Epidemiological investigation of Pseudomonas aeruginosa isolates including multidrug-resistant serogroup O12 isolates, by use of a rapid and simplified multiple-locus variable-number of tandem repeats analysis and whole genome sequencing.

BACKGROUND: Clustered cases of Pseudomonas aeruginosa infection in immunocompromised patients' wards require rapid characterization of a potential epidemic to guide investigations and identify the potential source of contamination. AIM: To design and evaluate a rapid and simple typing method for P. aeruginosa in comparison to whole genome sequencing (WGS). METHODS: A simplified polymerase chain reaction based on multiple-locus variable-number of tandem repeats analysis (MLVA) was designed and used to investigate cases of P. aeruginosa infection and colonization in a paediatric haematology department. The method was compared to WGS by using the Illumina method. FINDINGS: On the 17 isolates recovered from 15 children (eight from blood cultures, three from urinary tract infections, one from sputum and five stool isolates), MLVA distinguished 10 different profiles, and seven isolates from six children shared the same profile. Analysis by WGS revealed that these seven isolates belonged to sequence type ST111 and serotype O12, allowing at least three different genotypes to be distinguished among them. Five environmental strains had three MLVA profiles; one was shared with a clinical isolate but WGS excluded any relationship. CONCLUSION: The simplified and inexpensive MLVA method enabled the exclusion, in less than 5 h, of most of the unrelated isolates and thus to focus investigations on a small number of cases, whereas WGS, taking several days of work, drew definitive conclusions concerning the outbreak and the genetic relationships of the ST111 isolates circulating in the department. We conclude that sequential use of both methods is the optimal strategy to investigate clustered cases of P. aeruginosa infections.

Outbreak of hemolytic uremic syndrome with unusually severe clinical presentation caused by Shiga toxin-producing Escherichia coli O26:H11 in France.

BACKGROUND: In spring 2019, an outbreak of Shiga toxin-producing Escherichia coli-associated hemolytic uremic syndrome (STEC HUS) occurred in France. Epidemiological investigations made by Santé publique France in connection with microbiological investigations at the national reference center for STEC promptly identified a common exposure to consumption of raw cow's milk cheese, and confirmed a cluster affiliation of the E. coli O26:H11 outbreak strain. Here, we report the clinical characteristics of the patients, the treatment used, as well as the outcome at 1 month. METHOD: Patients with STEC HUS linked to the E. coli O26:H11 outbreak strain were identified from the national surveillance network of pediatric STEC HUS cases coordinated by Santé publique France. Clinical data were analyzed from the patients' hospital records obtained from the treating physicians. RESULTS: Overall, 20 pediatric cases of STEC HUS linked to the outbreak strain were identified. Their median age of the patients was 16 months (range: 5-60). Most of them presented with diarrhea but none had received prior antibiotherapy. A total of 13 patients required dialysis; 10 patients and four patients had central nervous system (CNS) and cardiac involvement, respectively. No deaths occurred. At the 1-month follow-up, only two patients had a decreased glomerular filtration rate, below 80 mL /min/1.73m2 and four had hypertension. One patient had neurological sequelae. CONCLUSION: The E. coli O26:H11 strain identified as the cause of an STEC HUS outbreak in France in spring 2019 is notable for the initial severe clinical presentation of the patients, with a particularly high frequency of CNS and cardiac involvement similar to the German E. coli O104:H4 outbreak described in 2011. However, despite the initial severity, the 1-month outcome was favorable in most cases. The patients' young age in this outbreak highlights the need to improve information and caregiver awareness regarding consumption of at-risk foods by young children as key preventive measures against STEC infections.

Hemolytic uremic syndrome due to Shiga toxin-producing Escherichia coli infection.

The leading cause of hemolytic uremic syndrome (HUS) in children is Shiga toxin-producing Escherichia coli (STEC) infection, which has a major outbreak potential. Since the early 2010s, STEC epidemiology is characterized by a decline of the historically predominant O157 serogroup and the emergence of non-O157 STEC, especially O26 and O80 in France. STEC contamination occurs through the ingestion of contaminated food or water, person-to-person transmission, or contact with ruminants or their contaminated environment. The main symptom is diarrhea, which is bloody in about 60% of patients and occurs after a median incubation period of three days. Shiga toxins released by STEC induce a cascade of thrombogenic and inflammatory changes of microvascular endothelial cells. HUS is observed in 5-15% of STEC infection cases, defined by the triad of mechanical hemolytic anemia, thrombocytopenia, and acute renal injury. The diagnosis of STEC infection relies on biological screening for Shiga toxins and STEC in stools and serology. Treatment of STEC-HUS is mainly symptomatic, as no specific drug has proved effective. The effect of antibiotics in STEC infection and STEC-HUS remains debated; however, some bacteriostatic antibiotics might have a beneficial effect. Proofs of evidence of a benefit from complement blockade therapy in STEC-HUS are also lacking. Clinical and bacteriological STEC-HUS surveillance needs to be continued. Ongoing prospective studies will document the role of bacteriostatic antibiotics in STEC infection and STEC-HUS, and of complement blockade therapy in STEC-HUS.

