Listeria Monocytogenes Strains Research Articles

Pediocin SA-1: A selective bacteriocin for controlling Listeria monocytogenes in maize silages.

In this study, we assessed the potential as silage additive of a bacteriocin produced by Pediococcus acidilactici Northern Regional Research Laboratory (NRRL) B-5627 (pediocin SA-1). Maize was inoculated either with a bacterial starter alone (I) or in combination with the bacteriocin (IP), and untreated silage served as control. We monitored the products of fermentation (ethanol, and lactic and acetic acids), the microbial population, and the presence of the indicator strain Listeria monocytogenes Colección Española de Cultivos Tipo (CECT) 4032 (1×10(5) cfu/g) after 1, 2, 5, 8, 16, and 30d of ensiling. Our results indicated antilisterial activity of the bacteriocin, anticipating the disappearance of L. monocytogenes in IP compared with I and control silages. The PCR-denaturing gradient gel electrophoresis analysis revealed the addition of the bacteriocin did not affect the bacterial communities of the spontaneous fermentation, and the inoculant-containing bacteria (Lactobacillus plantarum, Lactobacillus buchneri, and Enterococcus faecium) were found in addition to the bacterial communities of untreated maize silages in I and IP silages. Both treatments increased the concentration of antimicrobial compounds (acetic acid, ethanol, and 1,2-propanodiol) and led to lower residual sugar contents compared with the control, which would provide enhanced aerobic stability. The fact that the identified species L. plantarum, L. buchneri, and E. faecium produce some of these inhibitory compounds, together with their persistence throughout the 30d of fermentation, suggest these bacteria could actively participate in the ensiling process. According to these results, pediocin SA-1 could be used as an additive to control the presence of L. monocytogenes in maize silages selectively, while improving their fermentative quality and eventually their aerobic stability.

Genome Sequence of Listeria monocytogenes Strain HPB2088 (Serotype 1/2a), an Environmental Isolate Collected in Canada in 1994.

Listeria monocytogenes is a foodborne pathogen that causes severe illness. Thus, ongoing efforts at real-time whole-genome sequencing are of utmost importance. However, it is also important that retrospective analyses that place these data into context be performed. Here, we present the genome sequence of strain HPB2088, which was collected in 1994.

Artemisiaarborescens Essential Oil Composition, Enantiomeric Distribution, and Antimicrobial Activity from Different Wild Populations from the Mediterranean Area.

Aerial parts of Artemisiaarborescens were collected from different sites of the Mediterranean area (southwestern Algeria and southern Italy) and the chemical composition of their essential oil (EO) extracted by hydrodistillation was studied by both gas chromatography (GC) equipped with an enantioselective capillary column and GC/mass spectrometry (GC/MS). The EOs obtained were tested against several Listeriamonocytogenes strains. Using GC and GC/MS, 41 compounds were identified, accounting for 96.0-98.8% of the total EO. All EOs showed a similar terpene profile, which was rich in chamazulene, β-thujone, and camphor. However, the concentration of such compounds varied among the EOs. A.arborescens EO inhibited up to 83.3% of the L.monocytogenes strains, but the inhibitory spectrum varied among the EOs, with those from Algeria showing a higher inhibition degree than the Italian EOs. Such effect likely depended on the ketone (β-thujone+camphor) content of the EO. The differences in the EO composition support the hypothesis that A.arborescens has at least two different chemotypes: a β-thujone and a chamazulene type. The EO inhibitory spectrum indicates the A.arborescens EO as a valuable option in the control of the food-borne pathogens.

The effect of mild preservation treatments on the invasiveness of different Listeria monocytogenes strains on Greenshell™ mussels

High pressure processing (HPP) and thermal treatments are food preservation technologies used by the food industry to inactivate Listeria monocytogenes. However, the safe shelf-life of treated products could be compromised by the presence and/or recovery of surviving bacteria. The aim of the present study was to evaluate the effect of these technologies on the invasion capacity of L. monocytogenes. Eleven L. monocytogenes strains (serovars 1/2a,1/2b,1/2c and 4b) isolated from food processing plants and food products were grown in culture media, associated with Greenshell™ mussel surfaces and subjected (or not) to HPP (350 MPa, 2 min) or mild heating (MH, 55 °C, 10 min). The invasiveness of the strains was compared immediately after processing and after 14 days of storage at 8 °C using human intestinal epithelial Caco-2 cells. All strains on mussels had decreased invasion capacity compared with those coming directly from culture media. Invasiveness of L. monocytogenes also decreased immediately after pressurization but returned to the rate in untreated samples after 14 days of refrigerated storage. In contrast, MH did not affect invasion capacity, and treated samples showed the same rates as non-MH treated ones. Lineage I strains were more invasive than lineage II strains after both mild processing treatments. Based on these results, the risk of L. monocytogenes infection by the consumption of treated mussels will not be increased, as the invasiveness of possible survivors is not increased.

