Listeriosis

Mol Microbiol, 1999 Mar, 31(6), 1709 - 22
Listeriolysin O-dependent activation of endothelial cells during infection with Listeria monocytogenes: activation of NF-kappa B and upregulation of adhesion molecules and chemokines; Kayal S et al.; The facultative intracellular bacterium Listeria monocytogenes is an invasive pathogen that crosses the vascular endothelium and disseminates to the placenta and the central nervous system . Its interaction with endothelial cells is crucial for the pathogenesis of listeriosis . By infecting in vitro human umbilical vein endothelial cells (HUVEC) with L . monocytogenes, we found that wild-type bacteria induced the expression of the adhesion molecules (ICAM-1 and E-selectin), chemokine secretion (IL-8 and monocyte chemotactic protein-1) and NF-kappa B nuclear translocation . The activation of HUVEC required viable bacteria and was abolished in prfA-deficient mutants of L . monocytogenes, suggesting that virulence genes are associated with endothelial cell activation . Using a genetic approach with mutants of virulence genes, we found that listeriolysin O (LLO)-deficient mutants inactivated in the hly gene did not induce HUVEC activation, as opposed to mutants inactivated in the other virulence genes . Adhesion molecule expression, chemokine secretion and NF-kappa B activation were fully restored by a strain of Listeria innocua transformed with the hly gene encoding LLO . The relevance in vivo of endothelial cell activation for listerial pathogenesis was investigated in transgenic mice carrying an NF-kappa B-responsive lacZ reporter gene . NF-kappa B activation was visualized by a strong lacZ expression in endothelial cells of capillaries of mice infected with a virulent haemolytic strain, but was not seen in those infected with a non-haemolytic isogenic mutant . Direct evidence that LLO is involved in NF-kappa B activation in transgenic mice was provided by injecting intravenously purified LLO, thus inducing stimulation of NF-kappa B in endothelial cells of blood capillaries . Our results demonstrate that functional listeriolysin O secreted by bacteria contributes as a potent inflammatory stimulus to inducing endothelial cell activation during the infectious process.

Mol Microbiol, 1999 Mar, 31(6), 1631 - 41
Delivery of protein to the cytosol of macrophages using Escherichia coli K-12; Higgins DE et al.; Listeriolysin O (LLO) is an essential determinant of pathogenicity whose natural biological role is to mediate lysis of Listeria monocytogenes containing phagosomes . In this study, we report that Escherichia coli expressing cytoplasmic recombinant LLO can efficiently deliver co-expressed proteins to the cytosol of macrophages . We propose a model in which subsequent or concomitant to phagocytosis the E . coli are killed and degraded within phagosomes causing the release of LLO and target proteins from the bacteria . LLO acts by forming large pores in the phagosomal membrane, thus releasing the target protein into the cytosol . Delivery was shown to be rapid, within minutes after phagocytosis . Using this method, a large enzymatically active protein was delivered to the cytosol . Furthermore, we demonstrated that the E . coli/LLO system is very efficient for delivery of ovalbumin (OVA) to the major histocompatibility (MHC) class I pathway for antigen processing and presentation, greater than 4 logs compared with E . coli expressing OVA alone . Moreover, the time required for processing and presentation of an OVA-derived peptide was similar to that previously reported when purified OVA was introduced directly into the cytosol by other methods . Using this system, potentially large amounts of any protein that can be expressed in E . coli can be delivered to the cytosol without protein purification . The potential use of this system for the delivery of antigenic protein in vivo and the delivery of DNA are discussed.

J Biol Chem, 1999 Apr 23, 274(17), 11459 - 62
Increased expression of Rab5a correlates directly with accelerated maturation of Listeria monocytogenes phagosomes; Alvarez-Dominguez C et al.; Previous studies have shown that Listeria monocytogenes (LM) modulates phagocytic membrane traffic . Here we explore whether Rab5a, a GTPase associated with phagosome-endosome fusion, is related to phagosome maturation and to the intracellular survival of LM . Stable transfection of Rab5a cDNA into macrophages accelerates intracellular degradation of LM . Morphological studies confirmed that phagosome maturation and phagosome-lysosome fusion is enhanced by overexpression of Rab5a . Down-regulation experiments using antisense oligonucleotides targeted to the Rab5a mRNA efficiently reduced Rab5a synthesis, reduced phagosome-endosome traffic, blocked phagosome-lysosome fusion, and extended intraphagosomal survival of LM . Down-regulation of Rab5a had no effect on LM internalization . Down-regulation of Rab5c had no effect on phagosome maturation and phagosome-lysosome fusion . The results indicate that Rab5a controls early phagosome-endosome interactions and governs the maturation of the early phagosome leading to phagosome-lysosome fusion.

J Immunol, 1999 Apr 15, 162(8), 4781 - 9
Enhancement of the Listeria monocytogenes p60-specific CD4 and CD8 T cell memory by nonpathogenic Listeria innocua; Geginat G et al.; The contact of T cells to cross-reactive antigenic determinants expressed by nonpathogenic environmental micro-organisms may contribute to the induction or maintenance of T cell memory . This hypothesis was evaluated in the model of murine Listeria monocytogenes infection . The influence of nonpathogenic L . innocua on the L . monocytogenes p60-specific T cell response was analyzed . We show that some CD4 T cell clones raised against purified p60 from L . monocytogenes cross-react with p60 purified from L . innocua . The L . monocytogenes p60-specific CD4 T cell clone 1A recognized the corresponding L . innocua p60 peptide QAAKPAPAPSTN, which differs only in the first amino acid residue . In vitro experiments revealed that after L . monocytogenes infection of APCs, MHC class I-restricted presentation of p60 occurs, while MHC class II-restricted p60 presentation is inhibited . L . innocua-infected cells presented p60 more weakly but equally well in the context of both MHC class I and MHC class II . In contrast to these in vitro experiments the infection of mice with L . monocytogenes induced a strong p60-specific CD4 and CD8 T cell response, while L . innocua infection failed to induce p60-specific T cells . L . innocua booster infection, however, expanded p60-specific memory T cells induced by previous L . monocytogenes infection . In conclusion, these findings suggest that infection with a frequently occurring environmental bacterium such as L . innocua, which is nonpathogenic and not adapted to intracellular replication, can contribute to the maintenance of memory T cells specific for a related intracellular pathogen.

Lett Appl Microbiol, 1999 Mar, 28(3), 216 - 20
Behaviour of Listeria monocytogenes under combined chilling processes; Membre JM et al.; The behaviour of Listeria monocytogenes under chilling processes was investigated . Growth kinetics were measured at 7 degrees C in TSBYE culture medium as a function of pH (7.2 and 6.2), pre-incubation temperatures (4 or 7 degrees C), cooling (0.05 or 0.1 degree C min-1) and freezing (0 and -5 degrees C) treatments . Growth curves generated were fitted by Gompertz and Baranyi functions . The Baranyi function gave better parameter estimation values than the Gompertz equation which over-estimated the specific growth rate values . Listeria monocytogenes grew at 7 degrees C without a lag phase, except when the sub-culture was performed at 37 degrees C, whereas the specific growth rate was affected by the chilling processes . In fact, L . monocytogenes grew slightly faster at 7 degrees C when a 4 degrees C pre-incubation treatment was applied than with a 7 degrees C pre-incubation treatment . These results suggest that to mimic the processes of contamination in industry, predictive microbiology studies with L . monocytogenes should be performed with organisms cultured at low temperatures.

J Appl Microbiol, 1999 Mar, 86(3), 544 - 56
Response to high-pressure, low-temperature treatment in vegetables: determination of survival rates of microbial populations using flow cytometry and detection of peroxidase activity using confocal microscopy; Arroyo G et al.; Application of high hydrostatic pressure (200, 300, 350 and 400 MPa) at 5 degrees C for 30 min to different micro-organisms, including Gram-positive and Gram-negative bacteria, moulds and yeasts, proved to be more effective in inactivating these organisms than treatments at 20 degrees C for 10 min and at 10 degrees C for 20 min . Moulds, yeasts, Gram-negative bacteria and Listeria monocytogenes were most sensitive, and their populations were completely inactivated at pressures between 300 and 350 MPa . The same conditions of pressure, temperature, and time were applied to different vegetables (lettuce, tomato, asparagus, spinach, cauliflower and onion), achieving reductions of from 2-4 log units in both viable mesophiles and moulds and yeasts at pressures of between 300 and 400 MPa . Sensory characteristics were unaltered, especially in asparagus, onion, tomato and cauliflower, though slight browning was observed in cauliflower at 350 MPa . Flow cytometry was applied to certain of the microbial populations used in the above experiment before and after the pressurization treatment . The results were indicative of differing percentage survival rates depending on micro-organism type, with higher survival rates for Gram-positive bacteria, except L . monocytogenes, than in the other test micro-organisms . Growth of survivors was undetectable using the plate count method, suggesting that micro-organisms suffering from pressure stress were metabolically inactive though alive . The pressurization treatments did not inactivate the peroxidase responsible for browning in vegetables . Confocal microscopic examination of epidermal tissue from onion showed that the enzyme had been displaced to the cell interior . Use of low temperatures and moderately long pressurization times yielded improved inactivation of micro-organisms and better sensorial characteristics of the vegetables, and should lower industrial costs.

J Appl Microbiol, 1999 Mar, 86(3), 469 - 76
Effect of osmotic, alkaline, acid or thermal stresses on the growth and inhibition of Listeria monocytogenes; Vasseur C et al.; Five strains of Listeria monocytogenes (a, b, c, d and e) isolated from industrial plants have been subjected to different osmotic, alkaline, acid or thermal stresses . The effects of these treatments on lag-phase (L) and growth rate (mu) of cells in mid-log phase have been followed using an automated optical density monitoring system . Increasing the osmotic pressure by the addition of different amounts of NaCl increased the lag phase and decreased the growth rate . The same phenomena were observed after decreasing the pH of the medium to 5.8, 5.6 or 5.4 by addition of acetic, lactic or hydrochloric acids . The inhibitory effect was: acetic acid > lactic acid > hydrochloric acid . The addition of NaOH to attain pH values of 9.5, 10.0, 10.5 or 11.0 in the medium produced a dramatic increase of the lag phase at pH 10.5 and 11 . Growth rates were also decreased while the maximal population increased with high pH values . These effects varied according to strains . Strains d and e were the most resistant to acidic and alkaline stresses, and e was the most affected by the addition of NaCl . A cold shock of 30 min at 0 degree C had limited effects on growth parameters . On the other hand, hyperthermal shocks (55 or 63 degrees C, 30 min) led to similar increased lag phases and to significant increases of the maximal population in all five strains.

J Virol, 1999 May, 73(5), 4120 - 6
CpG-containing oligonucleotides are efficient adjuvants for induction of protective antiviral immune responses with T-cell peptide vaccines; Oxenius A et al.; Synthetic nonmethylated oligonucleotides containing CpG dinucleotides (CpG-ODNs) have been shown to exhibit immunostimulatory activity . CpG-ODNs have the capacity to directly activate B cells, macrophages, and dendritic cells, and we show here that this is reflected by cell surface binding of oligonucleotides to these cell subsets . However, T cells are not directly activated by CpG-ODNs, which correlates with the failure to bind to the T-cell surface . Efficient competition for CpG-induced B-cell activation by non-CpG-containing oligonucleotides suggests that oligonucleotides might bind to an as yet undefined sequence-nonspecific receptor prior to cellular activation . Induction of protective T-cell responses against challenge infection with lymphocytic choriomeningitis virus (LCMV) or with recombinant vaccinia virus expressing the LCMV glycoprotein was achieved by immunizing mice with the immunodominant major histocompatibility complex class I-binding LCMV glycoprotein-derived peptide gp33 together with CpG-ODNs . In these experiments, B cells, potentially serving as CpG-ODN-activated antigen-presenting cells (APCs), were not required for induction of protective immunity since CpG-ODN-gp33-immunized B-cell-deficient mice were equally protected against challenge infection with both viruses . This finding suggested that macrophages and/or dendritic cells were sufficiently activated in vivo by CpG-ODNs to serve as potent APCs for the induction of naive T cells . Furthermore, treatment with CpG-ODN alone induced protection against infection with Listeria monocytogenes via antigen-independent activation of macrophages . These data suggest that CpG activation of macrophages and dendritic cells may provide a critical step in CpG-ODN adjuvant activity.

Mol Gen Genet, 1999 Mar, 261(2), 323 - 36
Differential expression of Listeria monocytogenes virulence genes in mammalian host cells; Bubert A et al.; We have used RT-PCR and GFP-mediated fluorescence to analyse the regulation of PrfA-dependent virulence genes of Listeria monocytogenes during proliferation in mammalian host cells . Our data show that most of the PrfA-regulated virulence genes are more efficiently expressed, as measured by transcript levels, when L . monocytogenes is grown in macrophages and macrophage-like cells rather than in epithelial cells, hepatocytes or endothelial cells . The promoters for hly and plcA are predominantly activated within the phagosomal compartment, while those for actA and inlC are predominantly activated in the host cell cytosol . Expression of actA and plcB precedes that of inlC after infection of epithelial cells and macrophages . Little transcription of inlA or inlB is observed in epithelial cells and there is only slightly more in macrophages . In both cell types the level of transcription of the inlAB operon is lower than is seen under extracellular growth conditions in rich media, which is compatible with the assumption that InlA and InlB are not required during intracellular growth of the bacteria . Activation of the PrfA-independent iap promoter is also low during intracellular growth, although the gene product (p60) is required for cell viability . The levels of the PrfA-dependent virulence gene transcripts do not correlate with the amount of prfA transcript present, which is low under all intracellular conditions analysed, suggesting that the prfA transcript is either highly unstable in bacteria that are growing intracellularly, or that the small amount of PrfA produced is highly activated by additional component(s).

Int J Food Microbiol, 1999 Feb 18, 46(3), 251 - 61
A model describing the relationship between regrowth lag time and mild temperature increase for Listeria monocytogenes; Breand S et al.; In order to comply with the consumer demand for ready-to-eat and look 'fresh' products, mild heat treatment will be used more and more in the agrofood industry . Nonetheless there is no tool to define the most appropriate mild heat treatment . In order to build this tool, it is necessary to study and describe the response of a bacterial population to a mild increase in temperature, from the dynamic point of view . The response to a mild increase in temperature, defined by stress duration and temperature, consisted in a mortality phase followed by the lag time of the survivors and their exponential growth . The effect of the mild increase in temperature on the mortality phase was described in a previous paper (Breand et al., Int . J . Food Microbiol., in press) . The effect of the stress duration on the lag was presented in a previous paper (Breand et al., Int . J . Food Microbiol . 38 (1997) 157-167) . In particular, the biphasic relationship between the lag and the stress duration was observed and modelled with a four parameter nonlinear model: the primary model (Breand et al., Int . J . Food Microbiol . 38 (1997) 157-167) . The study presented in this paper deals with the effect of the stress temperature on the biphasic relationship between the lag time and the stress duration . The secondary models describing the effect of the stress temperature on this biphasic relationship, were empirically built from our experimental data concerning Listeria monocytogenes . This work pointed out that the higher the stress temperature, the narrower the range of stress duration for which the lag time increased . Since the primary and the secondary models of the lag time were available, the global model describing the effect of the mild increase duration and temperature directly on the lag was fitted . This model allowed an improvement of the parameter estimator precision . The potential contribution in mild heat treatment optimization of this global model and the one built for the mortality phase (Breand et al., Int . J . Food Microbiol., in press) is discussed.

Int J Food Microbiol, 1999 Feb 18, 46(3), 187 - 92
Characterization of Listeria monocytogenes from an ice cream plant by serotyping and pulsed-field gel electrophoresis; Miettinen MK et al.; One dominating strain of serotype 1/2b was found when serotyping and pulsed-field gel electrophoresis (PFGE) patterns were used for the characterization of 41 Listeria monocytogenes isolates originating from an ice cream plant . Samples were taken from the production environment, equipment and ice cream during the years 1990-1997 . Serotyping divided the isolates into two serovars, 1/2b and 4b . Three rare-cutting enzymes (ApaI, AscI and SmaI) were used in the creation of PFGE patterns . AscI resulted in the best restriction enzyme digestion patterns (REDPs) for visual comparison . Eight different AscI REDPs were obtained, whereas ApaI produced six and SmaI seven banding patterns . When one-band differences are taken into account, 12 different PFGE types were distinguished based on information obtained with all three enzymes . The dominant PFGE type was found to have persisted in the ice cream plant for seven years . Improved and precisely targeted cleaning and disinfection practices combined with structural changes making for easier cleaning of the packaging machine, resulted in eradication of L . monocytogenes from this plant.

J Clin Periodontol, 1999 Mar, 26(3), 164 - 8
Comparative antiplaque effectiveness of an essential oil and an amine fluoride/stannous fluoride mouthrinse; Riep BG et al.; The adjunctive use of antimicrobial mouthrinses to help control supragingival plaque and gingivitis has been shown to contribute significantly to patients' daily oral hygiene regimens . This controlled clinical study used an observer-blind, randomized, cross-over design in a 4-day plaque regrowth model to determine the relative efficacies of an essential oil-containing mouthrinse (Listerine Antiseptic) and an amine fluoride/stannous fluoride-containing mouthrinse (Meridol) in inhibiting the development of supragingival plaque . A 0.1% chlorhexidine mouthrinse (Chlorhexamed-Fluid) was used as a positive control, and a 5% hydroalcohol solution was used as a negative control . Dosing for each of the test mouthrinses was based on the manufacturers' label directions . Because the volume and rinse time for each of the test mouthrinses were different, each test mouthrinse had its own negative control group . On day 1 of each test period, subjects received an oral soft and hard tissue examination and a dental prophylaxis to remove all plaque, calculus, and extrinsic stain . Starting the same day, subjects refrained from all mechanical oral hygiene procedures for the next 4 days and rinsed 2x daily under supervision with their randomly-assigned mouthrinse . On day 5, each subject received a plaque assessment as well as an oral examination to assess side effects . Each test period was separated by a 2-week washout period . 23 volunteers with a median age of 26 years completed the study . Compared to the respective placebos, the median percent plaque reductions at 5 days were 23.0%, 12.2%, and 38.2% for the essential oil, amine/stannous fluoride, and chlorhexidine rinses, respectively . The plaque reductions seen in the essential oil and chlorhexidine rinse groups were statistically significant (p < 0.001), while the plaque reduction in the amine/stannous fluoride rinse group was not statistically significant (p > 0.05) . Additionally, the essential oil rinse was significantly more effective (p < 0.001) than the amine/stannous fluoride rinse in inhibiting plaque accumulation in this clinical model.

Zentralbl Bakteriol, 1999 Feb, 289(1), 31 - 6
Detection of Listeria monocytogenes in milk by the polymerase chain reaction; Salmanzadeh Ahrabi S et al.; A on the polymerase chain reaction (PCR) method based was developed for detection of Listeria monocytogenes in milk samples after enrichment culture . It consists of culturing samples in Listeria enrichment broth, followed by DNA extraction and detection of the organism using PCR . Dilutions of L . monocytogenes in milk were subjected to PCR amplification after enrichment culture . When determining the sensitivity of the method, it was found to be possible to detect 37 CFU (colony forming unit gl/ml) of the bacterium in milk . The method was assessed as a sensitive, specific, times-saving and practical way of detecting L . monocytogenes in milk samples.

Acta Crystallogr D Biol Crystallogr, 1999 Feb, 55 ( Pt 2), 552 - 3
Crystallization and preliminary X-ray crystallographic analysis of the unusual ferritin from Listeria innocua; Ilari A et al.; Single crystals of ferritin extracted from Listeria innocua have been obtained by the vapour-diffusion method using PEG 1000 as precipitant . The crystals are orthorhombic, space group P212121, with unit-cell dimensions a = 87.7, b = 137.5, c = 173.1 A . The crystals diffract to 2.9 A resolution on a rotating-anode X-ray source and to 2.35 A resolution on a synchrotron X-ray source . The asymmetric unit contains one molecule formed by 12 subunits, corresponding to a packing density of 2.41 A3 Da-1

J Leukoc Biol, 1999 Feb, 65(2), 265 - 9
LPS down-regulates the expression of chemokine receptor CCR2 in mice and abolishes macrophage infiltration in acute inflammation; Zhou Y et al.; Interactions between chemokines and their specific receptors are important for leukocyte trafficking . The CC-chemokine monocyte chemoattractant protein-1 (MCP-1) and its specific receptor CCR2 are essential in monocytic infiltration and have been associated with several inflammatory diseases . It has been reported that several endotoxin and proinflammatory cytokines inhibit CCR2 expression in vitro in human monocytes . We report here that lipopolysaccharides (LPS) down-regulated CCR2 expression both in vitro and in vivo . Injection of LPS into mice dramatically reduced the expression of CCR2 on the surface of peripheral blood cells and completely blocked macrophage infiltration into the peritoneal cavity in response to thioglycollate elicitation . In addition, treatment of mice with LPS reduced their efficiency to clear Listeria monocytogenes infection . These results suggest that down-regulation of CCR2 and blockage of monocyte infiltration may contribute to the inhibition of macrophage function in vivo by a low dose of LPS.

J Leukoc Biol, 1999 Feb, 65(2), 256 - 64
Functional deficiencies of peritoneal cells from gene-targeted mice lacking G-CSF or GM-CSF; Zhan Y et al.; Gene-targeted mice lacking the hemopoietic growth factors, granulocyte colony-stimulating factor (G-CSF) or granulocyte-macrophage (GM)-CSF, show increased susceptibility to infection with the facultative intracellular bacterium, Listeria monocytogenes . The resident peritoneal cell populations from G-CSF(-/-) and GM-CSF(-/-) mice showed reduced production of the bactericidal molecule nitric oxide . Macrophage-mediated tumoricidal activity and phagocytosis of Listeria were reduced in G-CSF(-/-), but not in GM-CSF(-/-), mice . In G-CSF(-/-) mice, there was an unexpected expansion (from 18% in WT to 38%) of a population of cells with morphology intermediate between typical macrophages and typical lymphocytes . These cells had some of the features of poorly differentiated macrophages, being adherent to plastic but poorly phagocytic, nonspecific esterase positive but myeloperoxidase negative . They were largely negative for the macrophage marker F4/80 and for Thy1, B220, and Gr1 . Their disproportionate presence, and the corresponding deficiency in typical macrophages, possibly accounts for some of the functional deficiencies observed in G-CSF(-/-) mice.

J Cell Biol, 1999 Mar 22, 144(6), 1245 - 58
Role of proteins of the Ena/VASP family in actin-based motility of Listeria monocytogenes; Laurent V et al.; Intracellular propulsion of Listeria monocytogenes is the best understood form of motility dependent on actin polymerization . We have used in vitro motility assays of Listeria in platelet and brain extracts to elucidate the function of the focal adhesion proteins of the Ena (Drosophila Enabled)/VASP (vasodilator-stimulated phosphoprotein) family in actin-based motility . Immunodepletion of VASP from platelet extracts and of Evl (Ena/VASP-like protein) from brain extracts of Mena knockout (-/-) mice combined with add-back of recombinant (bacterial or eukaryotic) VASP and Evl show that VASP, Mena, and Evl play interchangeable roles and are required to transform actin polymerization into active movement and propulsive force . The EVH1 (Ena/VASP homology 1) domain of VASP is in slow association-dissociation equilibrium high-affinity binding to the zyxin-homologous, proline-rich region of ActA . VASP also interacts with F-actin via its COOH-terminal EVH2 domain . Hence VASP/ Ena/Evl link the bacterium to the actin tail, which is required for movement . The affinity of VASP for F-actin is controlled by phosphorylation of serine 157 by cAMP-dependent protein kinase . Phospho-VASP binds with high affinity (0.5 x 10(8) M-1); dephospho-VASP binds 40-fold less tightly . We propose a molecular ratchet model for insertional polymerization of actin, within which frequent attachment-detachment of VASP to F-actin allows its sliding along the growing filament.

Infect Immun, 1999 Apr, 67(4), 1901 - 9
Effects of interleukin-1 receptor antagonist overexpression on infection by Listeria monocytogenes; Irikura VM et al.; Interleukin-1 receptor antagonist (IL-1ra) is a naturally occurring cytokine whose only known function is the inhibition of interleukin-1 (IL-1) . Using a reverse genetic approach in mice, we previously showed that increasing IL-1ra gene dosage leads to reduced survival of a primary listerial infection . In this study, we characterize further the role of endogenously produced IL-1ra and, by inference, IL-1 in murine listeriosis . IL-1ra overexpression inhibits, but does not eliminate, primary immune responses, reducing survival and increasing bacterial loads in the target organs . We demonstrate that IL-1ra functions in the innate immune response to regulate the peak leukocyte levels in the blood, the accumulation of leukocytes at sites of infection, and the activation of macrophages during a primary infection . Reduced macrophage class II major histocompatibility complex expression was observed despite increased gamma interferon (IFN-gamma) levels, suggesting that IL-1 activity is essential along with IFN-gamma for macrophage activation in vivo . We also show that IL-1ra plays a more limited role during secondary listeriosis, blunting the strength of the delayed-type hypersensitivity response to listerial antigen while not significantly altering cellular immunity to a second infectious challenge . When these results are compared to those for other mutant mice, IL-1ra appears to be unique among the cytokines studied to date in its regulation of leukocyte migration during primary listeriosis.

Infect Immun, 1999 Apr, 67(4), 1844 - 52
Examination of Listeria monocytogenes intracellular gene expression by using the green fluorescent protein of Aequorea victoria; Freitag NE et al.; The ActA protein of Listeria monocytogenes is an essential virulence factor and is required for intracellular bacterial motility and cell-to-cell spread . plcB, cotranscribed with actA, encodes a broad-specificity phospholipase C that contributes to lysis of host cell vacuoles and cell-to-cell spread . Construction of a transcriptional fusion between actA-plcB and the green fluorescent protein gene of Aequorea victoria has facilitated the detailed examination of patterns of actA/plcB expression within infected tissue culture cells . actA/plcB expression began approximately 30 min postinfection and was dependent upon entry of L . monocytogenes into the host cytosol . L . monocytogenes Deltahly mutants, which are unable to escape from host cell vacuoles, did not express actA/plcB at detectable levels within infected tissue culture cells; however, complementation of the hly defect allowed entry of the bacteria into the host cytoplasm and subsequent actA/plcB expression . These results emphasize the ability of L . monocytogenes to sense the different host cell compartment environments encountered during the course of infection and to regulate virulence gene expression in response.

Infect Immun, 1999 Apr, 67(4), 1770 - 8
Listeria monocytogenes phospholipase C-dependent calcium signaling modulates bacterial entry into J774 macrophage-like cells; Wadsworth SJ et al.; Listeria monocytogenes secretes several proteins that have been shown to contribute to virulence . Among these is listeriolysin O (LLO), a pore-forming hemolysin that is absolutely required for virulence . Two other virulence factors are phospholipases: a phosphatidylinositol-specific phospholipase C (PI-PLC {plcA}) and a broad-range PLC (plcB) . Although mutations in plcA or plcB resulted in small increases in mouse 50% lethal dose (LD50), deletions in both genes resulted in a 500-fold increase in LD50 . We have examined the role of these secreted proteins in host intracellular signaling in the J774 macrophage-like cell line . Measurements of cytosolic free calcium ({Ca2+}i) have revealed a rapid spike upon exposure of these cells to wild-type L . monocytogenes . This is followed by a second peak at 5 min and a third prolonged peak with a maximal {Ca2+}i of 800 to 1,000 nM . The pattern of calcium changes was greatly altered by deletion of any of the three virulence factors . An LLO mutant produced none of these elevations in {Ca2+}i; however, a transient elevation was observed whenever these bacteria entered the cell . A PI-PLC mutant produced a diminished single elevation in {Ca2+}i at 15 to 30 min . A broad-range PLC mutant produced only the first calcium spike . Studies with inhibitors suggested that the first elevation arises from influx of calcium from the extracellular medium through plasma membrane channels and that the second and third elevations come from release of Ca2+ from intracellular stores . We observed that internalization of wild-type bacteria and the broad-range PLC mutant was delayed for 5 to 10 min, but the LLO and PI-PLC mutants were internalized rapidly upon infection . Inhibitors that affected calcium signaling changed the kinetics of association of wild-type bacteria with J774 cells, the kinetics of entry, and the efficiency of escape from the primary phagosome.

Immunobiology, 1999 Feb, 200(1), 120 - 7
Effect of fetal calf serum on cytokine release by bone marrow-derived macrophages during infection with intracellular bacteria; Flesch IE et al.; Bone marrow-derived macrophages (BMM) comprise a population of quiescent cells which can be activated by defined signals . Here, we directly compare the release of chemokines and monokines by BMM raised either in serum-supplemented or in serum-free medium in response to Listeria monocytogenes EGD or Mycobacterium bovis BCG infection . We focused on this issue because there have been several controversial reports on the production of cytokines by BMM due to different in vitro culture conditions . Culture in serum-supplemented medium primed BMM for release of monocyte chemoattractant protein (MCP)-1, interleukin (IL)-6, and IL-12, but had no effect on macrophage inflammatory protein (MIP)-1alpha and tumor necrosis factor (TNF)-alpha production in response to L . monocytogenes infection . After challenge infection with M . bovis, BMM raised and stimulated in serum-supplemented medium secreted higher levels of MCP-1, MIP-1alpha, IL-6, and TNF-alpha but not of IL-12 as compared to BMM cultured and infected in a serum-free medium . The effects of serum could be partially mimicked by interferon-gamma . Because the serum components responsible for BMM priming are undefined, BMM cultured under serum-free conditions provide an appropriate cell population for defining macrophage activating signals.

Immunopharmacol Immunotoxicol, 1999 Feb, 21(1), 109 - 24
Petiveria alliacea L . extract protects mice against Listeria monocytogenes infection--effects on bone marrow progenitor cells; Quadros MR et al.; In this study we have investigated the effects of Petiveria alliacea on the hematopoietic response of mice infected with Listeria monocytogenes . Our results demonstrate a protective effect of the crude extract of P . alliacea since the survival of the treated/infected was higher than that in the infected group . Moreover, the number of granulocyte/macrophage colonies (CFU-GM) and the serum colony stimulating activity levels were increased in the treated/infected mice in relation to the infected group . These results suggest an immunomodulation of Petiveria alliacea extract on hematopoiesis, which may be responsible, at least in part, for the increased resistance of mice to Listeria monocytogenes infection.

FEMS Microbiol Lett, 1999 Feb 15, 171(2), 209 - 14
Evaluation of the API test, phosphatidylinositol-specific phospholipase C activity and PCR method in identification of Listeria monocytogenes in meat foods; Paziak-Domanska B et al.; The aim of this work was to compare the possibility of identifying Listeria monocytogenes strains isolated from meat and sausage on the basis of the API-Listeria test, production of phosphatidylinositol-specific phospholipase C (PI-PLC) and polymerase chain reaction (PCR) for a DNA fragment of the hlyA gene encoding listeriolysin O . Forty-six strains were isolated and examined . The lethality of some Listeria isolates for BALB/c mice was also determined . In this study, all isolates identified as L . monocytogenes in the API test gave a positive signal in the PCR . Listeriae identified as L . innocua or L . welshimeri in the API test were negative in the PCR conducted with the primers for listeriolysin O . All strains identified as L . monocytogenes on the basis of the API test and the PCR produced PI-PLC . However, this activity was not limited to the bacteria of this species . Four out of 17 L . innocua and three out of 10 L . welshimeri isolates were PI-PLC-positive . None of the L . innocua or L . welshimeri isolates (neither PI-PLC+ or PI-PLC-) showed lethality for BALB/c mice . In contrast, two L . monocytogenes isolates as well as a reference L . monocytogenes strain killed all mice used for the experiment.

Tidsskr Nor Laegeforen, 1999 Jan 30, 119(3), 375 - 9
{Listeria monocytogenes--the perfect parasite?}; Wyller VB et al.; The pathogen Listeria monocytogenes has an extraordinary intracellular life cycle, based on adaption to and exploitation of normal cellular mechanisms . Extracellular organisms induce their own phagocytosis, followed by destruction of the phagosomal membrane . Cytoplasmatic bacteria organize the intracellular protein actin into "comet tails", and thus gain motility . In contact with the plasma membrane, motile bacteria induce a pseudopodium, corresponding to an invagination of the plasma membrane of the neighbouring cell . Eventually, the pseudopodium is engulfed by the neighbouring cell, creating a double membrane vacuole . L . monocytogenes destroys the double membrane, and escapes into the cytoplasm . This article reviews the molecular biology of Listeria infection, and how research in this field has yielded increased insight into normal cellular processes . Finally, we propose that the neuroinvasive properties of L . monocytogenes is caused by actin-dependent transport within axons from the periphery to the central nervous system.

Tidsskr Nor Laegeforen, 1999 Jan 30, 119(3), 373 - 4
{Brain stem infection caused by Listeria monocytogenes}; Antal EA et al.; Rhombencephalitis due to Listeria monocytogenes is a serious form of brainstem inflammation . It is difficult to diagnose on the basis of clinical and laboratory findings alone . We describe a fatal case of Listeria monocytogenes rhombencephalitis in a previously healthy female teenager . Clinical and pathogenetic aspects of this condition are discussed.

J Vasc Surg, 1999 Mar, 29(3), 554 - 6
Infrarenal endoluminal bifurcated stent graft infected with Listeria monocytogenes; Heikkinen L et al.; Prosthetic graft infection as a result of Listeria monocytogenes is an extremely rare event that recently occurred in a 77-year-old man who underwent endoluminal stent grafting for infrarenal abdominal aortic aneurysm . The infected aortic endoluminal prosthesis was removed by means of en bloc resection of the aneurysm and contained endograft with in situ aortoiliac reconstruction . At the 10-month follow-up examination, the patient was well and had no signs of infection.

Vet Res Commun, 1998 Dec, 22(8), 505 - 16
Kinetics of interferon-gamma production and its comparison with anti-listeriolysin O detection in experimental bovine listeriosis; Barbuddhe SB et al.; The kinetics of the production of interferon gamma (IFN-gamma) in whole blood culture and its comparison with anti-listeriolysin O (ALLO) detection by ELISA were studied during oral infection of calves with Listeria monocytogenes . Culture filtrate antigen (CFA), listeriolysin O (LLO), and sonicated antigen (SA) were used to prime the peripheral blood mononuclear cells (PBMCs) and the plasma from orally infected calves . IFN-gamma and ALLO appeared as early as day 7 of an oral infection . IFN-gamma was detected earlier with LLO than with SA . The Max50 interleukin (IL-2) activity and IFN-gamma estimated in the culture supernatant from PBMCs primed in vitro with different antigens of L . monocytogenes revealed high induction of IL-2 and IFN-gamma by CFA, LLO and live antigen . IFN-gamma assay and ALLO detection were used for testing cases of repeat breeding in dairy cattle . It appeared that detection of IFN-gamma employing LLO can be used to diagnose listerial infections.

Immunol Lett, 1999 Feb, 65(3), 167 - 73
Dietary omega-3 polyunsaturated fatty acids from fish oil reduce interleukin-12 and interferon-gamma production in mice; Fritsche KL et al.; The objective of this study was to investigate the impact of feeding mice a diet rich in n-3 polyunsaturated fatty acids (PUFA) from fish oil on the interleukin-12 (IL-12) and interferon-gamma (IFNgamma) production during the early stage of an infectious challenge with Listeria monocytogenes . Weanling female C3H/HeN mice were fed AIN-93G experimental diets containing 20%, by weight one of three fat sources: lard (low PUFA), soybean oil (n-6 PUFA) or a mixture (9:1) of menhaden fish oil and corn oil (n-3 PUFA) . After 4 weeks, mice were injected intraperitoneally with 10(5) Listeria monocytogenes and the concentration of IL-12(p70) and IFNgamma in serum was determined 24 h post-infection by ELISA . IL-12p35, IL-12p40 mRNA, and IFNgamma mRNA in the spleen were quantified by RNase protection assay . The number of IFNgamma-producing cells in the spleen was determined by flow cytometry using an intracellular staining procedure . We found that n-3 PUFA-fed mice had lower levels of circulating IL-12 at 24 h post-infection than n-6 PUFA- or low PUFA-fed mice (9.7+/-3.4 pg/ml vs . 61.6+/-10.6, and 44.4+/-12.5 pg/ml, respectively; P=0.002, n = 10/trt) . The level of IL-12 p35 mRNA did not significantly differ among dietary treatment groups . However, IL-12p40 mRNA was significantly lower in n-3 PUFA- and n-6 PUFA-fed mice compared to low-PUFA-fed mice . Further, the n-3 PUFA group also had the lowest circulating IFNgamma (4.4+/-1.8 ng/ml vs . 9.1+/-1.0, and 9.7+/-2.1 ng/ml, respectively; P = 0.007 . n = 8-10/trt) . The n-3 PUFA-fed mice had significantly lower IFNgamma mRNA in their spleens compared to the mice fed the other fat sources . In agreement with having lower circulating IFNgamma and lower splenic IFNgamma mRNA, n-3 PUFA-fed mice had a significantly lower percentage of IFNgamma-producing cells in their spleens compared with the n-6 PUFA-fed group (2.1+/-0.6 vs . 4.2+/-0.7%; P = 0.037, n = 10/trt) . In summary, feeding mice a diet rich in n-3 PUFA from fish oil significantly lowered the production of both IL-12 and IFNgamma during the early phase of a Listeria infection.

