scholarly journals comKProphage Junction Fragments as Markers for Listeria monocytogenes Genotypes Unique to Individual Meat and Poultry Processing Plants and a Model for Rapid Niche-Specific Adaptation, Biofilm Formation, and Persistence

2011 ◽  
Vol 77 (10) ◽  
pp. 3279-3292 ◽  
Author(s):  
Bindhu Verghese ◽  
Mei Lok ◽  
Jia Wen ◽  
Valentina Alessandria ◽  
Yi Chen ◽  
...  
Keyword(s):  
Listeria Monocytogenes ◽  
Food Processing ◽  
Biofilm Formation ◽  
Content Type ◽  
Niche Adaptation ◽  
Processing Plants ◽  
Poultry Processing ◽  
Conditioning Film ◽  
Conditioning Films ◽  
Different Strains

ABSTRACTDifferent strains ofListeria monocytogenesare well known to persist in individual food processing plants and to contaminate foods for many years; however, the specific genotypic and phenotypic mechanisms responsible for persistence of these unique strains remain largely unknown. Based on sequences incomKprophage junction fragments, different strains of epidemic clones (ECs), which included ECII, ECIII, and ECV, were identified and shown to be specific to individual meat and poultry processing plants. ThecomKprophage-containing strains showed significantly higher cell densities after incubation at 30°C for 48 h on meat and poultry food-conditioning films than did strains lacking thecomKprophage (P< 0.05). Overall, the type of strain, the type of conditioning film, and the interaction between the two were all highly significant (P< 0.001). Recombination analysis indicated that thecomKprophage junction fragments in these strains had evolved due to extensive recombination. Based on the results of the present study, we propose a novel model in which the concept of defectivecomKprophage was replaced with the rapid adaptation island (RAI). Genes within the RAI were recharacterized as “adaptons,” as these genes may allowL. monocytogenesto rapidly adapt to different food processing facilities and foods. If confirmed, the model presented would help explainListeria's rapid niche adaptation, biofilm formation, persistence, and subsequent transmission to foods. Also,comKprophage junction fragment sequences may permit accurate tracking of persistent strains back to and within individual food processing operations and thus allow the design of more effective intervention strategies to reduce contamination and enhance food safety.

scholarly journals Biofilm Formation by Listeria monocytogenes 15G01, a Persistent Isolate from a Seafood-Processing Plant, Is Influenced by Inactivation of Multiple Genes Belonging to Different Functional Groups

2021 ◽  
Vol 87 (10) ◽  
Author(s):  
Jessika Nowak ◽  
Sandra B. Visnovsky ◽  
Andrew R. Pitman ◽  
Cristina D. Cruz ◽  
Jon Palmer ◽  
...  
Keyword(s):  
Listeria Monocytogenes ◽  
Food Processing ◽  
Biofilm Formation ◽  
Processing Plant ◽  
Wild Type ◽  
Content Type ◽  
Multiple Genes ◽  
Processing Plants ◽  
Biofilm Structure ◽  
Seafood Processing

