Whole-Genome Sequencing Analysis of Listeria monocytogenes from Rural, Urban, and Farm Environments in Norway: Genetic Diversity, Persistence, and Relation to Clinical and Food Isolates.

Listeria monocytogenes is a ubiquitous environmental bacterium associated with a wide variety of natural and human-made environments, such as soil, vegetation, livestock, food processing environments, and urban areas. It is also among the deadliest foodborne pathogens, and knowledge about its presence and diversity in potential sources is crucial to effectively track and control it in the food chain. Isolation of L. monocytogenes from various rural and urban environments showed higher prevalence in agricultural and urban developments than in forest or mountain areas, and that detection was positively associated with rainfall. Whole-genome sequencing (WGS) was performed for the collected isolates and for L. monocytogenes from Norwegian dairy farms and slugs (218 isolates in total). The data were compared to available data sets from clinical and food-associated sources in Norway collected within the last decade. Multiple examples of clusters of isolates with 0 to 8 whole-genome multilocus sequence typing (wgMLST) allelic differences were collected over time in the same location, demonstrating persistence of L. monocytogenes in natural, urban, and farm environments. Furthermore, several clusters with 6 to 20 wgMLST allelic differences containing isolates collected across different locations, times, and habitats were identified, including nine clusters harboring clinical isolates. The most ubiquitous clones found in soil and other natural and animal ecosystems (CC91, CC11, and CC37) were distinct from clones predominating among both clinical (CC7, CC121, and CC1) and food (CC9, CC121, CC7, and CC8) isolates. The analyses indicated that ST91 was more prevalent in Norway than other countries and revealed a high proportion of the hypovirulent ST121 among Norwegian clinical cases. IMPORTANCE Listeria monocytogenes is a deadly foodborne pathogen that is widespread in the environment. For effective management, both public health authorities and food producers need reliable tools for source tracking, surveillance, and risk assessment. For this, whole-genome sequencing (WGS) is regarded as the present and future gold standard. In the current study, we use WGS to show that L. monocytogenes can persist for months and years in natural, urban, and dairy farm environments. Notably, clusters of almost identical isolates, with genetic distances within the thresholds often suggested for defining an outbreak cluster, can be collected from geographically and temporally unrelated sources. The work highlights the need for a greater knowledge of the genetic relationships between clinical isolates and isolates of L. monocytogenes from a wide range of environments, including natural, urban, agricultural, livestock, food production, and food processing environments, to correctly interpret and use results from WGS analyses.

The prevalence of Campylobacter spp., Listeria monocytogenes and Shiga toxin-producing Escherichia coli in Norwegian dairy cattle farms: A comparison between free stall and tie stall housing systems.

AIMS: This study explored how dairy farm operating systems with free-stall or tie-stall housing and cow hygiene score influence the occurrence of zoonotic bacteria in raw milk. METHODS AND RESULTS: Samples from bulk tank milk (BTM), milk filters, faeces, feed, teats and teat milk were collected from 11 farms with loose housing and seven farms with tie-stall housing every second month over a period of 11 months and analysed for the presence of STEC by culturing combined with polymerase chain reaction and for Campylobacter spp. and L. monocytogenes by culturing only. Campylobacter spp., L. monocytogenes and STEC were present in samples from the farm environment and were also detected in 4%, 13% and 7% of the milk filters, respectively, and in 3%, 0% and 1% of BTM samples. Four STEC isolates carried the eae gene, which is linked to the capacity to cause severe human disease. L. monocytogenes were detected more frequently in loose housing herds compared with tie-stalled herds in faeces (p = 0.02) and feed (p = 0.03), and Campylobacter spp. were detected more frequently in loose housing herds in faeces (p < 0.01) and teat swabs (p = 0.03). An association between cow hygiene score and detection of Campylobacter spp. in teat milk was observed (p = 0.03). CONCLUSION: Since some samples collected from loose housing systems revealed a significantly higher (p < 0.05) content of L. monocytogenes and Campylobacter spp. than samples collected from tie-stalled herds, the current study suggests that the type of housing system may influence the food safety of raw milk. SIGNIFICANCE AND IMPACT OF THE STUDY: This study highlights that zoonotic bacteria can be present in raw milk independent of hygienic conditions at the farm and what housing system is used. Altogether, this study provides important knowledge for evaluating the risk of drinking unpasteurized milk.

