Listeriosis

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 and the carboxy-terminal domains are not essential . However, in vitro motility assays have demonstrated that mutants lacking the proline-rich repeats domain of ActA moved two times slower (6+/-2 micrometers min(-1)) than the wild type (13 +/-3 micrometers min(-1)) . In addition, phosphatase treatment of protein extracts of cells infected with the L . monocytogenes strains expressing the ActA variants suggested that phosphorylation may not be essential for ActA activity.

Bull Acad Natl Med, 1995 Nov, 179(8), 1613 - 24
{Listeriosis 1985-1995: microbiologic and epidemiologic aspects}; Rocourt J; Listeriosis is an emerging foodborne infection caused by L . monocytogenes (L . m.), mainly identified in industrialized countries . It is a severe disease (meningitis, septicaemia, abortion) which preferentially affects individuals whose immune system is perturbed (pregnant women, newborns, immunocompromized patients and the elderly) . Epidemiology is characterized by a background of sporadic cases on which may be surimposed outbreaks . Progresses in microbiology during the last decade (detection and typing of L . m., better understanding of L.m . ecology...) and epidemiological investigations (increased use of case-control studies) demonstrated that all kind of foods, at each step of the food chain, can transmit the disease . In many respects, L.m . differs from most recognized foodborne pathogens: it is ubiquitously present in nature, resistant to various kind of environments, microaerophilic and psychrophilic . Its tenacity in industrial environment and its capability to survive in food over extended period of time under adverse conditions made this bacterium the hottest topic for industrials during the last decade.

Minerva Med, 1995 Nov, 86(11), 499 - 502
{Listeria monocytogenes meningitis . Description of a clinically significant case}; Sbalzarini G et al.; Meningitis caused by Listeria monocytogenes is a rare affection: it develops from close contagion as professional illness (veterinarians, butchers) or in newborns by infected mothers; in indirect way for ingestion of contaminated food in subjects at high risk: elderly, immunosuppressed patients, alcoholics, diabetics . Clinically it is not diversified from the other bacterial meningitises . In this paper we present a case of Listeria monocytogenes meningitis in an adult female, without a sure occasion of infection and in absence of the factors of typical risk.

Kekkaku, 1995 Nov, 70(11), 651 - 7
{The mechanism of induction and expression of protective immunity with special reference to the role of cytokines}; Mitsuyama M; The mechanism of induction of protective T cells mediating the anti-tuberculous immunity is not well elucidated . Using viable and killed cells of Mycobacterium bovis BCG and Listeria monocytogenes, which differ in the ability to induce specific protection in mice, we have found that the difference is mainly due to the different ability to induce several cytokines especially IFN-rho not due to the difference in antigenic property . It was shown that the induction of protective T cells is highly dependent upon various cytokines produced by the host at the early stage of immunization . Based on our own experimental result in mice, the relative contribution of several cytokines to the induction and expression of cell-mediated immune protection was discussed.

Clin Infect Dis, 1995 Nov, 21(5), 1289 - 90
Listeriosis in bone marrow transplant recipients: incidence, clinical features, and treatment; Chang J et al.; Cultures of blood and/or cerebrospinal fluid from four of 1,013 bone marrow transplant recipients treated at our center between January 1972 and April 1994 were positive for Listeria monocytogenes . The overall occurrence of listeriosis was 0.39 case per 100 transplantations . Allograft recipients had received prior treatment with parenteral methylprednisolone, thus supporting an association between listeriosis and corticosteroids . Treatment with parenteral ampicillin (200 mg/{kg.d}) and gentamicin is recommended for a minimum of 3 weeks before oral therapy . Two patients with penicillin allergies in this study failed to respond to chloramphenicol-based therapeutic regimens . Recurrent meningitis occurred in two patients, and the therapeutic use of intrathecal gentamicin/vancomycin did not confer a survival advantage (i.e., the patients did not survive).

FEMS Microbiol Lett, 1995 Nov 1, 133(1-2), 77 - 83
An iron-dependent mutant of Listeria monocytogenes of attenuated virulence; Rouquette C et al.; A bank of Tn917-insertional mutants from the facultative intracellular pathogen Listeria monocytogenes was screened by an original method based on bacterial growth on synthetic medium under iron-limiting conditions . One mutant, whose in vitro growth in synthetic medium was specifically dependent upon the availability of iron in its environment, was isolated and characterized . The insertional event occurred in a non-coding region, upstream of a rrn operon and located within a 1100-kb NotI fragment of the physical map, where the virulence genes already identified in L . monocytogenes were also present . Protein analysis by SDS-PAGE revealed a pleiotropic effect of the insertional event on cell-associated proteins, suggesting a polar effect of the transposon on adjacent unknown gene(s) . The virulence in the mouse of this mutant was strongly impaired, although it was capable in vitro of growing intracellularly and of spreading from cell to cell, as shown by the production of lytic plaques on cell culture.

J Neurol Neurosurg Psychiatry, 1995 Nov, 59(5), 524 - 7
Serial MRI in listeria mesenrhombencephalitis: a case report; Lever N et al.; Listeria mesenrhombencephalitis is a rare disorder, especially in the non-immunocompromised host . The predilection for brain stem involvement is unexplained . The anatomical correlation of brain stem signs with serial MRI over three months is shown in a 64 year old white man . The pathological lesions seemed to be a combination of micro-abscesses and associated oedema.

Appl Environ Microbiol, 1995 Nov, 61(11), 3872 - 4
Division of Listeria monocytogenes serovar 4b strains into two groups by PCR and restriction enzyme analysis; Ericsson H et al.; Altogether, 133 strains of Listeria monocytogenes serovar 4b were investigated . A segment of 2,916 bp containing parts of the two genes inlA and inlB in L . monocytogenes was amplified by the PCR technique . The PCR product obtained was cleaved with the restriction enzyme AluI, and the fragments generated were separated by gel electrophoresis, leading to two distinct groups: PCR-restriction enzyme analysis groups I and II, containing 37 and 96 strains, respectively . The PCR-restriction enzyme analysis method described in this paper could be a useful tool for the subtyping of L . monocytogenes serovar 4b strains.

J Leukoc Biol, 1995 Nov, 58(5), 556 - 62
Differential regulation of TNF-alpha production by listeriolysin-producing versus nonproducing strains of Listeria monocytogenes; Vazquez MA et al.; Only Listeria monocytogenes that produce listeriolysin O (LLO) elicit protective immunity . Given the importance of tumor necrosis factor alpha (TNF-alpha) in anti-Listeria immunity, we have investigated TNF-alpha production by macrophages after they ingested live LLO-producing compared to LLO-non-producing bacteria . We used two genetically engineered strains of Listeria that differed only in their ability (Ly+) or inability (Ly-) to produce LLO . Ly+ and Ly- caused the same kinetics of increased mRNA abundance for TNF-alpha during the first 90 min after phagocytosis . However, only Ly+ caused sustained transcription of TNF-alpha mRNA, and this may account for the increased release of TNF-alpha . The transcriptional inhibitor 5,6-dichloro-1-beta-D-ribofuranosylbenzimidazole (DRB) prevented the sustained abundance of cytokine mRNA 20 h after ingestion of Ly+ . In addition, nuclear run-on assays indicated sustained transcription of TNF-alpha genes only after ingestion of Ly+ . LLO itself was not responsible for the ability of Ly+ to stimulate the sustained transcription of the TNF-alpha genes . Instead, LLO may allow Listeria to survive within macrophages so that other bacterial products cause sustained TNF-alpha gene transcription . Both Ly+ and Ly- produced molecules, isolated by 50% ammonium sulfate, that induced cytokine production . In conclusion, we now report that Ly+ causes sustained transcription of the TNF-alpha gene and production of TNF-alpha by macrophages in vitro . We speculate that the TNF-alpha may activate endothelium and thus allow the recruitment of T cells to sites of infection . This may contribute to the ability of only LLO-producing Listeria to induce protective immunity.

J Immunol, 1995 Nov 1, 155(9), 4367 - 75
Experimental Listeria meningoencephalitis . Macrophage inflammatory protein-1 alpha and -2 are produced intrathecally and mediate chemotactic activity in cerebrospinal fluid of infected mice; Seebach J et al.; In bacterial meningitis, the recruitment of leukocytes across the blood-brain barrier into the central nervous system may be crucial for both elimination of pathogens and tissue injury . In addition to bacterial cell wall products, host factors including chemokines may lead to accumulation of phagocytes within the central nervous system . As shown by Northern analysis, brains of mice infected intracerebrally with Listeria monocytogenes (LM) express mRNA for three chemokines, the macrophage inflammatory protein (MIP)-1 alpha, MIP-1 beta, and MIP-2 . The cellular sources of these chemokines comprise both the blood-derived polymorphonuclear leukocytes (PMNs) and monocytes infiltrating the meninges, the ventricular system and the periventricular area . In the course of meningitis a time-dependent increase of MIP-1 alpha and MIP-2 was found in the cerebrospinal fluid (CSF) by ELISA . CSF taken 24 h after infection (CSF-LM24) induced migration of human leukocytes when treated in chemotactic chambers in vitro . Neutralizing Abs to chemokines identified MIP-1 alpha and MIP-2 to be responsible for CSF-LM24 mediated chemotaxis of monocytes and PMNs, respectively . CSF obtained from mock-infected animals contained no MIP-1 alpha or MIP-2 and did not lead to migration of leukocytes . When testing CSF-LM24 on mouse spleen cells, the chemotactic activity detected for mononuclear cells was only partly inhibited by Abs to MIP-1 alpha and -1 beta . Thus, in addition to MIP-1 and -2 other not yet defined chemotactic factors are of importance for recruitment of leukocytes in bacterial meningitis.

J Immunol, 1995 Nov 1, 155(9), 4355 - 66
Mechanisms of T cell epitope immunodominance analyzed in murine listeriosis; Safley SA et al.; We demonstrate conclusively that the bacterial exotoxin listeriolysin O (LLO) is a target Ag for eliciting CD4+ T cell responses following infection with Listeria monocytogenes . The minimal I-Ek-restricted immunodominant CD4+ T cell epitope was identified as peptide 215-226 (p215-226) . Most LLO-specific T cell hybridomas recognized p203-226, p208-226, p215-226, and p215-234, although each exhibited a characteristic pattern of preferential reactivity . One hybridoma (IIIC5) reacted to p203-226 but not to p208-226 or any other LLO peptide tested . With APCs from B10 congenic mice and cells transfected with either I-Ak or I-Ek, IIIC5 recognized p203-216 with I-Ak, while a different hybridoma (IB5) recognized p215-226 with I-Ek . Competitive binding studies demonstrated that of 15 LLO peptides tested, only p203-226, p215-226, and p215-234 had high affinity for I-Ek, while p203-226 could also bind to isolated I-Ak . Of nine LLO peptides tested, only p215-234 bound multiple class II MHC alleles . These findings suggest that the immunodominance of p203-226 may be due in part to the presence of multiple T cell epitopes with I-Ek- and I-Ak-binding capability . Many of the rules of immunodominance observed with model Ags are also operative in our murine model of bacterial infectious disease . Furthermore, a novel mechanism of immunodominance based on newly defined structural features of MHC molecules is implicated . This information is crucial for rational vaccine development.

J Immunol, 1995 Nov 1, 155(9), 4347 - 54
IL-1 beta is required for IL-12 to induce production of IFN-gamma by NK cells . A role for IL-1 beta in the T cell-independent mechanism of resistance against intracellular pathogens; Hunter CA et al.; Mice with the severe combined immunodeficiency (SCID) possess an IFN-gamma-dependent mechanism of resistance to the intracellular pathogens Toxoplasma gondii and Listeria monocytogenes that is dependent on IL-12-induced production of IFN-gamma by NK cells . In this report we demonstrate that IL-1 beta is required for IL-12 to stimulate production of IFN-gamma by NK cells, and that IL-1 is important in IL-12-mediated resistance to T . gondii in vivo . Stimulation of SCID mouse splenocytes with tachyzoites of T . gondii resulted in production of IFN-gamma . Addition of neutralizing Ab specific for IL-1 beta to these cultures inhibited completely the production of IFN-gamma . Similar results were obtained when LPS or L . monocytogenes were used to stimulate production of IFN-gamma by SCID mouse splenocytes . Addition of a neutralizing Ab to IL-1 alpha did not affect production of IFN-gamma by SCID mouse splenocytes stimulated with T . gondii, L . monocytogenes, or LPS . Stimulation of SCID mouse splenocytes with IL-1 beta or IL-1 alpha did not result in production of IFN-gamma but enhanced remarkably the ability of T . gondii or IL-12 to stimulate production of IFN-gamma . Furthermore, production of IFN-gamma by SCID mouse splenocytes stimulated with IL-12 plus TNF-alpha was completely ablated by anti-IL-1 beta, but not by anti-IL-1 alpha . Analysis of the culture supernatants of spleen cells from SCID mice stimulated with T . gondii or IL-12 plus TNF-alpha detected low levels of IL-1 beta; addition of a neutralizing Ab to IFN-gamma resulted in a 5- to 10-fold increase in levels of IL-1 beta . Furthermore, stimulation of SCID mouse splenocytes with IL-12, in the presence of anti-IFN-gamma, resulted in an increase in detectable levels of IL-1 beta . To determine the in vivo relevance of our in vitro data, SCID mice were infected with T . gondii and treated with IL-12 alone or IL-12 in combination with an Ab specific for the type I IL-1 receptor . This Ab reduced production of IFN-gamma by SCID mouse splenocytes stimulated with either T . gondii, LPS, L . monocytogenes, or IL-12 plus IL-1 beta . In vivo administration of this Ab antagonized significantly the ability of exogenous IL-12 to delay the time to death of SCID mice infected with T . gondii.(ABSTRACT TRUNCATED AT 400 WORDS)

J Bacteriol, 1995 Nov, 177(22), 6601 - 9
Organization and transcriptional analysis of the Listeria phage A511 late gene region comprising the major capsid and tail sheath protein genes cps and tsh; Loessner MJ et al.; A511 is a broad-host-range, virulent myovirus for Listeria monocytogenes . The genes encoding major structural proteins of the capsid (cps) and tail sheath (tsh) were mapped to a 10.15-kb late gene fragment . We have determined the complete nucleotide sequence of this region and confirmed the identities of Cps (48.7 kDa) and Tsh (61.3 kDa) by N-terminal amino acid sequencing of both proteins . In addition, nine other open reading frames were identified . On the basis of amino acid sequence homologies to known phage-encoded proteins, some putative functions and locations could be assigned to some of the deduced gene products . We present evidence that the cps product is proteolytically cleaved between Lys-23 and Ser-24 to yield the 444-residue polypeptide found in the mature viral capsid . We also found that the N-terminal methionine is absent from the mature tail sheath protein . cps and tsh are late genes; mRNAs first appear 15 to 20 min after infection of L . monocytogenes . Northern (RNA) hybridizations of total late mRNA with specific oligonucleotide probes were used to determine the sizes of respective transcripts . Primer extension analyses enabled the positive identification of six late promoters, which were found to differ from those identified in the chromosome of Listeria spp . The bulk of transcripts from cps and tsh arise from two phage promoters with identical 13-nucleotide sequences (TGCTAGATTATAG {core region underlined}) in the -10 region which we speculate determines specific and timed expression of these genes . A 123-nucleotide leader sequence at the 5' end of the cps transcript was predicted to form a strong secondary structure (deltaG=-40.7 kcal {-170.3 kJ}/mol) . Out results show that the strongly expressed A511 cps and tsh genes are included in two separate gene clusters and are independently regulated at the transcriptional level.

Infect Immun, 1995 Nov, 63(11), 4531 - 4
The broad-range phospholipase C and a metalloprotease mediate listeriolysin O-independent escape of Listeria monocytogenes from a primary vacuole in human epithelial cells; Marquis H et al.; Intracellular growth of Listeria monocytogenes begins after lysis of the primary vacuole formed upon bacterial entry into a host cell . Listeriolysin O (LLO), a pore-forming hemolysin encoded by hly, is essential for vacuolar lysis in most cell types . However, in human epithelial cells, LLO- mutants are capable of growth, suggesting that gene products other than LLO are capable of mediating escape from a vacuole . In this study, we investigated the role of other bacterial gene products in lysis of the primary vacuole in the human epithelial cell line Henle 407 . Double internal in-frame deletion mutants were constructed by introducing a mutated hly allele into strains harboring deletions in either of the phospholipase C (PLC)-encoding genes or a metalloprotease-encoding gene . Bacterial escape from the primary vacuole, intracellular growth, and cell-to-cell spread were evaluated in Henle 407 cells . The results indicated that, in the absence of LLO, the broad-range PLC and the metalloprotease were both required for lysis of the primary vacuole in Henle 407 cells . Although phosphatidylinositol-specific PLC was not required, the efficiency of escape was reduced in an LLO phosphatidylinositol-specific PLC double mutant . These observations suggest that the relative importance of LLO, the phospholipases, and the metalloprotease may vary in different cell types or in cells from different species . In addition, these studies provide insight into the mechanisms of action of virulence determinants involved in the lysis of vacuolar membranes.

Infect Immun, 1995 Nov, 63(11), 4268 - 76
Listeria monocytogenes infects human endothelial cells by two distinct mechanisms; Drevets DA et al.; Infection of endothelial cells by bacteria may be an important component of the bacteria's ability to escape host defenses and cause disease . Listeria monocytogenes cause sepsis and central nervous system infection in domesticated animals and immunocompromised humans, suggesting that this bacterium interacts with endothelial cells in a significant fashion . The experiments presented here tested the hypothesis that L . monocytogenes can invade and replicate within human endothelial cells . We found that L . monocytogenes grows readily in umbilical vein endothelial cells and that its intracellular life cycle involves phagosomal escape, F-actin-based motility, and cell-to-cell spread . We found that L . monocytogenes invaded endothelial cells by cell-to-cell spread from adherent mononuclear phagocytes which were previously infected by this bacterium . Interestingly, L . monocytogenes mutants lacking the invasion protein, internalin, bound less well to endothelial cells than did wild-type bacteria in the absence, but not the presence, of serum, and their invasion of endothelial cells was diminished under both conditions . Thus, endothelial cell infection by L . monocytogenes can occur by two distinct mechanisms: direct bacterial invasion of the endothelial cells in an internalin-mediated fashion or cell-to-cell spread from adherent, infected mononuclear phagocytes . These data support the idea that endothelial cell infection by L . monocytogenes is an important event in the pathogenesis of listeriosis.

Infect Immun, 1995 Nov, 63(11), 4231 - 7
The two distinct phospholipases C of Listeria monocytogenes have overlapping roles in escape from a vacuole and cell-to-cell spread; Smith GA et al.; Listeria monocytogenes secretes two distinct phospholipases C, a phosphatidylinositol-specific phospholipase C (PI-PLC) and a broad-range phospholipase C (PC-PLC) . In this study, single in-frame deletion mutants with mutations in each PLC and a double mutant lacking both PLCs were characterized with regard to virulence in mice, escape from a primary vacuole, and cell-to-cell spread in cell culture . The mutant lacking PI-PLC, previously shown to be twofold less virulent than the wild type in mice, had a minor defect in escape from a primary vacuole but was not notably affected in cell-to-cell spread . The mutant lacking PC-PLC was 20-fold less virulent in mice and was defective in cell-to-cell spread but had no measurable defect in escape from a primary vacuole . The mutant lacking both PLCs was 500-fold less virulent in mice and was severely diminished in its ability to escape from the primary vacuole and to spread cell to cell . Cellular levels of diacylglycerol and ceramide, products of PLC activity, accumulated beginning 3 to 4 h after infection of cells with wild-type bacteria . The bacterial PLCs were partially responsible for this activity, since cells infected with the mutant lacking both PLCs had a reduced increase in diacylglycerol and no increase in ceramide . Elevation of diacylglycerol in the absence of bacterial PLCs indicated that host cell phospholipase(s) was activated during infection . The results of this study were consistent with the two bacterial PLCs having overlapping functions throughout the course of intracellular infection . Furthermore, the PC-PLC, and possibly PI-PLC, appeared to be enzymatically active intracellularly.

Cancer Res, 1995 Nov 1, 55(21), 4776 - 9
Regression of established tumors in mice mediated by the oral administration of a recombinant Listeria monocytogenes vaccine; Pan ZK et al.; We have shown previously that Listeria monocytogenes, a gram-positive, facultative intracellular bacterium, is a potent vector for targeting tumor-specific antigens to the immune system . After parenteral administration, we observed protection against both renal and colorectal mouse tumors and regression of established renal tumors . In the present study, we have exploited the fact that the normal route of infection of this organism is through the gut . We show that an L . monocytogenes recombinant that expresses a model tumor antigen is an effective cancer immunotherapeutic agent when delivered orally in that it causes regression of established, macroscopic mouse renal and colorectal tumors expressing the same antigen.

J Comp Pathol, 1995 Oct, 113(3), 263 - 75
Phenotypic characterization of the cells of the inflammatory response in ovine encephalitic listeriosis; Krueger N et al.; The brainstem (pons cerebri and medulla oblongata) of 22 sheep aged between 6 months and 3 years which had developed clinical signs of central nervous system dysfunction were examined . Histopathological changes characterized by microabscesses, focal gliosis and perivascular cuffing compatible with natural infection with Listeria monocytogenes were present . The brains were examined by lectin histochemistry and immunohistochemistry with markers for T lymphocytes (CD4+ and CD8+ subsets), B lymphocytes, mononuclear phagocytes (including macrophages, ramified microglia, activated microglia and amoeboid microglia), astroglia and L . monocytogenes . These methods allowed semiquantitative analyses of the frequency of the different cell types in the brain lesions . The distribution of listerial antigen in the lesions was variable but always sparse . Mononuclear phagocytes and neutrophils appeared to be the most numerous inflammatory cells in the affected areas of the brainstem . T lymphocytes (CD8+ and CD4+ subsets) and B lymphocytes also played a part in the inflammatory process, in addition to activated astrocytes.

FEMS Immunol Med Microbiol, 1995 Oct, 12(2), 143 - 52
Listeria monocytogenes infection in pregnant mice: abnormalities in the function of non-adherent accessory light density dendritic cells; Klink M et al.; Pregnant A/J mice were found to be more susceptible to the lethal effect of Listeria monocytogenes bacteria than virgin females . However, during the first four days of post-infection there was no difference in the elimination of Listeria from the spleens of pregnant and virgin mice . This suggests that the increase in the susceptibility of pregnant mice to pathogenic activity of L . monocytogenes was related to the diminution in Listeria-specific cellular reactions . Indeed, we found that non-adherent light density dendritic cells (DCs) from pregnant mice showed a marked reduction in the ability to form clusters with L . monocytogenes immune T lymphocytes and it is known that cell cluster formation between antigen presenting cells (APC) and responding T cells is required for antigen recognition as well as for cell proliferation . DCs from pregnant mice also demonstrated the decrease and an instability in the expression of H-2 class II molecules which play a crucial role in the recognition of exogenous antigens . The abnormalities demonstrated in the function of the light density dendritic cells from the spleens of pregnant mice could compromise cellular reactions to L . monocytogenes bacteria possibly resulting in increased susceptibility of pregnant mice to experimental listeriosis.

Int J Food Microbiol, 1995 Oct, 27(2-3), 245 - 52
Typing of Listeria monocytogenes by random amplified polymorphic DNA (RAPD) analysis; O'Donoghue K et al.; The aim of the study was to determine the effectiveness of random amplified polymorphic DNA analysis in typing Listeria monocytogenes from human infections . Twenty-five L . monocytogenes serogroup 1/2 and 70 serogroup 4 including 14 serovar 4b(x) were typed by RAPD-PCR analysis . Six primers were used to type each L . monocytogenes isolate and the DNA amplification performed with supertaq DNA polymerase in a Hybaid Thermal Reactor . Each bacterial strain was analysed separately with all primers and the profiles were judged by eye and designated to a group by comparison to other strains . Bands were classified as major or minor . Based on analysis of major band patterns, the 25 serogroup 1/2 isolates gave rise to 12 different groups . The groups only contained serovar 1/2a or 1/2b with a single exception . Using minor bands all isolates could be distinguished . All the serogroup 4 isolates gave the same major band patterns . The 14 serovar 4b(x) isolates which were epidemiologically related gave identical profiles with the exception of one isolate . Of the remaining strains, 41 produced individual patterns on minor band analysis . RAPD analysis with multiple primers is low cost, discriminatory and is most ideally suited to testing small (< 50) numbers of strains . We have shown that serogroup 1/2 L . monocytogenes strains are a more diverse group than serovar 4b strains and RAPD-PCR will provide a technique of considerable value in typing L . monocytogenes in the future.