ABSTRACT Listeria monocytogenes is a ubiquitous foodborne pathogen that results in a high rate of mortality in sensitive and immunocompromised people. Contamination of food with L. monocytogenes is thought to occur during food processing, most often as a result of the pathogen producing a biofilm that persists in the environment and acting as the source for subsequent dispersal of cells onto food. A survey of seafood-processing plants in New Zealand identified the persistent strain 15G01, which has a high capacity to form biofilms. In this study, a transposon library of L. monocytogenes 15G01 was screened for mutants with altered biofilm formation, assessed by a crystal violet assay, to identify genes involved in biofilm formation. This screen identified 36 transposants that showed a significant change in biofilm formation compared to the wild type. The insertion sites were in 27 genes, 20 of which led to decreased biofilm formation and seven to an increase. Two insertions were in intergenic regions. Annotation of the genes suggested that they are involved in diverse cellular processes, including stress response, autolysis, transporter systems, and cell wall/membrane synthesis. Analysis of the biofilms produced by the transposants using scanning electron microscopy and fluorescence microscopy showed notable differences in the structure of the biofilms compared to the wild type. In particular, inactivation of uvrB and mltD produced coccoid-shaped cells and elongated cells in long chains, respectively, and the mgtB mutant produced a unique biofilm with a sandwich structure which was reversed to the wild-type level upon magnesium addition. The mltD transposant was successfully complemented with the wild-type gene, whereas the phenotypes were not or only partially restored for the remaining mutants. IMPORTANCE The major source of contamination of food with Listeria monocytogenes is thought to be due to biofilm formation and/or persistence in food-processing plants. By establishing as a biofilm, L. monocytogenes cells become harder to eradicate due to their increased resistance to environmental threats. Understanding the genes involved in biofilm formation and their influence on biofilm structure will help identify new ways to eliminate harmful biofilms in food processing environments. To date, multiple genes have been identified as being involved in biofilm formation by L. monocytogenes; however, the exact mechanism remains unclear. This study identified four genes associated with biofilm formation by a persistent strain. Extensive microscopic analysis illustrated the effect of the disruption of mgtB, clsA, uvrB, and mltD and the influence of magnesium on the biofilm structure. The results strongly suggest an involvement in biofilm formation for the four genes and provide a basis for further studies to analyze gene regulation to assess the specific role of these biofilm-associated genes.

scholarly journals Persistent Listeria monocytogenes Isolates from a Poultry-Processing Facility Form More Biofilm but Do Not Have a Greater Resistance to Disinfectants than Sporadic Strains

Pathogens ◽  
2019 ◽  
Vol 8 (4) ◽  
pp. 250 ◽  
Author(s):  
Daniel Rodríguez-Campos ◽  
Cristina Rodríguez-Melcón ◽  
Carlos Alonso-Calleja ◽  
Rosa Capita
Keyword(s):  
Listeria Monocytogenes ◽  
Food Processing ◽  
Biofilm Formation ◽  
Benzalkonium Chloride ◽  
Bacterial Persistence ◽  
Crystal Violet Assay ◽  
Processing Facility ◽  
Poultry Processing ◽  
Sessile Cells ◽  
Selection Of

Some strains of Listeria monocytogenes can persist in food-processing environments, increasing the likelihood of the contamination of foodstuffs. To identify traits that contribute to bacterial persistence, a selection of persistent and sporadic L. monocytogenes isolates from a poultry-processing facility was investigated for biofilm-forming ability (crystal violet assay). The susceptibility of sessile cells to treatments (five minutes) with sodium hypochlorite having 10% active chlorine (SHY: 10,000 ppm, 25,000 ppm, and 50,000 ppm) and benzalkonium chloride (BZK: 2500 ppm, 10,000 ppm, and 25,000 ppm) was also studied. All isolates exhibited biofilm formation on polystyrene. Persistent strains showed larger (p < 0.001) biofilm formation (OD580 = 0.301 ± 0.097) than sporadic strains (OD580 = 0.188 ± 0.082). A greater susceptibility to disinfectants was observed for biofilms of persistent strains than for those of sporadic strains. The application of SHY reduced biofilms only for persistent strains. BZK increased OD580 in persistent strains (2500 ppm) and in sporadic strains (all concentrations). These results indicate that the use of BZK at the concentrations tested could represent a public health risk. Findings in this work suggest a link between persistence and biofilm formation, but do not support a relationship between persistence and the resistance of sessile cells to disinfectants.

scholarly journals Heavy Metal and Disinfectant Resistance of Listeria monocytogenes from Foods and Food Processing Plants

2012 ◽  
Vol 78 (19) ◽  
pp. 6938-6945 ◽  
Author(s):  
Shakir S. Ratani ◽  
Robin M. Siletzky ◽  
Vikrant Dutta ◽  
Suleyman Yildirim ◽  
Jason A. Osborne ◽  
...  
Keyword(s):  
Heavy Metals ◽  
Listeria Monocytogenes ◽  
Food Processing ◽  
Cadmium Resistance ◽  
Content Type ◽  
Ecological History ◽  
Resistance Determinants ◽  
Processing Plants ◽  
History Of ◽  
Serotype 4B