Growth differences and competition between Listeria monocytogenes strains determine their predominance on ham slices and lead to bias during selective enrichment with the ISO protocol

Listeria monocytogenes strains are widespread in the environment where they live well mixed, often resulting in multiple strains contaminating a single food sample. The occurrence of different strains in the same food might trigger strain competition, contributing to uneven growth of strains in food and to bias during selective procedures.We tested the growth of seven L. monocytogenes strains (C5, 6179, ScottA, PL24, PL25, PL26, PL27) on ham slices and on nutrient-rich agar at 10°C, singly and in combinations. Strains were made resistant to different antibiotics for their selective enumeration. In addition, growth of single strains (axenic culture) and competition between strains in xenic cultures of two strains was evaluated in enrichment broth and on selective agar. According to ISO 11290-1:1996/Amd 1:2004 standard protocol for detection of L. monocytogenes, two enrichment steps both followed by streaking on ALOA were performed. Strain cultures were directly added in the enrichment broth or used to inoculate minced beef and sliced hams which were then mixed with enrichment broth. 180–360 colonies were used to determine the relative percentage of each strain recovered on plates per enrichment step.The data showed a significant impact of co-cultivation on the growth of six out of seven strains on ham and a bias towards certain strains during selective enrichment. Competition was manifested by: (i) cessation of growth for the outcompeted strain when the dominant strain reached stationary phase, (ii) reduction of growth rates or (iii) total suppression of growth (both on ham and in enrichment broth or ALOA). Outgrowth of strains by their competitors on ALOA resulted in limited to no recovery, with the outcompeting strain accounting for up to 100% of the total recovered colonies. The observed bias was associated with the enrichment conditions (i.e. food type added to the enrichment broth) and the strain-combination. The outcome of growth competition on food or nonselective agar surface did not necessarily coincide with the results of competition during enrichment. The results show that certain strains present in foods may be missed during classical detection due to strain competition and such likelihood should be taken into consideration when resolving a listeriosis outbreak.

Screening of the Enterocin-Encoding Genes and Antimicrobial Activity in Enterococcus Species

In the current study, a total of 135 enterococci strains from different sources were screened for the presence of the enterocin-encoding genes entA, entP, entB, entL50A, and entL50B. The enterocin genes were present at different frequencies, with entA occurring the most frequently, followed by entP and entB; entL50A and L50B were not detected. The occurrence of single enterocin genes was higher than the occurrence of multiple enterocin gene combinations. The 80 isolates that harbor at least one enterocin-encoding gene (denoted "Gene(+) strains") were screened for antimicrobial activity. A total of 82.5% of the Gene(+) strains inhibited at least one of the indicator strains, and the isolates harboring multiple enterocin-encoding genes inhibited a larger number of indicator strains than isolates harboring a single gene. The indicator strains that exhibited growth inhibition included Listeria innocua strain CLIP 12612 (ATCC BAA-680), Listeria monocytogenes strain CDC 4555, Enterococcus faecalis ATCC 29212, Staphylococcus aureus ATCC 25923, S. aureus ATCC 29213, S. aureus ATCC 6538, Salmonella enteritidis ATCC 13076, Salmonella typhimurium strain UK-1 (ATCC 68169), and Escherichia coli BAC 49LT ETEC. Inhibition due to either bacteriophage lysis or cytolysin activity was excluded. The growth inhibition of antilisterial Gene+ strains was further tested under different culture conditions. Among the culture media formulations, the MRS agar medium supplemented with 2% (w/v) yeast extract was the best solidified medium for enterocin production. Our findings extend the current knowledge of enterocin-producing enterococci, which may have potential applications as biopreservatives in the food industry due to their capability of controlling food spoilage pathogens.