Immunol Lett, 1999 Jan, 65(1-2), 93 - 8
T lymphocyte dynamics during Listeria monocytogenes infection; Busch DH et al.; Infection of mice with Listeria monocytogenes results in a robust T lymphocyte response that clears the pathogen and provides long term immunity from reinfection . The number of splenic CD4+ and CD8+ T cells and natural killer cells increases during primary and recall infection with L . monocytogenes, however the proportional increase is greatest for CD8+ T cells . The proportion of CD8 T cells expressing low levels of CD62L, a sign of activation, was increased among immune splenocytes, suggesting a substantial expansion of L . monocytogenes specific CTL . Analysis of CTL specific for the immunodominant LLO 91-99 epitope showed that essentially all were CD62Llo during the primary response, but that many upregulated expression of CD62L during the memory phase . Interestingly, the antigen specificity of nearly all additional CD62Llo CTL detected in spleens during recall L . monocytogenes infection can be accounted for with MHC class I tetramers complexed with four different epitopes . These studies demonstrate the complex T lymphocyte dynamics during infection with intracellular pathogens.

Eur J Immunol, 1999 Feb, 29(2), 581 - 91
The resistance against Listeria monocytogenes and the formation of germinal centers depend on a functional death domain of the 55 kDa tumor necrosis factor receptor; Plitz T et al.; The biological functions mediated by the death domain of the 55-kDa tumor necrosis factor receptor (TNFRp55) in vivo are still elusive . TNFRp55 mutants lacking a functional death domain were expressed in TNFRp55-/- and in TNFRp55+/- mice as transgenes under control of the H-2Kb promoter . Analysis of these animals revealed that signals originating from the TNFRp55 death domain are indispensable for the protection against Listeria monocytogenes, the expression of mucosal addressin cell adhesion molecule-1 (MAdCAM-1) in the spleen and the development of splenic germinal centers . Furthermore, the transgenic coexpression of the TNFRp55 mutants in TNFRp55+/- mice exerts a dominant negative effect on the signaling of the endogenous receptor chains in vivo.

Vet Microbiol, 1999 Feb 12, 64(4), 333 - 41
Cytotoxic T-cell, delayed type hypersensitive and listeriolysin O responses in experimental bovine listeriosis; Barbuddhe SB et al.; Five different antigens of Listeria monocytogenes NCTC 11994 were assayed for their lymphoproliferative, cytotoxic, humoral and delayed type hypersensitivity (DTH) responses in experimentally infected calves . The antigens used were culture filtrate antigen (CFA), listeriolysin O (LLO), sonicated antigen (SA), live antigen (LA) and heat killed antigen (HKA) . CFA was most lymphoproliferative when assayed in vitro . In an autologous monocyte cytotoxicity assay, soluble protein as well as bacterial cells elicited cytolytic responses, however, LA and LLO primed monocytes showed a higher cytotoxic effect . The expression of cell surface markers of lymphocyte subpopulations was almost identical in experimentally infected as well as non-infected calves . SA elicited the highest humoral and DTH responses . LLO being a major virulence factor with appreciable humoral as well as cellular responses may be used as a candidate antigen to develop a reliable diagnostic immunoassay.

Can Vet J, 1998 Feb, 39(2), 100 - 2
Listeria monocytogenes and Escherichia coli septicemia and meningoencephalitis in a 7-day-old llama; Frank N et al.; Listeria monocytogenes and Escherichia coli were isolated from blood collected on presentation and tissues samples taken postmortem . Listeria monocytogenes was isolated from cerebrospinal fluid collected antemortem . The importance of passive transfer of immunity, the subtlety of neurologic signs in early meningitis, and considering blood-CSF penetration in antimicrobial selection are discussed.

Proc Natl Acad Sci U S A, 1999 Mar 2, 96(5), 2274 - 8
Dopamine beta-hydroxylase deficiency impairs cellular immunity; Alaniz RC et al.; Norepinephrine, released from sympathetic neurons, and epinephrine, released from the adrenal medulla, participate in a number of physiological processes including those that facilitate adaptation to stressful conditions . The thymus, spleen, and lymph nodes are richly innervated by the sympathetic nervous system, and catecholamines are thought to modulate the immune response . However, the importance of this modulatory role in vivo remains uncertain . We addressed this question genetically by using mice that lack dopamine beta-hydroxylase (dbh-/- mice) . dbh-/- mice cannot produce norepinephrine or epinephrine, but produce dopamine instead . When housed in specific pathogen-free conditions, dbh-/- mice had normal numbers of blood leukocytes, and normal T and B cell development and in vitro function . However, when challenged in vivo by infection with the intracellular pathogens Listeria monocytogenes or Mycobacterium tuberculosis, dbh-/- mice were more susceptible to infection, exhibited extreme thymic involution, and had impaired T cell function, including Th1 cytokine production . When immunized with trinitrophenyl-keyhole limpet hemocyanin, dbh-/- mice produced less Th1 cytokine-dependent-IgG2a antitrinitrophenyl antibody . These results indicate that physiological catecholamine production is not required for normal development of the immune system, but plays an important role in the modulation of T cell-mediated immunity to infection and immunization.

Protein Expr Purif, 1999 Mar, 15(2), 243 - 5
A method for purification of listeriolysin O from a hypersecretor strain of Listeria monocytogenes; Walton CM et al.; A simple and convenient method for the purification of the hemolytic toxin listeriolysin O (LLO) from Listeria monocytogenes is described . Supernatants from bacteria cultures were purified by application to a CH2 spiral cartridge concentrator (Amicon) and ion exchange chromatography . A critical step is removal of contaminating RNA . The purified proteins had characteristics described for bacterial thiol-activated hemolysins: activation by a reducing agent (DTT) and inactivation by cholesterol . In addition, the molecular weight of 58, 000 and pH-dependent hemolytic activity of this purified protein are consistent with the previously published characteristics of LLO .

Curr Opin Cell Biol, 1999 Feb, 11(1), 117 - 21
The Arp2/3 complex: a multifunctional actin organizer; Machesky LM et al.; The actin-related proteins (Arps) constitute a recently characterized family of proteins, many of which function as members of multiprotein complexes . The discovery that two family members, Arp2 and Arp3, act as multifunctional organizers of actin filaments in all eukaryotes has generated much excitement . Over the past two years, newly discovered properties of the Arp2/3 complex have suggested a central role in the control of actin polymerization . First, it promotes actin assembly on the surface of the motile intracellular pathogen Listeria monocytogenes . Second, it can nucleate and cross-link actin filaments in vitro . Third, it localizes with dynamic actin-rich spots of mammalian cells suggesting a role in protrusion; it is found in cortical actin patches in the budding and fission yeasts where it may control patch movement and cortical actin function . Clearly, the complex has a central role in actin cytoskeletal function and will be the subject of much research in the coming years.

J Food Prot, 1999 Feb, 62(2), 170 - 6
Development and validation of a dynamic growth model for Listeria monocytogenes in fluid whole milk; Alavi SH et al.; Listeria monocytogenes, a psychrotrophic microorganism, has been the cause of several food-borne illness outbreaks, including those traced back to pasteurized fluid milk and milk products . This microorganism is especially important because it can grow at storage temperatures recommended for milk (< or =7 degrees C) . Growth of L . monocytogenes in fluid milk depends to a large extent on the varying temperatures it is exposed to in the postpasteurization phase, i.e., during in-plant storage, transportation, and storage at retail stores . Growth data for L . monocytogenes in sterilized whole milk were collected at 4, 6, 8, 10, 15, 20, 25, 30, and 35 degrees C . Specific growth rate and maximum population density were calculated at each temperature using these data . The data for growth rates versus temperature were fitted to the Zwietering square root model . This equation was used to develop a dynamic growth model (i.e., the Baranyi dynamic growth model or BDGM) for L . monocytogenes based on a system of equations which had an intrinsic parameter for simulating the lag phase . Results from validation of the BDGM for a rapidly fluctuating temperature profile showed that although the exponential growth phase of the culture under dynamic temperature conditions was modeled accurately, the lag phase duration was overestimated . For an alpha0 (initial physiological state parameter) value of 0.137, which corresponded to the mean temperature of 15 degrees C, the population densities were underpredicted, although the experimental data fell within the narrow band calculated for extreme values of alpha0 . The maximum relative error between the experimental data and the curve based on an average alpha0 value was 10.42%, and the root mean square error was 0.28 log CFU/ml.

Lett Appl Microbiol, 1999 Jan, 28(1), 71 - 5
Resistance of heat-shocked cells of Listeria monocytogenes to mano-sonication and mano-thermo-sonication; Pagan R et al.; Heat shocks did not increase the resistance of Listeria monocytogenes to an ultrasonication treatment under pressure (Mano-Sonication; MS) . While heat-shocked cells (180 min, 45 degrees C) became sixfold more heat resistant than native cells (D62 = 1.8 min vs D62 = 0.24 min), the resistance of native and heat-shocked cells to MS (200 kPa, 117 microns) was the same (DMS = 1.6 min) . The inactivation rate of non-heat-shocked cells of L monocytogenes by a combined heat/ultrasonication treatment under pressure (Mano-Thermo-Sonication; MTS) was an additive effect . On the contrary, on heat-shocked cells, the inactivation rate of MTS was greater than that of heat added to the inactivation rate of MS (synergistic effect) in the range 62-68 degrees C.

Lett Appl Microbiol, 1999 Jan, 28(1), 45 - 8
Sub-lethal damage of Listeria monocytogenes after long-term chilled storage at 4 degrees C; Dykes GA et al.; The possibility that long term in vitro chilled storage may result in sub-lethal damage to Listeria monocytogenes cells was investigated by comparing growth of chill-stored (starvation at 4 degrees C) and fresh cultures on selective and non-selective media . Growth of freshly grown cells was minimally (3-8%) affected by selective LSAMM agar compared with non-selective Brain Heart Infusion agar . In contrast, numbers of chill-stored strains were reduced by greater than 99% after direct plating on the same selective and non-selective media . Furthermore, chill-stored strains were able to grow in standard selective broth (Listeria Selective broth and Fraser broth) only if undiluted inocula (approximately 10(5)-10(6) cfu ml-1) were used, whereas they were capable of growth in Brain Heart Infusion broth even when the lowest dilutions were used (approximately 10(1) cfu ml-1) . The potential public health consequences of this finding for the isolation of Listeria monocytogenes from foods is considered.

Infect Immun, 1999 Mar, 67(3), 1303 - 9
Noncompetitive expansion of cytotoxic T lymphocytes specific for different antigens during bacterial infection; Vijh S et al.; Listeria monocytogenes is an intracellular bacterium that elicits complex cytotoxic T-lymphocyte (CTL) responses in infected mice . The responses of CTL populations that differ in antigen specificity range in magnitude from large, dominant responses to small, subdominant responses . To test the hypothesis that dominant T-cell responses inhibit subdominant responses, we eliminated the two dominant epitopes of L . monocytogenes by anchor residue mutagenesis and measured the T-cell responses to the remaining subdominant epitopes . Surprisingly, the loss of dominant T-cell responses did not enhance subdominant responses . While mice immunized with bacteria lacking dominant epitopes developed L . monocytogenes-specific immunity, their ability to respond to dominant epitopes upon rechallenge with wild-type bacteria was markedly diminished . Recall responses in mice immunized with wild-type or epitope-deficient L . monocytogenes showed that antigen presentation during recall infection is sufficient for activating memory cells yet insufficient for optimal priming of naive T lymphocytes . Our findings suggest that T-cell priming to different epitopes during L . monocytogenes infection is not competitive . Rather, T-cell populations specific for different antigens but the same pathogen expand independently.

Infect Immun, 1999 Mar, 67(3), 1125 - 30
Role of Listeria monocytogenes exotoxins listeriolysin and phosphatidylinositol-specific phospholipase C in activation of human neutrophils; Sibelius U et al.; Polymorphonuclear leukocytes (PMN) are essential for resolution of infections with Listeria monocytogenes . The present study investigated the role of the listerial exotoxins listeriolysin (LLO) and phosphatidylinositol-specific phospholipase C (PlcA) in human neutrophil activation . Different Listeria strains, mutated in individual virulence genes, as well as purified LLO were used . Coincubation of human neutrophils with wild-type L . monocytogenes provoked PMN activation, occurring independently of phagocytosis events, with concomitant elastase secretion, leukotriene generation, platelet-activating factor (PAF) synthesis, respiratory burst, and enhanced phosphoinositide hydrolysis . Degranulation and leukotriene formation were noted to be solely dependent on LLO expression, as these features were absent when the LLO-defective mutant EGD- and the avirulent strain L . innocua were used . These effects were fully reproduced by a recombinant L . innocua strain expressing LLO (INN+) and by the purified LLO molecule . LLO secretion was also required for PAF synthesis . However, wild-type L . monocytogenes was more potent in eliciting PAF formation than mutants expressing LLO, suggesting the involvement of additional virulence factors . This was even more obvious for phosphoinositide hydrolysis and respiratory burst: these events were provoked not only by INN+ but also by the LLO-defective mutant EGD- and by a recombinant L . innocua strain producing listerial PlcA . We conclude that human neutrophils react to extracellularly provided listerial exotoxins by rapid cell activation . Listeriolysin is centrally involved in triggering degranulation and lipid mediator generation, and further virulence factors such as PlcA apparently contribute to trigger neutrophil phosphoinositide hydrolysis and respiratory burst . In this way, listerial exotoxins may influence the host defense against infections with L . monocytogenes.

Immunity, 1999 Jan, 10(1), 29 - 38
Phenotype of mice and macrophages deficient in both phagocyte oxidase and inducible nitric oxide synthase; Shiloh MU et al.; The two genetically established antimicrobial mechanisms of macrophages are production of reactive oxygen intermediates by phagocyte oxidase (phox) and reactive nitrogen intermediates by inducible nitric oxide synthase (NOS2) . Mice doubly deficient in both enzymes (gp91(phox-/-)/NOS2(-/-)) formed massive abscesses containing commensal organisms, mostly enteric bacteria, even when reared under specific pathogen-free conditions with antibiotics . Neither parental strain showed such infections . Thus, phox and NOS2 appear to compensate for each other's deficiency in providing resistance to indigenous bacteria, and no other pathway does so fully . Macrophages from gp91(phox-/-)/NOS2(-/-) mice could not kill virulent Listeria . Their killing of S . typhimurium, E . coli, and attenuated Listeria was markedly diminished but demonstrable, establishing the existence of a mechanism of macrophage antibacterial activity independent of phox and NOS2.

J Exp Med, 1999 Feb 15, 189(4), 657 - 62
Impaired antibacterial host defense in mice lacking the N-formylpeptide receptor; Gao JL et al.; N-formylpeptides derive from bacterial and mitochondrial proteins, and bind to specific receptors on mammalian phagocytes . Since binding induces chemotaxis and activation of phagocytes in vitro, it has been postulated that N-formylpeptide receptor signaling in vivo may be important in antimicrobial host defense, although direct proof has been lacking . Here we test this hypothesis in mice lacking the high affinity N-formylpeptide receptor (FPR), created by targeted gene disruption . FPR-/- mice developed normally, but had increased susceptibility to challenge with Listeria monocytogenes, as measured by increased mortality compared with wild-type littermates . FPR-/- mice also had increased bacterial load in spleen and liver 2 d after infection, which is before development of a specific cellular immune response, suggesting a defect in innate immunity . Consistent with this, neutrophil chemotaxis in vitro and neutrophil mobilization into peripheral blood in vivo in response to the prototype N-formylpeptide fMLF (formyl-methionyl-leucyl-phenylalanine) were both absent in FPR-/- mice . These results indicate that FPR functions in antibacterial host defense in vivo.

J Med Microbiol, 1999 Feb, 48(2), 117 - 24
Surface protein p104 is involved in adhesion of Listeria monocytogenes to human intestinal cell line, Caco-2; Pandiripally VK et al.; Adhesion of Listeria monocytogenes to intestinal endothelial cells is an important initial event in the pathogenesis of infection which is not well understood . The suggestion has been made that some proteins, including internalin and actin polymerisation protein (ActA), and carbohydrate molecules mediate, at least in part, the adhesion of listeria to certain cultured mammalian cells . This study investigated the role of a L . monocytogenes cell-surface protein of 104 kDa (p104) in adhesion to human intestinal enterocyte-like Caco-2 cell lines by transposon (Tn916) mutagenesis and a p104-specific monoclonal antibody (MAb-H7) . Genotypic and phenotypic characteristics of Tn916-transformed L . monocytogenes strains, AAMU530 and AAMU572, revealed that these strains did not express p104, and the transposon had been inserted at a single locus in the structural gene . Strains AAMU530 and AAMU572 yielded only 10 and 6.3% adhesion to Caco-2 cells . Coating of L . monocytogenes and L . innocua wild-type strains with MAb-H7 reduced adhesion to Caco-2 cells from 100% to 50 and 45%, respectively, whereas on isotype control MAb EM-7G1 had no effect . Western blot analysis with MAb-H7 indicated that p104 is present in all Listeria spp . except in L . grayi . Furthermore, p104 is also present in internalin (BUG8) and ActA (LUT12) deficient strains, suggesting that p104 is indeed different from internalin or ActA proteins . Cytotoxicity analysis of strains AAMU530 and AAMU572 demonstrated that these strains, although haemolytic and phospholipase-positive, were avirulent when tested with a hybridoma B-lymphocyte cell line . Loss of virulence could be attributed to the interruption of adhesion of mutant strains to the hybridoma cell line . These results strongly suggest that p104 is an adhesion factor in L . monocytogenes and possibly in other Listeria species and is involved in adhesion to intestinal cells.

Mol Gen Mikrobiol Virusol, 1998, (4), 18 - 22
{Ultrastructural and immunocytochemical study of Listeria monocytogenes with varying levels of pathogenicity factor production}; Didenko LV et al.; Wild and mutant strains of Listeria monocytogenes are examined by electron and immunoelectron microscopy . The mutant strain was characterized as a strain with a high level of production of pathogenic factors . No essential morphological criteria permitting the differentiation between wild and mutant Listeria strains were detected . Addition of activated charcoal to nutrient medium resulted in similar morphological changes in both strains . Enlargement of cells, a thicker cell wall, and changes in the cytoplasm structure are objective morphological signs of functional activity of bacteria with a higher level of pathogenic factor production . Indirect immunocytochemical method demonstrated the localization of specific phosphatidyl inosityl phospholipase C.

J Immunol, 1999 Feb 15, 162(4), 2368 - 74
Immunostimulatory CpG-oligodeoxynucleotides cause extramedullary murine hemopoiesis; Sparwasser T et al.; Bacterial DNA and the synthetic CpG-oligodeoxynucleotides (ODNs) derived thereof have attracted attention because they activate cells of the adaptive immune system (lymphocytes) and the innate immune system (APCs) in a sequence-dependent manner . Here, we addressed whether CpG-ODNs affect hemopoiesis . Challenging mice with immunostimulatory CpG-ODN sequences led to transient splenomegaly, with a maximum increase of spleen weight at day 6 . The induction of splenomegaly by CpG-ODNs was sequence-specific, dose-dependent, and associated with an increase in splenic cell count, in numbers of granulocyte-macrophage CFUs (GM-CFUs), and early erythroid progenitors (burst-forming units-erythroid) . The transfer of spleen cells from CpG-ODN-pretreated animals into lethally irradiated syngeneic mice yielded an increase of spleen CFUs . Furthermore, the challenge of sublethally irradiated mice with CpG-ODNs caused radioprotective effects, in that recovery of GM-CFUs and cytotoxic T cell function was enhanced . The increase in GM-CFU and CTL function correlated with an enhanced resistance to Listeria infection in irradiated mice . We conclude from these data that CpG-ODNs trigger extramedullary hemopoiesis, and that this finding could be of therapeutic relevance in myelosuppression.

J Immunol, 1999 Feb 15, 162(4), 2291 - 8
Bacterial DNA containing CpG motifs stimulates lymphocyte-dependent protection of mice against lethal infection with intracellular bacteria; Elkins KL et al.; Bacterial DNA containing unmethylated CpG motifs activates mammalian lymphocytes and macrophages to produce cytokines and polyclonal Ig . These include IFN-gamma, IL-12, TNF-alpha, and IL-6, which are important in the control of intracellular bacterial infection . Here, we show that bacterial DNA, as well as synthetic oligonucleotides containing CpG motifs, induce protection against large lethal doses of Francisella tularensis live vaccine strain (LVS) and Listeria monocytogenes . Methylation of DNA at CpG dinucleotides or inversion of the motif abolished this protection . Surprisingly, DNA-mediated protection was highly dependent on lymphocytes, particularly B cells, as well as the production of IFN-gamma . Optimal protection was elicited 2-3 days after inoculation with DNA and persisted for up to 2 wk . Further, animals surviving lethal challenge developed pathogen-specific secondary immunity . These findings indicate that host innate immune responses to bacterial DNA may contribute to the induction of protective immunity to bacteria and the subsequent development of memory.

J Immunol, 1999 Feb 1, 162(3), 1723 - 9
A transgenic model to analyze the immunoregulatory role of IL-10 secreted by antigen-presenting cells; Groux H et al.; IL-10 is a cytokine secreted by a wide variety of cells type that has pleiotropic stimulatory and suppressive activities on both lymphoid and myeloid cells in vitro . To analyze the consequences of high IL-10 secretion by APCs in immune responses, we produced transgenic mice expressing human IL-10 directed by the MHC class II Ea promoter . Despite alterations in the development of T and B cells, no gross abnormalities were detected in peripheral lymphocyte populations or serum Ig levels . However, when immunized using conditions that give either a Th2-type or a Th1-type response, IL-10 transgenic mice failed to mount a significant T or B cell immune response to OVA . IL-10 transgenic mice were also highly susceptible to infection with intracellular pathogens like Listeria monocytogenes or Leishmania major, in contrast to IL-10 transgenic mice, where the transgene was express in T cells . Finally, the recently described stimulatory effect of IL-10 on CD8+ T cells was confirmed by the ability of IL-10 transgenic mice to limit the growth of immunogenic tumors by a CTL-mediated mechanism . These results demonstrate, that, depending on the type of immune response, IL-10 can mediate immunosuppressive or immunostimulatory activities in vivo.

J Toxicol Environ Health A, 1999 Feb 12, 56(3), 205 - 19
Xenobiotic-metabolizing enzymes in carp (Cyprinus carpio) liver, spleen, and head kidney following experimental Listeria monocytogenes infection; Chambras C et al.; Infection of carp with Listeria monocytogenes 4b resulted in decreased liver, spleen, and head kidney enzyme activities, involved in the metabolism of xenobiotics . After infection, cytochrome P-450 levels and ethoxyresorufin O-deethylase (EROD) activity were decreased while conjugation enzymes remained unaffected . The maximum decrease for phase I enzymes occurred on d 3 . This loss of monooxygenase levels and activity could not be directly correlated with an increase in the number of organisms, as consistently high bacterial counts were observed in all three organs during infection . The effect of L . monocytogenes infection was also measured in carp exposed to 3-methylcholanthrene (MCA) . Cytochrome P-450 levels and EROD activity were significantly reduced, especially on d 3 . A significant decreased activity of conjugation enzymes such as glutathione S-transferase (GST) and UDP-glucuronosyltransferase (UDPGT) was also observed for all days studied . Listeria infection inhibited MCA-induced increases in xenobiotic-metabolizing enzyme activities . These results indicate that infection may have deleterious effects on basal cytochrome P-450 monooxygenase levels . Furthermore, MCA treatment aggravates the insult to xenobiotic biotransformation enzymes by L . monocytogenes infection, by impairing a number of detoxification enzymes . These findings could result in significant changes in the susceptibility of fish to pollutants.

FEMS Microbiol Lett, 1999 Jan 15, 170(2), 349 - 53
Characterisation of Listeria ivanovii isolates from the UK using pulsed-field gel electrophoresis; Ramage CP et al.; Forty-three Listeria ivanovii isolates were collected in the UK between 1991 and 1997 from: 35 animal infections; two human infections; five foods; and one environmental source . A further two type strains of L . ivanovii (subsp . ivanovii and subsp . londoniensis) were obtained from a culture collection . These bacteria were characterised by conventional phenotypic methods and by pulsed-field gel electrophoresis (PFGE) using ApaI and SmaI . Forty-two of the isolates from the UK were identified as L . ivanovii subsp . ivanovii and the remaining culture as L . ivanovii subsp . londoniensis . Six and four PFGE profiles were obtained using ApaI and SmaI digestion respectively; six composite profiles were obtained combining the results for both enzymes . The PFGE profile of the UK L . ivanovii subsp . londoniensis (isolated from processed shrimps) was similar to the type strain of this subspecies and differed from all of the L . ivanovii subsp . ivanovii tested . The majority of isolates (38 out of 45) belonged to one profile showing that the UK population of this bacterium is much less genetically diverse than similar studies have shown for Listeria monocytogenes.

Biochem J, 1999 Feb 15, 338 ( Pt 1), 71 - 5
Incorporation of iron by the unusual dodecameric ferritin from Listeria innocua; Stefanini S et al.; The polypeptide chain that assembles into the unusual dodecameric shell of Listeria innocua apoferritin lacks the ferroxidase centre characteristic of H-type mammalian chains, but is able to catalyse both Fe(II) oxidation and nucleation of the iron core . A cluster of five carboxylate residues, which correspond in part to the site of iron core nucleation typical of L-type mammalian ferritins, has been proposed to be involved in both functions . The features of the iron uptake kinetics and of Fe(II) autoxidation in the presence of citrate followed spectrophotometrically confirm this assignment . In Listeria the kinetics of iron uptake is hyperbolic at low Fe(II)-to-dodecamer ratios and becomes sigmoidal when iron exceeds 150 Fe(II) atoms per dodecamer, namely when a fast crystal growth phase follows a slow initial nucleation step . Iron autoxidation in the presence of citrate displays a similar behaviour . Thus the time course is sigmoidal at low citrate-to-Fe ratios at which Fe(III) polymerization is predominant, but is hyperbolic at ligand concentrations high enough to prevent polymerization . The marked inhibitory effect of Tb(III) on the kinetics of iron incorporation confirms that carboxylates provide the iron ligands in L . innocua apoferritin . Iron uptake followed in steady-state fluorescence experiments allows one to distinguish Fe(II) binding and oxidation from the subsequent movement of Fe(III) into the apoferritin cavity as in mammalian ferritins despite the different localization of the tryptophan residues.

Gastroenterol Hepatol, 1998 Dec, 21(10), 489 - 91
{Spontaneous bacterial peritonitis caused by Listeria monocytogenes}; Jorquera Plaza F et al.; We present a 68 year old male with alcoholic cirrhosis that was admitted with abdominal pain and fever . Hepatocarcinoma and spontaneous bacterial peritonitis by Listeria monocytogenes was diagnosed . The patient was treated with ampicillin and tobramycin during 25 days following a favorable course although ascitic fluid remained abnormal during 21 days . It is noted the rarity of Listeria as a cause of bacterial peritonitis in cirrhotic patients although they are immunodeficient . It is also important to establish the etiological origin because standard treatment of spontaneous bacterial peritonitis is cefotaxime and Listeria is resistant to this antibiotic . The 66% of spontaneous bacterial peritonitis secondary to Listeria monocytogenes infection in cirrhotic patients has been reported in Spain and this might be due to a higher incidence of human listeriosis in this country.

J Food Prot, 1999 Jan, 62(1), 46 - 50
Combined effect of nisin and moderate heat on destruction of Listeria monocytogenes in cold-pack lobster meat; Budu-Amoako E et al.; The combined effect of nisin and moderate heat to increase the killing of Listeria monocytogenes in cans of "cold-pack" lobster was investigated . Adding nisin at a level of 25 mg/kg of can contents to the brine surrounding the lobster, in combination with a heat process giving internal can temperatures of 60 degrees C for 5 min and 65 degrees C for 2 min, resulted in decimal reductions of inoculated L . monocytogenes of 3 to 5 logs, whereas heat or nisin alone resulted in decimal reductions of 1 to 3 logs . Such a reduced heat process to that currently commercially used (65.5 degrees C for 13 to 18 min, depending on the can size) results in significant reduction in drained weight loss, thus allowing considerable cost savings to the lobster-processing industry.

J Food Prot, 1999 Jan, 62(1), 40 - 5
High-pressure destruction kinetics of Listeria monocytogenes on pork; Mussa DM et al.; Packaged fresh pork chops (30-g samples) containing an indigenous bacterial population of approximately 10(7) CFU/g were inoculated with 10(7) CFU of Listeria monocytogenes Scott A per g, heat sealed, and subjected to high-pressure processing at 200 to 400 MPa for up to 90 min . Total counts and the number of surviving L . monocytogenes cells were determined by a spread plate technique on tryptic soy agar and modified Oxford medium, respectively . The pressure destruction was characterized by a dual-behavior, consisting of a step change in the number of survivors (Pk0) with the application of a pressure pulse and a first-order rate drop in the number of survivors during the pressure hold period . Higher pressures resulted in higher rates of microbial inactivation, as indicated by their associated lower D values (and higher k values) . The pressure sensitivities of the kinetic parameters were evaluated on the basis of Arrhenius and pressure death time (PDT)-type models . The results suggested that L . monocytogenes was more resistant to pressure inactivation than the indigenous microflora (the volume change of activation, deltaV* {Arrhenius model}), and Zp values (PDT model) were -4.17 x 10(-5) m3 mole(-1) and 134 MPa for indigenous microflora and -3.43 x 10(-5) m3 mole(-1) and 163 MPa for L . monocytogenes respectively.

MMWR Morb Mortal Wkly Rep, 1999 Jan 8, 47(51-52), 1117 - 8
Update: multistate outbreak of listeriosis--United States, 1998-1999; Restraint stress-induced immunosuppression by inhibiting leukocyte migration and Th1 cytokine expression during the intraperitoneal infection of Listeria monocytogenes; Department of Immunology, Medical Institute of Bioregulation, Kyushu University, Fukuoka, JapanIn this study, a murine model of Listeria monocytogenes infection was used to investigate effects of restraint stress (RST) on host defense . We observed that the L . monocytogenes infection as well as RST induced an elevation of endogenous corticosterone (CORT) levels and RST synergistically enhanced endogenous CORT levels during the listerial infection . RST suppressed the migration of leukocytes including macrophages, neutrophils, NK cells and lymphocytes into the peritoneal cavities after the intraperitoneal inoculation of L . monocytogenes . RST also suppressed the increase of the surface MHC class II antigen expression in both peritoneal macrophages and B cells during the listerial infection . Interestingly, gene expression of iNOS, MCP-1 (JE) and Th1-type cytokines including IFN-gamma and IL-12 was down-regulated but Th2-type cytokine (IL-4 and IL-6) gene expression in the PEC was rather up-regulated on day 7 after infection, indicating that Th2-type immune response is more resistant to the elevated endogenous CORT levels than Th1-type response . Treatment of mice with RU486, a glucocorticoid receptor antagonist, restored the immune responses suppressed by RST to their normal levels in the infected mice, suggesting that the RST-induced elevation of endogenous corticosterone levels is mainly responsible for the induction of the immunosuppressive events during L . monocytogenes infection.

J Immunol, 1999 Jan 15, 162(2), 980 - 8
Perforin-deficient CD8+ T cells: in vivo priming and antigen-specific immunity against Listeria monocytogenes; White DW et al.; CD8+ T cells require perforin to mediate immunity against some, but not all, intracellular pathogens . Previous studies with H-2b MHC perforin gene knockout (PO) mice revealed both perforin-dependent and perforin-independent pathways of CD8+ T cell-mediated immunity to Listeria monocytogenes (LM) . In this study, we address two previously unresolved issues regarding the requirement for perforin in antilisterial immunity: 1) Is CD8+ T cell-mediated, perforin-independent immunity specific for a single Ag or generalizable to multiple Ags? 2) Is there a deficiency in the priming of the CD8+ T cell compartment of PO mice following an immunizing challenge with LM? We used H-2d MHC PO mice to generate CD8+ T cell lines individually specific for three known Ags expressed by a recombinant strain of virulent LM . Adoptive transfer experiments into BALB/c host mice revealed that immunity can be mediated by PO CD8+ T cells specific for all Ags examined, indicating that perforin-independent immunity is not limited to CD8+ T cells that recognize listeriolysin O . Analysis of epitope-specific CD8+ T cell expansion by MHC class I tetramer staining and ELISPOT revealed no deficiency in either the primary or secondary response to LM infection in PO mice . These results demonstrate that the perforin-independent pathway of antilisterial resistance mediated by CD8+ T cells is generalizable to multiple epitopes . Furthermore, the results show that reduced antilisterial resistance observed with polyclonal PO CD8+ T cells is a consequence of a deficiency in effector function and not a result of suboptimal CD8+ T cell priming.

J Immunol, 1999 Jan 15, 162(2), 965 - 73
IL-12 is not required for induction of type 1 cytokine responses in viral infections; Oxenius A et al.; To investigate the physiological role of IL-12 in viral infections in terms of T cell cytokine responses involved in virus-specific Ig isotype induction and in antiviral protection, immune responses elicited upon infection of IL-12-deficient mice with lymphocytic choriomeningitis virus (LCMV) or vesicular stomatitis virus (VSV) were studied . Infection of IL-12-deficient mice with LCMV induced a virus-specific type 1 cytokine response as determined by in vitro cytokine secretion patterns as well as by in vivo intracellular cytokine staining of LCMV-specific CD4+ TCR transgenic T cells that had clonally expanded in LCMV-infected IL-12-deficient recipient mice . In addition, LCMV- and VSV-specific IgG responses exhibited normal serum IgG2a/IgG1 ratios, demonstrating again virus-specific CD4+ T cell induction of type 1 phenotype in IL-12-deficient mice upon viral infection . LCMV and VSV immune mice were found to be protected against challenge immunization with recombinant vaccinia viruses expressing either the LCMV- or the VSV-derived glycoprotein, respectively . This protection is known to be mediated by T cell-secreted type 1 cytokines IFN-gamma and TNF-alpha . In contrast, IL-12-deficient mice showed impaired abilities to control infection with the facultative intracellular bacterium Listeria monocytogenes at early time points after infection . However, at later time points of infection, IL-12-deficient mice were able to clear infection . These findings may indicate that viruses are able to induce type 1 T cell responses in the absence of IL-12 as opposed to some bacterial or parasitical infections that are crucially dependent on the presence of IL-12 for the induction of type 1 immune responses.

Infect Immun, 1999 Feb, 67(2), 568 - 75
Induction of protective T cells against Listeria monocytogenes in mice by immunization with a listeriolysin O-negative avirulent strain of bacteria and liposome-encapsulated listeriolysin O; Tanabe Y et al.; Only listeriolysin O (LLO)-producing strains of Listeria monocytogenes generate protective immunity in mice . Based on the findings that endogenous gamma interferon (IFN-gamma) production was induced only by such strains and that purified LLO could induce IFN-gamma from NK cells, we have postulated that LLO may play a pivotal role in the induction of Th1-type protective T cells, which are highly dependent on IFN-gamma . In this study, mice were immunized with L . monocytogenes ATCC 15313, an LLO-nonproducing avirulent strain, along with LLO encapsulated in liposome (LLO-liposome) . LLO-liposome was highly potent in the induction of various cytokines, including IFN-gamma . Immunization of mice with either LLO-liposome or the viable strain ATCC 15313 alone did not induce protection against challenge infection . In contrast, the combination of LLO-nonproducing bacteria plus LLO-liposome induced a significant level of protective immunity mediated mainly by Th1-type cells capable of producing a large amount of IFN-gamma in an antigen-specific manner . The protection afforded by the combination was not dependent on LLO-specific cytotoxic T cells . These results support the idea that the inability of an LLO-nonproducing avirulent strain or killed bacteria to induce the generation of protective T cells is due not to the lack of a central T-cell epitope(s) but to the lack of ability to induce the production of endogenous cytokine during the early stage of immunization; the results also suggest that an appropriate use of LLO at least in an animal model may be effective in the induction of antigen-specific Th1-dependent protective immunity to various kinds of intracellular parasitic bacteria.

