Proteome analysis of Campylobacter jejuni poultry strain 2704 survival during 45 min exposure to peracetic acid.

Peracetic acid (PAA) applied to whole poultry carcasses can reduce the number of Campylobacter, a leading cause of human gastroenteritis. However, previous modelling experiments indicated that Campylobacter survived in greater numbers when pre-treated with a thermal stress equivalent to poultry processing scalding prior to chilling with PAA than when subject to chilling with PAA only. To better understand how Campylobacter responds to PAA, proteomes of C. jejuni poultry strain 2704 were measured after exposure to PAA (60 ppm, pH 4.0) for 45 min under laboratory ambient conditions (approximately 23 °C) to establish a foundational map of survival mechanism before combining with other stresses. Analysis of 580 quantified proteins did not indicate a triggered "peroxide shock" response, nor were common heat shock responses detected. Thioredoxin, iron homeostatic, peroxiredoxins and cytochrome c peroxidases became more abundant suggesting that PAA disturbed cytoplasmic redox homeostasis resulting in antioxidant activation and increased prioritisation of iron homeostasis. The PAA treatment led to responses that included an increased priority for oxidative phosphorylation and a simultaneous decrease in central metabolism associated protein abundances. Lon protease was induced suggesting it has a role in maintaining homeostasis during non-thermal stress. Proteins in flagella and chemotaxis became more abundant though whether PAA has a chemorepellent effect requires further investigation. Overall, the proteome data suggests there was a rapid cellular response to applied PAA stress in the first 15 min with the adaptation to the stress completing between 30 and 45 min. The findings will help guide PAA implementation in commercial poultry processing in terms of processing location and length of application.

Clostridium perfringens associated with dairy farm systems show diverse genotypes.

Clostridium perfringens is a bacterial species of importance to both public and animal health. Frequently found in food system environments, it presents a risk to food animal health such as dairy herds, and may cross contaminate associated ingredients or food products, with potential to cause sporadic and outbreaks of disease in human populations, including gastroenteric illness. In this study, we characterized C. perfringens isolated from bovine, caprine, and ovine dairy farm systems (n = 8, 11 and 4, respectively). Isolates were phenotypically screened for antimicrobial sensitivity profiling, and subjected to whole genome sequencing to elucidate related genetic markers, as well as examine virulence gene markers, mobile genetic elements, and other features. Both toxin type A and type D isolates were identified (78 % and 22 % of isolates, respectively), including 20 novel sequence types. Resistance to clindamycin was most prevalent among antibiotics screened (30 %), followed by erythromycin (13 %), then penicillin and tetracycline (4 %), although an additional 3 isolates were non-susceptible to tetracycline. Most isolates harboured plasmids, which mobilised virulence markers such as etx, cpb2, and resistance markers tetA(P), tetB(P), and erm(Q), on conjugative plasmids. The presence of type D isolates on caprine farms emphasizes the need for control efforts to prevent infection and potential enterotoxemia. Clostridium perfringens enterotoxin (cpe) was not identified, suggesting lower risk of gastrointestinal illness from contaminated foods, the presence of other virulence and antimicrobial resistance markers suggests farm hygiene remains an important consideration to help ensure food safety of associated dairy foods produced.

Co-culture with Acinetobacter johnsonii enhances benzalkonium chloride resistance in Salmonella enterica via triggering lipid A modifications.

