Danish Broiler Research Articles

Prevalence and Transmission of Extended-Spectrum Cephalosporin (ESC) Resistance Genes in Escherichia coli Isolated from Poultry Production Systems and Slaughterhouses in Denmark.

The emergence of extended-spectrum cephalosporin (ESC)-resistant Escherichia coli is a global concern. This study aimed to assess the prevalence and transmission of ESC-resistant E. coli in the Danish broiler production system. Samples from two vertically integrated Production Systems (1 and 2) and two slaughterhouses (A and B) were analyzed (n = 943) for the occurrence of ESC-resistant E. coli from 2015 to 2018. ESC-resistant E. coli isolates were whole-genome sequenced (WGS) for characterization of the multi-locus sequence type (MLST), antibiotic resistance genes, virulence genes, and plasmid replicon types. An ad hoc core genome (cg) MLST based on 2513 alleles was used to examine the genetic relatedness among isolates. The prevalence of ESC-resistant E. coli in the conventional Production System 1 was 2.7%, while in Production System 2 the prevalence was 26.7% and 56.5% for samples from the conventional and organic production, respectively. The overall prevalence of ESC-resistant E. coli in broiler thigh and fecal samples ranged from 19.3% in Slaughterhouse A to 22.4% in Slaughterhouse B. In total, 162 ESC-resistant E. coli were isolated and shown to belong to 16 different sequence types (STs). The most prevalent STs were ST2040 (n = 85) and ST429 (n = 22). Seven ESC resistance genes were detected: blaCMY-2 (n = 119), blaTEM-52B (n = 16), blaCTX-M-1 (n = 5), blaTEM-52C (n = 3), blaCTX-M-14 (n = 1), blaSHV-12 (n = 1), and up-regulation of ampC (n = 16), with an unknown resistance gene in one isolate (n = 1). The carriage of blaCMY-2 in 119 isolates was primarily associated with IncI1 (n = 87), and IncK plasmids (n = 31). Highly similar blaCMY-2 carrying E. coli isolates from ST429 were found in production systems as well as in slaughterhouses. In conclusion, findings from this study indicate that ESC-resistant E. coli are transferred vertically from farms in the production systems to slaughterhouses with the potential to enter the food supply.

Genomic traits of Aeromonas veronii isolated from slaughtered Danish broilers

Aeromonas veronii is a Gram-negative opportunistic bacterium found in fish, poultry and humans and has occasionally been associated with disease although not generally considered a poultry pathogen. A. veronii was recently isolated from both healthy and condemned broiler carcasses at a major Danish abattoir. In this study, we did a whole genomic analysis of 24A. veronii strains from the abattoir to determine their potential sources and relatedness as well as their pathogenic potential, antimicrobial resistance determinants and associated mobile elements. No strains were multi-drug resistant, but all strains carried the beta-lactam resistance genes cphA3 and blaOXA-12 without being phenotypically resistant to carbapenems. One strain carried an IncA plasmid with tet(A), tet(B) and tet(E) genes. A phylogenetic tree including public A. veronii sequences showed that our isolates were not clonal but were dispersed around the phylogenetic tree, suggesting a diffuse spread of A. veronii across human, aquatic and poultry samples. Strains carried different virulence factors known to be associated with pathogenesis and severity of disease in animals and humans, e.g. type II (aerolysin, amylases, proteases, and cytotoxic enterotoxin Act) and III secretion systems where the latter has been associated with mortality in hospitalized patients. Although our genomic analysis of A. veronii shows zoonotic potential, epidemiological studies of human gastro-enteritis cases of A. veronii associated with consumption of broiler meat are needed. It remains to be proven if A. veronii is a true poultry pathogen and part of the established microflora in abattoirs and the gut-intestinal microflora of poultry.

