Status quo: Levels of Campylobacter spp. and hygiene indicators in German slaughterhouses for broiler and turkey.

Poultry is a common reservoir for Campylobacter and a main source for human campylobacteriosis. With broiler being the predominant poultry for food production, most food safety related research is conducted for this species, for turkey, few studies are available. Although animals are typically colonized at the farm level, the slaughtering process is considered an important factor in re- and cross-contamination. We examined the development of Campylobacter, E. coli and total colony counts (TCC) after several processing steps in three broiler and one turkey slaughterhouses. Whole carcass rinsing and neck skin sampling was applied for broilers resulting in 486 samples in total, while 126 neck skin samples were collected for turkeys. A decrease in the loads of the different bacterial groups along the broiler slaughtering process was observed. Campylobacter mean counts dropped from 4.5 ± 1.7 log10 CFU/ml after killing to 1.6 ± 0.4 log10 CFU/ml after chilling. However, an increase in Campylobacter counts was evident after evisceration before the values again decreased by the final processing step. Although the Campylobacter prevalence in the turkey samples showed a similar development, the bacterial loads were much lower with 1.7 ± 0.3 log10 CFU/g after killing and 1.7 ± 0.4 log10 CFU/g after chilling compared to those of broilers. The loads of E. coli and total colony count of turkey were higher after killing, were reduced by scalding and remained stable until after chilling. This study highlights trends during the slaughtering process in reducing the levels of Campylobacter, E. coli, and total colony counts for broiler and turkey carcasses, from the initial step to after chilling. These results contribute to our understanding of microbial dynamics during meat processing.

Growth of methicillin-resistant Staphylococcus aureus during raw milk soft cheese-production and the inhibitory effect of starter cultures.

The consumption of raw milk or raw milk products might be a potential risk factor for the transmission of methicillin-resistant Staphylococcus aureus (MRSA). Therefore, we studied MRSA growth during raw milk soft cheese-production. Furthermore, we investigated the inhibitory effect of four starter cultures (Lactococcus lactis, Lacticaseibacillus rhamnosus, Lactiplantibacillus plantarum, Lactobacillus helveticus) on the growth of MRSA in a spot-agar-assay and in raw milk co-culture following a cheesemaking temperature profile. During the initial phases of raw milk cheese-production, MRSA counts increased by 2 log units. In the ripening phase, MRSA counts only dropped slightly and remained high up to the end of the storage. Comparable MRSA counts were found in the rind and core and strain-specific differences in survival were observed. In the spot-agar-assay, all four starter cultures showed strong or intermediate inhibition of MRSA growth. In contrast, in raw milk, only Lactococcus lactis strongly inhibited MRSA, whereas all other starter cultures only had minor inhibitory effects on MRSA growth. Our results indicate that MRSA follow a similar growth pattern as described for other S. aureus during raw milk soft cheese-production and illustrate the potential use of appropriate starter cultures to inhibit MRSA growth during the production of raw milk cheese.

Quantitative assessment of Campylobacter spp. levels with real-time PCR methods at different stages of the broiler food chain.

The importance of thermotolerant Campylobacter spp. as a food-borne pathogen continues to increase and there is a great need for rapid quantitative results in routine diagnostics. However, currently, only the culture-based ISO method is authorized for use in the context of official food control. The present study therefore aimed to assess the suitability of a qPCR method for a rapid quantitative determination of Campylobacter spp. at different stages in the poultry production chain and its equivalence with the culture-based method. Samples from two processors were collected and evaluated both separately and together. Censored regression (Tobit) models have been used to establish a relationship between Campylobacter qPCR counts on the carcasses and explanatory variables of processor and meat counts. Further, correlations of qPCR Campylobacter spp. counts at the different stages of production were calculated. In addition, the comparative data between microbiological enumeration and qPCR results were statistically analyzed. In the correlation calculation of the qPCR results, a highly significant relationship between the Campylobacter spp. counts of the neck skin samples to breast fillet and leg samples could be calculated, indicating a good prediction of Campylobacter spp. loads in these samples. The intercalating dye ethidium monoazide (EMA) was used to see whether the correlations between microbiological counts and qPCR results were improved by pretreating fecal and cecal samples before qPCR analysis. It was shown that the observed values of scatter plots between the qPCR-based and the culture-based methods were strongly correlated. However, on average, the qPCR results were two log10 CFU/mL levels higher than the microbiological counts. However, the classical culture-based method for food hygiene risk assessment cannot be replaced one-to-one by the qPCR or EMA-qPCR. The qPCR method can rather be used for the rapid identification of particularly highly contaminated flocks.

