Listeria Monocytogenes Isolates Research Articles

Oxford Nanopore's 2024 sequencing technology for Listeria monocytogenes outbreak detection and source attribution: progress and clone-specific challenges.

Whole genome sequencing is an essential cornerstone of pathogen surveillance and outbreak detection. Established sequencing technologies are currently being challenged by Oxford Nanopore Technologies (ONT), which offers an accessible and cost-effective alternative enabling gap-free assemblies of chromosomes and plasmids. Limited accuracy has hindered its use for investigating pathogen transmission, but recent technology updates have brought significant improvements. To evaluate its readiness for outbreak detection, we selected 78 Listeria monocytogenes isolates from diverse lineages or known epidemiological clusters for sequencing with ONT's V14 Rapid Barcoding Kit and R10.4.1 flow cells. The most accurate of several tested workflows generated assemblies with a median of one error (SNP or indel) per assembly. For 66 isolates, the cgMLST profiles from ONT-only assemblies were identical to those generated from Illumina data. Eight assemblies were of lower quality, with more than 20 erroneous sites each, primarily caused by methylations at the GAAGAC motif (5'-GAAG6mAC-3'/5'-GT4mCTTC-3'). This led to inaccurate clustering, failing to group isolates from a persistence-associated clone that carried the responsible restriction-modification system. Out of 50 methylation motifs detected among the 78 isolates, only the GAAGAC motif was linked to substantially increased error rates. Our study shows that most L. monocytogenes genomes assembled from ONT-only data are suitable for high-resolution genotyping, but further improvements of chemistries or basecallers are required for reliable routine use in outbreak and food safety investigations.

Nanozyme-catalyzed and zwitterion-modified swabs based for the detection of Listeria monocytogenes in complex matrices

In recent years, nanozymes have been widely used in the field of biosensing and food safety testing due to their advantages of low cost, high stability, easy modification and adjustable catalytic activity. However, how to reduce the signal interference generated by reducing substances, macromolecules and colored substances in the food matrix in nanozymes-based colorimetric sensing is still a major challenge. In this paper, using Listeria monocytogenes as a model analyte, sodium sulfonyl methacrylate (SBMA) polymers were modified onto cotton swabs by photothermal polymerization and combined with Listeria monocytogenes-specific aptamer (Apt1) to prepare swabs that can specifically capture and isolate Listeria monocytogenes from complex matrices (SBMA/Apt1 cotton swab). In addition, in combination with the inhibitory effect of the aptamer (Apt2) on the oxidase activity of Mn3O4 NPs, a colorimetric biosensor based on nanozymes that can quantitatively, sensitively, and specifically identify Listeria monocytogenes in food products was constructed. The results showed that the colorimetric signal of the method was linear with the concentration of Listeria monocytogenes in the range of 2.83–2.83 × 105 CFU/mL, and the limit of detection was 2.64 CFU/mL, which can be used for the detection of Listeria monocytogenes in complex environments and food samples.

Colorimetric aptasensor for Listeria monocytogenes detection using dual functional Fe3O4@MIL-100(Fe) with magnetic separation and oxidase-like activities in food samples.

A novel colorimetric aptasensor assay based on the excellent magnetic responsiveness and oxidase-like activity of Fe3O4@MIL-100(Fe) was developed. Fe3O4@MIL-100(Fe) absorbed with aptamer and blocked by BSA served as capture probe for selective isolation and enrichment of Listeria monocytogenesone of the most common and dangerous foodborne pathogenic bacteria. The aptamer absorbed on Fe3O4@MIL-100(Fe) was further used as signal probe that specifically binds with target bacteria conjugation of capture probe for colorimetric detection of Listeria monocytogenes, taking advantages of its oxidase-like activity. The linear range of the detection of Listeria monocytogenes was from 102 to 107CFUmL-1, with the limit of detection as low as 14CFUmL-1. The approach also showed good feasibility for detection of Listeria monocytogenes in milk and meat samples. The spiked recoveries were in the range 81-114% with relative standard deviations ranging from 1.28 to 5.19%. Thus, this work provides an efficient, convenient, and practical tool for selective isolation and colorimetric detection of Listeria monocytogenes in food.

