Polar-Region Soils as Novel Reservoir of Lactic Acid Bacteria from the Genus Carnobacterium.

Polar habitats offer excellent sites to isolate unique bacterial strains due to their diverse physical, geochemical, and biological factors. We hypothesize that the unique environmental conditions of polar regions select for distinct strains of lactic acid bacteria (LAB) with novel biochemical properties. In this study, we characterized ten strains of psychrotrophic LAB isolated from hitherto poorly described sources-High Arctic and maritime Antarctic soils and soil-like materials, including ornithogenic soils, cryoconites, elephant seal colonies, and postglacial moraines. We evaluated the physiological and biochemical properties of the isolates. Based on 16S rRNA and housekeeping genes, the four LAB strains were assigned to three Carnobacterium species: C. alterfunditum, C. maltaromaticum, and C. jeotgali. The remaining strains may represent three new species of the Carnobacterium genus. All isolates were neutrophilic and halophilic psychrotrophs capable of fermenting various carbohydrates, organic acids, and alcohols. The identified metabolic properties of the isolated Carnobacterium strains suggest possible syntrophic interactions with other microorganisms in polar habitats. Some showed antimicrobial activity against food pathogens such as Listeria monocytogenes and human pathogens like Staphylococcus spp. Several isolates exhibited unique metabolic traits with potential biotechnological applications that could be more effectively exploited under less stringent technological conditions compared to thermophilic LAB strains, such as lower temperatures and reduced nutrient concentrations. Analysis of extrachromosomal genetic elements revealed 13 plasmids ranging from 4.5 to 79.5 kb in five isolates, featuring unique genetic structures and high levels of previously uncharacterized genes. This work is the first comprehensive study of the biochemical properties of both known and new Carnobacterium species and enhances our understanding of bacterial communities in harsh and highly selective polar soil ecosystems.

Carnobacterium as a bioprotective and potential probiotic culture to improve food quality, food safety, and human health - a scoping review.

It is well-known that some bacteria can promote human and animal health. Bacteria of the genus Carnobacterium, while underexplored, have demonstrated significant probiotic and bioprotective potential. In this review, the recent scientific advances in this area are discussed. There are several requirements for a strain to be considered a probiotic or bioprotective agent, including the absence of antimicrobial resistance and the ability to colonize the gastrointestinal tract. Several researchers have reported such features in Carnobacterium bacteria, especially with regard to the production of antimicrobial substances. Research into animal production has advanced, especially in the aquaculture field, wherein inhibitory activity has been demonstrated against several important pathogens (for example Vibrio), and improvement in zootechnical indexes is evident. With respect to human health-related applications, research is still in the early stages. However, excellent in vitro results against pathogens, such as Candida albicans and Pseudomonas aeruginosa, have been reported. Carnobacterium bacteria have been assessed for a variety of applications in food, including direct application to the matrix and application to smart packaging, with proven effectiveness against Listeria monocytogenes. However, there is a lack of in vivo studies on Carnobacterium applications, which hinders its applications in various industries despite its high potential.

Identification of novel Carnobacterium maltaromaticum strains in bone marrow samples of patients with acute myeloid leukemia using a metagenomic binning approach.

The aim of this study aimed to examine the existence of a bacterial metagenome in the bone marrow of patients with acute myeloid leukemia (AML). We re-examined whole-genome sequencing data from the bone marrow samples of seven patients with AML, four of whom were remitted after treatment, for metagenomic analysis. After the removal of human reads, unmapped reads were used to profile the species-level composition. We used the metagenomic binning approach to confirm whether the identified taxon was a complete genome of known or novel strains. We observed a unique and novel microbial signature in which Carnobacterium maltaromaticum was the most abundant species in five patients with AML or remission. The complete genome of C. maltaromaticum "BMAML_KR01," which was observed in all samples, was 100% complete with 8.5% contamination and closely clustered with C. maltaromaticum strains DSM20730 and SF668 based on single nucleotide polymorphism variations. We identified five unique proteins that could contribute to cancer progression and 104 virulent factor proteins in the BMAML_KR01 genome. To our knowledge, this is the first report of a new strain of C. maltaromaticum in patients with AML. The presence of C. maltaromaticum and its new strain in patients indicates an urgent need to validate the existence of this bacterium and evaluate its pathophysiological role.

