northeastern Pakistan. A total of 262 serum samples were collected from persons of different occupational groups: veterinary personnel, milkers, abattoir workers, livestock farmers, and others (drivers, security guards, housewives). Data related to gender, age, occupation, contact with animals, brucellosis-related symptoms, consumption of raw milk, and geographical region were collected. The Rose Bengal plate test and the serum agglutination test were performed to determine the seroprevalence of brucellosis. The overall seroprevalence was found to be 6.9% (95% confidence interval [CI]: 4.1, 10.6). Real-time polymerase chain reaction assay showed that all cases were affected by Brucella abortus. Individuals who consumed raw milk had higher odds of brucellosis seropositivity. This is the first report of human brucellosis related to B. abortus in high-risk professionals from Pakistan by the combined use of serological and molecular methods.
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Quantitative analysis of the Brucella suis proteome reveals metabolic adaptation to long-term nutrient starvation.
BMC Microbiol.
PUBLISHED: 03-30-2013
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During the infection process, bacteria are confronted with various stress factors including nutrient starvation. In an in vitro model, adaptation strategies of nutrient-starved brucellae, which are facultative intracellular pathogens capable of long-term persistence, were determined.
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Detection of genetic diversity in Campylobacter jejuni isolated from a commercial turkey flock using flaA typing, MLST analysis and microarray assay.
PLoS ONE
PUBLISHED: 02-20-2013
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Campylobacter is genetically highly diverse and undergoes frequent intraspecific recombination. Turkeys have been identified as an important reservoir for Campylobacter jejuni which is of public health significance. The assessment of the genetic diversity among Campylobacter population is critical for our understanding of the epidemiology of this bacterium. The genetic profiles were different according to the molecular typing methods used. The performance of established flaA genotyping, multilocus sequencing typing (MLST) and DNA microarray assay based on the ArrayTube™ technology was evaluated using 14 Campylobacter jejuni isolated from a commercial turkey flock. The flaA typing was performed using PCR-RFLP with restriction enzymes Sau3AI, AluI, a composite flaA analysis of AluI and Sau3AI and DdeI. The 14 isolates were differentiated into 3, 5, 7 and 9 genotypes, respectively. Entire flaA gene and short variable region (SVR) sequences were analysed. Sequencing of the entire flaA provided 11 different genotypes. flaA-SVR sequence analysis detected 8 flaA alleles and 4 flaA peptides. One new flaA allele type (528) was identified. MLST analysis represented 10 different sequence types (STs) and 5 clonal complexes (CCs). The microarray assay recognised 14 different genotypes. The discriminatory indices were 0.560, 0.802, 0.857, and 0.912 for flaA-RFLP depending on the used enzymes, 0.890 for flaA-SVR, 0.967 for entire flaA sequencing, 0.945 for MLST and 1.00 for the DNA microarray assay. The flaA gene was genetically stable over 20 passages on blood agar. In conclusion, the different typing tools demonstrated a high level of genetic heterogeneity of Campylobacter jejuni in a turkey flock, indicating that a single flock can be infected by multiple genotypes within one rearing cycle. DNA microarray-based assays had the highest discriminatory power when compared with other genotyping tools.
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Why longer song elements are easier to detect: threshold level-duration functions in the Great Tit and comparison with human data.
J. Comp. Physiol. A Neuroethol. Sens. Neural. Behav. Physiol.
PUBLISHED: 01-22-2013
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Our study estimates detection thresholds for tones of different durations and frequencies in Great Tits (Parus major) with operant procedures. We employ signals covering the duration and frequency range of communication signals of this species (40-1,010 ms; 2, 4, 6.3 kHz), and we measure threshold level-duration (TLD) function (relating threshold level to signal duration) in silence as well as under behaviorally relevant environmental noise conditions (urban noise, woodland noise). Detection thresholds decreased with increasing signal duration. Thresholds at any given duration were a function of signal frequency and were elevated in background noise, but the shape of Great Tit TLD functions was independent of signal frequency and background condition. To enable comparisons of our Great Tit data to those from other species, TLD functions were first fitted with a traditional leaky-integrator model. We then applied a probabilistic model to interpret the trade-off between signal amplitude and duration at threshold. Great Tit TLD functions exhibit features that are similar across species. The current results, however, cannot explain why Great Tits in noisy urban environments produce shorter song elements or faster songs than those in quieter woodland environments, as detection thresholds are lower for longer elements also under noisy conditions.