Monitoring the effect of active packaging films with silver-kaolinite using different packaging systems on the quality of beef meat.

Active packaging films based on gelatin with silver-kaolinite (Ag-Kln) were developed and their effects on the quality and microbial growth of beef meat stored in different packaging systems (cling film, vacuum and modified atmosphere packaging) at 4 °C for 13 days were evaluated. The analysis revealed that Ag-Kln packaging films had no adverse effects on the pH and colour of the beef samples irrespective of the packaging system used. Beef meat in packaging with active films lost more weight (P < 0.05) than without active films for meat in the vacuum and modified systems on day 13. In general, these gelatin films with Ag-Kln showed the potential as antibacterial films and could enhance the shelf life of food products, however, further studies are required to establish the release rate of silver from packaging films, as well as test the efficiency of these materials under different storage conditions. In conclusion, this study revealed that gelatin film with silver-kaolinite is a promising antibacterial agent and preservation material for food shelf life extension.

Suitability of lactic acid bacteria and deriving antibacterial preparations to enhance shelf-life and consumer safety of emulsion type sausages.

Ready-to-eat (RTE) sliced emulsion type sausages are sensitive to recontamination with Listeria (L.) monocytogenes during processing and packaging steps. Since Listeria spp. are able to grow on those products under cold storage conditions, taking steps to reduce the recontamination risk and implementing antibacterial hurdles contribute to consumer safety and increase the product quality. With this study data about the suitability of culture broth, cell-free supernatant (CFS) or concentrated bacteriocin preparations (CFSconc) of bacteriocin-producing lactic acid bacteria (LAB) obtained from fermented sausages from Germany as protective culture or antibacterial additive were provided. In different challenge tests, the potential of selected LAB or their preparations were investigated for their potential to reduce growth of L. monocytogenes and/or Brochothrix (B.) thermosphacta on sliced RTE emulsion type sausages under modified atmosphere or vacuum during refrigerated storage for a 21-day period. Applied LAB culture broth and CFS could not reduce the growth of L. monocytogenes or B. thermosphacta. On the other hand, samples treated with CFSconc obtained from Pediococcus spp. strains showed a significant inhibition (p < 0.05) of more than 1.5 log10 of the applied L. monocytogenes strains during the storage period. The growth of B. thermosphacta could not be influenced. Thereby, the need for concentrating preparations was shown to be important to obtain a suitable antibacterial preparation that would contribute to consumer safety and food quality when applied as a protective additive.

Management factors influencing the occurrence of cellulitis in broiler chickens.

One of the main reasons for condemning fattening broiler chickens during meat inspection is cellulitis, which demonstrates the great economic issue concerning this topic. The aim of this epidemiological study was therefore to identify risk factors in order to draw conclusions on how to prevent the occurrence of cellulitis in broilers by implementing management changes. The data were collected between April and November 2018 on conventional broiler farms (n = 100) in the north of Germany with one to fourteen poultry houses per farm. In total, data were collected from 199 broiler flocks with a total of 5,332,767 broilers. Data on the type of management (feeding- and drinking management, housing, lighting management, litter type and animal health) were collected via a questionnaire, with additional data on condemnation rates being provided by the abattoirs. It was found that litter additives like fennel, eucalyptus and probiotics as well as a moist litter quality were associated with lower cellulitis condemnation rates. Flocks fattened in windowless barns, but with relatively higher lux-values as well as those broilers examined in a lower number of housing inspections had significantly lower cellulitis condemnation rates compared to other husbandry systems. In addition, lower cellulitis rates were seen when housing capacities were smaller, regardless of stocking density. The source of the breeders and hatchery also had a significant influence on the occurrence of cellulitis. No correlation was found between the condemnation rates due to cellulitis and the performance of thinning, the water source used, the use of drinking additives, observational skills and number of herd managers monitoring the broilers, participation in an animal welfare programme, the technique of heating and ventilation systems used, the feed supplier, litter material, the broiler breed, the length of darkness periods and chick losses during the first seven days. We concluded that management decisions that lead to stress reduction in the broiler flocks are beneficial in terms of chicken welfare and occurrence of cellulitis.

Characterisation of mecA gene negative Staphylococcus aureus isolated from bovine mastitis milk from Northern Germany.