Dramatic increase in slaughter condemnations due to Escherichia coli ST23 and ST101 within the Danish broiler production

Escherichia coli constitutes a major challenge to poultry even when the prevalence of colibacillosis is low. Additionally, specific E. coli strains can severely enhance the detrimental effects on productivity, animal welfare and antimicrobial use.In 2019–2020, a dramatic increase in colibacillosis occurred among Danish broilers causing late-onset mortality and high slaughter condemnations. In the present study, the pathology and causative E. coli-types were characterised. Furthermore, the outbreak-related strains were compared to isolates from concurrent “background” colibacillosis.During the study, 1039 birds were subjected to a comprehensive post-mortem examination, and a total of 349 E. coli isolates were sequenced and characterised by multi-locus sequence typing, virulence and resistance gene presence, plasmid replicon content and phylogenetic analysis.Productivity data from outbreak flocks revealed a mortality of 6.34% ± 3.74 and a condemnation of 5.04% ± 3.67. Contrary, the numbers were 3.18% ± 1.57% and 1.02% ± 0.4 among non-outbreak flocks, respectively. Major lesions were cellulitis (46.82%), airsacculitis (67.63%), pericarditis (55.49%), perihepatitis (41.04%) and femoral head necrosis with physeal/metaphyseal involvement (44.51%). Among non-outbreak broilers, the prevalence was 4.46%, 7.64%, 7.01%, 3.82% and 8.28%, respectively. ST23 and ST101 dominated heavily in outbreak flocks, whereas non-outbreak related isolates consisted of various other STs. A low level of resistance markers was evident, except in few multidrug-resistant isolates. Within ST23 and ST101, 13 and 12 virulence genes were significantly over-represented compared to non-outbreak isolates.In conclusion, clonal lineages were documented as the cause of a devastating outbreak of colibacillosis with great prospects for future interventions.

Prevalence of Potential Pathogenic and Antimicrobial Resistant Escherichia coli in Danish Broilers.

Avian pathogenic Escherichia coli (APEC) are important bacteria in broiler production in terms of economy, welfare, and use of antibiotics. During a previous outbreak of APEC in the Nordic countries, it was suggested that the pathogenic clones of E. coli causing the outbreak originated from grandparent stock and were transmitted to the offspring, causing increased first week mortality. This study investigated whether the pathogenic potential of E. coli at the parent and broiler level differs in relation to pathogenic potential described by the level of virulence-associated genes and pattern of antimicrobial resistance. The hypothesis was that, due to higher biosecurity at the parent level, the E. coli population will show a lower level of antimicrobial resistance and carry fewer virulence-associated genes, as a result of fewer E. coli infections observed. From four parent flocks and eight broiler flocks, 715 E. coli were isolated from cloacal swabs of newly hatched chickens (Ross 308). The isolated E. coli were characterized by eight virulence-associated genes and phenotypic resistance against six antimicrobials. It was found that the prevalence of virulence-associated genes and phenotypic antimicrobial resistance varied significantly between flocks, and the virulence-associated genes papC and irp2 and resistance against ampicillin were significantly more prevalent in breeder flocks compared to broiler flocks.

A model using an inter-sectorial data integration process indicates that reducing Campylobacter cross-contamination at slaughter mitigates the risk of human campylobacteriosis effectively

The risk of human campylobacteriosis due to Danish broiler flocks cross-contaminated (CC) at slaughter with Campylobacter spp. was assessed. National surveillance data (2018) on flock Campylobacter status (called animal level component (AL)) and on contamination of chilled carcasses ready for consumption (food safety component (FS)), were compared. The AL component consisted of PCR testing results on pools of cloacal swabs collected from 3,012 conventional flocks, while the FS component presented culture testing of leg skins from 999 (of the 3,012) randomly sampled flocks. Datasets were integrated on flocks tested in both components, by combinations of farm-ID, house-unit and sampling date. The CC flocks were those entering the slaughterhouse as AL-negative, but resulting FS-positive. All remaining carcass positive flocks were instead classified as Non-CC flocks. The apparent prevalence (AP) of carcass positive flocks and the colony forming units per gram (cfu/g), measured by the FS component, were fed into a published simulation model, to assess under three simulation scenarios: the mean monthly risk per serving during 2018, relative (RR) to that of 2013 (reference year in the current Danish Action Plan). In the baseline scenario, the original AL status and the FS cfu/g were maintained. In the alternative scenarios I and II, the FS cfu/g were set = 0 (i.e. negative) for the Non-CC and for the CC flocks, respectively. Thus, scenario I and the differences between the other two scenarios, provided the contribution of the CC flocks to the AP and to the RR. The (overall) annual median log10 cfu/g was ≈ 2.8 (min. = 1.0; max. = 4.0) for the Non-CC flocks and 1.4 (1.0; 3.9) for the CC flocks. The median monthly difference in AP, between the baseline scenario and scenario II was 7% (min = 2% in January; max = 19% in August), while the difference in risk was 0.04% (0.001%; 0.11%), which was similar to the mean monthly risk under scenario I. If cross-contamination had not occurred (scenario II), the annual AP would have reduced from 24.3% to 16.1% and the RR would have reduced from 0.92 to 0.77. Therefore, ≈16% of the public health risk posed by Danish conventional broiler meat, appeared attributable to CC flocks. Reducing cross-contamination could mitigate the risk of human campylobacteriosis notably. This study illustrates how inter-sectorial surveillance data integration, can be used to optimize National Action Plans against Campylobacter spp. and other similar foodborne pathogens.