ABSTRACTThe persistence ofListeria monocytogenesin food processing plants and other ecosystems reflects its ability to adapt to numerous stresses. In this study, we investigated 138 isolates from foods and food processing plants for resistance to the quaternary ammonium disinfectant benzalkonium chloride (BC) and to heavy metals (cadmium and arsenic). We also determined the prevalence of distinct cadmium resistance determinants (cadA1,cadA2, andcadA3) among cadmium-resistant isolates. Most BC-resistant isolates were resistant to cadmium as well. Arsenic resistance was encountered primarily in serotype 4b and was an attribute of most isolates of the serotype 4b epidemic clonal group ECIa. Prevalence of the known cadmium resistance determinants was serotype associated:cadA1was more common in isolates of serotypes 1/2a and 1/2b than 4b, whilecadA2was more common in those of serotype 4b. A subset (15/77 [19%]) of the cadmium-resistant isolates lacked the known cadmium resistance determinants. Most of these isolates were of serotype 4b and were also resistant to arsenic, suggesting novel determinants that may confer resistance to both cadmium and arsenic in these serotype 4b strains. The findings may reflect previously unrecognized components of the ecological history of different serotypes and clonal groups ofL. monocytogenes, including exposures to heavy metals and disinfectants.

scholarly journals Conservation and Distribution of the Benzalkonium Chloride Resistance CassettebcrABCin Listeria monocytogenes

2013 ◽  
Vol 79 (19) ◽  
pp. 6067-6074 ◽  
Author(s):  
Vikrant Dutta ◽  
Driss Elhanafi ◽  
Sophia Kathariou
Keyword(s):  
Listeria Monocytogenes ◽  
Food Processing ◽  
Benzalkonium Chloride ◽  
Clinical Environment ◽  
Outbreak Strain ◽  
Content Type ◽  
Chloride Resistance ◽  
Processing Plants ◽  
Hot Dog ◽  
Widespread Dissemination

ABSTRACTAnalysis of a panel of 116Listeria monocytogenesstrains of diverse serotypes and sources (clinical, environment of food processing plants, and food) revealed that all but one of the 71 benzalkonium chloride-resistant (BCr) isolates harboredbcrABC, previously identified on a large plasmid (pLM80) of the 1998-1999 hot dog outbreak strain H7858. In contrast,bcrABCwas not detected among BC-susceptible (BCs) isolates. ThebcrABCsequences were highly conserved among strains of different serotypes, but variability was noted in sequences flankingbcrABC. The majority of the BCrisolates had either the pLM80-type of organization of thebcrABCregion or appeared to harborbcrABCon the chromosome, adjacent to novel sequences. Transcription ofbcrABCwas induced by BC (10 μg/ml) in strains of different serotypes and diversebcrABCregion organization. These findings reveal widespread dissemination ofbcrABCacross BCrL. monocytogenesstrains regardless of serotype and source, while also suggesting possible mechanisms ofbcrABCdissemination acrossL. monocytogenesgenomes.

Formation of Biofilm at Different Nutrient Levels by Various Genotypes of Listeria monocytogenes

2006 ◽  
Vol 69 (4) ◽  
pp. 826-834 ◽  
Author(s):  
JAMES P. FOLSOM ◽  
GREGORY R. SIRAGUSA ◽  
JOSEPH F. FRANK
Keyword(s):  
Listeria Monocytogenes ◽  
Food Processing ◽  
Hoechst 33258 ◽  
Genetic Subtype ◽  
Processing Plants ◽  
Serotype 1 ◽  
Serotype 4B ◽  
Different Strains ◽  
Element Sequence ◽  
Range Test