Int J Food Microbiol, 1995 Oct, 27(2-3), 161 - 74
Genomic subtraction in combination with PCR for enrichment of Listeria monocytogenes-specific sequences; Wu FM et al.; Genomic DNA from Listeria innocua and Listeria ivanovii was used in subtractive hybridization with DNA from Listeria monocytogenes involving two amplification strategies . Subtraction was accomplished by labelling the subtracting DNA with biotin and removal after liquid hybridization with tester DNA (L . monocytogenes) by reaction with streptavidin and phenol extraction . In one strategy, L . monocytogenes DNA was poly(A)-tailed with terminal transferase and amplified asymmetrically after subtraction . In another procedure, adapters ligated to the target DNA allowed symmetrical amplification after subtraction using an adapter-specific primer; in both amplifications, the amplified products were labelled with biotin-modified dUTP . Southern hybridization of the amplified/subtracted probes with tester- and subtractor-related strains demonstrated numerous L . monocytogenes-specific sequences . The genome-subtracted mixed probe identified 7 RFLP patterns among 13 strains of L . monocytogenes representing 11 L . monocytogenes serovars . Southern blot analysis demonstrated that the subtracted probe cross-hybridized to two bands among L . welshimeri strains but had little or no hybridization with five other species of Listeria including L . innocua, L . ivanovii, L . seeligeri, L . grayi, and L . murrayi . These data demonstrate that genomic subtraction via subtractive hybridization is a powerful method to enrich for specie-specific sequences in L . monocytogenes; the enriched sequences in the subtracted probe may be useful for typing L . monocytogenes strains by specific RFLP patterns or for cloning L . monocytogenes-specific sequences.

Med Microbiol Immunol (Berl), 1995 Oct, 184(3), 105 - 8
A simple colony-blot method for identification of Listeria in food samples; Belyi YF et al.; A method for identification of Listeria in food samples was developed . It consisted of cultivating of suspected specimen on standard agar medium, direct absorption of grown colonies onto nitrocellulose membrane and processing of the latter with rabbit serum raised against purified cell wall protein Lm79/39 of L . monocytogenes . Analysis using anti-rabbit peroxidase conjugate and 4-chloro-1-naphthol and H2O2 solutions allowed direct detection of Listeria colonies which remained readily available for subsequent isolation.

Neurologia, 1995 Oct, 10(8), 342 - 5
{Wallenberg syndrome as a sign of rhombencephalitis-meningitis due to Listeria monocytogenes}; Caminero Rodriguez AB et al.; We report a case of rhombencephalitis with meningitis in a 36-years-old previously healthy man; neurological signs and symptoms were initially consistent with a diagnosis of Wallenberg syndrome . Analysis of cerebrospinal fluid showed predominantly lymphocytic pleocytosis and elevated protein levels . A CT brain scan was normal . MRI of the brain showed a hypertensive type lesion in T2, in the right pontomedullary region that suggested inflammation . A blood culture grew Listeria monocytogenes . The patient improved and fully recovered with appropriate antibiotic treatment . Listeria monocytogenes is a recognized cause of acute brainstem meningoencephalitis . Differential diagnoses that must be considered are other forms of purulent meningitis, viral meningoencephalitis, granulomatosis infections of the central nervous system and, occasionally, stroke.

Eur J Immunol, 1995 Oct, 25(10), 2967 - 71
Listeriolysin generates a route for the presentation of exogenous antigens by major histocompatibility complex class I; Darji A et al.; We have exploited the pore forming activity of listeriolysin, the hemolysin of Listeria monocytogenes, to activate CD8+ T cells with soluble proteins in vivo and in vitro . Immunization with soluble, hemolytically active listeriolysin induces both cytotoxic CD8+ T cells and CD4+ T cells, and the CD8+ T cells can be propagated with soluble listeriolysin in vitro . Moreover, conventional antigens like ovalbumin mixed together with listeriolysin are also efficiently introduced into the MHC class I pathway in vitro and in vivo . Hence, listeriolysin effectively directs itself and passenger molecules into the intracellular compartment that leads to the cytotoxic T cell response . In this way, we circumvent the bias of CD8+ T cells to recognize intracellular antigens presented by major histocompatibility complex class I molecules . As cytotoxic CD8+ T cells are of pivotal importance in eliminating viral and microbial pathogens, the findings reported here could prove to be useful in vaccine development.

Microbiology, 1995 Oct, 141 ( Pt 10), 2577 - 84
Characterization of cryptic prophages (monocins) in Listeria and sequence analysis of a holin/endolysin gene; Zink R et al.; Monocins in Listeria were induced by UV-irradiation of liquid cultures, and defective phage particles were purified from the lysates . Electron microscopy showed flexible, non-contractile bacteriophage-tail-like particles, consisting of specific proteins of molecular mass 20-45 kDa and pI 4.6-6.7 . These particles were able to lyse listerial cells . DNA sequence homologies between chromosomal DNA of monocin-producing strains and labelled Listeria phage DNAs were inferred from DNA/DNA hybridizations, suggesting that most of the prophage DNA is still present in the listerial chromosome . An endolysin gene cpl2438 was cloned from listerial chromosomal DNA and was identified by its expression of lytic activity against Listeria cells in a bioassay . The gene consists of 864 nt encoding a protein of 287 aa with a calculated molecular mass of 32975 Da (CPL2438) . This is in good agreement with the size of a protein observed in SDS-PAGE after overexpression of the lytic protein in Escherichia coli . The nucleotide sequence of a putative holin gene (hol2438, 291 nt) upstream of cpl2438 was determined after PCR-amplification of listerial DNA and it shows typical features common to the holin gene family . Expression of the encoded protein (HOL2438, 95 aa, 10.1 kDa) in E . coli was found to be lethal for the host cells . The results underline the close relationship between monocins and intact Listeria bacteriophages, indicating that monocins are incompletely assembled phage particles derived from cryptic prophages of Listeria, probably including the phage lysin.

J Immunol, 1995 Oct 1, 155(7), 3427 - 32
Secondary response to Listeria infection requires IFN-gamma but is partially independent of IL-12; Tripp CS et al.; During a secondary immune response to Listeria monocytogenes (LM), the production of IFN-gamma was still required for resistance, but it was considerably less dependent on IL-12 production . When IL-12 was neutralized in vivo using specific hamster antimurine IL-12 mAbs, there was a dramatically increased susceptibility to infection during primary listeriosis but much less during a secondary infection . However, neutralization of IFN-gamma in vivo resulted in a similar increased susceptibility during both primary and secondary listeriosis . In culture, splenocytes isolated from unimmunized mice produced IFN-gamma in response to heat-killed L . monocytogenes (hk-LM) that was absolutely dependent upon IL-12 production . However, directly stimulating the TCR with anti-CD3-epsilon mAbs resulted in IFN-gamma production that was unaffected by neutralizing IL-12 in vitro . In contrast, splenocytes isolated from LM-immune mice produced IFN-gamma in response to hk-LM, part of which was independent on IL-12 production . However, anti-CD3-epsilon Ab-stimulated IFN-gamma production remained independent of IL-12 production . The source of hk-LM-induced, IL-12-independent IFN-gamma production was the T cell because anti-Thy1.2 Ab plus complement treatment in vitro completely abolished it . Together, these data support a model of memory T cells being produced during the primary infection with LM that can be stimulated to produce IFN-gamma during the secondary response to LM, partially independent of macrophage IL-12 production.

J Cell Biol, 1995 Oct, 131(1), 179 - 89
Myosin is involved in postmitotic cell spreading; Cramer LP et al.; We have investigated a role for myosin in postmitotic Potoroo tridactylis kidney (PtK2) cell spreading by inhibitor studies, time-lapse video microscopy, and immunofluorescence . We have also determined the spatial organization and polarity of actin filaments in postmitotic spreading cells . We show that butanedione monoxime (BDM), a known inhibitor of muscle myosin II, inhibits nonmuscle myosin II and myosin V adenosine triphosphatases . BDM reversibly inhibits PtK2 postmitotic cell spreading . Listeria motility is not affected by this drug . Electron microscopy studies show that some actin filaments in spreading edges are part of actin bundles that are also found in long, thin, structures that are connected to spreading edges and substrate (retraction fibers), and that 90% of this actin is oriented with barbed ends in the direction of spreading . The remaining actin in spreading edges has a more random orientation and spatial arrangement . Myosin II is associated with actin polymer in spreading cell edges, but not retraction fibers . Myosin II is excluded from lamellipodia that protrude from the cell edge at the end of spreading . We suggest that spreading involves myosin, possibly myosin II.

Infect Immun, 1995 Oct, 63(10), 3896 - 903
Expression of the Listeria monocytogenes EGD inlA and inlB genes, whose products mediate bacterial entry into tissue culture cell lines, by PrfA-dependent and -independent mechanisms; Lingnau A et al.; Internalization of Listeria monocytogenes into nonphagocytic cell lines in vitro requires the products of the inlAB locus (J.-L . Gaillard, P . Berche, C . Frehel, E . Gouin, and P . Cossart, Cell 65:1127-1141, 1991) . By generating isogenic mutants with a chromosomal in-frame deletion in either inlA or inlB, we have identified InlA and InlB as surface-bound proteins of L . monocytogenes with molecular weights of 88,000 and 65,000, respectively . These results were obtained with monoclonal antibodies raised against either protein and corroborated by N-terminal end sequencing of InlA and InlB . By immunoblot analysis, the production of both polypeptides was found to be strongly dependent on growth temperature and, particularly for InlB, on the presence of the PrfA regulator protein . Expression of InlA was not strictly dependent on the presence of the PrfA regulator protein . Transcription analysis of the inlAB locus revealed that the inlA gene was transcribed by several promoters, of which only one is PrfA dependent . This PrfA-dependent inlA promoter, which contains two base substitutions within its putative PrfA DNA-binding palindrome, is responsible for transcription of both inlA and inlB genes . A hitherto unrecognized promoter located 51 bp upstream of the GTG start codon of the inlB gene was also detected . Hence, inlA and inlB are transcribed both individually and in an operon by PrfA-dependent and -independent mechanisms . Tissue culture invasion assays employing various epithelial cell lines demonstrated that both InlA and InlB are required for invasion . In vivo studies using the mouse infection model revealed that both internalin mutants were attenuated for virulence.

Immunology, 1995 Oct, 86(2), 256 - 62
Tumour necrosis factor, but not interferon-gamma, is essential for acquired resistance to Listeria monocytogenes during a secondary infection in mice; Samsom JN et al.; Mice with a secondary Listeria monocytogenes infection eliminate the bacteria much faster and more efficiently from their organs than mice with a primary infection . During the course of a secondary infection, serum concentrations of interferon-gamma (IFN-gamma) and tumour necrosis factor-alpha (TNF) are higher than during a primary infection . The aim of the present study was to determine whether these cytokines are involved in the acquired resistance to L . monocytogenes during a secondary infection in mice . In order to neutralize cytokines, alginate-encapsulated cells, which form anti-cytokine monoclonal antibodies, were injected into the nuchal region of mice during a Listeria infection . Mice recovered from a sublethal primary Listeria infection, which acquired cell-mediated immunity, received a subcutaneous injection of anti-IFN-gamma-forming cells, or anti-TNF-forming cells, and 4 days later received an intravenous injection with 10 50% lethal dose (LD50) L . monocytogenes . The number of bacteria recovered from the liver and spleen of immune mice treated with anti-IFN-gamma-forming cells was slightly larger (approximately 1 log10) than that found for immune mice treated with anti-beta-galactosidase-forming cells, called immune control mice . The organs of immune mice treated with anti-TNF-forming cells yielded significantly more (approximately 4 log10) bacteria than those of immune control mice, more than those of immune mice treated with anti-IFN-gamma-forming cells, and comparable numbers to those of non-immune mice . Taken together, these results demonstrate that TNF is essential in acquired resistance to L . monocytogenes during a secondary infection in mice, while IFN-gamma plays a minor role.

Immunology, 1995 Oct, 86(2), 199 - 205
A protective role of gamma delta T cells in primary infection with Listeria monocytogenes in autoimmune non-obese diabetic mice; Usami J et al.; We investigated the host defense mechanism in primary infection with Listeria monocytogenes in non-obese diabetic (NOD) mice at pre-diabetic stage showing an impaired responsiveness of the alpha beta T cells to T-cell receptor (TCR) triggering . The NOD mice showed a deteriorated resistance at the late stage after an intraperitoneal infection with L . monocytogenes compared with BALB/c and C57BL/6 mice as assessed by bacterial growth in organs . Consistent with our previous findings, a prominent increase in number of gamma delta T cells was evident at the early stage after infection, while generation of Listeria-specific alpha beta T cells was impaired in these mice . In vivo administration of anti-TCR gamma delta monoclonal antibody (mAb) allowed L . monocytogenes to grow exaggeratedly in the NOD mice . These results imply that gamma delta T cells may be mainly involved in protection against primary infection with L . monocytogenes in NOD mice.

Immunology, 1995 Oct, 86(2), 190 - 8
The induction of renal autoantigen-specific T cells by a local Listeria monocytogenes infection; Sonoda K et al.; In order to examine the possibility that a local chronic infection can induce organ-specific autoimmune disease, we inoculated unilateral kidneys with viable Listeria monocytogenes (intrarenal infection) . The delayed footpad reaction against syngeneic kidney homogenate (KH) became positive from 1 week after initiating the intrarenal infection . A proliferative response of the spleen T cells from the infected mice was also observed against KH from 1 week after initiating the intrarenal infection, but no such response was seen against liver homogenate (LH) . In contrast, an intravenous Listeria infection did not induce a delayed footpad reaction or proliferative response against KH, suggesting that these autoimmune responses were not caused by molecular mimicry between renal antigens and Listeria antigens . Furthermore, the ability to transfer the autoimmune response of spleen T cells from intrarenally infected mice was examined . The transferred mice showed a positive delayed footpad reaction against KH and an interstitial infiltration of mononuclear cells in their kidneys . These results demonstrate that the intrarenal Listeria infection induced renal autoantigen-specific T cells, which subsequently induced an autoimmune interstitial nephritis (AIN) . The autoreactive T cells were all induced without immunization by autoantigens mixed with complete Freund's adjuvant . Based on these findings, we propose that a local bacterial infection may induce an autoimmune response against autoantigens in the infected organ and subsequently trigger organ-specific autoimmune disease.

Appl Environ Microbiol, 1995 Oct, 61(10), 3745 - 7
A new method for direct detection of Listeria monocytogenes from foods by PCR; Makino S et al.; The preparation of DNA by alcohol precipitation in the presence of Na1 was used to enable the direct detection of Listeria monocytogenes in the amount of 10(3) CFU/0.5 g of sample . This procedure produces PCR-quality DNA directly from foods, such as soft cheese and minced meat.

Appl Environ Microbiol, 1995 Oct, 61(10), 3724 - 8
Use of a fluorogenic probe in a PCR-based assay for the detection of Listeria monocytogenes; Bassler HA et al.; A PCR-based assay for Listeria monocytogenes that uses the hydrolysis of an internal fluorogenic probe to monitor the amplification of the target has been formatted . The fluorogenic 5' nuclease PCR assay takes advantage of the endogenous 5' --> 3' nuclease activity of Taq DNA polymerase to digest a probe which is labelled with two fluorescent dyes and hybridizes to the amplicon during PCR . When the probe is intact, the two fluorophores interact such that the emission of the reporter dye is quenched . During amplification, the probe is hydrolyzed, relieving the quenching of the reporter and resulting in an increase in its fluorescence intensity . This change in reporter dye fluorescence is quantitative for the amount of PCR product and, under appropriate conditions, for the amount of template . We have applied the fluorogenic 5' nuclease PCR assay to detect L . monocytogenes, using an 858-bp amplicon of hemolysin (hlyA) as the target . Maximum sensitivity was achieved by evaluating various fluorogenic probes and then optimizing the assay components and cycling parameters . With crude cell lysates, the total assay could be completed in 3 h with a detection limit of approximately 50 CFU . Quantification was linear over a range of 5 x 10(1) to 5 x 10(5) CFU.

J Antimicrob Chemother, 1995 Sep, 36(3), 527 - 30
Activity of CI-960 alone and in combination with amoxycillin against Listeria monocytogenes, and comparison with other quinolones; Boisivon A et al.; In-vitro activity of CI-960 against seven strains of Listeria monocytogenes was compared to that of other fluoroquinolones . MICs/MBCs were: ciprofloxacin and sparfloxacin: 0.5-2/1-4 mg/L;PD 131628: 0.125-0.5/0.25-2 mg/L; CI-960: 0.06-0.25/0.25-0.5 mg/L . At 4 h, CI-960 (4 x MIC) reduced the inoculum (log10 cfu) by 2.88 +/- 0.61 compared with 1.78 +/- 0.48 for ciprofloxacin, 1.5 +/- 0.46 for sparfloxacin, and 1.35 +/- 0.47 for PD 131628 (P < 0.05) . At 24 h, the bactericidal effects were similar (-4 log10 cfu) . The amoxycillin/CI-960 (8 x MIC) combination was bactericidal, without emergence of resistant mutants.

Int J Food Microbiol, 1995 Sep, 27(1), 77 - 89
Development of NASBA, a nucleic acid amplification system, for identification of Listeria monocytogenes and comparison to ELISA and a modified FDA method; Uyttendaele M et al.; NASBA, an isothermal nucleic acid amplification system was used for identification of Listeria monocytogenes . A primer set and a species-specific probe were selected from the 16S rRNA sequence . The probe was shown to hybridize specifically to the amplified single-stranded RNA of L . monocytogenes . No hybridization occurred with amplification product of L . seeligeri, L . innocua, L . ivanovii, and L . welshimeri . Detection sensitivity for the NASBA assay was determined at 10(6) cfu/ml . The possibility of using the NASBA assay for detection of L . monocytogenes in foods after a 2-day enrichment procedure was explored . NASBA was compared to a modified FDA method and ELISA for detection of L . monocytogenes artificially inoculated (1-100 cfu/25 g) in eight food products . False-negative results were obtained with the modified FDA method (6.75%) . NASBA and ELISA were shown in this study to detect the pathogen with equal efficiency (no false negative or false positive results) . Both methods allowed detection of less than 10 cfu/25 g within 3 days but ELISA can only be used for diagnosis of Listeria spp . while the NASBA procedure permitted specific identification of the human pathogen L . monocytogenes.

EMBO J, 1995 Sep 1, 14(17), 4230 - 9
Granzyme A-deficient mice retain potent cell-mediated cytotoxicity; Ebnet K et al.; Granzyme A, a granule-associated serine proteinase of activated cytotoxic T cells and natural killer cells, has been reported to play a critical role in DNA fragmentation of target cells . To address the question of the biological role of granzyme A, we have now generated a granzyme A-deficient mouse mutant by homologous recombination . Western blot analysis, enzyme assays and reverse transcription-PCR confirmed the absence of granzyme A in activated T cells . In addition, deletion of granzyme A does not alter the expression patterns of other granule components, such as granzymes B-G and perforin . Granzyme A-deficient mice are healthy and show normal hematopoietic development . Most notably, their in vitro- and ex vivo-derived cytotoxic T cells and natural killer cells are indistinguishable from those of normal mice in causing membrane disruption, apoptosis and DNA fragmentation in target cells . Furthermore, granzyme A-deficient mice readily recover from both lymphocytic choriomeningitis virus and Listeria monocytogenes infections and eradicate syngeneic tumors with kinetics similar to the wild-type strain . These results demonstrate that granzyme A does not play a primary role in cell-mediated cytotoxicity, as has been assumed previously.

Microbiology, 1995 Sep, 141 ( Pt 9), 2053 - 61
Listeria monocytogenes exists in at least three evolutionary lines: evidence from flagellin, invasive associated protein and listeriolysin O genes; Rasmussen OF et al.; Regions of the genes encoding flagellin (flaA), the invasive associated protein (iap), listeriolysin O (hly) and 23S rRNA were sequenced for a range of Listeria monocytogenes isolates of different origin and serotypes . Several nucleotide sequence variations were found in the flaA, iap and hly genes . No differences were found for the rRNA genes, but our approach does not exclude the existence of differences between single copies of these genes . Based on the sequence differences, the L . monocytogenes strains can be divided into three distinct sequence types . Further, the presence of only a small number of sequence differences within each group indicates a strong degree of conservation within the groups . There was a complete correspondence among the groups of strains formed according to the analysis of the flaA, iap and hly genes, and the grouping correlates with serotype, pulsed field gel electrophoretic and multilocus enzyme electrophoretic data . Analysis of the region encoding the threonine-asparagine repeat units in the iap gene revealed some striking features . Sequence type 1 strains were found to have 16-17 repeats, sequence type 2 strains had 16-20 repeats whereas the two sequence type 3 strains analysed had only 11 repeats . Furthermore, within a 19 bp segment there was a 37% difference between the sequences of type 1 and 2 strains and that segment was absent in type 3 strains . Within the threonine-asparagine repeat region the nucleotide differences gave rise to four amino acid changes; however, all were changes among the three amino acids present in the repeat structure indicating a strong selective pressure on the composition of this region.

Dtsch Med Wochenschr, 1995 Aug 25, 120(34-35), 1161 - 4
{T-cell lymphoma under immunosuppressive treatment in minimal change glomerulopathy with nephrotic syndrome}; Spath M et al.; HISTORY AND CLINICAL FINDINGS: A 43-year-old woman was admitted for treatment of a nephrotic syndrome with increasing oedema, ascites, dyspnoea and nausea during the preceding 2 weeks . INVESTIGATIONS: There was a proteinuria of 25 g daily . Renal biopsy (seven glomeruli) showed a minimal-change glomerulopathy . TREATMENT AND COURSE: Prednisone administration (1 mg/kg daily) achieved complete remission . In the course of this treatment the patient contracted listeriosis with encephalitis, which quickly responded to ampicillin and netilmicin . When the glucocorticoid dosage was reduced or the drug discontinued, the nephrotic syndrome recurred . Cyclosporin treatment was begun (5 mg/kg daily) and quickly led to another remission . 4 weeks after start of the treatment a reddish-blue nodule, about 2 cm in diameter, was noted on the right upper arm . Histologically it showed cutaneous infiltration of a medium-sized T-cell lymphoma . Other efflorescences were found on the trunk and the limbs . Cyclosporin treatment was at once discontinued and the cutaneous changes gradually regressed . There were no other manifestations of a lymphoma . A further recurrence of the nephrotic syndrome responded well to a short course of chlorambucil (cumulative dose 420 mg) and of prednisone with slow dose reduction . There has been no recurrence in the last year.

J Immunol, 1995 Aug 15, 155(4), 2047 - 56
Bacterial infection of the testis leading to autoaggressive immunity triggers apparently opposed responses of alpha beta and gamma delta T cells; Mukasa A et al.; The mechanisms that lead to the breakdown of self-tolerance and testis-specific immune reactivity in the murine orchitis model are understood only in part . We investigated the histopathologic and immunologic consequences of a unilateral bacterial (Listeria monocytogenes) infection of the testis . Both infected and contralateral sides of this bilateral organ suffered severe inflammatory responses despite a conspicuous absence of bacteria in the contralateral tissue . Also, in both testicles, T cell populations increased, involving both alpha beta and gamma delta T cell subsets . Concomitant with the bilateral orchitis, testis-specific delayed type hypersensitivity and Ab responses developed . Ab depletion experiments indicated that in this orchitis model, as in others, alpha beta T cells are initiators of the autoaggressive reactivity . In contrast, Ab depletion of gamma delta T cells accelerated the inflammatory response in both testicles, suggesting a regulatory role for this type of T cells in both infection-induced and autoimmune orchitis.

J Occup Environ Med, 1995 Aug, 37(8), 962 - 5
Biological agents and pregnancy; Ekblad U; Pregnant women are exposed to many biological, eg microbial, agents, which are potentially harmful to the fetus . The reported rates of vertical transmission of hepatitis B and human immunodeficiency virus vary between 3 to 90% and 0 to 65%, respectively . The susceptibility to hepatitis B and human immunodeficiency infection is increased in pregnant physicians, midwives, and nurses because of the bloodborne nature of these viruses . Also, TORCH (toxoplasmosis-rubella-cytomegalovirus-herpes) infections, acquired during pregnancy, may result in congenital infection, and serious sequelae in the neonatal period or years after birth . Schoolteachers and daycare personnel have an increased risk of perinatal varicella, "fifth disease," and mumps . Perinatal listeriosis affects one in 20,000 births and may result in fetal wastage . Because of the risk of the possibility of vertical transmission, immunization during pregnancy with live virus vaccines is not recommended.