First isolation of Carnobacterium maltaromaticum from farmed Rainbow Trout in Virginia.

OBJECTIVE: Carnobacterium maltaromaticum is considered an emerging pathogen of salmonids in the United States and around the world. METHODS: Bacterial cultures obtained from the posterior kidney and skin of moribund Rainbow Trout Oncorhynchus mykiss from a commercial aquaculture facility in Virginia, USA, grew C. maltaromaticum, which was confirmed by additional phenotypic and molecular characterization. RESULT: A presumptive diagnosis based on the clinical signs, necropsy observations, histopathology, and bacterial cultures was bacterial septicemia due to C. maltaromaticum. CONCLUSION: This represents the first documentation of C. maltaromaticum in Rainbow Trout from Virginia.

Characterization of novel bacteriocin produced by bacteriocinogenic Carnobacterium maltaromaticum isolated from raw milk.

This study was designed to assess newly isolated bacteriocin-producing strain as potential food preservative. A bacteriocin producing lactic acid bacterium, named Carnobacterium maltaromatium KCA018, was screened from raw milk using deferred and spot-on-the-lawn assays. The crude cell free supernatant (CFS) was purified to obtain proteinaceous bacteriocin by ammonium sulfate precipitation (assigned as bacteriocin KCA) and tested for bacteriocin production, physical stability, antimicrobial activity, and bacteriocin-encoding gene detection. The growth curves of C. maltaromatium KCA018 reached late exponential phase after 15 h of incubation at 25 °C and 30 °C (Fig. 2). The maximum production of bacteriocin KCA was reached after 12 h of incubation at 25 °C, showing the antimicrobial activity of more than 3000 AU/ml against Listeria monocytogenes. The purified bacteriocin KCA was stable up to 67 °C for 30 min of exposure and between pH 4 and 7, showing more than 6000 AU/ml. The antibacterial activity of bacteriocin KCA was lost in the presence of pronase, proteinase K, and trypsin. Purified bacteriocin KCA showed higher antibacterial activity against Gram-positive bacteria than against Gram-negative bacteria. The CFS and purified bacteriocin KCA effectively inhibited the growth of L. monocytogenes ATCC 1911, E. faecalis ATCC 19433, and E. feacium ATCC 11576. The molecular weight of purified bacteriocin KCA was estimated at approximately 5 kDa. The positive amplification was observed for pisA and cbnBM1 with approximately between 100 and 200 bp. The newly identified bacteriocin can be a promising preservative for application in food.

Characterization of bacterial communities in Coregonus peled fillets during chilled storage and interactions between selected bacterial strains.

AIM: Coregonus peled fillets were used as a model to evaluate the dominant bacterial growth of chilled fish during storage after shipping and interactions of selected bacterial strains. METHODS AND RESULTS: Coregonus peled fillets were transported by air and land in ice boxes about 48 h from aquatic products company in Xinjiang, China, to the laboratory located in Dalian, China. Both culture-dependent (plate counts on nonselective media) based on 16S rRNA gene sequencing and culture-independent (Illumina-MiSeq high-throughput sequencing) methods were used. To detect interactions among bacterial populations from chilled fish, the influence of 18 test strains on the growth of 12 indicator isolates was measured by a drop assay and in liquid culture medium broth. The results showed that bacterial counts exceeded 7.0 log CFU/g following storage for 4 days at 4 °C. When the bacterial counts exceeded 8.5 log CFU/g after 12 days, the predominant micro-organisms were Aeromonas, Pseudomonas, Carnobacterium, Psychrobacter and Shewanella, as measured by the culture-independent method. All test strains showed inhibiting effects on the growth of other strains in liquid culture. Pseudomonas isolates showed antibacterial activity for approximately 60% of the indicator strains on nutritional agar plates. The majority of test isolates enhancing indicator strain growth were the strains isolated on day 0. CONCLUSIONS: High-throughput sequencing approach gives whole picture of bacterial communities in chilled C. peled fillets during storage, while growth interferences between selected bacterial strains illustrate the complexity of microbial interactions. SIGNIFICANCE AND IMPACT OF THE STUDY: We determined the bacterial communities and growth interferences in chilled Coregonus peled after shipping and these are the first data concerning microbiota in C. peled using a culture-independent analysis. The present study will be useful for manufacture and preservation of C. peled products by providing with valuable information regarding microbiological spoilage of C. peled.