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Biotyping and Genotyping (MLVA16) of Brucella abortus Isolated from Cattle in Brazil, 1977 to 2008.
PLoS ONE
PUBLISHED: 01-01-2013
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Brucellosis is a worldwide distributed zoonosis that causes important economic losses to animal production. In Brazil, information on the distribution of biovars and genotypes of Brucella spp. is scarce or unavailable. This study aimed (i) to biotype and genotype 137 Brazilian cattle isolates (from 1977 to 2008) of B. abortus and (ii) to analyze their distribution. B. abortus biovars 1, 2 and 3 (subgroup 3b) were confirmed and biovars 4 and 6 were first described in Brazil. Genotyping by the panel 1 revealed two groups, one clustering around genotype 40 and another around genotype 28. Panels 2A and 2B disclosed a high diversity among Brazilian B. abortus strains. Eighty-nine genotypes were found by MLVA16. MLVA16 panel 1 and 2 showed geographic clustering of some genotypes. Biotyping and MLVA16 genotyping of Brazilian B. abortus isolates were useful to better understand the epidemiology of bovine brucellosis in the region.
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Comparison of diagnostic tests for the detection of Brucella spp. in camel sera.
BMC Res Notes
PUBLISHED: 12-06-2011
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Brucellosis in livestock causes enormous losses for economies of developing countries and poses a severe health risk to consumers of dairy products. Little information is known especially on camel brucellosis and its impact on human health. For surveillance and control of the disease, sensitive and reliable detection methods are needed. Although serological tests are the mainstay of diagnosis in camel brucellosis, these tests have been directly transposed from cattle without adequate validation. To date, little information on application of real-time PCR for detection of Brucella in camel serum is available. Therefore, this study was performed to compare the diagnostic efficiency of different serological tests and real-time PCR in order to identify the most sensitive, rapid and simple combination of tests for detecting Brucella infection in camels.
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An improved model for the rate-level functions of auditory-nerve fibers.
J. Neurosci.
PUBLISHED: 10-28-2011
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Acoustic information is conveyed to the brain by the spike patterns in auditory-nerve fibers (ANFs). In mammals, each ANF is excited via a single ribbon synapse in a single inner hair cell (IHC), and the spike patterns therefore also provide valuable information about those intriguing synapses. Here we reexamine and model a key property of ANFs, the dependence of their spike rates on the sound pressure level of acoustic stimuli (rate-level functions). We build upon the seminal model of Sachs and Abbas (1974), which provides good fits to experimental data but has limited utility for defining physiological mechanisms. We present an improved, physiologically plausible model according to which the spike rate follows a Hill equation and spontaneous activity and its experimentally observed tight correlation with ANF sensitivity are emergent properties. We apply it to 156 cat ANF rate-level functions using frequencies where the mechanics are linear and find that a single Hill coefficient of 3 can account for the population of functions. We also demonstrate a tight correspondence between ANF rate-level functions and the Ca(2+) dependence of exocytosis from IHCs, and derive estimates of the effective intracellular Ca(2+) concentrations at the individual active zones of IHCs. We argue that the Hill coefficient might reflect the intrinsic, biochemical Ca(2+) cooperativity of the Ca(2+) sensor involved in exocytosis from the IHC. The model also links ANF properties with properties of psychophysical absolute thresholds.
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Development of a diagnostic multiplex polymerase chain reaction microarray assay to detect and differentiate Brucella spp.
Diagn. Microbiol. Infect. Dis.
PUBLISHED: 08-01-2011
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Brucellosis is a worldwide zoonosis leading to tremendous economic losses and severe human illness. Fast and reliable laboratory tests are needed to detect disease in both humans and animals and to monitor the production of safe food products and feed. For rapid identification of the genus Brucella and differentiation of its species, a multiplex polymerase chain reaction microarray assay based on 11 signature sequences and redundant oligonucleotide probes was developed. The gene targets included genus-specific sequences in bcsp31, perA, cgs, and omp2b, as well as chromosomal regions displaying species-specific hybridization patterns. Brucella reference strains and a representative panel of 102 field isolates were unambiguously identified by their hybridization patterns. The differentiation of species, however, was limited in members of the groups B. suis bv 3/4/B. canis and B. neotomae/B. microti. In summary, the newly developed Brucella ArrayTube® assay is an easy-to-handle molecular test for high-throughput and parallel analysis.