[Nosocomial rotavirus gastroenteritis]. / Gastro-entérites nosocomiales à rotavirus : étude rétrospective dans un service de pédiatrie générale.

Rotavirus is the most common cause of gastroenteritis in children requiring hospitalization. It is a very resistant and contagious virus causing nosocomial gastroenteritis. In France, the vaccine against rotavirus has been available since 2006, but the vaccine is not recommended for infant vaccination. The aim of this retrospective study was to describe nosocomial rotavirus gastroenteritis (NRGE) and to assess its impact on children hospitalized in the General Pediatrics Department of Robert-Debré Hospital (Paris) between 1 January 2009 and 31 December 2013. We analyzed the demographic characteristics of children (age, term birth, underlying diseases) and the severity of the NRGE (oral or intravenous hydration), and assessed whether these children could benefit from vaccination against rotavirus. RESULTS: One hundred thirty-six children presented nosocomial rotavirus infection, with an incidence of 2.5 NRGE per 1000 days of hospitalization. The incidence of NRGE was stable between 2009 and 2013 despite the introduction of specific hygiene measures. The average age of the children was 7 months (range: 0.5-111 months). Most often NRGE occurred in children hospitalized for respiratory diseases (65% of cases) and requiring prolonged hospitalization (median: 18 days). One-third of children were born premature (25%). Hydration was oral in 80 patients (59%), by intravenous infusion in 18 patients (13%), and intraosseous in one patient. Half of the patients were aged less than 5 months and could benefit from the protection afforded by vaccination. CONCLUSION: NRGE are common. Rotavirus mass vaccination should have a positive impact on the incidence of NRGE by reducing the number of children hospitalized for gastroenteritis, therefore indirectly reducing the number of hospital cross-infections of hospitalized children who are too young to be vaccinated.

Shiga toxin-producing Escherichia coli strains isolated from dairy products - Genetic diversity and virulence gene profiles.

Shiga toxin-producing Escherichia coli (STEC) are widely recognized as pathogens causing food borne disease. Here we evaluate the genetic diversity of 197 strains, mainly STEC, from serotypes O157:H7, O26:H11, O103:H2, O111:H8 and O145:28 and compared strains recovered in dairy products against strains from human, meat and environment cases. For this purpose, we characterized a set of reference-collection STEC isolates from dairy products by PFGE DNA fingerprinting and a subset of these by virulence-gene profiling. PFGE profiles of restricted STEC total DNA showed high genomic variability (0.9976 on Simpson's discriminatory index), enabling all dairy isolates to be differentiated. High-throughput real-time PCR screening of STEC virulence genes were applied on the O157:H7 and O26:H11 STEC isolates from dairy products and human cases. The virulence gene profiles of dairy and human STEC strains were similar. Nevertheless, frequency-wise, stx1 was more prevalent among dairy O26:H11 isolates than in human cases ones (87% vs. 44%) while stx2 was more prevalent among O26:H11 human isolates (23% vs. 81%). For O157:H7 isolates, stx1 (0% vs. 39%), nleF (40% vs 94%) and Z6065 (40% vs 100%) were more prevalent among human than dairy strains. Our data point to differences between human and dairy strains but these differences were not sufficient to associate PFGE and virulence gene profiles to a putative lower pathogenicity of dairy strains based on their lower incidence in disease. Further comparison of whole-genome expression and virulence gene profiles should be investigated in cheese and intestinal tract samples.

Haemolytic-uraemic syndrome with bacteraemia caused by a new hybrid Escherichia coli pathotype.

We describe a new atypical Shiga-toxin-producing Escherichia coli (STEC) responsible for a severe episode of haemolytic-uraemic syndrome in an adult with a relapse associated with bacteraemia. This STECs train of serotype O80:H2 harboured stx2c and stx2d gene subtypes, the rare eae ξ variant and a ColV plasmid with a conserved virulence plasmidic region involved in virulence of human and avian extraintestinal pathogenic E. coli. This atypical hybrid pathotype, which represents a new threat, is a further demonstration that STEC may be a recipient for extraintestinal virulence factors and raises again the question of antibiotic therapy during STEC infection.

Foodborne transmission of sorbitol-fermenting Escherichia coli O157:[H7] via ground beef: an outbreak in northern France, 2011.