Salmonella enterica is one of the leading causes of foodborne gastroenteritis worldwide. In the food production environment, many bacterial species co-exist on surfaces in biofilm structures, which can act as reservoirs of microbial contamination of food products. Polymicrobial biofilms have been shown to have greater tolerance to antimicrobials, such as disinfectants, however the mechanistic basis of this is poorly understood. In this study, S. enterica subsp. enterica serovar Liverpool was co-cultured in mixed-species biofilms with bacteria isolated from the food production environment and challenged with the cationic biocide disinfectant, benzalkonium chloride (BC). Co-culture with the common environmental bacterium Acinetobacter johnsonii resulted in >200-fold higher resistance of S. Liverpool to BC, compared to mono-culture biofilms. The transcriptional response of S. enterica to biofilm co-culture was determined using a dual RNA-seq strategy. Genes controlled by the PhoPQ and PmrAB two-component systems, involved in lipid A modification and associated with cationic antimicrobial peptide resistance (CAMP) of S. Liverpool, were significantly upregulated. Deletion of either the phoP or pmrA genes resulted in an increase in susceptibility to BC, suggesting that activation of their regulons during co-culture enhances BC resistance. S. Liverpool lipid A profiles changed significantly upon co-culturing, with greater incorporation of both phosphoethanolamine and palmitate, which was dependent upon activation of PhoPQ and PmrAB. We conclude that when grown in the presence of A. johnsonii, S. Liverpool increases its tolerance to cationic BC disinfection by remodelling its cell envelope including reducing the net negative charge of lipid A and increasing lipid A acyl density.

Natural Anti-Microbials for Enhanced Microbial Safety and Shelf-Life of Processed Packaged Meat.

Microbial food contamination is a major concern for consumers and food industries. Consumers desire nutritious, safe and "clean label" products, free of synthetic preservatives and food industries and food scientists try to meet their demands by finding natural effective alternatives for food preservation. One of the alternatives to synthetic preservatives is the use of natural anti-microbial agents in the food products and/or in the packaging materials. Meat and processed meat products are characteristic examples of products that are highly perishable; hence natural anti-microbials can be used for extending their shelf-life and enhancing their safety. Despite several examples of the successful application of natural anti-microbial agents in meat products reported in research studies, their commercial use remains limited. This review objective is to present an extensive overview of recent research in the field of natural anti-microbials, covering essential oils, plant extracts, flavonoids, animal-derived compounds, organic acids, bacteriocins and nanoparticles. The anti-microbial mode of action of the agents, in situ studies involving meat products, regulations and, limitations for usage and future perspectives are described. The review concludes that naturally derived anti-microbials can potentially support the meat industry to provide "clean label", nutritious and safe meat products for consumers.

Survival of Salmonella Under Heat Stress is Associated with the Presence/Absence of CRISPR Cas Genes and Iron Levels.

Clustered regularly interspaced short palindromic repeats (CRISPR) cas genes have been linked to stress response in Salmonella. Our aim was to identify the presence of CRISPR cas in Salmonella and its response to heat in the presence of iron. Whole genomes of Salmonella (n = 50) of seven serovars were compared to identify the presence of CRISPR cas genes, direct-repeats and spacers. All Salmonella genomes had all cas genes present except S. Newport 2393 which lacked these genes. Gene-specific primers were used to confirm the absence of these genes in S. Newport 2393. The presence/absence of CRISPR cas genes was further investigated among 469 S. Newport genomes from PATRIC with 283 genomes selected for pan-genome analysis. The response of eleven Salmonella strains of various serovars to gradual heat in ferrous and ferric forms of iron was investigated. A total of 32/283 S. Newport genomes that lacked all CRISPR cas genes clustered together. S. Newport 2393 was the most heat-sensitive strain at higher iron levels (200 and 220 pm) in ferrous and ferric forms of iron. The absence of CRISPR cas genes in S. Newport 2393 may contribute to its increase in heat sensitivity and iron may play a role in this. The high reduction in numbers of most Salmonella strains exposed to heat makes it unfeasible to extract RNA and conduct transcription studies. Further studies should be conducted to validate the survival of Salmonella when exposed to heat in the presence/absence of CRISPR cas genes and different iron levels.

Comparison between cage and free-range egg production on microbial composition, diversity and the presence of Salmonella enterica.