Staphylococcus aureus (S. aureus) is an important causative agent of contagious intermammary infections in dairy cattle. S. aureus is also considered as an important foodborne pathogen and cause of food poisoning cases and outbreaks worldwide. In order to understand the molecular ecology of S. aureus, the present study compared phenotypic and genotypic characteristics of 70 S. aureus isolates from bovine mastitis milk samples collected during the period from August 2001 to March 2014 in different regions of Northern Germany. The S. aureus isolates were characterised phenotypically, as well as genotypically for their genetic diversity using multi-locus sequence typing (MLST), spa typing and the presence of virulence genes encoding 16 staphylococcal enterotoxins (sea-selu), toxic shock syndrome toxin (tst), thermonuclease (nuc), clumping factor (clfA and clfB), coagulase (coa) and the methicillin resistance gene mecA. A total of 16 sequence types were grouped into eight clonal complexes (CCs), and 17 spa types were identified. These included six novel sequence types and one novel spa type. The majority of bovine mastitis milk-associated sequence types belonged to the clonal complex CC5, CC97, CC133, and CC151 and showed closely related genotypes or lineages with sequence types of human origin. The genotype CC133 (ST133-t1403) was predominant, constituting 27.1% of the isolates. In addition, the S. aureus isolates displayed nine different enterotoxigenic profiles. All S. aureus were methicillin-susceptible (MSSA). The current study provides new information on phenotypic and genotypic traits of S. aureus isolates from bovine mastitis. The comparison of characteristics of isolates from the present study originating from mastitis milk showed similarities with human isolates. This might help to better understand the distribution of S. aureus in the one health context.

Genome Sequences of Malonate-Positive Cronobacter sakazakii Serogroup O:2, Sequence Type 64 Strains CDC 1121-73 and GK1025, Isolated from Human Bronchial Wash and a Powdered Infant Formula Manufacturing Plant.

We introduce draft genome sequences of strains CDC1121-73 (human bronchial wash isolate) and GK1025 (powdered infant formula manufacturing facility isolate), which are both malonate-positive Cronobacter sakazakii serogroup O:2, sequence type 64. Assemblies for these strains have sizes of 4,442,307 and 4,599,266 bp and % G+C contents of 56.9 and 56.7, respectively.

Influence of process parameter on Campylobacter spp. counts on poultry meat in a slaughterhouse environment.

Campylobacter spp. are the most important food-borne pathogens in broilers. Exposure of the consumer can be influenced by the reduction of contaminated broiler meat at various steps along the production line. This study was performed at a poultry slaughterhouse in Germany. Steps within the slaughter process were defined by the slaughterhouse quality control for potential Campylobacter reduction. Their impact was tested for two process variations. The first process variation was the increase of the temperature of the scalding water from 53.0 to 53.9 °C. The second step was the application of an additional outside sprayer which was placed after plucking. The increase of the scalding water temperature was the most effective measure (>2 log reduction), but resulted in defects to the broiler skin. This would limit marketing of fresh broiler meat with skin. The additional water spray after plucking had no additional effect. In fact, numbers of Campylobacter were lower before introduction of the sprayer. In conclusion, modifications of the processing technology have to be evaluated carefully, but can have additional effects for Campylobacter reduction.

Large-scale feasibility of organic acids as a permanent preharvest intervention in drinking water of broilers and their effect on foodborne Campylobacter spp. before processing.

AIMS: Evaluating the effect of a commercially available organic acid water additive in conventional broiler production on Campylobacter spp. METHODS AND RESULTS: The organic acid water additive was added to the drinking water from chick housing to catching in three consecutive rearing cycles. The broiler performance data were evaluated, and the load of thermophilic Campylobacter spp. was analysed in water, feed and the environment as well as determined in caecum content and on carcasses at the abattoir according to ISO 10272:1.2-2002. The results indicated that permanent application of acidified drinking water did not have detrimental effects on production parameters or animal welfare. The quantitative results obtained at slaughter were ambiguous, but suggested a reduced carriage of Campylobacter spp. by the flock and in caecum content. Such reduction did not result in lower Campylobacter carriage of the carcasses after slaughter. CONCLUSIONS: Organic acids in drinking water of broilers can partly reduce the caecal Campylobacter spp. load, but this did not reduce carcass contamination. SIGNIFICANCE AND IMPACT OF THE STUDY: Broiler meat is a major source of foodborne campylobacteriosis. The public health would considerably benefit from controlling Campylobacter in the food chain. The addition of organic acid to drinking water of broilers can potentially lower the caecal carriage in primary production. However, in this field trial, a commercial product failed to have an impact on the bacterial load after slaughter.

Prevalence of Cronobacter spp. in a powdered infant formula processing environment.