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Brucellosis in camels.
Res. Vet. Sci.
PUBLISHED: 04-26-2011
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Camels are highly susceptible to brucellosis caused by Brucella melitensis and Brucella abortus. Difficulties can arise in diagnosis of camel brucellosis, especially as this disease provokes only few clinical signs in contrast to its clinical course in cattle. Because none of the commonly used serological test can be perceived as a perfect test for Brucella diagnosis in camel and most serological tests used for camels have been directly transposed from cattle without adequate validation, an incorrect diagnosis may occur when diagnosis is based on serology alone. Of imminent concern is the fact that brucellosis can be easily transmitted from animals or their products to humans mainly via milk. In many developing countries in the arid areas of Asia and Africa, camels are still the most important productive livestock for nomadic populations. Therefore, we reviewed the literatures on camel brucellosis to highlight the epidemiologic, economic and public health impact of camel brucellosis as a basis for designing effective control strategies.
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Isolation and characterization of Ochrobactrum anthropi and Ochrobactrum pecoris from caecal content of commercial turkeys.
Vet. Microbiol.
PUBLISHED: 03-08-2011
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Two Gram negative, micro-aerophilic, non-motile and non-spore-forming coccoid bacteria were isolated from female turkey caecal samples collected from a slaughterhouse. The biochemical reaction profiles (API 20 E and API 20 NE) typed both strains as Ochrobactrum anthropi. On the basis of 16S rRNA gene and recA gene sequence similarities the strains were identified as O. anthropi and Ochrobactrum pecoris, respectively. Both strains were highly resistant against beta-lactam antibiotics, chloramphenicol and sulphonamides but variable in susceptibility to ciprofloxacin, gentamicin and tetracycline. This is the first time that Ochrobactrum species were isolated from an avian host, i.e. turkey.
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Assessment of a novel multiplex real-time PCR assay for the detection of the CBPP agent Mycoplasma mycoides subsp. mycoides SC through experimental infection in cattle.
BMC Vet. Res.
PUBLISHED: 02-17-2011
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Mycoplasma mycoides subsp. mycoides SC is the pathogenic agent of contagious bovine pleuropneumonia (CBPP), the most important disease of cattle in Africa causing significant economic losses. The re-emergence of CBPP in Europe in the 1980s and 1990s illustrates that it is still a threat also to countries that have successfully eradicated the disease in the past. Nowadays, probe-based real-time PCR techniques are among the most advanced tools for a reliable identification and a sensitive detection of many pathogens, but only few protocols have been published so far for CBPP diagnosis. Therefore we developed a novel TaqMan®-based real-time PCR assay comprising the amplification of two independent targets (MSC_0136 and MSC_1046) and an internal exogenous amplification control in a multiplex reaction and evaluated its diagnostic performance with clinical samples.
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Rapid identification of Burkholderia pseudomallei and Burkholderia mallei by fluorescence in situ hybridization (FISH) from culture and paraffin-embedded tissue samples.
Int. J. Med. Microbiol.
PUBLISHED: 02-15-2011
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We evaluated newly developed probes for rapid identification of Burkholderia (B.) pseudomallei and B. mallei and differentiation from B. thailandensis by fluorescence in situ hybridization (FISH). FISH correctly identified 100% of the tested B. pseudomallei (11), B. mallei (11), and B. thailandensis (1) strains, excluded 100% of all tested negative controls (61), and allowed demonstration of B. pseudomallei infection in a paraffin-embedded spleen tissue sample of an experimentally infected mouse.
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Use of a Western blot technique for the serodiagnosis of glanders.
BMC Vet. Res.
PUBLISHED: 01-19-2011
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The in vivo diagnosis of glanders relies on the highly sensitive complement fixation test (CFT). Frequently observed false positive results are troublesome for veterinary authorities and cause financial losses to animal owners. Consequently, there is an urgent need to develop a test with high specificity. Hence, a Western blot assay making use of a partly purified lipopolysaccaride (LPS) containing antigen of three Burkholderia mallei strains was developed. The test was validated investigating a comprehensive set of positive and negative sera obtained from horses and mules from endemic and non endemic areas.