Chitosan grafted monomethyl fumaric acid as a potential food preservative

The present study aims at in vitro antibacterial and antioxidant activity evaluation of chitosan modified with monomethyl fumaric acid (MFA) using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) as mediator. Three different kinds of chitosan derivatives Ch-Ds-1,Ch-Ds-2 and Ch-Ds-3 were synthesized by feeding different concentration of MFA. The chemical structures of resulting materials were characterized by 1H NMR, 13C NMR, HR-XRD, FT-IR and TNBS assay. The results showed that Ch-Ds-1, Ch-Ds-2 and Ch-Ds-3 were successfully synthesized. The % amino groups of chitosan modified by MFA were evaluated by TNBS assay and ranging from 1.82±0.05% to 7.88±0.04%. All the chitosan derivatives are readily soluble in water and swelled by dimethyl sulfoxide (DMSO), toluene and dimethyl formamide (DMF). The antioxidant activity for all the chitosan derivatives have been significantly improved (P<0.05) compared to the chitosan. Upon antibacterial activity at pH 4.0, all the chitosan derivatives showed significant (P<0.05) antibacterial activity against Gram positive Staphylococcus aureus, Listeria monocytogenes strains and Gram negative Escherichia coli and Salmonella enteritidis strains compared to chitosan. In conclusion, MFA modified chitosan has shown enhanced activities along with solubility, and could be used as a novel food preservative and packaging material for long time food safety and security.

Antimicrobial and Physicochemical Characterization of Biodegradable, Nitric Oxide-Releasing Nanocellulose-Chitosan Packaging Membranes.

Biodegradable composite membranes with antimicrobial properties consisting of nanocellulose fibrils (CNFs), chitosan, and S-nitroso-N-acetyl-d-penicillamine (SNAP) were developed and tested for food packaging applications. As a nitric oxide donor, SNAP was encapsulated into completely dispersed chitosan in 100 mL of 0.1 N acetic acid and was thoroughly mixed with CNFs to produce a composite membrane. The fabricated membranes had a uniform dispersion of chitosan and SNAP within the CNFs, which was confirmed through scanning electron microscopy (SEM) micrographs and a chemiluminescence nitric oxide analyzer. The membranes prepared without SNAP showed lower water vapor permeability than that of the membranes with SNAP. The addition of SNAP resulted in a decrease in Young's modulus for both two- and three-layer membrane configurations. Antimicrobial property evaluation of SNAP-incorporated membranes showed an effective zone of inhibition against bacterial strains of Enterococcus faecalis, Staphylococcus aureus, and Listeria monocytogenes and demonstrated its potential applications for food packaging.

CD8+ T cells specific to a single Yersinia pseudotuberculosis epitope restrict bacterial replication in the liver but fail to provide sterilizing immunity

CD8+ T cells use contact-dependent cytolysis of target cells to protect the host against intracellular pathogens. We have previously shown that CD8+ T cells and perforin are required to protect against the extracellular pathogen Yersinia pseudotuberculosis. Here we establish an experimental system where CD8+ T cells specific to a single model antigen are the only memory response present at time of challenge. Using mice immunized with a vaccine strain of Listeria monocytogenes that expresses secreted ovalbumin (Lm-OVA), we show that OVA-specific CD8+ T cells are generated and provide limited protection against challenge with virulent OVA+Y. pseudotuberculosis. Perforin expression by OVA-specific CD8+ T cells was required, as Lm-OVA-immunized perforin-deficient mice showed higher bacterial burden as compared to Lm-OVA-immunized perforin-sufficient mice. Surprisingly, antigen-specific T cell protection waned over time, as Lm-OVA-immune mice eventually succumbed to Yersinia infection. Kinetic analysis of infection in mice with and without OVA-specific CD8+ T cells revealed that bacterial numbers increased sharply in OVA-naïve mice until death, while OVA-immune mice held bacterial burden to a lower level throughout the duration of illness until death. Clonal analysis of bacterial populations in OVA-naïve and OVA-immune mice at distinct time points revealed equivalent and severe bottle-neck effects for bacteria in both sets of mice immediately after intravenous challenge, demonstrating a dominant role for other aspects of the immune system regardless of CD8+ T cell status. These studies indicate that CD8+ T cells against a single antigen can restrict Y. pseudotuberculosis colonization in a perforin-dependent manner, but ultimately are insufficient in their ability to provide sterilizing immunity and protect against death.

Population Genetic Structure of Listeria monocytogenes Strains as Determined by Pulsed-Field Gel Electrophoresis and Multilocus Sequence Typing.