Some virulence genes and biofilm formation capabilities of Listeria monocytogenes isolates from different sources

In this study, it was aimed to determine the biofilm-forming abilities of both clinical and food-borne isolates of Listeria monocytogenes, to investigate the presence of nine different virulence genes, and to consider the current threat status of this agent. A total of 28 isolates, 21 from food and seven from clinical origin, were used in the study. To determine the biofilm formation abilities of isolates, two different methods namely “tube adherence” and “microplate” were used respectively. For the determination of nine different virulence genes of Listeria monocytogenes (inlA, inlC, inlJ, hylA, luxS, flaA, prfA, inlB, actA), the method of polymerase chain reaction (PCR) was used. As a result, all isolates were found to be able to form a biofilm to varying degrees by both tube and microplate methods. These two methods were similar in terms of their results. All nine different virulence gene regions were detected at various rates in the isolates. Whereas the genes directly related to biofilm formation for the isolates weren't detected, to form biofilm was observed. The virulence genes detected in clinical origin isolates were proportionally higher than in food-borne isolates (except for flaA and prfA gene regions). It was concluded that bacteria of Listeria monocytogenes continue to form biofilm and carry virulence genes regardless they are from food or clinical origin. Also, food-borne contaminations continue to be a severe threat to human health. So, to prevent listeriosis cases of both humans and animals should be taken required precautions and all cases should be considered carefully.

Prevalence, antibiogram and molecular characterization of Listeria monocytogenes from ruminants and humans in New Valley and Beheira Governorates, Egypt

BackgroundListeriosis is a global health threat to both animals and humans, especially in developing countries. This study was designed to isolate Listeria monocytogenes from faeces; environmental samples; and cow, sheep and goat milk, as well as human stool, to study its molecular characteristics and antibiotic sensitivity in the New Valley and Beheira Governorates, Egypt. The isolation and identification of L. monocytogenes were carried out using traditional culture and biochemical methods, followed by antibiography, genus confirmation of some isolates and detection and sequencing of InlB genes via PCR.ResultsOut of 2097 examined samples, the prevalence of L. monocytogenes was 13.4% in animals; the prevalence was 9.2%, 2.4%, 25.4%, 4%, 42.4%, and 6.4% in cattle faeces, cattle milk, sheep faeces, sheep milk, goat faeces, and goat milk, respectively. However, the prevalence of L. monocytogenes was 8.3% in human samples. Both animal and human isolates showed 100% resistance to trimethoprim-sulfamethoxazole, and the isolates showed the highest sensitivity to flumequine (100%), amikacin (99.2%), gentamicin (97.6%), and levofloxacin (94.6%). Multidrug resistance (MDR) was detected in 86.9% of the tested isolates. The 16 S rRNA and inlB genes were detected in 100% of the randomly selected L. monocytogenes isolates. Phylogenetic analysis of three isolates based on the inlB gene showed 100% identity between faecal, milk and human stool isolates.ConclusionsFaeces and milk are major sources of listeriosis, and the high degree of genetic similarity between animal and human isolates suggests the possibility of zoonotic circulation. The high prevalence of MDR L. monocytogenes in both animal and human samples could negatively impact the success of prevention and treatments for animal and human diseases, thereby imposing serious risks to public health.

Biofilm formation by Listeria monocytogenes from the meat processing industry environment and the use of different combinations of detergents, sanitizers, and UV-A radiation to control this microorganism in planktonic and sessile forms.