MMWR Morb Mortal Wkly Rep, 1998 Dec 25, 47(50), 1085 - 6
Multistate outbreak of listeriosis--United States, 1998; Intramedullary brain stem cyst and trapped IV ventricle after infection with Listeria monocytogenes; Neurochirurgische Klinik im Neurozentrum, Albert-Ludwigs-Universitat, Freiburg i.Br., GermanyWe present the rare case of a girl surviving intrauterine listeria brain stem meningoencephalitis, who subsequently developed hydrocephalus, a trapped IV ventricle and an intramedullary cyst . Such cases have been reported only infrequently, and in earlier cases modern imaging studies were not available . Magnetic resonance imaging has been helpful in our patient to delineate the lesions and plan further treatment.

Prev Vet Med, 1998 Dec 1, 37(1-4), 129 - 45
Quantitative risk assessment of human listeriosis from consumption of soft cheese made from raw milk; Bemrah N et al.; Microbial hazards have been identified in soft cheese made from raw milk . Quantification of the resulting risk for public health was attempted within the frame of the Codex Alimentarius Commission, 1995 approach to quantitative risk assessment, using Monte Carlo simulation software . Quantitative data could only be found for Listeria monocytogenes . The complete process of cheese making was modeled, from milking to consumption . Using data published on the different sources of milk contamination (environment and mastitis) and bacterial growth, distributions were assumed for parameters of the model . Equations of Farber, J.M., Ross, W.H., Harwing, J . (1996) for general and at-risk populations were used to link the ingested dose of L . monocytogenes to the occurrence of listeriosis . The probability of milk contamination was estimated to be 67% with concentration ranging from 0 to 33 CFU ml-1 . The percentage of cheese with a predicted concentration of L . monocytogenes greater than 100 CFU g-1 was low (1.4%) . The probability of consuming a contaminated cheese serving was 65.3% . Individual annual cumulative risk of listeriosis, in a population each consuming 50 servings of 31 g, ranged from 1.97 x 10(-9) to 6.4 x 10(-8) in a low-risk sub-population and 1.04 10(-6) to 7.19 10(-5) in a high-risk sub-population . The average number of expected cases of listeriosis per year was 57 for a high-risk sub-population and one for a low-risk healthy sub-population . When the frequency of environmental milk contamination was reduced in the model and L . monocytogenes mastitis was eliminated, the expected incidence of listeriosis decreased substantially; the average number of expected cases was reduced by a factor of 5 . Thus the usefulness of simulation to demonstrate the efficiency of various management options could be demonstrated, even if results should be interpreted with care (as many assumptions had to be made on data and their distributions.

J Vet Med Sci, 1998 Dec, 60(12), 1341 - 3
Detection of Listeria monocytogenes in humans, animals and foods; Iida T et al.; The Listeria monocytogenes-carrying rates were 100% for listeriosis patients and 1.3% for healthy humans . The L . monocytogenes contamination rates for retail sliced beef (34.2%) and pork (36.4%) were significantly higher (p < 0.05) than those for cattle (2.0%) and pigs (0.8%) and for cattle (4.9%) and swine (7.4%) carcasses . The percentages of serotypes 1/2a, 1/2b and 4b which are most dominant in human patients were high in isolates from fresh (90.0%) and processed (100%) fish and shellfish and imported natural cheese (96.7%).

Physiol Behav, 1998 Dec 1, 65(3), 535 - 43
2-deoxy-D-glucose-induced metabolic stress enhances resistance to Listeria monocytogenes infection in mice; Miller ES et al.; Exposure to different forms of psychological and physiological stress can elicit a host stress response, which alters normal parameters of neuroendocrine homeostasis . The present study evaluated the influence of the metabolic stressor 2-deoxy-D-glucose (2-DG; a glucose analog, which when administered to rodents, induces acute periods of metabolic stress) on the capacity of mice to resist infection with the facultative intracellular bacterial pathogen Listeria monocytogenes . Female BDF1 mice were injected with 2-DG (500 mg/kg b . wt.) once every 48 h prior to, concurrent with, or after the onset of a sublethal dose of virulent L . monocytogenes . Kinetics of bacterial growth in mice were not altered if 2-DG was applied concurrently or after the start of the infection . In contrast, mice exposed to 2-DG prior to infection demonstrated an enhanced resistance to the listeria challenge . The enhanced bacterial clearance in vivo could not be explained by 2-DG exerting a toxic effect on the listeria, based on the results of two experiments . First, 2-DG did not inhibit listeria replication in trypticase soy broth . Second, replication of L . monocytogenes was not inhibited in bone marrow-derived macrophage cultures exposed to 2-DG . Production of neopterin and lysozyme, indicators of macrophage activation, were enhanced following exposure to 2-DG, which correlated with the increased resistance to L . monocytogenes . These results support the contention that the host response to 2-DG-induced metabolic stress can influence the capacity of the immune system to resist infection by certain classes of microbial pathogens.

Appl Environ Microbiol, 1999 Jan, 65(1), 150 - 5
Sources of Listeria monocytogenes contamination in a cold-smoked rainbow trout processing plant detected by pulsed-field gel electrophoresis typing; Autio T et al.; Sites of Listeria monocytogenes contamination in a cold-smoked rainbow trout (Oncorhynchus mykiss) processing plant were detected by sampling the production line, environment, and fish at different production stages . Two lots were monitored . The frequency of raw fish samples containing L . monocytogenes was low . During processing, the frequency of fish contaminated with L . monocytogenes clearly rose after brining, and the most contaminated sites of the processing plant were the brining and postbrining areas . A total of 303 isolates from the raw fish, product, and the environment were characterized by pulsed-field gel electrophoresis (PFGE) . PFGE yielded nine pulsotypes, which formed four clusters . The predominating L . monocytogenes pulsotypes of the final product were associated with brining and slicing, whereas contaminants of raw fish were not detected in the final product . Air-mediated contamination in the plant could not be proved . In accordance with these results, an L . monocytogenes eradication program was planned . The use of hot steam, hot air, and hot water seemed to be useful in eliminating L . monocytogenes . None of the control samples taken in the 5 months after the eradication program was implemented contained L . monocytogenes.

Lett Appl Microbiol, 1998 Dec, 27(6), 362 - 8
Natural milk fatty acids affect survival and invasiveness of Listeria monocytogenes; Petrone G et al.; The effects of 12 fatty acids, naturally occurring in milk from several mammalian species, on the survival and invasion ability of Listeria monocytogenes, a food-borne pathogen, were determined . The survival was tested in the presence of 200 micrograms ml-1 fatty acids suspended in brain hearth infusion broth or in storage conditioning solution (NaCl 1%) of Mozzarella cheese, an Italian soft unripened cheese, at pH 7.0 or 5.0 . Lauric (C12:0), linoleic (C18:2) and linolenic (C18:3) acids exerted the strongest bactericidal activity . The invasive efficiency of L . monocytogenes, determined in the Caco-2 enterocyte-like cell line, was strongly decreased in the presence of the fatty acids tested (from about 20 to 500-fold) . This research suggests that naturally occurring fatty acids of milk, supplemented in milk derivatives, could affect both bacterial growth and invasiveness and consequently, could serve as barriers towards L . monocytogenes infection.

Kekkaku, 1998 Nov, 73(11), 639 - 44
{Mechanisms of induction and expression of anti-tuberculous immunity}; Mitsuyama M; The induction of anti-tuberculous immunity highly depends on the cytokines produced endogenously at the initial stage of immunization . Among several cytokines, IFN-gamma appears to be the most important to generate antigen-specific Th1 type of protective T cells in mice . IL-12 and IL-18, which are produced by macrophages in response to virulent mycobacteria, are responsible for stimulating NK cells to produce IFN-gamma . Once antigen-specific Th1 cells are generated, Th1-dependent macrophage activation was effective in the elimination of infected bacteria through enhanced production of reactive oxygen intermediates and reactive nitrogen intermediates . In Listeria monocytogenes, one of the intracellular bacteria, listeriolysin O (LLO) appeared to be responsible for the induction of endogenous IFN-gamma from NK cells . The possible mechanisms operating in the induction and expression of anti-tuberculous immunity are discussed with special reference to cytokine responses . An application of LLO to the induction of protective immunity is also discussed.

Clin Ter, 1998 Jul-Aug, 149(4), 307 - 11
{A case of maternal and neonatal infection due to Listeria monocytogenes}; Tridente V et al.; A Listeria monocytogenes infection may develop during pregnancy by eating sausages, fresh meats and milk products derived from infected animals . According to the period in which the infection starts, the pregnancy outcome can be abortion or pre-term or at term delivery . The infection can pass from mother to fetus and can cause a serious neonatal sepsis . Listeriosis in pregnant women can be asymptomatic or may present as an influenza-like syndrome . This case report, along with other published cases, demonstrates how hard is to make a correct diagnosis of listeriosis during pregnancy . Since this is mainly related to the aspecificity of symptoms, it is very important to have a high suspicion and to take a careful patient history.

Infect Immun, 1999 Jan, 67(1), 253 - 8
Existing antilisterial immunity does not inhibit the development of a Listeria monocytogenes-specific primary cytotoxic T-lymphocyte response; Bouwer HG et al.; Infection of BALB/c mice with Listeria monocytogenes stimulates an antilisterial immune response evident by the appearance of H2-Kd-restricted CD8(+) cytotoxic T lymphocytes (CTLs) specific for the nanomer peptides amino acids (aa) 91 to 99 of listeriolysin O (LLO 91-99) and aa 217 to 225 of the p60 molecule (p60 217-225) . We have introduced point mutations at anchor residues within LLO 91-99 (92F) or p60 217-225 (218F), and BALB/c mice infected with L . monocytogenes strains containing these point mutations do not develop CTLs specific for LLO 91-99 or p60 217-225, respectively . We have used these strains to test whether primary CTL responses against L . monocytogenes-derived determinants can be stimulated within an environment of existing antilisterial immunity . We found that the development of a primary L . monocytogenes-specific CTL response is not altered by existing immunity to L . monocytogenes . For example, primary immunization with the p60 218F strain of L . monocytogenes followed by a secondary immunization with wild-type L . monocytogenes results in stimulation of p60 217-225-specific CTLs at primary response levels and LLO 91-99-specific effectors at levels consistent with a memory CTL response . Similarly, primary immunization with the 92F strain of L . monocytogenes followed by a secondary immunization with wild-type L . monocytogenes results in stimulation of LLO 91-99-specific CTLs at primary response levels and p60 217-225-specific effectors at levels consistent with a memory CTL response . These results provide additional support for the use of L . monocytogenes as a recombinant vaccine vector and show that antivector immunity does not inhibit the development of a primary CTL response when the epitope is delivered by L . monocytogenes as the vaccine strain.

Infect Immun, 1999 Jan, 67(1), 182 - 6
Mutagenesis of active-site histidines of Listeria monocytogenes phosphatidylinositol-specific phospholipase C: effects on enzyme activity and biological function; Bannam T et al.; Listeria monocytogenes, a gram-positive facultative intracellular pathogen, produces two distinct phospholipases C . PC-PLC, encoded by plcB, is a broad-range phospholipase, whereas PI-PLC, encoded by plcA, is specific for phosphatidylinositol . It was previously shown that PI-PLC plays a role in efficient escape of L . monocytogenes from the primary phagosome . To further understand the function of PI-PLC in intracellular growth, site-directed mutagenesis of plcA was performed . Two potential active-site histidine residues were mutated independently to alanine, serine, and phenylalanine . With the exception of the activity of the enzyme containing H38F, which was unstable, the PI-PLC enzyme activities of culture supernatants containing each mutant enzyme were <1% of wild-type activity . In addition, the levels of expression of the mutant PI-PLC proteins were equivalent to wild-type expression . Derivatives of L . monocytogenes containing these specific plcA mutations were found to have phenotypes similar to that of the plcA deletion strain in an assay for escape from the primary vacuole, in intracellular growth in a murine macrophage cell line, and in a plaquing assay for cell-to-cell spread . Thus, catalytic activity of PI-PLC is required for all its intracellular functions.

J Immunol, 1998 Dec 15, 161(12), 6715 - 23
Two families of GTPases dominate the complex cellular response to IFN-gamma; Boehm U et al.; IFN-gamma induces a number of cellular programs functional in innate and adaptive resistance to infectious pathogens . It has recently become clear that the complete cellular response to IFN-gamma is extraordinarily complex, with >500 genes (i.e., approximately 0.5% of the genome) activated . We made suppression-subtractive hybridization differential libraries from IFN-gamma-stimulated primary mouse embryonic fibroblasts and from a mouse macrophage cell line, ANA-1, in each case with reference to unstimulated cells . Of approximately 250 clones sequenced at random from the two libraries, >35% were representatives of one or the other of two small unrelated families of GTPases, the 65-kDa and 47-kDa families . These families dominate the IFN-gamma-induced response in both cell types . We report here the full-length sequences of one new 65-kDa and two new 47-kDa family members . The 65-kDa family members are under transcriptional control of IRF-1, whereas the 47-kDa family members are inducible in embryonic fibroblasts from IRF-1(-/-) mice . Members of both GTPase families are strongly up-regulated in livers of wild-type mice infected with the pathogenic bacterium, Listeria monocytogenes, but not in IFN-gammaR(0/0) mice . These GTPases appear to be dedicated to the IFN-gamma response, since resting levels are negligible and since neither family shows any significant relationship to any other described family of GTPases . Understanding the role of these GTPases in IFN-gamma-mediated resistance against pathogens is the task for the future.

Mol Gen Genet, 1998 Nov, 260(2-3), 144 - 58
The gene cluster inlC2DE of Listeria monocytogenes contains additional new internalin genes and is important for virulence in mice; Raffelsbauer D et al.; In this work we identified and characterized a gene cluster containing three internalin genes of Listeria monocytogenes EGD . These genes, termed inlG, inlH and inlE, encode proteins of 490, 548 and 499 amino acids, respectively, which belong to the family of large, cell wall-bound internalins . The inlGHE gene cluster is flanked by two listerial house-keeping genes encoding proteins homologous to the 6-phospho-beta-glucosidase and the succinyl-diaminopimelate desuccinylase of E . coli . A similar internalin gene cluster, inlC2DE, localised to the same position on the L . monocytogenes EGD chromosome was recently described in a different isolate (Dramsi S, Dehoux P, Lebrun M, Goossens PL, Cossart P (1997) Infect Immun 65: 1615-1625) . Sequence comparison of the two inl gene clusters indicates that inlG is a new internalin gene, while inlH was generated by a site-specific recombination, leading to an in-frame deletion which removed the 3'-terminal end of inlC2 and the 5'-terminal part of inlD . The third gene of the inlGHE cluster, inlE, is almost identical to the previously reported inlE gene . Our data show that the inlGHE gene cluster is probably transcribed from a major PrfA-independent promoter located upstream of inlG . PCR analysis revealed the presence of the newly identified inl genes inlG and inlH in most L . monocytogenes isolates tested . A mutant which has lost inlG, inlH and inlE by an in-frame deletion exhibited, after oral infection of mice, a significant loss in virulence and shows drastically reduced numbers of viable bacteria in both liver and spleen when compared to the wild-type strain.

J Bacteriol, 1998 Dec, 180(24), 6655 - 60
Functional similarities between the Listeria monocytogenes virulence regulator PrfA and cyclic AMP receptor protein: the PrfA* (Gly145Ser) mutation increases binding affinity for target DNA; Vega Y et al.; Most Listeria monocytogenes virulence genes are positively regulated by the PrfA protein, a transcription factor sharing sequence similarities with cyclic AMP (cAMP) receptor protein (CRP) . Its coding gene, prfA, is regulated by PrfA itself via an autoregulatory loop mediated by the upstream PrfA-dependent plcA promoter . We have recently characterized prfA* mutants from L . monocytogenes which, as a result of a single amino acid substitution in PrfA, Gly145Ser, constitutively overexpress prfA and the genes of the PrfA virulence regulon . Here, we show that about 10 times more PrfA protein is produced in a prfA* strain than in the wild type . Thus, the phenotype of prfA* mutants is presumably due to the synthesis of a PrfA protein with higher promoter-activating activity (PrfA*), which keeps its intracellular levels constantly elevated by positive feedback . We investigated the interaction of PrfA and PrfA* (Gly145Ser) with target DNA . Gel retardation assays performed with a DNA fragment carrying the PrfA binding site of the plcA promoter demonstrated that the PrfA* mutant form is much more efficient than wild-type PrfA at forming specific DNA-protein complexes . In footprinting experiments, the two purified PrfA forms interacted with the same nucleotides at the target site, although the minimum amount required for protection was 6 to 7 times lower with PrfA* . These results show that the primary functional consequence of the Gly145Ser mutation is an increase in the affinity of PrfA for its target sequence . Interestingly, similar mutations at the equivalent position in CRP result in a transcriptionally active, CRP* mutant form which binds with high affinity to target DNA in the absence of the activating cofactor, cAMP . Our observations suggest that the structural similarities between PrfA and CRP are also functionally relevant and support a model in which the PrfA protein, like CRP, shifts from transcriptionally inactive to active conformations by interaction with a cofactor.

Int J Food Microbiol, 1998 Oct 20, 44(1-2), 141 - 4
Initial numbers, serovars and phagevars of Listeria monocytogenes isolated in prepared foods in the city of Barcelona (Spain); de Simon M et al.; A total of 1100 samples of prepared foods purchased in restaurants and delicatessen shops in the city of Barcelona was examined for the presence of Listerio . L . monocytogenes was more frequently isolated from foods intended to be consumed without further cooking (9.3%) than from foods intended to receive further cooking prior to consumption (2.9%) . A quantitative study, carried out with 773 samples, yielded 1% of samples with numbers of L . monocytogenes higher than 100 CFU/g . Strains of L . monocytogenes belonged to serovar 4b (36.3%), 1/2a (29.5%), 1/2b (22.7%) and 1/2c (11.3%) . L . monocytogenes serovar 4b phagevar 3274: 2671 (6 of 11) was prevalent among the strains studied.

Int J Food Microbiol, 1998 Oct 20, 44(1-2), 83 - 92
The effect of the growth environment on the lag phase of Listeria monocytogenes; Robinson TP et al.; The duration of lag in Listeria monocytogenes was examined in relation to the physico-chemical properties of the growth environment . It was supposed that lag would be determined by two hypothetical quantities, the amount of work that a cell has to perform to adapt to new conditions and the rate at which it can perform that work . If the rate at which the cell can perform the necessary work is a function of the maximum specific growth rate in the new environment, the hypothesis predicts that lag time should be related in some way to growth rate, provided cells are initially in approximately the same physiological state . Literature data suggest this is true for many organisms when temperature is the sole growth limiting factor . However, lag times of L . monocytogenes displayed an unusual response to temperature in which lag times of cells precultured at 37 degrees C were shorter at 15 degrees C than at 20 degrees C or 25 degrees C . Analysis of data from the Food Micromodel in which growth of L . monocytogenes was controlled by combinations of pH, NaCl concentration and temperature, showed that there was a linear relationship between lag time and mean generation time although there was much scatter in the data . When the effects of pH, solute type and concentration were investigated individually in this work the correlation between lag time and mean generation time was often poor . It would thus appear that the relationship between growth environment and lag time is more complex than the corresponding relationship between growth environment and maximum specific growth rate.

Dig Surg, 1998, 15(4), 364 - 8
Listeria monocytogenes causing solitary liver abscess . Case report and review of the literature; Bronnimann S et al.; The authors report on a case of a solitary liver abscess due to Listeria monocytogenes in a 53-year-old diabetic white male and review all published cases of solitary listerial abscesses of the liver . L . monocytogenes is a rare cause of solitary liver abscess which occurs in elderly patients with diabetes mellitus . The clinical signs are variable and often mimic malignancy, with epigastric pain, night sweats and weight loss . Prevalent features are poor control of glycemia, temperature up to 38.5 degrees C and elevated alkaline phosphatase . Optimal treatment includes percutaneous drainage of the hepatic abscess and antibiotic therapy with an aminopenicillin or trimethoprim/sulfamethoxazole . Outcome of the reviewed patients was favourable with no mortality and no relapse of the disease.

Eur J Cell Biol, 1998 Oct, 77(2), 81 - 90
The suitability and application of a GFP-actin fusion protein for long-term imaging of the organization and dynamics of the cytoskeleton in mammalian cells; Choidas A et al.; The product of a GFP-actin gene fusion, permanently or transiently transfected in diverse mammalian cell lines, was shown to be a suitable, intrinsic probe of both the organization and dynamics of the actin cytoskeleton . In live Swiss 3T3 and NIH 3T3 cells, the fusion protein was found to accumulate in lamellipodia, filopodia, focal contacts and stress fibers . Furthermore, comparisons of fluorescence images of GFP-actin and Cy3.5-phalloidin, an independent marker of F-actin, in permeabilized cells showed a complete overlap of the two fluorescence signals . In GFP-actin-transfected Hela cells that had been infected with Listeria monocytogenes, the fluorescence of the fusion protein was shown to dynamically associate in the F-actin rich comet tail that formed behind a motile bacterium . In stable transfectants of PC12 cells, GFP-actin constituted on the average 5% of the total actin - these cells exhibited normal growth behavior and responded to treatment with nerve growth factor by extending neurite-like extensions, the filopodia-like tips of which were densely packed with filamentous GFP-actin . Finally, the photobleaching decay time of GFP-actin in live cells of 63 seconds was much longer than that of fluorescein-labeled actin conjugates and little or no damage to the cytoskeleton was found during the photobleaching of GFP-actin . Having shown the suitability of GFP-actin as a probe of the cytoskeleton, its fluorescence was used in long-term imaging studies aimed at documenting changes in the cytoskeleton of rat bladder NBT-II carcinoma cells during the 24-hour growth factor-mediated epithelia to mesenchyme transformation . The intrinsic fluorescent probe was also used to investigate the organization of the actin cytoskeleton and behavior of individual mesenchyme NBT-II cells slowly migrating through a colony of epithelia cells.

Vet Immunol Immunopathol, 1998 Oct 23, 65(2-4), 125 - 38
Effect of feline immunodeficiency virus on cytokine response to Listeria monocytogenes in vivo; Dean GA et al.; Feline immunodeficiency virus (FIV) is a lentivirus that induces an acquired immunodeficiency in domestic cats . The objective of this study was to compare the immune response of chronically FIV-infected cats and specific pathogen free (SPF) cats to Listeria monocytogenes, a facultative intracellular bacterium . Regional lymph nodes were removed at various times after subcutaneous inoculation with L . monocytogenes and evaluated . Lymph nodes of chronically FIV-infected cats enlarged more slowly and to a lesser degree than SPF cats . This was due to delayed and blunted lymphoid follicle formation and markedly diminished histiocyte influx . The cellular response correlated with a marked upregulation in IL10 transcription and delayed increase in TNF-alpha upregulation in FIV-infected cats . Transcriptional upregulation of IFN-gamma, IL4, and the p40 chain of IL12 was similar in lymph nodes of FIV-infected and SPF cats . Clinically, FIV-infected cats had a more severe response at the site of L . monocytogenes injection and showed signs of systemic bacterial dissemination while SPF cats remained clinically normal . FIV-infected cats generated a delayed hypersensitivity response similar to SPF cats but also had a significantly greater antibody response . Taken together, these data suggest excessive IL10 production may be responsible for the deficiency observed in the innate immune response of chronically FIV-infected cats challenged with L . monocytogenes.

Appl Environ Microbiol, 1998 Dec, 64(12), 4883 - 90
Purification and genetic characterization of enterocin I from Enterococcus faecium 6T1a, a novel antilisterial plasmid-encoded bacteriocin which does not belong to the pediocin family of bacteriocins; Floriano B et al.; Enterocin I (ENTI) is a novel bacteriocin produced by Enterococcus faecium 6T1a, a strain originally isolated from a Spanish-style green olive fermentation . The bacteriocin is active against many olive spoilage and food-borne gram-positive pathogenic bacteria, including clostridia, propionibacteria, and Listeria monocytogenes . ENTI was purified to homogeneity by ammonium sulfate precipitation, binding to an SP-Sepharose fast-flow column, and phenyl-Sepharose CL-4B and C2/C18 reverse-phase chromatography . The purification procedure resulted in a final yield of 954% and a 170,000-fold increase in specific activity . The primary structure of ENTI was determined by amino acid and nucleotide sequencing . ENTI consists of 44 amino acids and does not show significant sequence similarity with any other previously described bacteriocin . Sequencing of the entI structural gene, which is located on the 23-kb plasmid pEF1 of E . faecium 6T1a, revealed the absence of a leader peptide at the N-terminal region of the gene product . A second open reading frame, ORF2, located downstream of entI, encodes a putative protein that is 72.7% identical to ENTI . entI and ORF2 appear to be cotranscribed, yielding an mRNA of ca . 0.35 kb . A gene encoding immunity to ENTI was not identified . However, curing experiments demonstrated that both enterocin production and immunity are conferred by pEF1.

Rev Med Chil, 1998 Jul, 126(7), 828 - 32
{Listeria monocytogenes rhomboencephalitis . Clinical case}; Bravo M et al.; We report a previously healthy 44 years old female, that presented with mild clouding of consciousness, a left cerebellar syndrome, involvement of V, X and XII left cranial nerves and an alteration of epicritic sensitivity in the left half of the body . Cerebrospinal fluid had inflammatory features . Cerebrospinal fluid and blood cultures were positive for Listeria monocytogenes . Magnetic resonance imaging disclosed a rhomboencephalitis . Antibiotics were started and the clinical condition of the patient improved progressively . After three months of follow up, the patient is notably recovered and there is a regression of hyperintense lesions of the brainstem in the magnetic resonance imaging . The diagnosis of Listeria monocytogenes infection must be born in mind in the presence of a rhomboencephalitis.

Lett Appl Microbiol, 1998 Nov, 27(5), 275 - 8
Variations in tolerance of Listeria monocytogenes to nisin, pediocin PA-1 and bavaricin A; Rasch M et al.; A collection of 381 Listeria monocytogenes strains was examined for strain variations in nisin and pediocin sensitivity at three different concentrations on Tryptic Soya Agar . Two of the strains were able to grow on agar plates containing 500 IU nisin ml-1 . These strains were characterized as having an enhanced nisin tolerance . Twenty strains had normal growth on agar plates containing 1600 AU pediocin ml-1 and higher . These strains were characterized as pediocin-resistant . Another 34 strains, characterized as having enhanced tolerance, exhibited inhibited growth at 1600 AU pediocin ml-1 . Bavaricin sensitivity, examined for 22 strains of L . monocytogenes, correlated with the pediocin sensitivity of the strains . It is therefore assumed that there is accordance between the pediocin and bavaricin sensitivity of the remaining strains . Cross-resistance between nisin and pediocin/bavaricin was not found.

J Food Prot, 1998 Nov, 61(11), 1475 - 9
Behavior of Listeria spp . in smoked fish products affected by liquid smoke, NaCl concentration, and temperature; Thurette J et al.; The growth, survival, and death of Listeria monocytogenes were studied in a synthetic medium as a function of temperature, NaCl content, and amount of liquid smoke, and the findings were validated in smoked fish products . The smoke preservative compound was simulated by adding liquid smoke, and the concentration was expressed as phenol concentration . The growth of L . monocytogenes was limited at a temperature as low as 4 degrees C or at a phenol concentration as high as 20 ppm . The predicted values were obtained using a mathematical model established in liquid medium in a previous study . They accurately fit values observed in L . monocytogenes challenge tests on smoked fish . After 21 days of storage the deviation between the predicted and experimental values was within 0.5 log for 60% of the data . This model may be useful in predicting Listeria contamination in smoked fish . Moreover, this study emphasizes the importance of phenol concentration to control the growth of Listeria spp . in smoked food products.

J Food Prot, 1998 Nov, 61(11), 1470 - 4
Organic acid dipping of catfish fillets: effect on color, microbial load, and Listeria monocytogenes; Bal'a MF et al.; Microbiological and color changes of catfish fillets were determined following dip treatment in solutions at 4 degrees C of 2% acetic, citric, hydrochloric, lactic, malic, or tartaric acid . Fillets were inoculated with an eight-strain mixture of Listeria monocytogenes prior to dipping . L . monocytogenes, coliform, and aerobic plate counts and surface pH and Hunter color were measured at 0, 2, 5, and 8 days of storage at 4 degrees C . Acid dipping reduced surface pH and L . monocytogenes, coliform, and aerobic microbial loads . Little microbial proliferation was observed on acid-treated fillets, however, controls had a distinct foul odor and microbial loads in excess of 10(6) CFU/g by day 8 . On untreated fillets, L . monocytogenes counts did not increase during storage, perhaps due to competitive inhibition by normal catfish microflora . Hunter color analysis revealed lighter and yellower acid-treated fillets than untreated controls, with malic acid producing the least bleaching . The shelf life of refrigerated fillets increased when fillets were acid dipped . It remains to be established if this enhanced microbial quality also parallels sensory acceptability.

J Food Prot, 1998 Nov, 61(11), 1465 - 9
Survival of Listeria spp . on raw whole chickens cooked in microwave ovens; Farber JM et al.; The prevalence of microwave ovens in North American homes has increased dramatically within the last decade . Although microwave ovens are primarily used for reheating of foods, they are now more commonly being applied to the cooking of raw foods . Although cooking of raw foods, according to manufacturers' instructions targets an organoleptically acceptable end product, the process does not address the microbiological safety of the cooked food . Seventeen microwave ovens from various commercial suppliers were used to cook naturally contaminated whole raw broilers (< or = 1.8 kg) and roasters (> 1.8 kg) according to manufacturers' instructions . Temperature probes (six per chicken) were used to measure the temperature of chickens immediately after cooking and during the holding period . Of 81 Listeria-positive raw broilers and 93 raw roasters, 1 (1.2%) and 9 (9.7%), respectively, yielded viable Listeria spp . after microwave cooking . Of these, two were undercooked (visual inspection), one was over the maximum weight stipulated by the oven manufacturer and another one was over the maximum weight and undercooked . A significantly greater proportion of contaminated cooked birds was observed with roasters than with broilers, where for one of these contaminated roasters, the temperature at all six measured sites was > or = 87 degrees C . Most of the postcook Listeria-positive birds were associated with 2 of the 17 microwave ovens . Factors such as wattage, cavity size, and the presence or absence of a turntable seemingly did not play a significant role in the survival of Listeria spp . in microwave-cooked chicken . However, the general inability of microwave ovens to uniformly heat chicken carcasses was noted . In order to promote greater safety of microwave-cooked foods, general recommendations for consumers are provided.

Prenat Diagn, 1998 Oct, 18(10), 1075 - 8
Ultrasound features of congenital listeriosis--a case report; Quinlivan JA et al.; We present a case of congenital listeriosis in a twin pregnancy . Presentation was prompted by decreased fetal movements and an ultrasound examination which demonstrated features similar to those observed in an adult with inflammatory conditions of the bowel, namely, small amounts of ascites, dilated loops of bowel and thickening of the bowel wall . Such ultrasound signs may be useful in consideration of the diagnosis of congenital infection and prompt diagnostic tests and appropriate interventions that assist in early therapy.

Infect Immun, 1998 Dec, 66(12), 5930 - 8
Phosphatidylcholine-specific phospholipase C from Listeria monocytogenes is an important virulence factor in murine cerebral listeriosis; Schluter D et al.; Meningoencephalitis is a serious and often fatal complication of Listeria monocytogenes infection . The aim of the present study was to analyze the role of internalin A (InlA) and B, which are involved in the invasion of L . monocytogenes into cultivated host tissue cells, and that of phosphatidylcholine-specific phospholipase C (PlcB), which mainly promotes the direct cell-to-cell spread of L . monocytogenes, in murine cerebral listeriosis by use of an InlA/B (DeltainlAB2)- and a PlcB (DeltaplcB2)-deficient isogenic deletion mutant strain and the wild-type (WT) L . monocytogenes EGD . Listeria strains were directly applied to the brain, a technique which has been employed previously to study the pathogenesis of cerebral listeriosis (D . Schluter, S . B . Oprisiu, S . Chahoud, D . Weiner, O . D . Wiestler, H . Hof, and M . Deckert-Schluter, Eur . J . Immunol . 25:2384-2391, 1995) . We demonstrated that PlcB, but not InlA or InlB, is an important virulence factor in cerebral listeriosis . Nonimmunized mice infected intracerebrally with the DeltaplcB2 strain survived significantly longer and had a reduced intracerebral bacterial load compared to mice infected with the DeltainlAB2 strain or WT bacteria . In addition, immunization with the WT prior to intracerebral infection significantly increased the survival rate of mice challenged intracerebrally with the DeltaplcB2 strain compared to that of mice infected with the WT or DeltainlAB2 strain . Histopathology revealed that the major difference between the various experimental groups was a significantly delayed intracerebral spread of the DeltaplcB2 mutant strain, indicating that cell-to-cell spread is an important pathogenic feature of cerebral listeriosis . Interestingly, irrespective of the Listeria mutant used, the apoptosis of hippocampal and cerebellar neurons and an internal hydrocephalus developed in surviving mice, indicating that these complications are not dependent on the virulence factors InlA/B and PlcB . In conclusion, this study points to PlcB as a virulence factor important for the intracerebral pathogenesis of murine L . monocytogenes meningoencephalitis.

Infect Immun, 1998 Dec, 66(12), 5677 - 83
Interleukin-15 may be responsible for early activation of intestinal intraepithelial lymphocytes after oral infection with Listeria monocytogenes in rats; Hirose K et al.; Exogenous interleukin-15 (IL-15) stimulates intestinal intraepithelial lymphocytes (i-IEL) from mice to proliferate and produce gamma interferon (IFN-gamma) in vitro . To determine whether endogenous IL-15 is involved in activation of i-IEL during intestinal infection, we examined IL-15 synthesis by intestinal epithelial cells (i-EC) after infection with Listeria monocytogenes in rats . In in vitro experiments, invasion of L . monocytogenes into IEC-6 cells, a rat small intestine epithelial cell line, evidently induced IL-15 mRNA expression coincident with nuclear factor kappaB (NF-kappaB) activation, which is essential for IL-15 gene expression . IL-15 synthesis was detected in rat i-EC on day 1 after an oral inoculation of L . monocytogenes in vivo . The numbers of T-cell receptor (TCR) gamma delta+ T cells, NKR.P1(+) cells, and CD3(+) CD8(+) alpha alpha cells in i-IEL were significantly increased on day 1 after oral infection . The i-IEL from infected rats produced larger amounts of IFN-gamma upon stimulation with immobilized anti-TCR gamma delta or anti-NKR.P1 monoclonal antibodies . These results suggest that IL-15 produced by i-EC may stimulate significant fractions of i-IEL to produce IFN-gamma at an early phase of oral infection with L . monocytogenes.

Immunology, 1998 Nov, 95(3), 395 - 401
Gamma delta T cells in infection-induced and autoimmune-induced testicular inflammation; Mukasa A et al.; In a previous report, we investigated inflammatory responses induced by injecting Listeria monocytogenes into one testis of a mouse . We demonstrated that the contralateral testis also developed an orchitis despite the absence of bacteria, indicating that the inflammation on the uninfected, contralateral side was of autoimmune character . In both infected and autoimmune testes, gammadelta and alphabeta T cells infiltrated during the inflammation . In this paper, we present the data of a comparison of the character of gammadelta T cells of the infected and autoimmune testes . In both testes, gammadelta T cells appeared to be activated, as assessed by high CD44 and low l-selectin expression . Analysis of T-cell receptor (TCR) usage in both inflammation types revealed the same gammadelta TCR repertoire . Finally, the semi-quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) demonstrated that gammadelta T cells in both types of inflammation were capable of producing interleukin-2 (IL-2), IL-4, interferon-gamma (IFN-gamma), IL-10 and transforming growth factor-beta (TGF-beta) . These results imply that gammadelta T cells present in infected-induced and autoimmunity-induced inflammation have the same characteristics and could work as immunoregulatory cells.

