Multi-locus variable-number tandem repeat analysis for outbreak studies of Salmonella enterica serotype Enteritidis

<p>Abstract</p> <p>Background</p> <p><it>Salmonella enterica </it>subsp. <it>enterica </it>serotype Enteritidis is known as an important and pathogenic clonal group which continues to cause worldwide sporadic cases and outbreaks in humans. Here a new multiple-locus variable-number tandem repeat analysis (MLVA) method is reported for highly-discriminative subtyping of <it>Salmonella </it>Enteritidis. Emphasis was given on the most predominant phage types PT4 and PT8. The method comprises multiplex PCR specifically amplifying repeated sequences from nine different loci followed by an automatic fragment size analysis using a multicolor capillary electrophoresis instrument. A total of 240 human, animal, food and environmental isolates of <it>S</it>. Enteritidis including 23 definite phage types were used for development and validation. Furthermore, the MLVA types were compared to the phage types of several isolates from two recent outbreaks to determine the concordance between both methods and to estimate their in vivo stability. The in vitro stability of the two MLVA types specifically for PT4 and PT8 strains were determined by multiple freeze-thaw cycles.</p> <p>Results</p> <p>Seventy-nine different MLVA types were identified in 240 <it>S</it>. Enteritidis strains. The Simpson's diversity index for the MLVA method was 0.919 and Nei diversity values for the nine VNTR loci ranged from 0.07 to 0.65. Twenty-four MLVA types could be assigned to 62 PT4 strains and 21 types to 81 PT8 strains. All outbreak isolates had an indistinguishable outbreak specific MLVA type. The in vitro stability experiments showed no changes of the MLVA type compared to the original isolate.</p> <p>Conclusion</p> <p>This MLVA method is useful to discriminate <it>S</it>. Enteritidis strains even within a single phage type. It is easy in use, fast, and cheap compared to other high-resolution molecular methods and therefore an important tool for surveillance and outbreak studies for <it>S</it>. Enteritidis.</p

Diversity of Salmonella enterica serovar Derby isolated from pig, pork and humans in Germany

Salmonella enterica serovar Derby (S. Derby) is one of the most prevalent serovars in pigs in Europe and in the U.S. and ranks among the 10 most frequently isolated serovars in humans. Therefore, a set of 82 epidemiologically unrelated S. Derby strains isolated between 2006 and 2008 from pigs, pork and humans in Germany was selected and investigated in respect to the transmission of clonal groups of the serovar along the food chain. Various phenotypic and genotypic methods were applied and the pathogenicity and resistance gene repertoire was determined. Phenotypically 72% of the strains were susceptible to all 17 antimicrobials tested while the others were monoresistant to tetracycline or multi-resistant with different resistance profiles. Four major clonal groups were identified based on PFGE, sequence data of the virulence genes sopA, sopB and sopD, VNTR-locus STTR5 and MLST revealing also the new sequence type ST774. Thirty different PFGE profiles were detected resulting in four clusters representing the four groups. The pathogenicity gene repertoire of 32 representative S. Derby strains analyzed by microarray showed six types with differences in the Salmonella pathogenicity islands, pathogenicity genes on smaller islets or prophages and fimbriae coding genes. The pathogenicity gene repertoire of the predominant types PAT DE1 and DE2 were most similar to the ones of S. Paratyphi B (dT+, O5−) and to a minor degree to S. Infantis and S. Virchow PATs. Overall this study showed that in Germany currently one major S. Derby clone is frequently isolated from pigs and humans. Contaminated pork was identified as one vehicle and consequently is a risk for human health. To prevent this serovar from entering the food chain, control measurements should be applied at the farm level

Comparative Analysis of ESBL-Positive Escherichia coli isolates from Animals and Humans from the UK, the Netherlands and Germany

