Disentangling outbreaks using whole-genome sequencing: concurrent multistate outbreaks of Salmonella Kottbus in Germany, 2017.

In June 2017, an outbreak of Salmonella Kottbus infection was suspected in Germany. We investigated the outbreak with whole-genome sequencing (WGS) and a case-control study. Forty-six isolates from 69 cases were subtyped. Three WGS clusters were identified: cluster 1 (n = 36), cluster 2 (n = 5) and cluster 3 (n = 3). Compared to controls, cluster 1 cases more frequently consumed raw smoked ham (odds ratio (OR) 10, 95% confidence interval (CI) 1.2-88) bought at supermarket chain X (OR 36, 95% CI 4-356; 9/10 consumed ham Y). All four cluster 2 cases interviewed had consumed quail eggs. Timely WGS was invaluable in distinguishing concurrent outbreaks of a rare Salmonella serotype.

Immaturity for gestational age of microvasculature and placental barrier in term placentas with high weight.

OBJECTIVE: Villous immaturity for gestational age is a multifactorial developmental deviation associated with unexpected placental insufficiency, fetal hypoxia and term fetal death. In our previous work we have shown that immature CD15+/CD31+/CD34+ endothelial cells were an important indicator of placental villous immaturity and chronic insufficiency. The aim of this study was to perform a comparative analysis of CD15-marked immaturity in the vessel walls between normal and pathological term placentas of clinically and structurally heterogenous groups with normal, low and high weight. STUDY DESIGN: 165 clinically normal and pathological placentas of gestational age 39-42 with normal weight (25-75 percentile), low weight (<10 percentile) and high weight (>90 percentile) were structurally and immunohistochemically analyzed. Excluded were placentas with a severe form of placental insufficiency associated with intrauterine fetal death, low APGAR-score, genetic and chromosomal diseases or placental inflammations. The distribution patterns of CD15, CD31 and CD34 were assessed separately in the macrovasculature, microvasculature and placental barrier (PB) - associated capillaries. RESULTS: All placental groups with normal weight, low weight and high weight include normal, accelerated villous maturation or villous immaturity independent of their weight. However, a significant increase of immature CD15+/CD31+/CD34+ endothelial cells was detected in microvasculature and PB -associated capillaries in high weight-placentas (63.5%/52.2%), compared to those of normal weight (13.8%/8.2%) and low weight (16.1%/17.8%). The distribution of macrovascular immature CD15+/CD31+/CD34+ endothelial cells did not show such marked differences. CONCLUSION: We have identified the immaturity of microvasculature and PB -associated capillaries with a pathological persistency of immature CD15+/CD31+/CD34+ endothelial cells and a reduction of terminally differentiated CD15-/CD31+/CD34+ endothelial cells in a structurally and clinically heterogeneous group of high weight-placentas. We assume that immaturity of placental vessels are part of prenatal adaptational processes that can be recruited in different emergency situations and may provide potential targets of therapeutic correction of placental growth and chronic insufficiency. We therefore recommend the use of CD15-based immunophenotyping as a method to identify latent unfavorable conditions of fetal development in the intrauterine life and individual risk of disease in the postnatal period.

[Female Preterm Monozygotic Twins Discordant for Fetal Megacystis due to Cloacal Dysgenesis After Conception by Intracytoplasmatic Sperm Injection - Case Report and Review of Literature]. / Monozygote, weibliche Zwillingsfrühgeborene diskordant für das seltene, schwere Fehlbildungssyndrom einer fetalen Megazystis mit Kloakenfehlbildung nach künstlicher Befruchtung ­ Fallbericht und Literaturübersicht.

