Int J Food Microbiol, 1996 Apr, 29(2-3), 403 - 10 Effect of monolaurin and lactic acid on Listeria monocytogenes attached to catfish fillets; Verhaegh EG et al.; The purpose of this study was to determine the effects of monolaurin and lactic acid, singly or combined, on Listeria monocytogenes attached to catfish fillets . Skinless catfish fillets were inoculated with L . monocytogenes and dip treated in monolaurin and/or lactic acid solution for various time periods . Results showed that monolaurin up to 400 micrograms/ml had no influence on counts . Conversely, lactic acid-treated fillets had reduced counts compared to controls . Dipping in 0.85, 1.70, or 2.55% lactic acid for 30 min reduced counts by 0.9, 1.4, or 1.3 logs, respectively . Extending the dipping time to 60 min resulted in little additional decrease in counts . Combining monolaurin with lactic acid yielded results similar to lactic acid alone . Hence, population reduction ability resides with lactic acid and not monolaurin. Int J Food Microbiol, 1996 Apr, 29(2-3), 201 - 11 The particular behaviour of Listeria monocytogenes under sub-optimal conditions; Bajard S et al.; Listeria monocytogenes is a ubiquitous pathogenic microorganism which has been described as growing at temperatures of interest to food production and especially at low temperatures (-2 degrees to 8 degrees C) in storage process . However, the general relationship between the maximum specific growth rate, mumax and temperature has not often been studied for L . monocytogenes in the whole temperature range from minimal to maximal growth temperature . A global analysis of this relationship for temperatures between -2 degrees C and 42 degrees C was therefore done . The global shape of this relationship was that usually observed for microorganisms, especially in the neighbourhood of the optimal temperature, Topt . But a more detailed study showed the existence of a so-called "change temperature", occurring between 10 degrees and 15 degrees C, below which L . monocytogenes grows faster than one would expect . This implies that the minimal growth temperature of both studied strains of L . monocytogenes is lower than expected. Immunol Lett, 1996 Apr, 50(1-2), 81 - 5 Activation of natural killer cells by heat-killed Listeria monocytogenes requires additional signals from lymphoid cells; Daugelat S et al.; Regulatory and protective functions have been attributed to murine natural killer (NK) cells in a number of infectious diseases including listeriosis . We have developed an in vitro model to study parameters underlying the activation of naive NK cells using heat-killed Listeria monocytogenes (HKL) as stimulator . Independent from expression of the cell surface marker NK1.1, NK cells lysed YAC-1 cells after in vitro stimulation with HKL or HKL + Interleukin (IL)-2, but not medium or IL-2 alone . In contrast, NK cells from severely immunocompromised SCID or RAG-1-/-mutant mice failed to respond to HKL alone, but required exogenous IL-2 . Using single-gene-disruption mutant mice, we show that NK-cell activation can be supported by either T-cell receptor (TCR) alpha beta cells, TCR- gamma delta cells . MHC class I or MHC class II gene products . We conclude from these data that recognition of listerial components alone is insufficient for activation of naive NK cells, and that additional costimulatory signals are necessary . These can be provided by various lymphoid cells and appear to be cytokines. Microb Pathog, 1996 Apr, 20(4), 247 - 53 Comparison of the infectivity of isolates of Listeria monocytogenes following intragastric and intravenous inoculation in mice; Barbour AH et al.; The infectivity of 19 haemolytic isolates of Listeria monocytogenes from different sources (clinical and environmental) and representative isolates from Listeria ivanovii and Listeria innocua was compared following intragastric (i.g.) and intravenous (i.v.) inoculation in immunocompetent male BALB/c mice . There was marked variation in the infectivity of the different isolates by either route but when isolates were ranked in descending order by spleen count, following i.g . administration, the strains fell into four groups . Infectivity of some isolates also differed when i.v . inoculation was compared with i.g . administration, so that assessment of virulence by spleen counts only following i.v . inoculation might fail to detect isolates of poor infectivity by the i.g . route . These results suggest that intragastric inoculation of normal immunocompetent mice is a useful model for detecting strains of L . monocytogenes that are poorly invasive via the gut even though they are relatively virulent by intravenous inoculation. Vet Microbiol, 1996 Apr, 49(3-4), 169 - 79 Studies on the cell tropism of Listeria monocytogenes in ovine fetal brain cell cultures; Peters M et al.; The uptake of Listeria monocytogenes by cells in primary dissociated brain cell cultures prepared from ovine fetuses at approximately 50 to 60 days of gestation was studied using a sequential double immunofluorescence technique with antibodies against cell type-specific markers and the bacterial pathogen . Cell cultures were inoculated with bacteria at day 4, 8, and 15 in vitro . Listeria monocytogenes was predominately internalized by CD68-positive macrophages, followed by astrocytes, fibronectin-expressing cells, and neurons . An uptake of the bacterium by galactocerebroside (GC)-positive oligodendrocytes, which were first detected at day 15 in vitro, did not occur . Although a tropism for neurons was not observed, the susceptibility of neurons for infection with Listeria monocytogenes is in accordance with the supposed intraaxonal migration of the bacterium in the pathogenesis of focal brain stem encephalitis . The pattern of the infection rates of ovine brain cell types was similar to that shown in murine fetal brain cell cultures, indicating that there is no species-specific brain cell tropism of the bacterium. J Chemother, 1996 Apr, 8(2), 107 - 12 RP 59500, a streptogramin derivative, is effective in murine listeriosis; Nichterlein T et al.; RP 59500 (Synercid) a streptogramin derivative, is a mixture of two water-soluble substances, RP 57669 (quinupristin), and RP 54476 (dalfopristin) . It was tested in vitro and in vivo against Listeria strains . All strains were sensitive in vitro . The MICs of 60 strains of Listeria monocytogenes, L . seeligeri, L . ivanovii, and L . innocua were between 0.156 and 0.625 mg/l . Strains of L . grayi were inhibited by 1.25 mg/l of RP 59500 . In contrast to its bactericidal effect against other gram-positive bacteria, RP 59500 was bacteriostatic against L . monocytogenes in all concentrations tested (up to 16 x MIC) . However, it exerted a pronounced postantibiotic effect . RP 59500 was ineffective against intracellular L . monocytogenes multiplying in L929 mouse fibroblast cells . When it was included in the supernatant of the cells in nontoxic concentrations of below 12.5mg/l alone or in combination with gentamicin (50mg/l) it was not able to inhibit the growth of the listeriae in the cells . However, when tested in peritoneal exudate cells, it was bacteriostatic against L . monocytogenes . In spite of its relatively poor effects on listeriae in vitro, RP 59500 was as active as erythromycin in a mouse model of listeriosis . When mice iv infected with L . monocytogenes were treated ip with 2mg/animal every 12 hours with either erythromycin or RP 59500, both substances prevented growth of the bacteria in the organs, but were unable to eradicate the listeriae . We conclude that RP 59500 is a candidate substance for the treatment of human listeriosis which might be used when therapy with ampicillin is not feasible. Proc Soc Exp Biol Med, 1996 Apr, 211(4), 346 - 52 Effects of amiodarone-induced phospholipidosis on pulmonary host defense functions in rats; Reasor MJ et al.; The effect of the induction of pulmonary phospholipidosis by amiodarone on selected pulmonary host defense functions was studied in male Fischer-344 rats . One week of daily amiodarone treatment resulted in a 4.5-fold increase in total phospholipid in alveolar macrophages recovered from the lungs by bronchoalveolar lavage . The presence of the phospholipidosis had no effect on the phagocytosis of heat-killed yeast cells, the induction of luminol-dependent chemiluminescence, or the spontaneous release of interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-alpha), or spontaneous and LPS-stimulated release of IL-1 by alveolar macrophages in vitro . In contrast, the LPS-stimulated release of IL-6 and TNF-alpha by phospholipidotic alveolar macrophages was enhanced compared with control cells . The pulmonary clearance of Listeria monocytogenes following intratracheal administration of the bacteria was not affected by the phospholipidotic condition . It appears that, in the context of the functions studied, the induction of pulmonary phospholipidosis by amiodarone does not impair pulmonary host defense processes in rats, and may actually be associated with the augmentation of some activities. J Med Microbiol, 1996 Apr, 44(4), 295 - 302 Early pathogenesis of Listeria monocytogenes infection in the mouse spleen; Conlan JW; Histological observations suggested that in the spleen, blood-borne Listeria monocytogenes bacteria were preferentially ingested by two morphologically distinct mononuclear phagocyte populations present in the marginal zone of the white pulp . The morphologies of these phagocytes corresponded to those of marginal zone macrophages or marginal zone dendritic cells . Moreover, during the first day of infection, the same phagocytes containing listeria apparently translocated from the marginal zone into the white pulp where they established secondary infectious foci . This event was associated with a large influx of neutrophil polymorphonuclear leucocytes (PMNLs) into infected white pulp, and with the disappearance of lymphocytes from this compartment . White pulp lymphocytopenia also occurred in the spleens of listeria-infected mice selectively depleted of neutrophil PMNLs, indicating that these phagocytes were not responsible for displacing or destroying lymphocytes . The implications of these findings for explaining the virulence and immunogenicity of L . monocytogenes are discussed. Infect Immun, 1996 Apr, 64(4), 1299 - 308 Effect of cell polarization and differentiation on entry of Listeria monocytogenes into the enterocyte-like Caco-2 cell line; Gaillard JL et al.; The entry of Listeria monocytogenes into the enterocyte-like Caco-2 cell line was studied as a function of cell polarization and differentiation . L . monocytogenes entered through the entire surface of nonpolarized cells and, predominantly, through the basolateral surface of polarized cells based on the following observations: (i) sites of L . monocytogenes invasion paralleled the distribution of the transferrin receptor, a well-known basolateral marker of polarization; (ii) numbers of internalized bacteria decreased dramatically when Caco-2 monolayers cultured beyond confluency were used (about 0.1% of the inoculated bacteria versus 1 to 2% with nonconfluent monolayers); and (iii) L . monocytogenes entry into postconfluent monolayers was greatly enhanced by treating cells with Ca(2)+ -free medium, a procedure that disrupts intercellular junctions and thus exposes the basolateral surface to bacteria . Ethylene glycol-bis (beta-aminoethyl ether)-N, N, N',N' -tetraacetic acid (EGTA) had contradictory effects on L . monocytogenes entry as this reagent opened intercellular junctions but inhibited binding and internalization of bacteria . Finally, the role of the inlAB locus in L . monocytogenes entry was confirmed because and inlAB mutant was 50- to 100-fold less invasive than the parental strain regardless of the monolayer's age . However, the inlAB mutant was still able to enter cells and to induce intracellular actin polymerization . Entry of inlAB bacteria into Caco-2 cells was not inhibited by EGTA. Infect Immun, 1996 Apr, 64(4), 1252 - 8 Endogenous interleukin-4, but not interleukin-10, is involved in suppression of host resistance against Listeria monocytogenes infection in interferon-depleted mice; Nakane A et al.; The production and roles of endogenous interleukin-4 (IL-4) and IL-10 in a sublethal infection with Listeria monocytogenes were studies in normal mice and anti-gamma interferon (IFN-gamma) monoclonal antibody (MAb)-pretreated mice . In normal mice, the expression of mRNAs for IL-4 and IL-10, which was amplified by reverse transcription-PCR, was induced in the spleens and livers either early or late in infection, although the serum IL-4 and IL-10 were not detectable by enzyme-linked immunosorbent assays . In vivo administration of anti-IL-4 MAb showed no effect on antilisterial resistance, whereas anti-IL-10 MAb partially diminished the defense . In anti-IFN-gamma MAb-pretreated mice, a delay in the bacterial elimination from the spleens and livers was observed and high titers of serum IL-4 and IL-10 were induced late in infection . Production of endogenous IL-4 and IL-10 was suppressed in both CD4+ cell-and CD8+ cell depleted mice . The suppression of antilisterial resistance in anti-IFN-gamma MAb-pretreated mice was canceled when anti-IL-4 MAb was injected late in infection, whereas anti-IL-10 MAb showed no effect . These results suggest that the host immune responses were polarized into the T-helper 2 phenotype in anti-IFN-gamma MAb-pretreated mice and that inhibition of host resistance against L . monocytogenes