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Coxiella burnetii and coinfections in Ixodes ricinus ticks in Central Germany.
Vector Borne Zoonotic Dis.
PUBLISHED: 12-13-2010
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A total of 1000 Ixodes ricinus ticks were collected in 2006 and 2007 in a forest region of Central Germany and investigated for Coxiella burnetii. The transposase element IS1111 and isocitrate dehydrogenase gene were targets of the real-time polymerase chain reaction. The pathogen was detected in 19 ticks (1.9%), and interestingly, in 10 of these samples, coinfections with Borrelia spp., spotted fever group rickettsiae, or Babesia spp. were present. Our study reports on C. burnetii infections in I. ricinus ticks in an area where cases of Q fever occur regularly and Dermacentor marginatus is not present. The broad spectrum of copathogens indicates interactions in transmission cycles and the possibility of coinfections in humans in areas where people are in close contact with infected ticks and domestic animals.
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Brucellosis - regionally emerging zoonotic disease?
Croat. Med. J.
PUBLISHED: 08-19-2010
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To gain deeper insight into the seroprevalence of brucellosis, which remains a zoonotic disease of worldwide public health concern, by reviewing studies from countries including North Africa, the Middle East, and India.
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Differential phenotyping of Brucella species using a newly developed semi-automated metabolic system.
BMC Microbiol.
PUBLISHED: 06-28-2010
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A commercial biotyping system (Taxa Profile™, Merlin Diagnostika) testing the metabolization of various substrates by bacteria was used to determine if a set of phenotypic features will allow the identification of members of the genus Brucella and their differentiation into species and biovars.
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Detection of Yersinia pestis using real-time PCR in patients with suspected bubonic plague.
Mol. Cell. Probes
PUBLISHED: 06-16-2010
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Yersinia (Y.) pestis, the causative agent of plague, is endemic in natural foci of Asia, Africa, and America. Real-time PCR assays have been described as rapid diagnostic tools, but so far none has been validated for its clinical use. In a retrospective clinical study we evaluated three real-time PCR assays in two different assay formats, 5-nuclease and hybridization probes assays. Lymph node aspirates from 149 patients from Madagascar with the clinical diagnosis of bubonic plague were investigated for the detection of Y. pestis DNA. Results of real-time PCR assays targeting the virulence plasmids pPCP1 (pla gene), and pMT1 (caf1, Ymt genes) were compared with an F1-antigen immunochromatographic test (ICT) and cultivation of the organism. Out of the 149 samples an infection with Y. pestis was confirmed by culture in 47 patients while ICT was positive in 88 including all culture proven cases. The best real-time PCR assay was the 5-nuclease assay targeting pla which was positive in 120 cases. In conclusion, the 5-nuclease assay targeting pla can be recommended as diagnostic tool for establishing a presumptive diagnosis when bubonic plague is clinically suspected.
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Comparison of commercial DNA preparation kits for the detection of Brucellae in tissue using quantitative real-time PCR.
BMC Infect. Dis.
PUBLISHED: 04-20-2010
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The detection of Brucellae in tissue specimens using PCR assays is difficult because the amount of bacteria is usually low. Therefore, optimised DNA extraction methods are critical. The aim of this study was to assess the performance of commercial kits for the extraction of Brucella DNA.
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Development and validation of a multiplex real-time PCR for detection of Clostridium chauvoei and Clostridium septicum.
Mol. Cell. Probes
PUBLISHED: 02-05-2010
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Clostridium chauvoei is the causative agent of blackleg in cattle and sheep. The clinical symptoms of this severe disease are very similar to that of malignant edema (Clostridium septicum), infections of other Clostridium species belonging to the gas edema complex, and anthrax (Bacillus anthracis). C. chauvoei and C. septicum are closely related taxa and share many phenotypic properties hampering diagnosis by using traditional microbiological methods. Thus, there is a need for a fast and reliable identification method for specific detection of both species in clinical samples. The multiplex real-time PCR assay presented here is based on the detection of the spo0A gene and enables the simultaneous identification of C. chauvoei and C. septicum. The assay design includes an amplification control DNA template for the recognition of PCR-inhibitors. Assay validation was performed using a collection of 29 C. chauvoei, 38 C. septicum strains and 26 strains of other Clostridium species. Furthermore, the real-time PCR assay was successfully tested on tissue samples from 19 clinical blackleg cases. The assay allowed the reliable detection of one picogram DNA which represents approximate 239 genome equivalents.