Strains of Listeria monocytogenes differ in their ability to form biofilms. The objectives of this study were to determine whether genetically related strains have similar biofilm-forming capacities and what effect nutrient concentration has on the ability of different strains to produce biofilms. Biofilms of 30 strains of L. monocytogenes, obtained from a variety of sources were grown on stainless steel in tryptic soy broth (TSB) or in a 1:10 dilution of TSB (DTSB) for 24 h at 32°C. The amount of biofilm formed was determined with image analysis after cells were stained with bisBenzimide H 33258 (Hoechst 33258). The strains were genetically subtyped by repetitive element sequence–based PCR (rep-PCR) with the primer set rep-PRODt and rep-PROG5. Data were analyzed with an analysis of variance and Duncan's multiple range test. Eleven strains produced the same amount of biofilm in both media. Fourteen strains produced more biofilm in TSB than in DTSB. Five strains produced more biofilm in DTSB than in TSB. Serotype 4b strains produced more biofilm in TSB than did serotype 1/2a strains, whereas serotype 1/2a strains produced more biofilm in DTSB than did serotype 4b strains. Growth in DTSB resulted in decreased biofilm accumulation for serotype 4b strains. There was no correlation between genetic subtype and the amount of biofilm accumulation. These results indicate that strains of serotype 1/2a and serotype 4b differ in the regulation of their biofilm phenotype. The poor biofilm accumulation of serotype 4b isolates when grown in DTSB could be a factor in the predominance of serogroup 1/2 strains in food processing plants, where nutrients may be limited.

scholarly journals Does Campylobacter jejuni Form Biofilms in Food-Related Environments?

2014 ◽  
Vol 80 (17) ◽  
pp. 5154-5160 ◽  
Author(s):  
Amy Huei Teen Teh ◽  
Sui Mae Lee ◽  
Gary A. Dykes
Keyword(s):  
Campylobacter Jejuni ◽  
Food Processing ◽  
Biofilm Formation ◽  
Food Production ◽  
Significant Risk ◽  
Primary Source ◽  
Content Type ◽  
Food Borne ◽  
Processing Plants ◽  
Atmosphere Temperature

ABSTRACTCampylobacter jejuniis one of the most frequent causes of bacterial gastrointestinal food-borne infection worldwide. This species is part of the normal flora of the gastrointestinal tracts of animals used for food production, including poultry, which is regarded as the primary source of humanCampylobacterinfections. The survival and persistence ofC. jejuniin food processing environments, especially in poultry processing plants, represent significant risk factors that contribute to the spread of this pathogen through the food chain. Compared to other food-borne pathogens,C. jejuniis more fastidious in its growth requirements and is very susceptible to various environmental stressors. Biofilm formation is suggested to play a significant role in the survival ofC. jejuniin the food production and processing environment. The aims of this minireview were (i) to examine the evidence thatC. jejuniforms biofilms and (ii) to establish the extent to which reported and largely laboratory-based studies ofC. jejunibiofilms provide evidence for biofilm formation by this pathogen in food processing environments. Overall existing studies do not provide strong evidence for biofilm formation (as usually defined) by mostC. jejunistrains in food-related environments under the combined conditions of atmosphere, temperature, and shear that they are likely to encounter. Simple attachment to and survival on surfaces and in existing biofilms of other species are far more likely to contribute toC. jejunisurvival in food-related environments based on our current understanding of this species.

scholarly journals Comparison of Listeria monocytogenes Exoproteomes from Biofilm and Planktonic State: Lmo2504, a Protein Associated with Biofilms

2013 ◽  
Vol 79 (19) ◽  
pp. 6075-6082 ◽  
Author(s):  
António Lourenço ◽  
Aitor de Las Heras ◽  
Mariela Scortti ◽  
Jose Vazquez-Boland ◽  
Joseph F. Frank ◽  
...  
Keyword(s):  
Listeria Monocytogenes ◽  
Food Processing ◽  
Biofilm Formation ◽  
Ruthenium Red ◽  
Matrix Remodeling ◽  
Wild Type ◽  
Content Type ◽  
Cell Wall Binding ◽  
Physiological Processes ◽  
Planktonic State