Monozygotic twins were previously regarded as "identical". By now an increasing number of case reports of monozygotic but discordant twins have been reported, and therefore discordance between monozygotic twins is being investigated intensively. We report a case of female preterm monozygotic twins who were discordant for fetal megacystis due to cloacal dysgenesis. Pregnancy was achieved after intracytoplasmatic sperm injection and transfer of 2 embryos. By the first trimester fetal megacystis with consecutive oligohydramnios and hypoplasia of the lungs was diagnosed. Both foetuses had normal karyotypes. After delivery at 25+3 weeks of gestation due to premature labour, the affected child was treated palliatively and died within 2 hours. In the postmortem physical examination, a cloacal dysgenesis was detected. In the male foetus, megacystis is typically caused by obstructive uropathy. In the rarely affected female foetus, it usually results from complex urogenital malformations like cloacal dysgenesis which originates from disruption during gastrulation. We identified 10 case series of mono- or dizygotic twins who were discordant either for fetal megacystis or for cloacal dysgenesis. Issues like conception, zygosity, sex, karyotype and aetiology of fetal megacystis were not reported in all cases. Discordance between monozygotic twins for structural birth defects is closely linked to the twinning process itself. Assisted reproduction is said to generate a higher rate of monozygotic twin pregnancies and to be responsible for a higher prevalence of chromosomal aberrations or congenital malformations. With regard to conception, zygosity, sex, karyotype and combination of malformations, our case is unique.

Epidemiological analysis of the dynamic and diversity of Salmonella spp. in five German pig production clusters using pheno- and genotyping methods: an exploratory study.

An exploratory study in five conventional pig production clusters was carried out to investigate the dynamic and diversity of Salmonella spp. within different production stages and sample site categories (pooled feces, direct and non-direct environment). Observing two production cycles per production cluster, a total of 1276 samples were collected along the pig production chain. Following a microbiological examination via culture, 2246 subcultures were generated out of 285 Salmonella positive samples and analysed by pheno- and genotyping methods. Based on a combination of serotyping, MLVA (multiple-locus variable-number tandem repeat (VNTR) analysis), PFGE (pulse-field gel electrophoresis) and MLST (multilocus sequence typing), an amount of 22.3% Salmonella positive samples were characterized in clonal lineages and its variants. Within each production cluster, one main clonal lineage could be identified and persisted over both production cycles with a large diversity of variants and a wide distribution in sample site categories and production stages. Results underline the importance of biosecurity with emphasis on the environment to prevent persistence and circulation of Salmonella within herds. Furthermore, the combined implementation of MLVA, PFGE and MLST with conventional culture techniques for isolate classification could be successfully applied as an effective and valuable tool for identifying similar pattern of Salmonella occurrence within pig production clusters.

Difficulties in diagnosing terminal ileitis due to Yersinia pseudotuberculosis.

We report three patients with terminal ileitis and positive fecal cultures with Yersinia pseudotuberculosis. From one patient, a virulence plasmid (pYV)-negative Y. pseudotuberculosis was isolated, which represents the second finding of a pYV-negative isolate associated with human disease. All patients were treated with ciprofloxacin and fully recovered. Since conventional culture methods for yersiniosis are gradually replaced with molecular tests not recognizing Y. pseudotuberculosis, we recommend to include a specific culture medium or to apply a specific polymerase chain reaction (PCR) assay on fecal samples from patients suspected of terminal ileitis.

Evidence for the transmission of Salmonella from reptiles to children in Germany, July 2010 to October 2011.

This study examines the Salmonella status in reptiles kept in households with children suffering from gastroenteritis due to an exotic Salmonella serovar, to obtain information on possible transmission paths. A number of affected households (n=79) were contacted, and almost half (34/79) comprised at least one reptile in the home. Of the households, 19 were further studied, whereby a total of 36 reptiles were investigated. Samples were taken from the reptiles including the oral cavity, the cloaca, the skin and, in the case of lizards, the stomach, and isolation of Salmonella strains was performed using repeated enrichment and typing. Where the Salmonella serovars of the infected child and the reptile were identical, typing was followed by pulsed-field gel electrophoresis (PFGE). Bearded dragons (Pogona vitticeps) constituted 19 of 36 examined reptiles. Altogether 319 Salmonella isolates were investigated and 24 different serovars identified in the reptiles. In 15 of 19 households, an identical serovar to the human case was confirmed in at least one reptile (including 16 of all 19 bearded dragons examined). The results demonstrate that reptiles and especially bearded dragons shed various Salmonella serovars including those isolated from infected children in the respective households. Hygiene protocols and parents' education are therefore highly necessary to reduce the risk of transmission. From a terminological point of view, we propose to call such infections 'Reptile-Exotic-Pet-Associated-Salmonellosis' (REPAS).

Campylobacter spp., Yersinia enterocolitica, and Salmonella enterica and their simultaneous occurrence in German fattening pig herds and their environment.