This study aimed to evaluate the ability of biofilm formation by L. monocytogenes from the meat processing industry environment, as well as the use of different combinations of detergents, sanitizers, and UV-A radiation in the control of this microorganism in the planktonic and sessile forms. Four L. monocytogenes isolates were evaluated and showed moderate ability to form biofilm, as well as carried genes related to biofilm production (agrB, agrD, prfA, actA, cheA, cheY, flaA, sigB), and genes related to tolerance to sanitizers (lde and qacH). The biofilm-forming isolates of L. monocytogenes were susceptible to quaternary ammonium compound (QAC) and peracetic acid (PA) in planktonic form, with minimum inhibitory concentrations of 125 and 75 ppm, respectively, for contact times of 10 and 5min. These concentrations are lower than those recommended by the manufacturers, which are at least 200 and 300 ppm for QAC and PA, respectively. Biofilms of L. monocytogenes formed from a pool of isolates on stainless steel and polyurethane coupons were subjected to 14 treatments involving acid and enzymatic detergents, QAC and PA sanitizers, and UV-A radiation at varying concentrations and contact times. All treatments reduced L. monocytogenes counts in the biofilm, indicating that the tested detergents, sanitizers, and UV-A radiation exhibited antimicrobial activity against biofilms on both surface types. Notably, the biofilm formed on polyurethane showed greater tolerance to the evaluated compounds than the biofilm on stainless steel, likely due to the material's surface facilitating faster microbial colonization and the development of a more complex structure, as observed by scanning electron microscopy. Listeria monocytogenes isolates from the meat processing industry carry genes associated with biofilm production and can form biofilms on both stainless steel and polyurethane surfaces, which may contribute to their persistence within meat processing lines. Despite carrying sanitizer tolerance genes, QAC and PA effectively controlled these microorganisms in their planktonic form. However, combinations of detergent (AC and ENZ) with sanitizers (QAC and PA) at minimum concentrations of 125 ppm and 300 ppm, respectively, were the most effective.

Whole Genome Sequence Analysis of Listeria monocytogenes Isolates Obtained from the Beef Production Chain in Gauteng Province, South Africa.

The study used whole-genome sequencing (WGS) and bioinformatics analysis for the genomic characterization of 60 isolates of Listeria monocytogenes obtained from the beef production chain (cattle farms, abattoirs, and retail outlets) in Gauteng province, South Africa. The sequence types (STs), clonal complexes (CCs), and the lineages of the isolates were determined using in silico multilocus sequence typing (MLST). We used BLAST-based analyses to identify virulence and antimicrobial genes, plasmids, proviruses/prophages, and the CRISPR-Cas system. The study investigated any association of the detected genes to the origin in the beef production chain of the L. monocytogenes isolates. Overall, in 60 isolates of Listeria monocytogenes, there were seven STs, six CCs, forty-four putative virulence factors, two resistance genes, one plasmid with AMR genes, and three with conjugative genes, one CRISPR gene, and all 60 isolates were positive for proviruses/prophages. Among the seven STs detected, ST204 (46.7%) and ST2 (21.7%) were the most prominent, with ST frequency varying significantly (p < 0.001). The predominant CC detected were CC2 (21.7%) and CC204 (46.7%) in lineages I and II, respectively. Of the 44 virulence factors detected, 26 (across Listeria Pathogenicity Islands, LIPIs) were present in all the isolates. The difference in the detection frequency varied significantly (p < 0.001). The two AMR genes (fosX and vga(G)) detected were present in all 60 (100%) isolates of L. monocytogenes. The only plasmid, NF033156, was present in three (5%) isolates. A CRISPR-Cas system was detected in six (10%), and all the isolates carried proviruses/prophages. The source and sample type significantly affected the frequencies of STs and virulence factors in the isolates of L. monocytogenes. The presence of fosX and vga(G) genes in all L. monocytogenes isolates obtained from the three industries of the beef production chain can potentially cause therapeutic implications. Our study, which characterized L. monocytogenes recovered from the three levels in the beef production chain, is the first time genomics was performed on this type of data set in the country, and this provides insights into the health implications of Listeria.

Profile of pathogenic bacteria isolated from cow's milk in N'Djamena: associated risk factors and antibiotic resistance

