Diversity of Staphylococcus aureus isolated from nares of ruminants.

AIMS: To examine the diversity of Staphylococcus aureus isolated from nasal swabs of ruminants in Rwanda. METHODS AND RESULTS: A total of 454 nasal swabs from 203 cows, 170 goats, and 81 sheep were examined for the presence of S. aureus, and 30 S. aureus isolates were detected and characterized pheno- and genotypically. Resistance to penicillin and/or tetracycline was observed. The isolates were assigned to eight different spa types (t21057 (novel), t10103, t18853, t20842, t318, t355, t458, and t9432) belonging to six clonal complexes (CCs) (CC152, CC30, CC3591, CC3666, CC522, and CC97). Panton-Valentine leukocidin (PVL) genes (lukF-PV/lukS-PV), the bovine leukocidin genes (lukM/lukF-P83), and the human and bovine variants of the toxic shock syndrome toxin gene tst-1 variants were detected. CONCLUSION: These findings demonstrate that the nares of ruminants in Rwanda are colonized with mastitis-associated S. aureus, including lineages that are also carried by humans, underscoring the zoonotic risk, especially for livestock keepers. These results highlight the crucial importance of hygiene measures when handling livestock.

Comparison of the population structure of Streptococcus uberis mastitis isolates from Austrian small-scale dairy farms and a Slovakian large-scale farm.

Streptococcus uberis, a major mastitis pathogen associated with intramammary infections (IMI), can be found ubiquitously in the cow's environment. Although Strep. uberis is reported to be susceptible to most antimicrobials, in practice poor responses to treatment and recurrent mastitis are observed. This can be explained by reinfection or by persistence of strains. We hypothesized that among a heterogeneous group of Strep. uberis mastitis isolates, some predominant host-adapted clones might be recurrently isolated from IMI. Therefore, the aim of this pilot study was to determine the Strep. uberis genotype variety found among small-scale dairy herds (127 Austrian dairy farms) and compare this with a large-scale herd (a Slovakian dairy farm). We determined the occurrence and strain diversity of Strep. uberis (n = 309) isolates using molecular analysis. Streptococcus uberis isolates from aseptically collected quarter milk samples were genotypically characterized using pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing. The Strep. uberis strain set covered isolates from 4 Austrian federal areas [Lower Austria (n = 67), Upper Austria (n = 8), Salzburg (n = 51), and Styria (n = 1)] and the Bratislava Region of Slovakia (n = 1). The PFGE analysis resulted in 187 SmaI profiles with 151 unique profiles. Simpson's index of diversity was 0.988. Individual cows (n = 17) harbored up to 3 different PFGE types in the udder. Dairy cows shared distinct PFGE types within a farm. Seven PFGE types were widely distributed among Austrian dairy farms. In the Slovakian farm, 10 predominant PFGE types were recurrently isolated from the same quarters; these genotypes were assigned as persisters. We identified novel sequence types (ST) using multilocus sequence typing related to the global clonal complexes ST5 and ST143. We concluded that Strep. uberis IMI are caused by strains with a wide heterogeneity of PFGE types. This large number of unique subtypes indicates a high diversity of Strep. uberis in the environment. In the large herd, molecular epidemiological results revealed that specific strains might be involved in contagious transmission events and potentially lead to persistence.

Escherichia coli isolates from femoral bone marrow of broilers exhibit diverse pheno- and genotypic characteristics that do not correlate with macroscopic lesions of bacterial chondronecrosis with osteomyelitis.

The pheno- and genotypic relatedness among Escherichia coli isolates from broilers with and without macroscopic lesions of the femoral head were investigated. In total, 219 isolates obtained from the bone marrow were characterized by serotyping, antimicrobial resistance (AMR) profiles, phylogenetic grouping, detection of virulence-associated genes (VAGs) and pulsed-field gel electrophoresis (PFGE). Serotyping revealed that 48.4% of the isolates were assigned to one of the three serotypes (O78:K80: 21.0%, O2:K1: 18.7%, O1:K1: 8.7%). Substantial phenotypic variation was also noticed in AMR testing as most of the birds harboured E. coli isolates with different AMR profiles, which is of high clinical relevance. The majority of isolates could be classified into phylogenetic groups D (54.3%) and B2 (25.6%), followed by A (11.4%) and B1 (8.7%). Virulotyping showed that the highest number of isolates contained genes iucD (86.8%) and iss (84.9%), whereas papC (16.0%) and astA (12.3%) were present in least number of isolates. PFGE resulted in 58 different profiles from 200 typeable isolates. No correlation was found between specific serotypes, AMR profiles, phylogenetic groups, PFGE types or VAG profiles of E. coli and the occurrence of bacterial chondronecrosis with osteomyelitis, contradicting the hypothesis of a specific bacterial pheno- or genotype being involved in the disease.