Carnobacterium maltaromaticum associated with meningoencephalitis and otitis in stranded common thresher sharks (Alopias vulpinus).

Juvenile common thresher sharks (Alopias vulpinus) have been recently stranding along the California coastline. Using Illumina sequencing of the bacterial 16S rRNA gene along with necropsy, cytological, bacteriological, and histological techniques, we screened microbial communities and described lesions characterizing affected sharks with the purpose of identifying potential pathogen sources and pathologic processes. Histopathological assessment of moribund sharks revealed severe meningoencephalitis, as previously described in stranded salmon sharks (Lamna ditropis), along with inflammation of the inner ear and subcutaneous tissues surrounding the endolymphatic ducts. Furthermore, inflamed areas were characterized by the prevalence of Carnobacterium maltaromaticum, suggesting this bacterium as a potential pathogen that gains access to the inner ear through the endolymphatic ducts, with subsequent spread into the brain. The absence or low abundance of this bacterium in the spiral valve in both healthy and infected sharks suggests that Carnobacterium is not a commensal member of their digestive communities and the spiral valve is unlikely to be the source of the pathogen. Furthermore, phylogenetic analysis suggests that C. maltaromaticum strains isolated from diseased sharks have minimal genetic variation and differ from other strains originating from food or diseased teleosts. While a C. maltaromaticum-like organism has previously been associated with meningoencephalitis in salmon shark strandings, this is the first study to report common thresher shark strandings associated with C. maltaromaticum, involving the endolymphatic ducts as portals of entry to the brain.

Development of duplex qPCR targeting Carnobacterium maltaromaticum and Vagococcus salmoninarum.

In November 2018, Vagococcus salmoninarum was identified as the causative agent of a chronic coldwater streptococcosis epizootic in broodstock brook trout (Salvelinus fontinalis) at the Iron River National Fish Hatchery in Wisconsin, USA. By February 2019, the epizootic spread to adjacent raceways containing broodstock lake trout (Salvelinus namaycush), whereby fish were found to be coinfected with Carnobacterium maltaromaticum and V. salmoninarum. To differentiate these two pathogens and determine the primary cause of the lake trout morbidity, a quantitative real-time PCR (qPCR) was developed targeting the C. maltaromaticum phenylalanyl-tRNA synthase alpha subunit (pheS) gene. The qPCR was combined with a V. salmoninarum qPCR, creating a duplex qPCR assay that simultaneously quantitates C. maltaromaticum and V. salmoninarum concentrations in individual lake trout tissues, and screens presumptive isolates from hatchery inspections and wild fish from national fish hatchery source waters throughout the Great Lakes basin. Vagococcus salmoninarum and C. maltaromaticum were co-detected in broodstock brook trout from two tribal hatcheries and C. maltaromaticum was present in wild fish in source waters of several national fish hatcheries. This study provides a powerful new tool to differentiate and diagnose two emerging Gram-positive bacterial pathogens.

Carnobacterium inhibens isolated in blood culture of an immunocompromised, metastatic cancer patient: a case report and literature review.

BACKGROUND: Carnobacterium species are lactic acid-producing Gram-positive bacteria that have been approved by the US Food and Drug Administration and Health Canada for use as a food bio-preservative. The use of live bacteria as a food additive and its potential risk of infections in immunocompromised patients are not well understood. CASE PRESENTATION: An 81-year-old male with a history of metastatic prostate cancer on androgen deprivation therapy and chronic steroids presented to our hospital with a 2-week history of productive cough, dyspnea, altered mentation, and fever. Extensive computed tomography imaging revealed multifocal pneumonia without other foci of infection. He was diagnosed with pneumonia and empirically treated with ceftriaxone and vancomycin. Blood cultures from admission later returned positive for Carnobacterium inhibens. He achieved clinical recovery with step-down to oral amoxicillin/clavulanic acid for a total 7-day course of antibiotics. CONCLUSIONS: This is the fourth reported case of bacteremia with Carnobacterium spp. isolated from humans. This case highlights the need to better understand the pathogenicity and disease spectrum of bacteria used in the food industry for bio-preservation, especially in immunocompromised patients.