Sorbitol-fermenting Escherichia coli O157:[H7] is a particularly virulent clone of E. coli O157:H7 associated with a higher incidence of haemolytic uraemic syndrome and a higher case fatality rate. Many fundamental aspects of its epidemiology remain to be elucidated, including its reservoir and transmission routes and vehicles. We describe an outbreak of sorbitol-fermenting E. coli O157:[H7] that occurred in France in 2011. Eighteen cases of paediatric haemolytic uraemic syndrome with symptom onset between 6 June and 15 July 2011 were identified among children aged 6 months to 10 years residing in northern France. A strain of sorbitol-fermenting E. coli O157:[H7] stx2a eae was isolated from ten cases. Epidemiological, microbiological and trace-back investigations identified multiply-contaminated frozen ground beef products bought in a supermarket chain as the outbreak vehicle. Strains with three distinct pulsotypes that were isolated from patients, ground beef preparations recovered from patients' freezers and from stored production samples taken at the production plant were indistinguishable upon molecular comparison. This investigation documents microbiologically confirmed foodborne transmission of sorbitol-fermenting of E. coli O157 via beef and could additionally provide evidence of a reservoir in cattle for this pathogen.

Shopper cards data and storage practices for the investigation of an outbreak of Shiga-toxin producing Escherichia coli O157 infections.

INTRODUCTION: An outbreak of shiga-toxin producing Escherichia coli infections occurred in southwest France in June 2012. The outbreak was investigated to identify the source of infection, and guide control measures. METHODS: Confirmed outbreak cases were patients who developed bloody diarrhoea or haemolytic uremic syndrome (HUS) between 28 May and 6 July 2012, with E. coli O157 isolates showing indistinguishable patterns on pulse field gel electrophoresis (PFGE). A standardized questionnaire was administered to patients to document food consumption and other risk exposures. Their purchase was checked through their supermarket shopper card data. RESULTS: Six patients (four with HUS and two with bloody diarrhea) were confirmed outbreak cases. Fresh ground beef burgers from one supermarket were the only common food exposure, identified by interviews and shopper card data. The PFGE profile of shiga toxin-producing E. coli O157 isolated from the suspected beef burgers was identical to those from the human cases. The suspected beef burgers were no longer on sale at the time of investigation but three patients confirmed as outbreak cases had deep-frozen some at home. CONCLUSION: Shopper card data was particularly useful to obtain precise and reliable information on the traceability of consumed food. Despite the expired use-by date, a recall was issued for the beef burgers. This contributed to preventing other cases among consumers who had deep-frozen the beef burgers.

Fusobacterium invasive infections in children: a retrospective study in two French tertiary care centres.

The purpose of this investigation was to describe the clinical and biological characteristics and evolution of invasive Fusobacterium infections in children admitted to two French paediatric tertiary care centres. Children who were admitted from 1998 to 2009 to two tertiary care centres for invasive Fusobacterium infection were included in a retrospective study. Thirty-one children with a median age of 5.7 years (interquartile range, IQR [2.3; 9.3]) were included. Nine children had an underlying condition, most commonly sickle cell disease (n = 3) or immunodeficiency (n = 3). Two children had skin effraction prior to the infection. The major sites of infection were the head and neck (n = 14) and abdomen (n = 10). Three children suffered from atypical Lemierre's syndrome. More than half of the children had a bacterial co-infection (58 %). Six children were hospitalised in an intensive care unit, and 67 % of them had a chronic underlying disease. None of the children died. Six children with negative cultures had Fusobacterium identified through 16S RNA-PCR. Fusobacterium is responsible for severe infection in children. Microbiological diagnosis might be improved by the wider use of molecular detection.

[Extended-spectrum beta-lactamase producing-enterobacteriaceae]. / Entérobactéries productrices de bêta-lactamases à spectre étendu en pédiatrie.

Extended-spectrum beta-lactamase (ESBLs) are defined as ß-lactamase able to hydrolyze all penicillins and cephalosporins with the exception of cephamycins (cefotixin, cefotetan), moxalactam and carbapenems and are encoded by mobile genes. The most frequently encountered ESBLs belong to the CTX-M, SHV, and TEM families. ESBLs were found first in Klebsiella pneumonia and then predominantly in E. coli. The incidence of patients with ESBLs E. coli increase since 2000 in Robert Debré Hospital in Paris. They were mainly implicated in urinary tract infections and less frequently in other infections such as materno-foetal infections or neonatal meningitis. An increase of consumption of carbapenems may lead to spread of carbapenem resistant organisms. Thus alternative to carbapenems for treatment of ESBL producers are needed.

[Escherichia coli 0104:H4: a hybrid pathogen]. / Escherichia coli O104:H4: un pathotype hybride.