Listeriosis in fattening pigs caused by poor quality silage - a case report.

BACKGROUND: Listeria (L.) monocytogenes as the causative agent of listeriosis in humans and different animal species, has its reservoir in the environment. It can be found in the gut and faeces of healthy pigs, but under certain circumstances it may cause clinical disease. Fatteners are usually not known to get affected by Listeria-associated septicaemia and enteritis. This case report shows, that L. monocytogenes should be part of the list of differential diagnoses, when fattening pigs suffer from haemorrhagic diarrhoea and septicaemia. CASE PRESENTATION: Here, we report of an episode of fatal listeriosis in fattening pigs in a piglet producing farm in Lower Austria, which was combined with a fattening unit with space for 450 fatteners. The mortality rate resulted in 7.8% among fattening pigs after suffering from clinical symptoms such as anorexia, bloody diarrhoea and increased body temperature. Two fattening pigs with clinical symptoms and maize silage samples were used for further diagnostics. L. monocytogenes were isolated from serosa samples of the pigs and in the corresponding fed maize silage. One animal was positively tested for Brachyspira hyodysenteriae, which may have also been involved in the development of colitis. Immunohistochemically, L. monocytogenes could be detected in high amounts in lymphatic tissue of the gut. Molecular biological characterisation of the L. monocytogenes isolates from pigs and maize silage resulted in an identical DNA-fingerprint assigned to sequence type (ST) 21. Additionally, a high content of deoxynivalenol (3000 parts per billion) was found in maize silage. Therefore, the maize silage produced under inappropriate ensilaging conditions in a silo, was most likely the source of infection. Antimicrobial therapy with amoxicillin led to a fast cure of the remaining affected fatteners. CONCLUSION: To conclude, we were able to show, that L. monocytogenes can cause clinical disease in finishing pigs, which may have been a result of immunosuppression due to high deoxynivalenol exposure. When feeding silage it is important that all ensilaging procedures occur under appropriate anaerobic conditions to guarantee suppression of listerial growth.

Diagnostic accuracy of a standardized scheme for identification of Streptococcus uberis in quarter milk samples: A comparison between conventional bacteriological examination, modified Rambach agar medium culturing, and 16S rRNA gene sequencing.

Bacteriological examination of milk samples is a prerequisite for pathogen-specific therapy and aids in limiting antimicrobial resistance. The aims of this study were to establish a standardized scheme for reliable Streptococcus uberis identification in routine diagnosis and to evaluate the accuracy of conventional tests and growing patterns of Strep. uberis on a selective medium (modified Rambach agar medium, MRAM) using 16S rRNA gene sequencing analysis as a reference method. We obtained isolates of presumptive Strep. uberis (n = 336) from quarter milk samples of dairy cows with intramammary infections and classified the isolates into 2 clusters using biochemical characterization. In cluster 1 (n = 280), cocci grew as non-hemolytic colonies, hydrolyzing esculin, carrying no Lancefield antigen (A/B/C/D/G) or Christie Atkins Munch-Petersen factor, and their growth was inhibited on an Enterococcus agar. Production of ß-d-galactosidase on MRAM was shown by 257 of the cluster 1 isolates (91.79%), and 16S rRNA gene sequencing verified 271 (96.79%) of the isolates to be Strep. uberis. In 264 isolates (94.29%), MRAM agreed with the sequencing results. In cluster 2 (n = 56), isolates showed different characteristics: 37 (66.07%) were ß-d-galactosidase-positive, and based on 16S sequencing results, 36 (64.29%) were identified correctly as Strep. uberis using biochemical methods. Identification success in this group differed significantly between routine diagnosis and MRAM application: MRAM agreed with sequencing results in 47 isolates (83.93%). To identify Strep. uberis and differentiate it from other lactic acid bacteria in routine diagnosis, we suggest using catalase reaction, hemolysis, esculin hydrolysis, and growth on enterococci agar. Isolates that show a typical biochemical profile can be identified satisfactorily with these tests. For Strep. uberis isolates with divergent patterns, application of MRAM as a follow-up test increased the diagnostic accuracy to 94.64%.