Raw milk and its derivatives are foodstuffs vulnerable to contamination by microorganisms including pathogenic bacteria and failure to comply with hygiene rules. The objective of this study was to determine the profile of pathogenic bacteria isolated from cow's milk and to evaluate the effectiveness of antibiotics commonly used in veterinary and human medicine against these bacteria in Chad. This was a prospective and analytical study based on bacteriological examination including 180 milk samples collected in 10 districts of N'Djamena. The isolation, identification and testing of sensitivity of isolated bacteria to antibiotics were carried out under standard food bacteriology conditions. Among 180 milk samples which were screened by bacteriological examination, 71 (34.44%) cases were positive. The bacterial strains isolated were: Staphylococcus aureus (38.03%), Streptococcus agalactiae (18.31%), Staphylococcus hyicus (11.27%), Streptococcus uberis (7.04%), Escherichia coliO157H7 (7.04%), Streptococcus pyogenes (5.63%), Aeromonas hydrophila (5.63%), Listeria monocytogenes (4.22%) and Mycobacterium tuberculosis (2.81%). The risk factors most frequently associated with milk contamination were hand milking (100%) followed by unsanitary environment (65%), 35% of milks were sold in reused bottles and 58% of vendors and producers were uneducated. The antibiogram of the 69 isolated bacterial agents showed reduced sensitivities to beta-lactams and aminoglycosides, and varied resistance to cyclins and fluoroquinolones. Streptomycin remains an aminoglycoside most sensitive to all strains of pathogenic bacteria with an average level of 95.7%. Staphylococci (Staphylococcus aureus and Staphylococcus hyicus) developed an average resistance of 80.15% to methycillin, 80.1% to vancomycin and all bacteria showed an average resistance of 85% to metronidazole. Streptococci strains (Streptococcus agalactiae, Streptococcus uberis and Streptococcus pyogenes) were very sensitive (76.3%) to vancomycin. The isolated Listeria monocytogenes strains were 100%sensitive to aminoglycosides and beta-lactams. The two-strains of Mycobacterium tuberculosis detected by GeneXpert had a profile of 50%sensitivity and 50%resistance to rifampicin. This study not only made it possible to know the high rate and frequency of pathogenic bacteria in cow's milk as well as the associated risk factors, but also showed a reduction in sensitivity of bacteria to aminopenicillins and aminoglycosides, a strong resistance of Staphylococcus aureus to methicillin and vancomycin and a high sensitivity of streptococci to vancomycin.

Characteristics of Clinical Isolates of Listeria monocytogenes in Sichuan, China, in 2022 Based on Whole Genome Sequencing Analysis.

Listeria monocytogenes is a foodborne pathogen. In 2022, we collected 15 strains of L. monocytogenes isolated from patients in some foodborne disease sentinel monitoring hospitals in Sichuan Province. Through whole genome sequencing (WGS), we obtained the virulence genes carried by the strains, multi-locus sequence typing (MLST), core genome MLST (cgMLST), clonal complex (CC), and serum groups and constructed a phylogenetic tree and minimum spanning tree with nonhuman strains. An analysis shows that all 15 strains of L. monocytogenes carry virulence genes LIPI-1 and LIPI-2, whereas the carrying rates of LIPI-3 and LIPI-4 virulence genes are relatively low. The MLST typing results showed a total of 10 sequence types (ST), including 10 CCs, with ST7 being the dominant type. The cgMLST clearly distinguishes strains of different lineages and CC types. The serum group is divided into three types: IIa, IIb, and IVb, with IIa being the dominant serum group. An analysis of antibiotic genes showed that all 15 strains carried FosX, lin, mprF, and norB with high carrying rates. The minimum inhibitory concentration results indicated that all were susceptible to eight antibiotics (ampicillin, penicillin, tetracycline, meropenem, erythromycin, vancomycin, ciprofloxacin, and trimethoprim-sulfamethoxazole). The analysis of strains isolated from different sources of Listeria revealed varying degrees of diversity, and the contamination of meat and environment within the province is closely related to clinical cases. L. monocytogenes isolated from clinical cases in Sichuan Province carry multiple virulence and antibiotic genes, with high potential pathogenicity. It is necessary to further strengthen the monitoring and control of food and environment by L. monocytogenes within Sichuan Province.

Investigation of the seasonal prevalence, phenotypic, and genotypic characteristics of Listeria monocytogenes in slaughterhouses in Burdur.