Immunology, 1998 Oct, 95(2), 226 - 33
The role of gammadelta T cells in induction of bacterial antigen-specific protective CD8+ cytotoxic T cells in immune response against the intracellular bacteria Listeria monocytogenes; Nomura A et al.; The role of T-cell receptor (TCR) gammadelta T cells in the induction of protective TCR alphabeta T cells against infection by the intracellular bacteria Listeria monocytogenes was analysed . We found that depletion of gammadelta T cells by anti-TCR delta monoclonal antibody treatment before intravenous immunization of mice with a sublethal dose of viable L . monocytogenes resulted in reduction of protection against secondary challenge infection in the immunized mice . The gammadelta T-cell depletion also reduced induction of protective alphabeta T cells capable of transferring the protection against challenge infection of L . monocytogenes into naive mice . Furthermore, the protective T cells that were affected by the gammadelta T-cell depletion were suggested to be CD8+ cytotoxic T cells rather than CD4+ T cells by the following observations . First, induction of cytotoxic T lymphocytes specific to a L . monocytogenes-derived H-2Kd-restricted peptide (listeriolysin O 91-99) was significantly suppressed by gammadelta T-cell depletion before immunization . Second, gammadelta T-cell depletion did not affect cytokine production and proliferation of T cells from immunized mice in response to in vitro stimulation with heat-killed Listeria which preferentially stimulates CD4+ T cells . Third, CD8+ alphabeta T cells from control immunized mice transferred protection against infection of L . monocytogenes into naive mice but only a limited degree of protection was transferred by CD8+ T cells from the gammadelta T-cell-depleted immunized mice; and fourth, CD4+ alphabeta T cells from the gammadelta T-cell-depleted mice transferred a similar level of protection as those from the control immunized mice . All these results suggest that gammadelta T cells participate in establishment of protective immunity against intracellular bacteria by supporting priming of bacterial antigen-specific CD8+ cytotoxic T cells.

Anesthesiology, 1998 Nov, 89(5), 1125 - 32
Intraoperative modulation of alveolar macrophage function during isoflurane and propofol anesthesia; Kotani N et al.; BACKGROUND: Alveolar macrophages are a critical part of the defense against pulmonary infection . Thus the authors determined time-dependent changes in alveolar macrophage functions in patients having surgery who were anesthetized with isoflurane or propofol . METHODS: Patients anesthetized with propofol (n = 30) or isoflurane (n = 30) during orthopedic surgery were studied . Alveolar macrophages were harvested by bronchoalveolar lavage immediately, and 2, 4, and 6 h after induction anesthesia and at the end of surgery . The fraction of aggregated and nonviable macrophages was determined . Then phagocytosis was measured by ingestion of opsonized and unopsonized particles . Finally, microbicidal activity was determined as the ability of the macrophages to kill Listeria monocytogenes directly . RESULTS: Demographic and morphometric characteristics of the patients given propofol and isoflurane were similar, as were their levels of pulmonary function and hemodynamic responses . The fraction of alveolar macrophages ingesting opsonized and unopsonized particles, and the number of particles ingested, decreased significantly over time, with the decrease slightly but significantly greater during isoflurane anesthesia . Microbicidal function decreased progressively during anesthesia and surgery, with the decrease almost twice as great during isoflurane compared with propofol anesthesia . The fraction of aggregated macrophages and recovered neutrophils increased over time in the patients given each anesthetic . CONCLUSIONS: Pulmonary immunologic function changed progressively during anesthesia and surgery . The data from this study suggest that pulmonary defenses are modulated by the type of anesthesia and by the duration of anesthesia and surgery.

J Immunol, 1998 Nov 15, 161(10), 5600 - 6
Early IFN-gamma production and innate immunity during Listeria monocytogenes infection in the absence of NK cells; Andersson A et al.; NK cells are believed to play a mandatory role during the early phases of Listeria monocytogenes infection by producing IFN-gamma, which is required for the activation of macrophage effector functions . Mice deficient in the common cytokine receptor gamma-chain (gamma(c)-/-), which completely lack NK cells, were used to examine whether NK cells were essential for resistance to Listeria infection in vivo . Surprisingly, infected gamma(c)-/- mice showed normal innate immunity and macrophage responses against sublethal Listeria infection 2 days postinfection . At this time point, gamma(c)-/- mice showed increased blood IFN-gamma levels compared with those in noninfected controls, demonstrating an NK-independent source of IFN-gamma, which explains early resistance . Listeria-infected gamma(c)-/- x recombinase-activating gene-2-/- double-deficient mice were unable to produce IFN-gamma and were highly susceptible to L . monocytogenes . Since T cells, but not B cells, are major IFN-gamma producers, and gamma(c)-/- T cells were found to be efficient IFN-gamma producers in vitro, we conclude from these results that T cells functionally replace NK cells for the early IFN-gamma production that is necessary for activating the innate immune system following infection with L . monocytogenes . This novel observation in listeriosis underscores how the adaptive immune response can maintain and influence innate immunity.

J Immunol, 1998 Nov 15, 161(10), 5594 - 9
Optimization of codon usage of plasmid DNA vaccine is required for the effective MHC class I-restricted T cell responses against an intracellular bacterium; Uchijima M et al.; In an attempt to study codon usage effects of DNA vaccines on the induction of MHC class I-restricted T cell responses against an intracellular bacterium, Listeria monocytogenes, we designed two plasmid DNA vaccines encoding an H-2Kd-restricted epitope of listeriolysin O (LLO) of L . monocytogenes, LLO 91-99 . One DNA vaccine, p91wt, carries the wild-type DNA sequence encoding LLO 91-99, and the other one, p91mam, possesses the altered DNA sequence in which the codon usage was optimized for murine system . Our read-through analyses with LLO 91-99/luciferase fusion genes confirmed that the optimized 91mam DNA sequence showed extremely higher translation efficiency than the wild-type sequence in murine cells . Consistent with this, i.m . injections of p91mam, but not of p91wt, into BALB/c mice were capable of inducing specific CTL- and IFN-gamma-producing CD8+ T cells able to confer partial protection against listerial challenge . Taken together, these observations suggest that optimization of codon should be taken into consideration in the construction of DNA vaccines against nonviral pathogens.

J Infect, 1998 Jul, 37(1), 77 - 8
Listeria monocytogenes meningitis in a penicillin-allergic paediatric renal transplant patient; Weston VC et al.; Currently in many centres the extended spectrum cephalosporins (e.g . cefotaxime and ceftriaxone) are being used empirically for patients with suspected bacterial meningitis . We present a case of meningitis in a penicillin allergic paediatric renal transplant patient from whose cerebrospinal fluid (CSF) Listeria monocytogenes was cultured, despite four days of cefotaxime therapy . The patient was successfully treated with meropenem but required neuro-endoscopic intervention for hydrocephalus.

J Infect Dis, 1998 Dec, 178(6), 1658 - 66
Listeria monocytogenes infection and activation of human brain microvascular endothelial cells; Wilson SL et al.; Listeria monocytogenes invasion of human brain microvascular endothelial cells (BMEC) and its role as a stimulus for endothelial cell activation were studied . Binding and invasion of intact BMEC monolayers were independent of the L . monocytogenes inlAB invasion locus . Cytochalasin D abrogated invasion of BMEC, whereas genistein effected only a 53% decrease in invasion, indicating a requirement for rearrangement of actin microfilaments but less dependence on tyrosine kinase activity . L . monocytogenes stimulated surface expression of E-selectin, ICAM-1, and to a lesser extent, VCAM-1, whereas L . monocytogenes prfA- and Deltahly mutants were severely compromised in this respect . Other experiments showed that BMEC infection stimulated monocyte and neutrophil adhesion and that CD18-mediated binding was the predominant mechanism for neutrophil adhesion to infected BMEC under static conditions . These data suggest that invasion of BMEC is a mechanism for triggering inflammation and leukocyte recruitment into the central nervous system during bacterial meningitis.

J Am Mosq Control Assoc, 1998 Sep, 14(3), 253 - 5
Bionomics and cytogenetics of Anopheles seretsei Abdulla-Khan, Coetzee, and Hunt, a new species from northern Botswana; Abdulla-Khan R et al.; Morphological analysis of anopheline mosquitoes from Kasane, Botswana, revealed a new species, Anopheles seretsei, that is closely related to Anopheles azevedoi and Anopheles listeri . A description of the type locality and biological characteristics of An . seretsei is given . Comparisons are made with An . listeri and An . azevedoi . The banding patterns of the giant polytene chromosomes of An . seretsei were compared with those of An . listeri and found to be homosequential.

J Am Mosq Control Assoc, 1998 Sep, 14(3), 248 - 52
Description of Anopheles (Cellia) seretsei sp . nov . from Kasane, Botswana; Abdulla-Khan R et al.; Anopheles (Cellia) seretsei, a new mosquito species from Kasane, northern Botswana, is described on the basis of the examination of a type series of 37 females and 18 males . Diagnostic features of the egg, larva, and pupa are presented . Comparisons of the new species with close relatives (Anopheles listeri and Anopheles azevedoi) are made.

Lett Appl Microbiol, 1998 Oct, 27(4), 198 - 202
Unidentified Listeria-like bacteria isolated from cheese; Rijpens N et al.; Unidentified Listeria-like bacteria, which lack only one of the phenotypic characteristics used to confirm Listeria spp., were isolated from cheese during routine analysis for Listeria monocytogenes . The VIDAS Listeria assay and the Listeria specific PCR or DNA probe assays used did not identify these strains as Listeria species . This group of bacteria was studied for its homogeneity using rep-PCR and PFGE . Sequence analysis of the 16S rRNA gene showed a homology of 94% to established Listeria spp., implicating a closer relationship than that between Listeria spp . and Brochothrix spp.

Zentralbl Bakteriol, 1995 Oct, 282(4), 384 - 8
Correlation between production of listeriolysin O by variants of Listeria monocytogenes and their virulence for rabbits; Waseem M et al.; L . monocytogenes serovar 1/2a NCTC 7973 was passaged through rabbits and the severity of infection at each passage was determined by counting viable bacteria from infested organs and recording the time of death . A comparative evaluation of the levels of hemolysin produced in vitro by the original and six variant cultures (V1-V6) was done by determination of hemolytic units (CHU) . While virulence of the cultures enhanced at each passage (2.2 x 10(9) CFU/g of the spleen for V6 as compared to 5.0 x 10(6) CFU/g spleen for the parent culture), the CHU decreased considerably, 3 CHU for the V6 as compared to 40 CHU for the parent strain . The results suggest that the level of in vitro production of listeriolysin may not parallel the pathogenicity of L . monocytogenes for rabbits.

Zentralbl Bakteriol, 1995 Nov, 283(1), 29 - 42
Simplified purification of Listeria monocytogenes listeriolysin O and preliminary application in the enzyme-linked immunosorbent assay (ELISA); Traub WH et al.; A simplified procedure was developed for purification of listeriolysin O (LLO) of Listeria monocytogenes, consisting of hydroxylapatite adsorption chromatography followed by Sepharose S ion exchange chromatography . The LLO (58 kDa) appeared pure in terms of sodium dodecylsulfate polyacrylamide electrophoresis and immunoblots with polyclonal rabbit immune sera . The purified LLO could be stored at -65 degrees C for 1 year without loss of immunoreactivity . Similarly, flat-bottom microtiter strips from two vendors that had been charged with LLO, could be stored at -65 degrees C for up to 3 months without loss of LLO . Three patients with documented listeriosis developed elevated IgG titres against LLO; 2 of the patients revealed minimally raised IgM titres, as determined with the enzyme-linked immunosorbent assay.

J Exp Med, 1998 Nov 2, 188(9), 1651 - 6
Interferon gamma derived from CD4(+) T cells is sufficient to mediate T helper cell type 1 development; Wakil AE et al.; Interferon gamma (IFN-gamma) has been implicated in T helper type 1 (Th1) cell development through its ability to optimize interleukin 12 (IL-12) production from macrophages and IL-12 receptor expression on activated T cells . Various systems have suggested a role for IFN-gamma derived from the innate immune system, particularly natural killer (NK) cells, in mediating Th1 differentiation in vivo . We tested this requirement by reconstituting T cell and IFN-gamma doubly deficient mice with wild-type CD4(+) T cells and challenging the mice with pathogens that elicited either minimal or robust IL-12 in vivo (Leishmania major or Listeria monocytogenes, respectively) . Th1 cells developed under both conditions, and this was unaffected by the presence or absence of IFN-gamma in non-T cells . Reconstitution with IFN-gamma-deficient CD4(+) T cells could not reestablish control over L . major, even in the presence of IFN-gamma from the NK compartment . These data demonstrate that activated T cells can maintain responsiveness to IL-12 through elaboration of endogenous IFN-gamma without requirement for an exogenous source of this cytokine.

Int J Food Microbiol, 1998 Sep 8, 43(3), 223 - 9
A small outbreak of listeriosis associated with smoked mussels; Brett MS et al.; Two perinatal listeriosis cases in Auckland, New Zealand, which were diagnosed during November and December 1992 gave histories of consuming Brand X smoked mussels . Listeria monocytogenes was isolated from an unopened packet of mussels collected from the refrigerator of one of the cases . Cultures of L . monocytogenes from the two patients were indistinguishable from those recovered from the mussels by serogrouping and DNA macrorestriction analysis using pulsed-field gel electrophoresis (PFGE) . PFGE analysis of isolates of an additional fifteen clinical cases caused by L . monocytogenes serogroup 1/2 which occurred in New Zealand during 1991 and 1992, revealed two more isolates with indistinguishable PFGE patterns . PFGE analysis of a further 222 L . monocytogenes serogroup 1/2 isolated in New Zealand from food and the environment did not reveal any more cultures with the Brand X PFGE type . A combination of serotyping, phage typing, DNA restriction fragment length polymorphism analysis (RFLP), cadmium and arsenic sensitivity testing, and PFGE analysis was used to subtype 38 isolates of L . monocytogenes . The isolates comprised: (a) the isolates from four patients; (b) 26 isolates from 15 packets of Brand X mussel products from retail sale, the processing factory, and the refrigerator of one of the patients; (c) seven isolates from environmental swabs taken in the processing factory; and (d) an isolate from Brand X mussel product from a wholesaler in the United Kingdom . The isolates from three of the clinical cases, 26 of the products and four from the factory environment were indistinguishable using all the subtyping systems . This is the first description from New Zealand of a link between cases of listeriosis, a contaminated food, and the food production environment which was microbiologically confirmed using a combination of subtyping methods for L . monocytogenes.

J Food Prot, 1998 Oct, 61(10), 1299 - 304
The contamination of pâté with Listeria monocytogenes--results from the 1994 European Community-Coordinated Food Control Program for England and Wales; Nichols G et al.; The Listeria monocytogenes contamination of 3,065 pate products sampled at the point of retail sale in England and Wales was examined . Ninety-seven percent of samples were free of contamination with L . monocytogenes, 2.0% (60) had levels of less than 200 CFU/g, and 0.6% (18) had levels of 200 CFU/g or more . Fish and seafood pate were significantly more commonly contaminated by L . monocytogenes than other pate types (chi 2 test, P = 0.001) . Pate obtained from small retail shops was significantly more likely to be contaminated at levels of > or = 200 CFU/g (chi 2 tests, P < 0.0005) than that obtained from supermarkets . L . monocytogenes was isolated significantly more often (chi 2 tests, P < 0.00002) from packs of pate that were open at the time of collection (3.8%) than those that were sold prepacked (1.2%) . There were also significantly more samples (chi 2 test, P = 0.0009) where L . monocytogenes was recovered at higher levels (> or = 200 CFU/g) in opened, as compared to prepacked, samples . There was a significant difference in the rates and levels of contamination of opened samples between shops and supermarkets (chi 2 tests, P < 0.0025) . Evidence from this study shows that most of the pate sold in England and Wales is not contaminated with L . monocytogenes, and we suggest that the main areas of concern are cross-contamination and the length of display of pate sold from opened packs.

J Food Prot, 1998 Oct, 61(10), 1293 - 8
Effect of environmental stress on the ability of Listeria monocytogenes Scott A to attach to food contact surfaces; Smoot LM et al.; Attachment of Listeria monocytogenes Scott A to Buna-N rubber and stainless steel under different temperature and pH conditions at the time of cell growth or at the time of attachment was investigated . All experiments were conducted using sterile phosphate buffer to avoid cell growth during exposure to the test surfaces . Numbers of attached cells increased with increasing attachment temperature (10 to 45 degrees C) and exposure time for both test surfaces . Maximum levels of attached cells were obtained when cell growth occurred at 30 degrees C . Downward, but not upward, shifts in the cell suspension holding temperature prior to attachment to Buna-N rubber resulted in reduced adhered cell populations . Maximum levels of adhered cells to Buna-N rubber were not affected by adjustments of the attachment medium pH between 4 and 9 . However, after short contact times (i.e., less than 30 min), levels of attached cells were lower when attachment occurred under alkaline conditions . Growth pH was also found to affect the levels of adhered cell populations to Buna-N rubber . L . monocytogenes Scott A attached to stainless steel at higher levels for all temperature and pH parameters evaluated in this study.

J Food Prot, 1998 Oct, 61(10), 1286 - 92
Influence of environmental stress on the kinetics and strength of attachment of Listeria monocytogenes Scott A to Buna-N rubber and stainless steel; Smoot LM et al.; Attachment and detachment of Listeria monocytogenes Scott A to Buna-N rubber and stainless steel under varying conditions of temperature and pH were investigated using model systems . Numbers of attached cells increased with increasing attachment temperature (10 to 45 degrees C) and time (up to 120 min) for both test surfaces . Compared to Buna-N rubber, the rate of attachment to stainless steel was markedly more rapid for all temperature and pH conditions studied and could not be calculated . Rate of attachment to Buna-N rubber was found to be significantly lower when cells were attached at 10 degrees C . Growth temperature did not significantly affect rates of adhesion to Buna-N rubber . Altering the medium pH during attachment between 4 and 9 demonstrated that rates of adhesion were slower under alkaline conditions . Growth pH was also found to significantly affect rates of attachment to Buna-N rubber . Detachment of cells adhered to Buna-N rubber was significantly affected by growth temperature but not growth pH . Significant differences in detachment were also found between Buna-N rubber and stainless steel, inferring stronger attachment to Buna-N rubber . Cell surface hydrophobicity was found to be affected by both growth temperature and growth pH . However, changes in hydrophobicity could not be correlated to differences in rates of attachment . Addition of 0.01% trypsin to the attachment medium during cell exposure to either test surface resulted in a 99.9% reduction in the adhered cell population when compared to controls . This would suggest that proteins play a role in the initial attachment process of L . monocytogenes.

J Food Prot, 1998 Oct, 61(10), 1281 - 5
Antimicrobial effects of D-3-phenyllactic acid on Listeria monocytogenes in TSB-YE medium, milk, and cheese; Dieuleveux V et al.; D-3-Phenyllactic acid is a compound with anti-Listeria activity which is produced and secreted by the yeastlike fungus, Geotrichum candidum . This compound has a bactericidal effect independent of the physiological state of Listeria monocytogenes when added at a concentration of 7 mg/ml to tryptic soy broth supplemented with yeast extract (TSB-YE) . An initial L . monocytogenes population of 10(5) CFU/ml was reduced 100-fold (2 log) after 4 days of culture at 25 degrees C in TSB-YE containing D-3-phenyllactic acid . The Listeria population was reduced 1,000-fold (3 log) when the compound was added during the exponential growth phase, and was reduced to less than 10 CFU/ml when it was added during the stationary phase . D-3-Phenyllactic acid had a bacteriostatic effect in UHT whole milk, reducing the population by 4.5 log, to give fewer cells than in the control after 5 days of culture . The results obtained with L . monocytogenes at concentrations of 10(5) and 10(3) CFU/ml in cheese curds were less conclusive . D-3-Phenyllactic acid was 10 times less active than nisin in our experimental conditions (TSB-YE at 25 degrees C).

Mol Microbiol, 1998 Oct, 30(2), 405 - 17
A novel PrfA-regulated chromosomal locus, which is specific for Listeria ivanovii, encodes two small, secreted internalins and contributes to virulence in mice; Engelbrecht F et al.; Several large, cell wall-associated internalins and one small, secreted internalin (InlC) have been described previously in Listeria monocytogenes . Using degenerate primers derived from sequenced peptides of an L . ivanovii major secreted protein, we identified a new 4.25 kb internalin locus of L . ivanovii, termed i-inlFE . The two proteins encoded by this locus, i-InlE and i-InlF, belong to the group of small, secreted internalins . Southern blot analyses show that the i-inlFE locus does not occur in L . monocytogenes . These data also indicate that six genes encoding small, secreted internalins are present in L . ivanovii, in contrast to L . monocytogenes, in which inlC encodes the only small internalin . The mature i-InlE protein (198 amino acids) is secreted in large amounts into the brain-heart infusion (BHI) culture medium in the stationary growth phase . In minimum essential medium (MEM), which has been used previously to induce PrfA-dependent gene transcription, i-inlE mRNA and i-InlE protein are expressed at high levels . As shown by Northern blot analysis and primer extension, transcription of the tandemly arranged i-inlF and i-inlE genes is dependent on the virulence regulator PrfA, and characteristic palindromic sequences ('PrfA-boxes') were identified in the promoter regions of i-inlF and i-inlE . Non-polar i-inlE and i-inlF deletion mutants and an i-inlFE double deletion mutant were constructed and tested in the mouse infection model . After intravenous infection, all three mutants entirely failed to kill C57BL/6 mice even at high infectious doses of 109 bacteria per mouse, whereas the LD50 for the parental strain was determined as 4 x 107 bacteria per mouse . These data suggest an important role for i-InlE and i-InlF in L . ivanovii virulence.

Am J Perinatol, 1998 Aug, 15(8), 461 - 7
Perinatal listeriosis: a population-based multicenter study in Barcelona, Spain (1990-1996); Nolla-Salas J et al.; The aim off this study was to describe the incidence, epidemiology, clinical presentation, and outcome of perinatal listeriosis for a 7-year period (1990-1996) based on data of an active population-based surveillance project implemented in the city of Barcelona, Spain . There were 30 cases (20.8%) associated with pregnancy (15 pregnant women, 13 neonates, and 2 fetal deaths) . The incidence of perinatal listeriosis varied from 4.1 to 0 per 10,000 live births . The proportion of perinatal cases in relation to the total number of cases of listeriosis varied between 0 and 42% . Early-onset neonatal sepsis accounted for 12 of 13 live births . The mean age of infected pregnant women with listeriosis was 30.1+/-2.0 years . Chorioamnionitis was the predominant clinical form (86.7%) . Only two mothers had primary bacteremia by L . monocytogenes in the second trimester of pregnancy . Both infants were born healthy, without signs of infection . One of these mothers was infected with the human immunodeficiency virus (HIV) . Since January 1994, 12 strains were available for serotyping and phagotyping; 9 belonged to serovar 4b, 2 to serovar 1/2b, and 1 to serovar 1/2a . No outbreaks of L . monocytogenes infection occurred during the study period.The overall neonatal mortality rate was 7.7% among infected live births . All pregnant women were treated with ampicillin and none died . Early antenatal treatment with ampicillin improves neonatal outcome and can result in the birth of healthy babies.

Cent Eur J Public Health, 1998 Aug, 6(3), 254 - 5
Detection of Listeria in raw and pasteurized milk; Ahrabi SS et al.; One hundred raw milk samples from different regions of Anatolia and 20 pasteurized milk samples from three different manufacturers in Ankara were analyzed for the presence of Listeria spp . L . monocytogenes was found in 1% of the raw milk samples and in 5% (1/20) of the pasteurized milk samples . L . innocua and L . seeligeri were found in 8 and 2% of the raw milk samples, respectively . No other species of Listeria was found . The overall incidence of Listeria spp . was 10% in the raw milk samples and 5% in the pasteurized milk samples.

Int J Gynecol Pathol, 1998 Oct, 17(4), 343 - 50
Immunohistochemical detection of Listeria antigens in the placenta in perinatal listeriosis; Parkash V et al.; Listeria monocytogenes, a worldwide pathogen, causes significant perinatal mortality and morbidity and has been implicated in spontaneous abortions, still-births, premature delivery, and neonatal sepsis, often with meningitis . Maternal symptoms are frequently minimal, and diagnosis is made only if the suspicion is high and diagnostic maternal blood or amniotic fluid cultures are performed . Because cultures are not routinely performed on spontaneously aborted fetuses, many authors feel that the true incidence of the disease may be underestimated . To date, the absence of a test to retrospectively diagnose Listeria infection has contributed to the lack of accurate estimates of the incidence of the disease . Seven cases in which immunohistochemical stains were used to confirm the diagnosis of placental listeriosis are described . All placentas showed the characteristic lesions with severe chorioamnionitis, numerous microabscesses, and focal necrotizing villitis . Immunohistochemical localization of Listeria antigen was made to the amnion (focally in areas with no inflammatory infiltrate), the abscesses, and the areas with villitis . In general, the antigen was extracellular and intracellular, predominantly within macrophages or the amnion epithelium . Listeria antigen was often found where definite identification of the organism was not possible on Brown-Hopps or Warthin-Starry stains . The immunohistochemical technique may therefore show an increase in sensitivity of detection of L monocytogenes compared with routine bacterial stains . Moreover, the ability to retrospectively evaluate placental specimens for evidence of this organism should permit the true incidence of perinatal listeriosis to be determined.

Infect Immun, 1998 Nov, 66(11), 5260 - 7
Interaction of Listeria monocytogenes with human brain microvascular endothelial cells: InlB-dependent invasion, long-term intracellular growth, and spread from macrophages to endothelial cells; Greiffenberg L et al.; Invasion of endothelial tissues may be crucial in a Listeria monocytogenes infection leading to meningitis and/or encephalitis . Internalization of L . monocytogenes into endothelial cells has been previously demonstrated by using human umbilical vein endothelial cells as a model system . However, during the crossing of the blood-brain barrier, L . monocytogenes most likely encounters brain microvascular endothelial cells which are strikingly different from macrovascular or umbilical vein endothelial cells . In the present study human brain microvascular endothelial cells (HBMEC) were used to study the interaction of L . monocytogenes with endothelial cells, which closely resemble native microvascular endothelial cells of the brain . We show that L . monocytogenes invades HBMEC in an InlB-dependent and wortmannin-insensitive manner . Once within the HBMEC, L . monocytogenes replicates efficiently over a period of at least 18 h, moves intracellularly by inducing actin tail formation, and spreads from cell to cell . Using a green fluorescent protein-expressing L . monocytogenes strain, we present direct evidence that HBMEC are highly resistant to damage by intracellularly growing L . monocytogenes . Infection of HBMEC with L . monocytogenes results in foci of heavily infected, but largely undamaged endothelial cells . Heterologous plaque assays with L . monocytogenes-infected P388D1 macrophages as vectors demonstrate efficient spreading of L . monocytogenes into HBMEC, fibroblasts, hepatocytes, and epithelial cells, and this phenomenon is independent of the inlC gene product.

Zh Mikrobiol Epidemiol Immunobiol, 1998 Jul-Aug, (4), 33 - 6
{Intrauterine zoonotic infections in Dagestan}; Sultanov GV et al.; As the result of the prospective examination of 863 pregnant women in urban and rural consultation clinics for women in Daghestan, a high proportion of them were found to be infected with toxoplasmosis (25.5%), brucellosis (1.85%) and listeriosis (12.2%) . The data on the contamination of 1325 women with aggravated obstetric history were confidently higher, constituting 52.0%, 3.3% and 22.2% respectively . The results of the examination of women working on live-stock farms (226 women) and poultry farms (106 women) demonstrated a significantly high frequency of contamination with the above-mentioned zoonotic infections . The data thus obtained were indicative of the necessity of organizing epidemiological surveillance on these infections; for their diagnostics a complex of laboratory methods could be used, though the effectiveness of these methods was different in different nosological forms.

Zh Mikrobiol Epidemiol Immunobiol, 1998 Jul-Aug, (4), 23 - 5
{The persistent characteristics of Listeria monocytogenes isolated from different sources}; Boiko OV; A number of biological properties of L . monocytogenes was studied . Significant differences in the lysozyme, "anti-interferon", RNAase and lipase activity between strains isolated from different warm-blooded animals and from environmental objects were shown . 75% of all strains under study were found to have "anti-interferon" activity, while no antilysozyme activity was detected.

J Immunol, 1998 Oct 15, 161(8), 4146 - 52
Heat-killed Listeria monocytogenes as an adjuvant converts established murine Th2-dominated immune responses into Th1-dominated responses; Yeung VP et al.; We investigated the capacity of heat-killed Listeria monocytogenes (HKL), a potent stimulator of the innate immune system, as a vaccine adjuvant to modify both primary and secondary Ag-specific immune responses . Mice immunized with the Ag keyhole limpet hemocyanin (KLH) mixed with HKL generated a KLH-specific primary response characterized by production of Th1 cytokines and large quantities of KLH-specific IgG2a Ab . Moreover, administration of KLH with HKL as an adjuvant reversed established immune responses dominated by the production of Th2 cytokines and high levels of KLH-specific IgE and induced a Th1-type response with high levels of IFN-gamma and IgG2a and low levels of IgE and IL-4 . Neutralization of IL-12 activity at the time of HKL administration blocked the enhancement of IFN-gamma and reduction of IL-4 production, indicating that IL-12, induced by HKL, was responsible for the adjuvant effects on cytokine production . These results suggest that HKL as an adjuvant during immunization can successfully bias the development of Ag-specific cytokine synthesis toward Th1 cytokine production even in the setting of an ongoing Th2-dominated response . Thus, HKL may be clinically effective in vaccine therapies for diseases such as allergy and asthma, which require the conversion of Th2-dominated immune responses into Th1-dominated responses.

J Immunol Methods, 1998 Aug 1, 217(1-2), 27 - 39
Bone marrow cellular composition in Listeria monocytogenes infected mice detected using ER-MP12 and ER-MP20 antibodies: a flow cytometric alternative to differential counting; de Bruijn MF et al.; Detailed assessment of bone marrow cellular composition is essential in the evaluation of various experimental in vivo systems, such as expression of transgenes, null mutations and stimulation of host defence in infection . Traditional morphological analysis of mouse bone marrow is laborious, requires specific cytological expertise, and is somewhat subjective . As an alternative, we have examined whether double labelling of bone marrow with the anti-precursor monoclonal antibodies ER-MP12 and ER-MP20 could be used for differential analysis by flow cytometry, as these antibodies define six relatively homogeneous cell populations in mouse bone marrow . Following a sublethal infection of mice with Listeria monocytogenes, we monitored changes in cellular composition of the bone marrow at various time points in three ways: differential morphological count; single-color flow cytometric analysis using markers for the myeloid, erythroid and lymphoid lineages; and double labelling with ER-MP12 and ER-MP20 . As expected, the bone marrow composition changed dramatically during infection, leading to an increase of myeloid cells which peaked after 1 week of infection . Data determined by ER-MP12/20 flow cytometric analysis appeared to be in close agreement with both morphology and lineage marker analysis . In addition, ER-MP12/20 analysis provided more detailed information with regards to the presence of early myeloid precursors compared to lineage marker analysis . These data show that flow cytometric analysis of bone marrow using ER-MP12 and ER-MP20 monoclonal antibodies provides a relatively simple, rapid and objective assay when evaluating cellular composition in the bone marrow of the mouse.

Medicine (Baltimore), 1998 Sep, 77(5), 313 - 36
Central nervous system infection with Listeria monocytogenes . 33 years' experience at a general hospital and review of 776 episodes from the literature; Mylonakis E et al.; We reviewed 776 previously reported and 44 new cases of CNS listeriosis outside of pregnancy and the neonatal period, and evaluated the epidemiologic, diagnostic, and therapeutic characteristics of this infection . Among patients with Listeria meningitis/meningoencephalitis, hematologic malignancy and kidney transplantation were the leading predisposing factors, but 36% of patients had no underlying diseases recognized . The infection occurred throughout life, with a higher incidence before the age of 3 and after the age of 45-50 years . Fever, altered sensorium, and headache were the most common symptoms, but 42% of patients had no meningeal signs on admission . Compared with patients with acute meningitis due to other bacterial pathogens, patients with Listeria infection had a significantly lower incidence of meningeal signs, and the CSF profile was significantly less likely to have a high WBC count or a high protein concentration . Gram stain of CSF was negative in two-thirds of cases of CNS listeriosis . One-third of patients had focal neurologic findings, and approximately one-fourth developed seizures over their course . Mortality was 26% overall, and was higher among patients with seizures and those older than 65 years of age . Relapse occurred in 7% of episodes . Ampicillin for a minimum of 15-21 days (with an aminoglycoside for at least the first 7-10 days) remains the treatment of choice . Cerebritis/abscess due to L . monocytogenes, without meningeal involvement, is less common but may be diagnosed by blood cultures and CNS imaging, or by stereotactic biopsy . Longer antibiotic therapy (at least 5-6 weeks) is needed in the presence of localized CNS involvement.

Toxicology, 1998 Aug 21, 129(2-3), 201 - 10
Effects of prolonged exposure to morphine and methadone on in vivo parameters of immune function in rats; De Waal EJ et al.; In rats, two 6-week repeated dose oral toxicity studies were performed with morphine (250 and 500 mg/kg food) and methadone (200 and 400 mg/kg food), respectively . Alterations in immune function were studied by assessing primary and secondary immune responses to sheep red blood cells . In addition, the ability to resist challenge with infectious agents was measured in host resistance models employing the parasite Trichinella spiralis and the bacterium Listeria monocytogenes . The primary and secondary antibody responses to sheep red blood cells were not affected by treatment with either morphine or methadone . The clearance of L . monocytogenes bacteria in the spleen was not affected either . Prolonged treatment with morphine, however, resulted in a decrease in host resistance to T . spiralis infection, as indicated by a 1.5-fold increase in numbers of muscle larvae counted in the carcass, but did not affect the T . spiralis-specific IgM, IgG and IgE antibody responses . In contrast to morphine, the methadone-treated animals did not show a significant change in host resistance to T . spiralis . Total serum IgG levels, however, were increased in high-dose methadone-treated animals . Apparently, prolonged administration of morphine to rats resulted in immune suppression, mediating a slight, though biologically relevant, exacerbation of the T . spiralis infection, whereas methadone did not.

Am J Gastroenterol, 1998 Oct, 93(10), 1942 - 4
Listeria infection after liver transplantation: report of a case and review of the literature; Limaye AP et al.; Listeria monocytogenes is a well-recognized cause of bacteremia and meningitis in immunocompromised individuals, including recipients of solid organ transplants, but has only rarely been reported following orthotopic liver transplantation (OLT) . Most previously reported cases of listeriosis occurred months to years following liver transplantation; we describe a case of listeriosis that occurred within 1 wk of liver transplantation, shortly after discontinuation of trimethoprim-sulfamethoxazole prophylaxis, and review the English literature on Listeria infection after OLT . The patient developed abdominal pain and fever that suggested a bile leak, but was definitively diagnosed with Listeria infection by blood culture . The infection was successfully treated with 3 wk of intravenous ampicillin . We conclude that serious systemic infection with Listeria monocytogenes is uncommon following OLT, may occur early in the postoperative period, and responds well to treatment with high dose ampicillin.

J Food Prot, 1998 Sep, 61(9), 1203 - 6
Inactivation of Listeria monocytogenes in milk by pulsed electric field; Reina LD et al.; Pasteurized whole, 2%, and skim milk were inoculated with Listeria monocytogenes Scott A and treated with high-voltage pulsed electric field (PEF) . The effects of milk composition (fat content) and PEF parameters (electric field strength, treatment time, and treatment temperature) on the inactivation of the bacterium were studied . No significant differences were observed in the inactivation of L . monocytogenes Scott A in three types of milk by PEF treatment . With treatment at 25 degrees C, 1- to 3-log reductions of L . monocytogenes were observed . PEF lethal effect was a function of field strength and treatment time . Higher field strength or longer treatment time resulted in a greater reduction of viable cells . A 4-log reduction of the bacterium was obtained by increasing the treatment temperature to 50 degrees C . Results indicate that the use of a high-voltage PEF is a promising technology for inactivation of foodborne pathogens.

