The influence of the cage system and colonisation of Salmonella Enteritidis on the microbial gut flora of laying hens studied by T-RFLP and 454 pyrosequencing

<p>Abstract</p> <p>Background</p> <p>In the EU conventional cages for laying hens are forbidden beginning in January 2012, however concerns about a higher transmission rate of <it>Salmonella </it>in alternative cages systems have been raised. The extent to which cage systems may affect the intestinal microbiota of laying hens is not known, and different microbiota may demonstrate different resistance towards colonization with <it>Salmonella</it>. To investigate this, ileal and caecal samples from two experimental studies where laying hens were inoculated with <it>Salmonella </it>Enteritidis and housed in different systems (conventional cage, furnished cage or aviary), were compared using Terminal Restriction Fragment Length Polymorphism (T-RFLP). The distribution of genera in the microbiota in caecum was furthermore described by next generation sequencing of 16S rDNA libraries.</p> <p>Results</p> <p>Hens in the same cage type developed similar T-RFLP fingerprints of the ileal and caecal microbiota, and these could be separated from layers in the other cages types. No significant difference in the fingerprint profiles could be observed between <it>Salmonella </it>positive and negative samples from same cage. By deep sequencing of 16S rDNA libraries from caecum, 197 different Operational Taxonomic Units (OTU) were identified, and 195 and 196 OTU respectively, were found in hens in aviary and furnished cages, but only 178 OTU of these were recovered from conventional cages. The ratio between the dominating phyla or families and genera in the microbiota remained fairly constant throughout the study. <it>Faecalibacterium </it>and <it>Butyricimonas </it>were the most prevalent genera found in the caecal microbiota of layers irrespective of the cage type.</p> <p>Conclusions</p> <p>Hens confined in the same cage group tend to develop similar microbiota in their ileum and caecum possibly due to isolation, while differences in the microbiota between cages may be caused by environmental or individual bird factors. Although the cages type had influence on composition of the microbiota in the layers by promoting higher diversity in furnished and aviary systems, we did not observe differences in colonization and excretion pattern of <it>Salmonella </it>from these groups. We suggest, that differences in group size and exposure to a more faecally contaminated environment in the alternative systems may explain the observed differences in diversity of the caecal microbiota.</p

Salmonella Enteritidis is superior in egg white survival compared with other Salmonella serotypes

Salmonella enterica subspecies enterica serotype Enteritidis is a major cause of egg-borne human salmonellosis. The ability to survive in egg albumen at chicken body temperature was hypothesized to be an important factor involved in the predominant contamination of eggs by this specific serotype. Eighty-nine Salmonella strains from different serotypes, belonging to 5 serogroups, were incubated for 24 h in egg white at 42 degrees C. The number of Salmonella Enteritidis strains that were able to survive in egg white was significantly higher compared with strains belonging to other serotypes and serogroups that were tested in this study. These data add evidence to the hypothesis that egg white survival is one of the reasons why Salmonella Enteritidis is more predominantly isolated from contaminated eggs, and helps explaining why most reported egg-borne Salmonella outbreaks in humans are caused by Salmonella Enteritidis

Horizontal transmission of Salmonella Enteritidis in groups of experimentally infected laying hens housed in different housing systems

Concerns regarding the welfare of laying hens have led to the ban of conventional battery cages in Europe from 2012 onward and to the development of alternative housing systems that allow hens to perform a broader range of natural behaviors. Limited information is available about the consequences of alternative housing systems on transmission of zoonotic pathogens such as Salmonella Enteritidis. However, Salmonella enterica serovar Enteritidis continues to be one of the leading causes of bacterial foodborne disease worldwide and this is mainly attributed to the consumption of contaminated eggs. A transmission experiment was performed to quantify the effect of the housing system on the spread of a Salmonella Enteritidis infection within a group of layers and on internal egg contamination. At the age of 16 wk, 126 birds housed on the floor were inoculated with Salmonella Enteritidis. Three weeks later, the inoculated hens were housed together with equal numbers of susceptible contact animals in 4 different housing systems: a conventional cage system, a furnished cage, an aviary, and a floor system. Transmission and egg contamination were followed during a 4-wk period. A trend toward increased bird-to-bird transmission was detected in the aviary and floor system compared with the cage systems. Also, significantly more contaminated eggs were found in the aviary compared with the cage systems and the floor system. These results suggest that there is a clear need to optimize and maintain Salmonella surveillance programs when laying hens will be moved from conventional cage systems to alternative housing systems

Effect of the housing system on shedding and colonization of gut and internal organs of laying hens with Salmonella enteritidis

As a result of welfare considerations, conventional cage systems will be banned in the European Union from 2012 onward. Currently, there is limited information on the level of contamination with zoonotic pathogens related to the laying hen housing system. Therefore, 2 studies were designed to investigate the effect of the housing system on colonization of layers with Salmonella. In both studies, layers were housed in 3 different housing systems: a conventional cage system, a furnished cage, and an aviary. At 18 wk of age, all birds were orally inoculated with Salmonella Enteritidis. Shedding and colonization were measured at regular time points. The results did not show an increased risk for alternative housing systems compared with the conventional battery cage system. In contrast, in one study, a faster decline in shedding was noted for layers housed in the alternative cage systems in comparison with the conventional cage system. This study does not give indications that housing layers in alternative systems will yield a risk for increased Salmonella contamination

Faecal sampling underestimates the actual prevalence of Salmonella in laying hen flocks

In all European Union member states, Salmonella monitoring in poultry flocks is obligatory. In these monitoring programmes, a limited number of pooled faeces and/or dust samples are collected to determine whether Salmonella is present in the flocks or not. Whether these limited sampling protocols are sufficiently sensitive to detect expected low within-flock prevalences of an intermittently shed pathogen is not yet clear. In this study, a comparison is made between different sampling procedures for the assessment of the between- and within-flock prevalence of Salmonella in laying hens. In total, 19 farms were sampled. Using a comparable sampling methodology as in the official surveillance programmes, Salmonella could not be detected in any of the flocks. After transportation of the hens to the laboratory and subsequent analysis of cloacal swabs and caecal contents, Salmonella Enteritidis was detected in laying hens from five of 19 farms. The observed within-flock prevalence ranged from 1% to 14%. Based on the results of this study, it can be expected that, depending on the sampling procedure, different estimates of the prevalence of Salmonella can be obtained and the proportion of Salmonella infected flocks is underestimated based on the results of the official monitoring programme