by depletion of IFN-gamma might be attributable to IL-4 produced by T cells polarized into the T-helper 2 phenotype as well as the inhibition of the IFN-gamma effects. Infect Immun, 1996 Apr, 64(4), 1197 - 202 Acquired resistance against a secondary infection with Listeria monocytogenes in mice is not dependent on reactive nitrogen intermediates; Samsom JN et al.; During an infection, inflammatory mediators can induce the production of nitric oxide, a reactive nitrogen intermediate (RNI) which plays a role in antimicrobial activity against a wide variety of pathogens . In vitro experiments have shown that release of RNI by macrophages is mediated by tumor necrosis factor alpha (TNF) . Since TNF is essential for acquired resistance during a secondary Listeria monocytogenes infection in mice, the aim of the present study was to determine whether RNI are also involved in the course of such an infection . Mice which had recovered from a sublethal primary infection with 0.1 50% lethal dose of (LD50) L . monocytogenes were infected intravenously with 10LD50 of L . monocytogenes . During a primary infection, the number of bacteria in the liver and spleen, as well as the concentration of RNI in plasma, increased . During a secondary infection, the number of bacteria in the liver and spleen decreased whereas no significant increase in the concentration of RNI in plasma was observed . Neutralization of endogenously produced TNF and gamma interferon by subcutaneous injection of alginate-encapsulated monoclonal antibody-forming cells during a secondary infection resulted in an increase in the number of bacteria in the liver and spleen an increase in the concentration of RNI in plasma . When the production of RNI was inhibited by treatment of mice with competitive NO-synthase inhibitor N omega-nitro-L-arginine methyl ester hydrochloride (L-Name) and an iota-arginine-deficient diet during a secondary infection, the proliferation of L . monocytogenes in the liver and spleen was not affected whereas the concentration of RNI in plasma of these mice was significantly reduced . Our findings that inhibition of RNI formation during a secondary infection does not affect the proliferation of L . monocytogenes in the liver and spleen and that enhanced elimination of bacteria from these organs is not accompanied by an increase in the concentration of RNI in plasma led to the conclusion that resistance against a secondary infection with L . monocytogenes is not dependent on RNI. J Clin Invest, 1996 Apr 1, 97(7), 1624 - 9 Intramolecular inhibition of human defensin HNP-1 by its propiece; Valore EV et al.; We examined mechanisms that protect host defense cells from their cytotoxic effector molecules . Human neutrophil peptides (HNP) 1-3 are microbicidal and cytotoxic defensins, initially synthesized as 94-amino acid preproHNP(1-94), cotranslationally proteolyzed to proHNP(20-94), then converted by removal of the anionic propiece to mature HNP(65-94)(HNP-1 and -3) and HNP(66-94) (HNP-2) . We hypothesized that during synthesis and subcellular sorting the anionic propiece inhibits the cytotoxicity of the cationic defensin . We expressed preproHNP-1 cDNA in recombinant baculovirus-infected insect cells that secreted the normally transient proHNP-1(20-94) into the medium . Cyanogen bromide cleaved proHNP-1(20-94) at the fortuitously located Met64 to yield mature recombinant HNP-1(65-94) and unlinked propiece . Recombinant and native HNP-1 purified from PMN were identical as judged by mass spectrometry, retention time in reverse-phase high performance liquid chromatography, migration on acid-urea polyacrylamide gels, and reaction with a conformation-specific antibody . Recombinant and native HNP-1 had comparable microbicidal activity towards Listeria monocytogenes and were similarly potent in permeabilizing K562 leukemia cells, but proHNP-1(20-94) was virtually inactive in both assays . Addition of unlinked propiece (proHNP-1(20-64) with Met64-->homoserine) inhibited the bactericidal and cell-permeabilizing activity of mature HNP-1 in a dose-dependent manner . Linked, and to a lesser extent unlinked, propiece interfered with the binding of HNP-1 to target cells . The propiece thus acts as an efficient intramolecular inhibitor of defensin HNP-1 cytotoxicity. J Biol Chem, 1996 Mar 29, 271(13), 7249 - 52 Delivery of macromolecules into cytosol using liposomes containing hemolysin from Listeria monocytogenes; Lee KD et al.; The cytosolic space of cells is an important but relatively inaccessible target for the delivery of therapeutic macromolecules . Here we describe the efficient delivery of macromolecules into the cytosolic space of macrophages from liposomes that contain listeriolysin O (LLO), the hemolytic protein of Listeria monocytogenes that normally mediates bacterial passage from phagosomes into cytosol . LLO was purified and encapsulated inside pH-sensitive liposomes, along with other molecules to be delivered . When internalized by bone marrow-derived macrophages, these liposomes rapidly released encapsulated fluorescent dye, first into endosomes and then into the cytosol, without measurably harming the cells . Furthermore, these liposomes efficiently delivered encapsulated ovalbumin to the cytosolic pathway of antigen processing and presentation, as measured by the major histocompatibility complex (MHC) class I-restricted presentation of peptides derived from ovalbumin . Delivery was significantly better than that obtained by other currently available liposome formulations . LLO-containing liposomes should therefore provide an efficient vehicle for delivery of antigens or therapeutic molecules in vivo. J Immunol, 1996 Mar 15, 156(6), 2214 - 20 Response of a gamma delta+ T cell receptor invariant subset during bacterial infection; Roark CE et al.; Murine gamma delta T cells can be divided into subsets based on the TCR gamma-chains they express . Most of these subsets have variable TCR junctions, but two, both associated with epithelia, express invariant TCRs . The absence of receptor variability in these cells implies uniformity of their ligands . This was previously taken as evidence to suggest that gamma delta T cells recognize host-derived, stress-induced ligands.We now demonstrate, for the first time, the response of a gamma delta TCR invariant subset during bacterial infection, a potential cause of stress . After infection with Listeria monocytogenes, absolute numbers of all T cells in the liver, including alpha beta and gamma delta T cell subsets, increased markedly . However, responses of a gamma delta T cell subset varied . We noted a decrease in the relative frequency of V delta 6.3+ cells, which are, for the most part, included in the V gamma 1+ subset . In contrast, cells bearing the invariant V gamma 6/V delta 1 TCR increased substantially in proportion to other gamma delta T cells, as determined by PCR analysis of liver T cell RNA and by comparing liver gamma delta T cell hybridomas made from normal mice to those from mice infected with Listeria . V gamma 6/V delta 1+ cells have been previously reported as a TCR invariant intraepithelial subset in the female reproductive tract and tongue . We show here that V gamma 6/V delta 1+ cells reactive in Listeria-infected liver are polyclonally derived, but still bear TCR chains with invariant junctional sequences, identical with those of the female reproductive tract . Although the Ag that stimulates these cells is unknown, our results indicate that only diverse, but also invariant, gamma delta T cell subsets can become involved in the host response to a bacterial infection. Appl Environ Microbiol, 1996 Mar, 62(3), 1116 - 9 Cold stress proteins induced in Listeria monocytogenes in response to temperature downshock and growth at low temperatures; Bayles DO et al.; Listeria monocytogenes is a food-borne pathogen with the ability to grow at refrigerator temperatures . Twelve cold shock proteins (Csps) with apparent M(r)s of 48,600, 41,000, 21,800, 21,100, 19,700, 19,200, 18,800, 18,800, 17,200, 15,500, 14,500, and 14,400 were induced by cold shocking L . monocytogenes 10403S from 37 to 5 degrees C, as revealed by labeling with L-{35S}methionine followed by two-dimensional gel electrophoresis . Strain SLCC53 showed a similar response . Cold acclimation proteins were observed in cultures of strain 10403S growing at 5 degrees C, and four of these proteins, with apparent M(r)s 48,000, 21,100, 19,700, and 18,800, were also Csps . Two cold-sensitive transposon-induced mutants were labeled less efficiently than the parent strain, but the Csp response of the mutant examined was very similar to that of the parent strain. Appl Environ Microbiol, 1996 Mar, 62(3), 1058 - 64 Changes in cell morphology of Listeria monocytogenes and Shewanella putrefaciens resulting from the action of protamine; Johansen C et al.; Protamine, which is an antibacterial basic peptide, was shown to alter the cell morphology of Listeria monocytogenes and Shewanella putrefaciens . Atomic force microscopy revealed that protamine smoothed the surface of cells, formed holes in the cell envelope, and caused fusion of S . putrefaciens cells . Immunoelectron microscopy of protamine-treated cells of both L . monocytogenes and S . putrefaciens showed great damage to the cell wall and condensation of the cytoplasm . Respiration of the cells was decreased due to treatment with sublethal concentrations of protamine, probably due to leakage or loss of cell envelope potential . It was concluded that protamine disrupted the outer surface structure and condensed the cytoplasm of sensitive cells and, in sublethal concentrations, altered membrane structures, thereby eliminating respiration. AJNR Am J Neuroradiol, 1996 Mar, 17(3), 593 - 6 MR findings in listerial rhombencephalitis; Alper G et al.; We describe a case of listerial rhombencephalitis in a previously healthy 40-year-old man . The diagnosis was based on the clinical findings, results of cerebrospinal fluid analysis, blood culture, and MR imaging findings . The treatment was started before culture results were available, and the patient had a full clinical recovery. FEMS Immunol Med Microbiol, 1996 Mar, 13(3), 211 - 3 Live tularemia vaccine confers protection against lethal Legionella and Listeria infections in experimental animals; Belyi YF et al.; The efficacy of a live Francisella tularensis vaccine strain to cause nonspecific immunity toward experimental legionellosis and listeriosis was studied . Immunisation with tularemia vaccine protected over 80% and 17% of experimental animals against subsequent lethal challenge with Legionella pneumophila and Listeria monocytogenes, respectively . The protection was maximal during the first month following immunisation and declined thereafter . In order to delineate the immunostimulatory moieties of the Francisella microbe, several cell wall proteins have been purified and characterized . However, isolated cell wall components failed to induce protection. J Appl Bacteriol, 1996 Mar, 80(3), 303 - 10 Quantitative structure activity relationship for the effect of benzoic acids, cinnamic acids and benzaldehydes on Listeria monocytogenes; Ramos-Nino ME et al.; The inhibition of a cocktail of 18 strains of Listeria monocytogenes by 24 mono-, di- and tri-substituted benzoic and cinnamic acids and 16 benzaldehydes was evaluated using the concentration (C) required to give a 50% growth inhibition under anaerobic conditions at 35 degree C and pH 6.2 as a measure of biological activity (BAV) . Using the method of least squares, multiple regression equations were obtained which described the contribution of some physiochemical and other structural properties of the compounds to their biological activity . The equation that best described the activity of benzoic and cinnamic acids was {formula: see text} where K is a lipophilicity parameter determined by RP-HPLC and the effect of ionization is represented by pKa . Benzaldehydes behaved differently, their activity being best described by the equation . {formula: see text} where the activity is controlled by a steric parameter, the van der Waals volume (Vw), and an electronic-steric parameter for ortho substituents . Absence of a lipophilicity parameter indicates that partitioning into the cell membrane might not be required for antimicrobial activity . The models were tested in several food systems which showed that in food with a high protein or lipid content antilisterial activity was much lower than predicted, making the models unacceptable in such circumstances. Int Immunol, 1996 Mar, 8(3), 367 - 78 H2-M3wt-restricted, Listeria monocytogenes-specific CD8 T cells recognize a novel, hydrophobic, protease-resistant, periodate-sensitive antigen; Nataraj C et al.; Mice infected with Listeria monocytogenes (LM) generate H2-M3wt-restricted CD8 effectors which recognize a heat-killed LM-associated antigen (HAA) presented by macrophages . To characterize HAA, we extracted a bioactive component from LM using SDS or NaOH . Extracted HAA aggregated in hydrophilic solvents but dissociated in the presence of SDS into a smaller subunit which migrated in Sephadex G-200 between chymotrypsinogen (25 kDa) and cytochrome c (12.5 kDa) . HAA bioactivity and size was unaffected by proteinase K under conditions which degraded virtually all detectable protein . HAA was also unaffected by other proteases, RNase and DNase, but HAA bioactivity was destroyed by periodate, an agent that degrades carbohydrates . These studies demonstrate that H2-M3wt can present a hydrophobic, non-peptide, microbial