This study examined Listeria monocytogenes isolates from two slaughterhouses in Burdur province, southern Turkey, over four seasons for antibiotic resistance, serogroups, virulence genes, in vitro biofilm forming capacity, and genetic relatedness. Carcass (540) and environment-equipment surface (180) samples were collected from two slaughterhouses (S1, S2) for 1 year (4 samplings). Of the 89 (12.4%) positive isolates, 48 (53.9%) were from animal carcasses, and 41 (46.1%) from the environment-equipment surfaces. Autumn was the peak season for Listeria monocytogenes compared to summer and spring (P<0.05). In addition, the most common serotype between seasons was 1/2c. Except for plcA and luxS genes, all isolates (100%) harbored inlA, inlC, inlJ, hlyA, actA, iap, flaA genes. Listeria monocytogenes isolates were identified as belonging to IIc (1/2c-3c; 68.5%), IVb (4b-4d-4e; 29.2%), and IIa (1/2a-3a; 2.2%) in the screening using multiplex polymerase chain reaction-based serogrouping test. A total of 65 pulsotypes and 13 clusters with at least 80% homology were determined by using pulsed field gel electrophoresis on samples that had been digested with ApaI. Thirty-four (38.2%) of the isolates were not resistant to any of the 14 antibiotics tested. The antibiotic to which the isolates showed the most resistance was rifampicin (44.9%). Serotype 1/2c was the most resistant serotype to antibiotics. Despite having biofilm-associated genes (inlA, inlB, actA, flaA, and luxS), a minority (11%) of isolates formed weak biofilm. This study revealed seasonal changes prevalence of Listeria monocytogenes, particularly higher in autumn, posing a greater risk of meat contamination. Notably, Serotype 1/2c showed significant prevalence and antibiotic resistance. Indistinguishable isolates indicated cross-contamination, underscoring the importance of prioritized training for slaughterhouse personnel in sanitation and hygiene protocols.

Draft genome of three isolates of Listeria monocytogenes isolated from Stenella coeruleoalba in Italy.

Listeria monocytogenes is the etiological agent of the listeriosis. Here, we described three draft genome sequences of L. monocytogenes isolated in Italy from stranded individuals of the striped dolphin Stenella coeruleoalba. All the genomes have been molecular typed through the multilocus sequence typing to identify the phylogenetic lineage, clonal complex, sublineage, and serogroup.

Heterogeneity of Antibiotic-Resistant Isolates of Listeria Monocytogenes Isolated from Food Products in Moscow

Relevance. Listeria monocytogenes is a ubiquitous bacterium that causes listeriosis, which represents a widespread infectious disease currently inflicting great damage to livestock production and posing a serious threat to human health.Aim. To analyze the population structure and assess the pathogenic potential of Listeria monocytogenes isolates isolated on the territory of the Russian Federation.Materials and methods. A total of 79 listeria isolates were isolated from food products. Species identification and phenotypic analysis for antibiotic resistance were performed using VITEK MS system (bioMerieux, Marcyl’toile, France). Thirty-five antibiotic-resistant isolates were characterized by analysis of whole-genome sequencing data.Results. Whole genome sequences of thirty-five antibiotic-resistant Listeria monocytogenes isolates of food origin were analyzed. We determined clonal structure of this population and revealed a small number of antibiotic resistance determinants (fosX, tetM и сlpL), extensive set of virulence factors, as well as the presence of CRISPR/Cas systems. Most of the isolates belonged to phylogenetic line II and were divided into nine clonal complexes with the prevalence of CC121, which was one of the epidemiologically significant genetic clones. Two CC2 isolates belonging to the most pathogenic phylogenetic lineage I were also found. Thirteen isolates were characterized by the presence of putative CRISPR/Cas systems of IB and IIA types. All ST 121 isolates contained two types of identified adaptive immunity systems simultaneously in their genomes. Correlation analysis confirmed their functionality.Conclusion. We believe that the whole genome data obtained for the foodborne Listeria monocytogenes isolates will facilitate and complement further epidemiological studies of this pathogen, as well as the investigations of its genome variability in terms of the acquisition of various genetic elements associated with adaptation, antimicrobial resistance, and virulence. Moreover, the results of such studies will help to develop preventive measures to effectively solve problems associated with the bacterial contamination of animal products and ensure food safety in production conditions and the «farm-to-table» chain.