Correlation between chemical composition and antimicrobial properties of essential oils against most common food pathogens and spoilers: In-vitro efficacy and predictive modelling.

Using disk diffusion assay and broth microdilution, we evaluated the antimicrobial activity of 38 commercially available essential oils (EOs) against 24 food pathogens and spoilers. These including E. coli O157: H7 (3 types), Listeria (3 types), Bacillus (2 types), Salmonella enterica (2 types), Staphylococcus aureus (3 types), Clostridium tyrobutiricum, Pseudomonas aeruginosa, Brochotrix thermosphacta, Campylobacter jejuni, Carnobacterium divergens, Aspergillus (2 types), and Penicillium (4 types). Correlation between EOs' chemical composition and antimicrobial properties was studied using R software. Moreover, statistical models representing the relationship were generated using Design Expert®. The predictive models identified the chemical attributes of EOs that drive their antimicrobial properties while providing an understanding of their interactions. Thyme (Aldrich, Novotaste), cinnamon (Aliksir, BSA), garlic (Novotaste), Mexican garlic blend N & A (Novotaste), and oregano (BSA) were the strongest antimicrobial. The most sensitive pathogens were P. solitum (MIC of 19.53 ppm) and L. monocytogenes (MIC of 39 ppm). The correlation analysis showed that phenols and aldehydes had the strongest positive effects on the antimicrobial properties followed by the sulfur containing compounds and the esters; while the effects of monoterpenes and ketones were negative. Different sensitivity of food pathogens to chemical families was observed. For instance, phenols and aldehydes exhibited a linear inhibitory effect on L. monocytogenes (LM1045, MIC), while sesquiterpene and ester showed a significant effect on S. aureus (ATCC 6538, MIC). The developed predictive models are expected to predict the antimicrobial properties based on the chemical families of essential oils.

Metatranscriptomic analysis of modified atmosphere packaged poultry meat enables prediction of Brochothrix thermosphacta and Carnobacterium divergens in situ metabolism.

In this study, in situ-expressed metabolic routes of Brochothrix (B.) thermosphacta and Carnobacterium (C.) divergens were evaluated based on a metatranscriptomic dataset from bacteria growing on MAP chicken meat (O2/CO2; N2/CO2). Both species exhibited no (C. divergens) or minor transcription regulation (B. thermosphacta) within their main metabolic routes in response to different atmospheres. Both employ pathways related to glucose and ribose. Gluconeogenesis from lipid-borne glycerol is active in the progressing lack of carbohydrates. Pyruvate fates in both species comprise lactate, ethanol, acetate, CO2, formate, C4-compounds and H2O2 (only B. thermosphacta). Both species express genes for a minimal aerobic respiratory chain, but do not possess the genetic setting for a functional citric acid cycle. While products of carbohydrate and glycerol metabolism display mild to medium sensorial off-characteristics, predicted end products of their amino acid metabolism comprise, e.g., isobutyrate and isovalerate (B. thermosphacta) or cadaverine and tyramine (C. divergens) as potent spoilage compounds.

Preliminary investigation of an idiopathic muscle disease in farmed burbot Lota lota.

The rearing of less established fish species in recirculating aquaculture systems (RASs) is increasing, but may require adaptations of the rearing facilities if health impairments occur. We observed several health issues in burbot Lota lota reared for up to 2 yr in a RAS and used microbiological, histological and molecular-biological methods to identify the causative agents. Minor skin trauma led to the development of ulcers. In addition, several fillets of burbot showed pronounced granulomatous inflammation and calcification with signs of muscle fiber degeneration which resembled a condition called 'sandy flesh disease' in North American walleye. Several infectious agents were able to be excluded as a cause for the disease. Carnobacterium maltaromaticum was isolated in high numbers in some of the affected muscle tissue. However, the role of this bacterium or other causative agents or husbandry conditions remains to be elucidated.

