Novel cost-efficient real-time PCR assays for detection and quantitation of Listeria monocytogenes

Listeriosis is a serious food-borne infection with mortality rates approaching 30%. Therefore, the rapid, cost-effective, and automated detection of Listeria monocytogenes throughout the food chain continues to be a major concern. Here we describe three novel quantitative real-time PCR assays for L. monocytogenes based on amplification of a target hlyA gene with SYBR Green I chemistry and hydrolysis probe (TaqMan MGB probe). In order to offer sensitive, rapid and robust tool of additional economical value the real-time PCR assays were designed and optimized to only 5 μl-reactions. All assays were evaluated by using different non-reference Listeria strains isolated from various food matrices. Results demonstrated specificity to L. monocytogenes with accurate quantification over a dynamic range of 5-6 log units with R² higher than 0.98 and amplification efficiencies reaching above 92%. The detection and quantification limits were as low as 165 genome equivalents. Comparison of novel assays to commercially available TaqMan® Listeria monocytogenes Detection Kit and previously published studies revealed similar specificity, sensitivity and efficiency, but greater robustness and especially cost-efficiency in the view of smaller reaction volumes and continuous increase in sample throughput.     

Metabolic Responses of Primary and Transformed Cells to Intracellular Listeria monocytogenes

The metabolic response of host cells, in particular of primary mammalian cells, to bacterial infections is poorly understood. Here, we compare the carbon metabolism of primary mouse macrophages and of established J774A.1 cells upon Listeria monocytogenes infection using ¹³C-labelled glucose or glutamine as carbon tracers. The ¹³C-profiles of protein-derived amino acids from labelled host cells and intracellular L. monocytogenes identified active metabolic pathways in the different cell types. In the primary cells, infection with live L. monocytogenes increased glycolytic activity and enhanced flux of pyruvate into the TCA cycle via pyruvate dehydrogenase and pyruvate carboxylase, while in J774A.1 cells the already high glycolytic and glutaminolytic activities hardly changed upon infection. The carbon metabolism of intracellular L. monocytogenes was similar in both host cells. Taken together, the data suggest that efficient listerial replication in the cytosol of the host cells mainly depends on the glycolytic activity of the hosts.     

Epigallocatechin gallate inhibits intracellular survival of Listeria monocytogenes in macrophages

Epigallocatechin gallate (EGCg), the major tea catechin, is known as a potent anti-microbial and anti-tumor compound. The effects of EGCg on host defense mechanisms against Listeria monocytogenes infection were examined in vitro using mouse peritoneal exudate cells. The study showed that EGCg inhibited the intracellular growth of L. monocytogenes in macrophages. The enhancement of in vitro anti-L. monocytogenes activity by EGCg is not due to the modulation of reactive oxygen intermediates or the production of reactive nitrogen intermediates but due to the inhibition of its escaping from the phagosome into cytosolic space. Anti-L. monocytogenes of EGCg is through the inhibition of hemolytic and cholesterol-binding activity of listeriolysin O, which usually disrupts the phagosomal membrane in the escaping phase of L. monocytogenes.     

Spontaneous bacteriocin resistance in Listeria monocytogenes as a susceptibility screen for identifying different mechanisms of resistance and modes of action by bacteriocins of lactic acid bacteria

A practical system was devised for grouping bacteriocins of lactic acid bacteria (LAB) based on mode of action as determined by changes in inhibitory activity to spontaneously-acquired bacteriocin resistance (Bac^R). Wild type Listeria monocytogenes 39-2 was sensitive to five bacteriocins produced by 3 genera of LAB: pediocin PA-1 and pediocin Bac3 (Pediococcus), lacticin FS97 and lacticin FS56 (Lactococcus), and curvaticin FS47 (Lactobacillus). A spontaneous Bac^R derivative of L. monocytogenes 39-2 obtained by selective recovery against lacticin FS56 provided complete resistance to the bacteriocin made by Lactococcus lactis FS56. The lacticin FS56-resistant strain of L. monocyotgenes 39-2 was also cross-resistant to curvaticin FS47 and pediocin PA-1, but not to lacticin FS97 or pediocin Bac3. The same pattern of cross-resistance was also observed with Bac^R isolates obtained with L. monocytogenes Scott A-2. A spontaneous mutation that renders a strain cross-resistant to different bacteriocins indicates that they share a common mechanism of resistance due to similar modes of action of the bacteriocins. Spontaneous resistance was acquired to other bacteriocins (in aggregate) by following the same procedure against which the Bac^R strain was still sensitive. In subsequent challenge assays, mixtures of bacteriocins of different modes of action provided greater inhibition than mixtures of bacteriocins of the same mode of action (as determined by our screening method). This study identifies a methodical approach to classify bacteriocins into functional groups based on mechanism of resistance (i.e., mode of action) that could be used for identifying the best mixture of bacteriocins for use as biopreservatives.     