Eur J Immunol, 1995 Aug, 25(8), 2401 - 7
Transgenic mice expressing high levels of soluble TNF-R1 fusion protein are protected from lethal septic shock and cerebral malaria, and are highly sensitive to Listeria monocytogenes and Leishmania major infections; Garcia I et al.; Mice bearing a transgene coding for a soluble tumor necrosis factor receptor type 1 (TNFR1)-FcIgG3 fusion protein and placed under the control of the alpha-1-antitrypsin gene promoter were generated . Depending on the mouse line, blood levels of the protein ranged from 25 ng/ml to over 100 micrograms/ml; this level of expression was most often transmitted to the transgene-bearing progeny as a relatively stable feature . High-expressor mice were completely resistant to lipopolysaccharide-induced shock and lethality, including after D-galactosamine sensitization, and mice expressing about 1 microgram of the fusion protein/ml were partially (60%) protected . In contrast, mice expressing less than 0.1 microgram of the protein/ml were more sensitive than controls with respect to incidence and time of death, even though the biological activity of serum tumor necrosis factor (TNF) was partially neutralized . High-expressor mice of the adequate genetic background were markedly, although not completely, protected from death by cerebral malaria after injection with Plasmodium berghei . They were highly susceptible to Listeria monocytogenes, dying from bacterial dissemination after sublethal infection, and to Leishmania major, displaying severe, non-healing lesions after local infection . Under the same conditions, mice expressing about 1 microgram protein/ml were only partially sensitive to these last agents, compared to non-transgenic littermate mice which were fully resistant . These transgenic mice represent a model of permanent, complete or partial, impairment of TNF use, which compares favorably, for ease of breeding and for the range of effects, to mice bearing gene disruptions.

Eur J Immunol, 1995 Aug, 25(8), 2384 - 91
Systemic immunization induces protective CD4+ and CD8+ T cell-mediated immune responses in murine Listeria monocytogenes meningoencephalitis; Schluter D et al.; The immune mechanisms underlying immunization-induced protection of mice from lethal central nervous system (CNS) listeriosis were evaluated by immunohistochemistry, flow cytometry of leukocytes isolated from the brain, reverse transcription-polymerase chain reaction analysis of intracerebral (i.c.) tumor-necrosis factor-alpha, interferon-gamma, interleukin (IL)-2, IL-1 beta, IL-10, granulocyte/macrophage colony-stimulating factor, and inducible nitric oxide synthase mRNA expression, and T cell depletion experiments . The data demonstrate that active immunization of mice prior to an i.c . infection with Listeria monocytogenes prevents the development of a fatal necrotizing encephalitis and accelerates the recruitment of an increased number of alpha beta T cell receptor (TcR)+ CD4+ and CD8+ T cells, gamma delta TcR+ T cells, B cells, granulocytes and macrophages to the brain compared to non-immunized animals . In addition, immunization induced a pronounced activation of i.c . macrophages and microglial cells as shown by an increased expression of MHC class II antigens . In parallel, transcript levels for all cytokine mRNA analyzed were higher in the brains of immunized mice . The protective effects of immunization were completely abolished by depletion of CD4+, CD8+, or both T cell subsets . All groups of T cell-depleted immunized mice developed a fatal necrotizing encephalitis with an increased i.c . bacterial load . In addition, cytokine mRNA synthesis was significantly impaired . The severity of disease was only slightly different between CD4+, CD8+ and CD4+/CD8+ T cell depleted mice, indicating that both subsets of T cells are required for an effective i.c . immune response to L . monocytogenes . This is in marked contrast to systemic listeriosis, and points to CNS-specific features of the immune response.

J Med Microbiol, 1995 Aug, 43(2), 125 - 32
Bovine mastitis caused by Listeria monocytogenes: characteristics of natural and experimental infections; Bourry A et al.; Experimental mastitis induced with a single intramammary injection of Listeria monocytogenes was compared with two naturally occurring cases . Four strains of L . monocytogenes, two of serotype 1/2a and two of serotype 4b were used for the experimental infections and two diametrically opposed quarters of four cows were inoculated with 300 cfu . Bacteriological examination and somatic cell counts of quarter foremilk samples were performed weekly for at least 6 months after challenge . All the inoculated quarters developed chronic subclinical mastitis with occasional clinical episodes . The results were similar to those observed in natural listeria mastitis . Four experimentally infected quarters were treated during lactation (gentamicin and cloxacillin) or at "drying-off" (cloxacillin), or at both times . Only one of four quarters was cured after treatment only at "drying-off" . All experimental and naturally infected animals were slaughtered and bacteriological examination was performed on liver, spleen and supramammary, iliac and mesenteric lymph nodes . L . monocytogenes strains were isolated from the supramammary lymph nodes of two experimentally and two naturally infected cows and from an iliac lymph node from one of the naturally infected cows . The epidemiological data were supported by serotyping, lysotyping and DNA macro-restriction analysis . The experimental model of listeria mastitis mimics spontaneous cases and should be useful in further studies of listeria mastitis.

J Hand Surg {Br}, 1995 Aug, 20(4), 509 - 10
Flexor tenosynovitis in the hand . An unusual aetiology; Aubert JP et al.; We report an unusual case of tenosynovitis in the hand . A female farm worker aged 66 developed swelling of the middle and little fingers . Extensive synovectomy of the flexor tendon sheaths was performed, and the synovitis was found to be due to Listeria monocytogenes, which is unusual in this area . Treatment with amoxycillin and gentamicin for 10 days was replaced by trimethoprim-sulfamethoxazole for 3 weeks . There was no recurrence of infection 3 months after surgery, and flexion and extension were complete at that time.

Immunology, 1995 Aug, 85(4), 562 - 7
Interaction of interleukin-6, tumour necrosis factor and interleukin-1 during Listeria infection; Liu Z et al.; Injected recombinant interleukin-6 (IL-6), tumour necrosis factor (TNF) and IL-1 all protect mice against experimental infection with Listeria monocytogenes . We have therefore investigated the interaction of these cytokines during infection . Treatment with recombinant (r)IL-6 enhanced TNF production by spleen cells during the first 2 days of infection . Anti-TNF antibody could totally abolish the protective effect of rIL-6, while the optimal protective function of TNF could not be achieved when IL-6 was neutralized by anti-IL-6 antibody . IL-1 induced a high level of IL-6 in the serum a short time after its administration, and neutralization of IL-6 totally abolished the protective function of rIL-1 . The results thus provide further evidence for the complexity of cytokine interaction.

Microbiology, 1995 Aug, 141 ( Pt 8), 1985 - 92
Reduction of exogenous ferric iron by a surface-associated ferric reductase of Listeria spp; Deneer HG et al.; The reduction of exogenous ferric iron by Listeria monocytogenes, a Gram-positive food-borne pathogen, was investigated . Using an assay incorporating the ferrous iron chelator ferrozine, we showed that intact cells of L . monocytogenes, when exposed to ferric iron, were able to rapidly reduce and solubilize the iron to the ferrous form . Reduction occurred only after direct contact between the bacteria and the iron source . A number of different ferric iron chelates, including transferrin and lactoferrin-bound iron, haemoglobin, ferritin, and iron complexed to siderophores, could be reduced . The ferric reductase activity was expressed by both reference strains and clinical isolates of L . monocytogenes and by all other species of Listeria, although significant quantitative differences were observed . In L . monocytogenes, the expression of ferric reductase was not affected by the growth phase of the bacteria nor by the presence or absence of iron in the growth medium . However, expression was greatly reduced in bacteria grown anaerobically and when cultured in media of reduced pH . In addition, bacteria grown at a cold temperature displayed greater ferric reductase activity than cells grown at higher temperatures . A surface-associated ferric reductase system may be one component of a general iron scavenging mechanism which can be used by Listeria growing in a variety of environments.

Appl Microbiol Biotechnol, 1995 Aug-Sep, 43(4), 717 - 24
Anti-DNA.RNA antibodies: an efficient tool for non-isotopic detection of Listeria species through a liquid-phase hybridization assay; Fliss I et al.; This study was undertaken to evaluate the potential of a new approach using anti-DNA.RNA monoclonal antibodies to detect Listeria in both pure culture and inoculated meat and meat products . A sensitive liquid-phase assay was first developed, based on the formation in solution of a hybrid between a 784-bp DNA probe, specific for the genus Listeria, and target rRNA . Monoclonal antibody and antisera raised against hybrid nucleic acids were then used in various immunoenzymatic assays to detect specific hybrids formed in solution . System 2, using a double sandwich enzyme-linked immunosorbent assay, and system 1, using a biotinylated probe, proved to be very effective . The method using biotin-streptavidin complex, however, resulted in a higher background signal . System 2 described here, using unlabeled probe, was more effective . This strategy allowed the detection of as little as 2.5 pg target RNA from pure culture and 500 cells from inoculated meat homogenate, even in the presence of other contaminating bacteria . The assay was more sensitive and could be completed within 3 h, as opposed to several days when conventional culture methods were used.

Infect Immun, 1995 Aug, 63(8), 3187 - 95
Restricted replication of Listeria monocytogenes in a gamma interferon-activated murine hepatocyte line; Szalay G et al.; The intracellular pathogen Listeria monocytogenes replicates mainly in resting macrophages and hepatocytes residing in infected tissues . Both innate and acquired resistance strongly depend on activation of listericidal capacities of macrophages by gamma interferon (IFN-gamma) produced by natural killer cells and T lymphocytes . In contrast to macrophages, hepatocytes have been considered to serve purely as a cellular habitat, prolonging survival of the pathogen in the host . By using an immortalized murine hepatocyte line, the relationship between L . monocytogenes and this cell type has been analyzed in more detail . Our data reveal that hepatocytes are able to eradicate listeriolysin-deficient (avirulent) L . monocytogenes but fail to control growth of listeriolysin-expressing (virulent) L . monocytogenes organisms . Following stimulation with IFN-gamma, hepatocytes gained the capacity to restrict growth of virulent L . monocytogenes, although less efficiently than the highly listericidal IFN-gamma-activated macrophages . Hepatocytes costimulated with a combination of IFN-gamma, interleukin 6 (IL-6), and tumor necrosis factor alpha (TNF-alpha) expressed the highest antilisterial activities . Although IFN-gamma-stimulated hepatocytes produced demonstrable levels of reactive nitrogen intermediates (RNI), the results of inhibition studies do not support a major role for these molecules in antilisterial hepatocyte activities . In contrast, inhibition of RNI produced by macrophages neutralized their antilisterial effects . IFN-gamma-stimulated, L . monocytogenes-infected hepatocytes expressed TNF-alpha mRNA, suggesting that they are a source of this cytokine during listeriosis . These studies suggest a novel function for hepatocytes in listeriosis: first, IFN-gamma-stimulated hepatocytes could contribute to listerial growth restriction in the liver, and second, through secretion of proinflammatory cytokines, they could promote phagocyte influx to the site of listerial growth.

Int J Food Microbiol, 1995 Aug, 26(3), 375 - 84
Comparison of the incidence of Listeria on equipment versus environmental sites within dairy processing plants; Pritchard TJ et al.; This study was undertaken to compare the incidence of Listeria contamination of processing equipment with that of the general dairy processing environment . A total of 378 sponge samples obtained from 21 dairy plants were analyzed for Listeria using three different enrichment media . Use of extended microbiological analysis allowed us to identify 26 Listeria positive sites which would have not been identified had a single test format been employed . Eighty (80) of 378 sites (21.2%) were identified as Listeria positive . Listeria innocua was isolated from 59 of the 80 (73.8%) positive samples, L . monocytogenes was identified in 35 (43.8%) of the positive samples, and L . seeligeri was isolated from 5 (6.3%) of the Listeria positive samples . Positive equipment samples were obtained from 6 of the 21 (28.6%) plants and 19 of the 21 (90.5%) plants had positive environmental sites . Seventeen of the 215 (7.9%) samples from equipment were positive for Listeria species . Eleven of these sites, including 3 holding tanks, 2 table tops, 3 conveyor/chain systems, a pasta filata wheel, a pint milk filler and a brine pre-filter machine, were positive for L . monocytogenes . Nineteen of the 21 (90.5%) plants had positive environmental sites . Sixty-three of the 163 (41.1%) samples from environmental sites were Listeria positive and 24 were positive for L . monocytogenes . Two-tailed student t-test analysis of the mean frequencies indicated that the level of contamination was significantly higher (p < 0.001) in 'environmental' (49.7%) as opposed to 'equipment' samples (7.0%) . Our study indicates that environmental contamination with Listeria does not necessarily translate into contamination of equipment within the same plant, and that greater emphasis needs to be placed on the cleaning and sanitizing of the plant environment.

Appl Environ Microbiol, 1995 Aug, 61(8), 3177 - 9
Use of pulsed-field gel electrophoresis to link sporadic cases of invasive listeriosis with recalled chocolate milk; Proctor ME et al.; Pulsed-field gel electrophoresis established the linkage between recalled chocolate milk and a multistate invasive listeriosis outbreak during a four-product recall period . Listeria monocytogenes isolates from four hospitalized patients and an environmental dairy sample displayed AscI restriction endonuclease digestion profiles identical to that of the chocolate milk isolate.

Mol Cell Probes, 1995 Aug, 9(4), 265 - 76
The sensitive detection of fluorescently labelled PCR products using an automated detection system; Maher M et al.; The polymerase chain reaction plays a central role in many detection assays and methods to improve the sensitivity and specificity of these detection systems are constantly being explored . In this study we investigated the use of an automated laser fluorescent system (ALF) in the context of DNA-based diagnostics for pathogenic bacteria . PCR products were generated using species-specific primer sets, one of which was labelled with a 5' fluorescein . PCR products with a fluorescent label were detected on line with an ALF DNA sequencer and the sensitivity of detection was found to be comparable to that for DNA probe hybridization with a radioactive probe . The technology was successfully applied to the detection of Mycobacterium tuberculosis supplemented into sputum samples and to the detection of listeria in paraffin-embedded tissue samples.

FEMS Immunol Med Microbiol, 1995 Jul, 11(4), 321 - 7
A monoclonal antibody against T-cell receptor alpha beta induces endogenous cytokines and prevents mice from a lethal infection with Listeria monocytogenes; Nakane A et al.; In vivo induction of cytokines by a monoclonal antibody (mAb) against T-cell receptor (TCR) alpha beta and the protective effect induced by the mAb on a lethal infection with Listeria monocytogenes were studied . Injection of anti-TCR alpha beta mAb induced rapid production of endogenous tumour necrosis factor in the spleens, and gamma interferon and interleukin-6 in the blood streams and spleens of mice . Administration of anti-CD4 mAb, anti-CD8 mAb, or anti-Thy1.2 mAb resulted in suppression of anti-TCR alpha beta mAb-induced endogenous cytokine production . Mice were protected against lethal L . monocytogenes infection when treated with anti-TCR alpha beta mAb . The protective effect was not demonstrated in CD4+ cell- or CD8+ cell-depleted mice . These results suggest that anti-TCR alpha beta mAb shows a protective effect on a lethal infection with L . monocytogenes in mice and that the mAb-induced endogenous cytokines might be involved in the effect of anti-TCR alpha beta mAb.

Infect Immun, 1995 Jul, 63(7), 2790 - 2
Role of interleukin-6 in T-cell activation during primary and secondary infection with Listeria monocytogenes; Liu Z et al.; Injection of recombinant interleukin-6 (IL-6) into mice enhances recovery from infection with Listeria monocytogenes . In this study, the role of IL-6 during primary and secondary Listeria infection was further tested . Neutralization of IL-6 by polyclonal antibody exacerbated primary infection and significantly delayed gamma interferon production by cultured spleen cells . In contrast, administration of anti-IL-6 antibody at the time of secondary infection did not affect the recovery of mice from infection or gamma interferon production, showing that activated T cells are not dependent on IL-6.

Infect Immun, 1995 Jul, 63(7), 2729 - 37
iactA of Listeria ivanovii, although distantly related to Listeria monocytogenes actA, restores actin tail formation in an L . monocytogenes actA mutant; Gouin E et al.; A gene homologous to the actA gene of Listeria monocytogenes was cloned from Listeria ivanovii (strain CLIP257) by chromosome walking starting from the ilo gene that encodes the pore-forming toxin ivanolysin . The nucleotide sequence revealed that this gene, named iactA, encodes a protein of 1,044 amino acids (IactA) comprising a central region with seven highly conserved tandem proline-rich repeats of 47 amino acids . Although IactA and ActA share an overall similar structure, these two proteins are only distantly related . Like ActA, IactA migrates aberrantly on sodium dodecyl sulfate gels . When expressed in an L . monocytogenes actA deletion mutant strain, iactA restored actin polymerization.

Lett Appl Microbiol, 1995 Jul, 21(1), 5 - 9
A rapid general method for the identification of PCR products using a fibre-optic biosensor and its application to the detection of Listeria; Strachan NJ et al.; A 200-mer fragment of the flaA gene from Listeria monocytogenes was amplified using the polymerase chain reaction (PCR) incorporating biotin- and fluorescein amadite (FAM-)labelled primers . Methods are described for isolating the single stranded FAM-labelled 200-mer . A central portion of this 200-mer was successfully hybridized onto a complementary sequence coated onto a fibre optic biosensor . Advantages in the specificity and speed of this approach compared to standard agarose gel electrophoresis and probing methods are discussed.

Can J Microbiol, 1995 Jul, 41(7), 572 - 7
Repression of motility and flagellin production at 37 degrees C is stronger in Listeria monocytogenes than in the nonpathogenic species Listeria innocua; Kathariou S et al.; Listeria monocytogenes and Listeria innocua differ markedly in virulence but are indistinguishable by classical taxonomic criteria . Both species are actively motile and produce abundant flagellin at 22 degrees C . We have found, however, noticeable differences between L . monocytogenes and L . innocua in motility and flagellin production at 37 degrees C . At this temperature, L . monocytogenes strains were virtually nonmotile and produced little or no detectable flagellin, whereas strains of L . innocua were frequently motile and produced substantial amounts of flagellin . This flagellin was recognized by a Listeria genus-specific monoclonal antibody that also recognized flagellin produced at 22 degrees C . These results suggest differential regulation of flagellin production between L . monocytogenes and L . innocua at 37 degrees C.

Eur J Immunol, 1995 Jul, 25(7), 1985 - 91
Early appearance of T cell receptor alpha beta + CD4- CD8- T cells with a skewed variable region repertoire after infection with Listeria monocytogenes; Matsuzaki G et al.; We found that the number of T cell receptor (TCR) alpha beta + CD4- CD8- T cells increased in the peritoneal cavity on day 5 after an intraperitoneal infection with Listeria monocytogenes strain EGD together with TCR gamma delta + CD4- CD8- T cells . Thereafter, the TCR alpha beta + CD4- CD8- T cells decreased to a normal level by day 14 . The TCR alpha beta + CD4- CD8- T cells showed an activated T cell phenotype (L-selectin CD44 +) and expressed CD45/B220 and interleukin-2 receptor beta, but did not express heat stable antigen, which is expressed by the immature CD4- CD8- thymocytes . Furthermore, 20-30% of the TCR alpha beta + CD4- CD8- T cells expressed the NK1.1 natural killer cell marker . Analysis of the TCR V region repertoire of the TCR alpha beta + CD4- CD8- T cells induced by L . monocytogenes infection showed that more than 80% of the TCR alpha beta + CD4- CD8- T cells expressed TCR V beta 8 detected by anti-TCR V beta 8.1 and 8.2 mAb, and a reverse transcription-polymerase chain reaction analysis of V alpha 14 relative to V alpha 11 expression revealed that the TCR alpha beta + CD4- CD8- T cells expressed a higher level of V alpha 14, which was reported to be preferentially expressed by TCR alpha beta + CD4- CD8- thymocytes rather than conventional CD4+ T cells . The TCR alpha beta + CD4- CD8-T cells showed a proliferative response to anti-TCR alpha beta mAb stimulation . In contrast, they showed no response to stimulation with either Listeria antigen or 65-kDa heat shock protein of Mycobacterium bovis, which do stimulate the Listeria-specific TCR alpha beta + CD4- CD8- T cells and the Listeria-induced TCR gamma delta + T cells, respectively . These results suggest that the TCR alpha beta + CD4- CD8- T cells may recognize a restricted set of self antigens induced by L . monocytogenes infection, and that they contribute to host protection at an early stage of infection.

Immunity, 1995 Jul, 3(1), 9 - 19
Resistance to fever induction and impaired acute-phase response in interleukin-1 beta-deficient mice; Zheng H et al.; We used gene targeting in embryonic stem cells to introduce an IL-1 beta null allele in mice . The IL-1 beta-deficient mice develop normally and are apparently healthy and fertile . The IL-1 beta null mice responded normally in models of contact and delayed-type hypersensitivity or following bacterial endotoxin LPS-induced inflammation . The IL-1 beta-deficient mice showed equivalent resistance to Listeria monocytogenes compared with wild-type controls . In contrast, when challenged with turpentine, which causes localized inflammation and tissue injury, the IL-1 beta mutant mice exhibited an impaired acute-phase inflammatory response and were completely resistant to fever development and anorexia . These results highlight a central role for IL-1 beta as a pyrogen and a mediator of the acute-phase response in a subset of inflammatory disease models, and support the notion that blocking the action of a single key cytokine can alter the course of specific immune and inflammatory responses.

Immunity, 1995 Jul, 3(1), 109 - 17
Specific immunity to Listeria monocytogenes in the absence of IFN gamma; Harty JT et al.; Cytokine and cytokine receptor gene knockout mice provide powerful experimental systems to characterize the functions of these molecules in resistance to infectious disease . Such mice may also provide unique models of immune deficiency to learn whether manipulation of the immune response can overcome the specific dysfunction . We demonstrate that resistance of IFN gamma gene knockout (GKO-/-) mice to the intracellular bacterium Listeria monocytogenes is severely impaired compared with wild-type mice . However, immunization of GKO-/- mice with an attenuated L . monocytogenes strain generates antigen-specific CD8 T cell responses that can transfer immunity to naive hosts . Furthermore, vaccinated GKO-/- mice themselves exhibit 20,000-fold increased resistance to challenge with virulent L . monocytogenes and this resistance appears to be CD8 T cell mediated . These studies demonstrate that vaccination-induced immunity can overcome the absence of a cytokine that is critical for resistance to acute infection.

J Cell Biol, 1995 Jul, 130(2), 331 - 43
Actin-based movement of Listeria monocytogenes: actin assembly results from the local maintenance of uncapped filament barbed ends at the bacterium surface; Marchand JB et al.; The thermodynamic basis for actin-based motility of Listeria monocytogenes has been investigated using cytoplasmic extracts of Xenopus eggs, initially developed by Theriot et al . (Theriot, J . A., J . Rosenblatt, D . A . Portnoy, P . J . Goldschmidt-Clermont, and T . J . Mitchison . 1994 . Cell . 76:505-517) as an in vitro cell-free system . A large proportion (75%) of actin was found unpolymerized in the extracts . The amount of unassembled actin (12 microM) is accounted for by the sequestering functions of T beta 4Xen (20 microM) and profilin (5 microM), the barbed ends being capped . Movement of Listeria was not abolished by depletion of over 99% of the endogenous profilin . The proline-rich sequences of ActA are unlikely to be the target of profilin . All data support the view that actin assembly at the rear of Listeria results from a local shift in steady state due to a factor, keeping filaments uncapped, bound to the surface of the bacterium, while barbed ends are capped in the bulk cytoplasm . Movement is controlled by the energetic difference (i.e., the difference in critical concentration) between the two ends of the filaments, hence a constant ATP supply and the presence of barbed end capped F-actin in the medium are required to buffer free G-actin at a high concentration . The role of membrane components is demonstrated by the facts that: (a) Listeria movement can be reconstituted in the resuspended pellets of high speed-centrifuged extracts that are enriched in membranes; (b) Actin-based motility of endogenous vesicles, exhibiting the same rocketing movement as Listeria, can be observed in the extracts.

Cell Immunol, 1995 Jul, 163(2), 260 - 7
Analysis of the interrelationship between IL-12, TNF-alpha, and IFN-gamma production during murine listeriosis; Liu W et al.; IL-12, a recently described cytokine, is an important mediator in the early production of IFN-gamma during infection . To evaluate the timing of IL-12 production, and its relationship to TNF-alpha, and IFN-gamma production during primary murine listeriosis, we measured cytokine mRNA and protein levels in C57B1/6 mice infected intravenously with Listeria monocytogenes (LM) . IL-12 is a disulfide-linked heterodimer containing two chains (designated P35 and P40); however, bioactive cytokine production has been more closely linked with P40 expression . Consequently, we monitored mRNA and protein levels of P40 in the spleen as a marker for IL-12 production in vivo . Splenic P40 mRNA levels (assayed using RNase protection methods) were low in uninfected animals, but increased markedly beginning 15 to 18 hr after LM infection . In sublethally infected animals, P40 mRNA levels remained elevated for 5 days, returning to baseline with the resolution of infection . P40 protein (assayed using an antibody capture ELISA) could be detected in the spleens of LM-infected animals beginning around 18 hr postinfection confirming linkage between P40 mRNA accumulation and the generation of a protein product . In comparing P40 and IFN-gamma mRNA levels in vivo, we found in each case that substantial increases in mRNA accumulation did not appear until 15-18 hr postinfection . In comparable studies using BALB/c animals, cytokine production began slightly earlier (between 12 and 15 hr) but once again P40 and IFN-gamma mRNA levels increased in a coordinated manner . P40 mRNA (like IFN-gamma and TNF-alpha mRNA) only accumulated in animals infused with live, virulent bacteria . Although we could detect no obvious lag between the time of onset of IL-12 and IFN-gamma accumulation in vivo, infusions of anti-IL-12 antibodies markedly reduced IFN-gamma expression implying that IL-12 production precedes and directs IFN-gamma production . TNF-alpha production, on the other hand, was not diminished by anti-IL-12 treatment . Our studies demonstrate that IL-12 generation is an essential step in normal IFN-gamma production during listeriosis, and suggest that IL-12, once produced, may begin enhancing IFN-gamma production in vivo in less than 3 hr.