ABSTRACTListeria monocytogenes is a ubiquitous bacterium that may cause the foodborne illness listeriosis. Only a small amount of data about the population genetic structure of strains isolated from food is available. This study aimed to provide an accurate view of the L. monocytogenes food strain population in France. From 1999 to 2014, 1,894 L. monocytogenes strains were isolated from food at the French National Reference Laboratory for L. monocytogenes and classified according to the five risk food matrices defined by the European Food Safety Authority (EFSA). A total of 396 strains were selected on the basis of different pulsed-field gel electrophoresis (PFGE) clusters, serotypes, and strain origins and typed by multilocus sequence typing (MLST), and the MLST results were supplemented with MLST data available from Institut Pasteur, representing human and additional food strains from France. The distribution of sequence types (STs) was compared between food and clinical strains on a panel of 675 strains. High congruence between PFGE and MLST was found. Out of 73 PFGE clusters, the two most prevalent corresponded to ST9 and ST121. Using original statistical analysis, we demonstrated that (i) there was not a clear association between ST9 and ST121 and the food matrices, (ii) serotype IIc, ST8, and ST4 were associated with meat products, and (iii) ST13 was associated with dairy products. Of the two major STs, ST121 was the ST that included the fewest clinical strains, which might indicate lower virulence. This observation may be directly relevant for refining risk analysis models for the better management of food safety.IMPORTANCE This study showed a very useful backward compatibility between PFGE and MLST for surveillance. The results enabled better understanding of the population structure of L. monocytogenes strains isolated from food and management of the health risks associated with L. monocytogenes food strains. Moreover, this work provided an accurate view of L. monocytogenes strain populations associated with specific food matrices. We clearly showed that some STs were associated with food matrices, such as meat, meat products, and dairy products. We opened the way to source attribution modeling in order to quantify the relative importance of the main food matrices.

A secret(e) weapon for food poisoning

Microbiology![Figure][1] Listeria monocytogenes secrete a toxin that allows them to overwhelm good gut microbes. CREDIT: SCIENCE PICTURE CO/SCIENCE SOURCE Trillions of microbes reside in our gut, producing essential nutrients and defending gut integrity. So how do a few incoming pathogens compete against these masses to establish an infection? Some strains of the bacteria Listeria monocytogenes cause gastroenteritis, which can be fatal in the immunocompromised and in pregnant women. Studying mice, Quereda et al. found that a virulent strain of L. monocytogenes produces a toxin called listeriolysin S, but only when it is in the gut. The toxin led to changes in the abundance of acetate- and butyrate-producing gut resident microbes in L. monocytogenes –infected mice. These short-chain fatty acids can inhibit L. monocytogenes growth, implying that L. monocytogenes expresses the toxin to overwhelm resident microbial competition. Proc. Natl. Acad. Sci. U.S.A. 10.1073/pnas.1523899113 (2016). [1]: pending:yes

Loss of SigB in Listeria monocytogenes Strains EGD-e and 10403S Confers Hyperresistance to Hydrogen Peroxide in Stationary Phase under Aerobic Conditions.

SigB is the main stress gene regulator in Listeria monocytogenes affecting the expression of more than 150 genes and thus contributing to multiple-stress resistance. Despite its clear role in most stresses, its role in oxidative stress is uncertain, as results accompanying the loss of sigB range from hyperresistance to hypersensitivity. Previously, these differences have been attributed to strain variation. In this study, we show conclusively that unlike for all other stresses, loss of sigB results in hyperresistance to H2O2 (more than 8 log CFU ml(-1) compared to the wild type) in aerobically grown stationary-phase cultures of L. monocytogenes strains 10403S and EGD-e. Furthermore, growth at 30°C resulted in higher resistance to oxidative stress than that at 37°C. Oxidative stress resistance seemed to be higher with higher levels of oxygen. Under anaerobic conditions, the loss of SigB in 10403S did not affect survival against H2O2, while in EGD-e, it resulted in a sensitive phenotype. During exponential phase, minor differences occurred, and this result was expected due to the absence of sigB transcription. Catalase tests were performed under all conditions, and stronger catalase results corresponded well with a higher survival rate, underpinning the important role of catalase in this phenotype. Furthermore, we assessed the catalase activity in protein lysates, which corresponded with the catalase tests and survival. In addition, reverse transcription-PCR (RT-PCR) showed no differences in transcription between the wild type and the ΔsigB mutant in various oxidative stress genes. Further investigation of the molecular mechanism behind this phenotype and its possible consequences for the overall phenotype of L. monocytogenes are under way. SigB is the most important stress gene regulator in L. monocytogenes and other Gram-positive bacteria. Its increased expression during stationary phase results in resistance to multiple stresses. However, despite its important role in general stress resistance, its expression is detrimental for the cell in the presence of oxidative stress, as it promotes hypersensitivity against hydrogen peroxide. This peculiar phenotype is an important element of the physiology of L. monocytogenes, and it might help us explain the behavior of this organism in environments where oxidative stress is present.