antigen, probably glycolipid in origin, to CD8 T cells. Infect Immun, 1996 Mar, 64(3), 1002 - 6 Identification and purification of novel internalin-related proteins in Listeria monocytogenes and Listeria ivanovii; Lingnau A et al.; Monoclonal antibodies were generated against a 30-kDa protein fraction derived from culture supernatants of a Listeria monocytogenes strain complemented with additional copies of the prfA regulator gene . Several of the antibodies reacted specifically with a hitherto unidentified, secreted 30-kDa polypeptide . By immunoblot analysis, the expression of this 30kDa polypeptide was found to be dependent on the presence of the PrfA regulator protein . Microsequencing of peptides derived from the partially purified 30-kDa protein revealed homologies to the InlA and InlB polypeptides of L . monocytogenes, which are required for the internalization of the bacteria into nonphagocytic cell lines . This prompted us to term the 30-kDa polypeptide internalin-related protein (Irp) . Irp-specific monoclonal antibodies cross-reacted with a 24-kDa polypeptide present in culture supernatants of Listeria ivanovii, indicating the existence of an Irp-related protein in this pathogenic Listeria species. J Immunol, 1996 Feb 15, 156(4), 1497 - 503 CTL epitope generation is tightly linked to cellular proteolysis of a Listeria monocytogenes antigen; Sijts AJ et al.; Listeria monocytogenes is a pathogenic intracellular bacterium that secretes proteins into the cytosol of host cells . A major secreted protein, p60, is processed by the host cell into the nonamer peptides p60 217-225 and p60 449-457, which are presented to CTL by H-2Kd MHC class I molecules . Herein, we use two membrane permeable peptide aldehyde protease inhibitors, LLnL and Z-LLF, to inhibit cytosolic proteolysis in L . monocytogenes-infected cells . These inhibitors, which have been shown to inhibit proteasomes, completely abrogate cytosolic p60 degradation . The effect of LLnL and Z-LLF on p60 epitope generation was determined by acid-eluting, HPLC-purifying, and quantifying p60 217-225 and p60 449-457 from infected cells . We show a direct linkage between p60 degradation and epitope generation . However, the two inhibitors have quantitatively different effects on the generation of the two epitopes . Our findings implicate proteasomes in the earliest stages of Ag degradation and suggest that different CTL epitopes can be generated by distinct proteolytic processes. Lett Appl Microbiol, 1996 Feb, 22(2), 153 - 8 Polymerase chain reaction (PCR) detection of Listeria monocytogenes in diluted milk and reversal of PCR inhibition caused by calcium ions; Bickley J et al.; DNA from Listeria monocytogenes was used as the model system from this investigation, with PCR primers based on the listeriolysin O gene . Under standard polymerase chain reaction (PCR) conditions and with no prior treatment, amplification failed in the presence of more than 5% milk . Since inhibition of the PCR occurred at the same milk concentrations with full fat, half fat and fat-free milk, inhibition was not attributed to the fat content of the milk . Calcium ions were, however, identified as a major source of PCR inhibition . The results demonstrated that the inhibitory effects of calcium ions and milk could be partially reversed by increasing the magnesium concentration in the reaction to well above the standard levels normally required for PCR . This work has important implications for the use of the PCR in the direct detection of food pathogens. Lett Appl Microbiol, 1996 Feb, 22(2), 121 - 4 The effect of temperature and growth rate on the susceptibility of Listeria monocytogenes to environmental stress conditions; Patchett RA et al.; The effect of growth temperature and growth rate on the susceptibility to heat and pH stress were investigated in Listeria monocytogenes grown in continuous culture where these two growth variables could be varied independently of each other, and in batch culture . After growth at 30 degrees C or 10 degrees C at constant growth rate, or at 30 degrees C at different growth rates, cells did not differ in their resistance to heat at 55 degrees C . Cells grown at 30 degrees C were more resistant to acid stress at pH 2.5 than cells grown at the same growth rates at 10 degrees C . Cells grown at low growth rate at 30 degrees C gave greater resistance to acid stress than those grown at high growth rate . Growth temperature and growth rate had independent effects on the susceptibility of L . monocytogenes to acid stress conditions . This may have implications for the survival of L . monocytogenes in acidic foods. Liver, 1996 Feb, 16(1), 67 - 9 Liver abscesses due to Listeria monocytogenes; Marino P et al.; Involvement of the liver in cases of Listeria monocytogenes is uncommon . We report a case of hepatitis due to L . monocytogenes, with a pathological finding of multiple abscesses . Blood cultures yielded L . monocytogenes . The patient died a few days after admission. Immunology, 1996 Feb, 87(2), 230 - 5 Human gamma/delta T-cell response to Listeria monocytogenes protein components in vitro; Munk ME et al.; Listeria monocytogenes is a facultative intracellular pathogen that replicates inside mononuclear phagocytes and induces specific cellular immunity . Listeriosis encompasses many clinical syndromes and meningitis is the most frequent clinical manifestation . Human alpha/beta and gamma/delta T cells have been shown to respond to L . monocytogenes antigens and to play an important role in resistance against listerial infection . We investigated the nature of listerial ligands and the influence of the major virulence factor, listeriolysin (hly), on the stimulation of human gamma/delta T cells from healthy individuals . We found that a listerial somatic protein ligand, which is sensitive to proteinase treatment, stimulated gamma/delta T cells in vitro; the majority of Listeria-responsive gamma/delta T cells expressed V gamma 9V delta 2 T-cell receptor chains and human leucocyte antigen-DR molecules; gamma/delta T-cell responses to hly+ and hly- Listeria strains were comparable; L . monocytogenes strains of different virulence stimulated gamma/delta T cells equally . Thus, protein components of L . monocytogenes unrelated to virulence activate human gamma/delta T cells in vitro. J Appl Bacteriol, 1996 Feb, 80(2), 216 - 24 Effect of pre- and post-heat shock temperature on the persistence of thermotolerance and heat shock-induced proteins in Listeria monocytogenes; Jorgensen F et al.; The effect of incubation temperature, before and after a heat shock, on thermotolerance of Listeria monocytogenes at 58 degrees C was investigated . Exposing cells grown at 10 degrees C and 30 degrees C to a heat shock resulted in similar rises in thermotolerance while the increase was significantly higher when cells were grown at 4 degrees C prior to the heat shock . Cells held at 4 degrees C and 10 degrees C after heat shock maintained heat shock-induced thermotolerance for longer than cells held at 30 degrees C . The growth temperature prior to inactivation had negligible effect on the persistence of heat shock-induced thermotolerance . Concurrent with measurements of thermotolerance were measurements of the levels of heat shock-induced proteins . Major proteins showing increased synthesis upon the heat shock had approximate molecular weights of 84, 74, 63, 25 and 19 kDa . There was little correlation between the loss of thermotolerance after the heat shock and the levels of these proteins . Thermotolerance of heat shocked and non-heat shocked cells was described by traditional log-linear kinetics and a model describing a sigmoidal death curve (logistic model) . Employing log-linear kinetics resulted in a poor fit to a major part of the data whereas a good fit was achieved by the use of a logistic model. J Appl Bacteriol, 1996 Feb, 80(2), 174 - 8 Differentiation of Listeria isolates by PCR amplicon profiling and sequence analysis of 16S-23S rRNA internal transcribed spacer loci; Drebot M et al.; The 16S-23S rRNA internal transcribed spacer region (ITS) in genomic DNA from Listeria species was amplified and sequenced so as to find sequence differences that would allow rapid species and strain differentiation . Agarose gel profiles of amplicons generated with primers designed to amplify ITS loci indicated that Listeria DNA can contain at least two distinct ITS regions . The direct sequencing of the smaller of these ITS amplicons (330 bp) was found useful for the rapid and accurate differentiation of various Listeria species . On the other hand analysis of ITS amplicons generated from a total of 27 L . monocytogenes strains indicated that 4/27 of these strains could be distinguished on the basis of their ITS profile (the presence of a unique 350 bp amplicon) . The lack of sequence heterogeneity in the small 333 bp amplicon did not permit rapid strain differentiation. J Exp Med, 1996 Feb 1, 183(2), 359 - 69 The inlAB locus mediates the entry of Listeria monocytogenes into hepatocytes in vivo; Gaillard JL et al.; The intracellular parasite Listeria monocytogenes is able to induce its internalization by cultured mammalian cells that are not normally phagocytic . This process requires the expression of the chromosomal locus inlAB . We studied the virulence of an inlAB mutant and of its parent in murine listeriosis . Irrespective of the route of inoculation, the inlAB mutant was severely attenuated for growth in the liver . The livers of mice inoculated with the inlAB mutant displayed much smaller infectious foci than the parent as early as 24 h after infection . Electron microscopy showed that these foci consisted of a few inflammatory cells, with few bacteria; bacteria were rarely found within hepatocytes . In contrast, foci in livers of mice inoculated with the parent consisted of islets of heavily infected hepatocytes that were infiltrated by numerous neutrophils; bacteria seemed intact within hepatocytes and damaged within neutrophils . A direct role of inlAB for the entry of L . monocytogenes into hepatocytes was confirmed in a cell infection system using the murine embryonic hepatocyte cell line TIB73 . The inlAB mutant was approximately 20-fold less invasive in trans . The "invasion locus" inlAB contributes to protect L . monocytogenes from the host's innate defense mechanisms by promoting its entry into hepatocytes. Eur J Immunol, 1996 Feb, 26(2), 356 - 64 Administration of interleukin-10 abolishes innate resistance to Listeria monocytogenes; Kelly JP et al.; We have used severe-combined immunodeficient (SCID) mice to examine the immunoregulatory effects of interleukin (IL)-10 on innate resistance to infection with Listeria monocytogenes . Addition of heat killed Listeria to spleen cells from naive SCID mice resulted in secretion of interferon (IFN)-gamma from natural killer cells in vitro . This response was enhanced up to 15-fold in the presence of exogenous IL-2, but was completely ablated by addition of IL-10 with IC50 of less than 0.5 U/ml . Infection of SCID mice with viable Listeria in vivo resulted in a prolonged course of infection eventually causing death by 12-14 days, whereas daily administration of IL-10 increased bacterial replication in the liver and spleen by up to 1000-fold resulting in death by day 4 post-infection . The immunosuppressive actions of IL-10 in vivo were also observed in immunocompetent BALB/c mice, whereas doses as low as 100 U/day converted a sublethal infection to 100% mortality . To study the events controlling expression of endogenous IL-10, peritoneal macrophage monolayers were challenged with Listeria after preincubation with a panel of recombinant cytokines . IFN-gamma primed macrophages for enhanced tumor necrosis factor (TNF) secretion, but inhibited IL-10 production, whereas granulocyte/macrophage colony-stimulating factor (CSF), macrophage CSF and also IL-4 enhanced macrophage IL-10 responses after ingestion of Listeria in vitro . Finally, monoclonal antibody neutralization of IFN-gamma during infection of SCID mice with Listeria inhibited TNF-alpha mRNA, but augmented expression of IL-10 mRNA in infected tissues . These results demonstrate that exogenous Il-10 is a potent immunosuppressive cytokine in the context of infection with an intracellular bacterium and that expression of endogenous IL-10 versus TNF is differentially regulated by the cytokine environment of the macrophage. Appl Environ Microbiol, 1996 Feb, 62(2), 705 - 11 Use of molecular typing methods to trace the dissemination of Listeria monocytogenes in a shrimp processing plant; Destro MT et al.; Molecular typing of bacteria has been widely used in epidemiological studies but not as extensively for tracing the transmission of pathogenic bacteria in food plants . This study was conducted to examine the potential use of two molecular typing methods, random amplified polymorphic DNA (RAPD) analysis and pulsed-field gel electrophoresis (PFGE), to trace Listeria monocytogenes contamination in a shrimp processing plant . Ribotyping and phase typing were also performed on a select number of strains . One hundred fifteen strains of L . monocytogenes collected in different areas of a shrimp processing plant were first serotyped and then subtyped by molecular typing . RAPD and PFGE showed great promise for typing L . monocytogenes isolates since distinguishable and reproducible DNA polymorphisms were obtained . When the composite profile from both (RAPD and PFGE) methods was generated, there was an increase in the