InlA Promotes Dissemination of Listeria monocytogenes to the Mesenteric Lymph Nodes during Food Borne Infection of Mice

Intestinal Listeria monocytogenes infection is not efficient in mice and this has been attributed to a low affinity interaction between the bacterial surface protein InlA and E-cadherin on murine intestinal epithelial cells. Previous studies using either transgenic mice expressing human E-cadherin or mouse-adapted L. monocytogenes expressing a modified InlA protein (InlA^m) with high affinity for murine E-cadherin showed increased efficiency of intragastric infection. However, the large inocula used in these studies disseminated to the spleen and liver rapidly, resulting in a lethal systemic infection that made it difficult to define the natural course of intestinal infection. We describe here a novel mouse model of oral listeriosis that closely mimics all phases of human disease: (1) ingestion of contaminated food, (2) a distinct period of time during which L. monocytogenes colonize only the intestines, (3) varying degrees of systemic spread in susceptible vs. resistant mice, and (4) late stage spread to the brain. Using this natural feeding model, we showed that the type of food, the time of day when feeding occurred, and mouse gender each affected susceptibility to L. monocytogenes infection. Co-infection studies using L. monocytogenes strains that expressed either a high affinity ligand for E-cadherin (InlA^m), a low affinity ligand (wild type InlA from Lm EGDe), or no InlA (ΔinlA) showed that InlA was not required to establish intestinal infection in mice. However, expression of InlA^m significantly increased bacterial persistence in the underlying lamina propria and greatly enhanced dissemination to the mesenteric lymph nodes. Thus, these studies revealed a previously uncharacterized role for InlA in facilitating systemic spread via the lymphatic system after invasion of the gut mucosa.     

Recombinant probiotic expressing Listeria adhesion protein attenuates Listeria monocytogenes virulence in vitro

Background

Listeria monocytogenes, an intracellular foodborne pathogen, infects immunocompromised hosts. The primary route of transmission is through contaminated food. In the gastrointestinal tract, it traverses the epithelial barrier through intracellular or paracellular routes. Strategies to prevent L. monocytogenes entry can potentially minimize infection in high-risk populations. Listeria adhesion protein (LAP) aids L. monocytogenes in crossing epithelial barriers via the paracellular route. The use of recombinant probiotic bacteria expressing LAP would aid targeted clearance of Listeria from the gut and protect high-risk populations from infection.

Methodology/Principal Findings

The objective was to investigate the ability of probiotic bacteria or LAP-expressing recombinant probiotic Lactobacillus paracasei (Lbp^LAP) to prevent L. monocytogenes adhesion, invasion, and transwell-based transepithelial translocation in a Caco-2 cell culture model. Several wild type probiotic bacteria showed strong adhesion to Caco-2 cells but none effectively prevented L. monocytogenes infection. Pre-exposure to Lbp^LAP for 1, 4, 15, or 24 h significantly (P<0.05) reduced adhesion, invasion, and transepithelial translocation of L. monocytogenes in Caco-2 cells, whereas pre-exposure to parental Lb. paracasei had no significant effect. Similarly, Lbp^LAP pre-exposure reduced L. monocytogenes translocation by as much as 46% after 24 h. Lbp^LAP also prevented L. monocytogenes-mediated cell damage and compromise of tight junction integrity. Furthermore, Lbp^LAP cells reduced L. monocytogenes-mediated cell cytotoxicity by 99.8% after 1 h and 79% after 24 h.

Conclusions/Significance

Wild type probiotic bacteria were unable to prevent L. monocytogenes infection in vitro. In contrast, Lbp^LAP blocked adhesion, invasion, and translocation of L. monocytogenes by interacting with host cell receptor Hsp60, thereby protecting cells from infection. These data show promise for the use of recombinant probiotics in preventing L. monocytogenes infection in high-risk populations.     