High genetic diversity among extraintestinal Escherichia coli isolates in pullets and layers revealed by a longitudinal study.

BACKGROUND: Various information about the genetic diversity of Escherichia coli isolates from chickens are available but a detailed epidemiological investigation based upon isolates obtained from interrelated pullet and layer flocks is still missing. Therefore, in the course of a longitudinal epidemiological study on pullets and layers, 144 E. coli isolates from chickens with or without pathological lesions of the reproductive tract were serotyped and genotyped with pulsed-field gel electrophoresis (PFGE). These isolates were collected during rearing, peak and at the end of production. The actual study is the first of its kind so as to elucidate genetic relatedness among extraintestinal E. coli isolated from chickens with varying pathological conditions in interrelated layer farms/flocks at different stages of rearing. RESULTS: Serotyping revealed that 63.19 % of the isolates could not be assigned to any of the three serotypes tested whereas 30.55 % of the isolates belonged to serotype O1:K1, 4.86 % to O2:K1 and 1.38 % to O78:K80. After macrorestriction digest with XbaI, 91.66 % of the isolates were typeable resulting in 96 distinct PFGE profiles. Among them, five PFGE types included isolates collected from diseased chickens as well as from birds without pathological lesions. This finding shows that pathogenicity of E. coli in layers seems to be largely influenced by concurrent susceptibility factors. Furthermore, in six out of eight cases where two isolates were collected from each of eight birds, different PFGE types were found in the same or different organs of the same bird. The existence of predominant or persistent E. coli genotypes was only observed in two cases. CONCLUSIONS: It is concluded that extraintestinal E. coli genotypes and serotypes in pullets and layers are heterogenous and also do not maintain a single clonality within the same bird. The facts that E. coli strains did not show any definite clonal population structure based on geographical region, age of the host and pathological lesions should have relevance in further epidemiological studies and control strategies.

Tracking Foodborne Pathogenic Bacteria in Raw and Ready-to-Eat Food Illegally Sold at the Eastern EU Border.

Food illegally brought into the European Union, mainly in the personal luggage of travelers, represents a potential threat to consumers' health. The aim of this study was to investigate the presence of five pathogens in food brought into the European Union by Moldavian citizens as personal goods and illegally sold in Romania in the vicinity of the border. The occurrence of Staphylococcus aureus and Listeria monocytogenes was 7.5% and 8%, while Campylobacter spp., Escherichia coli O157:H7, and Salmonella spp. were absent in all samples. L. monocytogenes sequence type 2, 9, 121, and 155, highly prevalent among foodstuffs worldwide, was also present among isolates from ready-to-eat food illegally sold in Romania, even at the same date of sampling, indicating cross-contamination during food handling. S. aureus spa types t449, t304, and t524 were most often isolated from raw-milk cheeses contaminated with 10(3)-10(5) colony-forming units per gram, evidencing a contamination at herd level or unhygienic conditions during processing. S. aureus t011 and t3625, both included in the livestock-associated CC398, were isolated from pork lard and poultry meat. This study shows that cross-border trade from nonmember states represents a neglected route of transmission of foodborne pathogens into the European Union that could lead to sporadic or family-associated cases of disease.

Reservoirs of listeria species in three environmental ecosystems.