Inter-strain variability on the cardinal parameters (pH and Aw) of clinical and food isolates of Listeria monocytogenes using turbidimetric measurements

Listeria monocytogenes is a foodborne pathogen which, in 2021, was considered the fifth most commonly reported zoonosis in humans in the European Union (EU). Ready-to-eat (RTE) fishery products, deli meats or soft cheeses have been mostly involved in food safety alerts and outbreaks in the last years. Hurdle technology by food industries has been widely used to enhance the safety of foods. Among the barriers, the application of acid and osmotic stress during processing is extensively used worldwide. This study aims to gain knowledge about the inter-strain variability of twenty-six clinical and food L. monocytogenes isolates with the estimation of their cardinal parameters using turbidimetric measurements. To analyse the data and to obtain the estimated cardinal values, a common statistical procedure was set up. The estimation of cardinal parameters showed a high inter-strain variability of L. monocytogenes, and no correlation was observed between Aw min and pHmin values for the studied strains. By grouping the strains in clinical, meat and fish origin, it was observed that strains from the meat group presented the lowest average pHmin values (4.57), thus showing potential acid adaptation. This work contributes to gain knowledge of the inter-strain variability of L. monocytogenes in relation with pH and Aw cardinal values, as well as provide a starting point for future validation studies in fish and meat food matrices.

Sensitivity of 240 Listeria monocytogenes isolates to common industrial biocides is more dependent on the presence of residual organic matter or biofilm than on genetic determinants

Listeria monocytogenes continues to be a problem in ready-to-eat foods. Tolerance and adaptation to biocides have been proposed to contribute to the persistence of L. monocytogenes in the food processing industry. This study aimed to determine if the sensitivity to common biocides varies among 240 L. monocytogenes isolates. The impact of residual organic matter (soils), biofilm, and ability to adapt to biocides was also evaluated. Food and food processing related isolates of L. monocytogenes (n = 240) were whole genome sequenced and assigned to sequence types (n = 53) and clonal complexes (n = 32). Isolates were screened for genetic elements associated with biocide tolerance and tested for their minimum inhibitory concentration (MIC) to benzalkonium chloride (BC), peracetic acid (PAA), sodium hypochlorite (SH) and ethanol (ET). Biocide tolerance under soiled conditions, in biofilm and in a broth suspension test (BST) was further tested for 19 representative isolates. Adaptation to sub-MICs of BC, PAA or SH (18 days, 15 °C) was tested for 15 isolates. MIC for PAA (62 mg/L) showed no variation, while for SH (47–94 mg/L) and ET (4.7–9.4% v/v) values differed 2 × among isolates. For BC, an 8 × difference (0.3–2.5 mg/L) was observed. Eighty-seven of 88 BC tolerant isolates (MIC of 2.5 mg/L) harbored known BC tolerance genes. Soiled conditions increased MIC-values 8–33 × for BC and 8 × for SH but not for PAA and ET. Biofilm increased minimum bactericidal concentrations 4–8 × for BC but not for PAA and SH. Survival of isolates with BC tolerance genes was not significantly (p > 0.05) improved in BSTs with PAA, SH or BC. Lineage I isolates (n = 7 of 19) showed significantly (p < 0.05) better survival than lineage II isolates (n = 12) when exposed to PAA (100 mg/L) in BSTs. Adaptation to BC (8 × MIC increase) occurred exclusively for originally sensitive isolates, while no adaptation to SH and PAA were observed. In conclusion, L. monocytogenes isolates showed limited variation in sensitivity to biocides in MIC assays. However, lineage I isolates were less sensitive to PAA than lineage II isolates in BSTs. The efficacy of SH and BC were affected by residual organic matter and/or biofilm, demonstrating the need for proper cleaning prior to biocide use with PAA being less perturbed.

Analysis of production levels of InlA and InlB invasion factors in Listeria monocytogenes isolates collected in the Russian Federation

