Comprehensive Contact Tracing, Testing and Sequencing Show Limited Transmission of SARS-CoV-2 between Children in Schools in Norway, August 2020 to May 2021.

The role of children in the spread of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in schools has been a topic of controversy. In this study among school contacts of SARS-CoV-2 positive children in 43 contact-investigations, we investigated SARS-CoV-2 transmission in Norway, August 2020-May 2021. All participants were tested twice within seven to ten days, using SARS-CoV-2 PCR on home-sampled saliva. Positive samples were whole genome sequenced. Among the 559 child contacts, eight tested positive (1.4%, 95% CI 0.62-2.80), with no significant difference between primary (1.0%, 95% CI 0.27-2.53) and secondary schools (2.6%, 95% CI 0.70-6.39), p = 0.229, nor by viral strain, non-Alpha (1.4%, 95% CI 0.50-2.94) and Alpha variant (B.1.1.7) (1.7%, 95% CI 0.21-5.99), p = 0.665. One adult contact (1/100) tested positive. In 34 index cases, we detected 13 different SARS-CoV-2 Pango lineage variants, with B.1.1.7 being most frequent. In the eight contact-investigations with SARS-CoV-2 positive contacts, four had the same sequence identity as the index, one had no relation, and three were inconclusive. With mitigation measures in place, the spread of SARS-CoV-2 from children in schools is limited. By excluding contact-investigations with adult cases known at the time of enrolment, our data provide a valid estimate on the role of children in the transmission of SARS-CoV-2 in schools.

Implications of screening and childcare exclusion policies for children with Shiga-toxin producing Escherichia coli infections: lessons learned from an outbreak in a daycare centre, Norway, 2012.

BACKGROUND: In Norway, it is recommended that children with Shiga-Toxin producing Escherichia coli (STEC) infections are excluded from daycare centers until up to five consecutive negative stool cultures are obtained. Children with gastrointestinal illness of unknown etiology are asked to remain home for 48 hours after symptoms subside. On 16 October 2012, two cases of STEC infection were reported from a daycare center, where other children were also symptomatic. Local health authorities temporarily closed the daycare center and all children and staff were screened for pathogenic E. coli. We present the results of the outbreak investigation in order to discuss the implications of screening and the exclusion policies for children attending daycare in Norway. METHODS: Stool specimens for all children (n = 91) and employees at the daycare center (n = 40) were tested for pathogenic E. coli. Information on demographics, symptoms and potential exposures was collected from parents through trawling interviews and a web-based questionnaire. Cases were monitored to determine the duration of shedding and the resulting exclusion period from daycare. RESULTS: We identified five children with stx1- and eae-positive STEC O103:H2 infections, and one staff member and one child with STEC O91:H- infections. Three additional children who tested positive for stx1 and eae genes were considered probable STEC cases. Three cases were asymptomatic. Median length of time of exclusion from daycare for STEC cases was 53 days (range 9 days-108 days). Survey responses for 75 children revealed mild gastrointestinal symptoms among both children with STEC infections and children with negative microbiological results. There was no evidence of common exposures; person-to-person transmission was likely. CONCLUSIONS: The results of screening indicate that E. coli infections can spread in daycare centres, reflected in the proportion of children with STEC and EPEC infections. While screening can identify asymptomatic cases, the implications should be carefully considered as it can produce unanticipated results and have significant socioeconomic consequences. Daycare exclusion policies should be reviewed to address the management of prolonged asymptomatic shedders.

Identification of the anti-terminator qO111:H)- gene in Norwegian sorbitol-fermenting Escherichia coli O157:NM.

Sorbitol-fermenting Escherichia coli O157:NM (SF O157) is an emerging pathogen suggested to be more virulent than nonsorbitol-fermenting Escherichia coli O157:H7 (NSF O157). Important virulence factors are the Shiga toxins (stx), encoded by stx1 and/or stx2 located within prophages integrated in the bacterial genome. The stx genes are expressed from p(R) (') as a late protein, and anti-terminator activity from the Q protein is necessary for read through of the late terminator t(R) (') and activation of p(R) (') . We investigated the regulation of stx2(EDL933) expression at the genomic level in 17 Norwegian SF O157. Sequencing of three selected SF O157 strains revealed that the anti-terminator q gene and genes upstream of stx2(EDL933) were identical or similar to the ones observed in the E. coli O111:H- strain AP010960, but different from the ones observed in the NSF O157 strain EDL933 (AE005174). This suggested divergent stx2(EDL933) -encoding bacteriophages between NSF O157 and the SF O157 strains (FR874039-41). Furthermore, different DNA structures were detected in the SF O157 strains, suggesting diversity among bacteriophages also within the SF O157 group. Further investigations are needed to elucidate whether the q(O111:H) (-) gene observed in all our SF O157 contributes to the increased virulence seen in SF O157 compared to NSF O157. An assay for detecting q(O111:H) (-) was developed.