Soil and water are suggested to represent pivotal niches for the transmission of Listeria monocytogenes to plant material, animals, and the food chain. In the present study, 467 soil and 68 water samples were collected in 12 distinct geological and ecological sites in Austria from 2007 to 2009. Listeria was present in 30% and 26% of the investigated soil and water samples, respectively. Generally, the most dominant species in soil and water samples were Listeria seeligeri, L. innocua, and L. ivanovii. The human- and animal-pathogenic L. monocytogenes was isolated exclusively from 6% soil samples in regions A (mountainous region) and B (meadow). Distinct ecological preferences were observed for L. seeligeri and L. ivanovii, which were more often isolated from wildlife reserve region C (Lake Neusiedl) and from sites in proximity to wild and domestic ruminants (region A). The higher L. monocytogenes detection and antibiotic resistance rates in regions A and B could be explained by the proximity to agricultural land and urban environment. L. monocytogenes multilocus sequence typing corroborated this evidence since sequence type 37 (ST37), ST91, ST101, and ST517 were repeatedly isolated from regions A and B over several months. A higher L. monocytogenes detection and strain variability was observed during flooding of the river Schwarza (region A) and Danube (region B) in September 2007, indicating dispersion via watercourses.

Collaborative survey on the colonization of different types of cheese-processing facilities with Listeria monocytogenes.

Cross-contamination via equipment and the food-processing environment has been implicated as the main cause of Listeria monocytogenes transmission. The aim of this study, therefore, was to determine the occurrence and potential persistence of L. monocytogenes in 19 European cheese-processing facilities. A sampling approach in 2007-2008 included, respectively, 11 and two industrial cheese producers in Austria and the Czech Republic, as well as six Irish on-farm cheese producers. From some of the producers, isolates were available from sampling before 2007. All isolates from both periods were included in a strain collection consisting of 226 L. monocytogenes isolates, which were then typed by serotyping and pulsed-field gel electrophoresis (PFGE). In addition, metabolic fingerprints from a subset of isolates were obtained by means of Fourier-transform infrared (FTIR) spectroscopy. PFGE typing showed that six processing environments were colonized with seven persistent PFGE types of L. monocytogenes. Multilocus sequence typing undertaken on representatives of the seven persisting PFGE types grouped them into distinct clades on the basis of country and origin; however, two persistent strains from an Austrian and an Irish food processor were shown to be clonal. It was concluded that despite the fact that elaborate Hazard Analysis and Critical Control Point concepts and cleaning programs are applied, persistent occurrence of L. monocytogenes can take place during cheese making. L. monocytogenes sanitation programs could be strengthened by including rapid analytical tools, such as FTIR, which allow prescreening of potentially persistent L. monocytogenes contaminants.

Staphylococcus aureus reservoirs during traditional Austrian raw milk cheese production.

Sampling approaches following the dairy chain, including microbiological hygiene status of critical processing steps and physicochemical parameters, contribute to our understanding of how Staphylococcus aureus contamination risks can be minimised. Such a sampling approach was adopted in this study, together with rapid culture-independent quantification of Staph. aureus to supplement standard microbiological methods. A regional cheese production chain, involving 18 farms, was sampled on two separate occasions. Overall, 51·4% of bulk milk samples were found to be Staph. aureus positive, most of them (34·3%) at the limit of culture-based detection. Staph. aureus positive samples >100 cfu/ml were recorded in 17·1% of bulk milk samples collected mainly during the sampling in November. A higher number of Staph. aureus positive bulk milk samples (94·3%) were detected after applying the culture-independent approach. A concentration effect of Staph. aureus was observed during curd processing. Staph. aureus were not consistently detectable with cultural methods during the late ripening phase, but >100 Staph. aureus cell equivalents (CE)/ml or g were quantifiable by the culture-independent approach until the end of ripening. Enterotoxin gene PCR and pulsed-field gel electrophoresis (PFGE) typing provided evidence that livestock adapted strains of Staph. aureus mostly dominate the post processing level and substantiates the belief that animal hygiene plays a pivotal role in minimising the risk of Staph. aureus associated contamination in cheese making. Therefore, the actual data strongly support the need for additional sampling activities and recording of physicochemical parameters during semi-hard cheese-making and cheese ripening, to estimate the risk of Staph. aureus contamination before consumption.

Diversity of Staphylococcus aureus associated with mastitis from dairy cows in Rwanda.