Campylobacter spp., Salmonella enterica, and Yersinia enterocolitica are common causes of foodborne infections in humans with pork as a potential source. Monitoring programs at farm level are, to date, only implemented for S. enterica, while epidemiological knowledge of the other two pathogens is still lacking. This study aimed to assess the pathogen load (in the pigs' environment) in fattening pig herds, their simultaneous occurrence, and the occurrence of Campylobacter spp. and Y. enterocolitica in herds in different Salmonella risk categories. In 50 fattening pig herds in northern Germany, four pooled fecal samples and 10 swab samples from the pigs' direct environment (pen walls, nipple drinkers), indirect environment (hallways, drive boards), and flies and rodent droppings were collected from each herd and submitted for cultural examination. Campylobacter spp. were detected in 38.1% of fecal, 32.7% of direct environment, 5.3% of indirect environment, and 4.6% of flies/pests samples collected, and Y. enterocolitica in 17.1, 8.1, 1.2, and 3.1% and S. enterica in 11.2, 7.7, 4.1, and 1.5%, respectively. For Campylobacter spp., Y. enterocolitica, and S. enterica, 80, 48, and 32% of herds were positive, respectively; 22 herds were positive for both Campylobacter spp. and Y. enterocolitica, 12 for Campylobacter spp. and S. enterica, and 7 for Y. enterocolitica and S. enterica. There was no significant association between the pathogens at herd level. Campylobacter spp. and Y. enterocolitica were found more often in samples from the low Salmonella risk category (odds ratio, 0.51; confidence interval, 0.36 to 0.73, and 0.3, 0.17 to 0.57), and this was also the case for Y. enterocolitica at herd level (odds ratio, 0.08; confidence interval, 0.02 to 0.3). This study provides evidence that the pigs' environment should be accounted for when implementing control measures on farms against Campylobacter spp. and Y. enterocolitica. An extrapolation from the current Salmonella monitoring to the other two pathogens does not seem feasible.

Unusual increase in reported cases of paratyphoid A fever among travellers returning from Cambodia, January to September 2013.

From January to September 2013, a marked increase in notifications of Salmonella Paratyphi A infections among travellers returning from Cambodia occurred in France. An investigation revealed 35 cases without a common source: 21 in France, five in Germany, three in the Netherlands, one in Norway, one in the United Kingdom, four in New-Zealand. Data suggest an ongoing event that should trigger further investigation. Travellers to Cambodia should observe preventive measures including good personal hygiene and food handling practices.

Subtype specific risk factor analyses for sporadic human salmonellosis: a case-case comparison in Lower Saxony, Germany.

With the intention to deepen the knowledge of the vertical transmission of particular subtypes of Salmonella enterica from "the stable to the table" a case1-case2 analysis in Lower Saxony, Germany, was conducted. The data collection was based on standardised telephone interviews with 1741 Salmonella case persons. Single-factor-analyses revealed statistically significant associations between S. Typhimurium infections and animal keeping (odds ratio (OR): 1.4; 95%-Confidence-interval (CI): 1.2-1.7), especially rodents (OR 1.5; CI 1.2-2.1), and with consumption of meat (OR 1.9; CI 1.3-2.8), raw ground pork (OR 3.0; CI 2.1-4.2) and uncooked pork sausage (OR 2.1; CI 1.6-2.9). The S. Typhimurium phage type DT 104 was associated most with consumption of uncooked pork sausage (OR 3.6; CI 1.3-8.5). Multiple logistic regression analyses confirmed the associations between S. Typhimurium infection and consumption of raw ground pork and with animal contact. The results circumstantiate the assumption of raw pork products still being a relevant source for S. Typhimurium infections in Germany. Therefore, it is recommended to intensify efforts to reduce salmonella infections caused by raw pork products. S. Enteritidis infection was associated statistically significantly with travelling abroad (OR 2.1; CI 1.6-3.3), consumption of raw tomatoes (OR 1.8; CI 1.5-2.1), dried herbs (OR 2.1; CI 1.0-1.8), and undercooked eggs (OR 1.3; CI 1.1-1.6) compared with other serovars. These results were confirmed in multiple logistic regression analyses, as well.

