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PubMed1988

Pathogenicity of Listeria monocytogenes in comparison to other Listeria species.

In different mouse models the pathogenicity of various strains of Listeria monocytogenes and of other Listeria spp. was evaluated. Although quantitative differences of virulence among strains of L. monocytogenes are found, all isolates have to be regarded pathogenic unless essential virulence factors, such as protein composition of the cell wall or hemolysin production, are lacking. Besides L. monocytogenes, only Listeria ivanovii is able to multiply in the murine host, though definitely to a lesser extent. The other Listeria spp., such as Listeria innocua, Listeria welshimeri and Listeria seeligeri, have to be regarded as non-pathogenic.

Animals↗
PubMed1993

Reduction of ferric iron by Listeria monocytogenes and other species of Listeria.

One mechanism by which Listeria monocytogenes is thought to obtain iron required for growth is through the extracellular reduction of a ferric iron source to the ferrous form. To better characterize this reductase activity we have developed a simple plate assay that allows detection of colonies of Listeria species able to reduce ferric iron. Cells are plated on an agar base medium containing a ferric iron source and ethylenediamine dihydroxyphenylacetic acid. Colonies are then overlain with soft agarose containing NADH, flavin mononucleotide, and Ferrozine, a chelator of ferrous iron. Colonies able to reduce the ferric iron source form a red-purple color as the ferrous iron is complexed with ferrozine. Using this qualitative assay we have shown that all species of Listeria are able to reduce ferric iron when presented as ferric ammonium citrate whereas most other species of Gram-positive and Gram-negative bacteria are not. Only Clostridium perfringens was able to reduce ferric iron to the same extent as Listeria. Listeria monocytogenes was further shown to be capable of reducing various ferric iron salts as well as iron bound to ferritin, transferrin, and 2,3-dihydroxybenzoic acid in the agar plate assay. The utility of this assay was demonstrated by using it to screen a bank of Tn916-derived mutants of L. monocytogenes for clones unable to reduce ferric iron. Four such mutants were identified and all were shown to have greatly decreased ferric reductase activity.

FMN Reductase↗
PubMed1994

Infection of murine fetal brain cell cultures with Listeria monocytogenes.

The uptake of Listeria monocytogenes by different brain cells was studied in primary dissociated brain cell cultures derived from murine fetuses. In respect to the supposed intraaxonal migration of Listeria monocytogenes in the pathogenesis of listeric focal brain stem encephalitis, it was examined whether the bacterium was internalized by neurons. Infection rates of distinct cell types were determined by double immunofluorescence with antibodies against cell type-specific markers and the bacterial pathogen. Because of the changing composition of the cultures and time-dependent expression of the oligodendrocyte marker galactocerebroside (GC), infections were carried out on day 4, 6, 8, and 15 in vitro. Listeria monocytogenes was detected predominantly within macrophages. Astrocytes, oligodendrocytes, and fibronectin-expressing cells were infected to a lesser extent. The lowest rates of infection were observed in neurons. A tropism of Listeria monocytogenes for neurons was not detected in vitro.

Animals↗
PubMed1992

Detection of genes coding for listeriolysin and Listeria monocytogenes antigen A (ImaA) in Listeria spp. by the polymerase chain reaction.

Two pairs of synthetic oligonucleotide primers were used in a polymerase chain reaction (PCR) protocol to detect targeted sequences in genes coding for listeriolysin O and Listeria monocytogenes antigen A (ImaA). Strains of Listeria spp. used in this study were isolated from clinical specimens, contaminated foods, and environmental sources. Primers were targeted to internal regions of the genes coding for listeriolysin (hlyA) and Listeria antigen (ImaA) and amplification fragments were detected after the PCR by agarose gel electrophoresis. PCR was performed using nucleic acids extracted from a collection of 74 strains of Listeria spp. including 18 reference strains, 41 L. monocytogenes, nine L. innocua, five L. seeligeri and one L. ivanovii, encompassing representative sources, serovars, and enzyme electrophoretic types. Although the listeriolysin gene was found exclusively in L. monocytogenes, some strains of serovar 4c were negative. Simultaneous presence of both genes was restricted to L. monocytogenes strains of serovars 1/2, 3, and 4. The ImaA gene was identified in five of 10 L. innocua strains and one L. ivanovii isolated from pork. Strains of L. seeligeri, L. welshimeri, and L. grayi were negative for both genes. The detection limits in the PCR were found to be 10 pg of nucleic acids for the hlyA gene and 1 pg for the ImaA gene.