From November 2005 to August 2006, microbiological analyses were performed in a processing plant for powdered infant formula (PIF) to estimate the occurrence and dissemination of Cronobacter spp. and the contamination levels of Enterobacteriaceae in the processing environment. In total 867 samples were collected from the dry processing environment including powders from vacuum cleaners and filtering (sieving) machines, fluids from the drains and swabs from contact surfaces. In total 35 samples in 14 out of 35 tested locations were positive for Cronobacter spp. Positive samples were found in environmental powders (94.3%) and two fluids (5.7%). The Cronobacter spp. prevalence was highest in powders from vacuum cleaners with 28.0% positive samples on a single location. While powders from the sensitive agglomeration section or production (sieving) had contamination rates of 10.7% or below 3.4%, respectively. In addition vacuum powder samples from the filling and can filling line were positive with 5.3 and 8.0%, respectively. High levels (>500 cfu/g) of Enterobacteriaceae were found in seven locations out of 21 tested powder locations. This fact emphasizes the importance of monitoring of the processing environment to implement effective measures to improve the safety and hygiene in PIF, a very sensitive product.

Microbiological quality of freshly shot game in Germany.

In the framework of a project on the hygiene status of freshly shot game 289 samples were microbiologically analysed: 127 samples from wild boars, 95 from roe deer and 67 from red deer. The microbiological parameters evaluated were the mesophilic aerobic count (APC), which showed mean log10-counts of 2.6cfu/cm(2) for roe deer, 2.9cfu/cm(2) for red deer and 3.2cfu/cm(2) for wild boars and the numbers of Enterobacteriaceae, which gave mean log10-values of 2.1cfu/cm(2) for all three species with differing ranges. The concentrations of coagulase positive staphylococci were >2.0 log10cfu/cm(2) between 3.2 and 6.3%, according to species. Listeria was found in 14 samples and three samples gave a positive result for Campylobacter. Salmonella was not found in any of the samples analysed.

Chemokine expression dynamics in mycobacterial (type-1) and schistosomal (type-2) antigen-elicited pulmonary granuloma formation.

Transcript expression of 24 chemokines (CKs) was examined throughout 8 days in mouse lungs with type-1 (Th1) or type-2 (Th2) cytokine-mediated granulomas induced by bead-immobilized mycobacterial purified protein derivative or Schistosoma mansoni egg antigens. Where possible, CK protein levels were also measured. In addition, we examined effects of in vivo cytokine depletions. Findings were as follows: 1) bead challenge induced increases in 18 of 24 CK transcripts with type-1 and type-2 responses displaying different patterns. CKs fell into four categories: a) type-1-dominant (gamma-interferon-inducible protein (IP-10), monokine induced by INF-gamma (MIG), macrophage inflammatory protein-2 (MIP-2), lipopolysaccharide-induced chemokine (LIX), rodent growth-related oncogene homologue (KP), macrophage inflammatory protein-1alpha (MIP-1alpha) and -1beta (MIP-1beta), lymphotactin), b) type-2-dominant (eotaxin, monocyte chemotactic protein-2 (MCP-2) and -3 (MCP-3), liver and activation-regulated chemokine (LARC), T cell activation protein-3 (TCA-3), c) type-1 and type-2 co-dominant (MCP-1, MCP-5, monocyte-derived chemokine (MDC), thymus and activation-related chemokine (TARC), C10), and d) constitutive (lungkine, secondary lymphoid-tissue chemokine (SLC), EBI1-ligand chemokine (ELC), fractalkine, macrophage inflammatory protein-1gamma (MIP1-gamma), and stromal cell derived factor-1alpha (SDF1-alpha). 2) CKs displayed characteristic temporal patterns. CXC (IP-10, MIG, MIP-2, LIX, KC) and certain CC (MCP-1, MCP-5, MIP-1alpha, MIP-1beta) CKs were produced maximally within 1 to 2 days. Others (MCP-2, MCP-3, eotaxin, lymphotactin, LARC, TCA-3) displayed peak expression later. 3) Interferon-gamma neutralization profoundly abrogated MIG, but had little effect on other CKs. Tumor necrosis factor-alpha neutralization caused up to 50% reduction in a range of CKs. These findings indicate that type-1 and type-2 granulomas display characteristic CK profiles with coordinated expression that is under cytokine-mediated regulation.

IL-13 transgene state impairs mycobacterial (type-1) and schistosomal (type-2) antigen-elicited responses.

