Enteromyxum leei in mediterranean gilthead sea bream farms: how much is out there?

Trabajo presentado en la International Conference & Exposition Aquaculture Europe, celebrada en Funchal, Maderia (Portugal) del 04 al 07 de octubre de 2021.[Introduction]: Enteromyxum leei (Myxozoa) is an important pathogen of gilthead seabream (GSB), Sparus aurata and other Sparidae fish cultured in the Mediterranean. In GSB, E. leei induces chronic enteritis followed by anorexia, cachexia, and even death. It can cause significant growth delay and mortality in marine cage farms as well as land-based mariculture systems. Water temperature and water recirculation are critical risk factors in the contagion and onset of enteromyxosis (Palenzuela et al., 2006, Sitjà-Bobadilla and Palenzuela 2012). However, the relevance of other risks factors like farm management practices, and the epidemiological situation in gilthead sea bream farms, have not been thoroughly determined. In this study, a risk assessment questionnaire and cross-sectional epidemiological study was designed to get insights in the distribution and incidence of E. leei in more than 50 Mediterranean and Atlantic GSB farms throughout Western Europe.[Methods]: A questionnaire for GSB cage farms was designed to ascertain information about farm background and self-awareness of risk relative to E. leei infection, as well as details on the facilities, production and farm management routines. In addition, the farms were invited to participate in a cross-sectional targeted epidemiological survey to test the presence of the parasite in their stock. The participant farms were recruited by local expert pathologists and consultants acting as nodes in each geographical region, and the samples and questionnaires were coded and studied blindly. For those farms participating in the biological sampling, a protocol was submitted to sample one batch of fish at harvest time. The sampling was designed to detect prevalence higher than 9.5% with a 95% confidence. The intestinal rectal ampoules (portion with maximum predictive value for E. leei detection in GSB) were taken from at least 30 randomly selected fish during routine harvest from one or several cages. For the validation of the survey protocol, a land-based farm which is enzootic for the parasite was enrolled in the study and two different GSB lots were sampled as positive controls. The samples were fixed in 80-90 º Ethanol and paired into 15 tubes, which were shipped to the central diagnosis laboratory at IATS-CSIC. The routine procedures for diagnosis of E. leei consist of a SYBR-green qPCR test with primers specific for the parasite against a standard curve containing known numbers of copies of the target gene. Upon reception at the laboratory the samples were individually processed and homogenised. DNA was extracted from an aliquot using a robotic system, its quantity and quality were evaluated, and samples were tested by qPCR at two different dilutions. They were considered positive when the cycle threshold (Ct) was