Background. Listeria monocytogenes is characterized by the presence of epidemic hypervirulent clones. A key feature of L. monocytogenes is its capacity to invade non-professional phagocytic cells. Hypervirulent clones are strongly associated with the increased production and/or the presence of certain isoforms of invasion factors InlA and InlB.&#x0D; The purpose of the study is to create a test system for InlA and InlB detection and to measure the InlA and InlB production levels in L. monocytogenes isolates belonging to clonal groups with different virulence potential.&#x0D; Materials and methods. The study was performed using 32 L. monocytogenes strains belonging to epidemic clones ECII, ECIV, ECVII (clonal complexes CC1, CC2, CC7) and hypovirulent clonal complex CC9. Sequencing of inlA and inlB genes was performed. The indirect enzyme-linked immunosorbent assay was used to analyze the production levels of InlA and InlB proteins.&#x0D; Results. The variability of InlA was revealed among strains belonging to the same clonal complex: 3 InlA isoforms were identified among strains belonging to CC7; out of 8 strains belonging to CC9, one strain had a stop codon in the inlA gene, leading to the loss of function of the InlA protein. The differences between inlB alleles correlated with the specificity of strains belonging to a certain clonal complex. Differences in production levels of invasion factors were measured. In strains belonging to CC9, the InlA production level was 2.5 times as low compared to strains belonging to CC1, CC2, and CC7. In strains belonging to phylogenetically related CC1 and CC2, the InlB production level was on average 4 times as high compared to strains belonging to CC7 and CC9.&#x0D; Conclusion. The obtained results confirm the variability of major invasion factors both among clonal complexes and strains of the same complex. The increased production of invasion factors InlA and InlB correlates with the potential virulence of strains.

Variability in Physicochemical Parameters and Its Impact on Microbiological Quality and Occurrence of Foodborne Pathogens in Artisanal Italian Organic Salami.

Artisanal salami is produced in small-scale production plants, where the lack of full automation might result in higher variability in food intrinsic properties. The aim of the present study was to evaluate the inter- and intra-batch variability in physicochemical parameters and its impact on microbial quality and occurrence of foodborne pathogens on 480 samples collected from six batches of an artisanal Italian production of organic salami. Relatively high total bacterial counts (TBC) were found on the surface of the table in the stuffing room (4.29 ± 0.40 log cfu/cm2). High loads of Enterobacteriaceae in the meat mixture of batch 2 and TBC in batch 5 were associated with a higher occurrence of bacterial pathogens. During ripening, water activity (aw) and pH failed to reach values lower than 0.86 and 5.3, respectively. Six Staphylococcus aureus and four Listeria monocytogenes isolates were collected from the salami meat mixture during ripening and the processing environment. A total of 126 isolates of Enterobacteriaceae were characterized at a species level, with Escherichia coli, Klebsiella pneumoniae, Enterobacter cloacae, and Citrobacter freundii isolated from the final products. Results suggest the relevance of first steps of production in terms of the hygiene of raw materials and handling during stuffing procedures, especially when the physicochemical parameters of the final products do not reach values that represent hurdles for foodborne pathogens.

Clinical Features and Antibiotic Treatment of Neonatal Listeriosis: A Hospital-Based Study.

To determine the clinical features, laboratory findings, antibiotic treatment, and outcomes of neonatal listeriosis in a specialized tertiary hospital in Wuhan, China. We retrospectively analyzed the medical records of patients diagnosed with neonatal listeriosis at Maternal and Child Health Hospital of Hubei Province from January 2015 to December 2022. Listeriosis was indicated by positive culture for Listeria monocytogenes (LM). A total of 11 cases were included in our study. The incidence rate of neonatal listeriosis was 2.06 per 100,000 live births. Seventy-three percent of the cases were born prematurely, caused early onset sepsis. Respiratory distress (100%) was the most common and earliest symptom, followed by fever (64%) and rashes (27%). The levels of C-reactive protein (CRP) and procalcitonin (PCT) were elevated in 100% of the cases. The median time-to-positivity (TTP) of the culture was 15 hours (range 9-28hours). Of the 11 neonates, 6 were cured, 2 showed improvement, and 3 died, with a mortality rate of 27%. There were statistically significant differences in Apgar score at 5 minutes (p=0.037) and CRP (p=0.025) between the survival group and fatality group. Ampicillin was sensitive to LM isolates and effective for therapy if initiated early. Neonatal listeriosis is a rare but severe infection with a high mortality rate. Early identification and appropriate use of effective antibiotics are particularly critical for achieving positive outcomes. Apgar score and CRP are relevant indices for prognosis. Ampicillin is the first-line therapy and can be empirically administered to neonates suspected of having listeriosis.

Phenotypic and Genotypic Characteristics of Non-Hemolytic L. monocytogenes Isolated from Food and Processing Environments.