Transgenic technology provides one approach for examining cytokine properties in vivo. This study directly tested the effect of a lung-targeted IL-13 transgene on the induction and elicitation of Th1 and Th2 cell-mediated immuno-inflammatory responses. Induction of Th1 (type 1) and Th2 (type 2) responses were tested by sensitization of IL-13 transgenics and littermates with purified protein derivative (PPD) of Mycobacterium bovis or Schistosoma mansoni eggs. Secondary elicitation of pulmonary granulomas was examined in adoptively sensitized transgenics and littermates challenged with bead-bound PPD or S. mansoni egg antigens. Parameters included lymphoid tissue cytokine profiles and granuloma sizes. Results showed that induction and elicitation of both type 1 and type 2 cytokines and granulomas were significantly abrogated in transgenics. Systemic effects were possible, as transgenic serum contained high levels of circulating IL-13. These findings support the concept that IL-13 impairs effector functions and provide novel information regarding its role in regulating Th2 cytokines.

Mitral valve prolapse in patients with coronary artery disease. Echocardiographic-angiographic correlation.

Echocardiography was performed in 25 consecutive patients with angina pectoris and angiographically demonstrable coronary artery disease. Left ventricular echograms detected late or pansystolic mitral valve bowing suggesting of mitral valve proplapse in 6/25 (24%). Left ventricular angiography showed prolapse of the posterior mitral leaflet in 15/25 (60%), including 5 detected by echocardiography. Significant triple vessel coronary disease was present in 11 of 15 patients with prolapsed mitralvalve. In each of the latter a greater than 90 per cent obstructive lesion was noted in at least one coronary artery: right coronary artery, 9 subjects (82%); left circumflex coronary artery, 5 patients (33%); and left anterior descending coronary artery, 4 patients (27%). Of 15 subjects with angiographic evidence of mitral valve prolapse, 13 had left ventricular asynergy-inferior or inferoposterior in 8 subjects (62%) and anterior or anteroapical in 5 subjects (38%). Eleven subjects had vectorcardiographic evidence of transmural myocardial infarction-inferior or inferoposterior in 9 (82%) and anteroseptal in 2 (18%). A single subject with mitral valve prolapse had mild mitral regurgitation. It is concluded that: (1) coexisting prolapse of the posterior mitral valve leaflet and coronary artery disease is usually associated with triple vessel obstructive lesions, (2) severe right coronary disease, inferior left ventricular wall asynergy, and inferior myocardial infarction are important angiographic and vectorcardiographic correlates, and (3) echocardiography will detect such mitral valve prolapse in only one-third of affected cases.

Correlation of electrocardiogram and vectorcardiogram with coronary occlusion and myocardial contraction abnormality.

Electrocardiograms and Frank vectorcardiograms were recorded in 156 consecutive patients with total occlusion of at least one coronary artery (on arteriography) and associated left ventricular contraction abnormality (on ventriculography). The angiograms and cardiograms were independently reviewed. In the presence of single vessel occlusion, appropriate vectorcardiographic diagnosis of myocardial infarction was determined in 118 of 156 cases (76 percent) compared with a lower electrocardiographic detection rate in 77 of 156 cases (49 percent). Findings diagnostic of two coexisting infarctions were observed in 71 percent of vectorcardiograms and 37 percent of electrocardiograms in 51 patients with double vessel occlusion and two areas of left ventricular dyskinesia. The vectorcardiographic detection rate was similarly superior to the electrocardiographic rate in the presence of subtotal coronary occlusion and myocardial asynergy in single (73 percent versus 53 percent) and double (53 percent versus 28 percent) vessel disease. The incidence rate of false positive diagnoses was 3 percent for electrocardiography and 4 percent for vectorcardiography. It is concluded that the vectorcardiogram is superior to the electrocardiogram in the diagnosis of obstructive coronary artery disease and left ventricular contraction abnormality.

Comparison of the electrocardiogram and vectorcardiogram for the diagnosis of left atrial enlargement.

Standard 12 lead electrocardiograms (ECG) and Frank vectorcardiograms (VCG) were recorded in 21 consecutive patients with mitral valvular disease and angiographically documented left atrial enlargement. Comparative sensitivities for the detection of left atrial enlargement were: diagnostic, ECG = 6/21 (29%), VCG = 14/21 (67%); suggestive, ECG = 3/21 (14%), VCG = 2/21 (9%); non-diagnostic, ECG = 12/21 (57%), VCG = 5/21 (24%). It is concluded that the Frank atrial vectorcardiogram is superior to the standard electrocardiogram for the diagnosis of left atrial enlargement.