Potential Impact of the Resistance to Quaternary Ammonium Disinfectants on the Persistence of Listeria monocytogenes in Food Processing Environments.

The persistence of certain strains of Listeria monocytogenes, even after the food processing environment has been cleaned and disinfected, suggests that this may be related to phenomena that reduce the concentration of the disinfectants to subinhibitory levels. This includes (i) the existence of environmental niches or reservoirs that are difficult for disinfectants to reach, (ii) microorganisms that form biofilms and create microenvironments in which adequate concentrations of disinfectants cannot be attained, and (iii) the acquisition of resistance mechanisms in L. monocytogenes, including those that lead to a reduction in the intracellular concentration of the disinfectants. The only available data with regard to the resistance of L. monocytogenes to disinfectants applied in food production environments refer to genotypic resistance to quaternary ammonium compounds (QACs). Although there are several well-characterized efflux pumps that confer resistance to QACs, it is a low-level resistance that does not generate resistance to QACs at the concentrations applied in the food industry. However, dilution in the environment and biodegradation result in QAC concentration gradients. As a result, the microorganisms are frequently exposed to subinhibitory concentrations of QACs. Therefore, the low-level resistance to QACs in L. monocytogenes may contribute to its environmental adaptation and persistence. In fact, in certain cases, the relationship between low-level resistance and the environmental persistence of L. monocytogenes in different food production chains has been previously established. The resistant strains would have survival advantages in these environments over sensitive strains, such as the ability to form biofilms in the presence of increased biocide concentrations.

The role of commensal microbiota on CD8 T cell response to Listeria monocytogenes infection

Abstract Commensal microbiota is composed of trillions of microorganisms including fungi, protozoans, viruses and bacteria, which have co-evolved with the host immune system. In particular, the gut is the largest reservoir of microorganisms that are involved in dynamic interaction with the immune system, including maintaining intestinal homeostasis, induction of several metabolic pathways, and protection from pathogenic microbes. To better understand the mechanisms involved microbiota-mediated protection from the pathogenic bacteria, we have studied oral infection of germ-free (GF) mice with a strain of Listeria monocytogenes (L. monocytogenes) that has been engineered to efficiently infect the intestinal epithelial cells of the mouse. We found that GF mice are more susceptible to orally administrated L. monocytogenes than conventional SPF mice, with an increased Listeria burden in the mLN, small intestine and feces. In addition, in the infected GF mice, L. monocytogenes-specific CD8 T cells were vigorously activated and induced to differentiate into memory precursor effector cells (MPECs) and tissue resident memory (CD69+ CD103+) T cells in the intestinal epithelium at a faster rate than in infected SPF mice. L. monocytogenes-specific CD8 T cells in GF mice also secrete more effector cytokines, such as interferon-γ and tumor necrosis factor-α. The mechanisms by how commensal microbiota confers resistance to infection by L. moncoytogenes in SPF mice is currently under investigation.

An Outbreak of Human Listeriosis in England between 2010 and 2012 Associated with the Consumption of Pork Pies

An outbreak of listeriosis in England affecting 14 people between 2010 and 2012 and linked to the consumption of pork pies was investigated. All 14 individuals were older than 55 years, 12 were men, and 10 reported the presence of an underlying condition. All were resident in or had visited either of two English regions and were infected with the same strain of Listeria monocytogenes. In interviews with 12 patients, 9 reported eating pork pies, and individuals that consumed pork pies were significantly more likely to be infected with an outbreak strain than were individuals with sporadic cases of listeriosis infections in England from 2010 to 2012. Pork pies were purchased from seven retailers in South Yorkshire or the East Midlands, and the outbreak strain was recovered from pork pies supplied by only the producer in South Yorkshire. The outbreak strain was also recovered from samples of finished product and from environmental samples collected from the manufacturer. The likely source of contamination was environmental sites within the manufacturing environment, and the contamination was associated with the process of adding gelatin to the pies after cooking. Inadequate temperature control and poor hygienic practices at one of the retailers were also identified as possible contributory factors allowing growth of the pathogen. Following improvements in manufacturing practices and implementation of additional control measures at the retailers' premises, L. monocytogenes was not recovered from subsequent food and environmental samples, and the outbreak strain was not detected in further individuals with listeriosis in England.