Haematologica, 1995 Jul-Aug, 80(4), 305 - 10
Combined autoimmune cytopenias; Martino R et al.; BACKGROUND . Autoimmune neutropenia (AIN) can occur in association with autoimmune hemolytic anemia (AIHA) and/or immune thrombocytopenic purpura (ITP), although these associations have not been studied in detail . METHODS AND RESULTS . Twenty cases of AIN were found in a group of 55 adults with unexplained neutropenia over a five-year period . Eight subjects with AIN had an associated AIHA and/or ITP (AIN+ITP, n = 2; AIN+AIHA, n = 2; AIN+ITP+AIHA, n = 4) . Thorough investigations identified no underlying disease in four patients, and none has appeared during follow-up . Of the other 4, one was found to have been suffering from systemic lupus erythematosus when the combined immunocytopenia was diagnosed, one patient from idiopathic myelofibrosis, one from a combined variable immunodeficiency and the other from disseminated tuberculosis . These last three conditions, while sometimes associated with autoimmune cytopenias, has not been previously reported together with combined immunocytopenias . All patients responded to immunosuppressors, although severe infectious complications occurred in two, leading to death from Pneumocystis carinii pneumonia and to irreversible neurologic damage from Listeria monocytogenes meningitis, respectively . CONCLUSIONS . We conclude that combined autoimmune cytopenias are frequently observed in patients with AIN, and a thorough search for associated conditions can lead to unsuspected diagnoses.

Int J Food Microbiol, 1995 Jul, 26(2), 245 - 50
Occurrence of Listeria monocytogenes in soft and semi-soft cheeses in retail outlets in Sweden; Loncarevic S et al.; Samples of 333 retail cheeses produced in or imported into Sweden were examined for the presence of Listeria monocytogenes . Listeria monocytogenes was isolated from 6% of the cheese samples . Cheeses made from raw milk were more frequently contaminated with L . monocytogenes (42%) than cheeses made from heat-treated milk (2%) . The incidence of the organism in whole cheeses and pre-cut wedges was similar (6%) . L . monocytogenes was only found in imported cheeses (18 from France, and one from Germany and Italy, respectively) . The numbers of L . monocytogenes varied from < 1 x 10(2) to 1 x 10(5) cfu/g . All L . monocytogenes strains belonged to serogroup 1/2, except isolates from two samples that belonged to serogroup 4.

Int J Food Microbiol, 1995 Jul, 26(2), 177 - 86
Factors affecting results obtained with European Community tests for pasteurized milk sampled at the heat treatment establishment; Burden P et al.; A total of 3000 samples of pasteurized milk taken at the heat treatment establishment over a 1-year period were examined for the presence of phosphatase and by a plate count at 30 degrees C, a coliform count at 30 degrees C, and a plate count at 21 degrees C after pre-incubation of the sample for 5 days at 6 degrees C, performed as prescribed in EC Directive 85/397/EEC . Samples were also examined for presence of Listeria species . Of 2690 samples (1713 from small dairies and 977 from large commercial premises) received at 4 degrees C or less, 96 (3.6%) has a plate count at 30 degrees C exceeding 30,000 organisms per ml, 608 (22.6%) contained 1 or more coliforms per ml, and in 1327 (49.3%) the pre-incubated count exceeded 100,000 organisms per ml . Thirty two samples from 23 dairies contained phosphatase . Listeria species were detected in 25 samples; only three of these strains were identified as Listeria monocytogenes . Results were analysed to determine relationships between factors which might affect test results . Samples from small dairies showed significantly higher coliform counts and pre-incubated plate counts than did those from large commercial premises . They were also more likely to contain phosphatase or Listeria species . Samples were significantly more likely to contain coliforms when taken in the July-September quarter or if received in glass containers.

Int J Food Microbiol, 1995 Jul, 26(2), 165 - 76
Enrichment in Fraser broth supplemented with catalase or Oxyrase, combined with the microcolony immunoblot technique, for detecting heat-injured Listeria monocytogenes in foods; Patel JR et al.; The microcolony immunoblot technique using monoclonal antibodies to Listeria monocytogenes was evaluated for its suitability to detect heat-injured cells . Pasteurized milk and filtrates of homogenized raw ground beef slurry and cabbage were inoculated with L . monocytogenes Scott A, heated, diluted, inoculated into Fraser broth (FB) supplemented with 400 micrograms of catalase ml-1 or 0.01 unit of Oxyrase ml-1, and incubated at 30 degrees C for 6 h . Three inoculum populations (high, medium, and low) were used . The extent of injury was dependent on the heating menstruum . Forty percent of the cells were injured in beef slurry filtrate, whereas 79 and 94% were injured in milk and cabbage filtrate, respectively, when foods were heated at 52 degrees C for 20 min . Populations of viable cells were determined using the immunoblot technique and by surface plating on modified Oxford (mMOX) agar . Recovery of cells from heated foods was enhanced in FB supplemented with catalase or Oxyrase compared to recovery in control broth . Essentially all unheated (control) cells could be detected within about 30 h using enrichment and the immunoblot technique; 54 h were required to easily detect colonies on mMOX . In most cases, the number of cells detected in heated milk or filtrates of homogenized beef after enrichment in FB supplemented with catalase or Oxyrase was significantly higher than populations detected using unsupplemented FB; however, enrichment in FB supplemented with catalase or Oxyrase did not significantly increase cell populations in heated cabbage filtrate . Within each heat treatment and level of inoculum, cell populations detected on mMOX agar after incubating plates for 48 h or on immunoblots after 24 h were not significantly different . Results indicate that the immunoblot technique in conjunction with enrichment in FB containing either catalase or Oxyrase can be successfully used to detect healthy and heat-injured cells of L . monocytogenes in diverse types of foods within 34 h.

J Interferon Cytokine Res, 1995 Jul, 15(7), 633 - 5
Interferon-gamma detection in cultures of newborn cells exposed to Listeria monocytogenes; Serushago B et al.; Clearance of Listeria monocytogenes in experimental models of infection has underscored the importance of interferon-gamma (IFN-gamma) in host resistance to intracellular pathogens . Because L . monocytogenes infections are more severe in newborns than adults, we compared IFN-gamma accumulation in the supernatants of mononuclear cells infected in vitro from newborns with those from adults . Supernatants were assayed for IFN-gamma using an enzyme-linked immunosorbent assay . Uninfected newborn and adult mononuclear cells had less than 50 pg/ml of IFN-gamma at all times tested . IFN-gamma levels in supernatants from infected adult mononuclear cells at 24 h of culture (1.15 x 10(3) +/- 0.92 pg/ml) were greater than supernatants from infected newborn mononuclear cells (0.19 x 10(3) +/- 0.33 pg/ml) . IFN-gamma concentrations in newborn cell cultures plateaued on day 3 of culture (1.6 x 10(3) +/- 1.1 pg/ml) and were not significantly less than concentrations from adult cells . However, adult cell IFN-gamma was further increased by day 5 (18.7 x 10(3) +/- 21.8 pg/ml) . Because IFN-gamma plays a critical role in the host defense against L . monocytogenes, this delay in the release of IFN-gamma may be a factor in the increased susceptibility and severity of infection in the neonate.

Vaccine, 1995 Jul, 13(10), 923 - 6
The efficacy of a live Listeria monocytogenes combined serotype 1/2a and serotype 4b vaccine; Linde K et al.; In each of two experiments, sheep and lambs were vaccinated by a subcutaneous injection of a test vaccine (consisting of a combined serotype 1/2a and serotype 4b live Listeria monocytogenes culture) and challenged 16 days later with a mixture of the homologous wild strains . After challenge, the mortality rate of vaccinated sheep was 28.1% and that of controls 71.9%; that of lambs was 25.0%, although these had been inoculated with the LD70 dose . Furthermore, in each of two field trials in Listeria-infected flocks, primiparous pregnant ewes were vaccinated . In the first field trial, 3 or 110 lambs died of listeriosis of those born of vaccinated (n = 564) or unvaccinated (n = 3345) ewes, respectively . In the second, the perinatal mortality rate of lambs born of vaccinated or unvaccinated ewes was 7.6 or 30.3%, respectively, and the mean birth weight of lambs born of vaccinated or control ewes was 2.2 or 1.8 kg, respectively; the mean milk production of vaccinated ewes was 106 and that of controls 83 l; no Listeria was isolated from milk samples of vaccinated ewes . It is concluded that the vaccine is efficacious for the protection of sheep from listeriosis.

EMBO J, 1995 Jun 15, 14(12), 2731 - 44
Targeting of Listeria monocytogenes ActA protein to the plasma membrane as a tool to dissect both actin-based cell morphogenesis and ActA function; Friederich E et al.; Actin assembly on the surface of Listeria monocytogenes in the cytoplasm of infected cells provides a model to study actin-based motility and changes in cell shape . We have shown previously that the ActA protein, exposed on the bacterial surface, is required for polarized nucleation of actin filaments . To investigate whether plasma membrane-associated ActA can control the organization of microfilaments and cell shape, variants of ActA, in which the bacterial membrane signal had been replaced by a plasma membrane anchor sequence, were produced in mammalian cells . While both cytoplasmic and membrane-bound forms of ActA increased the F-actin content, only membrane-associated ActA caused the formation of plasma membrane extensions . This finding suggests that ActA acts as an actin filament nucleator and shows that permanent association with the inner face of the plasma membrane is required for changes in cell shape . Based on the observation that the amino-terminal segment of ActA and the remaining portion which includes the proline-rich repeats cause distinct phenotypic modifications in transfected cells, we propose a model in which two functional domains of ActA cooperate in the nucleation and dynamic turnover of actin filaments . The present approach is a new model system to dissect the mechanism of action of ActA and to further investigate interactions of the plasma membrane and the actin cytoskeleton during dynamic changes of cell shape.

Mol Microbiol, 1995 Jun, 16(6), 1231 - 41
Heterogeneous endolysins in Listeria monocytogenes bacteriophages: a new class of enzymes and evidence for conserved holin genes within the siphoviral lysis cassettes; Loessner MJ et al.; Listeria monocytogenes bacteriophages A118, A500 and A511 are members of three distinct phage groups with characteristic host ranges . Their endolysin (ply) genes were cloned and expressed in Escherichia coli as demonstrated by the conferred lytic phenotype when colonies of recombinant cells were overlaid with a lawn of Listeria cells . The nucleotide sequences of the cloned DNA fragments were determined and the individual enzymes (PLY118, 30.8 kDa; PLY500, 33.4 kDa; PLY511, 36.5 kDa) were shown to have varying degrees of homology within their N-terminal or C-terminal domains . Transcriptional analysis revealed them to be 'late' genes with transcription beginning 15-20 min post-infection . The enzymes were overexpressed and partially purified and their individual specificities examined . When applied exogenously, the lysins induced rapid lysis of Listeria strains from all species but generally did not affect other bacteria . Using hydrolysis of purified listerial cell walls, PLY511 was characterized as an N-acetylmuramoyl-L-alanine amidase (EC 3.5.1.28) and shows homology in its N-terminal domain to other enzymes of this type . In contrast, PLY118 and PLY500 were shown to represent a new class of cell wall lytic enzymes which cleave between the L-alanine and D-glutamate residues of listerial peptidoglycan; these were designated as L-alanoyl-D-glutamate peptidases . These two enzymes share homology in the N-terminal domain which we propose determines hydrolytic specificity . Highly conserved holin (hol) gene sequences are present upstream of ply118 and ply500 . They encode proteins of structural similarity to the product of phage lambda gene S, and are predicted to be membrane proteins which form pores to allow access of the lysins to their peptidoglycan substrates . This arrangement of conserved holin genes with downstream lysin genes among the siphoviral lysis cassettes explains why the cytoplasmic endolysins alone are not lethal, since they require a specific transport function across the cell membrane.

Microb Pathog, 1995 Jun, 18(6), 417 - 22
Vaccination of mice with attenuated mutants of Listeria monocytogenes: requirement for induction of macrophage la expression; Gahan CG et al.; Isogenic mutant strains of Listeria monocytogenes demonstrating graded attenuation in mice were used to analyse the correlation between bacterial virulence, ability to induce class II MHC (la) molecules in antigen presenting cells and ability to vaccinate against secondary infection . The mutants used differed only in the amino acid sequence of the thiol-activated hemolysin, Listeriolysin O (LLO) . The results indicate that L . monocytogenes mutants of reduced virulence have the potential to act as vaccines only if they are sufficiently persistent to induce la expression in antigen presenting cells . The findings also suggest that specific mutagenesis of virulence factors, including LLO, could provide an approach for creating Listeria monocytogenes strains with potential for use as attenuated live vaccines.

Appl Environ Microbiol, 1995 Jun, 61(6), 2242 - 6
Investigations related to the epidemic strain involved in the French listeriosis outbreak in 1992; Jacquet C et al.; Two hundred seventy-nine cases of human listeriosis (92 pregnancy-related cases and 187 non-pregnancy-related cases) caused by a serovar 4b and phagovar 2389:2425:3274:2671:47:108:340 strain were identified in France between March and December 1992 . Epidemiological investigations included a case-control study (not described here) and microbiological analyses of foods . Results of the case-control study and characterization of food isolates identified pork tongue in jelly, a ready-to-eat meat product, as the major vehicle of this outbreak, and to a lesser extent, delicatessen products contaminated secondarily during handling in food stores . As far as serotyping, phage typing, DNA macrorestriction pattern analysis (obtained by pulsed-field gel electrophoresis {PFGE}), and ribotyping are concerned, this epidemic strain is phenotypically and genomically closely related to strains responsible for major outbreaks of listeriosis previously observed in Europe and North America . The epidemic strain sensu stricto as defined by PFGE (2/1/3) displayed the same serovar, phagovar, ribovar, and ApaI and NotI PFGE patterns as the epidemic strains from outbreaks in Switzerland, California, and Denmark, but it consistently showed differences in the SmaI PFGE profile . This information greatly contributed to the identification of the major food vehicle (pork tongue in jelly) and further allowed exclusion of other foods (cheese) as possible sources of this major listeriosis epidemic.

Appl Environ Microbiol, 1995 Jun, 61(6), 2139 - 44
Characterization of Listeria monocytogenes isolated from poultry products and from the poultry-processing environment by random amplification of polymorphic DNA and multilocus enzyme electrophoresis; Lawrence LM et al.; A total of 289 Listeria monocytogenes strains isolated from a poultry-processing environment and poultry products over a 6-month period were characterized by random amplification of polymorphic DNA, (RAPD) to pinpoint sources of contamination within the plant and gain some measure of the persistence of individual genotypes within this environment . Eighteen RAPD profiles (A through R) were identified within this group, with 64% (184 of 289) of all strains displaying a single RAPD profile, RAPD type A . This genotype was more prevalent in the raw-poultry-processing environment, where, although its origin within this environment appeared to be the incoming birds, it was also widespread on food contact surfaces, floors, and drains . This was the only genotype which persisted throughout the entire 6-month period, and it and RAPD type B were the only two genotypes found in both the raw- and cooked-poultry-processing environments . L . monocytogenes strains isolated from cooked poultry products and the cooked-poultry-processing environment up to 1 year later (17 strains) contained only RAPD types A and B, highlighting the potential which exists for persistent strains to cross-contaminate foods processed in that environment . The other genotypes (C through R) occurred more sporadically, suggesting varied sources of contamination . These were confined to either the raw- or the cooked-poultry-processing environment and were relatively short-lived . Further characterization of a selection of RAPD type A strains, together with strains of RAPD types B through R, was carried out by multilocus enzyme electrophoresis . Strains of RAPD type A contained two electrophoretic types, one of which was serotype 1/2a and the other was 1/2c.(ABSTRACT TRUNCATED AT 250 WORDS)

J Leukoc Biol, 1995 Jun, 57(6), 827 - 31
Neutrophils are essential for resolution of primary and secondary infection with Listeria monocytogenes; Rakhmilevich AL; The role for neutrophils in the resolution of primary and secondary infection with Listeria monocytogenes was studied . The results show that although control mice started to clear Listeria from their spleens and livers between days 2 and 4 of sublethal primary infection and eradicated bacteria in 2 weeks, mice given a specific granulocyte-depleting antibody (RB6-8C5) on days 4 or 6 of infection developed lethal listeriosis . Likewise, treatment of immunized mice with RB6-8C5 monoclonal antibody abolished their acquired ability to resolve a lethal challenge infection . The results demonstrate that neutrophils are necessary for the resolution of secondary and primary Listeria infection.

J Bacteriol, 1995 Jun, 177(11), 3205 - 12
An ATP-dependent L-carnitine transporter in Listeria monocytogenes Scott A is involved in osmoprotection; Verheul A et al.; Listeria monocytogenes is a gram-positive, psychotrophic, food-borne pathogen which is able to grow in osmotically stressful environments . Carnitine (beta-hydroxy-L-tau-N-trimethyl aminobutyrate) can contribute significantly to growth of L . monocytogenes at high osmolarity (R . R . Beumer, M . C . te Giffel, L . J . Cox, F . M . Rombouts, and T . Abee, Appl . Environ . Microbiol . 60:1359-1363, 1994) . Transport of L-{N-methyl-14C}carnitine in L . monocytogenes was shown to be energy dependent . Analysis of cell extracts revealed that L-carnitine was not further metabolized, which supplies evidence for its role as an osmoprotectant in L . monocytogenes . Uptake of L-carnitine proceeds in the absence of a proton motive force and is strongly inhibited in the presence of the phosphate analogs vanadate and arsenate . The L-carnitine permease is therefore most likely driven by ATP . Kinetic analysis of L-carnitine transport in glucose-energized cells revealed the presence of a high-affinity uptake system with a Km of 10 microM and a maximum rate of transport (Vmax) of 48 nmol min-1 mg of protein-1 . L-{14C}carnitine transport in L . monocytogenes is significantly inhibited by a 10-fold excess of unlabelled L-carnitine, acetylcarnitine, and tau-butyrobetaine, whereas L-proline and betaine display, even at a 100-fold excess, only a weak inhibitory effect . In conclusion, an ATP-dependent L-carnitine transport system in L . monocytogenes is described, and its possible roles in cold adaptation and intracellular growth in mammalian cells are discussed.

Infect Immun, 1995 Jun, 63(6), 2288 - 94
Human T-cell recognition of Listeria monocytogenes: recognition of listeriolysin O by TcR alpha beta + and TcR gamma delta + T cells; Guo Y et al.; The cell-mediated immune response to Listeria monocytogenes has been well characterized in the mouse . Listeriolysin O (LLO) is a major antigen in murine T-cell recognition of L . monocytogenes . In this study, we show that LLO is also recognized by human TcR alpha beta T cells and TcR gamma delta T cells . Human peripheral blood mononuclear cells (PBMC) cultured in vitro with live listeriae and then expanded with interleukin 2 were shown to respond to purified LLO . The generation of LLO-responsive T cells was dependent on the use of live bacteria during the initial in vitro challenge . LLO-induced proliferation of T cells expanded by exposure of PBMC to live listeriae was major histocompatibility complex restricted . PBMC cultured with formalin-fixed listeriae and subsequently expanded by interleukin 2 gave high proliferative responses to fixed bacteria but failed to respond to LLO . PBMC stimulated in vitro with fixed listeriae contained predominantly TcR alpha beta + T cells . In contrast, PBMC obtained from 85% of the donors studied generated high numbers of TcR gamma delta + T cells following in vitro culture with live listeriae . Using a panel of synthetic amphipathic LLO peptides, we found that LLO-specific T cells from different individuals recognized both common and unique peptides . LLO 470-508 was recognized by three of five individuals, while LLO 203-226 and LLO 107-126 were recognized by two of six individuals . A TcR gamma delta + T-cell line was established from PBMC stimulated with live listeriae and was shown to recognize LLO 470-508 . Proliferative responses could be induced in this cell line by peptide-pulsed autologous PBMC but not by peptide-pulsed allogeneic PBMC . Our results establish the importance of LLO in human T-cell recognition of listeriae and show that both TcR alpha beta + T cells and TcR gamma delta + T cells recognize this antigen . Finally, since LLO 470-508 has a high degree of homology with other gram-positive bacterial toxins, the recognition of this peptide by TcR gamma delta + T cells suggests that an important role of these T cells in host defense is the recognition of bacterium-derived toxins.

Infect Immun, 1995 Jun, 63(6), 2262 - 8
Interleukin-6-deficient mice are highly susceptible to Listeria monocytogenes infection: correlation with inefficient neutrophilia; Dalrymple SA et al.; We have produced interleukin-6 (IL-6)-deficient mice to examine, in vivo, the wide variety of biological activities attributed to this multifunctional cytokine . To investigate the role of IL-6 during infectious disease, IL-6-deficient mice were challenged with sublethal doses of Listeria monocytogenes, a facultative intracellular bacterium . While normal control animals were able to clear the infection, mutant animals exhibited a high mortality rate and showed uncontrolled replication of the bacteria in the spleen and liver at 2 and 3 days postinfection . Sections of infected tissues showed an increase in the number and severity of inflammatory foci . All aspects of this phenotype in the mutant animals were completely reverted upon administration of recombinant murine IL-6 (rIL-6) . Various parameters of natural killer (NK) cell and macrophage function were unaffected in the challenge of the mutant animals . However, IL-6-deficient animals failed to mount peripheral blood neutrophilia in response to listeriosis, whereas control animals displayed a prominent neutrophilia in the blood at 24 and 48 h postinfection . Additionally, we analyzed the efficacy of rIL-6 in protecting animals devoid of lymphocytes or devoid of neutrophils during listeriosis . Administration of rIL-6 was protective to animals devoid of lymphocytes, suggesting that the rIL-6 protective effect was not mediated through lymphocytes . In contrast, control and mutant animals depleted of neutrophils were refractory to the rIL-6 protective effect . These data suggest that IL-6 is critical early during listeriosis, perhaps acting by stimulating neutrophils either directly or indirectly . Additionally, these data show a promising therapeutic potential for rIL-6 administration during opportunistic infection.

Int J Food Microbiol, 1995 Jun, 26(1), 1 - 13
Culture media and methods for the isolation of Listeria monocytogenes; Curtis GD et al.; The recovery of low numbers of Listeria monocytogenes from foods and environmental samples requires the use of enrichment cultures followed by selective plating and, where injured organisms are likely to be present, a pre-enrichment step . The development of selective and enrichment media for L . monocytogenes is traced and currently used media are discussed . Comparisons of media and methods for the culture of L . monocytogenes are reported but no single method can be recommended for all situations . Guidance is given on the choice of media and methods which is governed by the type of sample, number and nature of competing flora and cost.

J Appl Bacteriol, 1995 Jun, 78(6), 636 - 46
Factors affecting the growth of Listeria monocytogenes on minimally processed fresh endive; Carlin F et al.; The influence of various factors on the fate of Listeria monocytogenes on cut leaves of broad-leaved endive has been studied . Factors considered were temperature, characteristics of the leaves (age, quantity and quality of the epiphytic microflora) and characteristics of the L . monocytogenes inoculum (concentration, strain) . The increases in numbers of L . monocytogenes were lower than those of the aerobic mesophilic microflora at 3 degrees, 6 degrees, 10 degrees and 20 degrees C . Doubling times of the populations of L . monocytogenes were in the same order of magnitude as those of aerobic bacteria at 10 degrees and 20 degrees C, but longer at 3 degrees and 6 degrees C . There were positive significant correlations between growth of L . monocytogenes and populations of aerobic bacteria, and between growth of L . monocytogenes and extent of spoilage on the leaves . Of 225 bacteria isolated from the leaves, 84% were identified as fluorescent pseudomonads; there was no difference in the species isolated from leaves that showed a low growth of L . monocytogenes and leaves that showed a high growth of L . monocytogenes . Populations of L . monocytogenes increased faster during the first 2 and 4 d of storage at 10 degrees C on leaves inoculated with 10-10(3) cfu g-1 than on leaves inoculated with about 10(5) cfu g-1, but the population reached after 7 d was lower . The behaviour of L . monocytogenes was similar among the three strains tested.