J Food Prot, 1998 Sep, 61(9), 1195 - 8
Reactivities of genus-specific monoclonal antibody EM-6E11 against Listeria species and serotypes of Listeria monocytogenes grown in nonselective and selective enrichment broth media; Nannapaneni R et al.; Depending on the growth medium used for enrichment of bacterial cells prior to assay, the monoclonal antibody (MAb) EM-6E11 recognizing Listeria genus-specific epitope on 43 and 94 to 97 kDa cell-surface antigens (A . K . Bhunia and M . G . Johnson, Appl . Environ . Microbiol . 58:1924-1929, 1992) exhibited extensive variability in the detection of Listeria species . MAb EM-6E11 strongly detected live cells of all Listeria species and all serotypes of L . monocytogenes by ELISA when cells were grown in nonselective brain heart infusion (BHI) broth, in selective Listeria enrichment broth (LEB), or in Listeria repair broth (LRB) . In contrast, EM-6E11 detected only four of the thirteen serotypes of L . monocytogenes (serotypes 1/2c, 3b, 4ab, and 7) when cells were grown in the UVM1 formulation of Listeria enrichment broth (UVM1) or Fraser broth (FRB) . This MAb failed to react with live cells of four other Listeria species, including L . ivanovii, L . welshimeri, L . grayi, and L . murrayi cells grown in UVM1 or FRB . Heating of Listeria cells at 100 degrees C for 20 min, irrespective of the enrichment media used, led to large losses of MAb EM-6E11 reactivity in ELISA, suggesting that the specific cell-surface epitopes involved may not be heat stable . Our results confirm that MAb EM-6E11 is suitable for detection of live cells but not heat-killed cells of Listeria spp . and can be used in conjunction with an enrichment step in BHI, LEB, or LRB but not in UVM1 or FRB.

Res Microbiol, 1997 Jul-Aug, 148(6), 485 - 90
A nested PCR method to detect Listeria monocytogenes in artificially contaminated blood specimens; Cocolin L et al.; A nested PCR-based test was developed for the detection of Listeria monocytogenes in blood specimens from patients with listeriosis . Two pairs of oligonucleotide primers were designed to amplify a 1395-bp and a 453-bp fragment of the iap gene of L . monocytogenes . Amplified products were analysed with gel electrophoresis and stained with ethidium bromide . The PCR method described could be routinely used to diagnose listeriosis.

Res Microbiol, 1997 May, 148(4), 305 - 13
Effect of saline concentration, pH and growth temperature on the invasive capacity of Listeria monocytogenes; Galdiero E et al.; The invasive ability of Listeria monocytogenes was monitored after treatment at different pH, temperature and salt concentrations . We found a complete loss of invasive ability in bacteria grown at pH < or = 4.5 independently of the incubation temperature (4, 22 and 30 degrees C) . Increasing salt concentrations at 22 and 30 degrees C had no effect at pH 7, while drastically affecting invasive ability at pH 5 . The expression of two proteins of 30 and 88 kDa, extracted from the culture supernatant and the cell wall, respectively, was detected only in cells grown under normal conditions, but not after low pH and high salt concentration treatment.

Acta Crystallogr D Biol Crystallogr, 1998 Jan 1, 54 ( Pt 1), 108 - 10
Crystallization and preliminary X-ray analysis of human platelet profilin complexed with an oligo proline peptide; Mahoney NM et al.; Profilin is an actin-monomer binding protein that regulates the distribution and dynamics of the actin cytoskeleton . Profilin binds poly-L-proline and proline-rich peptides in vitro and co-localizes with proline-rich proteins in focal adhesions and at the site of actin tail assembly on the surface of intracellular parasites such as Listeria monocytogenes . The crystallization of the complex between human platelet profilin (HPP) and an L-proline decamer {(Pro)10} is reported here . Diffraction from these crystals is consistent with the space group P21212 with unit-cell constants a = 68.25, b = 97.64, c = 39.10 A . The crystals contain two HPP molecules per asymmetric unit and diffract to 2.2 A.

Int J Food Microbiol, 1998 Aug 18, 43(1-2), 61 - 71
Comparison of five typing methods for the epidemiological study of Listeria monocytogenes; Kerouanton A et al.; Five typing methods were compared in a study designed to adapt a strategy for epidemiologically typing large numbers of Listeria monocytogenes strains . The methods studied were serotyping, electrophoretic typing of esterases (zymotyping), restriction fragment length polymorphism of ribosomal DNA (ribotyping), random amplified polymorphic DNA (RAPD) and pulsed-field gel electrophoresis (PFGE) . Data were analysed by computer-assisted statistical analysis . Included in the analysis were 35 strains of L . monocytogenes, including 14 epidemic strains isolated during outbreaks in France in 1992 and 1993, and 21 strains isolated from food and the environment . Five serotypes, eight zymotypes, ten ribotypes, 13 RAPD patterns and 12 PFGE patterns were identified among the 35 strains . The most discriminating combination of typing methods was ribotyping and PFGE typing {27 types, discriminatory index (D.I.) = 0.978} . A factorial analysis of correspondence for each method differentiated the epidemic strains from the environmental strains . This study shows that computer-assisted statistical treatment of the data, combined with the use of discriminating typing methods, is a powerful tool for the epidemiological analysis of Listeria monocytogenes.

Int J Food Microbiol, 1998 Aug 18, 43(1-2), 21 - 31
The combined affects of modified atmosphere, temperature, nisin and ALTA 2341 on the growth of Listeria monocytogenes; Szabo EA et al.; A cocktail of seven Listeria monocytogenes isolates of food, human and environmental origin was used to assess the antilisterial activity of the bacteriocins nisin and ALTA 2341 in combination with various atmospheres: air, 100% N2, 40% CO2:60% N2, or 100% CO2 . Buffered tryptone soya broth (pH 6.0) was used as the growth medium and incubation was at 4 degrees C (21 days) or 12 degrees C (7 days), or when temperature fluctuated between these values for defined periods . It was observed that atmosphere alone influenced the growth rate of L . monocytogenes, with 100% CO2 exerting the greatest inhibition . A 5 log population increase was observed in all atmospheres after 7 days at 12 degrees C . At 4 degrees C a 4-5 log population increase was observed in air, 100% N2 and 40% CO2:60% N2 within 21 days . Growth was prevented by 100% CO2 . In the presence of nisin (400 IU/ml), an increase in the lag phase was observed before growth (5 log population increase after 7 days) in all atmospheres at 12 degrees C . This effect was enhanced at 4 degrees C where a maximum 2 log population increase was observed in all atmospheres except 100% CO2, in which growth was prevented . Increasing the concentration of nisin to 1250 IU/ml prevented L . monocytogenes growth in all atmosphere combinations at 4 and 12 degrees C . Two concentrations of ALTA 2341 were also tested . In the presence of 0.1% ALTA 2341 and at 12 degrees C, a 3-5 log population increase was observed in all atmospheres with the exception of 100% CO2, which prevented L . monocytogenes growth . At 4 degrees C, growth was observed in the combination of 0.1% ALTA 2341 and 100% N2 only (3 log population increase) . Use of a higher concentration of ALTA 2341 (1.0%) resulted in a population decrease below the detection level within 24 h in all atmosphere/temperature combinations . Re-growth occurred in the presence of 1.0% ALTA 2341 in all atmospheres at 12 degrees C, and in combination with air or 100% N2 at 4 C . When the effectiveness of either nisin or ALTA 2341 and atmosphere was tested against L . monocytogenes as temperature fluctuated for periods between 4 and 12 degrees C, only the combination of 100% CO2 and 1.0% ALTA 2341 prevented growth . Cells surviving exposure to nisin or ALTA 2341 were recovered from 28 of the 32 combinations tested that contained bacteriocin . Nisin survivors remained sensitive to the bacteriocin . ALTA 2341 survivors had become resistant to the bacteriocin.

Int J Food Microbiol, 1998 Aug 18, 43(1-2), 15 - 9
Combined effect of nisin and high hydrostatic pressure on destruction of Listeria innocua and Escherichia coli in liquid whole egg; Ponce E et al.; High hydrostatic pressure inactivation of Escherichia coli and Listeria innocua inoculated in liquid whole egg was improved significantly (P < 0.05) with nisin addition at concentrations of 1.25 and 5 mg/1 . A reduction of almost 5 log10 units in E . coli counts and more than 6 log10 units for L . innocua was obtained at 450 MPa and 5 mg/l of nisin . For this treatment, the two microorganisms were not detectable after 1 month of storage at 4 degrees C . The amount of nisin added did not affect E . coli inactivation at 300 MPa . For L . innocua, 5 mg/l of nisin was more effective than 1.25 mg/l . Nisin showed no effect when samples were stored at 20 degrees C after pressurization, except for samples with L . innocua containing 5 mg/l of nisin and treated with 450 MPa.

Novartis Found Symp, 1998, 215, 159 - 67; discussion 167-71, 186-90
A two-step model for the induction of organ-specific autoimmunity; Limmer A et al.; Peripheral tolerance is considered to be a safeguard against autoimmunity but the mere existence of anergic T cells renders them potentially dangerous . Using transgenic mice that were tolerant to a foreign MHC class I antigen (Kb) exclusively expressed in the liver, we investigated whether reversal of tolerance in vivo would directly result in autoimmunity . Breaking of tolerance was achieved by application of tumour cells expressing both Kb and interleukin 2 . Despite the fact that the respective mice were now able to reject Kb-positive grafts, the reversed T cells did not infiltrate and attack the Kb-positive liver . However, when the liver was 'conditioned' through an inflammatory reaction either by irradiation or by infection with Listeria, massive T cell infiltration and liver damage were observed in the reversed mice . The results show that at least two steps are required for autoimmunity: (1) activation of antigen-specific T cells, and (2) conditioning of the target organ . It will be important to determine the factors leading to conditioning but it is likely that adhesion molecules are involved . These experiments are not only of relevance for treatment of autoimmune disease but also for tumour therapy.

Biotechnol Prog, 1998 Sep-Oct, 14(5), 782 - 90
Single- and dual-fractal analysis of hybridization binding kinetics: biosensor applications; Sadana A et al.; The diffusion-limited hybridization kinetics of analyte in solution to a receptor immobilized on a biosensor or immunosensor surface is analyzed within a fractal framework . The data may be analyzed by a single- or a dual-fractal analysis . This was indicated by the regression analysis provided by Sigmaplot . It is of interest to note that the binding rate coefficient and the fractal dimension both exhibit changes in the same direction for both the single-fractal and the dual-fractal analysis examples presented . For example, for a single-fractal analysis and for the hybridization of 10 nM 16CFl (oligonucleotide) to 16B immobilized via sulfosuccinimidyl-6-(biotinamido)hexanoate and streptavidin using chemical and thermal regeneration (Abel, A . P.; Weller, M . G.; Duveneck, G . L.; Ehrat, M . Widmer, H . M . Anal . Chem . 1996, 68, 2905-2912), an increase in the fractal dimension, Df from 1.211 (chemical regeneration) to 1.394 (thermal regeneration), leads to an increase in the binding rate coefficient, k, from 86.53 (chemical regeneration) to 100.0 (thermal regeneration) . An increase in the degree of heterogeneity on the biosensor surface leads to an increase in the binding rate coefficient . When a dual-fractal analysis was utilized, an increase in the fractal dimension value from Df1 to Df2 leads to an increase in the binding rate coefficient value from k1 to k2 . The fractional order of dependence of the binding rate coefficient, k1, on (a) the analyte (rRNA) concentration in solution and (b) on the fractal dimension, Df1, for the hybridization kinetics to detect Listeria species (Fliss, R.; St-Laurent, M.; Emond, E.; Simard, R . E.; Lemieux, R.; Ettriki, A.; Pandian, S . Appl . Microbiol . Biotechnol . 1995, 43, 717-724.) further reinforces the fractal nature of the system . The binding rate coefficient(s) expressions developed as a function of the analyte concentration in solution and the fractal dimension are of particular value since they provide a means to better control of biosensor or immunosensor performance.

Ann Intern Med, 1998 Oct 1, 129(7), 559 - 66
Infections in patients with chronic lymphocytic leukemia treated with fludarabine; Anaissie EJ et al.; BACKGROUND: Fludarabine, a purine analogue with activity in chronic lymphocytic leukemia, is usually well tolerated . Although serious infections after fludarabine therapy have been described, a systematic analysis of the risk factors for such infections in chronic lymphocytic leukemia is lacking . OBJECTIVE: To determine the risk factors for major infection in patients with chronic lymphocytic leukemia treated with fludarabine . DESIGN: Retrospective review of medical records . SETTING: Cancer center . PATIENTS: 402 patients with chronic lymphocytic leukemia not previously treated or treated with chlorambucil (with or without prednisone) who received fludarabine (30 mg/m2 of body surface area per day for 5 days) with or without prednisone at 4-week intervals . RESULTS: Infections occurred more often in previously treated (144 of 248 {58%}) than in previously untreated (53 of 154 {34%}) patients (P < 0.001) . Listeriosis or pneumocystosis occurred in 12 of 170 (7%) previously treated patients receiving fludarabine plus prednisone, 0 of 78 previously treated patients receiving fludarabine alone, and 2 of 154 (1%) previously untreated patients receiving fludarabine plus prednisone (P = 0.003) . Univariate analysis identified previous chemotherapy, advanced disease, failure to respond to fludarabine, elevated serum beta2-microglobulin level (P < 0.001), low serum albumin level (P = 0.024), elevated serum creatinine concentration (P = 0.008), and low granulocyte count (P = 0.003) as risk factors for infection . Multivariate analysis identified Rai stage III or IV (odds ratio, 1.98 {95% CI, 1.17 to 3.94}), previous treatment (odds ratio, 2.24 {CI, 1.43 to 3.51}), and elevated serum creatinine concentration (odds ratio, 1.98 {CI, 1.09 to 3.67}) as statistically significant independent risk factors for major infection . A baseline granulocyte count of more than 1000 cells/microL was protective (odds ratio, 0.54 {CI, 0.29 to 0.99}) . Five (26%) of 19 patients with a CD4 count less than 50 cells/mL developed cutaneous zoster compared with 9 (6%) of 139 patients with a CD4 count greater than 50 cells/mL (P = 0.01) . CONCLUSIONS: Fludarabine used in previously treated patients with chronic lymphocytic leukemia may be associated with infections involving T-cell dysfunction, such as listeriosis, pneumocystosis, mycobacterial infections, and opportunistic fungal and viral infections . Prophylaxis or presumptive therapy should be initiated in the appropriate setting.

Eur J Immunol, 1998 Sep, 28(9), 2630 - 9
Efficient induction of cytotoxic CD8+ T cells against exogenous proteins: establishment and characterization of a T cell line specific for the membrane protein ActA of Listeria monocytogenes; Bruder D et al.; The property of listeriolysin (LLO) to introduce soluble passenger proteins into the cytosol of antigen-presenting cells allows the induction of CD8+ cytotoxic T cells against such antigens . To overcome the potential problem of presentation of the immunodominant epitope LL091-99 by H-2Kd, a variant LLO92A was established in which Tyr 92 was replaced by Ala . Immunization of BALB/c mice with purified LLO92A failed to stimulate cytotoxic T cells specific for either the epitope LLO91-99 or for any other LLO-derived peptide . Injection of mixtures of purified LLO92A and soluble nucleoprotein (NP) of influenza virus into mice resulted in a strong cytotoxic T cell response exclusively directed against NP . The LLO92A variant was successfully used to generate, propagate and characterize a CD8 T cell line specific for the membrane-bound virulence factor ActA of Listeria monocytogenes . Interestingly, wildtype ActA bound to the surface of live L . monocytogenes was not presented by MHC class I molecules to the CD8+ T cell line.

Immunopharmacology, 1998 Jun, 39(3), 215 - 23
Effect of a traditional Chinese medicine, Bu-zhong-yi-qi-tang on the protection against an oral infection with Listeria monocytogenes; Yamaoka Y et al.; The protective effect against an oral infection with Listeria monocytogenes was observed in BALB/c mice who were orally administered a traditional Chinese medicine, Bu-zhong-yi-qi-tang (Japanese name: Hochu-ekki-to, HOT) daily for 7 days . Bacterial numbers in the Peyer's patch (PP) at 18 h, in the mesenteric lymph nodes (MLN) at 18 h, 1 day and 3 days and in the liver at 3 days after infection were significantly suppressed in HOT-treated mice, although there was no difference in the bacterial number in the small intestinal contents . The enhanced bactericidal activities of PP and liver macrophages by pretreatments of HOT were observed . The protective effect of HOT was not observed in athymic nu/nu and IFN-gamma deficient mice . The administration of HOT increased IFN-gamma-producing cells in the intestinal intraepithelial lymphocytes (IEL) but did not in the PP, MLN and liver . HOT exerts effects mainly on CD8alphabeta+ IEL which are thymus-dependent, and induced IFN-gamma production from their cells . These results suggest that HOT acts on the gut-associated lymphoid tissues and induces IFN-gamma from CD8alphabeta+ IEL, which activates PP and liver macrophages and consequently the resistance to L . monocytogenes is augmented in the mice.

Arch Latinoam Nutr, 1998 Mar, 48(1), 68 - 70
{Presence of total coliforms, Escherichia coli and Listeria sp . in enteral formulas}; Arias ML et al.; The presence of total coliforms, Escherichia coli and Listeria sp . was evaluated in 65 samples of enteral nutrition formulas . In more than the 75% of the samples made up from cooked vegetables, fruits or meat broth, the score level of total coliforms was of 10(4) UFC/g . In 12-31% of the different enteral food formula, E . coli was isolated in levels ranging form 3.0 x 10(2) to 2.1 x 10(4) UFC/g . Enteral nutrition formulas made out of fruits and those elaborated with meal broth presented this agent more frequently and in bigger quantities . Listeria sp . was isolated in 17% of the fruit preparations and in enteral formulas made with milk . L . grayi, L welshimeri and L . innocua were the species found.

Lett Appl Microbiol, 1998 Aug, 27(2), 67 - 70
Light inactivation of food-related pathogenic bacteria using a pulsed power source; MacGregor SJ et al.; The effects of high intensity light emissions, produced by a novel pulsed power energization technique (PPET), on the survival of bacterial populations of verocytotoxigenic Escherichia coli (serotype 0157:H7) and Listeria monocytogenes (serotype 4b) were investigated . Using this PPET approach, many megawatts (MW) of peak electrical power were dissipated in the light source in an extremely short energization time (about 1 microsecond) . The light source was subjected to electric field levels greater than could be achieved under conventional continuous operation, which led to a greater production of the shorter bacteriocidal wavelengths of light . In the exposure experiments, pre-determined bacterial populations were spread onto the surface of Trypone Soya Yeast Extract Agar and were then treated to a series of light pulses (spectral range of 200-530 nm) with an exposure time ranging from 1 to 512 microseconds . While results showed that as few as 64 light pulses of 1 microsecond duration were required to reduce E . coli 0157:H7 populations by 99.9% and Listeria populations by 99%, the greater the number of light pulses the larger the reduction in cell numbers (P < 0.01) . Cell populations of E . coli 0157:H7 and Listeria were reduced by as much as 6 and 7 log10 orders at the upper exposure level of 512 microseconds, respectively . Survival data revealed that E . coli 0157:H7 was less resistant to the lethal effects of radiation (P < 0.01) . These studies have shown that pulsed light emissions can significantly reduce populations of E . coli 0157:H7 and L . monocytogenes on exposed surfaces with exposure times which are 4-6 orders of magnitude lower than those required using continuous u.v . light sources.

J Appl Microbiol, 1998 Aug, 85(2), 337 - 46
Tissue culture assays using Caco-2 cell line differentiate virulent from non-virulent Listeria monocytogenes strains; Van Langendonck N et al.; Within the group of Listeria sp., only L . monocytogenes is pathogenic for humans and numerous studies of L . monocytogenes strains have described non-virulent isolates . In this study, the potential value of two tissue culture assays (TCA) was analysed to ascertain the virulence properties of L . monocytogenes strains, initially typed for virulence using the immunocompromised mouse model (ICMM) . The first assay assessed both the penetration into, and multiplication within, Caco-2 cells (PM assay): the second was a plaque-forming assay (PF assay) . All the clinical isolates (nine strains) were virulent in both TCA . Conversely, all the non-pathogenic species (seven strains) were non-virulent in PM and PF assays . Compared with the virulence obtained in the ICMM with 29 Listeria strains, including 12 non-virulent L . monocytogenes strains, the sensitivity of both TCA was equal to1 . Specificity was 0.89 and 0.84 for the PF and PM assays, respectively . However, a study of strains exhibiting virulence differences in three other in vivo virulence models showed that ICMM only detected highly virulent strains . The specificity of the PF test could, therefore, be higher, and close to that obtained by the enumeration of viable bacteria in the spleen of mice infected by subcutaneous injection in the footpad and by intravenous injection . Taken together, this study confirms the existence of low-virulence L . monocytogenes strains and shows that the virulence status of some non-clinical L . monocytogenes isolates depends on the virulence models used . The data suggest that the PF assay could be used as a primary test to evaluate the virulence of Listeria strains in order to reduce the cost of testing all strains in vivo.

J Appl Microbiol, 1998 Aug, 85(2), 287 - 92
Bacteriocins inhibit glucose PEP:PTS activity in Listeria monocytogenes by induced efflux of intracellular metabolites; Waite BL et al.; Glucose transport by the phosphoenolpyruvate (PEP)-dependent phosphotransferase system (PTS) of listeria monocytogenes is inhibited by the bacteriocins, nisin, pediocin JD and leuconocin S . To investigate the mechanism of inhibition, PTS activity assays were performed with permeabilized, bacteriocin-treated L . monocytogens Scott A cells . In the presence of exogenous PEP, nisin stimulated the PTS while both pediocin JD and leuconocin S partially inhibited its activity . These results suggested that PTS enzymes were still active in bacteriocin-treated cells and the bacteriocin-induced PEP efflux may be a mechanism for inhibition of the PTS . To verify that PEP did efflux from bacteriocin-treated L . monocytogens Scott A cells, intracellular and extracellular PEP were measured by HPLC . All three bacteriocins induced efflux of PEP . Nisin, pediocin JD and leuconocin S also induced efflux of AMP, ADP and ATP . These studies indicate that bacteriocin inhibition of the glucose PEP:PTS in L . monocytogenes is due to efflux of intracellular metabolites, particularly

J Appl Microbiol, 1998 Sep, 85(3), 545 - 53
Survival of Listeria monocytogenes in sea water and effect of exposure on thermal resistance; Bremer PJ et al.; Survival, recoverability and sublethal injury of two strains of Listeria monocytogenes, Scott A and an environmental strain KM, on exposure to sea water at 12.8 or 20.8 degrees C was determined using in situ diffusion chambers . Plate counts were used to assess recoverability and injury while 5-cyano-2,3-ditolyl tetrazolium chloride (CTC) reduction was used to determine respiratory activity . T90 values (times for 10-fold decreases in numbers of recoverable cells) on non-selective medium (trypticase soya agar with 0.6% yeast extract) at 12.8 and 20.8 degrees C were 61.7 and 69.2 h for L . monocytogenes Scott A, and 103.0 and 67.0 h for L . monocytogenes KM, respectively . On selective medium (Oxford agar), T90 values at 12.8 and 20.8 degrees C were 60.6 and 56.9 h for L . monocytogenes Scott A, and 83.0 and 65.9 h for L . monocytogenes KM, respectively . With Scott A, the percentage of sublethally injured cells at 12.8 and 20.8 degrees C was 1.7 and 17.7%, respectively, while for KM the values were 19.0 and 1.6%, respectively . The fraction of cells reducing CTC but which were not recoverable on plating progressively increased on exposure to sea water . Listeria monocytogenes KM challenged at 58 degrees C showed an apparent increase in heat resistance after exposure to sea water at 20.8 degrees C for 7 d (D58 = 2.64 min) compared with before exposure (D58 = 1.24) . This increase in thermal resistance was not apparent at temperatures greater than 63 degrees C, and analysis of the best-fit regression lines fitted to the thermal data obtained from the two cell populations indicated that their thermal resistance was not significantly different (P > 0.05) over the temperature range tested (58-62 degrees C).

J Appl Microbiol, 1998 Sep, 85(3), 521 - 6
Antilisterial activity of enterocin 81, a bacteriocin produced by Enterococcus faecium WHE 81 isolated from cheese; Ennahar S et al.; Enterocin 81, a bacteriocin produced by Enterococcus faecium WHE 81 previously isolated from cheese, exhibited a very narrow spectrum of activity, which is mainly directed against enterococci and Listeria spp . including Listeria monocytogenes . Enterocin 81 activity, which was extremely rapid with maximal effect achieved within 30 min, could not be detected after treatment with various proteolytic enzymes . This activity was bactericidal in nature and induced an important efflux of intracellular material, which was visualized under electron microscopy as filaments coming out of L . monocytogenes cells . However, enterocin 81 did not display bacterial lysis on sensitive cells, as no changes in cell morphology were detected following the bactericidal action . Furthermore, this bacteriocin was shown to be equally active at pH values ranging from 4.0 to 8.0, which, along with the narrow activity spectrum, are two factors of paramount interest with regards to possible use of this bacteriocin in fermented foods.

East Afr Med J, 1998 Apr, 75(4), 249 - 51
Listeria septicaemia and meningitis in a neonate: case report; Ashiru JO et al.; We report the first case of neonatal septicaemia and meningitis in Trinidad due to Listeria three days after blood transfusion . It is important to be aware of the organism in foods and patients . Modern methods of isolation and identification will be of invaluable assistance in future recognition of the organism.

J Immunol, 1998 Sep 15, 161(6), 3010 - 8
Two distinct phospholipases C of Listeria monocytogenes induce ceramide generation, nuclear factor-kappa B activation, and E-selectin expression in human endothelial cells; Schwarzer N et al.; Infection of endothelial cells by Listeria monocytogenes is an essential step in the pathogenesis of listeriosis . We recently reported that L . monocytogenes induces up-regulation of E-selectin and other endothelial adhesion molecules and subsequent polymorphonuclear leukocyte (PMN) adhesion into cultured human endothelial cells . In the present study, we characterized the mechanisms of enhanced E-selectin expression using L . monocytogenes wild type (EGD), the isogenic in-frame deletion mutants for phosphatidylcholine (PC)- and phosphatidylinositol (PI)-specific phospholipases EGD delta plcA and EGD delta plcB, as well as the nonvirulent control strain Listeria innocua . Infection of endothelial cells with EGD delta plcA or EGD delta plcB for 6 h induced, as compared with EGD wild type, intermediate levels of E-selectin mRNA and protein as well as PMN rolling and adhesion at a shear rate of 1 dyne/cm2, indicating that both bacterial phospholipases are required for a maximal effect . Similarly, ceramide content and NF-kappa B activity were increased in L . monocytogenes-exposed endothelial cells, but only to intermediate levels for PC- or PI-phospholipase C (PLC)-deficient listerial mutants . Phospholipase effects could be mimicked by exogenously added ceramides or bacterial sphingomyelinase . The data presented indicate that PI-PLC and PC-PLC are important virulence factors for L . monocytogenes infections that induce accumulation of ceramides that in turn may act as second messengers to control host cell signal-transduction pathways leading to persistent NF-kappa B activation, increased E-selectin expression, and enhanced PMN rolling/adhesion . The ability of L . monocytogenes to stimulate PMN adhesion to endothelial cells may be an important mechanism in the pathogenesis of severe listeriosis.

J Immunol, 1998 Sep 15, 161(6), 2985 - 93
The MHC class I-restricted immune response to HIV-gag in BALB/c mice selects a single epitope that does not have a predictable MHC-binding motif and binds to Kd through interactions between a glutamine at P3 and pocket D; Mata M et al.; Using a strain of Listeria monocytogenes that stably expresses and secretes HIV gag to deliver this Ag to the MHC class I pathway of Ag processing, we have identified the immunodominant CTL epitope to gag in the BALB/c mouse and shown that it is Kd restricted . The specific motif for the peptides that bind the MHC class I molecule H-2 Kd is believed to be a nonamer with residues tyrosine or phenylalanine in the second amino acid position and leucine or isoleucine in the carboxyl-terminal or ninth amino acid position as dominant anchoring positions . Surprisingly, the identified gag peptide, AMQMLKETI, does not contain an anchoring aromatic residue in position two although competition assays with other Kd-restricted epitopes indicated that it binds to Kd with comparable affinity . Using a theoretical molecular dynamics approach to probe the stability of peptide binding to MHC class I molecules, we show that the absence of an appropriate anchor residue at P2 in AMQMLKETI is compensated by favorable interactions of the glutamine at P3 with pocket D of Kd . These findings were verified experimentally, demonstrating the predictive power of this theoretical approach in analyzing MHC class I/peptide interactions . These studies also indicate that CTL epitope prediction that relies on dominant peptide motifs may not always identify the correct epitope.

J Lipid Res, 1998 Sep, 39(9), 1740 - 3
Apolipoprotein E-deficient mice have impaired innate immune responses to Listeria monocytogenes in vivo; Roselaar SE et al.; Apolipoprotein E (apoE) influences both innate and acquired immunity in cultured cells . To determine whether apoE affects the immune system in vivo, Listeria monocytogenes (LM) was administered intraperitoneally (10(4) c.f.u.) to congenic C57BL/6 apoE-/- and +/+ mice (n = 12 in each group) . Survival was assessed daily for 5 days . Deficiency of apoE significantly increased death by day 5 (P = 0.03) . The majority of deaths occurred at day 4 . Extent of infection after LM administration was assessed at day 3 by determining colony counts in hepatic and splenic extracts . ApoE+/+ mice had very low colony counts in both spleen and liver {mean +/- SE: 2.0 +/- 0.5 and 0.7 +/- 0.2 (x 10(4)), respectively, n = 8 in each group}; while apoE-/- mice had significantly increased counts in both spleen and liver {64 +/- 51 and 98 +/- 93 (x 10(4)), P = 0.05 and 0.03} . Serum concentrations of TNF-alpha were significantly increased in apoE-/- mice at day 3 compared to apoE+/+ mice (127 +/- 43 pg/ml versus 20 +/- 17, P = 0.003) . LM induced more hepatic damage in apoE-/- mice compared to apoE+/+ mice as judged by increased serum concentrations of alanine aminotransferase at day 1 (apoE-/- 301 +/- 45 U/ml, apoE+/+ 101 +/- 9 U/ml, P = 0.01) . The increased proliferation and mortality from LM in apoE-/- mice occurred prior to the initiation of acquired immune responses . Therefore, apoE-deficient mice have an impaired innate response to infection by LM.

Curr Biol, 1998 Sep 10, 8(18), R654 - 7
Actin cytoskeleton: the Arp2/3 complex gets to the point; Zigmond SH; Actin filament polymerization results primarily from the addition of monomers to pre-existing filaments . Recent studies have revealed that the Arp2/3 protein complex, which includes two actin-related proteins, can nucleate new actin filaments, and this capacity can be enhanced by ActA, a protein used by Listeria to polymerize actin.

Am J Vet Res, 1998 Sep, 59(9), 1125 - 8
Characterization of Listeria monocytogenes isolated from channel catfish (Ictalurus punctatus); Wang C et al.; OBJECTIVE: To characterize Listeria monocytogenes from tissues of channel catfish for their ability to cause hemolysis and grow intracellularly in mouse macrophages . SAMPLES: 15 isolates from processed fillets and 15 isolates from the brain, spleen, and kidneys . PROCEDURE: Serotype and hemolytic activity of L monocytogenes isolates were evaluated, using plate agglutination and CAMP tests, respectively . Invasiveness of L monocytogenes was determined by inoculating each strain or isolate on J774A.1 macrophage cells . Infected cells were incubated for 0 or 3 hours and lysed; then 100 gli of the lysate was plated onto a brain heart infusion agar plate . Colony counts for each strain or isolate were analyzed statistically . RESULTS: Of 30 isolates, 19 were serotype 1 and 11 were serotype 4 . Mouse J774A.1 macrophages were inoculated with catfish isolates, a wild-type (EGD) or a nonhemolytic strain of L monocytogenes . Seventy-three percent (11/15) of isolates originating from catfish organs and 100% (15/15) of isolates originating from fillets were not significantly different from the wild-type EGD strain . The nonhemolytic L monocytogenes strain used as a negative control failed to replicate . Intracellular growth of all L monocytogenes isolates decreased after an additional 3-hour incubation period with medium containing 50 {microg/ml of gentamicin . CONCLUSIONS: Similar to the wild-type EGD strain, most channel catfish L monocytogenes isolates were hemolytic, serotype 1 or 4, and were invasive for mouse J774A.1 macrophages . CLINICAL RELEVANCE: monocytogenes growth in mouse macrophages may serve as an in vitro model for determining virulence of isolates from food products or environments.

Am J Gastroenterol, 1998 Sep, 93(9), 1556 - 8
Spontaneous bacterial peritonitis caused by infection with Listeria monocytogenes: a case report and review of the literature; Jayaraj K et al.; Spontaneous bacterial peritonitis is a frequent and often serious complication of long-standing ascites in the presence of advanced liver disease . Coliform bacteria account for the infection in most cases and are thought to be related to translocation of bacteria from the bowel into the peritoneal cavity . The empiric use of cefotaxime is well established as most of the causative organisms are sensitive to this antibiotic . However, we report on a case of spontaneous bacterial peritonitis in a patient with hepatitis C related cirrhosis who was awaiting liver transplantation caused by infection with Listeria monocytogenes, in which the patient did not improve with empiric antibiotic therapy . This case adds to the 23 others reported in the literature since 1966 . Our case raises some concerns about the universal empiric usage of cefotaxime for spontaneous bacterial peritonitis because it does not offer adequate coverage against organisms such as Listeria, enterococci, Pasturella, and anaerobes.

Cell Immunol, 1998 Aug 1, 187(2), 88 - 94
Host resistance against Listeria monocytogenes is reciprocal during the course of infection in alymphoplastic aly mutant mice; Nishikawa S et al.; The aly is a unique spontaneous autosomal recessive mutation in mice that causes a systemic defect of lymph nodes and Peyer's patches . We investigated host resistance against Listeria monocytogenes infection in the mutant . The 50% lethal dose of L . monocytogenes in aly/aly mice was 10-fold higher than their heterozygotes, termed aly/+mice, or their wild-type C57BL/6 mice . The bacterial growth in the spleens and livers of aly/aly mice was more efficient early in infection, and their listericidal activity of peritoneal macrophages was higher than those of aly/+mice . In contrast, the complete elimination of bacteria from the spleens and livers of infected mice in the late stage of infection, in which a T-cell-dependent mechanism is required, was delayed in the aly/aly mice . Moreover, an acquired resistance against secondary infection with L . monocytogenes was markedly diminished in the aly/aly mice . The production of endogenous interferon-gamma (IFN-gamma) and tumor necrosis factor-alpha (TNF-alpha), which are critical in antilisterial resistance, was reduced in the aly/aly mice during the infection . The production of IFN-gamma, and interleukin-4 was also diminished in the spleen cell cultures of aly/aly mice when stimulated with heat-killed L . monocytogenes or the T-cell receptors were directly stimulated with anti-CD3-epsilon monoclonal antibody . These results suggest that acquired immunity against L . monocytogenes infection is attenuated in aly/aly mice, and that the insufficient production of IFN-gamma and TNF-alpha might be involved in the immunodeficiency.

J Cell Sci, 1998 Oct, 111 ( Pt 19), 2877 - 84
Identification of cofilin, coronin, Rac and capZ in actin tails using a Listeria affinity approach; David V et al.; Actin assembly is involved in cell motility and intracellular movement of Listeria monocytogenes . Induction of Listeria actin tails is mediated by the surface protein ActA . The N-terminal domain of ActA is sufficient for this function . Cell components known to play a role in the actin-based motility of Listeria are VASP (vasodilatator-stimulated phosphoprotein), the multiprotein Arp2/3 complex and cofilin . VASP interacts with the central domain of ActA . Proteins interacting with the N-terminal domain of ActA have not been identified . To identify novel host cell components of ActA-induced actin tails, we used bovine brain extracts and an affinity approach with Listeria as matrix . Several known components of Listeria tails were isolated including VASP, Arp3 and cofilin . Cofilin was identified by peptide sequencing, and cofilin recruitment and Listeria tail length were found to be pH-dependent, in agreement with its recently reported role in enhancing actin filament turnover . In addition, three proteins not previously known to be associated with Listeria tails, coronin, Rac and capZ, were identified in our affinity approach . In infected cells, the localization of the identified proteins was studied by immunofluorescence . Our findings suggest that these latter proteins, which are known to play critical roles in cellular actin rearrangements, may also be involved in the dynamics of Listeria-induced actin assembly.