Antigens, Bacterial↗
PubMed1999

Evaluation of an rDNA Listeria probe for Listeria monocytogenes typing.

A Listeria monocytogenes DNA fragment, identified as part of the 23S rRNA gene and called B17, was used to type 266 L. monocytogenes strains and 43 strains of other Listeria species. Results were compared with those obtained: i) with pBA2 (which consists of a 2.3 kb Bacillus subtilis DNA fragment encoding 16S rRNA, inserted into the HindIII site of pBR322), a probe previously used for Listeria and L. monocytogenes ribotyping, and ii) with DNA macrorestriction profiles analysis. Twenty profiles were identified for L. monocytogenes using pB 17, three of which accounted for 87% of strains. This new rDNA probe had greater discriminatory power for serogroups 1/2 or 3 strains than for serogroup 4 strains. The number of varieties and the discrimination index were higher with this new probe than with pBA2, but DNA macrorestriction patterns analysis gave better discrimination between strains.

Animals↗
PubMed1995

Repression of motility and flagellin production at 37 degrees C is stronger in Listeria monocytogenes than in the nonpathogenic species Listeria innocua.

Listeria monocytogenes and Listeria innocua differ markedly in virulence but are indistinguishable by classical taxonomic criteria. Both species are actively motile and produce abundant flagellin at 22 degrees C. We have found, however, noticeable differences between L. monocytogenes and L. innocua in motility and flagellin production at 37 degrees C. At this temperature, L. monocytogenes strains were virtually nonmotile and produced little or no detectable flagellin, whereas strains of L. innocua were frequently motile and produced substantial amounts of flagellin. This flagellin was recognized by a Listeria genus-specific monoclonal antibody that also recognized flagellin produced at 22 degrees C. These results suggest differential regulation of flagellin production between L. monocytogenes and L. innocua at 37 degrees C.

Antibodies, Bacterial↗
PubMed1987

Antigen-specific augmentation factor involved in murine delayed-type footpad reaction. II. Augmentation of delayed-type footpad reaction and acquired resistance to Listeria monocytogenes by transfer of Listeria-immune serum.

We previously found an antigen-specific factor capable of augmenting delayed-type hypersensitivity (DTH) in the culture supernatant of the mixture of immune spleen cells and erythrocyte antigen, or in the serum of mice immunized with heterologous erythrocytes and exhibiting delayed-type footpad reaction. To elucidate whether this kind of factor (DTH-augmentation factor; DAF) participates in the establishment of DTH to various kinds of antigen besides erythrocyte antigen, we chose a bacterial antigen, Listeria monocytogenes, which is a facultative intracellular bacterium. In the present study, we demonstrated that the immune serum from mice immunized with viable Listeria augmented the delayed-type footpad reaction to Listeria. Furthermore, acquired resistance against Listeria was also augmented by the transfer of such immune serum. Such augmentation of acquired resistance was observed in sites infected locally and in the spleen of mice infected systemically. This effect was also seen in sera from mice immunized with heat-killed Listeria emulsified with complete Freund's adjuvant.

Animals↗
PubMed1988

Minimum growth temperatures of Listeria monocytogenes and non-haemolytic Listeria.

Minimum growth temperatures and those of decreased growth were determined for 100 strains of listerias. The ability of 78 strains of Listeria monocytogenes isolated from animals and 22 non-haemolytic strains to grow at low temperatures was studied, using a flooding technique, in a plate-type continuous temperature gradient incubator at temperatures between -1.6 and 14.5 degrees C. The mean minimum temperature for L. monocytogenes was +1.7 +/- 0.5 degrees C. The growth of non-haemolytic listerias was unobservable at +1.7 +/- 0.5 degrees C. The L. monocytogenes strains grew at about 0.6 degrees C lower than the non-pathogenic strains. No differences in growth temperatures were observed among L. monocytogenes strains isolated from different sources. The serovars with the OI antigen grew at lower temperatures (+1.0 +/- 0.3 degrees C) than the other common serovar 4b (+1.3 +/- 0.4 degrees C). The results indicate that L. monocytogenes grows better than non-haemolytic strains under cold conditions. The possible role of haemolysins as growth factors is also discussed.

