In April 2015, Finnish public health authorities alerted European Union member states of a possible multi-country Salmonella enteritidis outbreak linked to an international youth ice-hockey tournament in Latvia. The European Centre for Disease Prevention and Control (ECDC), Finnish and Latvian authorities initiated an outbreak investigation to identify the source. The investigation included a description of the outbreak, retrospective cohort study, microbiological investigation and trace-back. We identified 154 suspected and 96 confirmed cases from seven countries. Consuming Bolognese sauce and salad at a specific event arena significantly increased the risk of illness. Isolates from Finnish, Swedish and Norwegian cases had an identical multiple-locus variable-number of tandem repeats analysis-profile (3-10-6-4-1). Breaches in hygiene and food storing practices in the specific arena's kitchen allowing for cross-contamination were identified. Riga Cup participants were recommended to follow good hand hygiene and consume only freshly cooked foods. This investigation demonstrated that the use of ECDC's Epidemic Intelligence Information System for Food- and Waterborne Diseases and Zoonoses platform was essential to progress the investigation by facilitating information exchange between countries. Cross-border data sharing to perform whole genome sequencing gave relevant information regarding the source of the outbreak.

Multi-locus variable number tandem repeat analysis (MLVA) is used in clinical and reference laboratories for subtyping verocytotoxin-producing Escherichia coli O157 (VTEC O157). However, as yet there is no common allelic or profile nomenclature to enable laboratories to easily compare data. In this study, we carried out an inter-laboratory comparison of an eight-loci MLVA scheme using a set of 67 isolates of VTEC O157. We found all but two isolates were identical in profile in the two laboratories, and repeat units were homogeneous in size but some were incomplete. A subset of the isolates (n = 17) were sequenced to determine the actual copy number of representative alleles, thereby enabling alleles to be named according to international consensus guidelines. This work has enabled us to realize the potential of MLVA as a portable, highly discriminatory and convenient subtyping method.

In a first study from India, a diverse collection of 140 environmental and clinical non-O157 Shiga-toxigenic Escherichia coli strains from a large geographical area in north India was typed by multi-locus variable number tandem repeat analysis (MLVA). The distribution of major virulence genes stx1, stx2 and eae was found to be 78%, 70% and 10%, respectively; 15 isolates were enterohaemorrhagic E. coli (stx1 +/stx2 + and eae +). By MLVA analysis, 44 different alleles were obtained. Dendrogram analysis revealed 104 different genotypes and 19 MLVA-type complexes divided into two main lineages, i.e. mutton and animal stool. Human isolates presented a statistically significant greater odds ratio for clustering with mutton samples compared to animal stool isolates. Five human isolates clustered with animal stool strains suggesting that some of the human infections may be from cattle, perhaps through milk, contact or the environment. Further epidemiological studies are required to explore these sources in context with occurrence of human cases.

The aim of the study was to investigate how the use of fresh cheese brines compared with used brines and various combinations of pH and NaCl concentrations affected the survival of Listeria monocytogenes. Cheese brines from five Norwegian small scale cheese producers were analysed and showed great variations in pH (4·54–6·01) and NaCl concentrations (14·1–26·9 %). The survival of five strains of List. monocytogenes (two clinical isolates, two food isolates and one animal isolate) in four different cheese brines (three used and one fresh) was investigated. Results showed significant differences in survival both depending on the strains and the brines. Strains of human outbreak listeriosis cases showed greater ability to survive in the brines compared with food isolates and a List. monocytogenes reference strain (1–2 log10 difference after 200 d). All strains showed highest survival in the freshly prepared brine compared with the used brines. Molecular typing by multiple locus variable number tandem repeats analysis (MLVA) showed that there were no detectable alterations in the examined variable number tandem repeats of the genome in five strains after 200 d storage in any of the salt brines. Combined effects of pH (4·5, 5·25 and 6·0) and NaCl (15, 20 and 25 %) in fresh, filter sterilised brines on the survival of List. monocytogenes were examined and results showed that pathogen populations decreased over time in all brines. Death rates at any given NaCl concentration were highest at low pH (4·5) and death rates at any given pH were highest at low NaCl concentrations (15 %). In conclusion, the use of used brines reduced the survival of List. monocytogenes and a combination of low pH (4·5) and low salt concentrations (15 %) decreased the risk of List. monocytogenes survival compared with higher pH (5·25 or 6·0) and higher NaCl concentrations (20 or 25 %).

The rickettsial bacterium Ehrlichia ruminantium is the causative agent of heartwater, a serious tick-borne disease in ruminants. The genetic diversity of organisms in the field will have implications for cross-protective capacities of any vaccine developed, and for an effective vaccine design strategy proper genotyping and understanding of existing genetic diversity in the field is necessary. We searched for variable-number tandem-repeat (VNTR) loci for use in a multi-locus VNTR analysis (MLVA). Sequencing analysis of 30 potential VNTRs using a panel of 17 reference strains from geographically diverse origins identified 12 VNTRs with allelic profiles differing between strains. Application of MLVA to 38 E. ruminantium-infected Amblyomma variegatum collected from indigenous cattle in 6 different districts of Uganda identified 21 MLVA types. The discriminatory power of MLVA was greater than that of map1 PCR-restriction fragment length polymorphism analysis, with which only 6 genotypes were obtained. The high discriminatory power as well as cost- effective performance of MLVA provide the potential for this technique to be applied in the future with respect to optimizing vaccine trials by identifying local strain diversity, and also raise the possibility of exploring the association between E. ruminantium genotypes and phenotypes such as pathological outcome in the ruminant host.

During a recent large tularemia outbreak in Bulgaria we found several cases that were remote from the main focus. One case had an unusual mode of transmission. A hunter acquired tularemia through a nail scratch from a buzzard (Buteo buteo) and consequently developed a typical ulceroglandular form of the disease. The diagnosis was confirmed by serological methods and successful cultivation. Comparative strain typing was performed by high-resolution multi-locus variable-number tandem repeat analysis (MLVA). The isolated strain was identical to one of the outbreak genotypes. We consider that this case represents a bird-to-human transmission of F. tularensis.

Coagulase gene restriction fragment length polymorphism (RFLP), six-locus variable-number tandem-repeat analysis patterns (MLVA) and detection of enterotoxin genes (se) (sea, sec, sed, seg, seh, sei, sej and sel) were used to determine the phylogenetic relationship among isolates of Staphylococcus aureus isolated from dairy products from different regions of Italy. A total of 25 Staph. aureus were subtyped into 16 coagulase genotypes by RFLP, and MLVA revealed marked genomic variability. Furthermore, 17 of the isolates harboured at least one toxin gene, with the predominance of sea, sed and sej among cow isolates and sec-sel among the goat and sheep strains. Combined RFLP, MLVA polymorphism and se genes were found to be useful techniques for discriminating several genetic variants in Staph. aureus isolates.