The microbial composition of the food production environment plays an important role in food safety and quality. This study employed both 16 S rRNA gene sequencing technology and culture-based techniques to investigate the bacterial microbiota of an egg production facility comprising of both free-range and conventional cage housing systems. The study also aimed to detect the presence of Salmonella enterica and determine whether its presence was positively or negatively associated with other taxa. Our findings revealed that microbiota profiles of free-range and cage houses differ considerably in relation to the relative abundance and diversity with a number of taxa unique to each system and to individual sampling sites within sheds. Core to each housing system were known inhabitants of the poultry gastrointestinal tracts, Romboutsia and Turicibacter, as well as common spoilage bacteria. Generally, free-range samples contained fewer taxa and were dominated by Staphylococcus equorum, differentiating them from the cage samples. Salmonella enterica was significantly associated with the presence of a taxa belonging to the Carnobacteriaceae family. The results of this study demonstrate that the diversity and composition of the microbiota is highly variable across egg layer housing systems, which could have implications for productivity, food safety and spoilage.

Survival of Salmonella on Red Meat in Response to Dry Heat.

ABSTRACT: Red meat is associated with Salmonella outbreaks, resulting in negative impacts for the processing industry. Little work has been reported on the use of dry heat as opposed to moist heat against Salmonella on red meat. We determined the effect of drying at 25°C and dry heat at 70°C with ∼10% relative humidity for 1 h against 11 Salmonella strains of multiple serovars on beef, lamb, and goat and rubber as an inert surface. Each strain at ∼108 CFU/mL was inoculated (100 µL) onto ±1 g (1 cm2) of each surface and allowed to attach for 15 min in a microcentrifuge tube. Samples were then exposed to 70 and 25°C with ∼10% relative humidity in a heating block. Surviving Salmonella numbers on surfaces were enumerated on a thin agar layer medium. If numbers were below the limit of detection (2.01 log CFU/cm2), Salmonella cells were enriched before plating to determine the presence of viable cells. Water loss (percent) from meat after at 25 and 70°C was determined. Whole genomes of Salmonella were interrogated to identify the presence-absence of stress response genes (n = 30) related to dry heat that may contribute to the survival of Salmonella. The survival of Salmonella at 25°C was significantly higher across all surfaces (∼6.09 to 7.91 log CFU/cm2) than at 70°C (∼3.66 to 6.33 log CFU/cm2). On rubber, numbers of Salmonella were less than the limit of detection at 70°C. Water loss at 70°C (∼17.72 to 19.89%) was significantly higher than at 25°C (∼2.98 to 4.11%). Salmonella cells were not detected on rubber, whereas survival occurred on all red meat at 70°C, suggesting its protective effect against the effect of heat. All Salmonella strains carried 30 stress response genes that likely contributed to survival. A multi-antibiotic-resistant Salmonella Typhimurium 2470 exhibited an increase in heat resistance at 70°C on beef and lamb compared with other strains. Our work shows that dry heat at 70°C for 1 h against Salmonella on red meat is not a practical approach for effectively reducing or eliminating them from red meat.

Applicability of Enterobacteriaceae and coliforms tests as indicators for Cronobacter in milk powder factory environments.

The emergence of Cronobacter as an important potential pathogen for newborn children and its occurrence in powdered infant formulae has generated a need to develop new management practices for this food group. This includes reduction of the prevalence of Cronobacter in manufacturing environments which can be a source of Cronobacter. This study was performed to assess the suitability of qualitative and quantitative Enterobacteriaceae and coliforms indicator tests for the presence and prevalence of Cronobacter. Environmental swabs (205) from five milk powder factories were examined. The qualitative indicator tests had good sensitivity but they lacked specificity for reliable routine use. Logistic regression analyses revealed a significant relationship between the quantitative indicator tests and Cronobacter prevalence, where the Enterobacteriaceae count was a slightly stronger predictor for Cronobacter than the coliforms count. The optimum test sensitivity (81%) and specificity (66%) was obtained when the indicator count thresholds were set at ≥1 cfu/cm2. However, since 11% of samples were Cronobacter positive when counts of Enterobacteriaceae and coliforms were less than 1 cfu/cm2, specific testing for Cronobacter is advised in addition to Enterobacteriaceae testing to minimise risk of transfer of Cronobacter from the factory environment into powdered infant formulae products.

