S. Van Hoorebeke

Determination of the within and between flock prevalence and identification of risk factors for Salmonella infections in laying hen flocks housed in conventional and alternative systems

Salmonella outbreaks in humans are often linked with the consumption of contaminated eggs. Therefore a profound knowledge of the actual prevalence of Salmonella spp. in laying hens and the factors that influence the presence and persistence of Salmonella on a farm is of utmost importance. The housing of laying hens in conventional battery cages will be forbidden in the European Union (EU) from 2012 onwards. There is an urgent need to evaluate whether this move to alternative housing systems will influence the prevalence of Salmonella in laying hens. Therefore, a cross-sectional study was performed in 5 European countries (Belgium, Germany, Greece, Italy and Switzerland) to determine the between and within flock prevalence of hens shedding Salmonella and to investigate whether there is an effect of the housing type on Salmonella prevalence. In total 292 laying hen farms were sampled in the month prior to depopulation. An on-farm questionnaire was used to collect information on general management practices and specific characteristics of the sampled flock. Twenty-nine flocks were found positive for at least 1 Salmonella-serotype. In these flocks the within flock prevalence of shedding hens, determined by individual sampling of 40 hens, varied between 0% and 27.50%. A wide variety of serotypes was isolated with Salmonella Enteritidis as the most common. Housing in conventional battery cages, the absence of dry cleaning in between production rounds and sampling in winter turned out to be risk factors for the shedding of Salmonella Enteritidis or Typhimurium (P<0.05).

The influence of the housing system on Salmonella infections in laying hens: a review.

From 2012 onwards, housing of laying hens in conventional battery cages will be forbidden in the European Union and only enriched cages and non‐cage housing systems such as aviaries, floor‐raised, free‐range and organic systems will be allowed. Although this ban aims at improving the welfare of laying hens, it has also initiated the question whether there are any adverse consequences of this decision, especially with respect to the spread and/or persistence of zoonotic agents in a flock. A zoonotic agent that is traditionally associated with the consumption of eggs and egg products is Salmonella enteritidis. This paper provides a summary of the current knowledge regarding the direct and indirect effects of different housing systems on the occurrence and epidemiology of Salmonella in laying hen flocks.

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.

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.

Faecal Sampling Underestimates the Actual Prevalence of Salmonella in Laying Hen Flocks

Summary 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.

Salmonella Gallinarum field isolates from laying hens are related to the vaccine strain SG9R

Salmonella enterica subspecies enterica serotype Gallinarum can cause severe systemic disease in chickens and a live Salmonella Gallinarum 9R vaccine (SG9R) has been used widely to control disease. Using whole-genome sequencing we found point mutations in the pyruvate dehydrogenase (aceE) and/or lipopolysaccharide 1,2-glucosyltransferase (rfaJ) genes that likely explain the attenuation of the SG9R vaccine strain. Molecular typing using Pulsed Field Gel Electrophoresis and Multiple-Locus Variable number of tandem repeat Analysis showed that strains isolated from different layer flocks in multiple countries and the SG9R vaccine strain were similar. The genome of one Salmonella Gallinarum field strain, isolated from a flock with a mortality peak and selected on the basis of identical PFGE and MLVA patterns with SG9R, was sequenced. We found 9 non-silent single-nucleotide differences distinguishing the field strain from the SG9R vaccine strain. Our data show that a Salmonella Gallinarum field strain isolated from laying hens is almost identical to the SG9R vaccine. Mutations in the aceE and rfaJ genes could explain the reversion to a more virulent phenotype. Our results highlight the importance of using well defined gene deletion mutants as vaccines.

The age of production system and previous Salmonella infections on-farm are risk factors for low-level Salmonella infections in laying hen flocks

An explorative field study was carried out to determine risk factors for Salmonella infections in commercial laying hen flocks. For this purpose, 29 laying hen farms, including farms using conventional and alternative housing systems, were intensively sampled. An on-farm questionnaire was used to collect information on general management practices and specific characteristics of the sampled flock such as flock size, age of the hens, and age of the infrastructure. Salmonella was detected in laying hens from 6 of the 29 sampled farms. Using multivariate logistic regression with the Salmonella status of the flock as an outcome variable, a previous Salmonella contamination on the farm and the age of the production system were identified as risk factors for the presence of Salmonella in laying hens (P<0.05). The housing system did not have a significant influence on the prevalence of Salmonella in the current study.

The dynamics of Salmonella occurrence in commercial laying hen flocks throughout a laying period

Contaminated eggs and egg products have been recognized for many years as an important source of Salmonella infections in humans in the European Union and in the United States. Longitudinal studies can help to increase our knowledge about the dynamics of the occurrence of Salmonella in the course of a laying period. The total of 41 laying hen flocks—18 in Belgium, six in Denmark and 17 in Germany—were followed during an entire laying period. Samples taken from the empty cleaned and disinfected poultry houses were all negative for Salmonella. After hens arrived on the farms, five pooled faecal samples, one pooled dust sample and 40 cloacal swabs (Belgium and Germany) or 40 swabs from fresh droppings (Denmark) were taken four times from 18 flocks, three times from 21 flocks and two times from two flocks in the course of the laying period. Ten flocks (two Belgian and eight German flocks) tested up to three times positive for Salmonella. Forty-three out of 50 positive samples contained Salmonella Enteritidis phage type 4 (29 isolates) or phage type 8 (14 isolates). The probability of subsequent Salmonella-positive findings increased significantly in Salmonella-positive flocks (P<0.05, odds ratio = 6.4). However, the probability of finding Salmonella did not depend on the time of sampling in the laying period or the season.

Antimicrobial resistance of Escherichia coli and Enterococcus faecalis in housed laying-hen flocks in Europe.

The aim of this study was to determine the potential association between housing type and multiple drug resistance (MDR) in Escherichia coli and Enterococcus faecalis isolates recovered from 283 laying-hen flocks. In each flock, a cloacal swab from four hens was collected and produced 1102 E. coli and 792 E. faecalis isolates. Broth microdilution was used to test susceptibility to antimicrobials. Country and housing type interacted differently with the MDR levels of both species. In the E. coli model, housing in a raised-floor system was associated with an increased risk of MDR compared to the conventional battery system [odds ratio (OR) 2·12, 95% confidence interval (CI) 1·13-3·97)]. In the E. faecalis model the MDR levels were lower in free-range systems than in conventional battery cages (OR 0·51, 95% CI 0·27-0·94). In Belgium, ceftiofur-resistant E. coli isolates were more numerous than in the other countries.

Clostridium novyi type B as a causative agent of bovine meat spoilage

A series of bovine meat spoilage cases in which meat from clinically healthy Belgian Blue cattle showed green discoloration are described. Histology of skeletal muscle revealed numerous spore-forming rods in the discolored areas of the meat. These organisms stained positively for Clostridium novyi by immunohistochemistry. A combination of 16S rDNA and fliC gene sequencing of bacterial DNA, isolated from the spoiled meat samples, revealed the unique presence of C. novyi type B. Although this bacterium has been implicated in clinical necrotic hepatitis in cattle, the cases described here are the first implicating C. novyi type B as a cause of bovine meat spoilage.