Int J Food Microbiol, 1998 Jul 21, 42(3), 207 - 12
A rapid method for the identification and partial serotyping of Listeria monocytogenes in food by PCR and restriction enzyme analysis; Manzano M et al.; Two highly specific primers for Listeria monocytogenes were used to yield from foods such as milk, soft cheese and meat, PCR products that were cleaved with the restriction enzyme HindIII . The fragments generated allowed a distinction between two groups of L . monocytogenes serovars: serovars 1/2a and 1/2c cluster in one group and serovars 1/2b, 3b and 4b in the other subgroup . Since this procedure can be completed in 24 h, an epidemiological association between human disease and suspected sources can be rapidly confirmed at the subgroup level in the laboratory.

Int J Food Microbiol, 1998 Jul 21, 42(3), 201 - 6
A comparison of Listeria monocytogenes serovar 4b isolates of clinical and food origin in Japan by pulsed-field gel electrophoresis; Nakama A et al.; Pulsed-field gel electrophoresis (PFGE) patterns of 102 L . monocytogenes serovar 4b isolates from patients and foods examined in Japan were compared with 16 isolates from foodborne listeriosis episodes which occurred in North America or Europe . Using a combination of PFGE patterns with the restriction enzymes SmaI, ApaI, AscI and Sse8387I, 82 clinical isolates from Japan were categorized into 45 PFGE types: the largest group of 17 isolates (20.7%) were of the same PFGE type as cultures from the large foodborne outbreaks which occurred in California (1985) and Switzerland (1983-1987) . Twenty cultures from foods on retail sale in Japan were classified into 12 PFGE types: four isolates were of three PFGE types also recognized among isolates of clinical origin from Japan, including the predominant clinical type.

J Immunol, 1998 Sep 1, 161(5), 2428 - 34
CpG DNA induces sustained IL-12 expression in vivo and resistance to Listeria monocytogenes challenge; Krieg AM et al.; Vertebrates have evolved innate immune defense mechanisms that recognize and respond to structural patterns that are specific to microbial molecules . One such pattern recognition system is based on unmethylated CpG dinucleotides in particular sequence contexts (CpG motifs); these motifs are common in bacterial DNA but are under-represented ("CpG suppression") and methylated in vertebrate DNA . Mice that are injected with bacterial DNA or synthetic oligodeoxynucleotides (ODNs) containing CpG motifs respond with a rapid production of IL-12 and IFN-gamma . The serum levels of IL-12 were increased for at least 8 days after a single injection of CpG ODNs, but IFN-gamma levels returned to baseline within 24 h . This Th1-like cytokine response to CpG motifs induces a state of resistance to infection by Listeria monocytogenes in susceptible specific pathogen-free BALB/c mice . Resistance developed within 48 h of pretreatment with CpG ODNs, persisted for at least 2 wk, and was dependent upon IFN-gamma secretion . These data support the hypothesis that CpG DNA motifs are a "danger signal" that activates protective innate immune defenses and may have therapeutic potential.

J Immunol, 1998 Sep 1, 161(5), 2414 - 20
The role of the bacterial membrane protein ActA in immunity and protection against Listeria monocytogenes; Darji A et al.; ActA, an essential virulence factor of Listeria monocytogenes, is an integral membrane protein that is required for intracellular motility, cell-to-cell spread, and rapid dissemination of the bacteria in the infected host . To reveal cytotoxic T cell responses against ActA we introduced a recombinant soluble form of ActA into the MHC class I-processing compartment of APC using a variant of listeriolysin mutated within its immunodominant MHC class I epitope . With this experimental system we demonstrate that T cells are induced against ActA during a sublethal infection with L . monocytogenes . However, adoptively transferred cytotoxic CD8+ T cells specific for ActA did not protect mice against a subsequent challenge with this pathogen . This was due to an inability of APC to present ActA by either MHC class I or class II molecules as long as ActA remained tethered to the surface of intracellular viable bacteria . ActA was only presented when L . monocytogenes were engineered to secrete ActA or when the bacteria were killed by antibiotics during the assay . These findings raise questions on the general use of membrane proteins of pathogens as candidates for subunit vaccines.

J Immunol, 1998 Sep 1, 161(5), 2339 - 47
Bacterial surface proteins recognized by CD4+ T cells during murine infection with Listeria monocytogenes; Campbell DJ et al.; Optimal immunity to the Gram-positive pathogen Listeria monocytogenes (LM) requires both CD8+ and CD4+ antigen-specific T cell responses . Understanding how CD4+ T cells function in an immune response to LM and how bacterial proteins are processed to peptide/MHC class II complexes in infected cells requires identification of these proteins . Using LacZ-inducible, LM-specific CD4+ T cells as probes, we identified two immunogenic LM proteins by a novel expression cloning strategy . The antigenic peptides contained within these proteins were defined by deletion analysis of the genes, and their antigenicity was confirmed with synthetic peptides . The nucleotide sequences of the genes showed that they encode previously unknown LM proteins that are homologous to surface proteins in other bacterial species . Consistent with their surface topology, mild trypsin treatment of LM protoplasts ablated T cell recognition of these Ags . These findings establish a general strategy for identifying unknown CD4+ T cell Ags and demonstrate that LM surface proteins can provide the peptides for presentation by MHC class II molecules that are specific targets for CD4+ T cells during murine LM infection.

J Struct Biol, 1998, 122(1-2), 223 - 35
Sequence profile of the parallel beta helix in the pectate lyase superfamily; Heffron S et al.; The parallel beta helix structure found in the pectate lyase superfamily has been analyzed in detail . A comparative analysis of known structures has revealed a unique sequence profile, with a strong positional preference for specific amino acids oriented toward the interior of the parallel beta helix . Using the unique sequence profile, search patterns have been constructed and applied to the sequence databases to identify a subset of proteins that are likely to fold into the parallel beta helix . Of the 19 families identified, 39% are known to be carbohydrate-binding proteins, and 50% belong to a broad category of proteins with sequences containing leucine-rich repeats (LRRs) . The most striking result is the sequence match between the search pattern and four contiguous segments of internalin A, a surface protein from the bacterial pathogen Listeria monocytogenes . A plausible model of the repetitive LRR sequences of internalin A has been constructed and favorable 3D-1D profile scores have been calculated . Moreover, spectroscopic features characteristic of the parallel beta helix topology in the pectate lyases are present in the circular dichroic spectrum of internalin A . Altogether, the data support the hypothesis that sequence search patterns can be used to identify proteins, including a subset of LRR proteins, that are likely to fold into the parallel beta helix .

Rev Med Chil, 1991 Dec, 119(12), 1419 - 22
{Rhomboencephalitis caused by Listeria monocytogenes: a case studies with magnetic nuclear resonance}; Yulis J et al.; Central nervous system infections due to Listeria monocytogenes result in a variety of clinical syndromes ranging from meningitis to rhomboencephalitis . We report the case of a previously healthy patient with rhomboencephalitis in whom the CT scan was normal, while magnetic resonance imaging (MRI) confirmed the diagnosis . We review the literature and emphasize the value of MRI for timely diagnosis.

Curr Opin Immunol, 1998 Aug, 10(4), 450 - 8
Listeria monocytogenes as a probe to study cell-mediated immunity; Shen H et al.; The intracellular bacterium Listeria monocytogenes continues to serve as a model to define general paradigms of cell-mediated immunity . Genetic manipulations of the bacterium and its murine host have allowed us to begin dissecting the intricate interactions between this bacterium and the immune system . As a result, we have gained new insights into the mechanisms of immune surveillance, achieved better understanding of bacterial tactics for immune evasion and developed novel strategies in vaccine development.

J Food Prot, 1998 Aug, 61(8), 979 - 87
Fate of gamma-irradiated Listeria monocytogenes during refrigerated storage on raw or cooked turkey breast meat; Thayer DW et al.; The radiation resistance and ability of Listeria monocytogenes ATCC 7644, 15313, 43256, and 49594 to multiply on irradiated, air-packed, refrigerated raw or cooked turkey breast meat nuggets (ca . 25 g) and ground turkey breast meat was investigated . Gamma-radiation D values for L . monocytogenes were significantly different on raw and cooked nuggets, 0.56 +/- 0.03 kGy and 0.69 +/- 0.03 kGy, respectively; but they were not significantly different (P < or = 0.05) on raw and cooked ground turkey meat . High populations (approximately 10(9) CFU/g) of L . monocytogenes declined during 14 days of storage at 4 degrees C in both irradiated and nonirradiated samples of raw but not of cooked ground turkey breast meat . A moderate inoculum (approximately 10(3) CFU/g) did not survive a radiation dose of 3 kGy . The population increased in cooked but not in raw samples of irradiated ground turkey meat stored at either 2 or 7 degrees C for 21 days . The D value changed significantly from 0.70 +/- 0.04 to 0.60 +/- 0.02 kGy when the product was cooked to an internal temperature of 80 degrees C before irradiation . Growth on either raw or cooked turkey meat did not alter the radiation resistance of L . monocytogenes . Analyses were performed for pH, aw, moisture, and reducing potential of raw and cooked turkey meat and for pH, amino acid profile, thiamine, and riboflavin contents of aqueous extracts of raw and cooked turkey meats without identifying the factor or factors involved in differences in the survival and multiplication of L . monocytogenes on raw and cooked meat.

Eur J Immunol, 1998 Aug, 28(8), 2498 - 507
Human macrophages induced in vitro by macrophage colony-stimulating factor are deficient in IL-12 production; Smith W et al.; IL-12 is important for Th1 differentiation . Myeloid-derived antigen-presenting cells (APC) such as monocytes, macrophages (Mphi) and dendritic cells (DC) are believed to be major sources of IL-12 in vivo . We have compared IL-12 production of fresh monocytes with Mphi differentiated in vitro using macrophage colony-stimulating factor (M-CSF) or human plasma, and in vitro generated dendritic cells, since these differentiated cell types represent APC at sites of antigen challenge . Macrophages stimulated with lipopolysaccharide (LPS) or heat-killed Listeria monocytogenes in the presence or absence of IFN-gamma produced minimal IL-12 p70 by comparison with DC or monocytes, despite comparable production of TNF-alpha . M-CSF-induced Mphi produced low levels of IL-10 constitutively and high levels after stimulation with LPS, but neutralization of IL-10 did not augment Mphi IL-12 production . Exposure of Mphi to TNF-alpha, granulocyte-macrophage CSF or IFN-gamma did not substantially up-regulate IL-12 . Therefore M-CSF induces a differentiated Mphi phenotype in which IL-12 production is down-regulated, perhaps irreversibly . This may be the default pathway for monocyte-Mphi development in the absence of inflammation.

Cancer, 1998 Aug 15, 83(4), 817 - 20
Listeriosis in pediatric oncology patients; Mora J et al.; BACKGROUND: Adult cancer patients are considered to be at an increased risk for Listeria monocytogenes infections, but, to the authors' knowledge, little information regarding this infection in the pediatric oncology population has been published . METHODS: The Memorial Sloan-Kettering Cancer Center microbiology laboratory's database was searched for cases of Listeria monocytogenes infection during the period from January 1981 to December 1996, and thorough chart reviews of the cases identified in patients age < 21 years were performed . RESULTS: Listerial infections occurred in 5 children (3 with leukemia, 1 with lymphoma, and 1 with a brain tumor) among 20,612 admissions to the pediatric department during this period . All five children were actively receiving therapy for their malignancy, and two also were receiving other potentially immunosuppressive therapies . None was receiving co-trimoxazole prophylaxis . All were treated successfully for the Listeria monocytogenes infection with ampicillin and gentamicin (four patients) or ampicillin alone (one patient) . At last follow-up two patients were long term, event-free survivors, two had died of their underlying malignancy, and one patient had died of cytomegalovirus pneumonitis . CONCLUSIONS: Listeria monocytogenes infections in pediatric oncology patients can be treated successfully with ampicillin-containing antibiotic regimens.

Immunobiology, 1998 Jul, 199(1), 87 - 104
Macrophages and hepatocytic cells as chemokine producers in murine listeriosis; Barsig J et al.; The major target organ of systemic infection with the intracellular bacterium Listeria monocytogenes is the liver, to where inflammatory leukocytes are rapidly recruited . We determined by reverse transcriptase polymerase chain reaction the early chemokine response in the liver after systemic infection of mice with listeriae, and in parallel compared chemokine release from macrophages and hepatocytic cells in vitro . Murine bone marrow-derived macrophages (BMM) grown in fetal calf serum-supplemented medium were used as macrophages and the TIB75 cell line as hepatocytic cells . Within 1-3 hours, gene expression of monocyte chemoattractant protein (MCP)-1, macrophage inflammatory protein (MIP)-1 alpha, MIP-2, KC, and interferon-gamma inducible protein-10 (IP-10) was upregulated in liver tissue of infected mice . BMM infected in vitro with L . monocytogenes showed a generalized chemokine response, and readily released MCP-1, MIP-1 alpha, MIP-2, and KC, as measured by enzyme-linked immunosorbent assay . In contrast, the chemokine response of hepatocytic cells was more restricted, and infection induced MCP-1 and KC, but not MIP-2 and MIP-1 alpha . Interferon gamma enhanced chemokine release from hepatocytic cells, but unexpectedly had either no or a negative effect on chemokine secretion by BMM cultured in serum-supplemented medium . Listeriolysin (Hly)-negative avirulent listeriae as well as listeriae killed by heat or gentamycin initiated a similar chemokine response in BMM and hepatocytic cells as did wild-type L . monocytogenes . Stimulation of hepatocytic cells with the monokines, tumor necrosis factor alpha and interleukin (IL-)1 alpha, but not IL-6, augmented liberation of chemokines . Together, our data demonstrate an early hepatic chemokine response to L . monocytogenes in murine listeriosis . Probably, not only macrophages but also parenchymal cells participate in chemokine production.

Lett Appl Microbiol, 1998 Jun, 26(6), 432 - 6
Reduction of caecal Listeria monocytogenes in Leghorn chicks following treatment with a competitive exclusion culture (PREEMPT)
Hume ME, Byrd JA, Stanker LH, Ziprin RL.
Day-of-hatch Leghorn chicks were treated by oral gavage with PREEMPT, a continuous-flow competitive exclusion culture containing broiler caecal bacteria, followed by an oral challenge with Listeria monocytogenes, to determine the effects of PREEMPT on L . monocytogenes caecal colonization . Increased (P < 0.001) concentrations of caecal propionic acid in control chicks compared with PREEMPT-treated chicks at 3 days of age were indicative of the establishment of the PREEMPT bacteria . Caeca from control chicks at 7 days after the oral challenge with L . monocytogenes contained mean 3.4 +/- 1.4 log cfu g-1 of caecal content, while caeca from PREEMPT-treated chicks contained no detectable Listeria . Enrichment for L . monocytogenes resulted in 100% of caeca from control chicks testing culture-positive for L . monocytogenes, while none of the caeca from PREEMPT-treated chicks were culture-positive . The results indicated that prophylactic treatment of newly hatched chicks with PREEMPT significantly reduced caecal colonization by L . monocytogenes.

Folia Microbiol (Praha), 1998, 43(3), 291 - 303
Interactions of the bacterial pathogen Listeria monocytogenes with mammalian cells: bacterial factors, cellular ligands, and signaling; Cossart P; Listeria monocytogenes is a food borne pathogen which has the very unique property of crossing three barriers during infection eliciting meningitis, meningo-encephalitis and abortions with a mortality rate of about 30% . Indeed, after crossing the intestinal barrier, Listeria disseminates via the lymph and the blood, to the brain and/or the placenta after crossing the brain-blood barrier and/or the placental barrier . During disease, this organism infects a variety of tissues and cell types in which it is mostly intracellular due to its capacity to induce its own phagocytosis into cells which are normally nonphagocytic . The strategies used by Listeria to enter cells are different from those used by other well known invasive pathogens . Listeria thus appears as a fine model to study the molecular and cellular basis of bacterial invasion . In addition, not only during entry into cells but also during intra- and intercellular movement, Listeria exploits mammalian cell functions and is thus a novel tool for elucidating some unsolved fundamental aspects of cell biology, such as ligand receptor signaling and actin cytoskeleton rearrangements . In this review, the molecular and cellular basis of entry of Listeria into cells and of its intracellular motility will be discussed.

Int J Antimicrob Agents, 1998 May, 10(2), 119 - 25
Reduced intracellular activity of antibiotics against Listeria monocytogenes in multidrug resistant cells; Nichterlein T et al.; Multidrug resistance is expressed not only by bacteria, but also by tumor cells and by some normal cells of the body . It enables eukaryotic cells to exclude not only cytostatic drugs but also non-cytostatic antibiotics . This was demonstrated in genetically engineered multidrug resistant (MDR) cells infected with the facultative intracellular bacterium Listeria monocytogenes for all macrolide antibiotics tested (azithromycin, clarithromycin, erythromycin, josamycin, roxithromycin and spiramycin) . In these cells and in conventionally selected MDR cells higher concentrations of the macrolides were necessary to inhibit the growth of L . monocytogenes than in the respective parental cells . This effect was due to a reduced intracellular accumulation, which was shown with a biological assay for all macrolides tested . For azithromycin, the results of this test were confirmed by measurement of the intracellular concentrations with high-performance liquid chromatography (HPLC) . Besides the macrolides, MDR cells excluded also antibiotics of other chemical groups which was shown for ciprofloxacin, clindamycin, rifampicin and the streptogramin derivative RP 59500 . In addition, in conventionally selected cells higher concentrations of chloramphenicol, doxycyclin, ofloxacin and trimethoprim than in the respective parental cells were necessary to inhibit the growth of L . monocytogenes . In contrast, when using genetically engineered cells, no significant differences were found for these antibiotics . These differences might be due to a higher expression of multidrug resistance in the conventionally selected cells because these cells were also more effective in excluding rhodamine 123 in a flow cytometric assay . In conclusion, expression of multidrug resistance by eukaryotic cells leads to a reduced concentration of macrolides and other antibiotics in these cells and to an impairment of activity against intracellular bacteria.

Liver, 1998 Jun, 18(3), 213 - 5
Listeria monocytogenes-associated acute hepatitis in a liver transplant recipient; Vargas V et al.; We report a case of Listeria monocytogenes bacteriemia with liver involvement mimicking acute viral hepatitis in a liver transplant recipient . The patient presented with fever, jaundice and alanine aminotransferase levels ten times the upper limit of normal . Liver biopsy showed signs of acute hepatitis and occasional granulomas . Both blood and liver biopsy cultures were positive for Listeria monocytogenes . The patient received a 3 week course of ampicillin and gentamicin with clinical and biochemical recovery . Liver involvement during Listeria infections is unusual, and most cases occur in patients with underlying hepatic disease and impaired cell-mediated immunity . To our knowledge only one such case has previously been reported in a liver transplant patient . Furthermore, in the present case we isolated the causal agent from the liver biopsy specimen.

Infect Immun, 1998 Sep, 66(9), 4461 - 8
Entry of Listeria monocytogenes into neurons occurs by cell-to-cell spread: an in vitro study; Dramsi S et al.; Listeria monocytogenes is an intracellular pathogen that causes severe central nervous system infection in humans and animals . The ability of this bacterium to penetrate nerve cells was investigated by using rat spinal cell cultures . Entry into distinct cell types, i . e., glial cells and neurons, was monitored by a differential immunofluorescence technique with antibodies against cell type-specific markers and the bacterial pathogen . L . monocytogenes was detected predominantly within macrophages constituting the microglia . Astrocytes and oligodendrocytes, the major components of macroglia, were infected to a lesser extent . Surprisingly, Listeria innocua, a noninvasive and nonpathogenic species, also has the capacity to enter into these three types of glial cells . Entry into neurons was a very rare event . In contrast, we found that L . monocytogenes could efficiently invade neurons when these latter cells were cocultivated with Listeria-infected mouse macrophages . In this case, infection of neurons occurs by cell-to-cell spread via an actA-dependent mechanism . These data support the notion that infected phagocytes can be vectors by which L . monocytogenes gains access to privileged niches such as the central nervous system.

Infect Immun, 1998 Sep, 66(9), 4143 - 50
Fas (CD95)-dependent cell-mediated immunity to Listeria monocytogenes; Jensen ER et al.; Two distinct and complementary pathways, one mediated by perforin and the other dependent upon CD95 (Fas), effect cell-mediated cytotoxicity . We examined the relative roles of these pathways in host defenses against the intracellular bacterial pathogen Listeria monocytogenes by using murine listeriosis as a model system . Mice which lacked both perforin and Fas (P0L0) were generated, and their responses to primary and secondary listeriosis were compared to those of wild-type (WT), Fas-deficient (L0), and perforin knockout (P0) mice . Relative to WT mice during primary listeriosis, P0 mice exhibited a reduced capacity to clear the infection from their spleens but not their livers whereas L0 mice had elevated bacterial titers in their livers and a modestly increased titer in their spleens . In contrast, bacterial titers in P0L0 mice were increased approximately 50- to 560-fold in their spleens and 230- to 1, 000-fold in their livers; eventual clearance of listeriae from both organs was significantly delayed . Furthermore, the resistance of P0L0 mice to secondary listeriosis was significantly reduced in their spleens and livers compared to that of WT, P0, or L0 mice . In vitro experiments indicated that immune cytotoxic T lymphocytes (CTL) lysed L . monocytogenes-infected hepatocytes primarily via a Fas-dependent, perforin-independent mechanism . The absence of Fas severely abrogated the lysis of infected hepatocytes by immune CD8(+) CTL . Taken together, these results provide the first evidence for Fas-dependent CTL-mediated lysis of L . monocytogenes-infected hepatocytes and demonstrate complementary roles for Fas and perforin in host defenses against an intracellular bacterial pathogen.

Infect Immun, 1998 Sep, 66(9), 4043 - 9
The contributions of reactive oxygen intermediates and reactive nitrogen intermediates to listericidal mechanisms differ in macrophages activated pre- and postinfection; Ohya S et al.; The contribution of reactive oxygen intermediates (ROI) and reactive nitrogen intermediates (RNI) to the killing of Listeria monocytogenes by macrophages activated by addition of spleen cells from listeria-immune mice plus specific antigen was examined . When macrophages were infected with L . monocytogenes and then spleen cells were added, there was not as big a difference in listericidal activity between macrophages cultured with normal spleen cells and those cultured with immune spleen cells as expected . In this culture system, RNI was mainly involved in the macrophage intracellular killing . In macrophages first activated and then infected, a significant level of enhanced killing was observed . Blockade of ROI production drastically affected the enhanced killing ability, while inhibition of RNI production had a negligible effect . Thus, the contributions of ROI and RNI to listericidal mechanisms of macrophages were different between macrophages activated at pre- and postinfection stages.

Biochem Biophys Res Commun, 1998 Aug 19, 249(2), 526 - 30
Effect of siderophores, catecholamines, and catechol compounds on Listeria spp . Growth in iron-complexed medium; Coulanges V et al.; Almost all bacteria require iron for growth and virulence expression . However, Listeria spp . do not produce any siderophore for iron acquisition . Representative strains of each of the six species of Listeria were examined for their ability to use various compounds as iron suppliers in iron-restricted medium . Here we show that L . monocytogenes, L . innocua, L . ivanovii, L . welshimeri, L . seeligeri, and L . grayi were able to use exogenous siderophores and various catechol ligands, including catecholamines, to overcome growth inhibition induced by tropolone, an iron chelating agent . In contrast, no growth promoting effect was observed with normetanephrine or 4-hydroxy-3-methoxyphenylglycol-piperazine salt, which indicates that the o-diphenol function of the ligand must be free to allow iron acquisition . Furthermore, we demonstrate that catecholamines do not act through specific bacterial receptors, because no difference in growth stimulation was observed between {+}- and {-}-norepinephrine . These results show that utilization of a variety of catechol compounds to acquire iron is a general phenomenon in the genus Listeria .

Scand J Gastroenterol, 1998 Jul, 33(7), 778 - 82
Listeria monocytogenes in the colon in a case of fulminant ulcerative colitis; Chiba M et al.; A case of ulcerative colitis in which the presence of Listeria monocytogenes was confirmed in the resected colon with polymerase chain reaction and subsequent Southern blot analysis and immunohistochemistry using antibody against Listeria is presented . The patient developed ulcerative colitis at the age of 59 years . Prednisolone, 50 mg/day, was given for severe ulcerative colitis . Later the disease became fulminating, indicating colectomy 4 months after the onset . Multiple sealed colonic perforations were observed . Numerous L . monocytogenes were found at the site of perforation, in fissures, and in cracks in the submucosa . This case indicates the possibility that L . monocytogenes contributes to the exacerbation of colitis to fulminating and colonic perforation.

J Dairy Sci, 1998 Jul, 81(7), 1810 - 7
Survival of bioluminescent Listeria monocytogenes and Escherichia coli O157:H7 in soft cheeses; Ramsaran H et al.; Pasteurized and raw milks that had been inoculated at 10(4) cfu/ml with bioluminescent strains of Listeria monocytogenes and Escherichia coli O157:H7 were used in the manufacture of Camembert and Feta cheeses with or without nisin-producing starter culture . Survival of both organisms was determined during the manufacture and storage of Camembert and Feta cheeses at 2 +/- 1 degree C for 65 and 75 d, respectively . Bacterial bioluminescence was used as an indicator to enumerate the colonies plated on selective Listeria agar and on MacConkey agar . Escherichia coli O157:H7 survived the manufacturing process of both cheeses and was present at the end of the storage period in greater numbers than in the initial inoculum . At the end of 75 d of storage, E . coli O157:H7 was found in the brine of Feta cheese . The counts of L . monocytogenes increased as the pH of the Camembert cheese increased, and there were significant differences between the counts from samples taken from the inside and the counts from samples obtained near the surface of the cheese . The Feta cheese that contained nisin was the only cheese in which L . monocytogenes was at the level of the initial inoculum after 75 d of storage.

J Food Prot, 1998 Jun, 61(6), 683 - 7
Heat resistance and fatty acid composition of Listeria monocytogenes: effect of pH, acidulant, and growth temperature; Juneja VK et al.; The objective of this study was to determine the influence of pH, acidulant, and growth temperature history on the heat resistance and fatty acid composition of Listeria monocytogenes Scott A . Cells were grown to late exponential phase (OD600 = 0.6) at 10, 19, or 37 degrees C in brain heart infusion broth acidified to pH 5.4 or 7 with either acetic or lactic acid . Thermal death times at 60 degrees C subsequently were determined by using a submerged-coil heating apparatus . The surviving cell population was enumerated by spiral-plating heated samples onto tryptic soy agar supplemented with 0.6% yeast extract and 1% sodium pyruvate . The thermal resistance of cells cultured at a particular temperature was significantly lower (P < 0.05) when lactic acid was used to acidify the medium of pH 5.4 . Regardless of acid identity, D values significantly decreased (P < 0.05) with increased growth temperature when the pH of the growth medium was 5.4, whereas D values significantly increased (P < 0.05) with increased temperature at pH 7 . At pH 5.4 adjusted with lactic acid, D values were 1.30, 1.22, and 1.14 min for cells grown at 10, 19, and 37 degrees C, respectively . At pH 5.4 adjusted with acetic acid, L . monocytogenes failed to grow at 10 degrees C; the D values were 1.32 and 1.22 min when the cells were grown at 19 and 37 degrees C, respectively . At pH 7, the D values were 0.95, 1.12, and 1.28 min with lactic acid and 0.83, 0.93, and 1.11 min with acetic acid at 10, 19, and 37 degrees C, respectively . The most abundant fatty acids (44 to 82%) were branched-chain saturated fatty acids (anteiso-and iso-C15:0 and iso-C17:0) regardless of pH, acidulant, or growth temperature . However, there was an increase in C15:0 isomers at the expense of iso-C17:0 when the growth temperature was lowered from 37 to 10 degrees C . While variable changes in longer-chain fatty acids were found, the percentage of longer-chain (C16 and C18) fatty acids was greatest when L . monocytogenes was grown at 37 degrees C regardless of pH or acidulant . This study demonstrates that the heat resistance of L . monocytogenes depends upon its growth conditions.

J Food Prot, 1998 Mar, 61(3), 354 - 6
Incidence of Listeria monocytogenes in cheese produced in Rio de Janeiro, Brazil; da Silva MC et al.; The present study evaluated the incidence of Listeria spp . in some Brazilian cheeses obtained from retail stores in Rio de Janeiro, Of 103 samples of various types of cheese examined as recommended in the Listeria isolation protocol of the Health Protection Branch of Canada, 11 (10.68%) were contaminated by Listeria monocytogenes, 13 (12.62%) by Listeria innocua, 6 (5.83%) by Listeria grayi, and 1 (0.97%) by Listeria welshimeri . A higher incidence of L . monocytogenes as observed mainly in the homemade Minas Frescal cheeses (a Brazilian soft white cheese, eaten fresh), 7 of 17 (41.17%), followed by ripened cheeses, 3 of 53 (5.67%), and industrially manufactured Frescal (Minas and Ricotta) cheeses, 1 of 33 (3.03%) . Three serotypes (1/2a, 1/2b and 4b) were observed among the strains of L . monocytogenes isolated, all of them being frequently involved in outbreaks of foodborne listeriosis and sporadic cases of the disease all over the world.

J Food Prot, 1998 Mar, 61(3), 313 - 7
Growth of inoculated psychrotrophic pathogens on refrigerated fillets of aquacultured rainbow trout and channel catfish; Fernandes CF et al.; Aquacultured rainbow trout (Oncorhynchus mykiss) and channel catfish (Ictalurus punctatus) fillets were inoculated with the psychrotrophic pathogens Listeria monocytogenes and Aeromonas hydrophila: cell populations were monitored during refrigerated storage at 2 to 4 degrees C . Fillets of both species were placed individually in sterile plastic bags and inoculated with cell suspensions (10(4.7) CFU/100 g of fish) of either A . hydrophila or L monocytogenes or of both A . hydrophila and L . monocytogenes, for a total of three treatments for each species of fish . Each inoculum and fillet were mixed to ensure uniform distribution and then stored at 2 to 4 degrees C . A . hydrophila, L . monocytogenes, and aerobic cell populations were determined on days 1, 3, 6, 8, 10, 13, and 15 . Individually inoculated A . hydrophila and L . monocytogenes grew on catfish and trout fillets during the 15-day study . There was no inhibition of either pathogen by the natural flora on the fillets . Both psychrotrophic pathogens grew equally well in catfish and trout fillets inoculated with a combination of A . hydrophila and L . monocytogenes . In all three treatments, the counts of the psychrotrophic pathogens were lower than the aerobic plate counts . The growth of the psychrotrophic pathogens L . monocytogenes and/or A . hydrophila during refrigerated storage on aquacultured fish fillets could increase the food hazard risk, particularly where there is a possibility of cross-contamination with ready-to-eat food products.

J Food Prot, 1998 Mar, 61(3), 307 - 12
Inhibition of Listeria monocytogenes and Aeromonas hydrophila by plant extracts in refrigerated cooked beef; Hao YY et al.; Refrigerated ready-to-eat foods are becoming increasingly popular but are often vulnerable to contamination and subsequent growth by psychrotrophic foodborne pathogens . Consequently, there is a need for additional methods to assure the safety of these foods . Beef slices prepared from roasted whole sirloin tips were used in the study . Nine plant extracts were evaluated for ability to inhibit the growth of two psychrotrophic pathogens (Aeromonas hydrophila and Listeria monocytogenes) in refrigerated cooked beef . Results indicated that only eugenol (clove extract) and pimento extract significantly inhibited the growth of A . hydrophila and L . monocytogenes . However, L . monocytogenes was not as sensitive as was A . hydrophila to both treatments, especially to pimento extracts . These results suggest that plant extracts might be useful as an antimicrobial in cooked ready-to-eat meat.

J Food Prot, 1998 Feb, 61(2), 244 - 8
Studies on the risk assessment of Listeria monocytogenes; Notermans S et al.; Humans are frequently exposed to Listeria monocytogenes, and high numbers may be ingested during consumption of certain types of food . However, epidemiological investigations show that listeriosis is a rare disease . Risk assessment studies using an animal mouse model indicate that almost all L . monocytogenes serovars present in food have clear virulent properties . The intravenous dose causing infection in 50% (IV ID50) of mice not previously exposed to L . monocytogenes (nonprotected mice) was 1.8 log(10) units . For mice previously exposed to L.monocytogenes (immunologically protected mice was >9.0 log10 5.6 log(10) units . The ID(50) of orally exposed nonprotected mice amounted to 6.5 log10 units, and no significant effects of type of food (water/milk) and storage time at 5 degrees C (milk) were observed . The oral ID50 of immunologically protected mice was >9.0 log10 units . Furthermore, there was approximately 1-2 log10 difference between the ID50 and the lethal dose causing death in 50% (LD50) . The results show that both the intestinal barrier and the specific immune defense mechanism are highly effective in preventing infection of mice orally exposed to L.monocytogenes . Delaying the immune defense had no effect on the protective activity of the intestinal barrier, indicating that these protective mechanisms independently . The risk assessment results obtained in the mouse model support the epidemiological finding that listeriosis is a rare disease in humans, despite frequent exposure to the organism.

J Food Prot, 1998 Feb, 61(2), 192 - 5
Changes in populations of Listeria monocytogenes inoculated on packaged fresh-cut vegetables; Farber JM et al.; A variety of wholesale and retail packaged vegetables and salads were inoculated with a mixture of strains of Listeria monocytogenes and incubated at 4 and 10 degrees C . Whole rutabagas, butternut squash, and onions, as well as packaged Caesar salad, carrots, coleslaw mix, and stir-fry vegetables were purchased from local supermarkets in the Ottawa area . L . monocytogenes population levels remained constant on all fresh-cut vegetables stored at 4 degrees C for 9 days, except for carrots and butternut squash: counts of cell numbers declined on carrots and increased on the butternut squash . Fresh-cut vegetables stored at 10 degrees C, however, supported good growth of L . monocytogenes on all vegetables tested, except for chopped carrots, where the population decreased approximately 2 log units over a 9-day storage period . As in the situation with the produce stored at 4 degrees C, butternut squash supported the highest rate of cell growth . In addition, Caesar salad and coleslaw mix were kept at 25 degrees C for 1 or 2 days before subsequent storage at 4 or 10 degrees C to stimulate extreme temperature-abuse conditions . In Caesar salad stored at 4 degrees C, by day 6 an initial 24- and 48-h temperature abuse at 25 degrees C led to a 1.21- and 2.55-log-unit population increase, respectively, over the control . Similar increases were observed on Caesar salads stored at 10 degrees C . Compared to Caesar salad, coleslaw mix temperature-abused at 25 degrees C and then stored at 4 degrees C supported slightly greater increases in the population of L . monocytogenes, i.e., a 3.22- and 3.83-log-unit increase over the control for the 1- and 2-day abused samples, respectively . Coleslaw mix samples temperature-abused and then stored at 10 degrees, however, only showed log unit increases of 1.75 and 1.94, respectively, compared to the controls . These results point to the importance of strict temperature control to prevent or reduce the growth of L . monocytogenes cells on fresh-cut vegetables.

J Food Prot, 1998 Jan, 61(1), 119 - 22
Inactivation of Listeria innocua inoculated in liquid whole egg by high hydrostatic pressure; Ponce E et al.; The resistance of Listeria innocua, as a model microorganism for Listeria monocytogenes, to high hydrostatic pressure in liquid whole egg was studied at several pressures (300, 350, 400, and 450 MPa),temperatures (- 15, 2, and 20 degrees C), and times (5, 10, and 15 min) . Listeria innocus was added to liquid whole egg at approximately 10(6) CFU/ml . Listeria innocua was not totally inactivated in any of the treatments . In general, reduction was better at 2 degrees than at room temperature, but the greatest inactivation was obtained at 450 MPa at 20 degrees C for 15 min (over 5 log of reduction), The results indicate that microbial inactivation was increased with prolonged exposure to pressure . D values for Listeria innocua were obtained at 400 MPa for two temperatures (2 and 20 degrees C), and different times (0 to 20 min) . The microbial inactivation followed apparent first-order kinetics, exhibiting a decimal reduction time of 7.35 min at 2 degrees C and 8.23 min at 20 degrees C.