NR4A3 controls CD8+ T cell metabolism and differentiation

Abstract Following an infection, naive CD8+ T cells expand and differentiate into two major populations of effectors: short-lived effector cells meant to die by apoptosis and memory precursor effector cells (MPECs) destined to survive as memory cells that will confer long-term protection. We postulated that the transcription factor NR4A3, a member of the orphan nuclear receptor family, whose expression is induced by TCR signalling, will regulate in vivo CD8+ T cell response. To elucidate the role of NR4A3 during CD8+ T cell response, we have adoptively transferred wild-type and Nr4a3−/− OT-I T cells (specific for the ovalbumin (OVA) peptide in the context of Kb) into naive recipients and analyzed their response following infection with a recombinant strain of Listeria monocytogenes encoding OVA. Although Nr4a3+/+ and Nr4a3−/− OT-I T cells expanded similarly, we observed an increased generation of MPECs at the peak of the T cell response, which led to an enhanced (2–3X) generation of memory T cells. Furthermore, Nr4a3−/− effector and memory T cells produce more cytokines than their wild-type counterpart. The analysis of the early T cell response demonstrated that NR4A3 controls the metabolic activity of activated T cells, as Nr4a3−/− OT-I T cells express less nutrient receptors and have reduced phosphorylation of S6. Altogether, these results suggest that NR4A3 controls effector and memory T cell differentiation by modulating their metabolic activity during the immune response.

Rifampicin- and Rifabutin-Resistant Listeria monocytogenes Strains Isolated from Food Products Carry Mutations in rpoB Gene.

The aim of this study was to investigate the mechanism of rifampicin resistance in Listeria monocytogenes strains isolated from different types of food and the impact of specific mutations in the rpoB gene on susceptibility to different antimicrobial agents and on fitness cost. Fifteen spontaneous rifampicin-resistant strains were selected. The DNA regions corresponding to clusters I-II, III, and N-terminal end of the rpoB gene of Escherichia coli were amplified and sequenced, leading to the identification of 10 different substitutions, nine of which (Ser466Pro, Gln470Lys Asp473Asn, Gly479Asp, His483Tyr/Arg/Asp, Arg486His, and Leu490Pro) were located in cluster I and one (Pro521Leu) in cluster II. From among these mutations, substitutions at positions 466, 470, 486, 490, and 521 have not been described for L. monocytogenes. Only substitutions at positions 470, 479, 483, and 486 lead to resistance to very high concentrations of rifampicin (minimum inhibitory concentration [MIC] ≥256 μg/mL) and rifabutin (MIC 128 μg/mL). Furthermore, mutations at positions 473, 490, and 521 had different effects on susceptibility to rifampicin compared to other bacterial species. A correlation between rifampicin resistance and susceptibility to a wide range of antimicrobials was determined. Substitutions in RpoB did not change the susceptibility of the mutants to different antimicrobials. The fitness of the mutants was assessed by paired competition experiments. Mutations at positions 470 and 479 were not associated with a reduction in fitness level. There was no correlation between the MIC of rifampicin and fitness cost. The risk of transmission of resistant strains through the food chain highlights the need for monitoring resistance, identifying mutant organisms, their genotypes, and their altered phenotypes to understand their dissemination.

Latex protein extracts from Calotropis procera with immunomodulatory properties protect against experimental infections with Listeria monocytogenes