Assessing Campylobacter cross-contamination of Danish broiler flocks at slaughterhouses considering true flock prevalence estimates and ad-hoc sampling

Campylobacter cross-contamination of Danish broiler flocks at slaughterhouses was investigated using data from two national surveillance components and from ad-hoc sampling. The animal level (AL) and food safety (FS) components from 2018 were compared. The AL component contained results of PCR on pools of cloacal swabs from 3,012 flocks processed at two Danish slaughterhouses (S1-S2), while the FS component regarded culture testing of leg skins from 999/3,012 flocks. The monthly “apparent” (AP) and “true” flock prevalence (TP) were estimated. Agreement between components was measured in percentage and in weighted-Kappa values. The relationship between the occurrence of cross-contamination (flock positive only in the FS component = cross-contaminated or CC, vs. flock negative in both components or NegBoth), slaughterhouse and surveillance period (quarter: Q1 to Q4) was evaluated by a generalized linear mixed effects (GLM) model. Thereafter, a linear mixed effects (LME) model was used to investigate the relationship between the level of meat contamination of carcass positive flocks (y = log10 colony forming units per gram, cfu/g), slaughterhouse, surveillance period, and flock type (CC vs. positive in both components or PosBoth). For both models, the farm was the random effect. Finally, in autumn 2019, ad-hoc field investigations were carried out testing caecal and neck skin samples, from two consecutive flocks at S1 and S2. Whole genome sequencing (WGS) was performed on isolates, for multilocus sequence typing (MLST) and single nucleotide polymorphisms (SNP) analysis. The monthly TP was always higher for the FS than for the AL component. Agreement between the components was substantial, but 8.1–8.6% of the flocks were CC. Those had median cfu/g 21–28 times lower than that of PosBoth flocks. In the GLM model, the explanatory variables were both significant (P-value <0.05). For example, the odds ratios (ORs) were 8.4 (95% CI: 4.0; 17.6) for Q3 vs. Q1, and 3.1 (1.8; 5.2) for S2 vs. S1. In the LME model, the flock type and the interaction between the other two variables, were significant. In the field study, a caecal positive flock was succeeded by an initially negative flock, in one out of five sampling sessions at S2. The cecal negative flock was positive in 58.3% of the neck skins with the isolate genetically similar to that from the caecal positive flock. Those results show that cross-contamination can be affected by surveillance periods and slaughterhouses, and it can contribute significantly to the TP of carcass positive flocks.

Identification of Novel Phage Resistance Mechanisms in Campylobacter jejuni by Comparative Genomics.

Phages infecting Campylobacter jejuni are considered a promising intervention strategy at broiler farms, yet phage sensitivity of naturally occurring poultry isolates is not well studied. Here, we investigated phage sensitivity and identified resistance mechanisms of C. jejuni strains originating from Danish broilers belonging to the most prevalent MLST (ST) types. Determining plaque formation of 51 phages belonging to Fletchervirus or Firehammervirus showed that 21 out of 31 C. jejuni strains were susceptible to at least one phage. While C. jejuni ST-21 strains encoded the common phase variable O-methyl phosphoramidate (MeOPN) receptor of the Fletchervirus and were only infected by these phages, ST-45 strains did not encode this receptor and were exclusively infected by Firehammervirus phages. To identify internal phage resistance mechanism in ST-21 strains, we performed comparative genomics of two strains, CAMSA2002 sensitive to almost all Fletchervirus phages and CAMSA2038, resistant to all 51 phages. The strains encoded diverse clustered regularly interspaced short palindromic repeats (CRISPR) spacers but none matched the tested phages. Sequence divergence was also observed in a predicted SspE homolog and putative restriction modification systems including a methyl-specific McrBC endonuclease. Furthermore, when mcrB was deleted, CAMSA2038 became sensitive to 17 out of 43 phages, three being Firehammervirus phages that otherwise did not infect any ST-21 strains. Yet, 16 phages demonstrated significantly lower efficiencies of plating on the mcrB mutant suggesting additional resistance mechanism still restricting phage propagation in CAMSA2038. Thus, our work demonstrates that C. jejuni isolates originating from broilers may have acquired several resistance mechanisms to successfully prevent phage infection in their natural habitat.