discriminatory power to discern differences between strains of L . monocytogenes . The results indicated that environmental strains all fell into composite profile groupings unique to the environment, while strains from both water and utensils shared another composite profile group . L . monocytogenes fresh shrimp isolates belonging to one profile group were found in different areas of the processing line . This same profile group was also present in food handlers from the processing and packaging areas of the plant. Appl Environ Microbiol, 1996 Feb, 62(2), 323 - 7 Physiochemical characterization of the nisin-membrane interaction with liposomes derived from Listeria monocytogenes; Winkowski K et al.; Mechanistic information about the bacteriocin nisin was obtained by examining the efflux of 5(6)-carboxy-fluorescein from Listeria monocytogenes-derived liposomes . The initial leakage rate (percentage of efflux per minute) of the entrapped dye was dependent on both nisin and lipid concentrations . At all nisin concentrations tested, 5(6)-carboxyfluorescein efflux plateaued before all of the 5(6)-carboxyfluorescein was released (suggesting that pore formation was transient), but efflux resumed when more nisin was added . Isotherms for the binding of nisin to liposomes constructed on the basis of the Langmuir isotherm gave an apparent binding constant of 6.2 x 10(5)M(-1) at pH 6.0 . The critical number of nisin molecules required to induce efflux from liposomes at pH 6.0 was approximately 7,000 molecules per liposome . The pH affected the 5(6)-carboxyfluorescein leakage rates, with higher pH values resulting in higher leakage rates . The increased leakage rate observed at higher pH values was not due to an increase in the binding affinity of the nisin molecules towards the liposomal membrane . Rather, the critical number of nisin molecules required to induce activity was decreased (approximately 1,000 nisin molecules per liposome at pH 7.0) . These data are consistent with a poration mechanism in which the ionization state of histidine residues in nisin plays an important role in membrane permeabilization. Am J Pathol, 1996 Feb, 148(2), 657 - 66 Sequestration of inhaled particulate antigens by lung phagocytes . A mechanism for the effective inhibition of pulmonary cell-mediated immunity; MacLean JA et al.; Dendritic cells (DCs) have emerged as the dominant antigen-presenting cells (APCs) of the lung, playing a vital role in the induction of cell-mediated immunity to inhaled antigens . We have previously demonstrated that an airway challenge with the soluble antigen hen egg lysozyme yields rapid acquisition of specific antigen-presenting cell activity by purified pulmonary DCs and a cell-mediated immune response in the lung upon secondary challenge . To examine how a particulate antigen leads to a cell-mediated response in vivo, graded concentrations of heat-killed Listeria (HKL) were injected intratracheally into Lewis rats . The bacteria were rapidly ingested by lung macrophages and polymorphonuclear leukocytes . The ability of purified pulmonary DCs pulsed in vivo by an airway challenge with HKL to subsequently stimulate HKL-specific responses ex vivo showed a threshold response, requiring a dose in excess of 10(9) organisms/rat . By contrast, all dosages of HKL yielded specific sensitization of lymphocytes in the draining bilar nodes . Pulmonary DCs purified from rats after a secondary in vivo airway challenge with HKL at day 14 were ineffective antigen-presenting cells except at high dosages of antigen . The generation of cell-mediated pulmonary inflammation paralleled the antigen-presenting cell activity of pulmonary DCs and was observed only at high antigen dosages . Hen egg lysozyme immobilized onto polystyrene beads and injected intratracheally yielded comparable results to those observed with HKL . We suggest that a pulmonary cellular immune response is generated to an inhaled particulate antigen when the protective phagocytic capacities of the lung are exceeded and antigen is able to interact directly with interstitial DCs . The diversion of particulate antigens by pulmonary phagocytes may help to limit undesirable pulmonary inflammation while allowing the generation of antigen-specific immune lymphocytes in vivo. J Immunol, 1996 Feb 1, 156(3), 1196 - 206 Regulation of murine macrophage IL-12 production . Activation of macrophages in vivo, restimulation in vitro, and modulation by other cytokines; Skeen MJ et al.; IL-12 is important in the host response to a variety of pathogens . It plays an adjuvant-like role in an initial immune response as well as a therapeutic role in established infections . Despite its well documented importance, comparatively little is known about the regulation of IL-12 production . In this study, we examined IL-12 production by cultured murine peritoneal macrophages from two perspectives: 1) macrophage activation in vivo, and 2) stimulation of IL-12 secretion in vitro . Macrophages were maximally activated within 48 h in vivo during infection with Listeria . Interestingly, although avirulent or heat-killed Listeria induced only minimal production of IL-12 by macrophages, the immunogenic combination of heat-killed bacteria and rIL-12 was highly stimulatory for IL-12 production . LPS and peritoneal inflammatory agents were also stimulatory, but latex beads were ineffective, indicating that microbial components were essential and phagocytosis alone was insufficient . Restimulation in vitro revealed similar patterns, in that infection and LPS were stimulatory but latex beads were not . A systematic survey of potential stimulatory agents showed that microbial heat shock proteins, crude bacterial extracts, bacterial superantigens, a yeast extract, and dsRNA induced IL-12 in vitro . Other cytokines also influenced IL-12 induction . IFN-gamma, which is up-regulated during infection, acted in synergy with other stimuli, suggesting an amplification loop for IL-12 production, whereas IL-4, IL-10, IL-13, and TGF-beta were inhibitory . The existence of a broad range of stimuli from a wide variety of pathogenic organisms underscores the fundamental importance of IL-12 in host defense. J Immunol, 1996 Feb 1, 156(3), 1143 - 50 Macrophages have cell surface IL-10 that regulates macrophage bactericidal activity; Fleming SD et al.; IL-10, which is secreted by multiple cell types, has regulatory effects on macrophages, including decreasing their ability to kill some microorganisms . The experiments presented here test the hypothesis that endogenous IL-10 inhibits the ability of macrophages to kill the facultative intracellular bacterium, Listeria monocytogenes . We show that the nonbactericidal macrophage hybrid, H36.12j (12j), can kill Listeria after incubation for 3 days with anti-IL-10 mAb . IL-10 was not detected in 12j macrophage supernatants by ELISA . However, flow cytometric analysis revealed high levels of IL-10 on the 12j cell surface . This indicates that macrophages that fail to secrete IL-10 may express IL-10 on the cell surface, and this IL-10 appears to suppress listericidal activity . Surface IL-10 is not unique to the 12j macrophage hybrid and may correlate with the absence of bactericidal activity in other macrophages . For instance, nonlistericidal resident and thioglycolate-elicited peritoneal exudate cells have 24 to 72% IL-10-positive macrophages . In contrast listericidal proteose peptone-elicited peritoneal exudate cells contained < 5% IL-10-positive macrophages . Whether this IL-10 is an integral membrane protein or is bound to IL-10 receptors is not yet known . However, the IL-10 does not elute with acid as other passively bound molecules do, nor does exogenous IL-10 bind to macrophages . In either case, since anti-IL-10 induces nonbactericidal macrophages to become bactericidal, the surface IL-10 is biologically active, and it appears to regulate macrophage bactericidal activity. Infect Immun, 1996 Feb, 64(2), 674 - 6 Listeriolysin is a potent inducer of the phosphatidylinositol response and lipid mediator generation in human endothelial cells; Sibelius U et al.; The impact of Listeria monocytogenes listeriolysin O (LLO) secretion on phosphoinositide metabolism and mediator (platelet-activating factor and prostaglandin I2) generation was investigated in human umbilical vein endothelial cells . Wild-type L . monocytogenes, purified LLO, and an L . innocua strain engineered to secrete LLO all elicited a strong response, whereas mutant strains defective in LLO production were ineffective . Thus, human umbilical vein endothelial cell stimulation by listeriae is linked to production of LLO. Infect Immun, 1996 Feb, 64(2), 569 - 75 Influence of beta 2-microglobulin expression on gamma interferon secretion and target cell lysis by intraepithelial lymphocytes during intestinal Listeria monocytogenes infection; Emoto M et al.; Numerous microbial pathogens, including Listeria monocytogenes, enter the host through the intestine . Although relatively little is known about the biological functions of intestinal intraepithelial lymphocytes (i-IEL), they are generally considered a first line of defense against intestinal infections . In the mouse, the vast majority of i-IEL express the CD8 coreceptor either as a CD8 alpha/alpha homodimer or as a CD8 alpha/beta heterodimer . The CD8 receptor of T-cell receptor TcR gamma/delta i-IEL is exclusively homodimeric, whereas the CD8-expressing TcR alpha/beta i-IEL segregate into equal fractions of CD8 alpha/alpha and CD8 alpha/beta cells . We infected beta 2-microglobulin (beta 2m)+/- mice (possessing all i-IEL populations) and beta 2m -/- mutant mice (lacking all CD8 alpha/beta + i-IEL and having few CD8 alpha/alpha + TcR alpha/beta i-IEL) with L . monocytogenes per os and determined their biological functions after TcR ligation with monoclonal antibodies . Cytolytic activities of TcR alpha/beta and TcR gamma/delta i-IEL from beta 2m +/- mice were not influenced by intestinal listeriosis . Cytolytic activities of TcR alpha/beta i-IEL were impaired in uninfected beta 2m -/- mice, but this reduction was reestablished as a consequence of intestinal listeriosis . Frequencies of gamma interferon (IFN-gamma)-producing TcR alpha/beta i-IEL in uninfected beta 2m -/- mice were reduced, compared with that in their heterozygous controls . Equally low frequencies of IFN-gamma-producing TcR gamma/delta i-IEL in beta 2M +/- and beta 2m-/- mutants were found . Listeriosis increased frequencies of INF-gamma-producing TcR alpha/beta and TcR gamma/delta i-IEL in both mouse strains . Most remarkably, the proportion of IFN-gamma-producing TcR gamma/delta i-IEL was elevated 10-fold in listeria-infected beta 2M -/- mice . Our findings show that the beta 2m-independent CD8 beta- i-IEL expressing either TcR alpha/beta or TcR gamma/delta are stimulated by intestinal listeriosis independent of regional beta 2m expression . We conclude that the three major CD8+ i-IEL populations are stimulated by intestinal listeriosis and that CD8 beta- i-IEL compensate for the total lack of CD8 beta+ i-IEL in beta 2M -/- mutant mice . Hence, in contrast to the peripheral immune system, which crucially depends on CD8 alpha/beta + TcR alpha/beta lymphocytes, the mucosal immune system can rely on additional lymphocytes expressing the CD8 alpha/alpha homodimer. J Immunol, 1996 Jan 15, 156(2), 683 - 92 Two Listeria monocytogenes CTL epitopes are processed from the same antigen with different efficiencies; Sijts AJ et al.; Listeria monocytogenes is an intracellular bacterium that elicits MHC class I-restricted CTL in infected mice . A major CTL specificity is the nonamer peptide p60 217-225, which is derived from the bacterial murein hydrolase p60 and presented by the H-2Kd MHC class I molecule . In this report, we identify a second H-2Kd presented peptide, encompassing residues 449-457 of p60, that is detected by L . monocytogenes-specific CTL . Both p60-derived CTL epitopes are good competitors for H-2Kd binding and TAP (transporter associated with Ag processing) transport . CTL clone WP11.12 lyses L . monocytogenes infected cells and recognizes naturally processed p60 449-457 acid eluted from L . monocytogenes-infected macrophages . Although both epitopes derive from the same Ag and bind the same allelic form of MHC class I, quantitative analysis reveals that the amount of p60 449-457 in infected cells is approximately 10-fold greater than the amount of p60 217-225 . Shuffling p60 217-225 into position 449-457 decreases its processing efficiency, indicating that the large number of p60 449-457 epitopes cannot be entirely attributed to epitope-flanking sequences . Our findings indicate that CTL epitopes can be processed from Ags with markedly different kinetics and efficiencies . Intrinsic qualities of an epitope and its location within a protein influence the efficiency of Ag processing. J Immunol, 1996 Jan 15, 156(2), 679 - 84 Listeria monocytogenes induces apoptosis of infected hepatocytes; Rogers HW et al.; Infection with blood-borne Listeria monocytogenes results in their early uptake by the liver . Foci of hepatocytes become heavily infected and develop into microabscesses . Infection results in apoptosis of the hepatocytes . This is particularly evident in the edge of the microabscess, where hepatocytes are not yet destroyed by the neutrophil . It is also apparent when neutrophils are depleted from the circulation . Infection of hepatocytes in culture induces their death by apoptosis with the release of neutrophil chemoattractants . Cytokines do