OBJECTIVES: The objective of the present study was to examine the diversity of Staphylococcus aureus from mastitis milk samples of cows in Rwanda. METHODS: A total of 1080 quarter milk samples from 279 dairy cows were collected in 80 different farms from all five provinces of Rwanda. In total, 135 S. aureus isolates were obtained and subjected to genotyping (spa typing, DNA microarray, whole-genome sequencing (WGS)), antimicrobial susceptibility testing (AST) and phenotypic profiling by Fourier Transform Infrared (FTIR) spectroscopy (including capsular serotyping). RESULTS: Resistance to penicillin and/or tetracycline was most frequently observed. Ten sequence types (STs) (ST1, ST151, ST152, ST5477, ST700, ST7110, ST7983, ST7984, ST8320, ST97) belonging to seven clonal complexes (CCs) (CC1, CC130, CC152, CC3591, CC3666, CC705, CC97) were detected. The Panton-Valentine leukocidin (PVL) genes (lukF-PV/lukS-PV), the bovine leukocidin genes (lukM/lukF-P83) and the human and bovine toxic shock syndrome toxin gene tst-1 variants were detected. FTIR-based capsular serotyping showed CC-specific differences. Most CC97 (cap5 allele) isolates were primarily nonencapsulated (82%), whereas isolates of CC3591 and CC3666 (cap8 allele) were mostly encapsulated (86.4% and 57.8%, respectively). Our results underline the widespread global distribution of cattle-adapted CC97. CONCLUSION: The presence of CC3591 and CC3666 in bovine mastitis suggests an important role in cattle health and dairy production in Rwanda. The results of the present study support the need for a rigorous One-Health Surveillance program of the bovine-human interface.

Persistence of microbiological hazards in food and feed production and processing environments.

Listeria monocytogenes (in the meat, fish and seafood, dairy and fruit and vegetable sectors), Salmonella enterica (in the feed, meat, egg and low moisture food sectors) and Cronobacter sakazakii (in the low moisture food sector) were identified as the bacterial food safety hazards most relevant to public health that are associated with persistence in the food and feed processing environment (FFPE). There is a wide range of subtypes of these hazards involved in persistence in the FFPE. While some specific subtypes are more commonly reported as persistent, it is currently not possible to identify universal markers (i.e. genetic determinants) for this trait. Common risk factors for persistence in the FFPE are inadequate zoning and hygiene barriers; lack of hygienic design of equipment and machines; and inadequate cleaning and disinfection. A well-designed environmental sampling and testing programme is the most effective strategy to identify contamination sources and detect potentially persistent hazards. The establishment of hygienic barriers and measures within the food safety management system, during implementation of hazard analysis and critical control points, is key to prevent and/or control bacterial persistence in the FFPE. Once persistence is suspected in a plant, a 'seek-and-destroy' approach is frequently recommended, including intensified monitoring, the introduction of control measures and the continuation of the intensified monitoring. Successful actions triggered by persistence of L. monocytogenes are described, as well as interventions with direct bactericidal activity. These interventions could be efficient if properly validated, correctly applied and verified under industrial conditions. Perspectives are provided for performing a risk assessment for relevant combinations of hazard and food sector to assess the relative public health risk that can be associated with persistence, based on bottom-up and top-down approaches. Knowledge gaps related to bacterial food safety hazards associated with persistence in the FFPE and priorities for future research are provided.

Characterization of Leuconostoc carnosum and Latilactobacillus sakei during Cooked Pork Ham Processing.

Cooked ham is a popular, ready-to-eat product made of pork meat that is susceptible to microbial growth throughout its shelf life. In this study, we aimed to monitor the microbial growth and composition of nine vacuum-packed cooked ham lots using plate counting until the microbial limit of 7.4 log10 AMC/LAB CFU/g was exceeded. Eight out of nine lots exceeded the microbial limit after 20 days of storage. Lactic acid bacteria strains, particularly Leuconostoc carnosum and Latilactobacillus sakei, prevailed in vacuum-packed cooked ham. Leuconostoc carnosum 2 (Leuc 2) and Latilactobacillus sakei 4 (Sakei 4) were isolated from raw meat and the post-cooking area of the food processing facility. Carbohydrate utilization patterns of Leuc. carnosum PFGE types isolated from raw meat and the food processing environment differed from those isolated from cooked ham. These findings demonstrate how raw meat and its processing environment impact the quality and shelf life of cooked ham.

Assessment of microbial quality in poultry drinking water on farms in Austria.