Risk-based control of Campylobacter spp. in broiler farms and slaughtered flocks to mitigate risk of human campylobacteriosis – A One Health approach

Effects of risk-based control of Campylobacter spp. in Danish broiler farms and flocks were simulated, to assess potential reductions of human risk of campylobacteriosis, associated to the consumption of poultry meat produced in Denmark. Two national data streams were used and represented: Flock status by testing cloacal swabs (CS, 2018–2019) and carcass status by testing leg skin samples (LS, 2019). In the CS surveillance component all flocks slaughtered at the two major Danish slaughterhouses were tested with a polymerase chain reaction (PCR), while in LS one third randomly selected flocks were tested by culture (results in colony forming units per gram, cfu/g). Each farm was identified by its Central Husbandry Register (CHR) number. Two risk farm classification strategies (I-II) were based on CS data from 2018. Farms were classified as: always negative (Neg-CHRs), low risk (LowR-CHRs) and high risk (HighR-CHRs) farms. In strategy I, HighR-CHRs had more than five positive flocks, while in strategy II; they had more than 27.8% of the slaughtered flocks positive. Those two cut-offs were the annual 3rd quartiles across positive farms. Thereafter, a risk assessment model was used to estimate the annual relative risk (RR) of human campylobacteriosis in 2019, compared to that of 2013. Three hypothetical levels of cfu/g reductions (A, B and C) were simulated on the LS positive flocks (> 10 cfu/g) slaughtered by HighR-CHRs and were pairwise combined with the two classification strategies, yielding six risk-mitigation scenarios (A I-II; B I-II; C I-II). In scenarios A I-II, zero cfu/g were simulated, while in scenarios B and C, the original cfu/g were divided by three and by two. For each scenario, RRs were compared to the RR of the original cfu/g (scenario O).In 2018, if all flocks from HighR-CHRs had been negative, the annual CS flock prevalence would have reduced from 19.7% to 7.6% (strategy I) or 9.6% (strategy II). Whereas in 2019, it would have reduced from 17.1% to 7.8% or 11.6%. In both years, HighR-CHRs delivered a high percentage of the total annual positive flocks (61.4–54.4% under strategy I and 51.2–32.6% with strategy II). In 2019, if HighR-CHRs had delivered only LS negative flocks, the RR would have reduced from 0.94 (scenario “O”) to 0.51 (A-I). Other scenarios showed smaller RR reductions. Targeting high risk farms/flocks for intensive control could improve One Health-ness of national action plans against Campylobacter spp.

Using stochastic dynamic modelling to estimate the sensitivity of current and alternative surveillance program of Salmonella in conventional broiler production

Since 2018, the EU commission has declared the Danish broiler industry to be Salmonella free. However, there is continuous Salmonella pressure from the environment, and a number of parent flocks and broiler flocks become infected annually. When a parent flock becomes infected, the infection can be transmitted vertically to the broiler flocks, before the parent flock is detected and destroyed, including the eggs at the hatchery. To address this issue, we developed stochastic dynamic modelling of transmission of Salmonella in parent flocks and combined that with the relation between flock prevalence and test sensitivity for environmental samples in the flock. Results suggested that after 10 and 100 infected hens were seeded, the likelihood of detecting an infected parent flock within the three first weeks after the infection was strongly influenced by the taking of five boot swabs (95% CI 70–100) instead of two (95% CI 40–100) or the supplementing of the two boot swabs by a dust sample (95% CI 43–100). Results suggest that the likelihood of detecting the broiler flock as infected in the program was estimated to at least 99% in broiler flock even if only one chicken was initially infected. These findings are of relevance for managing parent flocks and eggs at the hatchery in case of Salmonella infection in parent flocks in the Danish poultry.