The Ability of Shiga Toxin-Producing Escherichia coli to Grow in Raw Cow's Milk Stored at Low Temperatures.

Despite the lack of scientific evidence, some consumers assert that raw milk is a natural food with nutritional and immunological properties superior to pasteurized milk. This has led to the increased popularity of unpasteurized cow milk (UPM) and disregard for the risks of being exposed to zoonotic infections. Dairy cattle are healthy carriers of Shiga toxin (Stx)-producing E. coli (STEC), and contaminated UPM has caused STEC outbreaks worldwide. The association between STEC, carrying the eae (E. coli attachment effacement) gene, and severe diseases is well-established. We have previously isolated four eae positive STEC isolates from two neighboring dairy farms in the Southeast of Norway. A whole genome analysis revealed that isolates from different farms exhibited nearly identical genetic profiles. To explore the risks associated with drinking UPM, we examined the ability of the isolates to produce Stx and their growth in UPM at different temperatures. All the isolates produced Stx and one of the isolates was able to propagate in UPM at 8 °C (p < 0.02). Altogether, these results highlight the risk for STEC infections associated with the consumption of UPM.

Occurrence of faecal endoparasites in reindeer (Rangifer tarandus) in two grazing areas in northern Norway.

BACKGROUND: Semi-domesticated reindeer represent an important livestock industry and livelihood for a proportion of the human population in northern Fennoscandia. Reindeer husbandry is considered an extensive animal husbandry, where the animals are kept mostly on natural pastures, although sometimes kept in fenced areas for shorter periods. These reindeer may harbour a variety of parasites that may affect animal health and production. The relatively limited close contact between herds and owners gives limited opportunities for diagnosis and treatment of diseases in general. Furthermore, the effects of subclinical parasitism in livestock are commonly expressed as a reduction in productivity rather than clinical disease and mortality. Thus, specific knowledge of endoparasites and parasitic infections in these herds is scarce. This study investigated the occurrence of various endoparasites in reindeer by analysis of a total of 114 faecal samples from winter-slaughtered reindeer from two different grazing areas in Troms and Finnmark, Norway. RESULTS: Using a McMaster method, a Baermann technique, and a direct immunofluorescent antibody test, the following parasites were identified in the faecal samples with the occurrence data given as percentages: Strongylid eggs (62%), Nematodirinae spp. eggs (24%), Capillaria sp. eggs (42%) and Moniezia spp. eggs (17%), Dictyocaulus spp. larvae (14%) protostrongylid larvae (40%), Eimera spp. oocysts (23%), and Giardia duodenalis cysts (5%). Cryptosporidium oocysts were not detected. Parasite eggs, oocysts, and cysts were quantified. Molecular analysis revealed G. duodenalis sub-assemblage AI, a potentially zoonotic genotype not previously reported in reindeer from this region. Morphological analyses of Eimeria oocysts identified two species, Eimeria mayeri and Eimeria rangiferis, and molecular analyses of the cytochrome C oxidase I (coi) gene and 18 s rRNA (18SSU) gene of Eimeria confirmed the presence of Eimeria species that are specific to reindeer. CONCLUSIONS: A high prevalence, but low burden, of endoparasites was detected in samples from these semi-domesticated reindeer. The samples were collected during winter, when adult gastrointestinal parasites usually produce only low numbers of transmission stages. Therefore, together with the low number of samples, detailed and definitive conclusions regarding parasite status of semi-domesticated reindeer are avoided. Nevertheless, these data provide a snapshot overview of parasite occurrence in a semi-domesticated animal group vulnerable to the various environmental changes to which they are exposed.