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Investigating an airborne tularemia outbreak, Germany.
Emerging Infect. Dis.
PUBLISHED: 02-02-2010
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In November 2005, an outbreak of tularemia occurred among 39 participants in a hare hunt in Hesse, Germany. Previously reported tularemia outbreaks in Germany dated back to the 1950s. We conducted a retrospective cohort study among participants and investigated the environment to identify risk factors for infection. Ten participants had serologic evidence of acute Francisella tularensis infection; 1 other participant died before laboratory confirmation was obtained. Presence within 5 meters of the place where disemboweled hares were rinsed with a water hose was the risk factor most strongly associated with infection (risk ratio 22.1; 95% confidence interval 13.2-154.3). Swabs taken at the game chamber and water samples were PCR negative for F. tularensis. Eleven of 14 hare parts showed low-level concentrations of F. tularensis, compatible with cross-contamination. More than half of case-patients may have acquired infection through inhalation of aerosolized droplets containing F. tularensis generated during rinsing of infected hares.
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Proteomic analysis of Brucella suis under oxygen deficiency reveals flexibility in adaptive expression of various pathways.
Proteomics
PUBLISHED: 06-16-2009
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Low oxygen tension was proposed to be one of the environmental parameters characteristic of the patho-physiological conditions of natural infections by Brucella suis. We previously showed that various respiratory pathways may be used by B. suis in response to microaerobiosis and anaerobiosis. Here, we compare the whole proteome of B. suis exposed to such low-oxygenated conditions to that obtained from bacteria grown under ambient air using 2-D DIGE. Data showed that the reduction of basal metabolism was in line with low or absence of growth of B. suis. Under both microaerobiosis and anaerobiosis, glycolysis and denitrification were favored. In addition, fatty acid oxidation and possibly citrate fermentation could also contribute to energy production sufficient for survival under anaerobiosis. When oxygen availability changed and became limiting, basic metabolic processes were still functional and variability of respiratory pathways was observed to a degree unexpected for a strictly aerobic microorganism. This highly flexible respiration probably constitutes an advantage for the survival of Brucella under the restricted oxygenation conditions encountered within host tissue.
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Spontaneous activity of auditory nerve fibers in the barn owl (Tyto alba): analyses of interspike interval distributions.
J. Neurophysiol.
PUBLISHED: 04-08-2009
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In vertebrate auditory systems, the conversion from graded receptor potentials across the hair-cell membrane into stochastic spike trains of the auditory nerve (AN) fibers is performed by ribbon synapses. The statistics underlying this process constrain auditory coding but are not precisely known. Here, we examine the distributions of interspike intervals (ISIs) from spontaneous activity of AN fibers of the barn owl (Tyto alba), a nocturnal avian predator whose auditory system is specialized for precise temporal coding. The spontaneous activity of AN fibers, with the exception of those showing preferred intervals, is commonly thought to result from excitatory events generated by a homogeneous Poisson point process, which lead to spikes unless the fiber is refractory. We show that the ISI distributions in the owl are better explained as resulting from the action of a brief refractory period ( approximately 0.5 ms) on excitatory events generated by a homogeneous stochastic process where the distribution of interevent intervals is a mixture of an exponential and a gamma distribution with shape factor 2, both with the same scaling parameter. The same model was previously shown to apply to AN fibers in the cat. However, the mean proportions of exponentially versus gamma-distributed intervals in the mixture were different for cat and owl. Furthermore, those proportions were constant across fibers in the cat, whereas they covaried with mean spontaneous rate and with characteristic frequency in the owl. We hypothesize that in birds, unlike in mammals, more than one ribbon may provide excitation to most fibers, accounting for the different proportions, and that variation in the number of ribbons may underlie the variation in the proportions.
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DNA microarray-based detection and identification of Burkholderia mallei, Burkholderia pseudomallei and Burkholderia spp.
