Medium-term storage of calf beddings affects bacterial community and effectiveness to inactivate zoonotic bacteria.

Land-spreading of animal faecal wastes -such as animal beddings- can introduce zoonotic enteropathogens into the food system environment. The study evaluated the effectiveness of animal beddings naturally contaminated by calf manure to reduce E. coli O157:H7 or Salmonella enterica. The two pathogens were introduced separately as a four strains-cocktail and at high (>6.5 Log10 g-1) concentration into bedding materials, and their inactivation over a 10 weeks-period was monitored by using a Most Probable Number (MPN) enumeration method. Inactivation of E. coli O157:H7 was more effective in the bedding inoculated immediately after collection from calf pens than in the beddings inoculated after a 2 months-pre-storage period: E. coli O157:H7 levels were reduced by 6.6 Log10 g-1 in unstored bedding (0.5 Log10 g-1 recovered; 95%CI: 0.0-1.2), and by 4.9 Log10 g-1 in pre-stored bedding (2.2 Log10 g-1 recovered; 95%CI: 1.5-2.8) with a significant (p<0.05) difference between unstored and pre-stored. S. enterica was inactivated less effectively as counts were reduced by one order of magnitude, with no significant difference in inactivation between unstored and pre-stored beddings. Low levels of naturally occurring E. coli O157 and Salmonella spp. were detected in the non-inoculated beddings, as well as in the straw prior to use in the animal facility. To better understand the possible biological processes involved, the bacterial community present in the beddings was characterised by short-read 16S rRNA sequencing. Pre-storage of the bedding affected the composition but not the diversity of the bacterial community. Analyses of the key bacterial phyla suggested that the presence of a diverse and stable bacterial community might facilitate inactivation of the introduced pathogens, and a possible role of bacterial orders associated with lignocellulolytic resources. Overall, the study contributed to the understanding of the fate of zoonotic bacteria introduced in animal beddings during storage and identified bedding storage practices pre-and post-use in animal facilities that could be important to prevent the risk of zoonosis dissemination to the environment or to the dairy herds.

Molecular detection and characterization of foodborne bacteria: Recent progresses and remaining challenges.

The global food demand is expected to increase in the coming years, along with challenges around climate change and food security. Concomitantly, food safety risks, particularly those related to bacterial pathogens, may also increase. Thus, the food sector needs to innovate to rise to this challenge. Here, we discuss recent advancements in molecular techniques that can be deployed within various foodborne bacteria surveillance systems across food settings. To start with, we provide updates on nucleic acid-based detection, with a focus on polymerase chain reaction (PCR)-based technologies and loop-mediated isothermal amplification (LAMP). These include descriptions of novel genetic markers for several foodborne bacteria and progresses in multiplex PCR and droplet digital PCR. The next section provides an overview of the development of clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated (Cas) proteins systems, such as CRISPR-Cas9, CRISPR-Cas12a, and CRISPR-Cas13a, as tools for enhanced sensitive and specific detection of foodborne pathogens. The final section describes utilizations of whole genome sequencing for accurate characterization of foodborne bacteria, ranging from epidemiological surveillance to model-based predictions of bacterial phenotypic traits through genome-wide association studies or machine learning.

Fecal excretion of Campylobacter jejuni by young dairy calves and the relationship with neonatal immunity and personality traits.

AIMS: Zoonotic pathogens in bovine herds are major concerns for human and animal health, but their monitoring in animals can be challenging in the absence of clinical signs. Our objective was to determine the association between fecal excretion of Campylobacter jejuni, neonatal immunity, and personality traits of calves. METHODS AND RESULTS: Forty-eight dairy calves were reared in three indoor pens from birth to 4 weeks of life. Microbial analyses of the fecal samples collected weekly revealed that the proportion of calves naturally contaminated with C. jejuni in each pen reached 70% after 3 weeks of life. High (>16 g l-1) levels of IgG levels in the serum of neonatal calves were negatively (P = .04) associated with fecal detection of C. jejuni over the trial period. Calves that spent more time interacting with a novel object tended to be positive (P = .058) for C. jejuni. CONCLUSIONS: Overall, the findings indicate that the immunity of neonatal dairy animals and possibly the animal's behavior may contribute to the fecal shedding of C. jejuni.

Importance of the Farm Environment and Wildlife for Transmission of Campylobacter jejuni in A Pasture-Based Dairy Herd.

