Subtyping of Listeria monocytogenes on the basis of plasmid profiles and arsenic and cadmium susceptibility

Brochothrix thermosphacta is a dominant but poorly studied meat spoilage organism. It is a close relative of the foodborne pathogen Listeria monocytogenes, and Brochothrix constitutes the second genus in the Listeriaceae family. Here, the genomes of 12 B. thermosphacta strains were sequenced, assembled into draft genomes, characterized, and compared with the genomes of Brochothrix campestris and L. monocytogenes. Phenotypic properties including biogenic amine production and antibiotic and heavy metal susceptibilities were tested. Comparative genomic analyses revealed a high degree of similarity among the B. thermosphacta strains, with bacteriophage genes constituting a significant proportion of the accessory genome. Genes for the production of the malodorous compounds acetate, acetoin, butanediol, and fatty acids were found, as were stress response regulatory genes, which likely play important roles in the spoilage process. Amino acid decarboxylases were not identified in the genomes, and phenotypic testing confirmed their absence. Orthologs of Listeria virulence proteins involved in virulence regulation, intracellular survival, and surface protein anchoring were found; however, key virulence genes were absent. Analysis of antibiotic susceptibility showed that strains were sensitive to the four tested antibiotics, except for one tetracycline-resistant isolate with plasmid-mediated tetracycline resistance genes. Strains tolerated higher levels of copper and cobalt than of cadmium although not at concentrations high enough to categorize the strains as being resistant. This study provides insight into the Brochothrix genome, links previous phenotypic data and data provided here to the gene inventory, and identifies genes that may contribute to the persistence of this organism in the food chain.IMPORTANCE Despite increasing knowledge and advances in food preservation techniques, microbial spoilage of foods causes substantial losses, with negative social and economic consequences. To better control the contamination and microbial spoilage of foods, fundamental knowledge of the biology of key spoilage bacteria is crucial. As a common meat spoilage organism, B. thermosphacta contributes substantially to spoilage-associated losses. Nonetheless, this organism and particularly its genome remain largely unstudied. This study contributes to improving our knowledge of the Brochothrix genus. Spoilage-relevant pathways and genes that may play a role in the survival of this organism in a food processing environment were identified, linking previous phenotypic data and data provided here to the gene inventory of Brochothrix and establishing parallels to and differences from the closely related foodborne pathogen L. monocytogenes.KEYWORDS Brochothrix, Listeria monocytogenes, meat spoilage, spoilage B rochothrix thermosphacta is one of the most abundant spoilage organisms of fresh and cured meats, fish, and fish products, due to its tolerance to high-salt and low-pH conditions, its ability to grow at refrigeration tempera...

Section: Resultsmentioning

confidence: 91%

Insight into the Genome of Brochothrix thermosphacta, a Problematic Meat Spoilage Bacterium

Stanborough

Fegan

Powell

et al. 2017

“…To determine resistance to cadmium, 3 l of the suspension was spotted in duplicate onto isosensitest agar (ISA) (Oxoid, Basingstoke, England) (control) and ISA containing 70 g/ml cadmium chloride anhydrous (Sigma, St. Louis, MO). The medium and concentration were based on those employed by McLauchlin et al (75 g/ml of cadmium chloride monohydrate) (20). For determination of resistance to arsenic, the cell suspension (3 l) was spotted in duplicate on ISA containing 500 g/ml sodium arsenite (Fluka, Buchs, Steinheim, Germany) (20).…”

Section: Methodsmentioning

confidence: 99%

“…80-kb) plasmid (7), identified through the genome sequencing of one of the strains, H7858 (23). Resistance to heavy metals, especially cadmium and arsenic, has been extensively used for strain-subtyping purposes in L. monocytogenes (10,20,31), and cadmium resistance has been found to be associated with a transposon (Tn5422) commonly harbored on plasmids (16, 18). However, no information has been available on possible associations between BC resistance and resistance to heavy metals in L. monocytogenes from the processing-plant environment.…”

mentioning

confidence: 99%

Heavy-Metal and Benzalkonium Chloride Resistance of Listeria monocytogenes Isolates from the Environment of Turkey-Processing Plants