ABSTRACTThe food-borne pathogenListeria monocytogenesis the causative agent of the severe human and animal disease listeriosis. The persistence of this bacterium in food processing environments is mainly attributed to its ability to form biofilms. The search for proteins associated with biofilm formation is an issue of great interest, with most studies targeting the whole bacterial proteome. Nevertheless, exoproteins constitute an important class of molecules participating in various physiological processes, such as cell signaling, pathogenesis, and matrix remodeling. The aim of this work was to quantify differences in protein abundance between exoproteomes from a biofilm and from the planktonic state. For this, two field strains previously evaluated to be good biofilm producers (3119 and J311) were used, and a procedure for the recovery of biofilm exoproteins was optimized. Proteins were resolved by two-dimensional difference gel electrophoresis and identified by electrospray ionization-tandem mass spectrometry. One of the proteins identified in higher abundance in the biofilm exoproteomes of both strains was the putative cell wall binding protein Lmo2504. A mutant strain with deletion of the gene for Lmo2504 was produced (3119Δlmo2504), and its biofilm-forming ability was compared to that of the wild type using the crystal violet and the ruthenium red assays as well as scanning electron microscopy. The results confirmed the involvement of Lmo2504 in biofilm formation, as strain 3119Δlmo2504showed a significantly (P< 0.05) lower biofilm-forming ability than the wild type. The identification of additional exoproteins associated with biofilm formation may lead to new strategies for controlling this pathogen in food processing facilities.

scholarly journals The Connection between Persistent, Disinfectant-Resistant Listeria monocytogenes Strains from Two Geographically Separate Iberian Pork Processing Plants: Evidence from Comparative Genome Analysis

2015 ◽  
Vol 82 (1) ◽  
pp. 308-317 ◽  
Author(s):  
Sagrario Ortiz ◽  
Victoria López-Alonso ◽  
Pablo Rodríguez ◽  
Joaquín V. Martínez-Suárez
Keyword(s):  
Listeria Monocytogenes ◽  
Food Processing ◽  
Pfge Type ◽  
Processing Plant ◽  
Content Type ◽  
Pork Products ◽  
Processing Plants ◽  
Pork Product ◽  
Ammonium Compounds ◽  
Insight Into

ABSTRACTThe aim of this study was to investigate the basis of the putative persistence ofListeria monocytogenesin a new industrial facility dedicated to the processing of ready-to-eat (RTE) Iberian pork products. Quaternary ammonium compounds, which included benzalkonium chloride (BAC), were repeatedly used as surface disinfectants in the processing plant. Clean and disinfected surfaces were sampled to evaluate if resistance to disinfectants was associated with persistence. Of the 14 isolates obtained from product contact and non-product contact surfaces, only five different pulsed-field gel electrophoresis (PFGE) types were identified during the 27-month study period. Two of these PFGE types (S1 and S10-1) were previously identified to be persistent and BAC-resistant (BACr) strains in a geographically separate slaughterhouse belonging to the same company. The remaining three PFGE types, which were first identified in this study, were also BACr. Whole-genome sequencing andin silicomultilocus sequence typing (MLST) analysis of five BACrisolates of the different PFGE types identified in this study showed that the isolate of the S1 PFGE type belonged to MLST sequence type 31 (ST31), a low-virulence type characterized by mutations in theinlAandprfAgenes. The isolates of the remaining four PFGE types were found to belong to MLST ST121, a persistent type that has been isolated in several countries. The ST121 strains contained the BAC resistance transposon Tn6188. The disinfection-resistantL. monocytogenespopulation in this RTE pork product plant comprised two distinct genotypes with different multidrug resistance phenotypes. This work offers insight into theL. monocytogenessubtypes associated with persistence in food processing environments.

scholarly journals An Open-Source Program (Haplo-ST) for Whole-Genome Sequence Typing Shows Extensive Diversity among Listeria monocytogenes Isolates in Outdoor Environments and Poultry Processing Plants

2020 ◽  
Vol 87 (1) ◽  
Author(s):  
Swarnali Louha ◽  
Richard J. Meinersmann ◽  
Zaid Abdo ◽  
Mark E. Berrang ◽  
Travis C. Glenn
Keyword(s):  
Listeria Monocytogenes ◽  
Open Source ◽  
Strain Identification ◽  
Specific Gene ◽  
Whole Genome ◽  
Content Type ◽  
Open Source Tool ◽  
Processing Plants ◽  
Poultry Processing ◽  
Reference Strains