The association between meat inspection codes, footpad lesions and thinning of broiler flocks in the Danish broiler production

The foundation of the condemnation practices in Post-Mortem Inspection (PMI) of poultry should be based on up-to-date scientific evidence about the cause of infection and hence whether the lesions observed are of food safety, animal health or welfare concerns. This study aimed to investigate the association between meat inspection codes, footpad lesions, and thinning of flocks in Danish broiler production. The data set was based on the delivery of chicken flocks to one of the two larger chicken slaughterhouses in Denmark, representing 71 farms, 174 houses, and 4,068 flocks over three years from January 2016 to December 2018. Post-mortem condemnation data of slaughtered chickens recorded and stored in the Danish Quality Assurance System (KIK) database was used in the study. Five potentially causal models were developed to investigate whether there was an association between dermatitis, arthritis, systemic infection, emaciation, mortality and possible explaining factors` (footpad lesion, age at slaughter, scratches, ascites, systemic infection and thinning of the flock). These five ecological logistic regression models were analyzed with the three levels: farm, house, and flock. Data from a total number of 126,137,002 (N) slaughtered chickens recorded in KIK databases were used for modeling and analyses. The prevalence of condemned carcasses was 1.1 % (n = 1,420,812). Overall, 12 individual reasons for condemnation of carcasses were recorded. The most frequently observed reason for condemnation was skin disease (scratches and dermatitis) with a prevalence of 0.5 %. Prevalence of ascites was 0.2 %, discoloration 0.09 %, emaciation 0.09 %, hepatitis 0.09 % and arthritis 0.07 %. In the first model, dermatitis was shown to be positively associated with age at slaughter with an OR = 1.04 (CI95 %: 1.02−1.05), while arthritis was considered an intervening variable. Moreover, there was a small protective effect of thinning of the flock for first and second delivery. There was a positive association between arthritis and age at the time of slaughter with an OR = 1.13 (CI95 %: 1.12−1.15). Systemic infections were associated with scratches with an OR = 24.5 (CI95 %: 16.6−36.3) and footpad lesions with an OR = 1.007 (CI95 %: 1.006−1.008). Further modelling of emaciation and mortality was not considered because of unbalanced groups in the data probably caused by the fact that some condemnation codes were rare. We observed that the most common causal factors of condemnation in the systemic infection models were scratches and footpad lesion, therefore preventing and controlling such lesions could reduce losses. Specific management and environmental etiological factors of the main infections causing condemnation in Danish broilers should be determined.

Longitudinal Study on Causes of Mortality in Danish Broiler Breeders.

Broiler production is highly dependent on good health in the parent flocks. The so-called normal mortality in these flocks remains to be addressed to further reduce mortality of the breeders and to improve the quality of broilers. The aim of the present study, therefore, was to investigate the etiology of this breeder mortality to map out possible critical periods during production in relation to possible risks of importance to the offspring. Dead birds from four flocks were subjected to postmortem and bacteriologic examination from onset of lay until slaughter (20-60 weeks). Causes of mortality were divided into noninfectious and infectious etiology. The infectious group could be subdivided into suppurative salpingitis/peritonitis caused by Escherichia coli and other infections (e.g., sepsis, endocarditis, and arthritis) mainly caused by Gram-positive cocci. Data analysis showed that 41% of the birds died from noninfectious causes, while 55% died from infectious causes, and 4% had no known cause of death. The prevalence of noninfectious mortality was highest in the youngest birds and lowest in the oldest birds. In contrast, the infectious mortality was lowest in the young birds and highest at the end of production. Within each age group, the prevalence of salpingitis/ peritonitis was 26% in young birds (20-29 weeks) and progressed throughout production to 41% in the oldest birds (≥50 weeks of age). Mortality due to other infections was low at onset of production (12%), peaking at 40-49 weeks of age (25%). Consequently, 40-49 weeks of age is identified as a critical period with regard to causes of mortality, possible vertical transmission of E. coli to the offspring, and increased risk of Gram-positive coccal infections.