not reduce the multiplication of Listeria in cultured hepatocytes . This study calls attention to an early program of inflammation induced in infected cells that are unresponsive to cytokines. Immunobiology, 1996, 196(4), 449 - 62 Down-regulation of Listeria monocytogenes-specific Th1 cytokine response by treatment of mice with goat antibody to mouse IgD; Song F et al.; Injection of goat anti-mouse IgD antibody (G alpha M IgD) to mice has been shown to induce polyclonal IgG1 and IgE production by B cells and IL-4 production by goat Ig-specific T cells . Surface IgD crosslinking also activates function of B cells as antigen presenting cells . Although the G alpha M IgD treatment is a well established system for regulation of immune response against antigens that bind to B cell receptor, we found that the G alpha M IgD treatment also influences immune response against irrelevant bacterial antigen . The T cells from the G alpha M IgD-treated Listeria monocytogenes-infected mice showed increased IL-4 production and decreased IFN-gamma and IL-2 production against listerial antigen compared with those from control L . monocytogenes-infected mice . Interestingly, changes were also found in antigen presenting cells in the G alpha M IgD-treated mice . MHC class II expression of both B cells and macrophages decreased significantly in the G alpha M IgD-treated mice, suggesting cytokine induced by G alpha M IgD-treatment may suppress MHC class II expression and modulate APC function in the G alpha M IgD-treated mice . In accordance with the assumption, T cells from the G alpha M IgD-treated mice produced high amount of IL-4 and IL-10 in in vitro culture with goat serum which contain goat Ig . These result suggest that G alpha M IgD treatment may modulate APC function in the G alpha M IgD-treated mice through Th2 type cytokine(s) produced by goat Ig-specific T cells, which results in changes of Th response against irrelevant antigen. Acta Vet Hung, 1996, 44(3), 277 - 85 Occurrence of Listeria and listeriosis in Hungary; Ralovich B et al.; Listeriosis is a rare human disease in Hungary . The number of cases is slowly increasing . Only sporadic events have been observed but the occurrence of epidemic listeriosis may be supposed . The Listeria monocytogenes (in abbreviation: L . m.) transmitter role of food in human infections has not yet been verified . The epidemiological character of animal listeriosis is different . Healthy carriers can be found among both humans and animals . Foodstuffs of animal as well as plant origin may be contaminated with Listeria . When the processing technology and/or hygienic conditions are not satisfactory, these microorganisms can be detected in food factories and in final products of the food industry. Gastroenterol Clin Biol, 1996, 20(8-9), 700 - 2 {Spontaneous Listeria monocytogenes peritoneal infection complicating hepatic transplantation}; Chapoutot C et al.; The incidence of listeriosis is increased in immunosuppressed patients . We report a case of spontaneous bacterial peritonitis with bacteraemia caused by Listeria monocytogenes in a 47-year old woman with liver transplantation . Complete recovery was achieved after amoxicillin and amikacin therapy . High doses of corticosteroids and OKT3 monoclonal therapy may have favoured the occurrence of infection . In liver transplant recipients, regular stool screening could be proposed, and trimethoprim-sulfamethoxazole antibioprophylaxy could be used when Listeria monocytogenes is isolated in stool culture or immunosuppressive therapy is increased. Brain Res Bull, 1996, 41(6), 327 - 33 Sorting signals and targeting of infectious agents through axons: an annotation to the 100 years' birth of the name "axon"; Kristensson K; A brief review is given on mechanisms by which axons may be initiated during development and by which the polarity of neurons is maintained by selective sorting and delivery of molecules to axons and dendrites . The use of viruses as tools to study targeting of newly synthesized proteins to axons is described . Emphasis is then given to the hazards that are presented to the individual by the retrograde transport of infectious agents in axons to the brain . Borna disease virus, prions, and Listeria monocytogenes are examined briefly as examples of these mechanisms . These agents have attracted interest previously in veterinary medicine for the most part, but they may present potential and substantial threats to human health . Such infectious agents also represent a new type of virus, a new principle for disease transmission, and a new mechanism for intracellular transport, respectively. Scand J Infect Dis, 1996, 28(4), 367 - 74 Listeriosis in 225 non-pregnant patients in 1992: clinical aspects and outcome in relation to predisposing conditions; Goulet V et al.; Clinical information was collected on 225 cases of Listeria monocytogenes (LM) infection not associated with pregnancy occurring in France in 1992 . Infections affected primarily men (62%) and persons over 65 years of age . Of the cases found, 81% occurred in persons with some underlying condition: 34% involved patients with severe immunosuppression; 37% were on dialysis or had diabetes mellitus, alcoholism, hepatic failure or malignancies without immunosuppressive therapy; and 10% had other underlying conditions not clearly associated with immunosuppression . The clinical presentation of listeriosis depended on the predisposing factors: in previously healthy adults central nervous system infection was the most frequent clinical form (80%), whereas the group characterized by severe immunosuppression or other immunosuppressive conditions tended to develop bacteraemia (52%) . The rate of hospital-associated cases (11%) was lower than that reported in other countries . Mortality directly related to LM infection was 24% . Predisposing disease was the major prognostic factor . No fatal outcome was observed in the group of adults < 65 years old without underlying conditions . In summary, although classification based on the degree of alteration of T-cell-mediated immunity enables determination of the role of predisposing conditions in cases of listeriosis outside of pregnancy, clinical aspects and outcome of listeriosis vary within these groups and depend on the severity of each underlying disease. Cell Motil Cytoskeleton, 1996, 34(4), 279 - 87 Novel form of actin-based motility transports bacteria on the surfaces of infected cells; Sanger JM et al.; Enteropathogenic Escherichia coli (EPEC) attach to cells (attachment) lining the intestine and induce a decrease in the number of the cells' microvilli (effacement) . This attachment and effacement is followed by diarrhea, which may be explained, at least in part, to the loss of microvilli and the decreased ability of the infected cells to absorb fluids . EPEC also attach to the surfaces of a number of cultured cells including CaCo-2, LLC-PK, and PtK2 cells . The extracellular, attached EPEC induce filaments of actin to form in the cytoplasm just underneath the EPEC surface attachment sites . Beneath some of the attached EPEC, the actin filaments become organized into membrane encased columns that extend up to 6 micrometers above the cell surface creating "pedestals" on which the EPEC rest . The raised pedestals can be readily observed in stereo pairs taken using the Intermediate Voltage Electron Microscope . The concentration of non-muscle isoforms of myosin II and tropomyosin near the base of the pedestals suggests a similarity of these structures to brush border microvilli . Video microscopy indicates that these EPEC pedestals can bend and undulate, alternately growing longer and shorter while remaining tethered in place on the cell surface . Some of the attached EPEC also translocate along the cell surface, reaching speeds up to 0.07 micrometers/sec . Both types of movement are inhibited by cytochalasin D, indicating that actin polymerization in the pedestals is required for the motility of EPEC on the host cell surface . In this respect, EPEC motility on host cells resembles the intracellular motility of Listeria, but there are differences in the actin filament bundles induced by the two different bacteria . The most obvious one is the interposition of the cell membrane between EPEC and the actin filaments in the pedestal in contrast to the close apposition of actin filaments to Listeria . The intensity of fluorescence of rhodamine phalloidin is nearly uniform along most of the length of the pedestals indicating a constant number of actin filaments, whereas the fluorescence intensity decreases along the length of Listeria tails reflecting the disassembly that occurs all along the tails . Epec's movements may be a hybrid of Listeria filopodia and Aplysia inductopodia movements . This paper is the first report of a microbe attached to the extracellular surface of an infected cell propelled by an intracellular actin polymerization-dependent mechanism. Microbiol Immunol, 1996, 40(2), 121 - 24 The prevalence of Listeria monocytogenes in the environment of dairy farms; Ueno H et al.; The prevalence of Listeria monocytogenes in the environment of dairy farms was surveyed from December 1993 to June 1994 in one city of Hokkaido . L . monocytogenes was isolated from 3 out of 5 farms investigated . Serovar 4b organism was isolated from the brain stem of a cow from one farm which was clinically diagnosed as having listeriosis . The same serovar of L . monocytogenes was also isolated from the rectal contents of a healthy cow, straw on the floor, straw in the barn, and silage scattered around the silo from the same farm . At another farm, with no reported cases of bovine listeriosis, serovar 1/2 organism was isolated from the same types of samples as the above mentioned farm except from straw on the floor . The difference in the isolation rates of the organism from straw on the floor between the two farms (22%:5/23 vs 0%:0/24) is considered to be caused by the different feeding methods of silage between the two farms. Rev Soc Bras Med Trop, 1996 Jan-Feb, 29(1), 41 - 5 {An anti-Listeria seroepidemiological study in Uberaba, MG}; Tavares-Neto J et al.; From a total of 445 individuals, 17.1% had antibodies against L . monocytogenes detected by the agglutination tube test . They were separated in seven groups: bloods donnors (n = 50), Hospital visitors (n = 40), frigorific workers (n = 28), aviculture workers (n = 87), herdsman (n = 31), agriculture students (n = 60) and street-sweepers (n = 51) . L1/2a serotype was predominant . Individuals from urban areas (19.5%) and those who had less contact with animals (21.7%) had significantly positive serology when compared with individuals from rural areas (9.4%) and those who had close contact with animals (13.2%) . The overall picture is individuals of more specialized occupations had more frequently (25.9%) anti listeria antibodies similar to the results observed in developed countries where listeriosis is a public health problem in urban areas. Eur Radiol, 1996, 6(2), 188 - 91 Cerebritis due to Listeria monocytogenes: CT and MR findings; Aladro Y et al.; Infections by Listeria monocytogenes are uncommon, with cerebritis being even rarer . We present three cases of cerebritis which occurred during an outbreak of listeriosis . The CT and MR findings at diagnosis and during follow-up are described . Predominant deep white matter lesions with nodular and ring enhancement were seen . The MR yielded a higher resolution of the lesions than CT. Neth J Med, 1996 Jan, 48(1), 15 - 7 Listeria monocytogenes endocarditis in a patient with an aortic prosthetic valve; Castro Cabezas M et al.; Patients with prosthetic cardiac valves have an increased risk of developing bacterial endocarditis . The causative micro-organism in bacterial endocarditis may be a guide to the portal of entry . In this case report, we describe a patient with a prosthetic cardiac valve who suffered from recurrent endocarditis with different micro-organisms from the gastrointestinal tract. J Clin Microbiol, 1996 Jan, 34(1), 15 - 9 Comparison of ribotyping, arbitrarily primed PCR, and pulsed-field gel electrophoresis for molecular typing of Listeria monocytogenes; Louie M et al.; Fifty-one clinical isolates of Listeria monocytogenes (15 isolates from two outbreaks and 36 epidemiologically unrelated isolates) were typed by conventional serotyping, ribotyping (RT), pulsed-field gel electrophoresis (PFGE), and arbitrarily primed PCR (AP-PCR) . Serotyping was unable to distinguish between related and unrelated strains of L . monocytogenes . Each of the three molecular methods showed excellent typeability and reproducibility . Restriction with EcoRI and PvuII gave 16 and 23 RT patterns, respectively . Restriction with ApaI or SmaI generated 22 and 26 PFGE profiles, respectively . ApaI profiles were easier to interpret, with 10 to 15 bands each, while SmaI profiles had 15 to 20 bands each . AP-PCR with two different primers yielded 29 and 31 randomly amplified polymorphic DNA patterns, respectively . Strains from the same outbreak shared concordant patterns by each of the three methods . Of the three techniques evaluated, RT was the least discriminating and could not distinguish between strains from the two outbreaks . The abilities of AP-PCR and PFGE to differentiate between strains were comparable . However, AP-PCR was more rapid and easier to perform . We conclude that the DNA profiles generated by either AP-PCR or PFGE can be used to differentiate outbreak strains from epidemiologically unrelated strains