Mol. Cell. Probes
PUBLISHED: 04-03-2009
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We developed a rapid oligonucleotide microarray assay based on genetic markers for the accurate identification and differentiation of Burkholderia (B.) mallei and Burkholderia pseudomallei, the agents of glanders and melioidosis, respectively. These two agents were clearly identified using at least 4 independent genetic markers including 16S rRNA gene, fliC, motB and also by novel species-specific target genes, identified by in silico sequence analysis. Specific hybridization signal profiles allowed the detection and differentiation of up to 10 further Burkholderia spp., including the closely related species Burkholderia thailandensis and Burkholderia-like agents, such as Burkholderia cepacia, Burkholderia cenocepacia, Burkholderia vietnamiensis, Burkholderia ambifaria, and Burkholderia gladioli, which are often associated with cystic fibrosis (CF) lung disease. The assay was developed using the easy-to-handle and economical ArrayTube (AT) platform. A representative strain panel comprising 44 B. mallei, 32 B. pseudomallei isolates, and various Burkholderia type strains were examined to validate the test. Assay specificity was determined by examination of 40 non-Burkholderia strains.
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Identification and antimicrobial susceptibilities of Ochrobactrum spp.
Int. J. Med. Microbiol.
PUBLISHED: 03-31-2009
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Ochrobactrum (O.) anthropi is an opportunistic emerging pathogen closely related to the genus Brucella. Identification and differentiation from brucellae and other Ochrobactrum spp. using routine biochemical test systems is not reliable due to the high phenotypic similarity. In this study, antibiotic susceptibilities of 103 Ochrobactrum isolates were determined using Etest for 19 clinically relevant antimicrobial agents. Ochrobactrum strains were highly resistant to beta-lactam antibiotics, susceptible to ciprofloxacin, and 97.1% were susceptible to trimethoprim/sulfamethoxazole. It was also demonstrated that biochemical reaction profiles of the API and BD Phoenix 100 systems for identifying Ochrobactrum isolates can only be used on the genus level. Our in vitro data suggest that combinations of antimicrobial agents including ciprofloxacin and/or trimethoprim/sulfamethoxazole may be useful for empirical treatment of Ochrobactrum infections.
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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.
BMC Microbiol.
PUBLISHED: 03-09-2009
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Since 1994, Brucella strains have been isolated from a wide range of marine mammals. They are currently recognized as two new Brucella species, B. pinnipedialis for the pinniped isolates and B. ceti 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 Brucella isolates and with reference to terrestrial mammal Brucella 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 Brucella strains from animal and human origin was used.
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Prevalence-dependent use of serological tests for diagnosing glanders in horses.
BMC Vet. Res.
PUBLISHED: 02-16-2009
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The internationally mandatory complement fixation test (CFT) for testing of equine sera for the absence of glanders has repeatedly led to discrepant results. Not only do "false positive" sera pose a problem for the diagnostician and the animal health authorities but they can also result in significant financial losses for the animal owners.Due to the very low prevalence of glanders in the horse population it is of major importance to use tests with a high specificity to overcome unreliable predictive values. We have compared formalin-fixed B. mallei whole cell antigen and a well characterised mouse monoclonal antibody with regard to their specificity and sensitivity for glanders serodiagnosis using CFT, an indirect (i) and a competitive (c) ELISA platform.
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Validation of ten new polymorphic tandem repeat loci and application to the MLVA typing of Burkholderia pseudomallei isolates collected in Singapore from 1988 to 2004.
J. Microbiol. Methods
PUBLISHED: 02-14-2009
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Multiple locus variable number of tandem repeat (VNTR) analysis (MLVA) has been shown to be very promising for the typing of Burkholderia pseudomallei and mallei. The currently available set of loci requires high resolution allele size measurement due to short repeat units. The present work was aimed at expanding the available set of VNTR loci, and generating data from a collection of 102 B. pseudomallei strains isolated in Singapore between 1988 and 2004 including few additional strains of various origins as references. Ten new VNTRs with a longer array size have been identified compatible with standard agarose gel separation, and a reference database of 72 genotypes was created which can be queried on the Internet.
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Determination of antimicrobial sensitivities of Campylobacter jejuni isolated from commercial turkey farms in Germany.
Avian Dis.