Comparison of multilocus variable-number tandem-repeat analysis and multilocus sequence typing for differentiation of hemolytic-uremic syndrome-associated Escherichia coli (HUSEC) collection strains.

Multilocus variable-number tandem-repeat analysis (MLVA) was compared to multilocus sequence typing (MLST) to differentiate hemolytic uremic syndrome-associated enterohemorrhagic Escherichia coli strains. Although MLVA--like MLST--was highly discriminatory (index of diversity, 0.988 versus 0.984), a low level of concordance demonstrated the limited ability of MLVA to reflect long-term evolutionary events.

Study of polymorphic variable-number of tandem repeats loci in the ECOR collection and in a set of pathogenic Escherichia coli and Shigella isolates for use in a genotyping assay.

The Escherichia coli (E. coli) reference collection, ECOR, consists of 72 strains that are representative of the genotypic diversity, as indexed by multilocus enzyme electrophoresis (MLEE), in the species as a whole. MLEE revealed 4 main phylogenetic groups designated A, B1, B2 and D. We present a study of the relationship between the ECOR strains as determined by polymorphisms in seven variable-number of tandem repeats (VNTR) loci. Seven tandem repeats that were present in more than one of the fully sequenced E. coli strains were selected, and primers were constructed in order to amplify the targets in all species where the loci were present. The combined result for all VNTR loci was adapted as a multiple-locus variable-number tandem repeats analysis (MLVA) and showed that the ECOR collection was divided into 63 distinct genotypes. The ECOR phylogenetic groups defined by MLEE were not well conserved by MLVA. A set of 61 pathogenic isolates of both E. coli and Shigella spp. was then tested with the same set of VNTR loci, and revealed 54 distinct genotypes. In addition, the MLVA method was used to genotype isolates from patients and suspected sources in a recent outbreak of E. coli O103 in Norway. The pathogenic E. coli isolates contained the diarrhea causing categories EIEC, EAEC, STEC, ETEC and EPEC. Shigella isolates were of species S. flexneri, S. boydii, S. sonnei and S. dysenteriae. The MLVA method rapidly genotyped all isolates in the study at a Simpson's index of diversity of D=0.98.

Octaplex PCR and fluorescence-based capillary electrophoresis for identification of human diarrheagenic Escherichia coli and Shigella spp.

A multiplex PCR assay, amplifying seven specific virulence genes and one internal control gene in a single reaction, was developed to identify the five main pathotypes of diarrheagenic Escherichia coli and Shigella spp. The virulence genes selected for each category were Stx1, Stx2, and eaeA for enterohemorrhagic E. coli (EHEC), eaeA for enteropathogenic E. coli (EPEC), STIb and LTI for enterotoxigenic E. coli (ETEC), ipaH for enteroinvasive E. coli (EIEC) and Shigella spp., and aggR for enteroaggregative E. coli (EAEC). Each forward primer was labelled with a fluorochrome and the PCR products were separated by multicolour capillary electrophoresis on an ABI PRISM310 Genetic Analyzer (Applied Biosystems). If present, several gene variants of each virulence gene were identified. The internal control gene rrs, encoding 16S rRNA, was amplified in all 110 clinical strains analyzed. Virulence genes were demonstrated in 103 (94%) of these strains. In the majority of the cases (98/103, 95%), classification obtained by the novel multiplex PCR assay was in agreement with that previously determined by phenotypic assays combined with other molecular genetic approaches. Numerous multiplex PCR assays have been published, but only a few of them detect all five E. coli pathotypes within a single reaction, and none of them has used multicolour capillary electrophoresis to separate the PCR products. The octaplex PCR assay followed by capillary electrophoresis presented in the present paper provides a simple, reliable, and rapid procedure that in a single reaction identifies the five main pathotypes of E. coli, and Shigella spp. This assay will replace the previous molecular genetic methods used in our laboratory and work as an important supplement to the more time-consuming phenotypic assays.