Study participants and data collection
Group 1
The first group consisted of 29 students and 3 teachers from a high school in the city of Lleida. The team arrived at the facility on April 9, 2018, the first case of gastroenteritis occurred in the afternoon of October 4, 2018, and an outbreak was reported on April 11, 2018, with 19 people affected. The clinical manifestations of those affected are vomiting, abdominal pain, diarrhea and, in some cases, low-grade fever. Food consumption information is as follows: Lunch on September 4, 2018, rice salad with mayonnaise, breaded meat with French fries and orange; dinner on the same day consisted of green beans, sausage, French fries and yogurt. Breakfast on April 10, 2018 consisted of toast and ham. Lunch that day consisted of cheese sandwiches, sausage sandwiches and an apple that they took out for a picnic, where they drank water from a fountain in Aigues Totes National Park.
Group 2
The second group consisted of 32 students and 3 teachers from the same high school, who arrived at the facility on April 16, 2018, the day after the first case emerged. The second outbreak was reported on April 18, 2018, with 12 affected individuals showing similar clinical manifestations to the previous group. Information on the food consumed is as follows: the lunch on April 16, 2018 was macaroni bolognese, hake fillet with green salad and ice cream, and the dinner was green beans with potatoes, cheese omelette and yogurt. Breakfast on 17 April 2018 includes bread and butter, jam, sausages and pastries. The same day we visited Aigues Totes National Park and had a picnic consisting of a ham sandwich, a sausage sandwich and an apple. They also drank water from a fountain in Aigues Totes National Park.
Based on the data collected, the study hypothesized a foodborne origin for the two acute gastroenteritis outbreaks. ESS designed a questionnaire to collect information on demographic variables, food consumption, clinical symptoms, and date of symptom onset. Questionnaires were sent to high school principals and distributed to panel members. Subsequently, questionnaires were collected and sent to ESS for analysis. The holiday camp is staffed by two food handlers. ESS technicians conducted telephone interviews with food handlers to collect medical history or presence of symptoms of acute gastroenteritis. The definition of an outbreak considered was the presence of two or more members of a group experiencing gastrointestinal symptoms. The following case definition was established: A student or teacher who developed vomiting and/or diarrhea or two or more of the following symptoms on or between 04/9/2018 and 04/11/2018 (Group 1): abdominal pain, fever and Disgusting April 16, 2018 and April 18, 2018 (Group 2). A confirmed case is considered when a microbiological test result is positive. The outbreak has been reported to the Health Protection Agency for investigation into the food handling procedures followed and for inspection of the facility’s food safety conditions.
Fecal samples were collected from each group of affected individuals and food handlers. Among food handlers, although they were identical, the first sample was taken during the first outbreak and the second sample was taken during the second outbreak. Fecal samples from the outbreak were analyzed by the microbiology laboratory at Hebron Valley University Hospital for use in studying community food poisoning outbreaks. The presence of the following enteropathogenic bacteria was studied: Salmonella, Shigella, Yersinia, Campylobacter and E. coli O157 by routine culture. Enteroviruses were detected using real-time PCR and the Allplex™ GI Virus Assay (Seegene, Inc.), which also detects NoV GI, NoV GII, adenovirus, rotavirus, sapovirus, and astrovirus. Samples positive for NoV GI and/or NoV GII were genotyped using GISKF-GISKR primers for GI and G2SKF-G2SKR primers for GII as described by Kojima et al.18. Amplification was performed by RT-PCR using a one-step RT-PCR kit (Qiagen, Hilden, Germany). Sequences were obtained using the ABI 3730 platform (Applied Biosystems, Foster City, California, USA) and assembled using the SeqMan 4.05 program (Dnastar, Madison, WI, USA). After obtaining the sequence, the norovirus typing tool version 2.0 (https://www.rivm.nl/mpf/typingtool/norovirus/) was used to obtain the genotype. Food samples were analyzed at the Girona Public Health Laboratory. NoV GI and NoV GII were detected by RT-PCR using a one-step RT-PCR kit (CeeramTools®, Biomérieux Spain). Using the Thermocycler 7500 Fast Real-Time PCR System (Applied Biosystems)19.
Statistical Analysis
Survey coverage and incidence rates, distribution by age and sex groups, prevalence curves, incubation periods and symptom frequencies were determined. The effect of each food consumption was determined by calculating the risk ratio (RR) and its 95% confidence interval (CI). When the value in a cell in the contingency table is zero and the RR cannot be calculated, 0.5 is added to each cell to calculate the RRtwenty. Fisher’s exact test was used to calculate p values.
Ethics Statement and Informed Consent Statement
The study was conducted in accordance with the guidelines of the Declaration of Helsinki, the regulations of the Catalan Directorate of Public Health and established ethical protocols. The study was approved by the Bioethics Committee of the University of Barcelona on April 12, 2016 (ethical approval number IRB00003099).
The authors declare that the exemption from informed consent was approved by the Bioethics Committee of the University of Barcelona. All data used in the analysis were collected during routine public health surveillance activities as part of the statutory mandate of the Catalan Ministry of Health, which is duly authorized to receive, process and temporarily store personal data in the context of infectious diseases . All data is completely anonymous. All research activities are part of the public health surveillance mission. The law regulates these activities and does not require informed consent.