Listeria monocytogenes-infected human umbilical vein endothelial cells: internalin-independent invasion, intracellular growth, movement, and host cell responses

The interaction of Listeria monocytogenes with human umbilical vein endothelial cells was studied. We show that L. monocytogenes invades human umbilical vein endothelial cells independently of internalin A, internalin B, internalin C, and ActA. L. monocytogenes replicates efficiently inside the cells and moves intracellularly by the induction of actin polymerization. We further show that L. monocytogenes-infection of human umbilical vein endothelial cells induces interleukin-6 and interleukin-8 expression during the first 6 h of infection. The expression of MCP-1 and the adhesion molecules VCAM-1 and ICAM-1 was not altered under the experimental conditions used here.     

Mitogen-activated protein kinases are required for effective infection of human choroid plexus epithelial cells by Listeria monocytogenes

Listeria monocytogenes, a Gram-positive bacterium, can cause meningitis after invading the human central nervous system. The blood-cerebrospinal fluid barrier (BCSFB), located at the epithelium of the choroid plexus, is a possible entry site for L. monocytogenes into the brain, and in vitro L. monocytogenes invades human choroid plexus epithelial papilloma (HIBCPP) cells. Although host cell signal transduction subsequent to infection by L. monocytogenes has been investigated, the role of mitogen-activated protein kinases (MAPK) is not clarified yet. We show that infection with L. monocytogenes causes activation of the MAPKs Erk1/2 and p38 preferentially when bacteria are added to the physiologically more relevant basolateral side of HIBCPP cells. Deletion of the listerial virulence factors Internalin (InlA) and InlB reduces MAPK activation. Whereas inhibition of either Erk1/2 or p38 signaling significantly attenuates infection of HIBCPP cells with L. monocytogenes, simultaneous inhibition of both MAPK pathways shows an additive effect, and Erk1/2 and p38 are involved in regulation of cytokine and chemokine expression following infection. Blocking of endocytosis with the synthetic dynamin inhibitor dynasore strongly abrogates infection of HIBCPP cells with L. monocytogenes. Concurrent inhibition of MAPK signaling further reduces infection, suggesting MAPKs mediate infection with L. monocytogenes during inhibition of dynamin-mediated endocytosis.     

Quantitative Detection of Listeria monocytogenes in Biofilms by Real-Time PCR

A quantitative method based on a real-time PCR assay to enumerate Listeria monocytogenes in biofilms was developed. The specificity for L. monocytogenes of primers targeting the listeriolysin gene was demonstrated using a SYBR Green I real-time PCR assay. The number of L. monocytogenes detected growing in biofilms was 6 × 10² CFU/cm².     

Src-dependent Tyrosine Phosphorylation of Non-muscle Myosin Heavy Chain-IIA Restricts Listeria monocytogenes Cellular Infection

Bacterial pathogens often interfere with host tyrosine phosphorylation cascades to control host responses and cause infection. Given the role of tyrosine phosphorylation events in different human infections and our previous results showing the activation of the tyrosine kinase Src upon incubation of cells with Listeria monocytogenes, we searched for novel host proteins undergoing tyrosine phosphorylation upon L. monocytogenes infection. We identify the heavy chain of the non-muscle myosin IIA (NMHC-IIA) as being phosphorylated in a specific tyrosine residue in response to L. monocytogenes infection. We characterize this novel post-translational modification event and show that, upon L. monocytogenes infection, Src phosphorylates NMHC-IIA in a previously uncharacterized tyrosine residue (Tyr-158) located in its motor domain near the ATP-binding site. In addition, we found that other intracellular and extracellular bacterial pathogens trigger NMHC-IIA tyrosine phosphorylation. We demonstrate that NMHC-IIA limits intracellular levels of L. monocytogenes, and this is dependent on the phosphorylation of Tyr-158. Our data suggest a novel mechanism of regulation of NMHC-IIA activity relying on the phosphorylation of Tyr-158 by Src.     