Effect of glucose, pH and lactic acid on Carnobacterium maltaromaticum, Brochothrix thermosphacta and Serratia liquefaciens within a commercial heat-shrunk vacuum-package film.

Carnobacterium maltaromaticum, Brochothrix thermosphacta and Serratia liquefaciens are common spoilage organisms found within the microbiome of refrigerated vacuum-packaged (VP) beef. Extending and predicting VP beef shelf-life requires knowledge about how spoilage bacteria growth is influenced by environmental extrinsic and intrinsic factors. Multifactorial effects of pH, lactic acid (LA) and glucose on growth kinetics were quantified for C. maltaromaticum, B. thermosphacta and S. liquefaciens within a heat shrink-wrapped VP commercial film containing a simulated beef medium. LA, pH, and undissociated lactic acid (UDLA) significantly affected bacterial growth rate (p < 0.001), whereas 5.55 mM glucose produced a marginal effect. At 1.12 mM UDLA, growth rate and maximum population density decreased 20.9 and 3.5%, 56 and 7%, and 11 and 2% for C. maltaromaticum, B. thermosphacta, and S. liquefaciens, respectively.

Genome-wide comparison of Carnobacterium maltaromaticum derived from diseased fish harbouring important virulence-related genes.

Although Carnobacterium maltaromaticum has been used as a probiotic in fish, it was reported to cause disease for the first time in Korea. The objective of this study was to understand the differences between pathogenic and non-pathogenic strains. Pathogenicity was tested by challenging rainbow trout with C. maltaromaticum ATCC35586 and 18ISCm isolated from diseased fish, and DSM20342 isolated from a dairy product. We also compared 24 genomes of C. maltaromaticum strains plus the genome of our isolate 18ISCm sequenced in this study. Only the strains from diseased fish caused high mortality with severe histopathological changes. Although all strains shared more than 90% of Ko_id, wecC and xtmA were found only in strains from diseased fish. Interestingly, only strains from diseased fish harboured two wecC paralogs involved in the production of D-mannosaminuronic acid which is a major component of a well-known virulence factor, teichuronic acid. Two wecC paralogs of 18ISCm were increased when they were co-cultured with trout blood cells, suggesting that wecC genes might play a role in virulence. The results of this study show that strains isolated from diseased fish are different from strains derived from food in terms of pathogenicity to fish and the presence of virulence-related genes.

Effect of milk bactofugation on the counts and diversity of thermoduric bacteria.

The objective of this work was to determine the effect of milk bactofugation on the counts and microbial diversity of mesophilic (MT), psychrotrophic (PT), and thermophilic (TT) thermoduric bacteria and its potential as a technological method to remove spoilage microorganisms resistant to pasteurization. Different batches of raw milk from 69 dairy farms divided into sets in 3 bulk tanks (A, B, C) were evaluated at different times during the technological process. As the raw milk was preheated (∼55°C) immediately before bactofugation (10,000 × g), the effect of bactofugation was estimated by comparing the counts in raw, preheated, and bactofuged milk. This centrifugation was sufficient to reduce the isolation of 88% of the MT in preheated milk. For PT, it was possible to verify a reduction of 72.5% in batch C. The TT were not recovered at higher detection limits (<5 cfu/mL). For diversity, 310 isolates were identified using a molecular approach; 15 species of contaminating thermoduric bacteria were identified from raw and preheated milk, and only 6 species were recovered in bactofuged milk. Only MT were recovered from the bactofuged milk, mainly the species Lysinibacillus fusiformis (61.7%) and Bacillus licheniformis (12.3%). Both species are known to be endospore-forming psychrotrophs and have proteolytic or lipolytic activity. The bactofugation of raw milk reduced the number of isolates of B. licheniformis, Bacillus toyonensis, Micrococcus aloeverae, and Aestuariimicrobium kwangyangense by 33, 43, 86, and 92%, respectively, and reduced the isolates of Macrococcus caseolyticus, Lysinibacillus varians, Carnobacterium divergens, Microbacterium hominis, Kocuria indica, Micrococcus yunnanensis, Gordonia paraffinivorans, Bacillus invictae, and Kocuria kristinae to undetectable levels. The results of this study indicate that bactofugation can be applied by the dairy industry to reduce pasteurization-resistant microorganisms in combination with prophylactic measures to prevent the contamination of raw milk by spores and vegetative forms of bacteria.