J Food Prot, 1998 Jan, 61(1), 116 - 8
Time and temperature of stretching as critical control points for Listeria monocytogenes during production of mozzarella cheese; Kim J et al.; Different heating times and temperatures commonly used during curd stretching were investigated to determine their effects on the viability of Listeria monocytogenes in mozzarella cheese . Pasteurized whole milk was inoculated with two levels of L . monocytogenes (7 and 3 log CFU/g) and coagulated with citric acid and rennet . The curd was stretched at 55, 66, and 77 degrees C for 1, 3, and 5 min . Results indicated that the majority of L . monocytogenes cells remained in the cheese curds at both inoculum levels . Stretching at 66 degrees C for 3 min reduced the number of L . monocytogenes by 5 log units, whereas stretching at 55 degrees C had a minimal effect . Stretching at 77 degrees C resulted in the complete demise of L . monocytogenes cells (from 7.6 log CFU/g to < 1.0 log CFU/g) in 1 min . If the stretching temperature partially reduced microbial counts, bring (4 degrees C for 12 h) usually had a lethal effect on the remaining microorganisms, but was less effective than the stretching temperature . These results show that stretching curd at 66 degrees C for 5 min or 77 degrees C for 1 min can effectively control L . monocytogenes during the production of mozzarella cheese.

J Food Prot, 1998 Jan, 61(1), 36 - 40
Bacterial growth in ground beef patties made with meat from animals fed diets without or with supplemental vitamin E; Cabedo L et al.; A study was designed to determine populations of aerobic bacteria, coliforms, sorbitol-negative bacteria, and Listeria monocytogenes during display at 4 and 12 degrees C of ground beef patties made with meat from animals fed diets supplemented daily (for 100 days) with 0, 1,000, or 2,000 IU of vitamin E . The patties (113.5 g) were either left uninoculated or were inoculated with Escherichia coli O157:H7 or L . monocytogenes and were tray-overwrapped and stored (at 4 or 12 degrees C for 8 to 10 or 4 to 6 days, respectively) while being continuously exposed to fluorescent light in a display setting . Patties were visually evaluated for overall appearance (based on color and/or discoloration) twice a day and analyzed for microbiological counts at 2-day intervals during display at 4 degrees C and at 0, 1, 2, 3, 4, and 6 days during display at 12 degrees C . Use of beef from animals fed supplemental vitamin E ("high-vitamin E beef") resulted in ground beef patties which, when stored at 4 degrees C, maintained visually acceptable color longer than did patties made from control beef (from animals not fed supplemental vitamin E), but effects on microbial growth were less pronounced . In general, use of high-vitamin E beef versus control beef in patty manufacture had no major effect on populations of aerobic bacteria, coliforms, sorbitol-negative bacteria, or L . monocytogenes in ground beef patties displayed at 4 or 12 degrees C . Listeria monocytogenes multiplied at 12 degrees C, but growth was similar among ground beef patties made from high-vitamin E beef versus control beef . Overall, changes in bacterial populations were similar in ground beef patties derived from meat from animals with or without added vitamin E in their diets, but control ground beef became visually unacceptable sooner.

Immunology, 1998 May, 94(1), 14 - 21
Administration of killed bacteria together with listeriolysin O induces protective immunity against Listeria monocytogenes in mice; Xiong H et al.; It is known that only listeriolysin O (LLO)-producing Listeria monocytogenes strains are able to induce protective immunity, but the underlining relationship between LLO produced by virulent strains and generation of protective immunity in the infected host remains poorly understood . In the present study, it was found that LLO gene expression was only detected in the mice infected with virulent strain which was able to induce protective immunity, while non-virulent strains or killed bacteria were not able to generate protective immunity . When mice were immunized with LLO plus killed bacteria in the presence of incomplete Freund's adjuvant, the protective immunity was partially generated, and adoptive transfer experiment confirmed that this protection was antigen specific . Reverse-transcription polymerase chain reaction revealed that LLO plus killed bacteria induced the expression of interferon-gamma (IFN-gamma) and interleukin-12 (IL-12) . Our results also showed CD4+ T cells were the principal cells constituting protective immunity . Taken together, it may be concluded that LLO produced from virulent strains of L . monocytogenes was essential for the generation of protective immunity, and that LLO plus killed bacteria induced IFN-gamma and IL-12 expression which resulted in the generation of protective immunity.

Rev Invest Clin, 1997 Jul-Aug, 49(4), 265 - 70
Report of 24 cases of Listeria monocytogenes infection at the University of Miami Medical Center; Qayyum QJ et al.; To examine the epidemiological spectrum of human listeriosis at a large municipal hospital in Miami, we reviewed the cases of Listeria monocytogenes infection seen at the University of Miami Medical Center over a nine-year period (1986-94) . Twenty-four patients (13 adults, 11 neonates) with bacteriologically proven Listeria monocytogenes infections were identified . The annual rate of listeriosis for the entire period 1986-1994 was 0.042 cases/1,000 hospital admissions . The rates of listeriosis during the first half of the study period (0.0628/1,000 admissions) were three times higher than the rates observed during the second half of the study (0.0214/1,000 admissions) . Four (57%) of the 7 adult cases of listeriosis seen after 1987 occurred in HIV-seropositive patients . Compared with the hospital population, the annual rates of listeriosis were several fold higher in patients post-renal transplant (4.65 cases/1,000 renal transplant-related admissions) and patients with HIV/AIDS (0.27 cases/1,000 HIV-related admissions) . No deaths were recorded . The decline in the annual rate of listeriosis noted in our study parallels national trends recently reported by the Centers for Disease Control and Prevention.

Int J Food Microbiol, 1998 Jun 30, 42(1-2), 127 - 31
Prevalence and growth of Listeria monocytogenes in naturally contaminated seafood; Jorgensen LV et al.; Listeria monocytogenes contamination of seafood varies with product category . The highest prevalence was found in cold-smoked fish (34-60%), while the lowest was found in heat-treated and cured seafood (4-12%) . The prevalence of L . monocytogenes differed greatly in cold-smoked salmon between production sites, ranging from < 1.4% (nil out of 70 samples) to 100% . The prevalence at the individual production sites was reproducible at repeated sampling . The results indicate that it is possible to produce cold-smoked salmon with a low prevalence of L . monocytogenes . The organism showed moderate growth in naturally contaminated cold-smoked, and 'gravad', fish while the growth appeared faster in hot smoked fish . Thus L . monocytogenes is not under control in these products . Finally, the prevalence and growth of L . monocytogenes in naturally contaminated cold-smoked salmon are discussed in relation to controlling this risk.

Int J Food Microbiol, 1998 Jun 30, 42(1-2), 71 - 7
Effects of pH or a(w) stress on growth of Listeria monocytogenes; Cheroutre-Vialette M et al.; The growth of three strains of Listeria monocytogenes at 20 degrees C in a meat broth of different pH or water activity was investigated . At inoculation or at the beginning of the exponential phase, cells were exposed to stress by the addition of NaOH or NH4+, acetic acid, NaCl or KCl, in order to reach a pH of either 9.0 or 5.6, or an a(w) of 0.950 or 0.965, respectively . The effects of the exposure to stress on the generation and lag times of each strain were analysed by turbidity measurements for cultures in micro-titer plates . Results were confirmed by conducting the same experiments in a fermentor, except for the maximal population reached . The three strains showed similar behaviour . Cells were able to overcome the alkaline stress rapidly whereas acid and osmotic shocks induced important changes of the growth parameters . Cells exposed to acid or osmotic conditions from the time of inoculation were less affected than cells exposed at the beginning of the mid-exponential phase.

J Clin Microbiol, 1998 Sep, 36(9), 2439 - 42
Evaluation of the RapID CB plus system for identification of coryneform bacteria and Listeria spp; Funke G et al.; In a stress test, the recently introduced RapID CB Plus system (Remel Inc . {formerly Innovative Diagnostic Systems}, Norcross, Ga.) was challenged with a diverse set of gram-positive rods comprising 345 strains of coryneform bacteria and 33 strains of Listeria spp . representing a total of 49 different taxa . Overall, within 4 h, the system correctly identified 80.9% of the strains on the species level and 12.2% of the strains on the genus level . Only 3.7% strains were misidentified, and for 3.2% of the strains no identification was provided . Difficulties with the system were mainly due to occasional uncertainties in reading reactions for acid production from carbohydrates and, to a lesser extent, aminopeptidase reactions . It is concluded that the system may also perform well under the conditions of a routine clinical laboratory.

J Intern Med, 1998 Jul, 244(1), 87 - 90
Septic arthritis with Listeria monocytogenes during low-dose methotrexate; Jansen TL et al.; We describe a 22-year-old female with systemic lupus erythematosus and lymphopenia, who developed septic arthritis of the right knee with Listeria monocytogenes type 1/2 A, whilst on low-dose methotrexate (MTX) . So far, septic arthritis due to this microorganism has been reported in two other patients treated with low-dose MTX, one having rheumatoid arthritis and the other psoriatic arthritis . No reports exist on patients treated with other cytotoxic antirheumatic therapies.

Ann Trop Paediatr, 1998 Mar, 18(1), 61 - 2
Perinephric abscess (presenting as abdominal pain) due to Listeria monocytogenes; Gomber S et al.; A 5-year-old malnourished child was admitted with a 1-week history of paroxysmal abdominal pain . Evaluation finally revealed a left-sided perinephric abscess caused by Listeria monocytogenes . The child was successfully treated by drainage of the abscess and antibiotic therapy . Renal abscess should be kept in mind in the differential diagnosis of abdominal pain, and Listeria, an unusual pathogen, should be considered as a possible aetiology.

Appl Environ Microbiol, 1998 Aug, 64(8), 3070 - 4
Unstable expression and thermal instability of a species-specific cell surface epitope associated with a 66-kilodalton antigen recognized by monoclonal antibody EM-7G1 within serotypes of Listeria monocytogenes grown in nonselective and selective broths; Nannapaneni R et al.; Conditions that resulted in unstable expression and heat instability of a cell surface epitope associated with a 66-kDa antigen in Listeria monocytogenes serotypes were identified with the probe monoclonal antibody (MAb) EM-7G1 in an enzyme-linked immunosorbent assay . This epitope appeared to be absent in three serotypes (serotypes 3b, 4a, and 4c), which did not react with MAb EM-7G1 irrespective of the enrichment broth tested . The remaining 10 serotypes were detected by MAb EM-7G1 only when cells were grown in nonselective brain heart infusion broth (BHI) or selective Listeria enrichment broth (LEB) . When cells were grown in Listeria repair broth (LRB), only 6 of the 13 serotypes were detected by MAb EM-7G1, and recognition of serogroup 4 was completely lost . None of the 13 serotypes was detected by MAb EM-7G1 when cells were grown in two other commonly used Listeria-selective media, UVM1 broth and Fraser broth (FRB), indicating that possible loss of epitope expression occurred under these conditions . MAb EM-7G1 maintained species specificity without cross-reacting with live or heat-killed cells of six other Listeria spp . (Listeria ivanovii, Listeria innocua, Listeria seeligeri, Listeria welshimeri, Listeria grayi, and Listeria murrayi) irrespective of the enrichment conditions tested . Due to heat instability of the cell surface epitope when it was exposed to 80 or 100 degrees C for 20 min, MAb EM-7G1 is suitable for detection of live cells of L . monocytogenes in BHI or LEB but not in LRB, UVM1, or FRB enrichment medium.

J Immunol, 1998 Aug 1, 161(3), 1447 - 53
Control of IL-12 and IFN-gamma production in response to live or dead bacteria by TNF and other factors; Zhan Y et al.; When mice were infected i.v . with either Listeria monocytogenes or Brucella abortus, bioactive IL-12 was briefly detected in serum and supernatants of spleen homogenates immediately ex vivo . Although the time scale was more prolonged for the more slowly growing B . abortus, in both instances IL-12 production ceased while bacteria still persisted in high numbers . Production of IL-12, detected in serum and spleen, was neither increased nor prolonged by injecting Abs to IL-10 or IL-4 . In contrast with live organisms, heat-killed bacteria did not induce detectable IL-12 in vivo and were less efficient when added in vitro to resident peritoneal cells or spleen cells . Mice lacking the receptors for TNF (TNFR-/- mice) were severely deficient in IL-12 production, suggesting a controlling role for TNF, which we have previously shown to be triggered by live, rather than dead, bacteria . Infection in the TNFR-/- mice was exacerbated, although in the Brucella-infected mice splenomegaly, the main indicator of immunopathology, was reduced . Production of NO by macrophages was deficient, but the TNFR-/- mice were not deficient in IFN-gamma production . In addition to being poor inducers of IL-12, killed bacteria actively suppressed IL-12 production in response to live bacteria, by mechanism(s) unknown . The implications of these findings are discussed in light of the fact that only live bacteria satisfactorily induce cell-mediated immunity to infection.

Cell Immunol . 1998 Jul 10;187(1):76.
Papers to Appear in Forthcoming Issues; Specificity of the BAX polymerase chain reaction system for detection of the foodborne pathogen Listeria monocytogenes; U.S . Food and Drug Administration, National Center for Food Safety and Technology, Summit-Argo, IL 60501, USAThe polymerase chain reaction (PCR) can be used for rapid and specific detection of foodborne pathogens . One commercial kit, the Qualicon BAX system uses PCR to detect Listeria monocytogenes in enrichment cultures derived from food and environmental samples . The specificity and sensitivity of the BAX system for detecting L . monocytogenes were characterized by using both pure and mixed cell cultures, and optimal conditions for production of cell lysates were determined . The BAX system was highly specific for L . monocytogenes, and no interference was seen in the presence of either other Listeria species or microbes from other genera . The assay detected L . monocytogenes at 10(5)-10(6) colony-forming units/mL . This sensitivity is adequate for detecting viable cells after enrichment but prevents false-positive signals from nonviable cells.

Mol Microbiol, 1998 Jun, 28(6), 1081 - 9
Listeriolysin O: cholesterol inhibits cytolysis but not binding to cellular membranes; Jacobs T et al.; Listeriolysin O (LLO) binds to cholesterol-containing membranes in which it oligomerizes to form pores . Preincubation of the toxin with cholesterol is known to inhibit haemolysis, whereas the oxidized form of cholesterol has no inhibitory effect . Using immunoblot analyses and flow cytometry we demonstrate that preincubation with cholesterol does not influence binding of the listeriolysin-cholesterol complex to red blood cells, eukaryotic cells or artificial membranes . Lytic activity of membrane-bound LLO inactivated by cholesterol can be restored by enzymatic treatment with cholesterol oxidase . To determine the step at which cholesterol inhibits lytic activity, we looked for pore formation using electron microscopy . Pores formed by purified listeriolysin could be directly visualized using erythrocyte ghosts . This property was lost upon incubation of the toxin with cholesterol . Quantitative analysis strongly suggest that inhibition of lysis by cholesterol is not due to decreased binding of listeriolysin to target membranes, but rather to an interference with a subsequent step leading to polymerization of the toxin.

Int Immunol, 1998 Jun, 10(6), 757 - 65
Differential chemokine response of murine macrophages stimulated with cytokines and infected with Listeria monocytogenes; Flesch IE et al.; During inflammatory processes the infected macrophage is a rich source of chemokines which induce infiltration of leukocytes to the site of infection . We investigated the regulation of chemokine production by murine macrophages in response to infection with the intracellular bacterial pathogen, Listeria monocytogenes . As a source of quiescent macrophages, murine bone marrow-derived macrophages (BMM) cultured under serum-free conditions were used . With RT-PCR, we detected induction of RNA message for the chemokines macrophage inflammatory protein (MIP)-2, KC, MIP-1alpha, MIP-1beta, IFN-gamma-inducible protein-10 and RANTES in L . monocytogenes-infected macrophages . Accordingly, ELISA-detectable MIP-1alpha, MIP-2 and KC protein was induced by infection with L . monocytogenes . In contrast, L . monocytogenes infection of BMM alone failed to induce considerable expression of monocyte chemoattractant protein (MCP)-1 at the mRNA or protein level, but co-treatment with IFN-gamma was necessary . Release of infection-triggered MIP-2, MIP-1alpha and KC was negatively regulated by IFN-gamma . Similarly, IL-4 stimulated MCP-1 release by infected macrophages but reduced production of MIP-1alpha, MIP-2 and KC . IL-10 turned out to be a general deactivator in terms of macrophage chemokine production . IL-13 had no effect on MIP-1alpha, MIP-2 and KC production by infected BMM, but slightly reduced MCP-1 release . By using IFN-gamma and IL-4 gene deletion mutant mice, in vivo regulation of these chemokines by IL-4 and IFN-gamma in listeriosis was studied . In summary, our results show that chemokines are produced by macrophages infected with L . monocytogenes, and that chemokine release is differentially regulated by the macrophage modulators IFN-gamma, IL-4, IL-10 and IL-13.

J Food Prot, 1998 Jul, 61(7), 899 - 902
Heat stability of virulence-associated enzymes from Listeria monocytogenes SLCC 5764; Zemser RB et al.; The enzyme activities of listeriolysin O (LLO), phosphatidylinositol-specific phospholipase C (PI-PLC), catalase (CA), and superoxide dismutase (SOD) from Listeria monocytogenes SLCC 5764 were examined after heat treatment, growth at elevated temperatures, and anaerobic growth . Growth at elevated temperatures may influence virulence as expressed by virulence enzyme activity . The enzymes were heated for 0 and 10 min at temperatures ranging from 40 to 100 degrees C . The production of LLO was examined when the bacterium was grown at elevated temperatures, and the production of LLO, CA, and SOD were examined after anaerobic growth . LLO was the most heat-labile factor, losing almost all activity when heated above 50 degrees C . CA was more heat stable, showing no decrease in activity until it was heated at 55 degrees C . The PI-PLC enzyme was most heat resistant: the activity decreased between 40 and 50 degrees C and continued to decrease when heated between 65 and 100 degrees C . When L . monocytogenes was grown at elevated temperatures, it produced less LLO than when grown at the optimum growth temperature of 37 degrees C . When the organism was grown under anaerobic conditions, the levels of CA and LLO decreased, while the SOD level remained unchanged.

J Food Prot, 1998 Jul, 61(7), 849 - 54
Reducing total aerobic counts and Listeria monocytogenes on the surface of king salmon (Oncorhynchus tshawytscha); Bremer PJ et al.; A trial industrial-scale fin-fish washing system was assessed for its effectiveness in removing bacteria associated with the skin of gilled and gutted king salmon (Oncorhynchus tshawytscha) . Exposure of the salmon to 200 ppm free chlorine at a turnover rate for the total volume of the wash solution of 2.25 cycles h-1 for 120 min resulted in decreases in the aerobic plate count (APC) recovered from the salmon ranging from 96.6 to 99.2% . In order to optimize the washing regime a laboratory-scale fin-fish washing system was developed . Twenty-six washing treatments were used to generate a model to relate efficacy of bacterial removal with chlorine concentration, flow rate, and duration of washing . The model gave two local maxima of percentage APC reduction, one of 99.3% at a concentration of 126.3 ppm chlorine with a turnover of 0.75 cycle h-1 and a duration of 71.3 min and a second of 100.6% at a concentration of 126.3 ppm chlorine with a turnover of 3.75 cycles h-1 and a duration of 120 min . In additional experiments, it was determined that washing could eliminate 99.79% of Listeria monocytogenes cells that had been artificially inoculated onto the surface of gilled and gutted salmon . It was concluded that while chlorinated wash regimens have the potential to reduce the carriage of bacteria, including L . monocytogenes, into fish-processing facilities, they will not ensure an L . monocytogenes-free product . Further, the use of such a system has to be assessed with regard to allowable chlorine levels (subject to regulation), the effect of washing on the quality of the finished product, and the cost of water purchase and disposal.

J Appl Microbiol, 1998 May, 84(5), 715 - 21
Bacteriocin inhibition of two glucose transport systems in Listeria monocytogenes; Waite BL et al.; Listeria monocytogenes transports glucose by proton motive force-mediated and phosphoenolpyruvate-dependent phosphotransferase systems (PEP-dependent PTS) . Inhibition of both systems by nisin, pediocin JD and leuconosin S is reported here for four strains of L . monocytogenes . Intracellular and extracellular adenosine triphosphate (ATP) and extracellular inorganic phosphate were measured in energized L . monocytogenes Scott A cells to determine whether inhibition of the PEP-dependent PTS might occur as a result of bacteriocin-induced leakage of intracellular components . Addition of nisin resulted in a decrease in intracellular ATP with an increase in extracellular ATP . Leuconosin S and pediocin JD induced a depletion of intracellular ATP . ATP efflux was low for the leuconosin S-treated cells and barely detectable for pediocin JD-treated cells . Addition of nisin, leuconosin S and pediocin JD induced efflux of inorganic phosphate . It appears that bacteriocin-mediated inhibition of the glucose PEP-dependent PTS occurs as a result of hydrolysis or efflux of ATP, PEP and other essential molecules from L . monocytogenes cells.

J Vet Med Sci, 1998 Jun, 60(6), 749 - 52
Molecular typing of Listeria monocytogenes isolated in Japan by pulsed-field gel electrophoresis; Nakama A et al.; A total of 40 strains of Listeria monocytogenes which have been demonstrated to be serovar 1/2a, 1/2b, and 4b were genotyped by pulsed-field gel electrophoresis (PFGE) after separate digestion with Apa I, Asc I, Sma I, and Sse 8387 I . Twenty-seven unrelated strains including four representative strains showed distinctly different genotypes according to their PFGE profiles . Then nine strains isolated from shredded cheese of different lots and four strains isolated from the cheese-processing environment were shown to display the same genotype . Therefore, it is suggested that the Listeria was spread in cheese by cross-contamination from the cheese-processing environment . Thus, PFGE analysis has a good typeability and excellent discriminatory power, and has provided a useful tool for investigation of the source of Listeria contamination.

Infect Immun, 1998 Aug, 66(8), 3775 - 82
Gelsolin, a protein that caps the barbed ends and severs actin filaments, enhances the actin-based motility of Listeria monocytogenes in host cells; Laine RO et al.; The actin-based motility of Listeria monocytogenes requires the addition of actin monomers to the barbed or plus ends of actin filaments . Immunofluorescence micrographs have demonstrated that gelsolin, a protein that both caps barbed ends and severs actin filaments, is concentrated directly behind motile bacteria at the junction between the actin filament rocket tail and the bacterium . In contrast, CapG, a protein that strictly caps actin filaments, fails to localize near intracellular Listeria . To explore the effect of increasing concentrations of gelsolin on bacterial motility, NIH 3T3 fibroblasts stably transfected with gelsolin cDNA were infected with Listeria . The C5 cell line containing 2.25 times control levels of gelsolin supported significantly higher velocities of bacterial movement than did control fibroblasts (mean +/- standard error of the mean, 0.09 +/- 0.003 micro(m)/s {n = 176} versus 0.05 +/- 0.003 micro(m)/s {n = 65}) . The rate of disassembly of the Listeria-induced actin filament rocket tail was found to be independent of gelsolin content . Therefore, if increases in gelsolin content result in increases in Listeria-induced rocket tail assembly rates, a positive correlation between gelsolin content and tail length would be expected . BODIPY-phalloidin staining of four different stably transfected NIH 3T3 fibroblast cell lines confirmed this expectation (r = 0.92) . Rocket tails were significantly longer in cells with a high gelsolin content . Microinjection of gelsolin 1/2 (consisting of the amino-terminal half of native gelsolin) also increased bacterial velocity by more than 2.2 times . Microinjection of CapG had no effect on bacterial movement . Cultured skin fibroblasts derived from gelsolin-null mice were capable of supporting intracellular Listeria motility at velocities comparable to those supported by wild-type skin fibroblasts . These experiments demonstrated that the surface of Listeria contains a polymerization zone that can block the barbed-end-capping activity of both gelsolin and CapG . The ability of Listeria to uncap actin filaments combined with the severing activity of gelsolin can accelerate actin-based motility . However, gelsolin is not absolutely required for the actin-based intracellular movement of Listeria because its function can be replaced by other actin regulatory proteins in gelsolin-null cells, demonstrating the functional redundancy of the actin system.

Infect Immun, 1998 Aug, 66(8), 3673 - 81
Listeria monocytogenes stimulates mucus exocytosis in cultured human polarized mucosecreting intestinal cells through action of listeriolysin O; Coconnier MH et al.; When the intracellular pathogen Listeria monocytogenes infects cultured human mucosecreting polarized HT29-MTX cells apically, it induces the stimulation of mucus exocytosis without cell entry . Using a set of isogenic mutants and purified listeriolysin O (LLO), we identified the L . monocytogenes thiol-activated exotoxin LLO as the agonist of mucus secretion . We demonstrated that the LLO-induced mucus exocytosis did not result from the LLO membrane-damaging activity . We found that LLO-induced mucus exocytosis is an event requiring the binding of LLO to a brush border-associated receptor and membrane oligomerization of the exotoxin . By a pharmacological approach, we demonstrated that no regulatory system or intracellular transducing signal known to be involved in control of mucin exocytosis was activated by LLO . Based on the present data, the stimulatory action of LLO on mucin exocytosis could be accounted for either by an unknown signaling system which remains to be determined or by direct action of LLO with the membrane vesicle components involved in the intracellular vesicular transport of mucins.

Infect Immun, 1998 Aug, 66(8), 3635 - 42
Regulation of hly expression in Listeria monocytogenes by carbon sources and pH occurs through separate mechanisms mediated by PrfA; Behari J et al.; Expression of the PrfA-controlled virulence gene hly (encoding the pore-forming cytolysin listeriolysin) is under negative regulation by readily metabolized carbon sources in Listeria monocytogenes . However, the hyperhemolytic strain NCTC 7973 exhibits deregulated hly expression in the presence of repressing sugars, raising the possibility that a defect in carbon source regulation is responsible for its anomalous behavior . We show here that the activity of a second glucose-repressed enzyme, alpha-glucosidase, is 10-fold higher in NCTC 7973 than in 10403S . Using hly-gus fusions, we show that the prfA allele from NCTC 7973 causes deregulated hly-gus expression in the presence of sugars in either the wild-type or the NCTC 7973 background, while the 10403S prfA allele restores carbon source regulation . However, the prfA genotype does not affect the regulation of alpha-glucosidase activity by repressing sugars . Of the two mutational differences in PrfA, only a Gly145Ser change is important for regulation of hly-gus . Therefore, NCTC 7973 and 10403S have genetic differences in at least two loci: one in prfA that affects carbon source regulation of virulence genes and another in an unidentified gene(s) that up-regulates alpha-glucosidase activity . We also show that the decrease in pH associated with utilization of sugars negatively regulates hly-gus expression, although sugars can affect hly-gus expression by another mechanism that is independent of pH.

Infect Immun, 1998 Aug, 66(8), 3611 - 7
The role of protegrins and other elastase-activated polypeptides in the bactericidal properties of porcine inflammatory fluids; Shi J et al.; The mammalian host response to infection includes the production and secretion of antimicrobial peptides from phagocytes and epithelial cells . Protegrins, a group of broadly microbicidal peptides isolated originally from porcine neutrophil lysates, were found to be stored as inactive proforms in porcine neutrophil granules but could be activated extracellularly by neutrophil elastase . We assessed the biological role of protegrins and other elastase-activated polypeptides in the microbicidal activity of neutrophil secretions and inflammatory fluids . When stimulated with phorbol myristate acetate (PMA), neutrophils generated stable microbicidal activity against both Escherichia coli and Listeria monocytogenes under normal-salt conditions and in the presence of 0 to 10% serum . The generation of these antimicrobial substances was dependent on neutrophil elastase, since it was inhibited by 1 mM N-methoxysuccinyl-Ala-Ala-Pro-Val chloromethyl ketone when it was present during activation, but not when this inhibitor was added afterwards . However, elastase-dependent activation of proprotegrins to protegrins in PMA-stimulated neutrophils was not inhibited by the presence of 1 to 2% serum . Porcine neutrophils also released antibacterial activity during phagocytosis of latex beads, and this too was dependent in large part on elastase-activated polypeptides, including protegrins . Moreover, protegrins were found at bactericidal concentrations in cell-free abscess fluid from naturally infected pigs . Taken together, these studies show that protegrins and other elastase-activated polypeptides are important stable antibacterial factors in porcine neutrophil secretions . The potential host defense role of elastase as an activating enzyme for the precursors of microbicidal peptides must be taken into account when therapeutic inhibitors of neutrophil elastase are evaluated for clinical use as anti-inflammatory agents.

Infect Immun, 1998 Aug, 66(8), 3552 - 61
Pathogenicity and immunogenicity of a Listeria monocytogenes strain that requires D-alanine for growth; Thompson RJ et al.; Listeria monocytogenes is an intracellular bacterial pathogen that elicits a strong cellular immune response following infection and therefore has potential use as a vaccine vector . However, while infections by L . monocytogenes are fairly rare and can readily be controlled by a number of antibiotics, the organism can nevertheless cause meningitis and death, particularly in immunocompromised or pregnant patients . We therefore have endeavored to isolate a highly attenuated strain of this organism for use as a vaccine vector . D-Alanine is required for the synthesis of the mucopeptide component of the cell walls of virtually all bacteria and is found almost exclusively in the microbial world . We have found in L . monocytogenes two genes that control the synthesis of this compound, an alanine racemase gene (dal) and a D-amino acid aminotransferase gene (dat) . By inactivating both genes, we produced an organism that could be grown in the laboratory when supplemented with D-alanine but was unable to grow outside the laboratory, particularly in the cytoplasm of eukaryotic host cells, the natural habitat of this organism during infection . In mice, the double-mutant strain was completely attenuated . Nevertheless, it showed the ability, particularly under conditions of transient suppression of the mutant phenotype, to induce cytotoxic T-lymphocyte responses and to generate protective immunity against lethal challenge by wild-type L . monocytogenes equivalent to that induced by the wild-type organism.

EMBO J, 1998 Jul 15, 17(14), 3797 - 806
Interactions of Listeria monocytogenes with mammalian cells during entry and actin-based movement: bacterial factors, cellular ligands and signaling; Cossart P et al.; Although <50 kb of its 3.3 megabase genome is known, Listeria monocytogenes has received much attention and an impressive amount of data has contributed in raising this bacterium among the best understood intracellular pathogens . The mechanisms that Listeria uses to enter cells, escape from the phagocytic vacuole and spread from one cell to another using an actin-based motility process have been analysed in detail . Several bacterial proteins contributing to these events have been identified, including the invasion proteins internalin A (InlA) and B (InlB), the secreted pore-forming toxin listeriolysin O (LLO) which promotes the escape from the phagocytic vacuole, and the surface protein ActA which is required for actin polymerization and bacterial movement . While LLO and ActA are critical for the infectious process and are not redundant with other listerial proteins, the precise role of InlA and InlB in vivo remains unclear . How InlA, InlB, LLO or ActA interact with the mammalian cells is beginning to be deciphered . The picture that emerges is that this bacterium uses general strategies also used by other invasive bacteria but has evolved a panel of specific tools and tricks to exploit mammalian cell functions . Their study may lead to a better understanding of important questions in cell biology such as ligand receptor signalling and dynamics of actin polymerization in mammalian cells.

Vet Immunol Immunopathol, 1998 Jul 8, 64(2), 149 - 59
Effect of in vitro monocyte activation by Listeria monocytogenes antigens on phagocytosis and production of reactive oxygen and nitrogen radicals in bovines; Barbuddhe SB et al.; Macrophages by virtue of their phagocytic and antibacterial activities play an important role in the host resistance to intracellular pathogens including Listeria monocytogenes . However, the precise killing mechanism adopted by macrophages in the case of L . monocytogenes and the role of macrophage activation in bacterial killing are still unclear . In the present investigation, different antigens of pathogenic L . monocytogenes and three non-specific activators, namely, Lipopoly-saccharide (LPS), Phorbol Myristate Acetate (PMA) and Concanavalin A (ConA) supernatant were studied to adjudge their efficacy with regard to in vitro activation of bovine monocytes in terms of the production of reactive nitrogen intermediates (RNI), reactive oxygen intermediates (ROI) and their phagocytic index (PI) . Of all the five antigens of L . monocytogenes, namely, viable bacteria used as live antigen (LA), heat-killed antigen (HKA), sonicated antigen (SA), culture filtrate antigen (CFA) and listeriolysin-O (LLO), LA turned out to be the best activator of monocytes for RNI as well as ROI production . In the PI assay, of the three antigens, that is, CFA, SA and LLO, CFA was found to be the best activator of phagocytosis followed by LLO and SA . Among non-specific activators, PMA induced the highest H2O2 production whereas LPS caused the maximum increase in RNI and PI production . On activation by different antigens of L . monocytogenes, the peripheral blood mononuclear cells (PBMCs) from infected animals produced significantly higher RNI than those from non-infected animals indicating the involvement of immune T-cells.

Proc Natl Acad Sci U S A, 1998 Jul 7, 95(14), 8233 - 8
Targeted disruption of the interferon-gamma receptor 2 gene results in severe immune defects in mice; Lu B et al.; To study the role of the interferon- (IFN) gammaR2 chain in IFN-gamma signaling and immune function, IFN-gammaR2-deficient mice have been generated and characterized . Cells derived from IFN-gammaR2 -/- mice are unable to activate either JAK/STAT signaling proteins or gene transcription in response to IFN-gamma . The lack of IFN-gamma responsiveness alters IFN-gamma-induced Ig class switching by B cells from these mice . In vitro cultures of T cells demonstrate that the T cells from the IFN-gammaR2 -/- mice have a defect in Th1 cell differentiation . The IFN-gammaR2 (-/-) mice also produce lower amounts of IFN-gamma in response to antigenic challenge . In addition, IFN-gammaR2 -/- mice are defective in contact hypersensitivity and are highly susceptible to infection by Listeria monocytogenes . These results demonstrate that the IFN-gammaR2 is essential for IFN-gamma-mediated immune responses in vivo.

J Exp Med, 1998 Jul 6, 188(1), 61 - 70
Evolution of a complex T cell receptor repertoire during primary and recall bacterial infection; Busch DH et al.; The mechanisms underlying the genesis and maintenance of T cell memory remain unclear . In this study, we examined the evolution of a complex, antigen-specific T cell population during the transition from primary effector to memory T cells after Listeria monocytogenes infection . T cell populations specific for listeriolysin O (LLO)91-99, the immunodominant epitope recognized by H2-Kd-restricted T lymphocytes, were directly identified in immune spleens using tetrameric H2-Kd-epitope complexes . The T cell receptor (TCR) Vbeta repertoire of specific T cells was determined by direct, ex vivo staining with a panel of mAbs . We demonstrate that LLO91-99-specific, primary effector T cell populations have a diverse TCR Vbeta repertoire . Analyses of memory T cell populations demonstrated similar TCR diversity . Furthermore, experiments with individual mice demonstrated that primary effector and memory T cells have indistinguishable TCR repertoires . Remarkably, after reinfection with L . monocytogenes, LLO91-99-specific T cells have a narrower TCR repertoire than do primary effector or memory T cells . Thus, our studies show that the TCR repertoire of primary effector T lymphocytes is uniformly transmitted to memory T cells, whereas expansion of memory T cells is selective.

Science, 1998 Jul 3, 281(5373), 105 - 8
Interaction of human Arp2/3 complex and the Listeria monocytogenes ActA protein in actin filament nucleation; Welch MD et al.; Actin filament assembly at the cell surface of the pathogenic bacterium Listeria monocytogenes requires the bacterial ActA surface protein and the host cell Arp2/3 complex . Purified Arp2/3 complex accelerated the nucleation of actin polymerization in vitro, but pure ActA had no effect . However, when combined, the Arp2/3 complex and ActA synergistically stimulated the nucleation of actin filaments . This mechanism of activating the host Arp2/3 complex at the L . monocytogenes surface may be similar to the strategy used by cells to control Arp2/3 complex activity and hence the spatial and temporal distribution of actin polymerization.