ABSTRACT A reliable and standardized classification of Listeria monocytogenes is important for accurate strain identification during outbreak investigations. Current whole-genome sequencing (WGS)-based approaches for strain characterization are either difficult to standardize, rendering them less suitable for data exchange, or are not freely available. Thus, we developed a portable and open-source tool, Haplo-ST, to improve standardization and provide maximum discriminatory potential to WGS data tied to a multilocus sequence typing (MLST) framework. Haplo-ST performs whole-genome MLST (wgMLST) for L. monocytogenes while allowing for data exchangeability worldwide. This tool takes in (i) raw WGS reads as input, (ii) cleans the raw data according to user-specified parameters, (iii) assembles genes across loci by mapping to genes from reference strains, and (iv) assigns allelic profiles to assembled genes and provides a wgMLST subtyping for each isolate. Data exchangeability relies on the tool assigning allelic profiles based on a centralized nomenclature defined by the widely used BIGSdb-Lm database. Tests of Haplo-ST’s performance with simulated reads from L. monocytogenes reference strains demonstrated high sensitivity (97.5%), and coverage depths of ≥20× were found to be sufficient for wgMLST profiling. We then used Haplo-ST to characterize and differentiate between two groups of L. monocytogenes isolates derived from the natural environment and poultry processing plants. Phylogenetic reconstruction identified lineages within each group, and no lineage specificity was observed with isolate phenotypes (transient versus persistent) or origins. Genetic differentiation analyses between isolate groups identified 21 significantly differentiated loci, potentially enriched for adaptation and persistence of L. monocytogenes within poultry processing plants. IMPORTANCE We have developed an open-source tool (https://github.com/swarnalilouha/Haplo-ST) that provides allele-based subtyping of L. monocytogenes isolates at the whole-genome level. Along with allelic profiles, this tool also generates allele sequences and identifies paralogs, which is useful for phylogenetic tree reconstruction and deciphering relationships between closely related isolates. More broadly, Haplo-ST is flexible and can be adapted to characterize the genome of any haploid organism simply by installing an organism-specific gene database. Haplo-ST also allows for scalable subtyping of isolates; fewer reference genes can be used for low-resolution typing, whereas higher resolution can be achieved by increasing the number of genes used in the analysis. Our tool enabled clustering of L. monocytogenes isolates into lineages and detection of potential loci for adaptation and persistence in food processing environments. Findings from these analyses highlight the effectiveness of Haplo-ST in subtyping and evaluating relationships among isolates in studies of bacterial population genetics.

Biofilm Formation and Associated Genes in Listeria Monocytogenes

2017 ◽  
Vol 12 (3) ◽  
Author(s):  
S. R. Warke ◽  
V. C. Ingle ◽  
N. V. Kurkure ◽  
P. A. Tembhurne ◽  
Minakshi Prasad ◽  
...  
Keyword(s):  
Public Health ◽  
Listeria Monocytogenes ◽  
Multiplex Pcr ◽  
Food Processing ◽  
Biofilm Formation ◽  
Milk Samples ◽  
Biofilm Production ◽  
Food Borne ◽  
Serious Infections ◽  
Microtiter Assay

Listeria monocytogenes, an opportunistic food borne pathogen can cause serious infections in immunocompromised individuals. L. monocytogenes is capable of producing biofilm on the surface of food processing lines and instruments.The biofilm transfers contamination to food products and impose risk to public health. In the present study biofilm producing ability of L. monocytogenes isolates were investigated phenotypically and genotypically by microtiter assay and multiplex PCR, respectively. Out of 38 L. monocytogenes isolates 14 were recovered from animal clinical cases, 12 bovine environment and 12 from milk samples. A total of 3 (21.42%) clinical, 2 (16.66%) environment and 3 (25%) milk samples respectively, revealed biofilm production in microtiter assay. Cumulative results showed that 23 (60.52%) out of 38 strains of L. monocytogenes were positive for luxS and flaA gene and 1 (2.63%) was positive only for the flaA gene.

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