Animals↗
PubMed2000

Identification of new loci involved in adhesion of Listeria monocytogenes to eukaryotic cells. European Listeria Genome Consortium.

Insertional mutagenesis was performed with Tn1545 in the genetic background of an inIAB deletion mutant to identify new adhesion determinants in Listeria monocytogenes. Four insertion mutants defective in adhesion to eukaryotic cells were identified. Insertion sites were cloned by inverse-PCR and sequenced. The genetic organization of insertion regions was further analysed by screening and sequencing DNA fragments from a HindIII library and by searching databases. Three adhesion-defective mutants each had one copy of Tn1545 inserted into their chromosome. The insertion sites were different in the three mutants: (i) upstream from two ORFs in tandem, similar to dfp and priA of Bacillus subtilis, respectively; (ii) within an ORF encoding a putative 126 amino-acid-polypeptide with no significant similarity to any known protein; (iii) within an ORF similar to a B. subtilis ORF with no known function, just upstream from an operon similar to an ABC (ATP-binding cassette) transporter operon from B. subtilis. The excisants obtained from these mutants using the excision reporter plasmid pTCR9 recovered full adhesion capacity. A fourth mutant was the most severely defective in adhesion. It had five Tn1545 insertions, one of which was upstream from dfp and priA, and another of which was upstream from ami, a gene encoding a surface-exposed autolysin with a C terminus similar to that of InIB. Ami was clearly involved because an ami null mutant constructed in an EGDdeltainIA-F background was adhesion-defective. Thus new regions involved in the adhesion of L. monocytogenes to eukaryotic cells were identified. Further study is required to define more accurately the roles of these regions in the adhesion process itself.

Bacterial Adhesion↗
PubMed1994

The virulence gene cluster of Listeria monocytogenes is also present in Listeria ivanovii, an animal pathogen, and Listeria seeligeri, a nonpathogenic species.

Most known Listeria monocytogenes virulence genes cluster within a 9.6-kb chromosomal region. This region is flanked on one end by two uncharacterized open reading frames (ORF A and ORF B) and ldh, an ORF presumably encoding the L. monocytogenes lactate dehydrogenase (J.-A. Vazquez-Boland, C. Kocks, S. Dramsi, H. Ohayon, C. Geoffroy, J. Mengaud, and P. Cossart, Infect. Immun. 60:219-230, 1992). We report here that the other end is flanked by prs, and ORF homologous to phosphoribosyl PPi synthetase genes. ORF B and prs were detected in all Listeria species and thus delimit the virulence region. This virulence gene cluster was detected exclusively in hemolytic Listeria species, Listeria ivanovii, an animal pathogen, and Listeria seeligeri, a nonpathogenic species.

Animals↗
PubMed2025

Characterization of a novel putative lantibiotic biosynthesis genomic island in emerging clones of Listeria monocytogenes serotype 4b.

Listeria monocytogenes is a Gram-positive facultative intracellular bacterium that is ubiquitous in nature and the causative agent of listeriosis. The outbreak-derived serotype 4b strain L. monocytogenes strain WS1, sequence type (ST) 558, sublineage (SL) 558, was previously found to have unusual pathogenicity, with ability to cause fetal damage in the first trimester of pregnancy. Search of the WS1 genome for novel and unique genomic features identified a putative lantibiotic island on the chromosome of WS1 and all tested strains of SL558 and two other putative emerging serotype 4b clones, clonal complex 554 (SL554 and SL555) and ST782 (SL782), but absent from all other major clones of L. monocytogenes. The island was deleted from four strains, including two each of ST558 and ST554. The deletions did not impact virulence in a Galleria mellonella model but consistently resulted in reduced hemolytic activity. In addition, we noted strain-dependent impacts on biofilm formation. Additional studies will be necessary to further elucidate the roles of this genomic island in the adaptive physiology and virulence of L. monocytogenes.

Listeria monocytogenes↗
PubMedDate not supplied

Sites of action by propionate on Listeria monocytogenes.