In 2011, an outbreak linked to a entero-haemorrhagic Escherichia coli strain, affecting adults more frequently, occurred in Germany, with 4320 bloody diarrhea cases, 850 cases of hemolytic uremic syndrome (HUS) and 82 deaths. Meanwhile, an epidemic affecting 24 patients took place in Bègle with similar epidemiological characteristics. These two strains were associated with consumption of contaminated seeds fenugreck by a particularly virulent strain belonging to a rare serotype, E. coli serotype O104:H4. This strain is a triple hybrid : it produces a shigatoxin, the adhesion at the gastrointestinal mucosa is related to the presence of fimbriae as enteroaggregants E. coli (ECAA), and has virulence factors of E. coli outer intestinal (EXPEC). In addition, it produced a ß-lactamasetype extended-spectrum CTXM15.

A new highly discriminatory multiplex capillary-based MLVA assay as a tool for the epidemiological survey of Pseudomonas aeruginosa in cystic fibrosis patients.

Multiple locus variable number of tandem repeats (VNTR) analysis (MLVA) has been shown to provide a high level of information for epidemiological investigations and the follow-up of Pseudomonas aeruginosa chronic infection. In the present study, an automatized MLVA assay has been developed for the analysis of 16 VNTRs in two multiplex polymerase chain reactions (PCRs), followed by capillary electrophoresis. The result in the form of a code is directly usable for clustering analyses. This MLVA-16(Orsay) scheme was applied to the genotyping of 83 isolates from eight cystic fibrosis patients, demonstrating that the same genotype persisted during eight years of chronic infection in the majority of cases. Comparison with pulsed-field gel electrophoresis (PFGE) analysis showed that both methods were congruent, MLVA providing, in some cases, additional informativity. The evolution of strains during long-term infection was revealed by the presence of VNTR variants.

Household transmission of haemolytic uraemic syndrome associated with Escherichia coli O104:H4, south-western France, June 2011.

Following the outbreak of haemolytic uraemic syndrome (HUS) on June 2011 in south-western France, household transmission due to Escherichia coli O104:H4 was suspected for two cases who developed symptoms 9 and 10 days after onset of symptoms of the index case. The analysis of exposures and of the incubation period is in favour of a secondary transmission within the family. Recommendations should be reinforced to prevent person-to-person transmission within households.

[Extended-spectrum beta-lactamase producing-enterobacteriaceae]. / Entérobactéries productrices de bêta-lactamases à spectre étendu.

Extended-spectrum beta-lactamase (ESBLs) are defined as ß-lactamase capable of hydrolyzine oximino-cephalosporins and aztreonam and are encoded by mobile genes. The most frequently encountered ESBLs belong to the CTX-M, SHV, and TEM families. ESBLs were found first in Klebsiella pneumonia and then predominantly in E. coli. The incidence of patients with ESBLs E. coli increase since 2000 in Robert Debré Hospital. They were responsible of cystitis or pyelopnephritis and rarely of materno-foetal infections or neonatal meningitis. These strains were susceptible to colimycin, carbapenems and fosfomycin.

[Streptococcus pyogenes or group A streptococcal infections in child: French national reference center data]. / Infections à Streptococcus pyogenes ou streptocoque du groupe A chez l'enfant : données du Centre national de référence (CNR).

Since the 1980s, infections due to Streptococcus pyogenes or group A streptococci (GAS) were marked by the increase in invasive infections and the emergence of clones which were resistant to macrolides. Those challenges led the French national reference center for streptococci to enhance the epidemiological survey and the characterization of GAS strains, in collaboration with the National Institute for Public Health Surveillance. Active surveillance is of major importance for implementation of therapeutic and prophylactic guidelines and for evaluation of future streptococcal vaccines.

Subtyping of emm1 group A Streptococci causing invasive infections in France.

By combining PCR amplification of toxin-encoding genes and sic gene sequencing, we distinguished 24 genotypes among 47 M/emm1 group A streptococci isolated from children and adults in France in 9 cases of infection comprising four clusters and 38 unrelated invasive infection cases used as controls.

Community-wide outbreak of Escherichia coli O157:H7 associated with consumption of frozen beef burgers.

On 24-25 October 2005 a cluster of five haemolytic uraemic syndrome (HUS) cases was reported in southwest France. An investigation was undertaken to identify the outbreak source and implement control measures. Cases were defined as individuals with HUS or diarrhoea with isolation of Escherichia coli O157:H7 in stools or a positive antibody response to E. coli O157 lipopolysaccharide, resident in southwest France with symptom onset after 19 September 2005. Sixty-nine identified patients had symptom onset between 5 October and 3 November 2005, including 17 cases of HUS. One brand of frozen beef burgers produced on 22 August 2005 was consumed by all patients in the week before symptom onset. E. coli O157:H7 strains from patients, patients' burgers and the manufacturing plant were genetically related. This is the largest community-wide outbreak of E. coli O157:H7 in France to date and the first associated with consumption of contaminated frozen beef burgers.