and to clearly identify unrelated strains as being distinct from one another . We recommend that at least two independent primers be used for AP-PCR typing in order to improve its discriminatory power. Annu Rev Immunol, 1996, 14, 207 - 32 Molecular mechanisms of lymphocyte-mediated cytotoxicity and their role in immunological protection and pathogenesis in vivo; Kagi D et al.; Studies with perforin-deficient mice have demonstrated that two independent mechanisms account for T cell-mediated cytotoxicity: A main pathway is mediated by the secretion of the pore-forming protein perforin by the cytotoxic T cell, whereas an alternative nonsecretory pathway relies on the interaction of the Fas ligand that is upregulated during T cell activation with the apoptosis-inducing Fas molecule on the target cell . NK cells use the former pathway exclusively . The protective role of the perforin-dependent pathway has been shown for infection with the noncytopathic lymphocytic choriomeningitis virus, for infection with Listeria monocytogenes, and for the elimination of tumor cells by T cells and NK cells . In contrast, perforin-dependent cytotoxicity is not involved in protection against the cytopathic vaccinia virus and vesicular stomatitis virus . LCMV-induced immunopathology and autoimmune diabetes have been found to require perforin-expression . A contribution of perforin-dependent cytotoxicity to the rejection of MHC class I-disparate heart grafts has also been observed . Its absence is efficiently compensated in rejection of fully allogeneic organ or skin grafts . So far, evidence for a role of Fas-dependent cytotoxicity as a T cell effector mechanism in vivo is lacking . Current data suggest that the main function of Fas may be in regulation of the immune response and apparently less at the level of an effector mechanism in host defense . Further analysis is necessary, however, to settle this point finally. Int Immunol, 1996 Jan, 8(1), 23 - 36 Multiple immune abnormalities in tumor necrosis factor and lymphotoxin-alpha double-deficient mice; Eugster HP et al.; To investigate the roles of tumor necrosis factor (TNF) and lymphotoxin (LT)-alpha in the development and function of the immune system, the Tnf and Ltalpha genes were simultaneously inactivated in mice by homologous recombination . These mutant mice are highly susceptible to Listeria monocytogenes infection and resistant to endotoxic shock induced by the combined administration of D-galactosamine (D-GaIN) and lipopolysaccharide (LPS) . Their splenic microarchitecture is disorganized, characterized by the loss of the clearly defined marginal zone, ill defined T and B cell areas, and absence of MAdCAM-1 and reduced ICAM-1, VCAM-1 and Mac-1 expression . They are devoid of peripheral lymph nodes and Peyer's patches, and show a strong reduction of IgA+ plasma cells in the intestinal lamina propria . The alymphoplasia is accompanied by a marked B lymphocytosis and reduced basal lg levels . Ig depositions in the renal glomerulus and a strong up-regulation of MHC class I antigen expression on endothelial cells of different tissues are observed . The primary humoral immune response towards sheep red blood cells reveals a defective IgG isotype switch, while that against vesicular stomatitis virus is normal . The cytotoxic T cell responses are attenuated, although still effective, against vaccinia, lymphocytic choriomeningitis virus (LCMV-ARM) and LCMV-WE . In conclusion, the combined inactivation of Tnf and Ltalpha confirms their essential role in the normal development and function of the immune system. Immunology, 1996 Jan, 87(1), 15 - 20 Suppression of T-helper type-1 immune response against Listeria monocytogenes by treatment of mice with goat antibodies to mouse IgD; Matsuzaki G et al.; Injection of goat anti-mouse IgD antibodies (GAM IgD) to mice has been shown to induce polyclonal IgG1 and IgE production by B cells and interleukin-4 (IL-4) production by goat Ig-specific T cells . Surface IgD cross-linking also activates B cells to function as antigen-presenting cells (APC) . Although the GAM IgD treatment is a well-established system for analysis of B-cell dependent antigen presentation, the influence of GAM IgD treatment on the immune response to irrelevant antigens is not known . To address this issue, we analysed effects of GAM IgD treatment on (1) the mitogen response of freshly isolated T cells, and (2) the listerial antigen-specific response after immunization with viable Listeria monocytogenes, which induces CD4+ interferon-gamma (IFN-gamma) producing protective T cells in normal mice . Spleen CD4+ T cells from the GAM IgD-treated mice produced higher levels of IL-4 but lower levels of IFN-gamma and IL-2 than those from the control mice when they were stimulated with concanavalin A (Con A) in vitro . When spleen T cells were stimulated with listerial antigen 10 days after a low dose (1/20 LD50) of L . monocytogenes infection, CD4+ T cells from the GAM IgD-treated mice showed increased IL-4 production and decreased IFN-gamma and IL-2 production compared with those from the control L . monocytogenes-infected mice . Furthermore, the GAM IgD treatment resulted in a reduction of the survival rate after a high dose (1/2 LD50) of L . monocytogenes infection . These results suggest that treatment of mice with GAM IgD suppresses the T-helper type-1 (Th1)-type T-cell response and induces a Th2-type response against irrelevant antigens, even when they are injected after GAM IgD treatment. Acta Vet Scand, 1996, 37(1), 13 - 8 Prevalence of Listeria monocytogenes and other Listeria spp.in smoked and 'gravad' fish; Loncarevic S et al.; Altogether 150 samples of vacuum-packed fish were examined for the presence of Listeria species . Listeria monocytogenes were isolated from 12 of 58 'gravad' fish samples, 3 of 26 cold-smoked and one of 66 hot-smoked fish samples . Ten of these 16 positive samples harboured more than 100 L . monocytogenes cfu/g . The highest level (132,000) was found in a sample of hot-smoked rainbow trout (Oncorhynchus mykiss) . Serogroup 1/2 was most frequently found, followed by 4 and 3 . One sample of gravad rainbow trout harboured more than one serogroup of L . monocytogenes . L . innocua and L . seeligeri were isolated from 12 and 1 samples, respectively. Int J Food Microbiol, 1996 Jan, 28(3), 351 - 9 Multilocus enzyme electrophoresis typing of New Zealand Listeria monocytogenes isolates; Avery SM et al.; Eighty-five strains of Listeria Monocytogenes, comprising 74 strains isolated from the environment, humans, animals and foods in New Zealand; 10 strains from similar sources in Denmark, and one reference strain, were studied using multilocus enzyme electrophoresis (MEE) . Fifty-five electrophoretic types (ETs) were identified among the 85 strains, most (71%) of which were represented by only one isolate . The 74 New Zealand isolates produced 45 ETs, most (67%) of which were represented by only one isolate . There was no link between the source of the isolates and ET, as most ETs contained only one isolate . One cluster of ETs, designated cluster 45, contained only environmental isolates . Of the five ETs that contained isolates from more than one source, four contained isolates that were obtained from clinical specimens and foods indicating a possible link between isolates from these two sources . The population of New Zealand isolates of L . monocytogenes studied appears to be genetically diverse when compared with the diversity found in similar studies in other countries. Int J Food Microbiol, 1996 Jan, 28(3), 333 - 40 Automated RNA probe assay for the identification of Listeria monocytogenes; Mabilat C et al.; Recent epidemics of human listeriosis have shown the importance of Listeria monocytogenes as a food-borne risk . We have developed an automated identification assay for L . monocytogenes using the specificity of the 16S rDNA probe described by Wang et al . (1991) . Bacterial cultures were lysed quickly by a chemical step releasing the target nucleic acids . Extracts were loaded into the VIDAS automate (bioMerieux) which performed a non-radio-active hybridisation reaction for 2 h . This assay recognised specifically 68 out of 69 L . monocytogenes isolates from clinical and food origins, including serovars 4b and 1/2, without cross-hybridising to other Listeria species (103 strains) or other bacterial species (15 strains) . The sensitivity of the assay was 10(7) bacteria . This automated technology is faster than conventional biochemical identification. J Immunol, 1996 Jan 1, 156(1), 232 - 7 In vivo cytokine production in murine listeriosis . Evidence for immunoregulation by gamma delta+ T cells; Hsieh B et al.; Differential cytokine production by gamma delta+ T cells influences Th1 and Th2 responses . Here, we describe the in vivo kinetics of peritoneal and splenic gamma delta+, CD4+, and CD8+ T cell cytokine responses during primary and secondary Listeria infections . The data show differences in the kinetics of IFN-gamma-producing alpha beta+ splenocytes consistent with immunologic memory . Most noteworthy, however, was the elevated production of IL-10 by splenic gamma delta+ T cells in the red pulp and marginal zones that coincided with maximal IFN-gamma production and with a decrease in inflammation and tissue damage . This result implies a role for gamma delta+ T cells in the control of Th1 responses. Lett Appl Microbiol, 1996 Jan, 22(1), 16 - 7 A comparison between starch and polyacrylamide gels for the analysis of Listeria monocytogenes using multilocus enzyme electrophoresis; Flint SH et al.; Forty-seven Listeria monocytogenes isolates were analysed using multilocus enzyme electrophoresis in two laboratories . Both assayed for the same six enzymes, but one used a starch gel method and the other polyacrylamide gels . The starch gel method distinguished six electrophoretic types whereas the polyacrylamide gel method produced 17 different electrophoretic types . The polyacrylamide gel method was more discriminatory than the starch gel method. Lett Appl Microbiol, 1996 Jan, 22(1), 13 - 5 Effects of benzylpenicillin on glucose utilization, macromolecule synthesis and cell wall proteins of Listeria monocytogenes; Chen LJ et al.; When growing cells were incubated in the presence of 100 ppm benzylpenicillin (BP) for 30 min and then harvested for metabolic studies, the initial rate of glucose utilization by resting cells was reduced by 19.6% compared to control cells, whereas the rates of protein, DNA and RNA synthesis were reduced by 40.7%, 55.0% and 87.5% respectively . Growth in the presence of 100 ppm BP for 2 h was found to result in a marked depletion of cell wall proteins. Lett Appl Microbiol, 1996 Jan, 22(1), 10 - 2 Effect of benzylpenicillin on the viability and osmotic sensitivity of Listeria monocytogenes; Chen LJ et al.; Growth in the presence of 100 ppm benzylpenicillin (BP) for 2 h failed to result in a detectable reduction in cfu . Cells grown and incubated for 24 h in 0, 1, 10 and 100 ppm BP underwent reductions in cfu of 0%, 0%, 90% and 99% respectively of maximum cfu values. Microbiology, 1996 Jan, 142 ( Pt 1), 173 - 80 The inlA gene required for cell invasion is conserved and specific to Listeria monocytogenes; Poyart C et al.; The Gram-positive bacterium Listeria monocytogenes can actively induce its own uptake by epithelial cells and fibroblasts through a surface-exposed 80 kDa protein, internalin (InIA), encoded by inIA . We studied the distribution and the DNA polymorphism of inIA sequences in a wide variety of wild strains of L . monocytogenes as compared to other Listeria species . This was done by PCR-amplifying inIA sequences encoding the fifteen repeats A and the three repeats B of InIA . inIA-repeated sequences were only found in L . monocytogenes . The amplified fragment of inIA encoding the repeats A displayed an AIuI DNA polymorphism which arises from point mutations . These results indicate that inIA required for cell invasion is specific to L . monocytogenes and that the intragenic repeats only exhibit a genetic heterogeneity due to point mutations and not to recombinations. Appl Environ Microbiol, 1996 Jan, 62(1), 304 - 7 Aerobic and anaerobic metabolism of Listeria monocytogenes in defined glucose medium; Romick TL et al.; A defined medium with glucose as the carbon source was used to quantitatively determine the metabolic end products produced by Listeria monocytogenes under aerobic and anaerobic conditions . Of 10 strains tested, all produced acetoin under aerobic conditions but not anaerobic conditions . Percent carbon recoveries of end products, typified by strain F5069, were as follows: lactate, 28%; acetate, 23%; and acetoin, 26% for aerobic growth and lactate, 79%; acetate, 2%; formate, 5.4%; ethanol, 7.8%; and carbon dioxide, 2.3% for anaerobic growth . No attempt to determine carbon dioxide under aerobic growth conditions was made . The possibility of using acetoin production to assay for growth of L . monocytogenes under defined conditions should be considered. J Neuropathol Exp Neurol, 1996 Jan, 55(1), 14 - 24 Intracerebral targets and immunomodulation of murine Listeria monocytogenes meningoencephalitis; Schluter D et al.; In humans, infection with Listeria monocytogenes (L . monocytogenes) can severely affect the central nervous system (CNS) . In the present study we have employed a murine model of CNS listeriosis to characterize the intracerebral distribution of L . monocytogenes . Following intracerebral application of a low dose of L . monocytogenes (serovar 