BackgroundThe latex from the medicinal plant Calotropis procera is often used in folk medicine against infectious and inflammatory diseases. PurposeIn this study, we investigate a protein fraction with immunomodulatory properties, named LPPI, against experimental infections, in vitro and in vivo, with a virulent strain of Listeria monocytogenes. Study designLPPI was exposed to cultured macrophages or Swiss mice and then challenged with L. monocytogenes. MethodsPeritoneal macrophages were obtained from Swiss mice, and cultured in 96-well microplates. Soluble latex proteins (LP) were subjected to fractionation by ion-exchange chromatography. The major peak (LPPI) was added into wells at 10 or 100µg/ml. Albumin (100µg/ml) was used for comparison between protein treatments. After incubation for 1h at 5% CO2/ 37°C, the supernatant was discarded and 0.2ml of L. monocytogenes overnight culture was added in the wells. Following 4h and 24h infection, the cytokine mRNA expression was evaluated as well as the number of intracellular colony forming units. Swiss mice (n=16) were injected intraperitoneally (i.p.) with LPPI (5 and 10mg/kg) while the control mice received albumin (10mg/kg) or LP (10mg/kg). After 24h, all animal groups were challenged with L. monocytogenes (106 CFU/ ml), also by i.p. route. ResultsLPPI was not toxic to uninfected macrophages (pMØ) and significantly increased mRNA expression of TNF-α, IL-6, IL-1β and iNOS. Following infection, cell viability was reduced by 50% in albumin-treated pMØ (control); but only 17% in pMØ treated with LPPI at 100µg/ml. In this case, LPPI increased expression of TNF-α and IL-6 whereas the number of bacterial colony-forming units was reduced 100-fold in comparison to control groups. Swiss mice pretreated with LPPI showed dose-dependent survival rates that reached 80%, while mice that received albumin died 1–3 days after infection. After 24h infection, leukocyte migration to the infectious foci was high in LPPI-treated mice whereas the number of viable bacteria in the peritoneal fluid, liver and bloodstream were significantly reduced. ConclusionWe conclude that LPPI present immunomodulatory properties that are beneficial for prevention of systemic bacterial infections caused by the intracellular bacteria L. monocytogenes.

Characterization of Antimicrobial Resistance of Listeria monocytogenes Strains Isolated from a Pork Processing Plant and Its Respective Meat Markets in Southern China.

A total of 78 Listeria monocytogenes isolates from a pork processing plant and the respective meat markets in southern China were examined. This number includes 60 isolates from pork at markets, 5 from cooked pork products at markets, 10 from pork at a processing plant, and 3 from food-contact surfaces at the processing plant. All isolates were subjected to serotyping, antimicrobial susceptibility testing, pulsed-field gel electrophoresis (PFGE), and molecular basis of antibiotic resistance. Four serogroups were identified among the 78 tested isolates, with serogroup I (serotypes: 1/2a and 3a) being predominant (42.3%, 33/78). Antimicrobial resistance was most frequently observed for tetracycline (20.5%, 16/78), streptomycin (9.0%, 7/78), cefotaxime (7.7%, 6/78), and gentamicin (6.4%, 5/78). Multiple resistances occurred among 10.2% (8/78) isolates. All strains were sensitive to ampicillin, ampicillin/sulbactam, imipenem, ciprofloxacin, levofloxacin, trimethoprim/sulfamethoxazole, and vancomycin. Two isolates were resistant to five antimicrobials. Twelve strains carried tet(M) and located on Tn916. PFGE analysis revealed genetic heterogeneity among individual serotypes. Two predominant PFGE types were found persistent from the processing plant to markets indicating that these two types of isolates were able to survive under environmental adverse conditions from the processing plant to markets, which need to be monitored. Compared to samples from the pork processing plant, the prevalence of L. monocytogenes in meat market samples tended to be higher, serovar was more variable, and the antibiotic resistance range was wider, probably due to secondary contamination. Therefore, stringent hygiene measures and bacteriological controls should be observed to reduce the risk of transmission of L. monocytogenes from food to humans.

Molecular Typing and Epidemiology of Human Listeriosis Cases, Denmark, 2002-2012.

Denmark has a high incidence of invasive listeriosis (0.9 cases/100,000 population in 2012). We analyzed patient data, clinical outcome, and trends in pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST) of Listeria monocytogenes strains isolated in Denmark during 2002-2012. We performed 2-enzyme PFGE and serotyping on 559 isolates and MLST on 92 isolates and identified some correlation between molecular type and clinical outcome and patient characteristics. We found 178 different PFGE types, but isolates from 122 cases belonged to just 2 closely related PFGE types, clonal complex 8 and sequence type 8. These 2 types were the main cause of a peak in incidence of invasive listeriosis during 2005-2009, possibly representing an outbreak or the presence of a highly prevalent clone. However, current typing methods could not fully confirm these possibilities, highlighting the need for more refined discriminatory typing methods to identify outbreaks within frequently occurring L. monocytogenes PFGE types.