Immunobiology, 1995 Jun, 193(1), 71 - 83
Prevention of mortality by in vivo depletion of alpha beta T cells in murine lethal listeriosis and involvement of gamma delta T cells in bacterial elimination; Matsutake T et al.; We previously demonstrated that in murine lethal listeriosis, death is mainly due to massive liver necrosis . In the present study we found that in vivo depletion of alpha beta T cells by administration of anti-TCR beta mAb (H57-597) protected recipient mice from acute mortality and converted lethal listeriosis to sublethal infection . Furthermore, our findings suggested that gamma delta T cells were not involved in the liver necrosis in this condition . After depletion of alpha beta T cells, the number of bacteria decreased gradually to the limit for their detection (10(2) CFU) in 4 wks . Depletion of both alpha beta T cells and gamma delta T cells by administration of anti-TCR beta mAb (H57-597) and anti-TCR delta mAb (3A10) resulted in increased multiplication and persistent presence of bacteria in the liver and spleen . These findings suggest that gamma delta T cells play a significant protective role during infection in mice depleted of alpha beta T cells . In these mice, gamma delta T cells appeared with a peak on day 13 in the liver and on day 20 in the lymph nodes . No increase of gamma delta T cells in the spleen was observed throughout the course of infection.

Rev Sci Tech, 1995 Jun, 14(2), 313 - 41
Cleaning and disinfection practice in the meat industries of Europe; Salvat G et al.; The application and efficacy of cleaning and disinfection methods are reviewed, together with the relevant European and French legislation . European Commission Hygiene Directive 93/43/EEC of 14 June 1993 proposes the adoption of hazard analysis and critical control points (HACCP) for the meat industry, and this includes cleaning and disinfection . It is necessary to organise a team for washing, cleaning, rinsing, disinfection and final rinsing; three different types of organisation are compared . Application of HACCP and its contribution to the shelf life of products and their contamination with Listeria monocytogenes is discussed in the light of practical experience with poultry meat and cured pork products . Various means of verifying the efficacy of cleaning and disinfection (turbidimetry, adenosine triphosphate assay and macroscopic observation) are compared with the techniques of conventional microbiology . The authors conclude that cleaning and disinfection are essential for application of HACCP to the meat industry.

J Chemother, 1995 Jun, 7(3), 184 - 8
Erythromycin is ineffective against Listeria monocytogenes in multidrug resistant cells; Nichterlein T et al.; Multidrug resistance of tumor cells is a well-known phenomenon in oncology . Among the substances excluded from the cells are not only antineoplastic drugs but also certain antibiotics, e.g . erythromycin . To prove the hypothesis that this might render infections with intracellular bacteria untreatable with these antibiotics we used erythromycin to treat intracellular infection of multidrug resistant (MDR) cells with Listeria monocytogenes . Erythromycin was unable to restrict the growth of L . monocytogenes in KBV-1 MDR cells in concentrations of up to 25 micrograms/ml . In contrast, 0.049 micrograms/ml of erythromycin were sufficient to restrict the growth of the bacteria in nonresistant KB 3-1 cells . When verapamil was added to the supernatant of KBV-1 cells, erythromycin regained its effectivity on L . monocytogenes multiplying in these cells . The fact that MDR cells may render intracellular bacteria inaccessible to certain antibiotics might have important implications for the persistence of these bacteria in the host and for the treatment of patients with genetically engineered MDR cells.

Clin Infect Dis, 1995 Jun, 20(6), 1548 - 50
Joint infections due to Listeria monocytogenes: case report and review; Ellis LC et al.; Joint infections of bone are unusual manifestations of listerial infection . Fourteen cases of septic arthritis due to Listeria monocytogenes have been previously described in adults . We report the 15th case of septic arthritis due to Listeria in which bilateral prosthetic hips in a renal transplant patient were involved . In addition, we review the literature on listerial joint infections.

Biologicals, 1995 Jun, 23(2), 135 - 43
Listeria monocytogenes: a live vector able to deliver heterologous protein within the cytosol and to drive a CD8 dependent T cell response; Goossens PL et al.; After an introduction on the entry and lifestyle of Listeria monocytogenes within mammalian eucaryotic cells, this chapter gives a brief overview of murine experimental listeriosis . Among the main characteristics of this murine model of infectious/pathogenic processes initiated by a facultative intracellular bacteria, we point out two main recent advances . One relates to Listeria monocytogenes-induced production of cytokines as local, and transient signals able to direct the immune responses along a type 1 pathway of CD4/CD8 T cell differentiation . The other relates (a) to the recognition of L . monocytogenes-reactive CD8+ T lymphocytes as effectors able, once recruited within infected loci, to critically contribute to the complete clearance of the bacteria, and (b) to the recently recognized specificity of some of these CD8 lymphocytes in BALB/c mice . In this paper, we also briefly review (a) the readout assays presently used to monitor the outcome of the infectious/pathogenic processes and the related development and expression of immune responses induced by intravenous inoculation of wild-type virulent or attenuated L . Monocytogenes, (b) why all this information allows us to consider the use of L monocytogenes of attenuated virulence as relevant live recombinant vectors in order to deliver heterologous proteins to the class I processing and presentation pathway, and to induce CD8 T cells along the type 1 pathway.

J Immunol, 1995 Jun 1, 154(11), 5832 - 41
Activation of gamma delta T cells for production of IFN-gamma is mediated by bacteria via macrophage-derived cytokines IL-1 and IL-12; Skeen MJ et al.; Gamma delta T cells are found at sites of microbial infection and have been reported to proliferate in response to bacterial Ags . We show here that although the response by Listeria-elicited peritoneal gamma delta T cells to heat-killed bacteria in the presence of macrophage accessory cells may be partially mediated via the TCR, it is predominantly mediated via cytokines produced by the macrophages . Macrophage cytokines IL-12 and IL-1 synergize to induce some proliferation and considerable IFN-gamma production by peritoneal gamma delta T cells . This cytokine synergy pattern differs from that reported for NK cells, in which IL-12 in combination with either IL-2 or TNF-alpha induces NK cells to produce IFN-gamma . The combination of IL-12 and IL-1 provides a strong stimulus for IFN-gamma production by gamma delta T cells, but a relatively weak signal for proliferation . This is in contrast to the strong proliferative signal from the combination of IL-7 and IL-1 and the relatively weak stimulation of IFN-gamma production by the IL-7/IL-1 combination . Thus, there is differential regulation of NK and gamma delta T cells by cytokines and differential regulation of activation functions within the gamma delta T cell population by combinations of cytokines . These data provide evidence for a potentially important pathway for augmentation of IFN-gamma secretion at sites of infection where gamma delta T cells are found in abundance and where IFN-gamma may play a major role in the control of the infection.

Int J Immunopharmacol, 1995 Jun, 17(6), 505 - 12
Hot water extracts of Chlorella vulgaris reduce opportunistic infection with Listeria monocytogenes in C57BL/6 mice infected with LP-BM5 murine leukemia viruses; Hasegawa T et al.; The bacterial elimination after infection with Listeria monocytogenes was impaired in mice with murine acquired immunodeficiency syndrome (MAIDS) by infection with LP-BM5 murine leukemia virus . Oral administration of hot water extracts of Chlorella vulgaris (CVE) restored the capacity of MAIDS mice to eliminate L . monocytogenes in association with improvement of the deteriorated immune response to L . monocytogenes . DTH response to Listeria in CVE-treated MAIDS mice was significantly higher than that of MAIDS mice after Listeria infection in association with increases in number of CD4+CD8- and CD4-CD8+ alpha beta T-cells in the infected sites . CVE might be effective in the treatment of opportunistic infection in retrovirus-induced immunodeficient patients.

Proc Natl Acad Sci U S A, 1995 May 23, 92(11), 5234 - 8
Types of Listeria monocytogenes predicted by the positions of EcoRI cleavage sites relative to ribosomal RNA sequences; Hubner RJ et al.; By using taxonomic characters derived from EcoRI restriction endonuclease digestion of genomic DNA and hybridization with a labeled rRNA operon from Escherichia coli, a polymorphic structure of Listeria monocytogenes, characterized by fragments with different frequencies of occurrence, was observed . This structure was expanded by creating predicted patterns through a recursive process of observation, expectation, prediction, and assessment of completeness . This process was applied, in turn, to normalized strain patterns, fragment bands, and positions of EcoRI recognition sites relative to rRNA regions . Analysis of 1346 strains provided observed patterns, fragment sizes, and their frequencies of occurrence in the patterns . Fragment size statistics led to the creation of unobserved combinations of bands, predicted pattern types . The observed fragment bands revealed positions of EcoRI sites relative to rRNA sequences . Each EcoRI site had a frequency of occurrence, and unobserved fragment sizes were postulated on the basis of knowing the restriction site locations . The result of the recursion process applied to the components of the strain data was an extended classification with observed and predicted members.

Proc Natl Acad Sci U S A, 1995 May 23, 92(11), 5229 - 33
Sets of EcoRI fragments containing ribosomal RNA sequences are conserved among different strains of Listeria monocytogenes; Bruce JL et al.; To classify Listeria monocytogenes using taxonomic characters derived from the rRNA operons and their flanking sequences, we studied a sample of 1346 strains within the taxon . DNA from each strain was digested with a restriction endonuclease, EcoRI . The fragments were separated by gel electrophoresis, immobilized on a membrane, and hybridized with a labeled rRNA operon from Escherichia coli . The pattern of bands, positions, and intensities of hybridized fragments were electronically captured . Software was used to normalize the band positions relative to standards, scale the signal intensity, and reduce the background so that each strain was reproducibly represented in a data base as a pattern . With these methods, L . monocytogenes was resolved into 50 pattern types differing in the length of at least one polymorphic fragment . Pattern types representing multiple strains were recorded as the mathematical average of the strain patterns . Pattern types were arranged by size polymorphisms of assigned rRNA regions into subsets, which revealed the branching genetic structure of the species . Subtracting the polymorphic variants of a specific assigned region from the pattern types and averaging the types within each subset resulted in reduced sets of conserved fragments that could be used to recognize strains of the species . Pattern types and reduced sets of conserved fragments were conserved among different strains of L . monocytogenes but were not observed in total among strains of other species.

Cell, 1995 May 19, 81(4), 641 - 50
Altered responses to bacterial infection and endotoxic shock in mice lacking inducible nitric oxide synthase; MacMicking JD et al.; Mice deficient in inducible nitric oxide synthase (iNOS) were generated to test the idea that iNOS defends the host against infectious agents and tumor cells at the risk of contributing to tissue damage and shock . iNOS-/-mice failed to restrain the replication of Listeria monocytogenes in vivo or lymphoma cells in vitro . Bacterial endotoxic lipopolysaccharide (LPS) caused shock and death in anesthetized wild-type mice, but in iNOS-/-mice, the fall in central arterial blood pressure was markedly attenuated and early death averted . However, unanesthetized iNOS-/-mice suffered as much LPS-induced liver damage as wild type, and when primed with Propionobacterium acnes and challenged with LPS, they succumbed at the same rate as wild type . Thus, there exist both iNOS-dependent and iNOS-independent routes to LPS-induced hypotension and death.

Lett Appl Microbiol, 1995 May, 20(5), 295 - 9
Listeriolysin O secretion by Listeria monocytogenes in the presence of cysteine and sorbate; Kouassi Y et al.; The influence of cysteine or cysteine-HCl and their combination with potassium sorbate on growth of Listeria monocytogenes and listeriolysin O (LLO) secretion, and on activation of LLO in the haemolysin assay system was studied . Both cysteine and cysteine-HCl (10 and 20 mmol l-1) enhanced LLO secretion in tryptic soy broth supplemented with yeast extract during 24 h incubation at 35 degrees C . While sorbate did not affect growth, it suppressed both LLO secretion and LLO activation by cysteine in the haemolysin activity assay . These findings provide further evidence that sulphydryl-containing enzymes are implicated in the mechanism of microbial inhibition of sorbate . Addition of sorbate to foods has the potential of inactivating listeriolysin and other thiol-dependent toxins.

Infect Immun, 1995 May, 63(5), 1876 - 86
Intracellular killing of Listeria monocytogenes in the J774.1 macrophage-like cell line and the lipopolysaccharide (LPS)-resistant mutant LPS1916 cell line defective in the generation of reactive oxygen intermediates after LPS treatment; Inoue S et al.; Listeria monocytogenes is a facultative intracellular pathogen and survives within phagocytic cells by escaping from phagosomes into the cytoplasm . It has been reported that, in vivo, L . monocytogenes is effectively eliminated through cell-mediated immunity, especially by macrophages which have been immunologically activated by cytokines such as gamma interferon (IFN-gamma) . However, this killing mechanism for L . monocytogenes and the role of macrophage activation in this bacterial killing are unclear . We demonstrated the listericidal effect of oxidative radicals induced by lipopolysaccharide (LPS) and IFN-gamma, using a macrophage-like cell line, J774.1, and a mutant cell line, LPS1916 . LPS1916 cells do not exhibit normal generation of O2- and H2O2 after treatment with 0.1 microgram of LPS per ml, although J774.1 cells generate 100 times the normal level of oxidative radicals with the same LPS treatment . The growth of L . monocytogenes was strongly inhibited in J774.1 cells pretreated with 0.1 microgram of LPS per ml or the combination of 0.1 microgram of LPS per ml and 10 U of IFN-gamma per ml . On the other hand, in LPS1916 cells, the growth of L . monocytogenes was not inhibited by treatment with LPS only, although LPS1916 cells pretreated with the combination of LPS and IFN-gamma showed moderate inhibition of listerial growth . This killing was not influenced by treatment with NG-monomethyl-L-arginine, which is a strong inhibitor of nitrite oxide generation . Interestingly, J774.1 cells treated with LPS did not show enhanced intraphagosomal killing of a nonhemolytic strain of avirulent L . monocytogenes that lacks the ability to escape from phagosomes, and this killing was not influenced by treatment with NG-monomethyl-L-arginine either . These results suggest that the reactive oxygen radicals are more important than nitric oxide in the mechanism underlying the intracellular killing of virulent L . monocytogenes and that there seem to be different killing mechanisms for virulent and avirulent strains of L . monocytogenes in activated-macrophage cell lines.

J Immunol, 1995 May 1, 154(9), 4642 - 50
CD8 T lymphocytes specific for the secreted p60 antigen protect against Listeria monocytogenes infection; Harty JT et al.; The ability of Listeria monocytogenes to gain access to the cytoplasm of infected cells results in the processing and presentation of bacterial Ags through the MHC class I pathway . As a result, CD8 T cells are the most effective mediators of acquired immunity in the mouse model of L . monocytogenes infection . CD8 T cells specific for a single nonamer epitope derived from the secreted virulence factor listeriolysin O (LLO) can protect H-2d mice against lethal infection . Bacteria lacking LLO are avirulent and do not elicit protective immunity in mice . Thus, the failure of LLO minus L . monocytogenes to elicit protective immunity could be caused either by their inability to enter the cytoplasm or to the lack of LLO-derived peptide epitopes . In this report we provide evidence that H-2d restricted CD8 T cells with specificity for another L . monocytogenes protein, the secreted p60 molecule, can protect against infection . Our studies further demonstrate that LLO-dependent induction of protective immunity results from access of the bacterium to the cytoplasm . In addition, these studies provide support for the hypothesis that secreted bacterial proteins are the most important targets for protective CD8 T cell-mediated immunity.

Appl Environ Microbiol, 1995 May, 61(5), 2013 - 5
Adhesion of Listeria monocytogenes to silica surfaces after sequential and competitive adsorption of bovine serum albumin and beta-lactoglobulin; al-Makhlafi H et al.; Adsorbed bovine serum albumin was resistant to exchange with beta-lactoglobulin, and when albumin was adsorbed from a mixture, its surface concentration increased with time . The passivating character of adsorbed albumin and its resistance to desorption were consistent with the level of Listeria monocytogenes adhesion evoked by albumin-containing protein films.

Pediatr Hematol Oncol, 1995 May-Jun, 12(3), 295 - 9
Listeria septicemia complicating bone marrow transplantation for Diamond-Blackfan syndrome; Lee AC et al.; Infection with Listeria monocytogenes is uncommon in patients receiving cytotoxic chemotherapy, and is even rarer among recipients of bone marrow transplantation . Hemosiderosis, either idiopathic or caused by transfusion, appears to be another risk factor . We report a 3-year-old Chinese girl with transfusion-dependent Diamond-Blackfan syndrome who had L . monocytogenes septicemia when she received an allogeneic bone marrow transplantation . She was treated successfully with intravenous ampicillin . Our case adds to the clinical evidence that patients with iron overload are susceptible to listeriosis, particularly when they are immunocompromised and do not receive iron-chelation treatment.

Rev Esp Enferm Dig, 1995 May, 87(5), 407 - 11
{Spontaneous bacterial peritonitis due to Listeria monocytogenes}; Vazquez J et al.; We present a case of spontaneous bacterial peritonitis (SBP) caused by Listeria monocytogenes in a patient previously diagnosed as alcoholic liver cirrhosis . The clinical presentation, biochemical data and outcome of the patient are compared with those of cases of SBP caused by Listeria monocytogenes in patients with cirrhosis published in the Spanish and English literature . Twelve out of 20 cases described in the literature were published by Spanish authors . This greater proportion could be related to dietary habits (greater consumption of fruits and vegetables), climatic or demographic factors . We underline the importance of pursuing a microbiological diagnosis since Listeria monocytogenes is intrinsically resistant to cefotaxime, the antimicrobial often selected to empirically treat SBP episodes.

Mt Sinai J Med, 1995 May, 62(3), 216 - 20
Listeria monocytogenes infections: laboratory investigation of an unexpected increase in incidence; Bottone EJ et al.; Listeria monocytogenes is a facultative intracellular pathogen causing mainly meningitis and septicemia in immunocompromised hosts . From July 1988 through December 1989, 16 patients at The Mount Sinai Hospital were diagnosed as having listeriosis shortly after admission, 14 within a one-year period (July 1988-July 1989) . Because this incidence was almost double the incidence in previous years (< 8 annually), an epidemiologic and microbiologic investigation was undertaken to determine a potential route of acquisition of L . monocytogenes . On the basis of plasmid profile, bacteriocin (enterocin) susceptibility pattern, and serotype, no single epidemic strain could be identified . Although direct evidence was lacking, we concluded that our patients may have acquired L . monocytogenes through transient contamination of food.

Arzneimittelforschung, 1995 May, 45(5), 614 - 6
Effects of leflunomide on the course of listeriosis in mice . Short communication; Hirschelmann R et al.; The intensity of an infection with Listeria monocytogenes depends on the immunological stab1p4f the body . Impairment of the specific or unspecific defence can lead to severe and partly fatal complications such as meningitis or septicaemia . In a normal defence situation affected individuals are often not diagnosed because the infection appears as an unspecific indisposition . The new immunomodulator leflunomide (n-(4-trifluoro-methyl-phenyl)-5-methyl-isoxazol-4-carboxamide, CAS 75706-12-6, HWA 486) acts mainly as immunosuppressive . Therefore, the influence of leflunomide was investigated on the number of spleen bacteria in the first phase and on the lethality in both phases of mouse listeriosis . The infected mice were either in a normal defence state or in an immunocompromised situation, which was produced by injection of cyclophosphamide or by dextran sulfate . A treatment with leflunomide, 20 mg/kg i.p., twice daily over a period of 3 days, caused no change of the lethality compared with control mice . Likewise, combined administration of leflunomide and ampicillin produced no alteration of the lethal outcomes in comparison with the ampicillin medication alone, the latter being highly effective in any case . However, the number of spleen bacteria was significantly higher in the group with combined treatment than in the ampicillin group . In summary, the treatment with leflunomide most likely will cause no additional risk regarding an infection such as listeriosis, although in the case of an infection an increased care should be given to diagnostic and therapy.

Immunol Lett, 1995 May, 46(1-2), 111 - 6
Listeria monocytogenes infection increases neutrophil adhesion and damage to a murine hepatocyte cell line in vitro; Boury NM et al.; Several studies have reported that Listeria monocytogenes multiples within hepatocytes and that inflammatory neutrophils inhibit this intracellular growth in vivo . In the present study, we used a murine embryonic hepatocyte cell line (ATCC TIB73) as an in vitro model to investigate neutrophil-hepatocyte interactions . Murine peritoneal exudate neutrophils adhered more readily to L . monocytogenes-infected hepatocyte monolayers than to uninfected monolayers or monolayers infected with actA- and hly- mutants of L . monocytogenes . L . monocytogenes-infected TIB73 cells increased their surface expression of ICAM-1 as compared with uninfected TIB73 cells . Neutrophil adherence and oxidative stress to TIB73 cells were reduced by pre-incubating the hepatocyte monolayers with anti-ICAM-1 monoclonal antibody and diminished further by pre-incubating the peritoneal exudate neutrophils with an anti-CR3 monoclonal antibody.

Nat Med, 1995 May, 1(5), 471 - 7
A recombinant Listeria monocytogenes vaccine expressing a model tumour antigen protects mice against lethal tumour cell challenge and causes regression of established tumours; Pan ZK et al.; Listeria monocytogenes is an intracellular organism that has the unusual ability to live in the cytoplasm of the cell . It is thus a good vector for targeting protein antigens to the cellular arm of the immune response . Here we use a model system, consisting of colon and renal carcinomas that express the influenza virus nucleoprotein and a recombinant L . monocytogenes that secrets this antigen, to test the potential of this organism as a cancer immunotherapeutic agent . We show that this recombinant organism can not only protect mice against lethal challenge with tumour cells that express the antigen, but can also cause regression of established macroscopic tumours in an antigen-specific T-cell-dependent manner.

Curr Biol, 1995 May 1, 5(5), 517 - 25
The bacterial actin nucleator protein ActA of Listeria monocytogenes contains multiple binding sites for host microfilament proteins; Pistor S et al.; BACKGROUND: Several intracellular pathogens, including Listeria monocytogenes, use components of the host actin-based cytoskeleton for intracellular movement and for cell-to-cell spread . These bacterial systems provide relatively simple model systems with which to study actin-based motility . Genetic analysis of L . monocytogenes led to the identification of the 90 kD surface-bound ActA polypeptide as the sole bacterial factor required for the initiation of recruitment of host actin filaments . Numerous host actin-binding proteins have been localized within the actin-based cytoskeleton that surrounds Listeria once it is inside a mammalian cell, including alpha-actinin, fimbrin, filamin, villin, ezrin/radixin, profilin and the vasodilator-stimulated phosphoprotein, VASP . Only VASP is known to bind directly to ActA . We sought to determine which regions of the ActA molecule interact with VASP and other components of the host microfilament system . RESULTS: We used the previously developed mitochondrial targeting assay to determine regions of the ActA protein that are involved in the recruitment of the host actin-based cytoskeleton . By examining amino-terminally truncated ActA derivatives for their ability to recruit cytoskeletal proteins, an essential element for actin filament nucleation was identified between amino acids 128 and 151 of ActA . An ActA derivative from which the central proline-rich repeats were deleted retained its ability to recruit filamentous actin, albeit poorly, but was unable to bind VASP . CONCLUSIONS: Our studies reveal the initial interactions that take place between invading Listeria and host microfilament proteins . The listerial ActA polypeptide contains at least two essential sites that are required for efficient microfilament assembly: an amino-terminal 23 amino-acid region for actin filament nucleation, and VASP-binding proline-rich repeats . Hence, ActA represents a prototype actin filament nucleator . We suggest that host cell analogues of ActA exist and are important components of structures involved in cell motility.