Exposure of Listeria monocytogenes to a solution of sodium propionate (8% w/v) for 60 min caused 87% of the population to be injured. Injury was evidenced by inability of the bacterium to tolerate 6% sodium chloride in tryptose agar as compared to ability to grow on tryptose agar with no added salt. Injured cells were allowed to repair in tryptose broth and the repair process was studied by addition to tryptose broth of sublethal amounts of metabolic or synthetic inhibitors. Repair of injured cells did not require electron transport or synthesis of cell wall components, mRNA or protein. No changes which may have occurred in the cell membrane of injured cells, allowed leakage of proteins or nucleotides into the medium. Exogenous cation salts enhanced the rate of recovery of injured cells. The specific activity of lactic dehydrogenase was reduced in propionate-injured L. monocytogenes.

Colony Count, Microbial↗
PubMed2003

The effect of processed meat and meat starter cultures on gastrointestinal colonization and virulence of Listeria monocytogenes in mice.

Listeria monocytogenes is a foodborne pathogen of major concern to the food industry in general and the meat industry in particular. The aim of this study was firstly to identify a strain of Listeria that was virulent in SPF BALB/c mice. Secondly, to investigate if a traditional meat starter culture (FloraCarn) and nontraditional meat starter (NTMS) cultures of dairy product and human origin (Lactobacillus and bifidobacteria) inhibit this pathogen in vivo. In addition, the inhibition of Listeria was investigated in vitro. In vitro inhibition was investigated using an agar inhibition assay, where soft agar containing the pathogen was laid over colonies of NTMS cultures, and inhibition expressed as the zones of inhibition developing around the colonies. For assessment of virulence, mice were intragastrically challenged with broth cultures of five strains of Listeria. For assessment of anti-listeria effect in vivo, the Listeria strain proven to be most pathogenic (LM3) was given to mice in salami batter containing no other added cultures (control) or batter inoculated with either (1) FloraCarn, (2) a NTMS culture, or (3) a combination of FloraCarn and a NTMS culture. The batter was given to mice after a 3-day fermentation and faecal levels of pathogen and body weight were monitored. Intragastric challenge with LM3, but no other strains, resulted in a significant weight loss (p<0.05) and up to 10(6) colony forming units (cfu) of LM3 per gram faeces. No weight loss was observed in animals fed with salami batter containing LM3. Consumption of salami batter fermented by a combination of NTMS culture (Lactobacillus acidophilus LAFTI(R) L10) and FloraCarn reduced faecal levels of the pathogen by 2.5 log units compared to the control. Consumption of salami batter fermented with FloraCarn and LAFTI(R) L10 (L10) alone reduced faecal levels by 0.5-1 and 1.5 log units, respectively. Of the NTMS cultures investigated here, L10 displayed the greatest inhibition of LM3 in vitro. These results indicate that the ability of pathogenic Listeria to cause listeriosis is dependent on the nature of the food in which the pathogen is present, and that a traditional meat starter culture (FloraCarn) and some NTMS cultures, particularly L10, inhibit growth of the pathogen during passage through the gastrointestinal tract.

Animals↗
PubMed2000

Comparative evaluation of adhesion and biofilm formation of different Listeria monocytogenes strains.

Thirteen Listeria monocytogenes strains were used to grow biofilms on glass surfaces in static conditions at 37 degrees C for up to 4 days. After the initial 3-h adhesion and in subsequent 1-day intervals, cell numbers were determined using standard plate count after swabbing the cells from the glass surface. The three-dimensional structure of in situ biofilms was determined by confocal scanning laser microscopy (CSLM). After 3 h incubation, bacterial cells for all 13 strains of L. monocytogenes were found attached to glass slides and all strains formed biofilms within 24 h. The strains varied significantly in their ability to adhere to the surface and significant differences for cell numbers after 24 h biofilm growth were found. Cell counts in biofilms formed by five L. monocytogenes strains were monitored over 4 days. The counts increased for the first 2 days reaching 10(5) cfu/cm2, except for L. monocytogenes 7148 (10(4) cfu/cm2). After 2 days, cell counts remained at 10(5) cfu/cm2 for four strains (tested on days 3 and 4), while L. monocytogenes 7148 continued to grow and reached 10(5) cfu/cm2 on day 4. This difference in biofilm growth was not related to variations in growth rates of planktonic cells suggesting that growth behaviour of Listeria in biofilms may be different from their planktonic growth. CSLM revealed that the biofilms grown under static conditions consisted of two distinct layers with 0.5 log10 higher cell numbers in the bottom layer as compared to the upper layer.