Complementary Antibacterial Effects of Bacteriocins and Organic Acids as Revealed by Comparative Analysis of Carnobacterium spp. from Meat.

Carnobacterium maltaromaticum and Carnobacterium divergens are often predominant in the microbiota of vacuum-packaged (VP) meats after prolonged storage at chiller temperatures, and more so in recent studies. We investigated the antibacterial activities of C. maltaromaticum and C. divergens (n = 31) from VP meats by phenotypic characterization and genomic analysis. Five strains showed antibacterial activities against Gram-positive bacteria in a spot-lawn assay, with C. maltaromaticum strains having an intergeneric and C. divergens strains an intrageneric inhibition spectrum. This inhibitory activity is correlated with the production of predicted bacteriocins, including carnobacteriocin B2 and carnolysin for C. maltaromaticum and divergicin A for C. divergens The supernatants of both species cultured in meat juice medium under anaerobic conditions retarded the growth of most Gram-positive and Gram-negative bacteria in broth assay in a strain-dependent manner. C. maltaromaticum and C. divergens produced formate and acetate but not lactate under VP meat-relevant conditions. The relative inhibitory activity by Carnobacterium strains was significantly correlated (P < 0.05) to the production of both acids. Genomic analysis revealed the presence of genes required for respiration in both species. In addition, two clusters of C. divergens have an average nucleotide identity below the cutoff value for species delineation and thus should be considered to be two subspecies. In conclusion, both bacteriocins and organic acids are factors contributing significantly to the antibacterial activity of C. maltaromaticum and C. divergens under VP meat-relevant conditions. A few Carnobacterium strains can be explored as protective cultures to extend the shelf life and improve the safety of VP meats.IMPORTANCE The results of this study demonstrated that both bacteriocins and organic acids are important factors contributing to the antibacterial activities of Carnobacterium from vacuum-packaged (VP) meats. This study demonstrated that formate and acetate are the key organic acids produced by Carnobacterium and demonstrated their association with the inhibitory activity of carnobacteria under VP meat-relevant storage conditions. The role of lactate, on the other hand, may not be as important as previously believed in the antimicrobial activities of Carnobacterium spp. on chilled VP meats. These findings advance our understanding of the physiology of Carnobacterium spp. to better explore their biopreservative properties for chilled VP meats.

Influence of culture conditions on the technological properties of Carnobacterium maltaromaticum CNCM I-3298 starters.

AIM: The aim of this study is to investigate the effect of a broad spectrum of culture conditions on the acidification activity and viability of Carnobacterium maltaromaticum CNCM I-3298, the main technological properties that determine the shelf-life of biological time-temperature integrator (TTI) labels. METHODS AND RESULTS: Cells were cultivated at different temperatures (20-37°C) and pH (6-9·5) according to a modified central composite design and harvested at increasing times up to 10 h of stationary phase. Acidification activity and viability of freeze-thawed concentrates were assessed in medium mimicking the biological label. Acidification activity was influenced by all three culture conditions, but pH and harvest time were the most influential. Viability was not significantly affected by the tested range of culture conditions. CONCLUSIONS: Carnobacterium maltaromaticum CNCM I-3298 must be cultivated at 20°C, pH 6 and harvested at the beginning of stationary phase to exhibit fastest acidification activities. However, if slower acidification activities are pursued, the recommended culture conditions are 30°C, pH 9·5 and a harvest time between 4-6 h of stationary phase. SIGNIFICANCE AND IMPACT OF THE STUDY: Quantifying the impact of fermentation temperature, pH and harvest time has led to a predictive model for the production of biological TTI covering a broad range of shelf-lives.