Characterization of Escherichia coli causing cellulitis in broilers

The aim of the study was to investigate cellulitis caused by Escherichia coli which has been responsible for economic and welfare problems in Danish broiler production between 2014 and 2016. The study included 13 flocks with unusually high condemnation rates due to cellulitis during a period of approximately one year. From six flocks, 126 condemned carcasses were collected at a Danish slaughterhouse. Further 272 broilers dead on their own were collected on nine broiler farms from flocks with increased mortality and cellulitis (2 farms included both birds from the rearing period and broilers subsequently condemned). All broilers were subjected to post mortem investigation including bacteriology and 247 E. coli isolates were obtained in pure culture from typical lesions of cellulitis. Two-hundred-thirty six E. coli isolates were investigated by pulsed field gel electrophoresis for clonality and 21 selected strains representing major clones were subsequently multi locus sequence typed allowing comparison to sequence types (ST) in the databases. One dominating PFGE type (A) was found to cause cellulitis on all 13 flocks (67% of all isolates). The clone belonged to ST117, which is well described as a pathogen in poultry, and was the primary agent responsible for cellulitis. Whole genome sequencing of eight E. coli isolates confirmed the close genetic relationship between isolates from the outbreaks and showed the presence of genes predicted to encode for the autotransporter proteins aatA, pic and upaG, reported to be of importance for adhesion of E. coli to eukaryotic cells.

Inter-rater agreement in visual assessment of footpad dermatitis in Danish broiler chickens

ABSTRACT1. The performance of the scoring in the Danish footpad dermatitis (FPD) surveillance system was evaluated by determining inter-rater agreement in visual inspection of FPD in broilers between two independent raters (R1 and R2) and the official scoring at a Danish slaughterhouse.2. FPD scores were evaluated in 1599 chicken feet. The two raters and the slaughterhouse scored equal proportions of score 0. So did R1 and R2 when assessing score 1 and the more severe lesion score 2, whereas the slaughterhouse scored a markedly higher proportion of score 1 and a lower proportion of score 2. Aggregated FPD flock scores ranged from 5 to 163 (R1 and R2) and from 8 to 107 (slaughterhouse).3. The level of agreement between the two raters was high for scores 0, 1 and 2 and for flock scores. Agreement between raters and the slaughterhouse was lower when R1 and R2 recorded score 2 than when they recorded scores 0 and 1.4. This study indicates that the occurrence and severity of lesions are underestimated in the official Danish FPD scoring system.

Risk factors for Campylobacter colonization in Danish broiler flocks, 2010 to 2011

The objectives of the two studies presented were to estimate the prevalence of Campylobacter-positive farms and flocks and to acquire updated knowledge about risk factors for the introduction of Campylobacter in Danish broiler flocks. In the first study, from September 2010 to September 2011, there were 25 Danish broiler farms visited, and a questionnaire was filled in by a veterinarian/consultant. In the second study, a similar questionnaire was distributed electronically to all Danish broiler farmers (n = 164) that were on record with an email address in the Quality Assurance System in the Danish Broiler Production (KIK) database. House- and flock-specific data collected in the surveys were supplemented with information obtained from the KIK database. Data obtained from the two studies were analyzed separately by logistic regression analysis. In both models, the dependent variable was “Campylobacter flock status (positive/negative),” which was based on real-time PCR testing of fecal material from the floor of each broiler house that had been collected preslaughter using a pair of tube gauze “socks.” This material was pooled into one sample. Of the 25 farms visited, 17 had delivered Campylobacter-positive flocks during the study period, and eight farms had no Campylobacter-positive flocks. Moreover, the flock prevalence of Campylobacter was 17% (n = 418). Data obtained from the electronically distributed survey revealed that 63% (n = 71) of the farms were Campylobacter-positive. Further, the flock prevalence of Campylobacter was 14% (n = 1,286). The multivariable models from the two sets of data identified the following statistically significant risk factors: summer vs. winter; if the previous flock in the house was positive for Campylobacter vs. if the previous flock in the house was negative; and litter delivered into the house close to the time of arrival of new chickens vs. storing litter on the farm. Furthermore, the data showed that a vertically based ventilation system (where most of the air is taken into the house through chimneys) constituted a higher risk than a horizontally based system (where the air is taken in through openings in the side walls).

Persistence of vancomycin resistance in multiple clones of Enterococcus faecium isolated from Danish broilers 15 years after the ban of avoparcin.