Mullapudi

Siletzky

Kathariou

2008

115

The resistance of Listeria monocytogenes to cadmium and arsenic has been used extensively for strain subtyping. However, limited information is available on the prevalence of such resistance among isolates from the environment of food-processing plants. In addition, it is not known whether the resistance of such isolates to heavy metals may correlate with resistance to quaternary ammonium compounds extensively used as disinfectants in the food-processing industry. In this study, we characterized 192 L. monocytogenes isolates (123 putative strains) from the environment of turkey-processing plants in the United States for resistance to cadmium and arsenic and to the quaternary ammonium disinfectant benzalkonium chloride (BC). Resistance to cadmium was significantly more prevalent among strains of serotypes 1/2a (or 3a) and 1/2b (or 3b) (83% and 74%, respectively) than among strains of the serotype 4b complex (19%). Resistance to BC was encountered among 60% and 51% of the serotype 1/2a (or 3a) and 1/2b (or 3b) strains, respectively, and among 7% of the strains of the serotype 4b complex. All BC-resistant strains were also resistant to cadmium, although the reverse was not always the case. In contrast, no correlation was found between BC resistance and resistance to arsenic, which overall was low (6%). Our findings suggest that the processing environment of turkeyprocessing plants may constitute a reservoir for L. monocytogenes harboring resistance to cadmium and to BC and raise the possibility of common genetic elements or mechanisms mediating resistance to quaternary ammonium disinfectants and to cadmium in L. monocytogenes.Listeria monocytogenes causes listeriosis, a relatively rare but serious food-borne disease with symptoms such as stillbirths or abortions, septicemia, and meningitis or encephalitis. It has a high mortality rate (ca. 20%) for at-risk populations, which include pregnant women and their fetuses, neonates, and those who are elderly and/or immunocompromised (25). Strains of three serotypes (1/2a, 1/2b, and 4b) are associated with most human listeriosis cases. A substantial fraction of sporadic cases and the majority of food-borne outbreaks are due to serotype 4b strains (12,25). However, strains of serotype 4b tend to be underrepresented in foods, and the majority of food isolates are of serotype 1/2a or 1/2b (11,14,27). The environment of food-processing plants is considered to play a key role in contamination of processed ready-to-eat foods, and substantial evidence points to the ability of certain strains to colonize the processing-plant environment and to persist there, frequently over extended periods of time (for reviews, see references 12 and 13).Benzalkonium chloride (BC) and other quaternary ammonium compounds (QACs) are extensively used as disinfectants in the food-processing industry (19,22). Frequent use of BC and other QACs in food-processing plants may constitute a selective pressure for the emergence and establishment of resistance to these compounds among L. monocytogenes isolates...

“…, and 8 g ml Ϫ1 , as previously described (42,69). Isolates were classified as resistant (r superscript) or sensitive (s superscript) to each compound, and results were combined to generate a unique resistance profile (e.g., if one isolate was resistant to cadmium and sensitive to arsenic and tetracycline, the results were expressed as Cd r As s Tet s ).…”

mentioning

confidence: 99%

Diverse Geno- and Phenotypes of Persistent Listeria monocytogenes Isolates from Fermented Meat Sausage Production Facilities in Portugal

Ferreira

Barbosa

Stasiewicz

et al. 2011

The persistence of Listeria monocytogenes in food-associated environments represents a key factor in transmission of this pathogen. To identify persistent and transient strains associated with production of fermented meat sausages in northern Portugal, 1,723 L. monocytogenes isolates from raw material and finished products from 11 processors were initially characterized by random amplification of polymorphic DNA (RAPD), PCR-based molecular serotyping, and epidemic clone characterization, as well as cadmium, arsenic, and tetracycline resistance typing. Pulsed-field gel electrophoresis (PFGE) typing of 240 representative isolates provided evidence for persistence of L. monocytogenes for periods of time ranging from 10 to 32 months for all seven processors for which isolates from different production dates were available. Among 50 L. monocytogenes isolates that included one representative for each PFGE pattern obtained from a given sample, 12 isolates showed reduced invasion efficiency in Caco-2 cells, including 8 isolates with premature stop codons in inlA. Among 41 isolates representing sporadic and persistent PFGE types, 22 isolates represented lysogens. Neither strains with reduced invasion nor lysogens were overrepresented among persistent isolates. While the susceptibility of isolates to lysogenic phages also did not correlate with persistence, it appeared to be associated with molecular serotype. Our data show the following.