1/2a, EGD strain) a severe fatal leptomeningitis, ventriculitis, and encephalitis developed . Listeria were detectable both intracellularly in different cell types of the CNS and extracellularly in the cerebrospinal fluid . Ultrastructural analysis revealed macrophages, granulocytes, plexus epithelial cells, ependymal cells, and neurons as target cells . An inflammatory reaction with macrophages and granulocytes developed in the brains of these animals but was not sufficient to prevent the fatal outcome of the disease . However, active immunization of mice prior to an intracerebral challenge infection significantly reduced the mortality . Immunized animals showed an early recruitment of a significant number of CD8+ and, to a lesser degree, CD4+ T cells within 24 hours p.i . as well as a strong activation of microglial cells and macrophages . These findings may provide an interesting model for studies on the pathogenesis of cerebral listeriosis. Proc Natl Acad Sci U S A, 1995 Dec 19, 92(26), 12389 - 92 Stimulation of protective CD8+ T lymphocytes by vaccination with nonliving bacteria; Szalay G et al.; Infectious diseases caused by intracellular microbes are responsible for major health problems, and satisfactory control will ultimately depend on efficient vaccination strategies . The general assumption is that activation of protective immune responses against intracellular microbes dominated by CD8+ T cells are achieved only by live vaccines . In contrast, we here demonstrate stimulation of protective immunity in mice against the intracellular pathogen Listeria monocytogenes by vaccination with heat-killed listeriae . Vaccine-induced immunity comprised cytolytic and interferon gamma-producing CD8+ T lymphocytes . CD8+ T cells from vaccinated donor mice transferred protection against listeriosis . Moreover, vaccination with heat-killed listeriae induced production in CD4+ T-cell-deficient, H2-A beta gene-disrupted mutant mice . We conclude that antigens from killed listeriae are introduced into the major histocompatibility complex class I pathway and thus are recognized by CD8+ T cells . The practicability of killed vaccines against human infectious diseases therefore should be reevaluated. J Biotechnol, 1995 Dec 15, 43(3), 205 - 12 Hyperexpression of listeriolysin in the nonpathogenic species Listeria innocua and high yield purification; Darji A et al.; Listeriolysin, the hemolysin of the pathogenic species Listeria monocytogenes, was expressed in the non-pathogenic species Listeria innocua . Coexpression of the positive regulatory factor prfA in the plasmid vector in conjunction with the structural gene hly increased the expression over 500-fold . Purification from supernatant fluids was achieved by two steps of ion exchange chromatography . The procedure resulted in over 60% yield of a hemolytically active, homogeneous 58 kDa protein which was used to produce monospecific antibodies . As shown by immunoblot the purified listeriolysin was free of p60, a highly immunogenic protein of similar size also produced by Listeria spp., which otherwise would interfere with immunoassays . Listeriolysin retained full activity for more than 6 months at -70 degrees C. Eur J Epidemiol, 1995 Dec, 11(6), 665 - 73 The use of PCR ribotyping for typing strains of Listeria spp; Sontakke S et al.; The potential of PCR ribotyping for discriminating between and within various species of Listeria, as well as strains of Listeria monocytogenes was examined . In total, 49 strains of Listeria monocytogenes and 12 isolates of Listeria spp . were analyzed . The genomic DNA isolated from these strains was subjected to PCR amplification in the regions between 16S and 5S rRNA . Amplifications were performed with both low and high concentrations of Taq polymerase . Length polymorphisms in the amplified DNA products enabled distinction between various strains of Listeria spp . and between various serotypes of L . monocytogenes . Six composite profiles for serotype 4b strains, 8 for 1/2a strains and 11 for 1/2b strains, were observed . In addition, several different PCR ribotyping strategies were evaluated . Restriction fragment length polymorphisms of the spacer region between the 16S and 23S rRNA genes of 16 strains of L . monocytogenes were not observed, except for two isolates . PCR ribotyping analysis displayed promise as an alternative to traditional L . monocytogenes molecular typing methods. Immunol Rev, 1995 Dec, 148, 5 - 18 Immune defence in mice lacking type I and/or type II interferon receptors; van den Broek MF et al.; Mice lacking the receptor for type I interferon (IFN-alpha beta, A129 mice), for type II interferon (IFN-gamma, G129 mice) or for both receptors (AG129 mice) have been generated by embryonic stem cell mediated gene targeting and inter-crossing A129 x G129, respectively . The role of the two IFN systems in controlling a range of infections has been studied using these mice . Type I IFN is shown to be responsible for the immune defence against most viral infections tested (Lymphocytic Choriomeningitis Virus, Semliki Forest Virus, Theiler's Virus, Vesicular Stomatitis Virus), type II IFN seems to be of little importance . In Vaccinia Virus and Theiler's Virus infection, however, both IFN systems were found to play a nonredundant role . IFN-gamma was critical for the defence against intracellular bacteria (Mycobacterium, Listeria) and parasites (Leishmania), whereas IFN-alpha beta was not . IFN-alpha beta is produced by virus-infected cells within hours and plays an important role in preventing virus spread early . Production of IFN-gamma on the other hand needs activation of the immune system and plays a major role later, i.e . mostly during the immune response . Data obtained with the mice described here show that both IFN systems seem to have evolved to complement each other in the host defence against a wide variety of infectious agents. Mol Microbiol, 1995 Dec, 18(5), 801 - 11 Modulation of DNA topology by flaR, a new gene from Listeria monocytogenes; Sanchez-Campillo M et al.; We report the identification of a previously unknown Listeria monocytogenes gene, flaR, which modulates DNA topology . Through the analysis of a Tn917 non-motile mutant, LOSC1, in which production of flagellin was abolished, we have identified a bacterial component involved in gene regulation . The transposon had inserted in flaR, an open reading frame of 531 bp, followed by a second open reading frame of 1252 bp in reverse orientation . On the L . monocytogenes physical map, flaR was located in a different region from that of the flaA gene encoding flagellin . Transcriptional analysis showed that the flaR gene product affects the flaA expression and negatively regulates its own expression . When expressed in Escherichia coli, flaR encodes a protein of 18 kDa (FlaR) whose transcription is osmoregulated . In addition, FlaR also influences the expression of reporter genes containing supercoiling-sensitive promoters such as proU or ompC . The data presented here suggest that FlaR is a histone-like bacterial protein which acts at specific sites to influence DNA topology and, therefore, transcription . flaR is the first gene of this class to be described in Gram-positive pathogenic bacteria. Mol Cell Probes, 1995 Dec, 9(6), 423 - 32 Simultaneous detection of Listeria spp . and Listeria monocytogenes by reverse hybridization with 16S-23S rRNA spacer probes; Rijpens NP et al.; Enzymatic amplification results showed that Listeria species have at least two 16S-23S rRNA spacer regions of different lengths . These spacer regions of L . monocytogenes, L . ivanovii and L . seeligeri were cloned after enzymatic amplification . Sequence analysis of the inserts revealed two spacers of 245-246 bp and 496-498 bp, respectively, of which the latter included tRNA(Ala) and tRNA(Ile) genes . One Listeria spp.-specific probe, LIS-ICG4, was deduced from the 245-bp spacer and a L . monocytogenes-specific probe, LMO-ICG5, was inferred from the 496-bp spacer . The specificity of both probes was tested in a reverse hybridization assay (Line Probe Assay, LiPA) . Both LIS-ICG4 and LMO-ICG5 proved to be highly specific when hybridized to a large collection of Listeria strains and strains from other relevant taxa . The LiPA test herein described for the simultaneous detection of Listeria spp . and L . monocytogenes can be expanded to detect other foodborne pathogens. Eur J Oral Sci, 1995 Dec, 103(6), 355 - 61 Plaque bacteria counts and vitality during chlorhexidine, meridol and listerine mouthrinses; Netuschil L et al.; The aim of this double-blind study was to enumerate the total number of living and dead bacteria on defined tooth areas during the application of antibacterial mouthrinses . After prophylaxis, 40 students refrained from all oral hygiene measures for 3 d, during which they rinsed with a phenolic compound (Listerine), an amine fluoride/stannous fluoride solution (Meridol), 0.2% chlorhexidine (CHX) or a control solution (0.02% quinine-hydrochloride) . The plaque index (P1I) was recorded at the start and the end of the investigation . Total bacterial counts (BC) and colony-forming units (CFU) of 1d-, 2d- and 3d-old dentogingival plaque were determined . The plating efficiency (PE) was calculated as a percentage of CFU/BC and the portion of vital microflora estimated by a vital fluorescence technique (VF) . All groups started with a P1I approximating 0.1 . On day 3, the P1I values were 1.21 in the control group and 0.51, 0.37 and 0.14 after Listerine, Meridol and CHX use, respectively . A tremendous variation existed between the numbers of viable bacteria found per mm2 on the enamel surface and day 3 (CHX: 0.2; Meridol: 300; Listerine; 6x10(4); control: 2x10(6)), while higher total numbers of bacteria were concomitantly present (CHX and Meridol: 1-2x10(4); Listerine: 2x10(5); control: 2x10(6)) . Both vitality parameters PE and VF reached 92% in the control group at day 3, but only 7% after CHX use . With Meridol and Listerine, the corresponding PE values were 3% and 43%, respectively, while the VF values reached 48% and 54% . The PII, BC, CFU and PE values of the CHX and the Meridol groups differed significantly from those of the control group . In contrast, Listerine showed no difference as compared to the control rinse . Due to the strong antibacterial action of CHX and Meridol during their use, almost only dead or non-proliferating bacteria were found on the tooth surfaces . Thus, only a thin plaque could develop . As a clinical consequence, both substances showed retardation of plaque development as reflected by significantly reduced plaque indices. Acta Paediatr, 1995 Dec, 84(12), 1434 - 5 Peripheral blood gamma delta T cells in human listeriosis; Bertotto A et al.; A phenotypical analysis carried out by direct immunofluorescence and two-colour cytofluorometry showed that the number of lymphocytes bearing the gamma delta T-cell receptor heterodimer was increased in the blood of eight children with Listeria monocytogenes infection, mainly due to an expansion of cells identified by monoclonal antibodies which recognize V delta 2 gene products . These findings are further evidence that gamma delta T cells are in some way involved in the immune response directed against human intracellular pathogens. J Clin Microbiol, 1995 Dec, 33(12), 3349 - 51 Frozen stored murine hybridoma cells can be used to determine the virulence of Listeria monocytogenes; Bhunia AK et al.; Murine hybridoma cells, designated Ped-2E9, when stored up to 60 days at -196 degrees C or up to 48 days at -80 degrees C, gave results equivalent to those for freshly grown murine hybridoma cells in an in vitro pathogenicity assay of Listeria species . Thus, laboratories do not need to have their own tissue culture facilities to maintain the hybridoma cells for the assay described. Eur J Immunol, 1995 Dec, 25(12), 3321 - 5 Interleukin-4-producing CD4+ NK1.1+ TCR alpha/beta intermediate liver lymphocytes are down-regulated by Listeria monocytogenes; Emoto M et al.; Experimental infection of mice with the intracellular bacterium, Listeria monocytogenes, provides a paragon model for immune defence dominated by T helper type 1 (Th1) responses . Potent production of interleukin (IL)-12 by infected macrophages is considered the determining factor in Th1 cell development . In contrast, it is assumed that IL-4 producers remain virtually unstimulated in listeriosis . In the liver, the major target organ of listeriosis, an unusual T lymphocyte population exists with the intriguing phenotype CD4+ NK1.1+ TCR alpha/beta intermediate (TCR alpha/beta int) . Here we show that IL-4-producing CD4+ NK1.1+ TCR alpha/beta int liver lymphocytes are down-regulated early in listeriosis . We assume that curtailment of IL-4-producing CD4+ NK1.1+ TCR alpha/beta int liver lymphocytes promotes unconstrained development of Th1 cells which are central to protection against intracellular bacteria. Rev Esp Enferm Dig, 1995 Dec, 87(12), 889 - 92 {Spontaneous bacterial peritonitis caused by Listeria monocytogenes}; Perez Roldan F et al.; Listeria monocytogenes is a gram-positive coccobacillus that produces infections in both the normal and the compromised host . Symptomatic bacteremia and pulmonary infection or meningitis are the most common clinical presentations in adults . According to a current review of the literature, Listeria is a rare bacteria that may produce spontaneous bacterial peritonitis (23 cases reported) . Listeria peritonitis occurs in more than two-thirds of the cases in patients