The Function of the Chemokine Receptor CXCR6 in the T Cell Response of Mice against Listeria monocytogenes

The chemokine receptor CXCR6 is expressed on different T cell subsets and up-regulated following T cell activation. CXCR6 has been implicated in the localization of cells to the liver due to the constitutive expression of its ligand CXCL16 on liver sinusoidal endothelial cells. Here, we analyzed the role of CXCR6 in CD8⁺ T cell responses to infection of mice with Listeria monocytogenes. CD8⁺ T cells responding to listerial antigens acquired high expression levels of CXCR6. However, deficiency of mice in CXCR6 did not impair control of the L. monocytogenes infection. CXCR6-deficient mice were able to generate listeria-specific CD4⁺ and CD8⁺ T cell responses and showed accumulation of T cells in the infected liver. In transfer assays, we detected reduced accumulation of listeria-specific CXCR6-deficient CD8⁺ T cells in the liver at early time points post infection. Though, CXCR6 was dispensable at later time points of the CD8⁺ T cell response. When transferred CD8⁺ T cells were followed for extended time periods, we observed a decline in CXCR6-deficient CD8⁺ T cells. The manifestation of this cell loss depended on the tissue analyzed. In conclusion, our results demonstrate that CXCR6 is not required for the formation of a T cell response to L. monocytogenes and for the accumulation of T cells in the infected liver but CXCR6 appears to influence long-term survival and tissue distribution of activated cells.     

Listeria monocytogenes Alters Mast Cell Phenotype,Mediator and Osteopontin Secretion in a Listeriolysin-Dependent Manner

Whilst mast cells participate in the immune defence against the intracellular bacterium Listeria monocytogenes, there is conflicting evidence regarding the ability of L. monocytogenes to infect mast cells. It is known that the pore-forming toxin listeriolysin (LLO) is important for mast cell activation, degranulation and the release of pro-inflammatory cytokines. Mast cells, however, are a potential source of a wide range of cytokines, chemokines and other mediators including osteopontin, which contributes to the clearing of L. monocytogenes infections in vivo, although its source is unknown. We therefore aimed to resolve the controversy of mast cell infection by L. monocytogenes and investigated the extent of mediator release in response to the bacterium. In this paper we show that the infection of bone marrow-derived mast cells by L. monocytogenes is inefficient and LLO-independent. LLO, however, is required for calcium-independent mast cell degranulation as well as for the transient and selective downregulation of cell surface CD117 (c-kit) on mast cells. We demonstrate that in addition to the key pro-inflammatory cytokines TNF-α and IL-6, mast cells release a wide range of other mediators in response to L. monocytogenes. Osteopontin, IL-2, IL-4, IL-13 and granulocyte macrophage colony-stimulating factor (GM-CSF), and chemokines including CCL2, CCL3, CCL4 and CCL5 are released in a MyD88-dependent manner. The wide range of mediators released by mast cells in response to L. monocytogenes may play an important role in the recruitment and activation of a variety of immune cells in vivo. The cocktail of mediators, however, is unlikely to skew the immune response to a particular effector response. We propose that mast cells provide a hitherto unreported source of osteopontin, and may provide an important role in co-ordinating the immune response during Listeria infection.     

Listeriolysin O Is Necessary and Sufficient to Induce Autophagy during Listeria monocytogenes Infection

Background

Recent studies have suggested that autophagy is utilized by cells as a protective mechanism against Listeria monocytogenes infection.

Methodology/Principal Findings

However we find autophagy has no measurable role in vacuolar escape and intracellular growth in primary cultured bone marrow derived macrophages (BMDMs) deficient for autophagy (atg5^−/−). Nevertheless, we provide evidence that the pore forming activity of the cholesterol-dependent cytolysin listeriolysin O (LLO) can induce autophagy subsequent to infection by L. monocytogenes. Infection of BMDMs with L. monocytogenes induced microtubule-associated protein light chain 3 (LC3) lipidation, consistent with autophagy activation, whereas a mutant lacking LLO did not. Infection of BMDMs that express LC3-GFP demonstrated that wild-type L. monocytogenes was encapsulated by LC3-GFP, consistent with autophagy activation, whereas a mutant lacking LLO was not. Bacillus subtilis expressing either LLO or a related cytolysin, perfringolysin O (PFO), induced LC3 colocalization and LC3 lipidation. Further, LLO-containing liposomes also recruited LC3-GFP, indicating that LLO was sufficient to induce targeted autophagy in the absence of infection. The role of autophagy had variable effects depending on the cell type assayed. In atg5^−/− mouse embryonic fibroblasts, L. monocytogenes had a primary vacuole escape defect. However, the bacteria escaped and grew normally in atg5^−/− BMDMs.

Conclusions/Significance

We propose that membrane damage, such as that caused by LLO, triggers bacterial-targeted autophagy, although autophagy does not affect the fate of wild-type intracellular L. monocytogenes in primary BMDMs.     