Salmonella survival after exposure to heat in a model meat juice system.

The effect of heat against eleven Salmonella strains in model meat juices was examined. Juices from beef, lamb and goat were made from either the fatty layer (FL), muscle (M) or a mixture of both (FLM). The pH of each FLM sample was altered to match the pH of PBS and vice versa to determine the pH effect on the survival of Salmonella against the effect of heat. Salmonella were exposed to either gradual heating to 70 °C in FLM, M and FL or heat shock at 70 °C for 5 min in FLM. Fat, fatty acid profile and iron content of the juices were determined. Gradual heat treatment significantly (p ≤ 0.05) reduced Salmonella as compared to the untreated controls (~1.92-7.61 log CFU ml-1) while heat shock significantly (p ≤ 0.05) reduced Salmonella as compared to the untreated controls (~5.80-7.36 log CFU ml-1). Survival of Salmonella was higher in lamb juices than other juices. The fat content in lamb FL (3.25%) was significantly higher (p ≤ 0.05) than beef (1.30%) and goat FL (1.42%). Iron content in lamb FLM (~127 mg kg-1) was significantly (p ≤ 0.05) lower than beef (~233 mg kg-1) and goat FLM (~210 mg kg-1). The omega 6 and linoleic acid content in goat FLM (~36.0% and ~34.4%) was significantly higher (p ≤ 0.05) than beef (~29.1% and ~27.1%). Fat, fatty acids and iron may differentially protect Salmonella against the effect of heat in these juices.

Impact of Poultry Processing Operating Parameters on Bacterial Transmission and Persistence on Chicken Carcasses and Their Shelf Life.

It is important for the poultry industry to maximize product safety and quality by understanding the connection between bacterial diversity on chicken carcasses throughout poultry processing to the end of shelf life and the impact of the local processing environment. Enumeration of total aerobic bacteria, Campylobacter and Pseudomonas, and 16S rRNA gene amplicon sequencing were used to evaluate the processing line by collecting 10 carcasses from five processing steps: prescald, postplucker, pre- and post-immersion chill, and post-air chill. The diversity throughout a 12-day shelf life was also determined by examining 30 packaged carcasses. To identify the sources of possible contamination, scald water tank, immersion chilling water tank, air samples, and wall surfaces in the air-chill room were analyzed. Despite bacterial reductions on carcasses (>5 log10 CFU/ml) throughout the process, each step altered the bacterial diversity. Campylobacter was a minor but persistent component in the bacterial community on carcasses. The combination of scalding, defeathering, and plucking distributed thermophilic spore-forming Anoxybacillus to carcasses, which remained at a high abundance on carcasses throughout subsequent processes. Pseudomonas was not isolated from carcasses after air chilling but was abundant on the wall of the air-chill room and became the predominant taxon at the end of shelf life, suggesting possible contamination through air movement. The results suggest that attention is needed at each processing step, regardless of bacterial reductions on carcasses. Changing scalding water regularly, maintaining good hygiene practices during processing, and thorough disinfection at the end of each processing day are important to minimize bacterial transmission.IMPORTANCE Culture-based and culture-independent approaches were utilized to reveal bacterial community changes on chicken carcasses at different processing steps and potential routes from the local processing environment. Current commercial processing effectively reduced bacterial loads on carcasses. Poultry processes have similar processes across facilities, but various processing arrangements and operating parameters could impact the bacterial transmission and persistence on carcasses differently. This study showed the use of a single tunnel incorporating scalding, defeathering and plucking may undesirably distribute the thermoduric bacteria, e.g., Campylobacter and Anoxybacillus, between the local environment and carcasses, whereas this does not occur when these steps are separated. The length of immersion and air chilling also impacted bacterial diversity on carcasses. Air chilling can transfer Pseudomonas from wall surfaces onto carcasses; this may subsequently influence chicken product shelf life. This study helps poultry processors understand the impact of current commercial processing and improve the chicken product quality and safety.