The putative virulence and antimicrobial resistance gene contents of extended spectrum β-lactamase (ESBL)-positive E. coli (n=629) isolated between 2005 and 2009 from humans, animals and animal food products in Germany, The Netherlands and the UK were compared using a microarray approach to test the suitability of this approach with regard to determining their similarities. A selection of isolates (n=313) were also analysed by multilocus sequence typing (MLST). Isolates harbouring blaCTX-M-group-1 dominated (66%, n=418) and originated from both animals and cases of human infections in all three countries; 23% (n=144) of all isolates contained both blaCTX-M-group-1 and blaOXA-1-like genes, predominantly from humans (n=127) and UK cattle (n=15). The antimicrobial resistance and virulence gene profiles of this collection of isolates were highly diverse. A substantial number of human isolates (32%, n=87) did not share more than 40% similarity (based on the Jaccard coefficient) with animal isolates. A further 43% of human isolates from the three countries (n=117) were at least 40% similar to each other and to five isolates from UK cattle and one each from Dutch chicken meat and a German dog; the members of this group usually harboured genes such as mph(A), mrx, aac(6’)-Ib, catB3, blaOXA-1-like and blaCTX-M-group-1. forty-four per cent of the MLST-typed isolates in this group belonged to ST131 (n=18) and 22% to ST405 (n=9), all from humans. Among animal isolates subjected to MLST (n=258), only 1.2% (n=3) were more than 70% similar to human isolates in gene profiles and shared the same MLST clonal complex with the corresponding human isolates. The results suggest that minimising human-to-human transmission is essential to control the spread of ESBL-positive E. coli in humans

The Emergence of Emotions

Emotion is conscious experience. It is the affective aspect of consciousness. Emotion arises from sensory stimulation and is typically accompanied by physiological and behavioral changes in the body. Hence an emotion is a complex reaction pattern consisting of three components: a physiological component, a behavioral component, and an experiential (conscious) component. The reactions making up an emotion determine what the emotion will be recognized as. Three processes are involved in generating an emotion: (1) identification of the emotional significance of a sensory stimulus, (2) production of an affective state (emotion), and (3) regulation of the affective state. Two opposing systems in the brain (the reward and punishment systems) establish an affective value or valence (stimulus-reinforcement association) for sensory stimulation. This is process (1), the first step in the generation of an emotion. Development of stimulus-reinforcement associations (affective valence) serves as the basis for emotion expression (process 2), conditioned emotion learning acquisition and expression, memory consolidation, reinforcement-expectations, decision-making, coping responses, and social behavior. The amygdala is critical for the representation of stimulus-reinforcement associations (both reward and punishment-based) for these functions. Three distinct and separate architectural and functional areas of the prefrontal cortex (dorsolateral prefrontal cortex, orbitofrontal cortex, anterior cingulate cortex) are involved in the regulation of emotion (process 3). The regulation of emotion by the prefrontal cortex consists of a positive feedback interaction between the prefrontal cortex and the inferior parietal cortex resulting in the nonlinear emergence of emotion. This positive feedback and nonlinear emergence represents a type of working memory (focal attention) by which perception is reorganized and rerepresented, becoming explicit, functional, and conscious. The explicit emotion states arising may be involved in the production of voluntary new or novel intentional (adaptive) behavior, especially social behavior