Repeated daily doses do not increase Listeria monocytogenes infection in ewes as shown by faecal excretion and serological monitoring

Ruminants fed contaminated forage may shed Listeria monocytogenes in their faeces, and prolonged low daily doses of L. monocytogenes could cause listerial infection [Maijala, R., Lyytikainen, O., Autio, T., Aalto, T., Haavisto, L., Honkanen-Buzalski, T., 2001. Exposure of Listeria monocytogenes within an epidemic caused by butter in Finland. Int. J. Food Microbiol. 70, 97–109]. To compare listerial infection following single or repeated doses and the contamination of the environment with the excreted bacteria, ewes were orally inoculated with either 10⁴, 10⁶ or 10¹⁰ cfu L. monocytogenes once, or daily for 10 days. Serological responses were monitored with indirect ELISAs using recombinant listeriolysin O (LLO), internalin A (InlA) and internalin A-related protein (IrpA). The 24 inoculated animals displayed no symptoms, except for a transient hyperthermia in two animals given 10¹⁰ cfu. One ewe died on day 9 after non-listerial mastitis followed by listerial septicaemia. L. monocytogenes was recovered from day 1 post-inoculation until day 17 from the faeces of ewes inoculated with 10⁶ or 10¹⁰ cfu. No antibodies were detected in ewes given 10⁴ or 10⁶ cfu. Anti-LLO and anti-IrpA antibodies were detected from day 15 in animals inoculated with 10¹⁰ cfu, and this strengthened the conclusion that these long-lasting shedders were infected but asymptomatic carriers. An anti-InlA response was detected only at a very low level. These results suggest that repeated daily doses are no more effective than a single dose in causing infection in ewes.     

Modeling the Growth of Listeria monocytogenes in Soft Blue-White Cheese

The aim of this study was to develop a predictive model simulating growth over time of the pathogenic bacterium Listeria monocytogenes in a soft blue-white cheese. The physicochemical properties in a matrix such as cheese are essential controlling factors influencing the growth of L. monocytogenes. We developed a predictive tertiary model of the bacterial growth of L. monocytogenes as a function of temperature, pH, NaCl, and lactic acid. We measured the variations over time of the physicochemical properties in the cheese. Our predictive model was developed based on broth data produced in previous studies. New growth data sets were produced to independently calibrate and validate the developed model. A characteristic of this tertiary model is that it handles dynamic growth conditions described in time series of temperature, pH, NaCl, and lactic acid. Supplying the model with realistic production and retail conditions showed that the number of L. monocytogenes cells increases 3 to 3.5 log within the shelf life of the cheese.     

Comparative Study of the Effects of Citral on the Growth and Injury of Listeria innocua and Listeria monocytogenes Cells

This study investigates the effect of citral on growth and on the occurrence of sublethal damage in Listeria innocua Serovar 6a (CECT 910) and Listeria monocytogenes Serovar 4b (CECT 4032) cells that were exposed to citral as a natural antimicrobial agent. Two initial inoculum concentrations were considered in this investigation: 10² and 10⁶ cfu/mL. Citral exhibited antilisterial activity against L. innocua and L. monocytogenes, and the observed effects were dependent on the concentration of citral present in the culture medium (0, 0.150 and 0.250 μL/mL) (p ≤ 0.05). L. innocua had a shorter lag phase than L. monocytogenes, and the two species had nearly identical maximum specific growth rates. These results indicate that L. innocua could be used as surrogate for L. monocytogenes when testing the effects of this antimicrobial. Significant differences in the lag phase and growth rate were observed between the small and large inoculum concentration (p ≤ 0.05). Citral-treated L. innocua and L. monocytogenes that were recovered on selective medium (i.e., TSA-YE-SC) had a shorter lag phase and a higher maximum specific growth rate than cells that were recovered on non-selective medium (i.e., TSA-YE) (p ≤ 0.05). This result suggests that damage occurs at sublethal concentrations of citral.     

