Chicken embryo lethality assay for determining the lethal dose and virulence of Enterococcus faecalis

Abstract

Enterococcus faecalis is the major pathogen found in field cases of amyloid arthropathy in chickens. Given the need for a better understanding of the virulence mechanisms of the causative strains, the embryo lethality assay (ELA) is proposed in the present study as a model to evaluate the virulence of E. faecalis strains, specifically the pathogenic avian strain K923/96, which was previously related with amyloid arthropathy. Hence, 0.2 ml of five doses of the cited strain (from 2.5 to 2500 colony-forming units (CFU) per ml) were inoculated into the allantoic cavity of 10-day-old embryos. The embryo mortality rate (EMR) was determined by daily candling of the eggs over a period of seven days and based on this information the median lethal dose (LD) was calculated. The ELA was repeated four times on a sample of 3443 eggs. The infectious dose showed a significant effect on the EMR. The EMR with the doses of 2.5, 5, 25, 250 and 2500 CFU/ml was 43%, 45%, 63%, 90% and 93%, respectively. The estimated dose at LD was 6.6 CFU/ml. As expected, the higher the infectious dose, the greater the EMR and the lower the embryo survival time. The highest EMR was recorded after three and four days post-inoculation in all doses. In conclusion, these results can be used as a basis for further researches on the E. faecalis virulence. In order to corroborate its model capacity to predict the virulence of this bacterium, more ELAs with different E. faecalis strains are required.

Presence of Virulence Genes inEnterococcusSpecies Isolated from Meat Turkeys in Germany Does Not Correlate with Chicken Embryo Lethality

“…Several factors (e.g., infection route, embryo quality, and age at inoculation) are known to influence embryo lethality results and impair comparability (summarized in [65]). Previous studies, however, were often not able to draw definite conclusions about whether the infective dose influences the response.…”

“…ese experiments included enterococci [37,66] but also a variety of other bacteria (e.g., [38,67,68]). A very recent study by Blanco et al [65] used E. faecalis K923/96 only and with different inoculum concentrations to define the median lethality dose in chicken embryos.…”

Assessment of listing and categorisation of animal diseases within the framework of the Animal Health Law (Regulation (EU) No 2016/429): antimicrobial‐resistant Enterococcus faecalis in poultry

“…E. faecalis is generally considered as an opportunistic pathogen, also in birds (Chadfield et al, 2004). Although considered an opportunistic pathogen, strains of E. faecalis may display variations in virulence, as shown by layer chick embryo lethality assays (Blanco et al, 2017(Blanco et al, , 2018.…”