Int Immunol, 1995 May, 7(5), 797 - 805
Attenuated Listeria monocytogenes as a live vector for induction of CD8+ T cells in vivo: a study with the nucleoprotein of the lymphocytic choriomeningitis virus; Goossens PL et al.; Listeria monocytogenes spends most of its intracellular life cycle in the cytosol of the infected eucaryotic cells . Within this cellular compartment originates the endogenous pathway of antigen processing and presentation . We thus assumed that recombinant L . monocytogenes expressing an heterologous protein, the nucleoprotein of the lymphocytic choriomeningitis virus (LCMV), should be able to induce antigen-specific CD8+ T cells in vivo . The LCMV nucleoprotein gene was inserted in phase with the sequence coding for the putative signal sequence of the hemolysin of L . monocytogenes in order to target its secretion into the cytosol of the infected cell . The ability of this recombinant bacterium to induce LCMV-reactive CD8+ T cells was then monitored in BALB/c mice . The immune status of the immunized BALB/c mice was studied on the seventh day after a single i.v . injection of a sublethal dose of the recombinant bacteria: (i) cytotoxic CD8+ T cells were detected in liver; (ii) using in vitro re-stimulation with PMA and ionomycin, secondary cytotoxic CD8+ T cells were detected in spleen; (iii) an early inflammatory reaction dependent on the presence of CD8+ T cells occurred in the footpad after intraplantar inoculation of live LCMV; and (iv) mice were protected against an otherwise lethal intracerebral LCMV challenge; the protection was accompanied by elimination of the virus . When the immune status of the immunized hosts was monitored for a longer period post-immunization, the balance between immune protection and immunopathology described for the anti-LCMV immune responses was observed; two phases of protection were detected, flanking a transitory phase of exacerbation of the lymphocytic choriomeningitis disease (weeks 2-5).(ABSTRACT TRUNCATED AT 250 WORDS)

Microb Pathog, 1995 May, 18(5), 355 - 64
Non-dystrophic 129 REJ mice are susceptible to i.p . infection with Listeria monocytogenes despite an ability to recruit inflammatory neutrophils to the peritoneal cavity; Gahan CG et al.; In this study we compared the host response to Listeria monocytogenes in 129 REJ mice with listeria-resistant (C57Bl/6j) and susceptible (Balb/c) mouse strains . In all experiments mice were inoculated by the i.p . route . 129 REJ mice and Balb/c mice were sensitive to listeriosis whilst C57Bl/6j mice were relatively resistant to i.p . infection . Relatively large numbers of viable bacteria could be detected in the spleens of 129 REJ mice as early as 6 h following i.p . inoculation suggesting that dissemination of listeria from the peritoneal cavity is rapid in this mouse strain . This contrasted with Balb/c mice which exhibited an early lag phase during which only low numbers of bacteria could be isolated from the spleens of infected animals . In response to both proteose peptone and live listeria, 129 REJ mice demonstrated a greater capacity to recruit neutrophils to the peritoneal cavity than Balb/c and C57Bl/6j mice . In addition, inflammatory phagocytes from 129 REJ mice were as bactericidal in vitro as phagocytes from the Balb/c and C57Bl/6j strains . However, in vivo, inflammatory neutrophils elicited by proteose peptone prior to i.p . infection with L . monocytogenes were not protective in the three mouse strains tested . Despite the apparent inadequacy of peritoneal neutrophils in controlling early bacterial proliferation, depletion of neutrophils in 129 REJ mice severely exacerbated i.p . infection with L . monocytogenes . The results indicate that neutrophils provide an inefficient but essential means of controlling early outgrowth of listeria in the peritoneal cavity of 129 REJ mice . The excessive inflammatory response seen in 129 REJ mice may facilitate the early dissemination of L . monocytogenes from the peritoneal cavity to peripheral sites.

Microb Pathog, 1995 May, 18(5), 323 - 36
Bacteremia is required for invasion of the murine central nervous system by Listeria monocytogenes; Berche P; The ability of the facultative intracellular pathogen Listeria monocytogenes to penetrate the central nervous system (CNS) was studied by following the kinetics of brain invasion and histological lesions during an acute intravenous (i.v.) infection in the mouse . CNS invasion occurred during the early phase of infection and produced severe meningoencephalitis characterized by multiple granulomatous foci predominantly located in the brainstem and associated with diffuse meningitis and an intense inflammatory reaction involving the choroid plexuses . Bacterial counts in the brain could reach 10(4.5)-10(5.8) by day 5 of infection with 1-2 x 10(6) bacteria i.v., depending upon the bacterial strain used . It was found that CNS invasion was highly dependent upon the level and the duration of bacteremia, thus indicating that persistent bacteremia is essential to induce meningoencephalitis to L . monocytogenes.

Proc Natl Acad Sci U S A, 1995 Apr 25, 92(9), 3987 - 91
Recombinant Listeria monocytogenes as a live vaccine vehicle for the induction of protective anti-viral cell-mediated immunity; Shen H et al.; Listeria monocytogenes (LM) is a Gram-positive bacterium that is able to enter host cells, escape from the endocytic vesicle, multiply within the cytoplasm, and spread directly from cell to cell without encountering the extracellular milieu . The ability of LM to gain access to the host cell cytosol allows proteins secreted by the bacterium to efficiently enter the pathway for major histocompatibility complex class I antigen processing and presentation . We have established a genetic system for expression and secretion of foreign antigens by recombinant strains, based on stable site-specific integration of expression cassettes into the LM genome . The ability of LM recombinants to induce protective immunity against a heterologous pathogen was demonstrated with lymphocytic choriomeningitis virus (LCMV) . LM strains expressing the entire LCMV nucleoprotein or an H-2Ld-restricted nucleoprotein epitope (aa 118-126) were constructed . Immunization of mice with LM vaccine strains conferred protection against challenge with virulent strains of LCMV that otherwise establish chronic infection in naive adult mice . In vivo depletion of CD8+ T cells from vaccinated mice abrogated their ability to clear viral infection, showing that protective anti-viral immunity was due to CD8+ T cells.

J Acquir Immune Defic Syndr Hum Retrovirol, 1995 Apr 15, 8(5), 461 - 5
Listeria monocytogenes infection and serotype distribution among HIV-infected persons in Los Angeles County, 1985-1992; Ewert DP et al.; Persons infected with human immunodeficiency virus (HIV) are at greater risk of infection with Listeria monocytogenes (LM) than the general population . We quantify the risk of listeriosis in persons with acquired immune deficiency syndrome (AIDS) and HIV infection in Los Angeles County (LAC) and report the LM serotype distribution among HIV-infected patients with listeriosis . Active surveillance for listeriosis was performed in LAC during most of the period from 1985 through 1992 . Thirty-four (10%) of 351 nonperinatal cases of listeriosis reported in LAC from 1985 through 1992 were in HIV-infected persons, 25 of whom met the 1987 AIDS case definition . The incidence of listeriosis was 95.8 and 8.8 cases per 100,000 person-years among persons with AIDS and all HIV-infected persons, respectively, but only 1.0 case per 100,000 person-years in the total population . Excluding cases from a 1985 listeriosis epidemic associated with consumption of contaminated Mexican-style cheese, 11 (65%) of 17 HIV-infected persons with available isolates were infected with LM serotype 1/2b, whereas only 64 (31%) of 208 other persons with listeriosis and available isolates were infected with LM serotype 1/2b (odds ratio = 4.1; 95% confidence interval = 1.3-14.1) . LM serogroup 1/2b may have been more common among HIV-infected persons in LAC than among other persons with listeriosis because of differences in diet or sexual practices, or to chance alone . Persons with HIV-infection, especially those with AIDS, should be educated in avoiding foods at high risk of listerial contamination, such as soft cheeses, foods sold from delicatessen counters, and undercooked chicken.

JAMA, 1995 Apr 12, 273(14), 1118 - 22
Reduction in the incidence of human listeriosis in the United States . Effectiveness of prevention efforts? The Listeriosis Study Group; Tappero JW et al.; BACKGROUND--Food-borne transmission is now recognized as a major cause of human listeriosis . OBJECTIVE--To assess the impact of prevention efforts, listeriosis rates before interventions were initiated in 1989 were compared with more recent rates (1990 through 1993) . DESIGN--From 1989 through 1993, multistate, laboratory-based active surveillance was conducted to identify all cases in which Listeria monocytogenes was isolated from cultures or ordinarily sterile sites in an aggregate population of more than 19 million . SETTING--All laboratories serving acute care hospitals in up to nine surveillance areas in the United States . INTERVENTIONS--In 1989, a well-publicized case report of listeriosis linked to processed poultry led US regulatory agencies to enforce aggressive food monitoring policies and prompted industry to invest in cleanup efforts . In May 1992, consumer guidelines for listeriosis prevention were disseminated . OUTCOME MEASURES--Cases of perinatal and nonperinatal listeriosis . RESULTS--The rate of listeriosis decreased in all surveillance areas . Projection of these rates to the US population suggests an estimated 1965 cases and 481 deaths occurred in 1989 compared with an estimated 1092 cases and 248 deaths in 1993, a 44% and 48% reduction in illness and death, respectively . Among adults 50 years of age and older, rates declined from 16.2 per 1 million in 1989 to 10.2 per 1 million in 1993 (P = .02) . Perinatal disease decreased from 17.4 cases per 100,000 births in 1989 to 8.6 cases per 100,000 births in 1993 (P = .003) . Three serotypes (1/2a, 1/2b, and 4b) of L monocytogenes accounted for more than 96% of cases during each year of the study (1989 through 1993) . CONCLUSIONS--The incidence of listeriosis in study areas was substantially lower in 1993 than in 1989 . The temporal association of this reduction with industry, regulatory, and educational efforts suggests these measures were effective.

Infect Immun, 1995 Apr, 63(4), 1595 - 7
Colony-stimulating factor 1 in the human response to neonatal listeriosis; Grieg A et al.; Immunity to Listeria monocytogenes, an important pathogen in human neonatal sepsis, is mediated by mononuclear phagocytes, which are regulated by the hematopoietic growth factor colony-stimulating factor 1 . Neonates with listeriosis had higher circulating colony-stimulating factor 1 levels and subsequent monocyte counts than those of both noninfected newborns and newborns infected with nonlisterial organisms.

J Rheumatol, 1995 Apr, 22(4), 786 - 7
Listeria monocytogenes infection in a patient treated with methotrexate for rheumatoid arthritis; McCambridge MM et al.; We describe a patient with rheumatoid arthritis who developed bacteremia from Listeria monocytogenes after treatment with low dose oral pulse methotrexate . We discuss possible immunologic mechanisms for susceptibility to Listeria infections . As the elderly population increases with more frequent use of immunosuppressive medications, clinical suspicion must be maintained to correctly diagnose and treat infections such as Listeria.

Lett Appl Microbiol, 1995 Apr, 20(4), 195 - 8
The inhibitory properties of various sponges on Listeria spp; Daley EF et al.; Various retail and environmental sponges were tested for inhibitory properties against Listeria species and several other bacterial genera . Sterile sponges, unrinsed and rinsed in sterile distilled water or sterile neutralizing buffer, were placed on seeded plates of tryptic soy agar with 0.6% yeast extract . Plates were incubated at 30 degrees C for 24 h and zones of inhibition measured . The Systems Plus environmental sponge and the Technical Service Consultants Ltd sponge (sTc) proved to be the only sponges which consistently demonstrated no inhibitory properties . Results using scanning electron microscopy showed considerable bacterial attachment to the Systems Plus sponge, further corroborating these findings.

Appl Environ Microbiol, 1995 Apr, 61(4), 1661 - 5
Characterization of five esterases from Listeria monocytogenes and use of their electrophoretic polymorphism for strain typing; Gilot P et al.; Esterases from Listeria monocytogenes strains isolated from cheeses were analyzed by starch gel electrophoresis . Five esterases, numbered from EST 1 to EST 5 in order of decreasing anodal migration, were identified . The EST 1, EST 3, EST 4, and EST 5 set was most active toward alpha-naphthyl propionate, while EST 2 was most active toward alpha-naphthyl acetate . Results from inhibitor studies suggest that all of these esterases were EC class 3.1.1.1 carboxylesterases, except that EST 1 and EST 3 also showed some sensitivity to parahydroxymercuribenzoate . Polymorphism of these five esterases was observed in the population . Twelve esterase patterns were defined and used to subdivide serotypes.

Appl Environ Microbiol, 1995 Apr, 61(4), 1643 - 5
Utilization of exogenous siderophores and natural catechols by Listeria monocytogenes; Simon N et al.; Listeria monocytogenes does not produce siderophores for iron acquisition . We demonstrate that a number of microbial siderophores and natural iron-binding compounds are able to promote the growth of iron-starved L . monocytogenes . We suggest that the ability of L . monocytogenes to use a variety of exogenous siderophores and natural catechols accounts for its ubiquitous character.

Appl Environ Microbiol, 1995 Apr, 61(4), 1514 - 9
Efficacy of marinades against Listeria monocytogenes cells in suspension or associated with green shell mussels (Perna canaliculus); Bremer PJ et al.; In order to determine the listericidal efficacies of three marinades used in the production of marinated green shell mussels (Perna canaliculus), decimal reduction times (D values) were determined for a mixture of seven strains of Listeria monocytogenes exposed to marinades in the presence and absence of mussels . With an acetic acid (1.5%, wt/vol) marinade, calculated D values in the presence and absence of mussels were 77.3 and 33.3 h, respectively . Likewise, for an acetic acid (0.75%)-lactic acid (0.75%) marinade and an acetic acid (1.5%)-Glucono Delta-Lactone (0.2%)-based marinade, the D values in the presence and absence of mussels were 125.5 and 26.9 h and 86.3 and 19.3 h, respectively . Various increases in decimal reduction times in the presence of mussels indicated that there was no simple relationship between the listericidal natures of these marinades and the presence of mussels . This result suggests that difficulties may occur in trying to relate acid inhibition studies carried out in model broth systems to "real food" systems.

J Appl Bacteriol, 1995 Apr, 78(4), 366 - 72
Evaluation of enrichment broths for their ability to recover heat-injured Listeria monocytogenes; Patel JR et al.; Listeria selective enrichment broth (LEB), University of Vermont (UVM) broth, modified UVM (MUVM) broth and Fraser broth (FB) were compared for their ability to recover cells of L . monocytogenes from heated tryptose phosphate broth . Three strains of L . monocytogenes were heated at 54 degrees C for 30 min, inoculated into enrichment broths supplemented with 400 micrograms catalase ml-1, and incubated for 8 h at 30 degrees C . After incubation for 4 h, the total viable cell populations either decreased or did not change, whereas the number of healthy (non-injured) cells of all strains increased significantly in all broths except FB inoculated with the LCDC strain . With an increase in incubation time to 8 h, the number of healthy cells of all strains increased in all broths . At 8 h, the difference between populations of total (injured plus healthy cells) and healthy cells detected in LEB inoculated with two strains was not significant . Overall, recovery of heat-treated cells was significantly higher in LEB, followed by MUVM broth, UVM broth and FB . The addition of catalase to enrichment broths significantly enhanced recovery of heat-injured cells . A slight reduction of catalase activity of heated cells of all test strains in all enrichment broths except FB was observed by extending the incubation period from 4 to 8 h . A test strain that produces relatively higher catalase activity compared to the other strains exhibited the greatest resistance to exogenous hydrogen peroxide . Enumeration of viable L . monocytogenes cells in heated foods should be done using LEB supplemented with 400 micrograms catalase ml-1 to maximize the recovery of injured cells.

J Clin Invest, 1995 Apr, 95(4), 1628 - 34
Immune complexes inhibit antimicrobial responses through interleukin-10 production . Effects in severe combined immunodeficient mice during Listeria infection; Tripp CS et al.; The presence of soluble antigen-antibody complexes renders mice highly susceptible to infection with the intracellular pathogen Listeria monocytogenes . In this report we show that this inhibition is manifest at the level of the innate immune response and is mediated by IL-10 . Like immuno-competent mice, mice with the severe combined immunodeficient mutation (SCID) injected with immune complexes died from a sublethal dose of L . monocytogenes . These mice were protected if pretreated with neutralizing antibodies to IL-10 . In vitro, immune complexes stimulated IL-10 production by SCID splenocytes and splenic macrophages . Likewise, immune complexes inhibited TNF and IFN-gamma production by SCID splenocytes cultured with heat-killed-L . monocytogenes . This inhibition was reversed by neutralization of IL-10 but not IL-4 or TGF-beta . Immune complexes and rIL-10 inhibited cytokine production by SCID splenocytes if added before or simultaneously with heat-killed-L . monocytogenes . These data support a model in which immune complexes modulate host defense and the immune response by stimulating the production of IL-10 from macrophages.

Vet Hum Toxicol, 1995 Apr, 37(2), 110 - 2
The influence of heavy metal emissions and Fasciola hepatica infestation on the immunogenicity of a Listeria vaccine; Pistl J et al.; The effect of oral dosing with emissions containing heavy metals (mercury, lead, copper, cadmium, zinc and chromium) on immune status was tested in 3 groups of sheep . Two groups (Groups I and II) were given emissions for 15 d . One of these groups (Group I) was then also infested with Fasciola hepatica metacercaria on day 16 . These 2 groups (Groups I and II) and a control group (Group C) were immunized with a Listeria vaccine on days 8 and 22 . A decreased index of metabolic response (IMR) of phagocytes and reduced responses of lymphocytes to mitogenic activation with phytohemagglutinin and L monocytogenes antigen stimulation in the leukocyte migration-inhibition test were recorded . Decreased agglutination titers of serum antibodies against L monocytogenes were observed . The F hepatica infestation had no significant effect on the migration index or IMR.

Oral Surg Oral Med Oral Pathol Oral Radiol Endod, 1995 Apr, 79(4), 442 - 8
The antiviral spectrum of Listerine antiseptic; Dennison DK et al.; The mechanism of activity and the antiviral spectrum of Listerine antiseptic have not been examined thoroughly . We therefore tested its effect on laboratory strains of herpes simplex type 1 and type 2 (enveloped DNA viruses), influenza A virus (enveloped RNA virus), rotavirus (nonenveloped RNA virus), and adenovirus type 5 (nonenveloped DNA virus) . Each virus was mixed with an equal volume of Listerine for 30 seconds to 5 minutes, and the residual infectivity of the virus was assessed . An antiviral effect was defined as greater than 95% reduction of infectivity . Exposure to Listerine for 30 seconds had an antiviral effect against herpes simplex type-1 and type-2 (96.3% and 100% reduction in infectious virus, respectively) and influenza A (100% reduction) . In contrast, rotavirus-induced plaque formation was reduced by 12.2% after 30 seconds of exposure to Listerine, whereas 5 minutes of exposure to Listerine resulted in a 21.5% increase in plaque formation . Exposure of adenovirus to Listerine had a minimal effect on the cytopathocity of the virus, with a 33.4% reduction in virus levels after 5 minutes . The antiviral activity of Listerine is thus not related to the viral genome but is probably directed to the viral envelope.

Mol Microbiol, 1995 Apr, 16(2), 251 - 61
Entry of Listeria monocytogenes into hepatocytes requires expression of inIB, a surface protein of the internalin multigene family; Dramsi S et al.; The intracellular bacterium Listeria monocytogenes can invade several types of normally non-phagocytic cells . Entry into cultured epithelial cells requires the expression of inIA, the first gene of an operon, comprising two genes: inIA, which encodes internalin, an 800-amino-acid protein, and inIB, which encodes a 630-amino-acid protein . Several genes homologous to inIA are detected in the genome of L . monocytogenes; InIB is one of them . We have assessed the role of inIB in invasiveness of L . monocytogenes by constructing isogenic chromosomal deletion mutants in the inIAB locus . Our findings indicate that: i) inIB is required for entry of L . monocytogenes into hepatocytes, but not into intestinal epithelial cells; ii) inIB encodes a surface protein; iii) internalin plays a role for entry into some hepatocyte cell lines . These results provide the first insight into the cell tropism displayed by L . monocytogenes.

Int J Food Microbiol, 1995 Apr, 25(2), 169 - 77
A method and medium for the electrical detection of Listeria spp . from food; Capell CJ et al.; The development of a liquid medium for the detection of Listeria spp . by capacitance monitoring of food samples previously enriched in UVM 1 broth is described . Rapid growth of Listeria monocytogenes was shown to occur in liquid media with selectivity based on antibiotics found in Oxford agar . The final capacitance medium contained higher concentrations of the Oxford selective agents than Oxford agar and did not require the esculin/ferric ammonium citrate reaction to be observed . The medium relied upon the ability of Listeria spp . to induce a greater than 30% change in capacitance within 30 h . When run in parallel with the Listeria spp . test samples of a large food company, the method gave far fewer false-positive results than Fraser broth.

J Vet Diagn Invest, 1995 Apr, 7(2), 223 - 8
Evaluation of laboratory tests for confirming the diagnosis of encephalitic listeriosis in ruminants; Johnson GC et al.; Retrospective analysis of 93 bovine, ovine, and caprine cases diagnosed as listerial encephalitis revealed positive bacterial isolations in only 63% of 59 cases in which bacterial culture was attempted . Only 42% of 41 attempted bovine brain cultures were successful, compared with 67% from 6 sheep brains and 92% from 12 goat brains . Gram stains and Listeria-specific immunohistochemistry were evaluated as tools for verifying the presence of bacteria or listerial antigens in 38 animals . Sixteen of 17 animals in this group with positive bacterial isolations were immunochemically positive for listerial antigens (including 5/6 cattle), but Gram stains detected only 9/17 positive animals (including 1/6 cattle) . Antigen was also detected in 15 of 21 animals (including 5/9 cattle) with unsuccessful or unattempted bacterial isolations . Of all 38 animals, the histologic diagnosis could be verified in 82% by immunohistochemistry, compared to 47% verified by Gram stains . Immunohistochemical testing was especially beneficial in locating antigen in lesions with few bacteria or bacterial antigens and is a rapid method of confirming the diagnosis of encephalitic listeriosis where inappropriate material is submitted for bacterial isolation or in culture-negative cases.

Zentralbl Veterinarmed B, 1995 Apr, 42(2), 84 - 8
Studies of the detection of Listeria monocytogenes by culture and PCR in cerebrospinal fluid samples from ruminants with listeric encephalitis; Peters M et al.; A total of 14 cerebrospinal fluid (CSF) samples from ruminants clinically suspected of suffering from listeric encephalitis were examined by polymerase chain reaction (PCR) for the detection of Listeria monocytogenes (L . m.) . Of these samples, 11 were examined bacteriologically . Although the clinical diagnosis was confirmed in eight of 11 ruminants by histological and/or bacteriological examination of the brains, L . m . was only detected in one of the CSF samples using PCR, and in none by culture . The PCR-positive CSF sample was obtained from a sheep which had been treated with antibiotics prior to CSF sampling . From these findings, it was concluded that L . m . only occasionally gains access to the meningoventricular system in the course of listeric encephalitis of ruminants and that a reliable aetiological in vivo diagnosis of listeric encephalitis generally cannot be based on the detection of L . m . in the CSF of affected ruminants.

FEBS Lett, 1995 Mar 27, 362(1), 65 - 9
Prophenin-1, an exceptionally proline-rich antimicrobial peptide from porcine leukocytes; Harwig SS et al.; We purified and characterized an unusual antimicrobial peptide, prophenin-1 (PF-1), from porcine leukocytes . The peptide had a mass of 8,683 and contained 79 residues, including 42 (53.2%) prolines and 15 (19.0%) phenylalanines . Its N-terminal 60 residues consisted of three perfect and three nearly perfect repeats of a decamer, FPPPNFPGPR . Prophenin-1 was encoded on a cathelin-containing precursor and showed substantially more activity against E . coli, a Gram-negative bacterium, than against Listeria monocytogenes, a Gram-positive organism, in vitro.

Ugeskr Laeger, 1995 Mar 20, 157(12), 1674 - 8
{Risk factors of listeriosis in Denmark 1989-1990}; Jensen A et al.; Risk factors for listeriosis are foods and underlying diseases . A case-control-study of listeriosis patients' dietary habits showed that unpasteurized milk was a risk food for sporadic listeriosis . A blue-mould cheese was significantly more often eaten by patients ill with an epidemic phage type . Leukaemia, AIDS and renal transplantation were found to be associated with a more than 1000 times higher risk of acquiring listeriosis, compared with healthy persons . As a part of prophylaxis, persons with a high risk of listeriosis should be informed individually about food hygiene and risk foods.

Vet Rec, 1995 Mar 4, 136(9), 211 - 6
Clinical and epidemiological correlates of the neurohistology of cases of histologically unconfirmed, clinically suspect bovine spongiform encephalopathy; Wells GA et al.; The associations between three major categories of the neurohistological diagnoses and the epidemiological data were examined in unconfirmed cases of clinically suspect bovine spongiform encephalopathy (BSE) . The diagnostic categories were focal spongiosis of white matter (37 cases), encephalic listeriosis (13 cases) and no significant lesions (78 cases) . An additional control category of 200 confirmed cases of BSE were included for comparison . Epidemiological variables were the frequencies of specific clinical signs, the season of clinical onset, the age, the duration of the clinical signs and the geographical origin of the cases . Discriminant analysis was used to assess the contribution of these variables to the distinction between the diagnostic categories . The analyses characterised the cases of listeriosis by their shortest clinical duration, the greater prevalences of certain clinical signs and their occurrence mainly in winter and spring, consistent with current understanding of the disease . Cases of focal spongiosis, a lesion of unknown significance, but potentially with a metabolic causation, were tentatively separable from cases with no significant lesions by their winter onset . The results also confirmed that among the categories, the cases of BSE had the longest clinical duration . Despite their statistical significance, the findings do not have sufficient predictive power to be of value in making clinical decisions.