Desperately seeking diarrhoea: outbreak of haemolytic uraemic syndrome caused by emerging sorbitol-fermenting shiga toxin-producing Escherichia coli O157:H-, Germany, 2009.

Infection with sorbitol-fermenting Shiga toxin-producing Escherichia coli O157:H- (sf STEC O157:H-) is rare, but emerging in Europe. The pathogen is typically isolated from paediatric patients with life-threatening haemolytic uraemic syndrome (HUS). It is unclear whether this observation primarily reflects the pathogen's virulence or its complex laboratory diagnosis, not routinely conducted in diarrhoeal patients. In summer 2009, four boys living in the same suburb in Germany developed diarrhoea-associated HUS: three were infected by sf STEC O157:H- and one died. We conducted two analytical epidemiological studies, an extensive search for diarrhoeal cases in potentially exposed groups, and an environmental investigation. Outbreak cases were residents of the suburb diagnosed with HUS, sf STEC O157:H- infection, or both between 24 July 2009 and 25 August 2009. Overall, we ascertained eight cases with a median age of 4 years (range: from 8 months to 9 years). Stool screening of 220 persons led to the identification of only four additional cases: two asymptomatic carriers and two diarrhoeal cases. HUS was strongly associated with visiting a local playground in July, particularly on 16th July (odds ratio = 42.7, P = 0.002). No other commonality, including food, was identified, and all environmental samples (n = 24) were negative. In this localized non-foodborne outbreak, the place of likely infection was a local playground. Sf STEC O157:H- infection apparently limits itself rarely to diarrhoeal illness and progresses frequently to HUS. Therefore, detection of and response to this hypervirulent pathogen primarily relies on HUS surveillance.

Large and ongoing outbreak of haemolytic uraemic syndrome, Germany, May 2011.

Since early May 2011, an increased incidence of haemolytic uraemic syndrome (HUS) and bloody diarrhoea related to infections with Shiga toxin-producing Escherichia coli (STEC) has been observed in Germany, with most cases in the north of the country. Cases reported from other European countries had travelled to this area. First results of a case­control study conducted in Hamburg suggest an association between the occurrence of disease and the consumption of raw tomatoes, cucumber and leaf salad.

Molecular analysis of virulence profiles and Shiga toxin genes in food-borne Shiga toxin-producing Escherichia coli.

In this study, 75 Shiga toxin (Stx)-producing Escherichia coli (STEC) strains originating from foods (n = 73) and drinking water (n = 2) were analyzed for their stx genotype, as well as for further chromosome-, phage-, and plasmid-encoded virulence factors. A broad spectrum of stx genes was detected. Fifty-three strains (70.7%) contained stx(2) or stx(2) variants, including stx(2d), mucus-activatable stx(2d), stx(2e), and stx(2g). Seven strains (9.3%) harbored stx(1) or stx(1c), and 15 strains (20.0%) carried both stx(2) and/or stx(2) variants and stx(1) or stx(1c). Beside stx, the most abundant accessory virulence markers in STEC food isolates were iha (57.3%), ehxA (40.0%), espP (28.0%), and subAB (25.3%). Only four strains were eae positive; three of these belonged to the serogroups O26, O103, and O157 and contained a typical enterohemorrhagic E. coli virulence spectrum. The results of this study show that a number of STEC strains that occur in foods appear to be pathogenic for humans, based on their virulence profiles. Analysis of stx subtypes and detection of additional virulence factors in eae-negative strains may help to better assess the risk of such strains for causing human infection.

[Reptile-associated Salmonellosis in Infants in Germany]. / Reptilien-assoziierte Salmonelleninfektionen bei Säuglingen und Kleinkindern in Deutschland.

Salmonellosis is an important disease in childhood. Increasing infections caused by meat and raw egg consumption starts in the second year of life. The transmission from reptiles to infants is an alternative route. Reptile-associated salmonellosis is becoming more significant for pediatricians and microbiologists due to the increasing numbers of household exotic pets. Investigations of routes of infections in connection with the local health authorities including microbiological, serological and molecular biological tests is possible in case of atypical salmonellosis. During recent years (2006-2008) an increasing number of salmonellosis cases caused by reptiles. Most of the 26 patients were infants of less than one year of age. Also Salmonella strains of subspecies II, IIIa, IIIb and IV play an important role. The US Centers for Disease Control and Prevention (CDC) has published recommendations which includes washing hands with soap and water after handling reptiles or the cages and keeping reptiles out of food preparation areas. The CDC has also adviced that children less than five years should not have reptiles as pets.