Bacterial Adhesion↗
PubMed2004

Methods for the isolation and identification of Listeria spp. and Listeria monocytogenes: a review.

Listeria monocytogenes is an important food-borne pathogen and is widely tested for in food, environmental and clinical samples. Identification traditionally involved culture methods based on selective enrichment and plating followed by the characterization of Listeria spp. based on colony morphology, sugar fermentation and haemolytic properties. These methods are the gold standard; but they are lengthy and may not be suitable for testing of foods with short shelf lives. As a result more rapid tests were developed based on antibodies (ELISA) or molecular techniques (PCR or DNA hybridization). While these tests possess equal sensitivity, they are rapid and allow testing to be completed within 48 h. More recently, molecular methods were developed that target RNA rather than DNA, such as RT-PCR, real time PCR or nucleic acid based sequence amplification (NASBA). These tests not only provide a measure of cell viability but they can also be used for quantitative analysis. In addition, a variety of tests are available for sub-species characterization, which are particularly useful in epidemiological investigations. Early typing methods differentiated isolates based on phenotypic markers, such as multilocus enzyme electrophoresis, phage typing and serotyping. These phenotypic typing methods are being replaced by molecular tests, which reflect genetic relationships between isolates and are more accurate. These new methods are currently mainly used in research but their considerable potential for routine testing in the future cannot be overlooked.

Animals↗
PubMed2005

Alternative sigma factor sigmaB is not essential for listeria monocytogenes surface attachment.

Listeria monocytogenes is a foodborne pathogen frequently isolated from the food processing environment. Multiple lines of evidence suggested a possible role for the L. monocytogenes alternative transcription factor sigma B (sigmaB) in surface attachment and biofilm formation. Therefore, through plate count and microscopic techniques, the L. monocytogenes 10403S strain and an otherwise isogenic deltasigB strain were tested for attachment to stainless steel. Analysis of microscopic images revealed that after 72 h at 24 degrees C under static conditions the tested L. monocytogenes strains attached uniformly to surfaces as single cells. Both strains were capable of rapid attachment (i.e., numbers of attached cells were essentially the same after either 5 min or 24 h of incubation). Numbers of attached deltasigB cells were significantly lower than those of the wild-type strain after 48 and 72 h of incubation at 24 degrees C (P = 0.001). Similar numbers of the deltasigB strain attached to stainless steel regardless of temperature (24 or 37 degrees C); however, deltasigB cells attached at higher relative numbers in the presence of 6% NaCl after 48 and 72 h. Furthermore, in the presence of Pseudomonas fluorescens, similarly high numbers of wild-type and deltasigB cells attached to the surfaces, forming mixed biofilms. Our data suggest that sigmaB is not required for initial surface attachment of L. monocytogenes.

Bacterial Adhesion↗
PubMed2003

Role of ctc from Listeria monocytogenes in osmotolerance.

Listeria monocytogenes is a food-borne pathogen with the ability to grow under conditions of high osmolarity. In a previous study, we reported the identification of 12 proteins showing high induction after salt stress. One of these proteins is highly similar to the general stress protein Ctc of Bacillus subtilis. In this study, induction of Ctc after salt stress was confirmed at the transcriptional level by using RNA slot blot experiments. To explore the role of the ctc gene product in resistance to stresses, we constructed a ctc insertional mutant. No difference in growth was observed between the wild-type strain LO28 and the ctc mutant either in rich medium after osmotic or heat stress or in minimal medium after heat stress. However, in minimal medium after osmotic stress, the growth rate of the mutant was increased by a factor of 2. Moreover, electron microscopy analysis showed impaired morphology of the mutant grown under osmotic stress conditions in minimal medium. Addition of the osmoprotectant glycine betaine to the medium completely abolished the osmotic sensitivity phenotype of the ctc mutant. Altogether, these results suggest that the Ctc protein of L. monocytogenes is involved in osmotic stress tolerance in the absence of any osmoprotectant in the medium.

Adaptation, Physiological↗