Infect Immun, 1995 Mar, 63(3), 926 - 33
Hepatocytes can serve as accessory cells in the response of immune T lymphocytes to heat-killed Listeria monocytogenes; Jiang X et al.; Previous findings in our laboratory indicated that the bulk of Listeria monocytogenes injected intravenously into mice and recovered in the liver is taken up and replicates within hepatocytes . Other investigators have shown that hepatocytes can display costimulatory adhesion molecules, express major histocompatibility complex class I and II molecules, and secrete a number of cytokines, including interleukin-1 (IL-1), IL-6, and IL-8 . These data suggest that hepatocytes may serve as accessory cells in the immune response to L . monocytogenes . The accessory function and capacity of hepatocytes to present listerial antigens, however, have never been explored . We undertook a series of experiments to examine the response of Listeria-immune T lymphocytes to murine hepatocytes preincubated with heat-killed listeriae (HKL) . Electron micrographs showing the organism within membrane-limiting vacuoles demonstrated the capacity of hepatocytes to internalize HKL . T cells cocultured with hepatocytes pulsed with HKL exhibited a 5- to 10-fold increase in {methyl-3H}thymidine incorporation relative to T cells cultured with either hepatocytes or HKL alone . Similarly, gamma interferon production by immune T cells was elevated significantly in cultures that contained both hepatocytes and HKL . The optimal response of T cells required lysosomal processing of HKL by hepatocytes and contact between the two cell populations . Furthermore, maximum T-cell proliferation and gamma interferon production were dependent upon the presence of CD4+ T lymphocytes and the expression of Ia antigens . Taken together, these findings demonstrate that hepatocytes pulsed with HKL can stimulate the antigen-specific response of immune T lymphocytes . These results suggest that hepatocytes can serve as accessory cells in host defenses to listerial infections of the liver.

Cell Immunol, 1995 Mar, 161(1), 112 - 24
Unique order of the lymphocyte subset induction in the liver and intestine of mice during Listeria monocytogenes infection; Ohtsuka K et al.; We investigated how NK cells, extrathymic T cells, and thymus-derived T cells are activated in mice during infection with an intracellular pathogen, Listeria (L.) monocytogenes . Although macrophages and granulocytes are known to be involved in the elimination of this pathogen in an early phase of infection, it was still controversial what type of lymphocytes are induced as effectors in subsequent phases . When mice were ip injected with 1 x 10(3) L . monocytogenes (a sublethal dose), a prominent increase in the number of mononuclear cells in the liver and spleen was induced . Phenotypic analysis revealed that serial induction of lymphocyte subsets, NK cells-->extrathymic T cells-->thymus-derived T cells, occurred in these organs . Extrathymic T cells were estimated to have intermediate CD3 and a high level of IL-2 receptor beta-chain on the surface (i.e., intermediate CD3 cells) . These mice became free from infection after 2 weeks . In the case of oral administration, 1 x 10(3) L . monocytogenes increased the number of cells in the liver and the number of intraepithelial and lamina propria lymphocytes in the intestine . Phenotypic analysis also showed a sequential induction of lymphocyte subsets in the liver and the induction of extrathymic T cells in the intestine . Preelimination of intermediate TCR cells and NK cells by in vivo treatment with anti-LFA-1 mAb made mice susceptible to an ip injected sublethal dose of L . monocytogenes . These results reveal a unique order of lymphocyte induction during listerial infection and indicate that extrathymic T cells might be one of the important cells in achieving resistance against L . monocytogenes.

Appl Environ Microbiol, 1995 Mar, 61(3), 992 - 7
Suppression of Listeria monocytogenes colonization following adsorption of nisin onto silica surfaces; Bower CK et al.; Nisin is an antimicrobial peptide proven to be an effective inhibitor of gram-positive bacteria . It is known that nisin can adsorb to various surfaces and still retain much of its original activity (M . A . Daeschel, J . McGuire, and H . Al-Makhlafi, J . Food Prot . 55:731-735, 1992) . In this study, nisin films were allowed to form on silanized silica surfaces and then exposed to medium containing Listeria monocytogenes . Representative areas were selected from each surface, and images of resident listeriae were obtained at 4-h intervals for 12 h . During this time, cells on surfaces that had been in contact with a high concentration of nisin (1.0 mg/ml) exhibited no signs of growth and many displayed evidence of cellular deterioration . Surfaces treated with a lower concentration of nisin (0.1 mg/ml) had a smaller degree of inhibition . In contrast, both protein-free surfaces and those with films of heat-inactivated nisin allowed attached L . monocytogenes cells to grow and reproduce . These studies, when repeated with a nisin-resistant strain of L . monocytogenes, resulted in no inhibition of growth on surfaces with adsorbed nisin . The bactericidal effect of adsorbed nisin was also studied with iodonitrotetrazolium violet, a tetrazolium salt, which is reduced to a red formazan crystal by viable bacteria . Crystals were visible in 95% of the cells adhered to control surfaces but were present in less than 20% of the cells on surfaces with adsorbed nisin . These data indicate that adsorbed nisin may have potential for use as a food grade antimicrobial agent on food contact surfaces.

Lett Appl Microbiol, 1995 Mar, 20(3), 188 - 90
Rapid RAPD analysis for distinguishing Listeria species and Listeria monocytogenes serotypes using a capillary air thermal cycler; Black SF et al.; Using a novel capillary thermal cycler, randomly amplified polymorphic DNA (RAPD) generated DNA fingerprints were obtained in 3 h . The RAPD profiles were produced using a random 10-mer primer (5'-ACCGCCTGCT-3') which discriminated between different Listeria spp . Unique fragment profiles of Listeria monocytogenes serotypes were produced from serotypes 1a, 2, 3a, 4ab, 4a and 4c but serotypes, 1/2a, 4b, 4d and 7 had similar profiles.

Rev Clin Esp, 1995 Mar, 195(3), 154 - 9
{Listeriosis in the nonpregnant adult during an epidemic outbreak on Grand Canary}; Aladro Benito Y et al.; Twenty-two cases of community-acquired epidemic listeriosis were recorded from December 31st, 1991, to May 15th, 1993, at the Nuestra Senora del Pino Hospital, Las Palmas . The incidence during this outbreak was 31 times higher than the corresponding incidence in the last few years . Twelve cases occurred in pregnant women and/or neonates and ten in non-pregnant adult individuals . Our aim was to study the clinical, biological, radiological, and evolutive issues in non-pregnant adult patients . Six patients had some immunosuppressive condition: cancer, chemotherapy, AIDS, diabetes, and alcoholism . Eight patients had documented involvement of central nervous system: 6 cases of meningitis and 3 of cerebritis (one case had both meningitis and cerebritis); in the remaining two patients associated with seizures and acute confusional states, respectively . A neurological involvement was not documented because of the fulminant clinical course . CSF examination revealed mononuclear predominance in half of meningitis cases and was normal in two of the three cerebritis cases . The mean time from admission to diagnosis was 3.5 days . All patients but the two who died in the first hours of the disease received ampicillin and an aminoglycoside . The response to therapy was excellent with exception of one patient with meningitis who died in the fourth day of therapy . The clustering of listeriosis cases should alert physicians about the possibility of an epidemic outbreak . Listeria infection in non-pregnant adult individuals in this outbreak showed a high rate of neurological involvement, with focal cerebritis and pleocytosis with a mononuclear predominance in meningitis.

J Appl Bacteriol, 1995 Mar, 78(3), 297 - 303
Antibacterial effect of protamine assayed by impedimetry; Johansen C et al.; Impedimetric measurements were used to assay the antibacterial effect of protamine . A good linear correlation between the impedance detection time and the initial cell counts was obtained (r = 0.99, n = 2) . As basic peptides may cause clumping of cells, this correlation curve was used when estimating the cell number after protamine treatment, rather than colony counts . Protamine from salmon killed growing Gram-positive bacteria and significantly inhibited growth of Gram-negative bacteria in Tryptone Soy Broth (TSB) at 25 degrees C . In general Gram-positive bacteria were more sensitive to protamine than Gram-negative bacteria; the minimum inhibitory concentrations (MIC) determined for Gram-positive strains varied from 20 to 1000 micrograms ml-1 and for Gram-negative strains from 500 micrograms ml-1 to more than 4000 micrograms ml-1 . The effect of protamine on non-growing Listeria monocytogenes Scott A suspended in buffer was not lethal as was the effect on growing cells; however, protamine (50-500 micrograms ml-1) killed the Gram-negative fish spoilage bacteria Shewanella putrefaciens when the live cells were suspended in buffer.

Zh Mikrobiol Epidemiol Immunobiol, 1995 Mar-Apr, (2), 15 - 8
{The isolation and partial characterization of the cell wall proteins of Listeria monocytogenes}; Belyi IuF et al.; The preparative scheme for the purification of proteins with molecular weights of 39 and 79 kD, obtained from L . monocytogenes membrane fractions, has been developed . This technology included the cultivation of bacteria in heart-brain broth, isolation of bacterial membranes, the extraction of their components with Triton-X-100 and chromatography on Superose columns . The purified proteins have been shown to form structures with a molecular weight of 500-100 kD and pl 4.7 in water solutions.

Eur J Clin Microbiol Infect Dis, 1995 Mar, 14(3), 165 - 75
Antimicrobial chemotherapy of human infection due to Listeria monocytogenes; Jones EM et al.; Listeriosis is an uncommon infection, but when it occurs it carries a high mortality rate . Early diagnosis is essential and thereafter appropriate antimicrobial chemotherapy . Ampicillin or penicillin plus gentamicin remains the treatment of choice for most manifestations of listeriosis, and adequate doses must be given, i.e . greater than 6g/day of ampicillin or penicillin . Co-trimoxazole appears to be an excellent alternative agent with good penetration into the cerebrospinal fluid . Vancomycin is an appropriate agent for the treatment of primary bacteraemia but does not cross the blood-brain barrier sufficiently well to be useful in meningitis, while erythromycin may be used to treat listeriosis in cases of pregnancy . Treatment of bacteraemia requires one to two weeks' therapy, while meningitis cases may need to be treated for longer; for example, it has been found that most patients with acute meningitis in the UK were treated for 20 days . Infective endocarditis needs treatment for six to eight weeks . Doses should be varied with patients' altered organ function and antimicrobial serum monitoring performed when appropriate.

Electrophoresis, 1995 Mar, 16(3), 444 - 50
Analysis of heat and cold shock proteins in Listeria by two-dimensional electrophoresis; Phan-Thanh L et al.; The proteins induced by heat and cold shock in Listeria monocytogenes (pathogenic for humans) and L . innocua (nonpathogenic) strains were analyzed by two-dimensional (2-D) electrophoresis with the help of a computerized 2-D gel analysis system . Heat (49 degrees C) and cold (4 degrees C) shock repressed roughly half the number of proteins synthesized at normal temperature (25 degrees C) and decreased the level of numerous other proteins . Conversely, the synthesis of a great number of proteins was enhanced and novel proteins appeared upon temperature stress . There were more proteins induced in the L . monocytogenes strain than in the L . innocua strain . Each stress induced a set of specific proteins . There was overlap between these sets of proteins induced by heat and cold shock . Furthermore, a number of heat or cold shock proteins were found to be induced in both Listeria species and by both heat and cold shock in both species . The induction by heat shock was more intense than that by cold shock . The most strongly induced common stress protein of Listeria had a molecular mass of 17.6 kDa and an isoelectric point of 5.1.

Int J Food Microbiol, 1995 Mar, 25(1), 75 - 81
Control of Listeria monocytogenes in the delicatessen industries: the lessons of a listeriosis outbreak in France; Salvat G et al.; During a recent outbreak of foodborne listeriosis which occurred in France in 1992, investigations were carried out in order to identify the plants associated with the production of contaminated products . These investigations were made in six 'suspect' delicatessen plants following the first epidemiological investigations, and in one 'control plant' . The first visits were made during working operations . Two hundred and seventy samples were taken ('environmental' swabs, air samples, products), with 68% of the swabs being positive for Listeria monocytogenes in raw product areas, and 33% positives in the finished product area . The epidemic phagovar was identified in a single plant preparing pork tongues in aspic . The major causes of contamination identified were contact of cooked products with soiled surfaces, cross-contamination between 'raw' and 'cooked' channels and the inadequacy of cleaning and disinfection procedures . A second visit was also made to five plants to provide validation of their cleaning and disinfection procedures . Among 112 swabs collected, 17% of the samples from raw product surfaces and 7% from finished product surfaces were found to be positive . These results suggest that cleaning and disinfection procedures were unable to eliminate sources of L . monocytogenes when not correctly applied.

Int J Food Microbiol, 1995 Mar, 25(1), 19 - 27
Contamination pattern of Listeria monocytogenes and other Listeria spp . in a salmon slaughterhouse and smoked salmon processing plant; Rorvik LM et al.; A smoked salmon processing plant including a smokehouse and a slaughterhouse was examined for the occurrence of Listeria monocytogenes and other Listeria spp . From a total of 475 samples the overall frequency of L . monocytogenes was 16%, while other Listeria spp . were found in 22% of the samples . L . monocytogenes was most often detected in samples from the smokehouse, where 29% of the environmental and 26% of the fish samples during processing contained the bacteria . 17% of the fish raw material to the smokehouse were contaminated, while 11% of the samples from vacuum-packed smoked salmon were positive for L . monocytogenes . The slaughterhouse was sporadically contaminated, but L . monocytogenes was not found in 50 samples of slaughtered fish . L . monocytogenes was found in the seawater outside the slaughterhouse . Multilocus enzyme electrophoresis divided the isolated L . monocytogenes strains into 11 electrophoretic types (ETs) . One ET, ET-6, which is the most common ET in Norway, seemed to have colonized the smokehouse . Isolates from the seawater, from the slaughterhouse and from fish coming into the smokehouse, before filleting, were other ETs.

Appl Environ Microbiol, 1995 Mar, 61(3), 1150 - 2
A new procedure for efficient recovery of DNA, RNA, and proteins from Listeria cells by rapid lysis with a recombinant bacteriophage endolysin; Loessner MJ et al.; A method for the rapid lysis of Listeria cells, employing a recombinant Listeria bacteriophage A118 lytic enzyme (PLY118), is described . The procedure can be used with all listerial species . It enables fast, efficient, and gentle recovery of DNA, RNA, or native cellular proteins from small-scale (2- to 5-ml) cultures . Moreover, this approach should be very useful in analytical detection and differentiation of Listeria strains when the release of native nucleic acids or proteins is required.

FEMS Microbiol Lett, 1995 Feb 15, 126(2), 113 - 21
The actin-polymerization protein from Listeria ivanovii is a large repeat protein which shows only limited amino acid sequence homology to ActA from Listeria monocytogenes; Kreft J et al.; Within infected eukaryotic cells the two pathogenic Listeria species, L . monocytogenes and L . ivanovii, induce polymerization of cellular actin and the formation of a propulsive actin tail at one bacterial pole . For L . monocytogenes it has been shown that the product of the listerial actA gene is required for this process which is regarded as a model for actin-based motility . We have now cloned and sequenced a functionally analogous gene from L . ivanovii; its product, as deduced from the DNA sequence, is considerably larger (108 kDa) than L . monocytogenes ActA (67 kDa) and shares only a limited amino acid sequence homology (46% similarity on average) with the latter protein . This is the first example of a virulence gene product from L . ivanovii which is significantly different from its L . monocytogenes counterpart . Comparison of the two ActA proteins gives new insight into the structure of this class of actin-polymerization proteins, in particular with respect to their proline-rich repeat region.

J Interferon Cytokine Res, 1995 Feb, 15(2), 105 - 14
Modulation of the antilisterial activity of human blood-derived macrophages by activating and deactivating cytokines; Blauer F et al.; A concept of macrophage deactivation by hormones and cytokines that opposes activation was recently proposed . Deactivation of the antilisterial activity of macrophages by IL-4, IL-10, and TGF-beta, as well as by dexamethasone, was studied here . IL-4, IL-10, and dexamethasone, but not TGF-beta, caused a complete loss of the competence of human blood-derived macrophages infected with Listeria monocytogenes to control or eliminate ingested bacteria . IL-10 and, to a lesser degree, dexamethasone lessened in parallel the capacity of macrophages to secrete H2O2 . The antilisterial activity of cells simultaneously exposed to deactivating agents could be significantly augmented by IFN-gamma . Likewise, TNF-alpha and to a limited degree GM-CSF increased the antilisterial activity of cells treated with IL-10 and dexamethasone but not that of cells treated with IL-4 . Suppression of TNF-alpha secretion in response to Listeria by TGF-beta, IL-10, dexamethasone, or pentoxifylline did not closely parallel antilisterial activity . Studies by transmission electron microscopy and actin staining suggested that deactivation by IL-10, IL-4, and dexamethasone of human blood-derived macrophages resulted in intraphagosomal multiplication of Listeria followed only then by an escape of bacteria into the cytoplasm . The antibacterial competence of human macrophages is lessened by IL-4 and IL-10 and augmented by IFN-gamma, TNF-alpha, and GM-CSF . The success of human macrophages in controlling intracellular pathogens appears to depend on the balance of activating and deactivating mediators modulating their activity.

J Clin Invest, 1995 Feb, 95(2), 603 - 10
Bactericidal properties of murine intestinal phospholipase A2; Harwig SS et al.; We purified a molecule from the murine small intestine that killed both Escherichia coli and Listeria monocytogenes, and identified it as intestinal phospholipase A2 (iPLA2) by NH2-terminal sequencing and enzymatic measurements . The ability of iPLA2 to kill . L . monocytogenes was greatly enhanced by 5 mM calcium, inhibited by EGTA and abolished after reduction and alkylation, suggesting that enzymatic activity was required for iPLA2-mediated bactericidal activity . A mouse-avirulent phoP mutant, S . typhimurium 7953S, was 3.5-fold more susceptible to iPLA2 than its isogenic virulent parent, S . typhimurium 14028S (estimated minimal bactericidal concentrations 12.7 +/- 0.5 micrograms/ml vs . 43.9 +/- 4.5 micrograms/ml P < 0.001) . Overall, these findings identify iPLA2 as part of the antimicrobial arsenal that equips Paneth cells to protect the small intestinal crypts from microbial invasion . Because iPLA2 is identical to Type 2 phospholipase A2 molecules found in other sites, including spleen, platelets and inflammatory exudate cells, this enzyme may also contribute to antibacterial defenses elsewhere in the body.

J Exp Med, 1995 Feb 1, 181(2), 607 - 17
Monoclonal antibodies specific for murine p55 and p75 tumor necrosis factor receptors: identification of a novel in vivo role for p75; Sheehan KC et al.; Monoclonal antibodies (mAbs) specific for the murine p55 and p75 tumor necrosis factor (TNF) receptors were produced after immunization of Armenian hamsters with the purified soluble extracellular domains of each receptor protein . Four p55- (55R) and five p75 (TR75)-reactive mAbs immunoprecipitated the appropriate receptor from the surface of L929 cells . None of the mAbs cross-reacted with the other TNF receptor form . The mAbs were functionally characterized by their ability to inhibit ligand binding and influence TNF-dependent L cell cytolytic activity or proliferation of the murine cytolytic T cell clone CT6 . One p55-specific mAb, 55R-593, displayed agonist activity, while two other p55-specific mAbs (55R-170 and -176) were found to be TNF antagonists . The fourth mAb (55R-286) had no functional effects on cells . Several antibodies specific for the p75 TNF receptor partially inhibited recombinant murine TNF-alpha-dependent cytolytic activity (60%) . Blocking mAbs specific for p75 but not anti-p55 inhibited TNF-mediated proliferation of CT6 T cells . When used in vivo, p55- but not p75-specific mAbs protected mice from lethal endotoxin shock and blocked development of a protective response against Listeria monocytogenes infection . In contrast, both p55 and p75 mAbs individually blocked development of skin necrosis in mice treated with murine TNF-alpha . These data thus demonstrate the utility of the two families of murine TNF receptor-specific mAbs and identify a novel function of the p75 TNF receptor in vivo.

Infect Immun, 1995 Feb, 63(2), 720 - 3
Differential induction of macrophage-derived cytokines by live and dead intracellular bacteria in vitro; Zhan Y et al.; Marked differences in the abilities of living and heat-killed Brucella abortus and Listeria monocytogenes organisms to induce production of tumor necrosis factor alpha by in vitro-cultured macrophages were observed . Interleukin-1 and interleukin-6 appeared to be under different control . The results are discussed in relation to the induction of gamma interferon-producing Th1 cells and acquired cellular resistance to infection by living vaccines but not killed vaccines.

Immunopharmacol Immunotoxicol, 1995 Feb, 17(1), 59 - 68
Polysaccharide (ANK-102) from Polianthes tuberosa cells deteriorates the resistance of mice to Listeria monocytogenes infection; Majima T et al.; Modulatory effect on the murine self defense system by a newly discovered acidic polysaccharide (ANK-102) produced by P . tuberosa cells in liquid culture was examined . Pretreatment with ANK-102 deteriorated the murine survival against lethal infection of Listeria monocytogenes, an intracellular gram-positive bacterium eliminated mainly by macrophages through T-cell mediated immune response . Pretreatment with ANK-102 resulted in the accumulation of Mac 1 and Mac 2 positive cells in the peritoneal cavity of the infected animals and the reduction of Thy1.2 expression on the surface of the thymocytes . A new type of immunosuppressive polysaccharide ANK-102 was introduced.

J Neurol, 1995 Feb, 242(3), 153 - 6
Spinal manifestation of neurolisteriosis; Pfadenhauer K et al.; Spinal symptoms in acute bacterial meningitis are rare . In a series of 10 cases of neurolisteriosis, we observed 2 spinal complications, one due to an acute intramedullary abscess, the other caused by chronic spinal arachnoiditis . Therefore, if spinal symptoms develop in acute bacterial meningitis, Listeria monocytogenes infection should be considered and early adequate antibiotic treatment be implemented.

APMIS, 1995 Feb, 103(2), 107 - 12
Live tularemia vaccine but not proteins purified from Francisella tularensis can confer protection against lethal Listeria infection in mice; Belyi YF et al.; Immunization of Balb/c mice with Francisella tularensis vaccine strain 15/10 conferred significant protection against subsequent listerial infection . Since immunostimulatory activities could apparently be relevant to surface components of the bacterium, a technique for purification of cell wall proteins was developed . The scheme designed consisted of Triton X-100 extraction with subsequent FPLC chromatography steps, and resulted in the isolation of homogeneous proteins with molecular masses of 54 kDa (pI = 6.7) and 82 kDa (pI = 5.3), and partially purified 50, 85 and 100 kDa components . It was shown that immunization with isolated proteins failed to protect mice against lethal Listeria monocytogenes infection . Possible reasons for failure are discussed.

Cell Immunol, 1995 Feb, 160(2), 211 - 6
In vitro generation of IFN-gamma-producing Listeria-specific T cells is dependent on IFN-gamma production by non-NK cells; Song F et al.; In vitro 5-day cultures of naive spleen cells with viable Listeria monocytogenes (VLM), but not heat-killed L . monocytogenes, induced CD4+ T cells that produced IFN-gamma upon secondary antigen stimulation . The VLM-induced Listeria-specific T cells produced IFN-gamma but lacked expression of IL-2 and IL-4 . To study the role of IFN-gamma in the induction of the IFN-gamma-producing T cells, we added anti-IFN-gamma mAb to the primary culture and analyzed IFN-gamma production upon secondary antigen stimulation . Addition of anti-IFN-gamma mAb to the culture suppressed generation of IFN-gamma-producing CD4+ T cells, suggesting that IFN-gamma is important in the induction of IFN-gamma-producing CD4+ T cells . Furthermore, our results showed that depletion of NK cells from spleen cells by anti-asialo GM1 antibody plus complement before culture enhanced induction of IFN-gamma-producing CD4+ T cells . Although NK cells are known to produce IFN-gamma, the results indicate that NK cell-derived IFN-gamma may not be important in induction of the Listeria-specific IFN-gamma-producing CD4+ T cells in the culture system . In addition, we demonstrated that IFN-gamma expression was high in CD4+ T cells from cultures of spleen cells with VLM at the primary culture level . These results suggest that IFN-gamma derived from T cells may enhance production of IFN-gamma by CD4+ T cells, while NK cells rather suppress the induction of IFN-gamma producing CD4+ T cells.

Can J Microbiol, 1995 Feb, 41(2), 152 - 6
The identification of bacterial gene expression differences using mRNA-based isothermal subtractive hybridization; Utt EA et al.; We describe a method for isolating and determining differences in gene expression between related bacterial strains . The method is based upon differences in mRNA expression . To demonstrate this procedure, cDNA generated from total RNA of Listeria monocytogenes serotype 1/2a was hybridized to total RNA from a Tn916 mutant of serogroup 1/2a (M3) that was deficient in the production of listeriolysin O, the product of the hly gene . The single-stranded cDNA fragments remaining after hybridization represent the difference in expressed genes between the two strains . These subtraction products were used as hybridization probes to identify the corresponding hly gene in a Southern hybridization.