Multilocus sequence typing of Carnobacterium maltaromaticum strains associated with fish disease and dairy products.

AIMS: Carnobacterium maltaromaticum is a lactic acid bacterium of technological interest in the field of dairy ripening and food bioprotection and is generally recognized as safe in the United States. As it is associated with fish infections, the European Food Safety Agency did not include this species in the qualified presumption safety list of micro-organisms. This implies that the risk assessment for the species has to be performed at the strain level. METHODS AND RESULTS: Multilocus sequence typing (MLST) is a tool that (i) potentially allows to discriminate strains isolated from diseased fish from apathogenic strains and (ii) to assess the genetic relatedness between both groups of strains. In this study, we characterized by MLST 21 C. maltaromaticum strains including 16 strains isolated from diseased fish and 5 apathogenic dairy strains isolated from cheese. The resulting population structure was investigated by integrating these new data to the previously published population structure (available at http://pubmlst.org), which represents an overall of 71 strains. CONCLUSIONS: This analysis revealed that none of the strains isolated from diseased fish is assigned to a clonal complex containing cheese isolates, and that 11 strains exhibit singleton genotypes suggesting that the population of diseased fish isolates is not clonal. SIGNIFICANCE AND IMPACT OF THE STUDY: This study thus provides a population structure of C. maltaromaticum that could serve in the future as a reference that could contribute to the risk assessment of C. maltaromaticum strains intended to be used in the food chain.

Tracking the sources of psychrotrophic bacteria contaminating chicken cuts during processing.

The major aim of the study was to establish the routes via which spoilage associated psychrotrophic bacteria contaminate poultry products at a large processing plant located in Belgium. Environmental samples were collected consisting of samples of air and swabs of food contact surfaces. Product samples were also collected consisting of modified atmosphere packaged (MAP) chicken wings and legs, which were analyzed microbiologically on the same day they were produced as well as after their sell-by date. Psychrotrophic bacteria from these samples were subsequently clustered and identified by means of MALDI-TOF MS and 16S rRNA gene sequencing. Carnobacterium maltaromaticum was determined to dominate the spoilage flora of both wings and legs. Other psychrotrophic bacteria able to grow on MRS which were identified on expired wings and legs included Carnobacterium divergens, Brocothrix thermosphacta, Lactobacillus curvatus, and Lactobacillus brevis. These were determined to arise from food contact surfaces such as cutting blades, leg hooks, Ertalon and polyurethane conveyor belts, working tables, and the hands of the operators. Importantly, it was determined that cleaning and disinfection was largely inadequate. Air was also determined to be an important vector of psychrotrophic bacteria in the processing environment, potentially contaminating the products directly or indirectly.

Carnobacterium antarcticum sp. nov., a psychrotolerant, alkaliphilic bacterium isolated from sandy soil in Antarctica.

A novel, alkaliphilic, psychrotolerant, facultative anaerobe, designated CP1T, was isolated from sandy soil near the Davis Station in Antarctica. The short-rod-shaped cells displayed Gram-positive staining and did not form spores. Strain CP1T was able to grow at temperatures between 4 and 36 °C, pH 6.0-9.5, and in the presence of up to 5.0 % (w/v) NaCl. 16S rRNA gene and multilocus (pheS, rpoA, and atpA) sequence analysis revealed Carnobacterium mobile DSM 4848T and Carnobacterium iners LMG 26642T as the closest relatives (97.4 and 97.1 % 16S rRNA gene sequence similarity, respectively). The genomic G+C content was 38.1 mol%, and DNA-DNA hybridization with DSM 4848T revealed 32.4±3.4 % similarity. The major fatty acid components were C14 : 0 and C16 : 1ω9c. The cell wall contained meso-diaminopimelic acid and was of peptidoglycan type A1γ. Based on physiological, genotypic and biochemical characteristics, strain CP1T represents a novel species of the genus Carnobacterium for which the name Carnobacterium antarcticum sp. nov. is proposed. The type strain is CP1T (=DSM 103363T=CGMCC 1.15643T).