Effects of heat stress on animal physiology, metabolism, and meat quality: A review.

Heat stress is one of the most stressful events in the life of livestock with harmful consequences for animal health, productivity and product quality. Ruminants, pigs and poultry are susceptible to heat stress due to their rapid metabolic rate and growth, high level of production, and species-specific characteristics such as rumen fermentation, sweating impairment, and skin insulation. Acute heat stress immediately before slaughter stimulates muscle glycogenolysis and can result in pale, soft and exudative (PSE) meat characterized by low water holding capacity (WHC). By contrast, animals subjected to chronic heat stress, have reduced muscle glycogen stores resulting in dark, firm and dry (DFD) meat with high ultimate pH and high WHC. Furthermore, heat stress leads to oxidative stress, lipid and protein oxidation, and reduced shelf life and food safety due to bacterial growth and shedding. This review discusses the scientific evidence regarding the effects of heat stress on livestock physiology and metabolism, and their consequences for meat quality and safety.

Changes of the bacterial community diversity on chicken carcasses through an Australian poultry processing line.

Understanding the bacterial community profile through poultry processing could help the industry to produce better poultry products. In this study, 10 chicken carcasses were randomly sampled from before and after scalding, before and after immersion chilling, and after air chilling each through a modern commercial processing line, along with the contents of 10 caeca. The sampled processing line effectively reduced the bacterial counts by > 4.6 Log10 CFU/ml for each of Total Viable Counts, Escherichia coli and Campylobacter. However, the metagenomics results suggested that Lactobacillus, Staphylococcus and unclassified Lachnospiraceae persisted at all sampling stages. Pseudomonas, Paeniglutamicibacter, Chryseobacterium and Pseudarthrobacter comprised 47.2% in the bacterial community on samples after air chilling compared to 0.3% on samples after immersion chilling, whereas TVCs were the same. Overall, the current interventions of the investigated poultry processing line were unable to eliminate persistence of certain foodborne pathogens, despite a significant reduction of the overall bacterial counts. Chilling is an important controlling point in contamination/cross-contamination, particularly extended air chilling. Lastly, the large presence of Pseudomonas on chickens after air chilling may lead to downstream spoilage related issues, which needs more investigation to explore quantitatively the effect on the shelf life of poultry products.

Draft Genome Sequences of Four Antibiotic-Resistant Salmonella Strains Isolated from Australian Red Meat Animal Species.

The genome sequences of four antibiotic-resistant Salmonella strains isolated from red meat animals in Australia are presented. Multidrug-resistant Salmonella enterica serovar Heidelberg 329 and Salmonella enterica serovar Typhimurium 2470 harbored an IncHI2 plasmid similar to the multidrug-resistant S. Heidelberg strain N13-01290 plasmid pN13-01290_23 previously isolated in Canada.

Comparative Genomics and Phenotypic Investigations Into Antibiotic, Heavy Metal, and Disinfectant Susceptibilities of Salmonella enterica Strains Isolated in Australia.

Salmonella enterica is recognized as a major contributor of gastrointestinal illness worldwide. Concerns have been raised over the increasing prevalence of antibiotic resistant strains of Salmonella isolated from animals and food, and the role of antibiotics and other antimicrobial agents such as biocides and heavy metals in the selection and dissemination of antibiotic resistant bacteria to human hosts. In this study the antibiotic, heavy metal and disinfectant resistance genotypes and phenotypes of 19 S. enterica isolates from food-producing animals were established using whole genome sequence analysis, disc diffusion, as well as broth or agar dilution methods. This study also investigated the genomic environment of resistance genes on mobile genetic elements and chromosomal DNA. An ampicillin and streptomycin resistant S. Infantis isolate in this study harbored a ß-lactamase (bla TEM-1 ), and two streptomycin resistance conferring genes (strA and strB) on a class 1 integron mobilized on a large conjugative plasmid. This plasmid also harbored two arsenic resistance gene cassettes. The arsenic resistance cassette, arsRCDAB, was also observed in two S. Singapore isolates with high tolerance to arsenate. A nalidixic acid resistant S. Typhimurium isolate was found to possess a mutation in gyrA resulting in amino acid change Asp87Gly and tetracycline resistant S. Typhimurium isolate was found to harbor efflux pump gene, tetA. No resistance (genotypic or phenotypic) was recorded to the disinfectants screened in this study. Taken together, results of this study showed a good correlation between predicted and measured resistances when comparing genotypic and phenotypic data, respectively. The findings of this study do not suggest resistance to clinically relevant antibiotics are widespread among Salmonella isolated from Australian food-producing animals.