Cattle are an established reservoir of the foodborne bacterial pathogen Campylobacter jejuni. Our six-month study aimed to evaluate sources and pathways governing long-term presence of C. jejuni in a pasture-based dairy herd. C. jejuni was detected in all sample types (soil, pasture, stock drinking water, bird, rodents and cow faeces). It was persistently detected from cow (54%; 49/90 samples) and bird (36%; 77/211) faeces. Genetic comparison of 252 C. jejuni isolates identified 30 Multi-Locus Sequence Types (ST). ST-61 and ST-42 were persistent in the herd and accounted for 43% of the cow isolates. They were also detected on pasture collected from fields both recently and not recently grazed, indicating that grazed pasture is an important pathway and reservoir for horizontal transmission among cows. ST-61 accounted for 9% of the bird isolates and was detected at four of the six sampling events, suggesting that bird populations might contribute to the cycling of ruminant-adapted genotypes on-farm. Overall, the results indicated that management of grazed pasture and supplementary feed contaminated by bird droppings could be targeted to effectively reduce transmission of C. jejuni to dairy herds, the farm environment and ultimately to humans.

Differences in the fecal concentrations and genetic diversities of Campylobacter jejuni populations among individual cows in two dairy herds.

Dairy cows have been identified as common carriers of Campylobacter jejuni, which causes many of the human gastroenteritis cases reported worldwide. To design on-farm management practices that control the human infection sourced from dairy cows, the first step is to acquire an understanding of the excretion patterns of the cow reservoir. We monitored the same 35 cows from two dairy farms for C. jejuni excretion fortnightly for up to 12 months. The objective was to examine the concentration of C. jejuni and assess the genetic relationship of the C. jejuni populations excreted by individual cows. Significant differences (P < 0.01) in C. jejuni fecal concentration were observed among the 35 cows, with median concentrations that varied by up to 3.6 log(10) · g(-1) feces. A total of 36 different genotypes were identified from the 514 positive samples by using enterobacterial repetitive intergenic consensus (ERIC)-PCR. Although 22 of these genotypes were excreted by more than one cow, the analysis of frequencies and distribution of the genotypes by model-based statistics revealed a high degree of individuality in the C. jejuni population in each cow. The observed variation in the frequency of excretion of a genotype among cows and the analysis by multilocus sequence typing (MLST) of these genotypes suggest that excretion of C. jejuni in high numbers is due to a successful adaptation of a particular genotype to a particular cow's gut environment, but that animal-related factors render some individual cows resistant to colonization by particular genotypes. The reasons for differences in C. jejuni colonization of animals warrant further investigation.

Lyophilization prior to direct DNA extraction from bovine feces improves the quantification of Escherichia coli O157:H7 and Campylobacter jejuni.

Lyophilization was used to concentrate bovine feces prior to DNA extraction and analysis using real-time PCR. Lyophilization significantly improved the sensitivity of detection compared to that in fresh feces and was associated with reliable quantification of both Escherichia coli O157:H7 and Campylobacter jejuni bacteria present in feces at concentrations ranging between 2 log(10) and 6 log(10) CFU g(-)(1).

Prion degradation in soil: possible role of microbial enzymes stimulated by the decomposition of buried carcasses.

This study is part of a European project focused on understanding the biotic and abiotic mechanisms involved in the retention and dissemination of transmissible spongiform encephalopathies (TSE) infectivity in soil in order to propose practical recommendations to limit environmental contamination. A 1-year field experiment was conducted with lamb carcasses buried in a pasture soil at three depths (25, 45, and 105 cm). Microbial community response to carcasses was monitored through the potential proteolytic activity and substrate induced respiration (SIR). Soil above carcasses and control soil exhibited low proteolytic capacity, whatever the depth of burial. Contrastingly, in soil beneath the carcasses, proteolysis was stimulated. Decomposing carcasses also stimulated SIR, i.e., microbial biomass, suggesting that proteolytic populations specifically developed on lixiviates from animal tissues. Decomposition of soft tissues occurred within 2 months at subsurface while it lasted at least 1 year at deeper depth where proteolytic activities were season-dependent. The ability of soil proteases to degrade the beta form of prion protein was shown in vitro and conditions of burial relevant to minimize the risk of prion protein dissemination are discussed.