[Reptile-associated Salmonellosis in a Breastfed Infant]. / Reptilien-assoziierte Salmonellenenteritis bei einem voll gestillten Säugling.

BACKGROUND: Although salmonellosis is one of the most common childhood illnesses associated with enteric bacteria, breastfed babies do not often get infected with Salmonella. In such cases it is therefore necessary to look carefully for the source of infection. PATIENT: It is reported an 8-week-old female infant with acute haemorrhagic diarrhoea and fever. With symptomatic therapy and breast-feeding her condition improved without antibiotic therapy. Microbiological diagnosis and subtyping identified infection with Salmonella enterica serovar Pomona. The source of the infection was a bearded dragon ( Pogona SPP.) living in a neighborhood household. Monitoring of the child showed shedding of the bacterium over a nine month period. CONCLUSION: In all cases of salmonellosis in breastfed babies, the source of infection should be carefully determined. Small numbers of bacteria can cause a symptomatic infection in this age group. Therefore an intensive search of the transmission route is indicated to develop prevention strategies.

Sorbitol-fermenting enterohaemorrhagic Escherichia coli O157:H- causes another outbreak of haemolytic uraemic syndrome in children.

An outbreak of haemolytic uraemic syndrome (HUS) among children caused by infection with sorbitol-fermenting enterohaemorrhagic Escherichia coli O157:H- (SF EHEC O157:H-) occurred in Germany in 2002. This pathogen has caused several outbreaks so far, yet its reservoir and routes of transmission remain unknown. SF EHEC O157:H- is easily missed as most laboratory protocols target the more common sorbitol non-fermenting strains. We performed active case-finding, extensive exploratory interviews and a case-control study. Clinical and environmental samples were screened for SF EHEC O157:H- and the isolates were subtyped by pulsed-field gel electrophoresis. We identified 38 case-patients in 11 federal states. Four case-patients died during the acute phase (case-fatality ratio 11%). The case-control study could not identify a single vehicle or source. Further studies are necessary to identify the pathogen's reservoir(s). Stool samples of patients with HUS should be tested with an adequate microbiological set-up to quickly identify SF EHEC O157:H-.

Enterobacterial autoinducer of growth enhances shiga toxin production by enterohemorrhagic Escherichia coli.

The addition of the enterobacterial autoinducer of growth to nutrient-poor minimal medium markedly accelerated the exponential growth rates of strains of enterohemorrhagic Escherichia coli but had little or no effect on maximal cell densities in stationary phase. Growth in the presence of the autoinducer resulted in an approximately twofold enhancement in Shiga toxin production.

[Risk factors for abundance of shiga toxin producing Escherichia coli in sewage water]. / Risikofaktoren der Belastung durch Shigatoxin-produzierende Escherichia coli (STEC) in Kläranlagen.

Shiga toxin producing Escherichia coli (STEC) in sewage influent into surface water are a potential source of human infections with STEC. Eight sewage treatment plants in Bavaria, Germany, were sampled at regular intervals from 2003 to 2004 in order to estimate STEC load and quantify risk factors. 95 of 378 samples (25 %) were tested positive for stx1and/or stx2 with PCR after enrichment culture. STEC elimination after treatment was 44 %. The following risk factors were analysed with logistic regression: location of sewage plant (rural vs. urban), treatment plant technology (two stage vs. three stage treatment) and sampling location (sewage input vs. sewage output). Rural plants had odds-ratios of 1,7 (95 % CI 1.03 - 2.69; p = 0.038) for a positive stx1 and/ or stx2 PCR result, sampling at sewage input of 2.1 (95 %CI 1.28 - 3.36; p = 0.003) and three stage plants of 1.51 (95 % CI 0.94 - 2.44; p = 0.087, not significant). Sampling after rain and after dry spells had no impact on STEC abundance (univariate Chi-square test = 0.01; df1; p = 0.920). Rural sewage plants had higher odds of STEC content. The influence of the sewage plant technology on the STEC load requires further clarification.