Transcriptomic response of Escherichia coli O157 isolates on meat: Comparison between a typical Australian isolate from cattle and a pathogenic clinical isolate.

The majority of foodborne illnesses associated with E. coli O157 are attributed to the consumption of foods of bovine origin. In this study, RNA-Seq experiments were undertaken with E. coli O157 to identify genes that may be associated with growth and survival on meat and the beef carcass at low temperature. In addition, the response of an E. coli O157 isolate representative of the general genetic 'type' found in Australia (E. coli O157:H- strain EC2422) was compared to that of a pathogenic clinical isolate (E. coli O157:H7 strain Sakai) not typically found in Australia. Both strains up-regulated genes involved in the acid stress response, cold shock response, quorum sensing, biofilm formation and Shiga toxin production. Differences were also observed, with E. coli O157:H7 Sakai up-regulating genes playing a critical role in the barrier function of the outer membrane, lipopolysaccharide biosynthesis, extracellular polysaccharide synthesis and curli production. In contrast, E. coli O157:H- EC2422 down-regulated genes involved in peptidoglycan biosynthesis and of the primary envelope stress response Cpx system. The unique gene expression profiles of the strains, indicate that these genotypes may differ in their ability to persist in the meat production environment and therefore also in their ability to cause disease.

Overexpressing ovotransferrin and avian ß-defensin-3 improves antimicrobial capacity of chickens and poultry products.

Zoonotic and foodborne diseases pose a significant burden, decreasing both human and animal health. Modifying chickens to overexpress antimicrobials has the potential to decrease bacterial growth on poultry products and boost chicken innate immunity. Chickens overexpressing either ovotransferrin or avian ß-defensin-3 (AvßD3) were generated using Tol-2 transposons. Transgene expression at the RNA and protein level was seen in egg white, breast muscle, and serum. There were significant differences in the immune cell populations in the blood, bursa, and spleen associated with transgene expression including an increased proportion of CD8+ cells in the blood of ovotransferrin and AvßD3 transgenic birds. Expression of the antimicrobials inhibited the in vitro growth of human and chicken bacterial pathogens and spoilage bacteria. For example, transgene expression significantly reduced growth of aerobic and coliform bacteria in breast muscle and decreased the growth of Salmonella enterica in egg white. Overall these results indicate that overexpression of antimicrobials in the chicken can impact the immune system and increase the antimicrobial capacity of poultry products.

Characterisation of the Brochothrix thermosphacta sortase A enzyme.

Gram-positive bacteria utilise class A sortases to coat the surface of their cells with a diversity of proteins that facilitate interactions with their environment and play fundamental roles in cell physiology and virulence. A putative sortase A gene was identified in the genome of the poorly studied meat spoilage bacterium Brochothrix thermosphacta. To understand how this bacterium mediates interactions with its environment, an N-terminal truncated, His-tagged variant of this protein (His6-BtSrtA) was expressed and purified. Catalytic activity of recombinant His6-BtSrtA was investigated, including sorting motif recognition of target proteins and bioconjugation activity. Further, the B. thermosphacta genome was examined for the presence of sortase A (SrtA) protein substrates. His6-BtSrtA readily formed intermediate complexes with LPXTG-tagged proteins. Although the reaction was inefficient, nucleophilic attack of the resultant thioacyl intermediates by tri-glycine was observed. Genome examination identified 11 potential SrtA substrates, two of which contained protein domains associated with adherence of pathogens to host extracellular matrix proteins and cells, suggesting the B. thermosphacta SrtA may be indirectly involved in its attachment to meat surfaces. Thus, further work in this area could provide crucial insight into molecular mechanisms involved in the colonisation of meat by B. thermosphacta.