Increasingly, Listeria monocytogenes (LM) with atypical phenotypic and genotypic characteristics are being isolated from food, causing problems with their classification and testing. From 2495 soil, food, and swab samples from the food industry, 262 LM isolates were found. A total of 30 isolates were isolated, mainly from soil and plant food, and were classified as atypical LM (aLM) because they lacked the ability to move (30/11.4%) and perform hemolysis (25/9.5%). The isolation environment affected aLM incidence, cell size, sugar fermentation capacity, antibiotic sensitivity, and the number of virulence genes. Therefore, despite several characteristics differentiating all aLMs/non-hemolytic isolates from reference LMs, the remaining phenotypic characteristics were specific to each aLM isolate (like a fingerprint). The aLM/non-hemolytic isolates, particularly those from the soil and meat industries, showed more variability in their sugar fermentation capacity and were less sensitive to antibiotics than LMs. As many as 11 (36.7%) aLM isolates had resistance to four different antibiotics or simultaneously to two antibiotics. The aLM isolates possessed 3-7 of the 12 virulence genes: prfA and hly in all aLMs, while iap was not present. Only five (16.7%) isolates were classified into serogroups 1/2c-3c or 4a-4c. The aLM/non-hemolytic isolates differed by many traits from L. immobilis and atypical L. innocua. The reference method of reviving and isolating LM required optimization of aLM. Statistical analyses of clustering, correlation, and PCA showed similarities and differences between LM and aLM/non-hemolytic isolates due to individual phenotypic traits and genes. Correlations were found between biochemical traits, antibiotic resistance, and virulence genes. The increase in the incidence of atypical non-hemolytic LM may pose a risk to humans, as they may not be detected by ISO methods and have greater antibiotic resistance than LM. aLM from LM can be distinguished based on lack of hemolysis, motility, growth at 4 °C, ability to ferment D-arabitol, and lack of six specific genes.

Molecular genetic studies of Listeria monocytogenes isolates from different animal species in Ukraine

The PCR-profiles of 12 archival cultures of listeria isolated from different species of animals and stored in the collection of microorganisms of the NSC IEСVM were determined. It was established that 10 isolates have genes prs, inlB, inlA, inlC, inlJ, actA, plcB, hly, iap, which characterizes them as highly virulent. The L. monocytogenes 61052 isolate did not contain the inlA gene, and the actA and hly genes were not detected in L. monocytogenes Varja. Using PCR, it was also established that 11 studied cultures belong to serotypes 4b, 4d, 4e, as well as one isolate to 1/2a. The obtained data can be used to improve the diagnosis of animal listeriosis in Ukraine due to the creation of highly specific antigens

Comparison of genetic variations between high- and low-risk Listeria monocytogenes isolates using whole-genome de novo sequencing.

In this study, genetic variations and characteristics of Listeria monocytogenes isolates from enoki mushrooms (23), smoked ducks (7), and processed ground meat products (30) were examined with respect to hemolysis, virulence genes, growth patterns, and heat resistance. The isolates that showed the highest pathogenicity and the lowest pathogenicity were analyzed to obtain the whole-genome sequence, and the sequences were further analyzed to identify genetic variations in virulence, low-temperature growth-related, and heat resistance-related factors. All isolates had β-hemolysis and virulence genes (actA, hlyA, inlA, inlB, and plcB). At low temperatures, isolates with high growth (L. monocytogenes strains SMFM 201803 SD 1-1, SMFM 201803 SD 4-2, and SMFM 201804 SD 5-3) and low growth (L. monocytogenes strains SMFM 2019-FV43, SMFM 2019-FV42, and SMFM 2020-BT30) were selected. Among them, L. monocytogenes SMFM 201804 SD 5-3 showed the highest resistance at 60°C and 70°C. The strains SMFM 201804 SD 5-3 (high-risk) and SMFM 2019-FV43 (low-risk) harbored 45 virulence genes; 41 single nucleotide variants (SNVs) were identified between these two isolates. A comparison of 26 genes related to low-temperature growth revealed 18 SNVs between these two isolates; a comparison of the 21 genes related to heat resistance revealed 16 SNVs. These results indicate that the differences in the pathogenicity of L. monocytogenes SMFM 201804 SD 5-3 and L. monocytogenes SMFM 2019-FV43 are associated with the SNVs identified in virulence genes, low-temperature growth-related genes, and heat resistance-related genes.