Res Microbiol, 1995 Feb, 146(2), 143 - 54
Identification and classification of Listeria by two-dimensional protein mapping; Gormon T et al.; The cellular proteins of 29 Listeria strains belonging to different species and serotypes were analysed by two-dimensional (2-D) electrophoresis with the help of a computerized 2-D gel analysis system . The protein patterns were similar among strains within a Listeria species, but were different from one species to another . The comparative analysis of these protein maps enabled us to find specific proteins and to determine the genetic relatedness among Listeria spp . strains . The cluster analysis based on protein mapping showed a division between species and a clear separation of L . monocytogenes from other Listeria species . Inside L . monocytogenes the strains were divided into two main clusters in correlation with flagellar antigenic structures; this is in concordance with the results that have been found on the basis of multilocus enzyme electromorphs or DNA-restriction patterns . This technique enabled us to subtype the strains sharing the same serovar or the same lysovar . Because of its independence and high discriminating capacity, the 2-D protein mapping technique might provide a powerful tool for the identification and classification of Listeria strains.

Appl Environ Microbiol, 1995 Feb, 61(2), 817 - 9
Direct detection of Listeria monocytogenes in 25 milliliters of raw milk by a two-step PCR with nested primers; Herman LM et al.; A method for direct detection of Listeria monocytogenes in 25 ml of raw milk is presented . The detection limit can be situated between 10 and 5 CFU . The detection method is based on chemical extraction of the milk components and PCR amplification with two nested pairs of primers specific for Listeria monocytogenes.

Infect Immun, 1995 Feb, 63(2), 486 - 97
CD14 is not involved in Rhodobacter sphaeroides diphosphoryl lipid A inhibition of tumor necrosis factor alpha and nitric oxide induction by taxol in murine macrophages; Kirikae F et al.; Taxol, a microtubule stabilizer with anticancer activity, mimics the actions of lipopolysaccharide (LPS) on murine macrophages in vitro . Recently, it was shown that taxol-induced macrophage activation was inhibited by the LPS antagonist Rhodobacter sphaeroides diphosphoryl lipid A (RsDPLA) . To investigate the mechanisms of taxol-induced macrophage activation, the present study focused on the interaction of LPS, RsDPLA, and taxol in the activation of and binding to macrophages . Taxol alone induced murine C3H/He macrophages to secrete tumor necrosis factor alpha (TNF) and to produce nitric oxide (NO) with kinetics similar to that of LPS . Macrophages from LPS-hyporesponsive C3H/HeJ mice, in contrast, did not yield any detectable TNF and NO production in response to LPS or taxol . RsDPLA inhibited taxol-induced TNF and NO production from C3H/He macrophages in a dose-dependent manner . The inhibition by RsDPLA was specific for LPS and taxol in that RsDPLA did not inhibit heat-killed Listeria monocytogenes- or zymosan-induced TNF production . Polymyxin B blocked the inhibitory effect of RsDPLA on taxol-induced TNF production . The inhibitory activity of RsDPLA appeared to be reversible since macrophages still responded to taxol in inducing TNF production after the RsDPLA was washed out with phosphate-buffered saline prior to the addition of taxol . Taxol-induced TNF production was not inhibited by colchicine, vinblastine, or 10-deacetylbaccatine III . A mutant cell line, J7.DEF3, defective in expression of a CD14 antigen, responded equally well to taxol by producing TNF as did the parent J774.1 cells . This suggested that the activation of macrophages by taxol does not require CD14 . Taxol-induced TNF production by the mutant cells was also inhibited by RsDPLA . 125I-labeled LPS and 3H-labeled taxol was reported to bind to J774.1 cells predominantly via CD14 and microtubules, respectively . The binding of 125I-labeled LPS to J7.DEF3 cells was about 30 to 40% of that to J774.1 cells . The binding of 125I-LPS to J774.1 cells was inhibited by unlabeled LPS and RsDPLA but not by taxol . On the other hand, 3H-labeled taxol bound to both J774.1 cells and J7.DEF3 cells in similar time- and dose-dependent manners . The binding of {3H}taxol to these cells was inhibited by taxol but not by LPS or RsDPLA . Although the binding studies failed to examine cross competition for binding to macrophages, a possible explanation of these results is that LPS, RsDPLA, and taxol share the same molecule(s) on murine macrophages for their functional receptor(s), which is neither CD14 nor tubulin.

Cell, 1995 Jan 27, 80(2), 353 - 61
Targeted disruption of the NF-IL6 gene discloses its essential role in bacteria killing and tumor cytotoxicity by macrophages; Tanaka T et al.; To investigate the role of NF-IL6 in vivo, we have generated NF-IL6 (-/-) mice by gene targeting . NF-IL6 (-/-) mice were highly susceptible to infection by Listeria monocytogenes . Electron microscopic observation revealed the escape of a larger number of pathogens from the phagosome to the cytoplasm in activated macrophages from NF-IL6 (-/-) mice . Furthermore, the tumor cytotoxicity of macrophages from NF-IL6 (-/-) mice was severely impaired . However, cytokines involved in macrophage activation, such as TNF and IFN gamma, were induced normally in NF-IL6 (-/-) mice . Nitric oxide (NO) formation was induced to a similar extent in macrophages from both wild-type and NF-IL6 (-/-) mice . These results demonstrate the crucial role of NF-IL6 in macrophage bactericidal and tumoricidal activities as well as the existence of a NO-independent mechanism of these activities . We also demonstrate that NF-IL6 is essential for the induction of G-CSF in macrophages and fibroblasts.

Nature, 1995 Jan 19, 373(6511), 255 - 7
Differential production of interferon-gamma and interleukin-4 in response to Th1- and Th2-stimulating pathogens by gamma delta T cells in vivo; Ferrick DA et al.; Exposure to various pathogens can stimulate at least two patterns of cytokine production by CD4-positive T cells . Responses that result in secretion of interferon-gamma (IFN-gamma), lymphotoxin and interleukin-2 (IL-2) are classified as T-helper-1 (Th1); CD4+ T-cell production of IL-4, IL-5, IL-9, IL-10 and IL-13 is called a T-helper-2 response (Th2) . Differentiation of CD4+ T cells into either Th1 or Th2 cells is influenced by the cytokine milieu in which the initial antigen priming occurs . Here we use flow cytometry to identify the presence of intracellular cytokines (cytoflow) and analyse T-cell production of IFN-gamma and IL-4 from mice infected with Listeria monocytogenes or Nippostrongylus brasiliensis . We show that T cells bearing gamma delta receptors discriminate early in infection between these two pathogens by producing cytokines associated with the appropriate T-helper response . Our results demonstrate that gamma delta T cells are involved in establishing primary immune responses.

Presse Med, 1995 Jan 7, 24(1), 23 - 5
{Listeriosis in patients with malignant hemopathy . 3 cases in the same hospital ward}; Fontan J et al.; During the listeriosis epidemic which occurred in France in the summer 1992, three patients with malignant haematopathies hospitalized in our service contracted the disease . Although the retrospective investigations were hindered by the variable incubation period and the impossibility of examining the foods eaten at the time of infection, there was a high probability that two of the patients had been infected by cooked ham and dairy products at home . The third patient was apparently infected in hospital with well-cooked food found to be contaminated . The hypothesis of coinfection has been raised.

Biol Cell, 1995, 85(1), 55 - 66
The loss of contact inhibition and anchorage-dependent growth are key steps in the acquisition of Listeria monocytogenes susceptibility phenotype by non-phagocytic cells; Velge P et al.; We have previously demonstrated that intestinal and kidney finite cell lines were resistant to L monocytogenes invasion (ie allowed low bacterial entry and no intracellular multiplication) in contrast to the continuous cell lines which were susceptible to Listeria invasion (ie allowed high bacterial entry and intracellular multiplication) (Velge et al (1994a) Med Microbial Immunol 183, 145) . The aim of this study was to discover whether epigenetic or genetic cellular modifications could convert L monocytogenes resistant cells into a susceptible phenotype and to determine the cellular steps involved in Listeria susceptibility . Among the 5-azacytidine treated finite cell lines, the untransformed immortal cell lines established remained resistant to L monocytogenes invasion whereas the weakly transformed continuous cell lines established were converted into a susceptible phenotype . Transfection of resistant cells by SV40 large T antigen induced only highly transformed continuous cell lines displaying a susceptible phenotype . Taken together these data show that cell transformation enhanced Listeria invasion . This conclusion was supported by the observation that L monocytogenes was able to induce cell foci within murine finite cell monolayers . This morphological cell transformation was completely reversible and required live bacteria inside cells . In conclusion, we may speculate that the L monocytogenes intracellular multiplication observed within cell foci could be explained by the loss of contact inhibition of the finite cell monolayer . Indeed, the loss of both contact inhibition and anchorage-dependent growth are the key steps involved in the L monocytogenes susceptibility phenotype.

J Inflamm, 1995, 45(4), 239 - 47
Expression of an adenovirally encoded lymphotoxin-beta inhibitor prevents clearance of Listeria monocytogenes in mice; Trueb R et al.; The lymphotoxin (LT)-beta heterotrimer was recently identified as a molecule containing LT-alpha subunits, tethered to the cell through non-covalent association with an integral plasma membrane protein, derived from the LT-beta gene . Since knockout mutations of the LT-alpha gene yield animals that lack lymph nodes, whereas animals lacking either or both of the receptors for tumor necrosis factor (TNF) and LT-alpha homotrimers have normal lymph nodes, it has been inferred that the association between the LT-beta heterotrimer and its cognate receptor is required for lymph node ontogeny . Similarly, LT-beta and its receptor are thought to be important for development of the spleen . Since LT-alpha deficient mice lack lymph nodes, it is difficult to assess the extradevelopmental contribution of LT-beta to immune competence . To this end, we employed a strategy for the conditional blockade of LT-beta heteromer activity in normal mice . The interaction between LT-beta and its receptor is essential for the destruction of intracellular Listeria monocytogenes.

Folia Microbiol (Praha), 1995, 40(6), 652 - 4
Accumulation of activated lymphocytes in liver of Listeria factor Ei treated rabbits; Janoutova J et al.; Factor Ei isolated from Listeria monocytogenes caused in rabbits 1 d after intravenous administration activation of peripheral blood lymphocytes in terms of rRNA biosynthesis . Increase of the number of these active lymphocytes was observed not only in peripheral blood, but also in spleen, lung, kidney, and liver . when the number of lymphocytes was related to the amount of the organ tissue cells, the increment appeared significant only in the liver where the number of active lymphocytes exceeded the control value by one order of magnitude . Based upon this observation we concluded that the accumulation of Listeria factor Ei activated lymphocytes occurred in liver . This characteristic is considered an additional immunomodulative property of Listeria factor Ei similar to the Escherichia coli lipopolysaccharide.

Arch Immunol Ther Exp (Warsz), 1995, 43(5-6), 239 - 45
Participation of IL-2 and direct cell cytotoxicity in the modulation of GvH reaction in mice infected with Listeria innocua; Goscicka T et al.; Listeria innocua can intensify the development of local GvH reaction in a semiallogeneic system in mice . This phenomenon is observed in (BALB/c x AKR)F1 mice intraperitoneally injected with the live bacteria on the 7th day before the transfer of BALB/c spleen cells . The local GvH reaction develops as strongly as in normal hybrid recipients given the parental cell graft in the presence of exogenous interleukin 2 (IL-2) . The development of the reaction is associated with an intensive increase of the direct cytotoxicity of lymph node cells which is also markedly enhanced after the bacteria injection itself . IL-2 seems to play a relevant role in the development of local GvH reaction in L . innocua-injected mice.

Scand J Rheumatol, 1995, 24(6), 392 - 4
Listeria monocytogenes arthritis of several joints; Ukkonen HJ et al.; A patient with rheumatoid arthritis (RA) developed an infection caused by Listeria monocytogenes in her left knee and both shoulder joints . The clinical presentation of the disease was rather indolent with relatively moderate joint symptoms . Moreover, the synovial fluid sample was only slightly turbid with a white blood cell count of 23.5 x 10(9)/1 . As compared to the earlier reported cases of L . monocytogenes septic arthritis, our patient is unique because she had infection in several joints . The polyarticular joint involvement combined with the clinical symptoms resembling the activation of RA posed us diagnostic difficulties.

J Mal Vasc, 1995, 20(4), 326 - 7
{A new case of arterial Listeria infection}; Poli P et al.; Listeria monocytogenes arterial infections are uncommon and usually occur on aneurysms of previously operated arteries, classically in very young and very old or immunodepressed subjects . Peroperative samples for bacteriology should always be taken as antibiotics generally used in vascular surgery are ineffective against listeria . We report a case of a patient who underwent emergency surgery . Bacteriologic analysis of the thrombus revealed listeria inoculation which was eradicated by adapted antibiotic treatment.

Artif Cells Blood Substit Immobil Biotechnol, 1995, 23(6), 665 - 79
Influence of steric stabilization of liposome-encapsulated hemoglobin on Listeria monocytogenes host defense; Sherwood RL et al.; Liposome-encapsulated hemoglobin (LEH) products are being investigated as potential blood substitutes . To determine if changes in LEH composition can modify the immune response, red blood cell substitutes based on conventional lipids containing phosphatidylinositol (LEH1) and sterically stabilized lipid vesicles containing polyethylene glycol phosphatidylethanolamine (LEH2) were tested for effects on host resistance . On Day 0, groups of 18 to 20 female CD-1 mice were given an intravenous (i.v.) infectious challenge with a 20% lethal dose of Listeria monocytogenes . Mice received a single i.v . dose of LEH1, LEH2, or albumin vehicle on Day +1 or Day -3 relative to infectious challenge . Mice dosed with LEH1 and LEH2 on Day +1 died rapidly from Listeria infection; but mice dosed with LEH2 lived significantly longer than did mice receiving LEH1 . By contrast, when administered on Day -3, LEH1 had no significant effect on host immunity, while LEH2 increased susceptibility to Listeria infection . In addition, LEH1 and LEH2 both caused significant reduction of phagocytic activity as measured by rat alveolar macrophage (AM) ingestion of latex microspheres . AM incubated 4 hr with either LEH1 or LEH2 prior to addition of microspheres ingested fewer beads in a dose-dependent manner . No difference in in vitro phagocytic activity was observed between LEH1 or LEH2 . The inability to differentiate LEH formulations based on in vitro phagocytic activity suggests that the in vivo Listeria infection model may be more relevant in discerning the immunotoxicity of the LEH formulations tested.

Med Dosw Mikrobiol, 1995, 47(1-2), 17 - 23
{Phosphatidylinositol specific phospholipase C (PI-PLC) in differentiation of Listeria monocytogenes and Listeria innocua}; Pachelska M et al.; Listeria strains were investigated as regard: the production of PI-PLC and lecytinase, haemolytic and CAMP activity survival in mouse tissues . Although all the examined L . monocytogenes strains produced PI-PLC, they showed a very different activity of this enzyme . It was surprising that one of the four L . innocua strains demonstrated a weak activity of PI-PLC . L . monocytogenes bacteria with strong PI-PLC activity survived in mouse spleens for a much longer period of time than L . monocytogenes microbes with only a weak production of this enzyme . However, L . innocua bacteria producing some PI-PLC were quickly eliminated by infected mice.

Microbiology, 1995 Jan, 141 ( Pt 1), 41 - 9
The role of peptide metabolism in the growth of Listeria monocytogenes ATCC 23074 at high osmolarity; Amezaga MR et al.; The growth of Listeria monocytogenes ATCC 23074 in defined medium is sensitive to high osmolarity when compared with its growth in complex media, such as brain heart infusion (BHI) . The two major contributors to this difference in growth rate are the availability in BHI of the osmoprotectant glycine betaine and peptides . Peptone plays two major roles: firstly as a nutritional supplement for protein synthesis, and secondly as a source of amino acids and peptides that serve as a mechanism of maintaining turgor . In the presence of peptone the total amino acid pool at high osmolarity is substantial and even in the presence of glycine betaine the amino acid pool makes a major contribution to turgor maintenance . At high osmolarity there is a general increase in amino acid pools, with particularly substantial pools of glutamate, aspartate, proline, hydroxyproline and glycine . Peptides are also accumulated by cells from the peptone supplied in the medium . Glycine-containing peptides are accumulated in the cytoplasm under all conditions . Specific glycine- and proline-containing peptides stimulate growth at high osmolarity . The peptide prolyl-hydroxyproline accumulates in cells to high levels in response to growth at high osmolarity, and the pools of the derived amino acids also show a dependence on the external osmotic pressure . However, proline only confers significant osmoprotection when supplied as peptides . The significance of these data in the context of the occurrence of L . monocytogenes in foods with high peptide content is discussed.

Arzneimittelforschung, 1995 Jan, 45(1), 104 - 7
Influence of ibuprofen on the infection with Listeria monocytogenes; Hockertz S et al.; Listeria monocytogenes is a bacterial infection, which is facultatively localized in monocytes and macrophages . The influence of ibuprofen (CAS 15687-27-1), a nonsteroidal anti-inflammatory drug (NSAID), on this bacterial infection in balb/c mice was investigated . One day prior to sublethal infection, balb/c mice were treated intravenously with various therapeutic concentrations of ibuprofen alone or ibuprofen in combination with a suboptimal dosage of murine recombinant interferon gamma, a lymphokine produced by T-helper cells . Three days post-infection, parasite burdens of the mainly infected organs, spleen and liver, were determined by the colony-forming unit assay . It was shown that the prophylactic treatment with ibuprofen in a concentration of 4 mg/kg body weight resulted in a more than 10-fold reduction of viable Listeria monocytogenes in the spleen, whereas in liver 12 mg/kg Ibuprofen was necessary for a comparable kill of viable bacteria . A higher concentration of ibuprofen did not resulted in a higher antibacterial efficacy . In order to clarify the mechanism of ibuprofen action, molecular-biological experiments were performed to measure the messenger RNA (mRNA) induced by ibuprofen . It is presented here that therapeutic concentrations of ibuprofen induced significant higher amounts of mRNA for interleukin-1 in human monocytes compared to untreated cells . These findings support the hypothesis that ibuprofen influences the complex immune system to overcome a bacterial infection.

Appl Environ Microbiol, 1995 Jan, 61(1), 377 - 8
Discrimination of species in the genus Listeria by Fourier transform infrared spectroscopy and canonical variate analysis; Holt C et al.; Infrared spectra of type cultures of the six recognized species of the genus Listeria and of Listeria grayi subsp . murrayi were recorded . By use of a library of 59 spectra, comprising at least six replicates of each type, discrimination by canonical variate analysis of the spectral amplitudes allowed all of the spectra to be correctly classified.

Appl Environ Microbiol, 1995 Jan, 61(1), 303 - 9
Proposals for optimization of the international phage typing system for Listeria monocytogenes: combined analysis of phage lytic spectrum and variability of typing results; Marquet-Van der Mee N et al.; Combined analysis of 5,179 serial phage reactions of 20 Listeria monocytogenes propagating strains over 14 years and phage typing results from 2,659 further L . monocytogenes strains allowed us to estimate lytic spectrum specificity and the variability of the lytic reactions of 35 phages . These included the 26 phages recommended for the international method for phage typing defined in 1985 by Rocourt et al . (J . Rocourt, A . Audurier, A . L . Courtieu, J . Durst, S . Ortel, A . Schrettenbrunner, and A . G . Taylor, Zentralbl . Bakteriol . Abt . 1 Orig . A 259:489-497, 1985) . The results are discussed individually for each phage . Proposals for modifying the present system are made with the aim of producing an optimal bacteriophage set for routine use.

J Appl Bacteriol, 1995 Jan, 78(1), 66 - 70
Comparative study of the growth of Listeria monocytogenes in defined media and demonstration of growth in continuous culture; Jones CE et al.; A basic requirement for physiological studies with Listeria monocytogenes is a chemically defined medium that supports growth of the bacterium in batch and continuous culture . A number of such media have been devised but comparative studies of their efficiency are few and none has been used in continuous culture . Six of the media were compared for their ability to sustain sequential growth of L . monocytogenes in static and aerated batch culture with glucose as sole carbon source . The most suitable, judged on the basis of ease of preparation, growth rate and yield, was that of Trivett and Meyer (1971) . This medium was shown to support growth of L . monocytogenes NCTC 7973 in continuous culture in a chemostat . A lytic phenomenon, noted with the same strain under anaerobic conditions and in batch culture in the chemostat, is discussed.

Infect Immun, 1995 Jan, 63(1), 182 - 90
Inhibition of Listeria locomotion by mosquito oostatic factor, a natural oligoproline peptide uncoupler of profilin action; Southwick FS et al.; Mosquito oostatic factor, a naturally occurring decapeptide (YDPAPPPPPP), strikingly resembles the primary structure of oligoproline-rich regions within the protein ActA, a bacterial surface protein required for Listeria motility in host cells . When microinjected into Listeria-infected PtK2 cells, the insect oostatic factor rapidly blocks Listeria-induced actin rocket tail assembly as well as intracellular locomotion of this pathogen . At intracellular concentrations of about 90 nM, transient inhibition of rocket tail formation and bacterial locomotion occurs, followed by full recovery of tail length and motility . However, at 0.9 microM oostatic factor, both processes are permanently arrested . Introduction of oostatic factor by microinjection also causes PtK2 peripheral membrane retraction in both Listeria-infected and uninfected cells . Epifluorescence microscopy with bodipy-phallacidin reveals that cells microinjected with the insect factor lose all actin stress fibers and accumulate F-actin in regions of membrane retraction . When the insect peptide is combined with profilin as an equimolar binary solution (1 microM {final concentration} each), intracellular addition fails to inhibit Listeria rocket-tail formation, fails to block intracellular bacterial movement, and no longer causes marked membrane retraction . The ability of profilin to neutralize the inhibitory action of oostatic factor is consistent with complex formation, and this finding suggests that profilin may interact directly with ActA peptide as well as a host cell peripheral membrane component to promote actin filament assembly by locally generating ATP-actin . Dispersal of profilin from such sites by oligoproline-rich peptide inhibitors suggests that profilin is directly involved in intracellular pathogen locomotion and reorganization of actin cytoskeleton of the host cell peripheral membrane.

Respiration, 1995, 62(2), 107 - 9
Pneumonia caused by Listeria monocytogenes; Garcia-Montero M et al.; Pneumonia due to Listeria monocytogenes is extremely uncommon . We report the case of an 87-year-old woman with no underlying immunosuppressive disease who presented with listerial pneumonia . Cutaneous anergy and a decrease in total lymphocyte count in this elderly woman could predispose her to listerial infection.

Lett Appl Microbiol, 1995 Jan, 20(1), 57 - 60
The incidence, numbers and types of Listeria monocytogenes isolated from farm bulk tank milks; Fenlon DR et al.; Bulk tank milk from 160 producers was tested for Listeria monocytogenes at three monthly intervals over 1 year . Twenty-five producers were positive, most on a single occasion, only seven were positive on three or more of the four samplings . Listeria monocytogenes numbers were low, usually < 1 ml-1, the highest was 35 ml-1 . All isolates were serotype 1, the use of multilocus enzyme electrophoresis on representative isolates gave nine different electrophoretic types, two have been associated with listeriosis in humans or animals, a further two had only been isolated from one other source (silage or faeces), while the majority (5) were unique to milk.

Cell Motil Cytoskeleton, 1995, 30(3), 229 - 46
Organization and structure of actin filament bundles in Listeria-infected cells; Zhukarev V et al.; During its motion inside host cells, Listeria monocytogenes promotes the formation of a column of actin filaments that extends outward from the distal end of the moving bacterium . The column is constructed of short actin filaments that polymerize at the bacteria-column interface . To get a measure of filament organization in the column, Listeria grown in cultured PtK2 cells were studied with steady state fluorescence polarization, confocal microscopy, and whole cell intermediate voltage electron microscopy . Although actin filament ordering was higher in nearby stress fibers than in the Listeria-associated actin, four distinct areas of ordering could be observed in fluorescence polarization ratio images of bacteria: 1) the surface of the bacteria, 2) the cytoplasm next to the bacteria, 3) the outer shell of the actin column, and 4) the core of the column . Filaments were preferentially oriented parallel to the long axis of the column with highest ordering along the long axis of the bacterial surface and in the shell of the tail . The lowest ordering was in the core (where filaments are possibly also shorter with respect to the cup and the shell), whereas in the adjacent cytoplasm, filaments were oriented perpendicular to the column . A mutant of Listeria that can polymerize actin around itself but cannot move intracellularly does not have its actin organized along the bacterial surface . Thus the alignment of the actin filaments along the bacterial surfaces may be important for the intracellular movement . These conclusions are also supported by confocal microscopy and whole mount electron microscopic data that also reveal that actin filaments can be deposited asymmetrically around the long axis of the bacteria, a distribution that may affect the direction of motility of Listeria monocytogenes inside infected cells.

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