The occurrence and diversity of vancomycin-resistant Enterococcus faecium (VREF) were investigated in 100 Danish broiler flocks 15 years after the avoparcin ban. VREF occurred in 47 flocks at low fecal concentrations detectable only by selective enrichment. Vancomycin resistance was prevalently associated with a transferable nontypeable plasmid lineage occurring in multiple E. faecium clones. Coselection of sequence type 842 by tetracycline use only partly explained the persistence of vancomycin resistance in the absence of detectable plasmid coresistance and toxin-antitoxin systems.

Systemic cost-effectiveness analysis of food hazard reduction – Campylobacter in Danish broiler supply

An integrated microbiological–economic framework for policy support is developed to determine the cost-effectiveness of alternative intervention methods and strategies to reduce the risk of Campylobacter in broilers. Four interventions at the farm level and four interventions at the processing stage are considered. Cost analyses are conducted for different risk reduction targets and for three alternative scenarios concerning the acceptable range of interventions. Results demonstrate that using a system-wide policy approach to risk reduction can be more cost-effective than a policy focusing purely on farm-level interventions. Allowing for chemical decontamination methods may enhance cost-effectiveness of intervention strategies further.

Combined steam and ultrasound treatment of broilers at slaughter: A promising intervention to significantly reduce numbers of naturally occurring campylobacters on carcasses

Steam or hot water decontamination treatment of broiler carcasses is hampered by process limitations due to prolonged treatment times and adverse changes to the epidermis. In this study, a combination of steam with ultrasound (SonoSteam®) was investigated on naturally contaminated broilers that were processed at conventional slaughter speeds of 8,500 birds per hour in a Danish broiler plant. Industrial-scale SonoSteam equipment was installed in the evisceration room, before the inside/outside carcass washer. The SonoSteam treatment was evaluated in two separate trials performed on two different dates. Numbers of naturally occurring Campylobacter spp. and TVC were determined from paired samples of skin excised from opposite sides of the breast of the same carcass, before and after treatments. Sampling was performed at two different points on the line: i) before and after the SonoSteam treatment and ii) before the SonoSteam treatment and after 80min of air chilling. A total of 44 carcasses were examined in the two trials. Results from the first trial showed that the mean initial Campylobacter contamination level of 2.35 log10 CFU was significantly reduced (n=12, p<0.001) to 1.40 log10 CFU after treatment. A significant reduction (n=11, p<0.001) was also observed with samples analyzed before SonoSteam treatment (2.64 log10 CFU) and after air chilling (1.44 log10 CFU).In the second trial, significant reductions (n=10, p<0.05) were obtained for carcasses analyzed before (mean level of 2.23 log10 CFU) and after the treatment (mean level of 1.36 log10 CFU). Significant reductions (n=11, p<0.01) were also found for Campylobacter numbers analyzed before the SonoSteam treatment (2.02 log10 CFU) and after the air chilling treatment (1.37 log10 CFU). The effect of air chilling without SonoSteam treatment was determined using 12 carcasses pre- and postchill. Results showed insignificant reductions of 0.09 log10 from a mean initial level of 2.19 log10 CFU.Numbers of TVC before treatments ranged between 3.47 and 4.79 log10 CFU. In all cases, TVC was significantly (p<0.001, n=45 in each trial) reduced by approximately 0.7 log10 CFU.An authorized sensory panel at the Danish Veterinary and Food Administration concluded that broiler carcasses treated with SonoSteam were acceptable for purchase. These conclusions were based on organoleptic differences (smell, skin/meat consistency, texture and color) of treated vs. untreated carcasses.Results obtained from this study suggest that steam–ultrasound treatment of carcasses at broiler processing plants can significantly reduce numbers of Campylobacter on naturally contaminated broilers.