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The emergence of antimicrobial resistance among Campylobacter isolates recovered from turkeys has increased dramatically. Monitoring the progress of this resistance becomes a growing public health issue. The aim of the present study was to provide information of the current status of antibiotic resistance patterns in Campylobacter jejuni from turkeys. Seventy-six C. jejuni isolates were recovered from 67 epidemiologically unrelated meat turkey flocks in different regions of Germany in 2010 and 2011. The isolates were typed by flaA genotyping and were investigated for antimicrobial susceptibility against 12 antibiotics by using a broth microdilution test as well as testing the genetic determination of ciprofloxacin, tetracycline, and erythromycin resistance. All isolates (n = 76) were sensitive to gentamicin and chloramphenicol. The numbers of isolates that were sensitive to streptomycin, erythromycin, neomycin, and amoxicillin were 69 (90.8%), 61 (80.2%), 58 (76.4%), and 44 (57.9%), respectively. Only one isolate was sensitive to all tested antibiotics. The emergence of a high resistance rate and multidrug resistance to three or more classes of antimicrobial agents were observed. The resistance against sulphamethoxazole/trimethoprim, metronidazole, ciprofloxacin, naladixic acid, and tetracycline was 58 (76.3%), 58 (76.3%), 53 (69.7%), 51 (67.1%), and 42 (55.3%), respectively. None of the isolates was resistant to all antibiotics. Multidrug resistance to three or more classes of antimicrobial agents was found and ranged from 3.9% to 40.8%. Replacement of the Thr-86-->Ile in gyrA gene and detection of the tet(O) gene were the main resistance mechanisms for fluoroquinolones and tetracycline, respectively, while the lack of mutation in position 2074 and 2075 on the 23S rRNA gene was responsible for macrolide resistance. The phenotypic and genotypic resistance profiles were compatible in the case of ciprofloxacin and tetracycline but were not completely congruent with respect to erythromycin.
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A review on camel brucellosis: a zoonosis sustained by ignorance and indifference.
Pathog Glob Health
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In many developing countries of Asia and Africa, camels are one of the most important sources of income for the nomadic population. With increasing urbanization, camel milk and meat have gained a wider market and commercialization and consumption of camel products are on the rise. Camel brucellosis can be encountered in all camel rearing countries with exception of Australia. High animal and herd prevalences have been reported from numerous countries, which not only pose a continuous risk for human infection, but also increase the spread of infection through uncontrolled trade of clinically inconspicuous animals. This short review aims at providing an overview on diagnostic investigations, as well as the public health and economic impact of brucellosis in old world camels.
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Effectiveness of an antimicrobial treatment scheme in a confined glanders outbreak.
BMC Vet. Res.
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Glanders is a contagious and fatal zoonotic disease of solipeds caused by the Gram-negative bacterium Burkholderia (B.) mallei. Although regulations call for culling of diseased animals, certain situations e.g. wild life conservation, highly valuable breeding stock, could benefit from effective treatment schemes and post-exposure prophylaxis.
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Rapid identification of Burkholderia mallei and Burkholderia pseudomallei by intact cell Matrix-assisted Laser Desorption/Ionisation mass spectrometric typing.
BMC Microbiol.
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Burkholderia (B.) pseudomallei and B. mallei are genetically closely related species. B. pseudomallei causes melioidosis in humans and animals, whereas B. mallei is the causative agent of glanders in equines and rarely also in humans. Both agents have been classified by the CDC as priority category B biological agents. Rapid identification is crucial, because both agents are intrinsically resistant to many antibiotics. Matrix-assisted laser desorption/ionisation mass spectrometry (MALDI-TOF MS) has the potential of rapid and reliable identification of pathogens, but is limited by the availability of a database containing validated reference spectra. The aim of this study was to evaluate the use of MALDI-TOF MS for the rapid and reliable identification and differentiation of B. pseudomallei and B. mallei and to build up a reliable reference database for both organisms.
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Prevalence and distribution of Clostridium difficile PCR ribotypes in cats and dogs from animal shelters in Thuringia, Germany.