The quality of poultry drinking water has a significant effect on broiler health and performance. This study conducted an analysis of aerobic mesophilic counts (AMC), Enterobacteriaceae (EB), Pseudomonadaceae (PS), and screened for the presence of Campylobacter spp. in water samples collected from a total of 14 farms in Austria, with either a public or private water source. The efficacy of two water line treatment methods was evaluated: a chemical treatment of the water lines with 4.0 ppm ClO2 (T1) and a combined chemical (4.0 ppm active ClO2 and 3.0% peracetic acid) and mechanical treatment (purging of the water lines with a high-pressure air pump; T2). However, both the T1 and T2 treatments failed to reduce the AMC counts below the maximum acceptable microbial limit of 4.0 log10 CFU/ml in water samples. In addition, no significant reduction in EB and PS counts was observed in water samples after either T1 or T2 water line treatment. The water samples showed a high level of microbial diversity with 18 to 26 different genera. The genus Pseudomonas was most frequently isolated across all poultry farms, while Campylobacter jejuni was identified in a single sample collected before water line treatment. Isolate analysis revealed the presence of opportunistic pathogens in water samples both before (T1 43.1%, T2 30.9%) and after (T1 36.3%, T2 33.3%) water line treatment. Opportunistic pathogens belonging to genera including Pseudomonas spp., Stenotrophomonas spp., and Ochrobactrum spp., were most frequently isolated from poultry drinking water. These isolates exhibited multidrug resistance and resistance phenotypes to antimicrobials commonly used in Austrian poultry farms. The findings of this study emphasize the potential risk of exposure to opportunistic pathogens for poultry and personnel, underscoring the importance of efficient water line management.

The Saprophytic Lifestyle of Listeria monocytogenes and Entry Into the Food-Processing Environment.

Listeria monocytogenes is an environmentally adapted saprophyte that can change into a human and animal bacterial pathogen with zoonotic potential through several regulatory systems. In this review, the focus is on the occurrence of Listeria sensu stricto and sensu lato in different ecological niches, the detection methods, and their analytical limitations. It also highlights the occurrence of L. monocytogenes genotypes in the environment (soil, water, and wildlife), reflects on the molecular determinants of L. monocytogenes for the saprophytic lifestyle and the potential for antibiotic resistance. In particular, the strain-specific properties with which some genotypes circulate in wastewater, surface water, soil, wildlife, and agricultural environments are of particular interest for the continuously updating risk analysis.

Listeria monocytogenes post-outbreak management - When could a food production be considered under control again?

In cases of outbreaks, food business operators face inspections, recall actions and delisting by retailers. This could have happened to an Austrian meat processor whose products have been associated with a cluster of seven cases of listeriosis spread over the years 2015-2017. Sequencing of clinical and foodborne isolates by public health specialists raised the suspect of a single source outbreak since all strains were of MLST 155, cgMLST 1234. Since the family-driven business was highly motivated to save their business, a crisis management scheme was applied that was agreed upon with national authorities. An end-product-based approach testing every single lot for L. monocytogenes was set into power and only negative lots were released for delivery. We combined the active food lot controls of food authorities with a Listeria environmental transmission mapping procedure. The environmental monitoring approach included 19 sampling activities during 3.5 years resulting in 1632 samples. This scheme allowed to trace and mitigate the Listeria contamination but did not jeopardize the processing of meat products. In total, 14 measures were set into power that reduced the overall Listeria occurrence after sanitation of 50-75 % (sampling event I, II) to 0.0-3.8 % (sampling events XIII to XIX). The outbreak-associated ST155/CT1234 clone was not detected in the third sampling event onwards but popped up during the sampling event VIII again. From then on, the outbreak clone ST155/CT1234 was no longer detected in the food business operator (FBO). We conclude that an intense combined investigation of food lots and environmental samples is needed to identify the source and verify that contamination levels are under control. Initially public health authorities suspected contamination of the slicer, but the monitoring approach has localized the source of ST155/CT1234 in a Schnitzel sorting machine. Other factors leading to the contamination scenario were inadequate conveyor belt hygiene. An inadequate crate washing system and an inadequate hygiene lock led to Listeria spreading between compartments. All transmission routes could be effectively interrupted. A root cause analysis and preventive maintenance program implemented in the FPE is mandatory for food processing facilities.

Performance Testing of Bacillus cereus Chromogenic Agar Media for Improved Detection in Milk and Other Food Samples.