Use of PCR-Restriction Fragment Length Polymorphism of inlA for Rapid Screening of Listeria monocytogenes Strains Deficient in the Ability To Invade Caco-2 Cells

A PCR-restriction fragment length polymorphism (RFLP) method was developed in order to screen a large number of strains for impaired adhesion to epithelial cells due to expression of truncated InlA. inlA polymorphism was analyzed by PCR-RFLP in order to correlate inlA PCR-RFLP profiles and production of truncated InlA. Thirty-seven Listeria monocytogenes strains isolated from various sources, including five noninvasive and two invasive reference strains, were screened. Two endonucleases (AluI and Tsp509I) were used, and they generated five composite profiles. Thirteen L. monocytogenes isolates were characterized by two specific PCR-RFLP profiles similar to PCR-RFLP profiles of noninvasive reference strains previously described as strains that produce truncated InlA. Ten of the 13 isolates showed low abilities to invade human epithelial Caco-2 cells. However, 4 of the 13 isolates were able to invade Caco-2 cells like reference strains containing complete InlA. Sequencing of inlA and Western blot analysis confirmed that truncated InlA was expressed in the 10 L. monocytogenes strains which were isolated from food. This PCR-RFLP method allowed us to identify 10 new strains expressing a truncated internalin. Based on the results obtained in this study, the PCR-RFLP method seems to be an interesting method for rapidly screening L. monocytogenes strains deficient in the ability to invade Caco-2 cells when a sizeable number of strains are studied.     

Amplified fragment length polymorphism analysis of Listeria monocytogenes by Experion™ automated microfluidic electrophoresis

Fifty Listeria monocytogenes strains were genotyped by sAFLP and PCR products were separated by agarose gel and automated chip-based microfluidic electrophoresis. A high congruency of results was observed comparing the two techniques, although for some cultures a better separation of sAFLP fragments was achieved with microfluidic system, which proved to be a highly reliable and reproducible tool to improve the molecular typing of L. monocytogenes, requiring lower volumes of samples and reducing significantly analysis time.     

Listeria monocytogenes Triggers the Cell Surface Expression of Gp96 Protein and Interacts with Its N Terminus to Support Cellular Infection

Listeria monocytogenes is an intracellular food-borne pathogen causing listeriosis in humans. This bacterium deploys an arsenal of virulence factors that act in concert to promote cellular infection. Bacterial surface proteins are of primary importance in the process of host cell invasion. They interact with host cellular receptors, inducing/modulating specific cellular responses. We previously identified Vip, a Listeria surface protein covalently attached to the bacterial cell wall acting as a key virulence factor. We have shown that Vip interacts with Gp96 localized at the surface of host cells during invasion and that this interaction is critical for a successful infection in vivo. To better understand the importance of Vip-Gp96 interaction during infection, we aimed to characterize this interaction at the molecular level. Here we demonstrate that, during infection, L. monocytogenes triggers the cellular redistribution of Gp96, inducing its exposure at the cell surface. Upon infection, Gp96 N-terminal domain is exposed to the extracellular milieu in L2071 fibroblasts and interacts with Vip expressed by Listeria. We identified Gp96 (Asp¹–Leu¹⁷⁰) as sufficient to interact with Vip; however, we also showed that the region Tyr¹⁷⁹–Leu³⁹⁰ of Gp96 is important for the interaction. Our findings unravel the Listeria-induced surface expression of Gp96 and the topology of its insertion on the plasma membrane and improve our knowledge on the Vip-Gp96 interaction during Listeria infection.     

Suppression of major histocompatibility complex class I and class II gene expression in Listeria monocytogenes-infected murine macrophages

Macrophage cells play a central role during infection with Listeria monocytogenes by both providing a major habitat for bacterial multiplication and presenting bacterial antigens to the immune system. In this study, we investigated the influence of L. monocytogenes infection on the expression of MHC class I and class II genes in two murine macrophage cell lines. Steady-state levels of I-Aβ chain mRNA were decreased in both resting J774A.1 and P388D₁ macrophages infected with L. monocytogenes whereas reduction of H-2K mRNA was only observed in P388D₁ cells. In addition, L. monocytogenes suppressed induction of MHC class I and class II mRNAs in response to γ-interferon as well as the maintenance of the induced state in activated P388D₁ macrophages. Exposure to the non-pathogenic species L. innocua or a deletion mutant of L. monocytogenes, which lacks the lecithinase operon, did not cause a reduction in H-2K and I-Aβ mRNA levels nor suppress expression of Ia antigens. Inhibition of MHC gene expression may represent an important part of the cross-talk between L. monocytogenes and the macrophage that probably influences the efficiency of a T cell-mediated immune response and thus the outcome of a listerial infection.