The role of meat in foodborne disease: Is there a coming revolution in risk assessment and management?

Meat has featured prominently as a source of foodborne disease and a public health concern. For about the past 20 years the risk management paradigm has dominated international thinking about food safety. Control through the supply chain is supported by risk management concepts, as the public health risk at the point of consumption becomes the accepted outcome based measure. Foodborne pathogens can be detected at several points in the supply chain and determining the source of where these pathogens arise and how they behave throughout meat production and processing are important parts of risk based approaches. Recent improvements in molecular and genetic based technologies and data analysis for investigating source attribution and pathogen behaviour have enabled greater insights into how foodborne outbreaks occur and where controls can be implemented. These new approaches will improve our understanding of the role of meat in foodborne disease and are expected to have a significant impact on our understanding in the coming years.

Salmonella enterica subsp. salamae serovar Sofia, a prevalent serovar in Australian broiler chickens, is also capable of transient colonisation in layers.

1. Salmonella enterica subsp. salamae serovar sofia (S. sofia) is a prevalent strain of Salmonella in Australian broilers and has been isolated from broiler chickens, litter, dust, as well as pre- and post-processing carcasses, and retail chicken portions but has never been reported in commercial Australian layers or eggs. 2. To investigate whether a S. sofia isolate from a broiler could colonise layers, one-month-old Hyline brown layers were orally inoculated with S. sofia and colonisation was monitored for 2-4 weeks. 3. Overall, 30-40% of the chickens shed S. sofia from the cloaca between 6 and 14 d post-inoculation which then declined to 10% by d 21. Necropsy at 2 weeks post-inoculation revealed 80% of birds harboured S. sofia in the caecum, whilst, by 4 weeks post-infection, no chickens were colonised with S. sofia in the gastrointestinal tract, liver or spleen. Additionally, no aerosol 'bird to bird' transfer was evident. 4. This study demonstrated that laying hens can be colonised by broiler-derived S. sofia; however, this colonisation was transient, reaching a peak at 14 d post-inoculation, and was completely cleared by 28 d post-inoculation. The transience of colonisation of S. sofia in layers could be a factor explaining why S. sofia has never been detected when screening for Salmonella serotypes found in Australian laying hens or eggs.

Genomic and metabolic characterization of spoilage-associated Pseudomonas species.

Pseudomonas are common spoilage agents of aerobically stored fresh foods. Their ability to cause spoilage is species- and may be strain-specific. To improve our understanding of the meat and milk spoilage agents Pseudomonas fragi and Pseudomonas lundensis, we sequenced the genomes of 12 P. fragi and seven P. lundensis isolates. These genomes provided a dataset for genomic analyses. Key volatile organic compounds (VOCs) produced or metabolised by the isolates were determined during their growth on a beef paste and where possible, metabolic activity was associated with gene repertoire. Genome analyses showed that the isolates included in this work may belong to more than two Pseudomonas species with possible spoilage potential. Pan-genome analyses demonstrated a high degree of diversity among the P. fragi and genetic flexibility and diversity may be traits of both species. Growth of the P. lundensis isolates was characterised by the production of large amounts of 1-undecene, 5-methyl-2-hexanone and methyl-2-butenoic acid. P. fragi isolates produced extensive amounts of methyl and ethyl acetate and the production of methyl esters predominated over ethyl esters. Some of the P. fragi produced extremely low levels of VOCs, highlighting the importance of strain-specific studies in food matrices. Furthermore, although usually not considered to be denitrifiers, all isolates generated molecular nitrogen, indicating that at least some steps of this pathway are intact.