Multilocus sequence typing and biocide tolerance of Arcobacter butzleri from Danish broiler carcasses

BackgroundArcobacter spp. have in recent years received increasing interest as potential emerging enteropathogens and zoonotic agents. They are associated with various animals including poultry and can be isolated from meat products. The possibilities of persistence and cross-contamination in slaughterhouses during meat processing are not well established. We have evaluated the occurrence and persistence of Arcobacter spp. in a Danish slaughterhouse and determined the sensitivity of isolates to sodium hypochlorite, a commonly used biocide.ResultsArcobacter contamination was examined in a broiler slaughterhouse by selective enrichment of 235 swabs from the processing line during two production days and after sanitizing in between. In total 13.6% of samples were positive for A. butzleri with the majority (29 of 32 isolates) originating from the evisceration machine. No Arcobacter spp. was isolated after cleaning. A. butzleri isolates confirmed by PCR were typed by multilocus sequence typing (MLST) resulting in 10 new sequence types (STs). Two sequence types were isolated on both processing days. Minimum inhibitory concentration (MIC) to sodium hypochlorite was determined to 0.5% hypochlorite biocide (500 ppm chlorine) for most isolates, which allows growth of A. butzleri within the working concentration of the biocide (0.2 - 0.5%).ConclusionsA. butzleri was readily isolated from a Danish broiler slaughterhouse, primarily in the evisceration machine. Typing by MLST showed high strain variability but the recurrence of two STs indicate that some persistence or cross-contamination takes place. Importantly, the isolates tolerated sodium hypochlorite, a biocide commonly employed in slaughterhouse sanitizing, at levels close to the disinfection concentration, and thus, A. butzleri may survive the disinfection process although this was not observed in our study.

Analysis of factors important for the occurrence of Campylobacter in Danish broiler flocks

For more than a decade human campylobacteriosis has been the leading zoonosis in many developed countries. Consumption of poultry or poultry products has been identified as the primary source of infection in humans. This study was conducted to identify risk factors for the occurrence of Campylobacter in Danish broiler flocks. The study was based on a large data set consisting of Campylobacter positive status for nearly 6000 broiler flocks and 43 explanatory variables. Data were obtained from the Danish Campylobacter surveillance programme in poultry and from the responses to a standardized questionnaire answered via interviews with broiler farm owners. Two hundred and forty broiler farms, comprising 539 broiler houses, were included in the study and their Campylobacter status was followed over a 2-year period (1999–2000). The large number of observations made it possible to carry out a multivariate analysis including all 43 variables. A multivariate analysis was conducted using a generalized linear model, and the correlations between the houses from the same farms were accounted for by adding a variance structure to the model. The procedures for analyses included backward elimination, forward selection and expanding of the number of observations used in the variance analysis along with the reduction of the number of parameters in the model. The unit of analysis was ‘broiler house’, meaning that all results from a broiler house were aggregated into one prevalence figure (number of positive flocks/total number of flocks delivered over the 2-year period).The following factors were found to be significantly associated with the occurrence of Campylobacter in the broiler flocks: old broiler houses, late introduction of whole wheat in the feed, relatively high broiler age at slaughter, improper rodent control, large number of chimneys on the broiler house, farm located in an area with a high density of cattle farms, having more than one broiler house on the farm, and improper storage of wheat. This large-scale study confirms several risk factors identified in previous studies. The results concerning chimneys may be explained by the easier access that flies have to the broiler houses, which seems in agreement with recent Danish studies on the significance of fly-screens to reduce Campylobacter in broiler flocks. The results of this study may be used in identification of effective interventions aimed at controlling Campylobacter in Danish broiler flocks.

Temporal changes and risk factors for foot-pad dermatitis in Danish broilers

Foot-pad dermatitis is a major welfare concern of broilers caused by ammonia irritation from the bedding material. In Denmark, an action plan to control the condition was implemented in 2002 with monitoring through a foot scoring system at slaughter and with predefined limits that trigger sanctions. The objective of the present study was to study time trends and to identify predisposing factors on the flock lesion scores. The analysis was carried out on a database created by merging abattoir lesion data with antemortem evaluation data, and the flock productivity database managed by the farmers' association. The database had a record for each flock and variables containing information on both flock foot-pad scores and a range of management factors. We observed a dramatic decline in flock lesion scores between the years 2002 and 2005 followed by a minimal decline hereafter. Mean flock lesion scores differed between abattoirs, and subsequent analysis was performed in a mixed effect model where abattoir was considered a random effect. The analysis showed that flock lesion scores increased when the litter quality was evaluated as poor during the on-site antemortem evaluation. Other significant risk factors were winter season as opposed to summer, low daily weight gain, straw as bedding material in contrast to wood shavings and sphagnum peat, and high age at slaughter. Stocking density was only weakly associated with flock lesion scores.