Anaerobe
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Clostridium difficile is an important cause of nosocomial diarrhoea in humans. Pet animals and livestock are discussed as potential natural reservoirs and sources of infection. In this study faecal samples from dogs and cats were collected at 10 animal shelters in Thuringia, Germany. C. difficile was isolated from 9 out of 165 (5.5%) canine and 5 out of 135 (3.7%) feline samples. Five PCR ribotypes (010, 014/020, 039, 045, SLO 066) were identified. PCR ribotypes 010 and 014/020 were detected in more than one shelter and PCR ribotypes 014/020 and 045 were isolated from dogs and cats. MLVA profiles of strains of a PCR ribotype from one shelter were identical or closely related, while strains of the same PCR ribotype from different shelters showed significant differences. This study shows that dogs and cats kept in animal shelters are a reservoir of C. difficile PCR ribotypes which can infect also humans.
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Q fever: single-point source outbreak with high attack rates and massive numbers of undetected infections across an entire region.
Clin. Infect. Dis.
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In early 2009, a dairy-goat annex care farm in South Limburg, the Netherlands, reported 220 Coxiella burnetii-related abortions in 450 pregnant goats. These preceded human cases and occurred in a region that was Q-fever free before 2009, providing a unique quasi-experimental setting for investigating regional transmission patterns associated with a Q-fever point source.
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Performance of complement fixation test and confirmatory immunoblot as two-cascade testing approach for serodiagnosis of glanders in an endemic region of South East Asia.
Berl. Munch. Tierarztl. Wochenschr.
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Various serological tests were used for the diagnosis of glanders in the past but still complement fixation test (CFT) is the internationally prescribed test for trading equines. A new immunoblot (IB) technique has recently been introduced to overcome the well known shortcomings of CFT i. e. a considerable number of false positive and negative results and anticomplementary effects of sera. The objective of this study was the comparative evaluation of two glanders CFT antigens commercially available at Central Veterinary Institute ofWageningen UR, Lelystad, NL (CIDC) and at c.c.pro GmbH, Oberdorla, DE (c.c.pro) in a glanders endemic area regarding specificity and sensitivity. A total of 1678 serum samples from the endemic region (Province Punjab, Pakistan) and a non-endemic area (Germany) were analysed. All sera tested positive or suspicious with CFT were analysed by the confirmatory IB to exclude CFT false positive results. Both CFT antigens showed 100% sensitivity. The use of CIDC or c.c.pro antigen resulted in specificities of 77.45% or 75.71% for sera from endemic area and 93.75% or 94.79% for sera from non-endemic areas, respectively. The results demonstrate the different performances of identical tests in different epidemiologically settings. Based on these results, the combined use of CFT and IB is highly suggestive for the serodiagnosis of glanders. Good agreement was calculated between CFT (using either c.c.pro or CIDC antigen) and immunoblot.
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Prevalence of Coxiella burnetii in clinically healthy German sheep flocks.
BMC Res Notes
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Current epidemiological data on the situation of Coxiella (C.) burnetii infections in sheep are missing, making risk assessment and the implementation of counteractive measures difficult. Using the German state of Thuringia as a model example, the estimated sero-, and antigen prevalence of C. burnetii (10% and 25%, respectively) was assessed at flock level in 39/252 randomly selected clinically healthy sheep flocks with more than 100 ewes and unknown abortion rate.
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Cross-border molecular tracing of brucellosis in Europe.
Comp. Immunol. Microbiol. Infect. Dis.
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To assess the general impact of endemic countries on the re-emergence of brucellosis in non-endemic regions of the European Union, the genetic fingerprints of Brucella melitensis strains imported to Germany were compared to ovine strains from Turkey in a molecular epidemiological study. Genotyping of 66 Brucella strains (based on Multiple Locus of Variable number of tandem repeats Analysis) isolated from German travellers and Turkish immigrants living in Germany revealed epidemiological concordance with 20 sheep isolates originating from Eastern Anatolia, Turkey. In summary, cross-border molecular tracing confirmed brucellosis being a zoonosis of concern for European public health.
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JoVE Visualize is a tool created to match the last 5 years of PubMed publications to methods in JoVE's video library.

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In developing our video relationships, we compare around 5 million PubMed articles to our library of over 4,500 methods videos. In some cases the language used in the PubMed abstracts makes matching that content to a JoVE video difficult. In other cases, there happens not to be any content in our video library that is relevant to the topic of a given abstract. In these cases, our algorithms are trying their best to display videos with relevant content, which can sometimes result in matched videos with only a slight relation.