with chronic liver disease, but may also occur in patients with malignancy or undergoing peritoneal dialysis . We describe two cases of SBP in cirrhotic patients, one with alcoholic cirrhosis and other due to HCV infection . One patient also presented with acute meningitis . Peritonitis due to Listeria was clinically and analytically similar to any SBP . Third-generation cephalosporins commonly used in the therapy of SBP, are ineffective in this infection . Ampicillin is the drug of choice, although it should be used in combination therapy usually with an aminoglycoside . The mortality from Listeria peritonitis is similar to that of other SBP (17%). Epidemiol Infect, 1995 Dec, 115(3), 519 - 26 Occurrence of Listeria species in ready to eat foods; Wilson IG; Over 8000 ready to eat foods were examined for the presence of Listeria species . Overall, 5% of foods were found to contain these organisms . Higher occurrence was found in some foods such as chicken (11%) and fish (14%) . Most of the Listeria species isolated were L . monocytogenes (49%) and L . innocua (36%) with lower numbers of other species . No seasonal pattern in the recovery of L . monocytogenes was found . Unsatisfactory or potentially hazardous levels of L . monocytogenes were found in 14 products (< 0.2%), mostly cooked meats . Undercooked chicken products appeared to present the greatest risk for the duration of this survey . The small number of samples which were potentially hazardous suggests that the risk to consumers is not high, and this is confirmed by the absence of clinical cases in the region during the period of study. Appl Environ Microbiol, 1995 Dec, 61(12), 4310 - 4 Differentiation of epidemic-associated strains of Listeria monocytogenes by restriction fragment length polymorphism in a gene region essential for growth at low temperatures (4 degrees C); Zheng W et al.; The growth of Listeria monocytogenes in food stored in the cold has often been implicated in outbreaks of listeriosis . Many subtyping schemes have suggested that epidemic-associated strains belong to a unique genetic group . It has not yet been possible, however, to identify molecular or bacteriologic markers unique to epidemic-associated strains . Recently we cloned three genes of L . monocytogenes, ltrA, ltrB, and ltrC, which are essential for growth at low temperatures (4 degrees C) . The use of a 1.2-kb PstI fragment derived from ltrB as a probe in Southern blots of HindIII-digested DNA revealed three hybridization patterns: the first (a 5.0-kb band) was observed in strains of serotypes 4b, 1/2b, and 3b; the second (a 3.1-kb band) was seen in strains of serotypes 1/2a, 3a, 1/2c, and 3c; and the third (a 9.5-kb band) was characteristic of epidemic-associated serotype 4b strains . These and other data suggest that probes derived from this gene region that is essential for growth at low temperatures can be useful molecular tools for the subtyping of strains implicated in food-borne listeriosis. J Immunol, 1995 Dec 1, 155(11), 5227 - 33 Listeriolysin is processed efficiently into an MHC class I-associated epitope in Listeria monocytogenes-infected cells; Villanueva MS et al.; Listeria monocytogenes is an intracellular pathogen that enters the cytoplasm of infected cells by secreting listeriolysin (LLO), a protein that destroys the phagosomal membrane . In infected mice, LLO is a major Ag detected by protective, MHC class I-restricted CTLs . Although the role of LLO in pathogenesis and host immunity is well established, its rate of intracellular synthesis has yet to be determined . Herein we show that cytosolic L . monocytogenes secrete LLO at a relatively low rate of approximately one molecule per bacterium per minute . Under extracellular labeling conditions, the rate of LLO secretion is approximately 50-fold higher . Intracellular LLO synthesis suffices, however, for the accumulation of 600 to 1000 H-2Kd-associated LLO 91-99 epitopes per cell . We calculate that between four and 11 LLO molecules are degraded for each LLO 91-99 epitope bound by H-2Kd . Our findings indicate that the antigenicity of LLO, with respect to MHC class I-restricted CTLs, cannot be attributed to high levels of intracellular secretion . Rather, LLO is a dominant Ag because it is rapidly degraded and very efficiently processed into an MHC class I-associated epitope. Infect Immun, 1995 Dec, 63(12), 4595 - 9 Listeria monocytogenes can grow in macrophages without the aid of proteins induced by environmental stresses; Hanawa T et al.; Listeria monocytogenes is a facultative intracellular pathogen which is able to survive and grow within phagocytic cells . Some facultative intracellular bacteria have been shown to respond to the hostile environment within phagocytic cells by producing a set of stress proteins . Since L . monocytogenes has a mechanism for intracellular survival that is distinct from those of other bacteria, we studied the phenotypic response of the bacterium to phagocytosis by macrophages . After phagocytosis of L . monocytogenes EGD by J774-1 macrophage cells, the microorganism rapidly increased in numbers about 20-fold during an incubation period of 5 h . In this phase of phagocytosis, the selective induction of 32 proteins was observed by two-dimensional gel electrophoresis . The responses to the environmental stresses of heat and hydrogen peroxide were also studied, and it was found that 14 heat shock proteins and 13 oxidative stress proteins were induced . Five of the induced proteins were common to both heat and oxidative stresses . By amino acid sequencing analysis, homologs of DnaK and GroEL were confirmed among the heat shock proteins . A comparison of the autoradiograms of the two-dimensional gels revealed that none of these stress proteins were among the proteins induced by L . monocytogenes within the macrophages . This behavior is entirely different from that shown by other facultative intracellular pathogens . Stress proteins known to be induced by environmental stresses were absent in intracellularly grown L . monocytogenes in the present study . This absence could be due to the mechanism by which the microorganisms rapidly escape from this stressful environment at a very early phase of phagocytosis. J Exp Med, 1995 Dec 1, 182(6), 1751 - 7 Identification of a CD4+ T cell-stimulating antigen of pathogenic bacteria by expression cloning; Sanderson S et al.; Identifying the immunogenic proteins that elicit pathogen-specific T cell responses is key to rational vaccine design . While several approaches have succeeded in identifying major histocompatibility complex (MHC) class I bound peptides that stimulate CD8+ T cells, these approaches have been difficult to extend to peptides presented by MHC class II molecules that stimulate CD4+ T cells . We describe here a novel strategy for identifying CD4+ T cell-stimulating antigen genes . Using Listeria monocytogenes-specific, lacZ-inducible T cells as single-cell probes, we screened a Listeria monocytogenes genomic library as recombinant Escherichia coli that were fed to macrophages . The antigen gene was isolated from the E . coli clone that, when ingested by the macrophages, allowed generation of the appropriate peptide/MHC class II complex and T cell activation . We show that the antigenic peptide is derived from a previously unknown listeria gene product with characteristics of a membrane-bound protein. J Immunol, 1995 Nov 15, 155(10), 4838 - 43 High level monocyte chemoattractant protein-1 expression in transgenic mice increases their susceptibility to intracellular pathogens; Rutledge BJ et al.; We have constructed transgenic mice in which the mouse mammary tumor virus long terminal repeat controls the expression of murine monocyte chemoattractant protein-1 (MCP-1) . Several independently derived lines of transgenic mice constitutively expressed MCP-1 protein in a variety of organs . Protein extracts from these organs had substantial in vitro monocyte chemoattractant activity that was neutralized by an anti-MCP-1 Ab, indicating that transgenic MCP-1 protein is biologically active . However, no transgenic mouse at any age displayed monocyte infiltrates in MCP-1-expressing organs . Two transgenic lines had circulating MCP-1 levels of 13 to 26 ng/ml, which is a concentration sufficient to induce maximal monocyte chemotaxis in vitro . These transgenic lines showed a 1 to 1.5 log greater sensitivity to infection with Listeria monocytogenes and Mycobacterium tuberculosis . A third transgenic line had lower serum levels of MCP-1 and was resistant to L . monocytogenes . The results suggest that this transgenic model is one of monocyte nonresponsiveness to locally produced MCP-1 due to either receptor desensitization or neutralization of a chemoattractant gradient by high systemic concentrations of MCP-1 . Regardless of the mechanism, the data indicate that constitutively high levels of MCP-1 expression do not induce monocytic infiltrates, and that MCP-1 is involved in the host response to intracellular pathogens. J Immunol, 1995 Nov 15, 155(10), 4817 - 28 Nonviable bacterial antigens administered with IL-12 generate antigen-specific T cell responses and protective immunity against Listeria monocytogenes; Miller MA et al.; The development of effective vaccine strategies for intracellular pathogens, including bacteria, viruses, and parasites, is one of the major frontiers of scientific research . For the studies described here, the murine model of Listeria infection was used to evaluate the adjuvant effects of IL-12 when used as an immunization component . These studies revealed that typically nonimmunogenic doses of heat-killed Listeria monocytogenes, or soluble listerial Ag preparations, elicit intense Th1-type Listeria-specific T cell responses when administered i.p . along with recombinant murine IL-12 . In addition to the Ag-specific production of IL-2 by CD4+ peritoneal cells that was elicited, several other correlates of protective responses were noted, including dramatic induction of CD3+ and alpha beta TCR+ cell populations in the peritoneal cavity and increased expression of class II MHC and production of IL-12 (upon in vitro restimulation) by peritoneal macrophages . Protection studies demonstrated that the T cell responses elicited by a IL-12-potentiated, heat-killed L . monocytogenes vaccine were sufficient to effectively protect mice against challenge with a large dose of virulent Listeria. J Immunol, 1995 Nov 15, 155(10), 4775 - 82 Induction of cell-mediated immune responses to human immunodeficiency virus type 1 Gag protein by using Listeria monocytogenes as a live vaccine vector; Frankel FR et al.; Cytolytic T cells, acting through cytokines or by direct lysis of infected target cells, have been shown to play a significant role in the control of viral infections and may be responsible for the prolonged asymptomatic phase following infection by HIV . Accordingly, methods that can generate strong cell-mediated immune responses may be useful in the development of prophylactic and therapeutic vaccines against HIV . Listeria monocytogenes is a Gram-positive intracellular microorganism that elicits strong cell-mediated immune responses against its own secreted proteins following infection . In this study we have modified the chromosome of L . monocytogenes so that it stably expresses and secretes the p55 HIV gag gene product and examined the cell-mediated immune response of BALB/c mice to infection with this recombinant organism . Infected animals were found to mount a specific, strong, long-lasting CD8+ cytolytic T cell response against a predominant epitope contained within the p24 fragment of the HIV Gag protein . This epitope previously has been shown to be recognized by CTLs obtained from some HIV-infected humans . Our results suggest that chromosomally modified strains of L . monocytogenes may provide valuable vaccine vectors for use against HIV. J Am Vet Med Assoc, 1995 Nov 15, 207(10), 1325 - 6 Listeria monocytogenes septicemia in a foal; Wallace SS et al.; Listeria monocytogenes is rarely reported as a cause of septicemia in foals . In this case, the foal had diarrhea 2 weeks prior to the onset of signs of lethargy, high rectal temperature, and leukopenia with a left shift . Listeria monocytogenes was isolated from the blood culture . The most commonly isolated organism causing septicemia in foals is Escherichia coli . Without the blood cultures, a definitive diagnosis would not have been possible. Mol Microbiol, 1995 Nov, 18(3), 425 - 36 The amino-terminal part of ActA is critical for the actin-based motility of Listeria monocytogenes; the central proline-rich region acts as a stimulator; Lasa I et al.; The intracellular bacterial pathogen Listeria monocytogenes moves inside the host-cell cytoplasm propelled by continuous actin assembly at one pole of the bacterium . This process requires expression of the bacterial surface protein ActA . Recently, in order to identify the regions of ActA which are required for actin assembly, we and others have expressed different domains of ActA by transfection in eukaryotic cells . As this type of approach cannot address the role of ActA in the actin-driven bacterial propulsion, we have now generated several L . monocytogenes strains expressing different domains of ActA and analysed the ability of the different domains to trigger actin assembly and bacterial movement in both infected cells and cytoplasmic extracts . We show here that the amino-terminal part is critical for F-actin assembly and movement . The internal proline-rich repeats a