Isolation of Listeria spp. from faecal samples of cracked egg fed chickens and RAPD analysis of Listeria monocytogenes strains.

At 56 weeks of age, 30 Hyline (W-377) laying pullets were randomly distributed in individual (1 pullet per cage) 45 x 45 x 35 cm cages. Cracked egg was used as protein supplement to the diet. Three groups of 10 Hyline laying pullets each were formed. Three isocaloric and isonitrogenous diets were prepared from different protein sources as followed: ration 1 (control) − soybean meal; ration 2 (experiment 1) – soybean meal + cracked egg (3.25%), ration 3 (experiment 2) – soybean meal + cracked egg (7.50%). A total of three feed and 30 faecal samples were obtained and examined for the presence of Listeria spp. Samples were enriched using Listeria Enrichment Broth and inoculated onto Listeria Selective Agar. The isolates were identified by both conventional methods and polymerase chain reaction (PCR). A 701 bp fragment of listeriolysin O sequence for Listeria monocytogenes was amplified using specific primers by PCR for confirmation of the identification. In ration 3 (soybean meal + cracked egg 7.50%) and ration 2 (soybean meal + cracked egg 3.25%), L. monocytogenes and Listeria innocua, were respectively isolated by culture. In the faeces from hens fed ration 3, L. monocytogenes (5 hens) and Listeria welshimeri (1 hen) were detected by culture. Birds fed ration 2 exhibited positive culture results for L. innocua (4 hens) and L. welshimeri (1 hen). All L. monocytogenes strains that were positive by culture were also detected to be positive by the PCR. Random amplified polymorphic DNA (RAPD) were used to detect the genetic variability among L. monocytogenes isolates. All L. monocytogenes isolates were typed by RAPD using a random primer (OPA-11). In the molecular epidemiological analysis using RAPD assay, the isolates produced three different band profiles using OPA-11. The results of this study demonstrated that multiple genotypes of L. monocytogenes isolates existed in faeces of hens fed rations supplemented with cracked eggs.