In this study, the performance of four alternative selective chromogenic B. cereus agar was compared to the reference mannitol-yolk polymyxin (MYP) agar (ISO 7932) using inclusion and exclusion test strains (n = 110) and by analyzing naturally contaminated milk and other food samples (n = 64). Subsequently, the panC group affiliation and toxin gene profile of Bacillus cereus senso lato (s.l.) isolates were determined. Our results corroborate that the overall best performing media CHROMagar™ B. cereus (93.6% inclusivity; 82.7% exclusivity) and BACARA® (98.2% inclusivity, 62.7% exclusivity) are more sensitive and specific compared to Brilliance™ B. cereus, MYP and ChromoSelect Bacillus Agar. Both media allow unequivocal detection of B. cereus with low risks of misidentification. Media containing ß-D-glucosidase for the detection of presumptive B. cereus may form atypical colony morphologies resulting in a false negative evaluation of the sample. Naturally contaminated samples presented high numbers of background flora, while numbers of presumptive B. cereus were below the detection limit (<10 CFU g-1 or mL-1). Recovery after freezing resulted in the highest detection of B. cereus s.l. on BACARA® (57.8%), CHROMagar™ B. cereus (56.3%) and MYP agar (54.7%). The panC/toxin profile combination IV/A was the most abundant (33.0%), followed by III/F (21.7%) and VI/C (10.4%). More panC and toxin combinations were present in 15.6% of samples when reanalyzed after freezing. In order to improve detection and confirmation of B. cereus s.l. in food samples, we recommend the parallel use of two complementary selective media followed by molecular characterization (e.g., panC typing combined with toxin gene profiling). When determining psychrotolerant or thermophilic members of the B. cereus group, the selective agar media should additionally be incubated at appropriate temperatures (5 °C, ≥45 °C). If high-risk toxin genes (e.g., ces or cytK-1) are detected, the strain-specific ability to produce toxin should be examined to decisively assess risk.

Autochthonous fungi are central components in microbial community structure in raw fermented sausages.

Raw meat sausage represents a unique ecological niche rich in nutrients for microbial consumption, making it particularly vulnerable to microbial spoilage. Starter cultures are applied to improve product stability and safety as well as flavour characteristics. However, the influence of starter cultures on microbial community assembly and succession throughout the fermentation process is largely unknown. In particular the effect on the fungal community has not yet been explored. We evaluate the microbiological status of four different raw meat sausages using high-throughput 16S rRNA gene and ITS2 gene sequencing. The objective was to study temporal changes of microbial composition during the fermentation process and to identify potential keystone species that play an important role within the microbial community. Our results suggest that fungi assigned to the species Debaryomyces hansenii and Alternaria alternata play a key role in microbial community dynamics during fermentation. In addition, bacteria related to the starter culture Lactobacillus sakei and the spoilage-associated genera Acinetobacter, Pseudomonas and Psychrobacter are central components of the microbial ecosystem in raw fermented sausages. Elucidating the exact role and interactions of these microorganisms has the potential to have direct impacts on the quality and safety of fermented foods.

Stress survival islet 1 (SSI-1) survey in Listeria monocytogenes reveals an insert common to listeria innocua in sequence type 121 L. monocytogenes strains.

Listeria monocytogenes strains (n = 117) were screened for the presence of stress survival islet 1 (SSI-1). SSI-1(+) strains (32.5%) belonged mainly to serotypes 1/2c, 3b, and 3c. All sequence type 121 (ST-121) strains included (n = 7) possessed homologues to Listeria innocua genes lin0464 and lin0465 instead of SSI-1.

Sampling the Food-Processing Environment: Taking Up the Cudgel for Preventive Quality Management in Food Processing (FP).

The Listeria monitoring program for Austrian dairies and cheese factories was established in 1988. The aim was to control the entrance of L. monocytogenes into the food-processing environment (FPE), preventing the contamination of food under processing. The Austrian Listeria monitoring program comprises four levels of investigation, dealing with routine monitoring of samples and consequences of finding a positive sample. Preventive quality control concepts attempt to detect a foodborne hazard along the food-processing chain, prior to food delivery, retailing, and consumption. The implementation of a preventive food safety concept provokes a deepened insight by the manufacturers into problems concerning food safety. The development of preventive quality assurance strategies contributes to the national food safety status and protects public health.