Epidemiology of resistance to quinolones in Salmonella

Fluoroquinolones account for about 11% of antimicrobial prescriptions in human medicine worldwide and represent the drug of choice for the treatment of a wide range of human infectious diseases. They were introduced into veterinary medicine in Europe in the late 1980s and early 1990s and in the USA in 1995. Following their introduction, resistant strains of bacteria, including Salmonella, started to emerge. Resistance to quinolones depends on chromosomal mutations and the subsequent spread of resistant clones. While the selective pressure caused by the use of quinolones facilitates their epidemic transmission, the resistant mutants may spread independently of quinolone use. In view of the key role of this group of antimicrobials in human medicine and the position of Salmonella as the leading cause of food-borne infections in many countries, the public health hazard posed by quinolone-resistant zoonotic Salmonella serovars has been a subject of concern. The fluoroquinolones are on the WHO list of drugs that should be reserved for human use. Considering the mounting evidence that quinolone-resistant zoonotic Salmonella are the cause of severe, sometimes fatal, infections in humans, the use of fluoroquinolones in food animals should be discontinued or severely restricted. Such an intervention should be accompanied by prudent use measures involving all other groups of antimicrobials to reduce the need for fluoroquinolones in veterinary medicine.Épidémiologie de la résistance aux quinolones chez Salmonella. Les fluoroquinolones représentent environ 11 % de la totalité des prescriptions d'antibiotiques en médecine humaine et sont les antibiotiques de choix pour le traitement d'une grande variété de maladies infectieuses humaines. En médecine vétérinaire, ils ont été introduits en Europe à la fin des années 80-début des années 90, et aux USA en 1995. Après leur introduction, des souches de bactéries résistantes, dont des Salmonella, ont commencé à émerger. La résistance aux quinolones est due à des mutations chromosomiques et à la propagation ultérieure de clones résistants. Alors que la pression de sélection par utilisation de quinolones facilite leur transmission épidémique, les mutants résistants peuvent se propager indépendamment de l'utilisation de quinolones. Étant donné le rôle-clé de ce groupe d'antibiotiques en médecine humaine, et le fait que Salmonella soit la cause principale des infections alimentaires dans de nombreux pays, le problème est posé du risque pour la santé publique dû à des sérovars de Salmonella zoonotiques résistants aux quinolones. Les fluoroquinolones sont sur la liste de l'OMS des antibiotiques qui devraient être réservés à la médecine humaine. Considérant qu'il est de plus en plus évident que les Salmonella zoonotiques résistantes aux quinolones sont la cause d'infections sévères, quelquefois fatales chez l'homme, l'utilisation de fluoroquinolones chez les animaux entrant dans la chaîne alimentaire devrait être interrompue ou sévèrement restreinte. Une telle intervention devrait être accompagnée de mesures pour une utilisation prudente de tous les autres antibiotiques afin de réduire la nécessité d'utiliser des fluoroquinolones en médecine vétérinaire

Discrimination of d-Tartrate-Fermenting and -Nonfermenting Salmonella enterica subsp. enterica Isolates by Genotypic and Phenotypic Methods

A multiplex PCR and an improved lead acetate test were developed to discriminate d-tartrate-fermenting and -nonfermenting Salmonella enterica subsp. enterica strains. Both methods showed an accuracy of 100% when 125 Salmonella strains belonging to 15 serovars were tested. Special emphasis was given to S. enterica subsp. enterica serovar Paratyphi B isolates because of the clinical importance of its d-tartrate-nonfermenting variant and the recently increasing numbers of cases of human outbreaks caused by its fermenting variant (formerly Salmonella serovar Java). The lead acetate test described previously (G. A. Alfredsson, R. M. Barker, D. C. Old, and J. P. Duguid, J. Hyg. 70:651-666, 1972) was modified in the inoculation and incubation procedure. The PCR assay was based on the genotypic difference of the presence (d-tartrate-fermenting strains) or absence (d-tartrate-nonfermenting strains) of the ATG start codon for the gene STM 3356, which encodes a putative cation transporter. Sequence data revealed a nucleotide exchange from G to A within the ATG start codon of gene STM 3356 in the d-tartrate-nonfermenting strains. In order to increase the reliability of the PCR assay, a positive control based on a Salmonella genus-specific primer set for the detection of Salmonella DNA was included. The PCR-based discrimination needs only several hours compared to 6 days needed by the improved lead acetate test to obtain reliable results. Consequently, the PCR d-tartrate assay should be the method of choice for the discrimination of d-tartrate-fermenting and -nonfermenting Salmonella strains in the future

Incidence of the Recently Described Sulfonamide Resistance Gene sul3 among German Salmonella enterica Strains Isolated from Livestock and Food

The sul3 gene recently described in Escherichia coli was found in 22 of 512 (4.3%) German Salmonella isolates from different regions and sources and of different serotypes, antimicrobial resistance phenotypes, and genomic groups. This is the first report on the prevalence of sul3 among Salmonella strains, and the findings support the strong potential of this determinant to spread within bacterial populations