Eur J Immunol, 1998 Jun, 28(6), 1807 - 14
Attenuated Listeria monocytogenes carrier strains can deliver an HIV-1 gp120 T helper epitope to MHC class II-restricted human CD4+ T cells; Guzman CA et al.; Listeria monocytogenes is a facultative intracellular pathogen which, following uptake by macrophages, escapes from the phagosome and replicates in the cytoplasm . This property has been exploited using recombinant L . monocytogenes as a carrier for the intracytoplasmic expression of antigens when MHC class I-restricted cytotoxic T lymphocyte responses are required . Much less is known of the ability of these bacteria to trigger MHC class II-restricted responses . Here, we demonstrate that after ingestion of L . monocytogenes expressing a T helper epitope from the gp120 envelope glycoprotein of HIV, human adherent macrophages and dendritic cells can process and present the epitope to a specific CD4+ T cell line in the context of MHC class II molecules . No significant differences were observed when the attenuated strains were trapped in the phagolysosome or impaired in the capacity to spread intracellularly or from cell to cell . Similar results were obtained using carrier proteins that were either secreted, associated with the bacterial surface, or restricted to the bacterial cytoplasm . A dominant expression of the TCR Vbeta 22 gene subfamily was observed in specific T cell lines generated after stimulation with the recombinant strains or with soluble gp120 . Our data show that in this in vitro system L . monocytogenes can efficiently deliver antigens to the MHC class II pathway, in addition to the well-established MHC class I pathway . The eukaryotic cell compartment in which the antigen is synthesized, and the mode of display seem to play a minor role in the overall efficiency of epitope processing and presentation.

Kansenshogaku Zasshi, 1998 May, 72(5), 548 - 52
{A case of Listeria monocytogenes sepsis in an elderly who survived}; Kobashi Y et al.; A 81-year-old man who complained of fever and disturbance of consciousness was admitted to our hospital . Listeria monocytogenes type 1/2aA was cultured from only the blood . He was treated with gamma-globrine and sensitive antibiotics (PAPM/BP, EM) immediately after admission, and recovered in spite of multiple organ failure due to septic shock . He was not an immunocompromised host and did not have complication of meningitis, but had rhabdomyolysis and liver dysfunction.

Infect Immun, 1998 Jul, 66(7), 3467 - 9
Nonspecific early protective immunity in Francisella and Listeria infections can be dependent on lymphocytes; Elkins KL et al.; Normal mice, but not lymphocyte-deficient or B-cell-deficient mice, given a sublethal infection of Francisella tularensis LVS survive a secondary lethal challenge of more than 10,000 50% lethal doses given 3 days later . In this work, we show that similar early protection that is also strongly lymphocyte dependent operates in Listeria monocytogenes infection . Since sublethal infection with either LVS or L . monocytogenes protects against heterologous lethal challenge, this early protection is nonspecific.

Infect Immun, 1998 Jul, 66(7), 3420 - 2
The inlA gene of Listeria monocytogenes LO28 harbors a nonsense mutation resulting in release of internalin; Jonquieres R et al.; Internalin is a surface protein that mediates entry of Listeria monocytogenes EGD into epithelial cells expressing the cell adhesion molecule human E-cadherin or its chicken homolog, L-CAM, which act as receptors for internalin . After observing that entry of L . monocytogenes LO28 into S180 fibroblasts, in contrast to that of EGD, did not increase after transfection with L-CAM, we examined both the expression and the structure of internalin in strain LO28 . We discovered a nonsense mutation in inlA which results in a truncated protein released in the culture medium . Mutations leading to release of internalin were also detected in clinical and food isolates . These results question the role of internalin as a virulence factor in murine listeriosis.

Infect Immun, 1998 Jul, 66(7), 3250 - 4
Construction and vaccine potential of acapsular mutants of Erysipelothrix rhusiopathiae: use of excision of Tn916 to inactivate a target gene; Shimoji Y et al.; We previously showed that acapsular transposon Tn916 mutants of Erysipelothrix rhusiopathiae are avirulent for mice . In this study, we constructed nonreverting acapsular mutants and examined the vaccine potential of the mutants in mice . A representative acapsular transposon mutant, 33H6, was plated on selective agar containing autoclaved chlortetracycline and quinaldic acid, and two tetracycline-sensitive mutants were obtained . Sequence analysis of chromosomal regions of the mutants in which Tn916 had flanked revealed that Tn916 had spontaneously excised from the region and that the six new nucleotides, which were presumably inserted with Tn916 into 33H6 chromosome, substituted for those present at the insertion site . The mutants were confirmed to be devoid of capsular antigen by Western immunoblotting and were nonvirulent for mice (subcutaneous 50% lethal dose {LD50}, >10(9) CFU) . The safety and efficacy of acapsular mutants for live vaccines was further studied by using one mutant strain, named YS-1 . The YS-1 bacteria were cleared from the skin sites of inoculation, livers, and spleens of the inoculated mice by 7 days after subcutaneous (s.c.) inoculation . Mice immunized s.c . with doses ranging from 2 x 10(4) to 2 x 10(8) CFU of strain YS-1 were completely protected against challenge with 100 LD50 of the homologous, highly virulent strain Fujisawa-SmR 21 days postimmunization, and protective immunity conferred by immunization with 2 x 10(8) CFU of the strain lasted for as long as the 3 months of the observation period . In passive immunization experiments, sera collected from mice immunized with strain YS-1 at days 14 and 21 postimmunization provided protection against challenge with Fujisawa-SmR, whereas sera collected at days 4 and 7 did not . Furthermore, specific spleen cell responses to E . rhusiopathiae antigens were observed in mice immunized with strain YS-1, and cross-protection against the antigenically heterologous bacterium Listeria monocytogenes was observed at 7 days after immunization in the mice, suggesting that cell-mediated immunity had been induced . These results suggest that E . rhusiopathiae YS-1 may be a suitable choice for further studies of vaccine efficacy in swine.

Infect Immun, 1998 Jul, 66(7), 3128 - 33
Purification of the inlB gene product of Listeria monocytogenes and demonstration of its biological activity; Muller S et al.; Entry of Listeria monocytogenes into nonphagocytic cells requires the inlAB gene products . InlA and InlB are bacterial cell wall-associated polypeptides that can be released by sodium dodecyl sulfate treatment . By applying more gentle extraction methods, we have purified InlB in its native form . Treatment of bacteria with various nondenaturating agents including mutanolysin, thiol reagents, sodium chloride, and detergents like Triton X-100 or 3-{(3-cholamidopropyl)-dimethylammonio}-1-propanesulfonate did not release substantial amounts of InlB from the bacterial cell wall . Instead, InlB was nearly quantitatively extracted in a solubilized form by treatment of bacteria with 1 M Tris-Cl or other protonated amines at pH 7.5 . However, the reduced solubility of the extracted InlB in low-salt buffers hampered further biochemical purification . A panel of monoclonal antibodies against listerial Tris-Cl extracts containing InlB was therefore produced to generate reagents for use in affinity chromatography . One of the monoclonal antibodies enabled purification of the InlB protein to homogeneity with relatively high yields . When added externally, purified InlB associated with the surface of noninvasive bacteria such as Listeria innocua or an L . monocytogenes inlB2 mutant, where it promoted entry of these strains into Vero cells >300- and 17-fold, respectively . This effect was even more dramatic for HeLa cells, where the observed invasion was increased about 9,000- and 4,000-fold, respectively . The availability of purified native, invasion-competent InlB will allow analysis of the molecular basis of InlB-mediated entry into tissue culture cell lines in greater detail.

J Immunol, 1998 Jun 15, 160(12), 6056 - 61
IL-6 produced by Kupffer cells induces STAT protein activation in hepatocytes early during the course of systemic listerial infections; Gregory SH et al.; Kupffer cells were the principal source of IL-6 produced in the livers of mice following i.v . inoculation of Listeria monocytogenes . IL-6 mRNA expression and the production of IL-6 were reduced drastically within the nonparenchymal liver cell population derived from mice rendered Kupffer cell depleted by pretreatment with liposome-encapsulated dichloromethylene diphosphonate . A sharp increase in the appearance of activated STAT3 occurred in extracts of purified hepatocytes derived from normal mice infected i.v . with Listeria . Remarkably, the kinetics of this increase overlapped IL-6 mRNA expression by Kupffer cells; each peaked at approximately 30 min postinfection . No increase in STAT3 activation was observed in IL-6-deficient or Kupffer cell-depleted animals . The results of these experiments indicate that the synthesis of IL-6 and the activation of STAT3 within hepatocytes are critical functions of Kupffer cells occurring very early during the course of systemic listerial infections.

J Immunol, 1998 Jun 15, 160(12), 6046 - 55
Th1 cells specific for a secreted protein of Listeria monocytogenes are protective in vivo; Geginat G et al.; In the present study we have investigated the role of the secreted p60 protein from Listeria monocytogenes as an Ag for CD4 T cells . The p60 protein is an abundant extracellular protein that is highly conserved within the members of the genus Listeria . Our results show that L . monocytogenes infection induces a potent p60-specific Th1 immune response . Remarkably, we found that p60-specific Th1 clones mediate significant protection against L . monocytogenes infection . For one p60-specific clone, the peptide epitope was defined . This clone recognized p60 301-312 (EAAKPAPAPSTN) in the context of the H-2Ad molecule . Despite the fact that acquired immunity against L . monocytogenes is primarily mediated by cytotoxic CD8 T lymphocytes, our data clearly demonstrate that secreted bacterial proteins are important CD4 T cell Ags and that Th1 clones specific for a secreted bacterial protein can contribute to the protection against an intracellular pathogen such as L . monocytogenes.

FEBS Lett, 1998 May 15, 427(3), 353 - 6
Cdc42 is required for membrane dependent actin polymerization in vitro; Moreau V et al.; In vitro actin based motility assays with bacterial pathogens have provided powerful systems to both understand and dissect actin dynamics as well as cell motility . Taking advantage of endogenous membrane vesicles in Xenopus extracts we have developed an in vitro assay to study membrane dependent actin polymerization . Our results demonstrate that membrane dependent actin polymerization, in contrast to Listeria stimulated actin filament assembly, is dependent on small GTPases of the Rho family . Using a combination of depletion and reconstitution experiments we have shown that Cdc42 but not Rac or Rho is required to stimulate actin polymerization from membranes . The in vitro system we have described here is amenable to identification of the downstream effectors of Cdc42 required for membrane dependent actin polymerization.

Zentralbl Hyg Umweltmed, 1997 Aug, 200(2-3), 189 - 95
Nonhemolytic strains of Listeria monocytogenes detected in milk products using VIDAS immunoassay kit; Allerberger F et al.; In December 1995 detection of Listeria monocytogenes Sv 1/2a in milk products that were routinely sampled for investigation at the Austrian Federal Food Inspection Laboratory (Vienna) led the food manufacturer in question to withdraw his product from the market . While one of seven Listeria strains isolated from this food product using "VIDAS L . monocytogenes kit" was undoubtedly L . monocyotogenes, six strains were nonhemolytic . In classical bacteriology haemolysin is used as an important phenotypic property to differentiate L . monocytogenes from the apathogenic L . innocua species . Species identification by PCR and by Western blotting confirmed that all nonhemolytic strains were indeed L . monocytogenes . Our confirmation of nonhemolytic isolates as true L . monocytogenes strains underlines the considerable potential of the VIDAS system as a specific immunological easy-to-use and automated test kit for detection of L . monocytogenes in food products . Whether or not the presence of non-hemolytic Listeria monocytogenes in food products justifies legal actions should be addressed by the proper authorities.

J Appl Microbiol, 1998 Apr, 84(4), 642 - 8
Sensitivity to commercial disinfectants, and the occurrence of plasmids within various Listeria monocytogenes genotypes isolated from poultry products and the poultry processing environment; Earnshaw AM et al.; The European Suspension Test was used to assess the relative resistance of 19 individual Listeria monocytogenes genotypes, isolated from the poultry processing environment, to three commercially used disinfectants employed in the plant at the time of their isolation . To establish the relative resistance between the strains, the concentration of each disinfectant was reduced until inter-strain variation became apparent . For Darasan 214 and 7058, variation was detected at 0.1% and 0.5% v/v, respectively, while Daraclean 7361 had to be reduced to only 2.5% v/v . At these concentrations, the mean microbiocidal effect (ME) of each disinfectant ranged between 4.3 and 3.1 log10 reduction in cfu ml-1 . Significant differences between the strains were obtained with respect to their resistance to the disinfectants employed (P < 0.01), but the overall log10 reduction for genotypes 'A1' and 'A2', which were found to persist in the poultry processing environment, were not found to be significantly different from the genotypes which had been isolated on a more sporadic basis (P > 0.05) . The L . monocytogenes strains fell into four groups with respect to incidence and size of plasmids isolated . The first group contained strains which carried two plasmids (5 and 40 MDa) and the other three (groups 2, 3 and 4) comprised strains which carried a single plasmid (14, 47 and 52 MDa, respectively) . There was no correlation between persistent and sporadic strains with respect to incidence and size of plasmids isolated . Moreover, the strains which carried no plasmids were found to be as resistant to the disinfectants as those which did carry plasmids, suggesting that the plasmids isolated did not confer resistance of L . monocytogenes planktonic cells to the disinfectants tested . Therefore, it is unlikely that the strains which had been found to persist in the poultry processing environment did so by means of plasmid-mediated resistance to the commercial disinfectants used.

J Appl Microbiol, 1998 Feb, 84(2), 255 - 62
Polymorphism of Listeria monocytogenes and Listeria innocua strains isolated from short-ripened cheeses; Margolles A et al.; Thirty isolates of Listeria monocytogenes and 18 of L . innocua obtained from different short-ripened cheeses manufactured in Asturias (northern Spain), were compared with each other and with reference strains using serotype, phage type and pulsed-field restriction endonuclease digestion profiles analysis of the total DNA . Restriction enzymes ApaI and SmaI defined five clusters in L . monocytogenes (m1 to m5) and two main clusters in L . innocua (i1 and i2) . Cluster i2 was further arranged into three subclusters (i2a, i2b and i2c) based on the different Eco52I (XmaIII) and Crf42I (SacII) patterns of its isolates . Clusters of L . innocua were clearly different whereas those of L . monocytogenes were more closely related to each other . In this latter species, serotype 4b isolates (m4 and m5) constituted a more homogeneous group than serogroup I isolates (m1, m2 and m3) . Cluster m3 contained two strains of serotype 1/2a whereas m1 and m2 harboured strains of both serotypes, 1/2a and 1/2b . Therefore, the combined use of restriction patterns and serotype may be useful to differentiate L . monocytogenes strains showing identical restriction profiles but differing in serotype . The cheese source of Listeria strains proved that isolates from cluster m1 were repeatedly detected as a contaminant in the same type of cheese . Comparison of L . monocytogenes ApaI profiles showed a genetic proximity of m4 and m5 to the recognized pathogenic strains ATCC 13932 and NCTC 11994, responsible for meningitis cases in other countries . Finally, bacteriophage typing data indicated that m4, the sole phage typable group, had a phage type resembling that of strains causing the Auckland (New Zealand) outbreak of listeriosis in 1969 . These data suggest a wide distribution of closely related types which might cause, under several circumstances, sporadic cases of listeriosis.

J Appl Microbiol, 1998 Feb, 84(2), 234 - 9
Heat resistance of Listeria monocytogenes in dairy products as affected by the growth medium; Casadei MA et al.; Listeria monocytogenes strains 1151 and Scott A were grown in broth at 30 degrees C and transferred to half cream, double cream and butter stored at 5 degrees C to determine the influence of dairy product composition on heat resistance at 52, 56, 60, 64 and 68 degrees C . Strain 1151 showed a higher heat resistance than strain Scott A . The heat resistance of both strains was higher in the dairy products than in broth, particularly at lower temperatures . A significant difference was observed between log 10 of the D-values in the different dairy products . The D-values obtained for both strains resuspended in all the dairy products would result in efficient elimination of the pathogen at 72.7 degrees C for 15 s . The highest D-value was 11.30 s at 68 degrees C and by using a z-value of 6.71 degrees C it can be determined that at 72.7 degrees C the D-value would be 1.5 s . The 15 s process would therefore achieve 10 log reductions . The effect of growth conditions on the heat resistance at 60 degrees C of L . monocytogenes Scott A was also investigated . When the cells were grown in the diary products themselves, and particularly butter, the heat resistance of Scott A was enhanced; for example, the D-values were 7.15 times higher than in broth . Further studies are required to investigate if this protection against heating exists at higher temperatures, in which case the efficiency of pasteurization treatments or other heat treatments would be considerably lowered.

J Appl Microbiol, 1998 Feb, 84(2), 185 - 91
Mathematical modelling of the heat resistance of Listeria monocytogenes; Augustin JC et al.; The heat resistance of Listeria monocytogenes phagovar 2389/2425/3274/2671/47/108/340 (1992 French outbreak strain) in broth was studied at 55, 60 and 65 degrees C . Experiments were carried out on bacterial cultures in three different physiological states: cultures at the end of the log phase, cultures heat-shocked at 42 degrees C for 1 h, and subcultures of cells resistant to prolonged heating . Survivor curves were better fitted using a sigmoidal equation than the classical log-linear model . This approach was justified by the existence of heat resistance distributions within the bacterial populations . Peaks (log10 of heating time) of heat resistance distributions of untreated, heat-shocked, and selected cultures at 55, 60 and 65 degrees C were 0.34, -0.90 and -1.84 min, 0.74, -0.51 and -1.24 min, and 0.17, -0.94 and -1.45 min, respectively . The widths of the distributions are proportional to 0.29, 0.36 and 0.41 min0.5, 0.26, 0.36 and 0.41 min0.5, and 0.34, 0.44 and 0.41 min0.5 . An increase in the thermal tolerance could then be induced by sublethal heat shock or by selection of heat resistant cells.

Lett Appl Microbiol, 1998 Apr, 26(4), 305 - 10
Alkaline phosphatase release assay to determine cytotoxicity for Listeria species; Bhunia AK et al.; A simple cytotoxicity assay for Listeria species was developed by assaying alkaline phosphatase (AP) release from an infected hybrid B lymphocyte (Ped-2E9) line . Eight of eight L . monocytogenes and six of 11 L . ivanovii strains induced significantly high AP release from Ped-2E9 cells compared to five other L . ivanovii strains and other Listeria spp . In contrast, all L . monocytogenes and L . ivanovii test strains showed high release of lactate dehydrogenase (LDH) activity from Ped-2E9 cells . The molecular mass of AP was estimated to be about 128-165 kDa, suggesting severe membrane damage in Ped-2E9 cells due to Listeria infection . The data presented here indicate that AP assay could be used over LDH assay to detect Listeria-induced cell cytotoxicity.

Mol Microbiol, 1998 May, 28(3), 403 - 12
Metal ion homeostasis and intracellular parasitism; Agranoff DD et al.; Bacteria possess multiple mechanisms for the transport of metal ions . While many of these systems may have evolved in the first instance to resist the detrimental effects of toxic environmental heavy metals, they have since become adapted to a variety of important homeostatic functions . The 'P'-type ATPases play a key role in metal ion transport in bacteria . A Cu+-ATPase from the intracellular bacterium Listeria monocytogenes is implicated in pathogenesis, and similar pumps in Mycobacterium tuberculosis and M . leprae may play a comparable role . Intracellular bacteria require transition metal cations for the synthesis of superoxide dismutases and catalases, which constitute an important line of defence against macrophage-killing mechanisms . The macrophage protein Nramp1, which confers resistance to a variety of intracellular pathogens, has also been shown recently to be a divalent amphoteric cation transporter . Mycobacterial homologues have recently been identified by genomic analysis . These findings suggest a model in which competition for divalent cations plays a pivotal role in the interaction between host and parasite.

Cell Immunol, 1998 Mar 15, 184(2), 92 - 104
Long-lived protective immunity to Listeria is conferred by immunization with particulate or soluble listerial antigen preparations coadministered with IL-12; Miller MA et al.; The ability of IL-12 to promote the development of Th1-type immune responses, and thus promote cellular immunity, has been well documented . In a previous report, we showed that coadministration of IL-12 with heat-killed Listeria monocytogenes elicited intense antigen-specific T cell responses that conferred protective listerial immunity . Herein, we have extended those studies by demonstrating that multiple injections of heat-killed L . monocytogenes and IL-12 elicit memory responses that confer long-lived (> or = 3 months) protective immunity and that immunity can be transferred adoptively with cells from immunized mice injected into naive mice . These studies have also demonstrated that the powerful adjuvanticity of IL-12 is observed with soluble as well as particulate immunogens and is operative in mouse strains that have different MHC haplotypes . These findings suggest that IL-12 may be a useful adjuvant component of vaccines for a wide variety of pathogens in animal and human systems.

J Neuroimmunol, 1998 May 1, 85(1), 33 - 43
Chemokines and chemotaxis of leukocytes in infectious meningitis; Lahrtz F et al.; Chemokines constitute a constantly growing family of small inflammatory cytokines . They have been implied in many different diseases of the CNS including trauma, stroke and inflammation, e.g., multiple sclerosis . In this review we focus on the role of chemokines in infectious meningitis of bacterial or viral origin . In experimental bacterial meningitis induced by Listeria monocytogeneses both CXC and CC chemokines namely MIP-1alpha, MIP-1beta and MIP-2 are produced intrathecally by meningeal macrophages and leukocytes which infiltrate into the CNS . In patients with bacterial meningitis, IL-8, GROalpha, MCP-1, MIP-1alpha and MIP-1beta are detectable in the CSF . These chemokines contribute to CSF mediated chemotaxis on neutrophils and PBMC in vitro . In viral meningitis IL-8, IP-10 and MCP-1 are identified in the CSF to be responsible for chemotactic activity on neutrophils, PBMC and activated T cells . Taken collectively these data indicate that the recruitment of leukocytes in infectious meningitis involves the intrathecal production of chemokines.

FEMS Immunol Med Microbiol, 1998 Apr, 20(4), 289 - 99
Suppression of major histocompatibility complex class I and class II gene expression in Listeria monocytogenes-infected murine macrophages; Schuller S et al.; Macrophage cells play a central role during infection with Listeria monocytogenes by both providing a major habitat for bacterial multiplication and presenting bacterial antigens to the immune system . In this study, we investigated the influence of L . monocytogenes infection on the expression of MHC class I and class II genes in two murine macrophage cell lines . Steady-state levels of I-Abeta chain mRNA were decreased in both resting J774A.1 and P388D1 macrophages infected with L . monocytogenes whereas reduction of H-2K mRNA was only observed in P388D1 cells . In addition, L . monocytogenes suppressed induction of MHC class I and class II mRNAs in response to gamma-interferon as well as the maintenance of the induced state in activated P388D1 macrophages . Exposure to the non-pathogenic species L . innocua or a deletion mutant of L . monocytogenes, which lacks the lecithinase operon, did not cause a reduction in H-2K and I-Abeta mRNA levels nor suppress expression of Ia antigens . Inhibition of MHC gene expression may represent an important part of the cross-talk between L . monocytogenes and the macrophage that probably influences the efficiency of a T cell-mediated immune response and thus the outcome of a listerial infection.

J Exp Med, 1998 May 18, 187(10), 1711 - 9
H2-M3 restricted presentation of a Listeria-derived leader peptide; Princiotta MF et al.; Protective immunity to infection by many intracellular pathogens requires recognition by cytotoxic T lymphocytes (CTLs) of antigens presented on major histocompatibility complex (MHC) class I molecules . To be presented for recognition by pathogen-specific CTLs, these antigens must gain access to the host cell class I processing pathway . In the case of intracellular bacterial pathogens, the majority of bacterial proteins are retained within the bacterial membrane and therefore remain inaccessible to the host cell for antigen processing . We have isolated a CTL clone from a C57BL/6 mouse infected with the intracellular gram-positive bacterium Listeria monocytogenes (LM) and have identified the source of the antigen . Using a genomic expression library, we determined that the clone recognizes an antigenic N-formyl peptide presented by the nonpolymorphic murine MHC class Ib molecule, H2-M3 . Several lengths of this peptide were able to sensitize cells for lysis by this CTL clone . The source of this antigenic peptide is a 23-amino acid polypeptide encoded at the start of a polycistronic region . Analysis of mRNA secondary structure of this region suggests that this polypeptide may be a leader peptide encoded by a transcriptional attenuator.

Microbiol Immunol, 1998, 42(4), 325 - 7
Systemic dissemination by intrarectal infection with Listeria monocytogenes in mice; Nishikawa S et al.; Orally ingested Listeria monocytogenes is known to penetrate into Peyer's patches (PP) and translocate to the spleen and liver . Herein, extraintestinal dissemination of the bacterium independent of PP was investigated . Dissemination of Listeriae to the spleen and liver was observed in intrarectally infected mice as well as in intragastrically infected animals in spite that no Listeriae were detected in the small intestines of mice infected intrarectally . Decreased numbers of intestinal intraepithelial lymphocytes (iIEL) and increased numbers of lymphocytes in the contents of the small and large intestines were observed after intragastric infection and in the large intestine after intrarectal infection, giving the assumption that the leakage of iIEL caused by injury of epithelial layers in intestines might occur during infection . These results suggest that L . monocytogenes might be able to disseminate through small and large intestines in part by a PP-independent mechanism.

Am J Vet Res, 1998 Jun, 59(6), 717 - 21
Induction of cytokine messenger RNA transcripts in mouse macrophages by Listeria monocytogenes isolated from channel catfish; Zhang LF et al.; OBJECTIVE: To determine whether differences exist in induction and quantity of tumor necrosis factor alpha (TNF-alpha), interleukin (IL)1 beta, and IL-10 mRNA transcripts when mouse J774A.1 macrophages are infected with Listeria monocytogenes, including 2 isolates originating from channel catfish, the wild-type virulent (EGD) strain, and a nonhemolytic strain (ATCC 15313) . SAMPLES: Listeria monocytogenes isolates from kidneys or fillets of channel catfish were used to stimulate cytokine production from mouse macrophages . The RNA from the infected macrophages was collected . PROCEDURE: Four hours after infection with L monocytogenes, total cellular RNA was extracted from the J774A.1 cells and reversed transcribed to cDNA, which was amplified, using specific primers for TNF-alpha, IL-1 beta, or IL-10 . The specific amplified DNA fragments were detected on polyacrylamide gels and quantified, using a reverse transcription polymerase chain reaction (PCR)-mediated ELISA . RESULTS: The wild-type hemolytic EGD strain and the 2 hemolytic catfish isolates of L monocytogenes induced higher amounts of TNF-alpha-, IL-1 beta-, and IL-10-specific mRNA in J774A.1 cells than did the nonhemolytic strain . CONCLUSIONS: Hemolysin-associated induction of TNF-alpha, IL-1 beta, and IL-10 cytokines may be related to survival and replication of L monocytogenes in macrophages . It also suggests that the PCR-mediated ELISA procedure is a sensitive test to quantify cytokines from cell cultures.

Int J Food Microbiol, 1998 Apr 14, 40(3), 185 - 95
Model of the influence of time and mild temperature on Listeria monocytogenes nonlinear survival curves; Breand S et al.; Heat treatment has long been regarded as one of the most widely used and most effective means of destroying pathogens in food . Up to now the linear relationship between the death rate and the temperature has been used when choosing the best heat treatment to apply . However, the information given by this linear relationship is no longer sufficient when nonlinear survival curves are observed . Consequently, the agri-food industry needs a tool to choose the best mild heat treatment to apply in the case of nonlinear survival curves . This study deals with the temperature-induced death of Listeria monocytogenes CIP 7831 in the stationary phase of growth . Eleven temperatures were tested . With the proposed primary and secondary models good fits of our data were obtained . A model describing both the effect of the duration of treatment and the temperature on the logarithm of the number of survivors was then built . A clear increase in the precision of the estimation of the parameters was obtained with this model . Moreover, with this model a new graphical strategy to choose a mild heat increase regarding a maximal survivor number has been proposed.

Int J Food Microbiol, 1998 Apr 14, 40(3), 149 - 57
Prediction of Listeria spp . growth as affected by various levels of chemicals, pH, temperature and storage time in a model broth; Razavilar V et al.; The effects of concentration of NaCl (0.5 to 12.5%), methyl paraben (0.0 to 0.2%), sodium propionate (0.3%), sodium benzoate (0.1%), potassium sorbate (0.3%), pH (> 5.9) temperature (4 to 30 degrees C), storage time (up to 58 d) and inoculum (> 10(5) to > 10(-2) per ml) on the log10 probability percentage of one cell of Listeria spp . to initiate growth in a broth system were evaluated in a factorial design study . At pH 5.96 and temperature ranging from 4 to 30 degrees C the concentrations of sodium propionate, potassium sorbate, and sodium benzoate examined allowed growth of L . monocytogenes with lag phases at 4 degrees C of 18, 27 and 21 days, respectively . For 0.1 and 0.2% methyl paraben growth of all Listeria spp . was initiated at 8 degrees C and 30 degrees C, respectively . At pH 6, concentration of 12% NaCl supported the growth of L . monocytogenes at 8 to 30 degrees C, whereas 12.5% inhibited all Listeria species . Four regression equations were derived relating probability of growth initiation to temperature, concentrations of NaCl and preservatives storage time, and Listeria species specific effects . From these equations, the number of cells needed for growth initiation can be calculated . The impact of this type of quantitative study and its possible application on the development of microbial standards for foods is discussed.

Regul Toxicol Pharmacol, 1998 Feb, 27(1 Pt 2), S40 - 54
Effects of Great Lakes fish consumption on the immune system of Sprague-Dawley rats investigated during a two-generation reproductive study; Tryphonas H et al.; The effects of Great Lakes fish contaminants on several quantitative and functional aspects of the immune system were investigated in the first (F1) and second (F2) generations of Sprague-Dawley rats . The F0 rats were fed either a control diet or diets containing 5 or 20% lyophilized chinook salmon from the Credit River of Lake Ontario (LO) and Owen Sound point of Lake Huron (LH) . The F1 and F2 pups were exposed to fish in utero, through the dam's milk to 21 days old, and through the dam's respective diets to 13 weeks of age . The study included an F1-reversibility (F1-R) phase in which rats at 13 weeks of exposure to fish or control diets were switched to the control diet for 3 months . The most outstanding finding was a statistically significant increase in absolute spleen leukocytes and absolute and percentage lymphocytes in the F2 male rats fed the LH fish diets compared to the control and to those fed the LO fish diets with the 20% fish diets having higher cell numbers compared to the LO-5% fish diets . A parallel increase in the T-helper/inducer T-lymphocyte subset numbers was observed . Increased but statistically insignificant plaque-forming cell (PFC) numbers were obtained in the F2 male rats fed the LH fish diets compared to those fed the LO fish diets and in the F1-R female group of rats fed the LH fish diet compared to those fed the LO fish diets . Phagocytosis by resident peritoneal macrophages was significantly increased in the F1 male and F2 female rats fed the fish diets compared to the control . The phagocytic activity was significantly higher in the F2-generation male and female rats fed the LO diets compared to those fed the LH diets . Other parameters including lymphocyte transformation in response to mitogens, the number of Listeria monocytogenes bacteria surviving in the rat spleens, and the natural killer cell activity were not affected significantly by any of the treatments . Overall, the effects of diets containing chinook salmon from the LO and LH sources on the immune system of rats were minimal and were on quantitative rather than on functional aspects of the system . Further focused research would be required in order to establish conclusively that the immune system of cohorts who ingest Great Lakes fish frequently is at a greater risk for adverse effects .

J Reprod Immunol, 1998 Apr, 38(1), 31 - 53
Docosahexaenoic acid, a constituent of fetal and neonatal serum, inhibits nitric oxide production by murine macrophages stimulated by IFN gamma plus LPS, or by IFN gamma plus Listeria monocytogenes; Lu CY et al.; Murine macrophage activation is deficient in the fetus and the neonate, and in areas of the placenta perfused by the fetal circulation . Fetal and neonatal serum concentrations of docosahexaenoic acid (DHA) are 150 microM, or approximately 50-fold higher than in the adult . We previously showed that DHA inhibits activation of the gene for inducible nitric oxide synthase (iNOS) in murine macrophages stimulated in vitro with interferon gamma (IFN gamma) plus lipopolysaccharide (LPS) . We have now pursued these observations in greater depth . An assay system was developed which separated the stimulation of macrophages by IFN gamma plus LPS, and the actual production of nitric oxide (NO) . It was found that macrophages do not produce NO until they have been stimulated by IFN gamma plus LPS for a period of 10 h . NO is produced during the subsequent 10 h, even though IFN gamma plus LPS are not longer present . DHA, if present, inhibited only during the initial 10 h stimulation; DHA did not inhibit the production of NO by macrophages which had previously been stimulated by IFN gamma plus LPS, and were already producing NO . It was also found that DHA was less inhibitory if given prior to the IFN gamma plus LPS stimulation . In a dose-responsive manner, DHA inhibited the increased abundance of iNOS mRNA by macrophages stimulated by IFN gamma plus LPS . NO contributes to the host defense against Listeria monocytogenes and other intracellular pathogens . We therefore investigated the ability of DHA to inhibit NO production by macrophages stimulated by IFN gamma plus Listeria monocytogenes in vitro; DHA inhibited transcription of the iNOS gene and also the listeriocidal activity of activated macrophages . Inhibition of NO production by DHA may contribute to the increased susceptibility of the fetoplacental unit and neonate to intracellular infections.

Proc Natl Acad Sci U S A, 1998 Apr 28, 95(9), 5299 - 304
Mycobacterium bovis Bacille Calmette-Guérin strains secreting listeriolysin of Listeria monocytogenes; Hess J et al.; Recombinant (r) Mycobacterium bovis strains were constructed that secrete biologically active listeriolysin (Hly) fusion protein of Listeria monocytogenes . The r-BCG strains pAT261:Hly or pMV306:Hly expressed plasmid multicopies or chromosomal single copies of the hly gene, respectively . Human and murine macrophage-like cell lines were infected with r-BCG pAT261:Hly and pMV306:Hly strains . Interestingly, intracellular persistence of both r-BCG strains was reduced in macrophages as compared with the parental BCG strain . By immunogold labeling Hly was detected in membrane structures and within the phagosomal space of macrophages . In addition, Hly was localized within cytoplasmic vacuoles outside the mycobacteria-containing phagosome of host cells infected with r-BCG pAT261:Hly or r-BCG pMV306:Hly . Hly fusions consistently colocalized with a lysosome-associated membrane glycoprotein, suggesting that membrane-attack conformation of Hly was not altered . Although r-BCG pAT261:Hly and r-BCG pMV306:Hly microorganims apparently did not egress into the cytoplasmic compartment of host cells, they both improved major histocompatibility complex class I presentation of cophagocytosed soluble protein as compared with wild-type BCG microbes . These data suggest that Hly secretion endows BCG with an improved capacity to stimulate CD8 T cells . Because CD8 T cells play a major role in protection against tuberculosis such Hly secreting r-BCG constructs are antituberculosis vaccine candidates.

Klin Monatsbl Augenheilkd, 1998 Jan, 212(1), 55 - 8
{Listeria monocytogenes-induced endogenous endophthalmitis in an otherwise healthy patient: PCR-assisted rapid diagnosis as the basis for successful therapy}; Lohmann CP et al.; BACKGROUND: Listeria monocytogenes is a rare cause of endogenous endophthalmitis . Only 14 cases are published in the literature so far . All eyes showed similar clinical features and profound visual loss . PATIENTS AND METHODS: We report on a case of an otherwise healthy 73-year-old male . He was referred to our hospital because of acute hypopyoniritis with secondary glaucoma . Within a few hours the severity of the intraocular infection increased dramatically resulting in the clinical picture of an acute endophthalmitis . RESULTS AND CONCLUSION: Early identification of the causative pathogen in the aqueous humor after anterior chamber paracenthesis using polymerase chain reaction (PCR) and the initiation of a specific, systemic antibiotic medication resulted in a complete recovery of visual acuity.

Int J Biochem Cell Biol, 1998 Mar, 30(3), 307 - 11
The focal adhesion phosphoprotein, VASP; Holt MR et al.; Vasodilator-stimulated phosphoprotein (VASP) is associated with focal adhesions and areas of dynamic membrane activity, where it is thought to have an important role in actin filament assembly and cell motility . VASP contains a central proline-rich sequence which recruits the G-actin binding protein profilin . Localization of VASP to the leading edge of a migrating cell can lead to local accumulation of profilin, which in turn can supply actin monomers to growing filament ends . VASP binds to the focal adhesion proteins vinculin and zyxin and this probably directs the phosphoprotein to focal adhesions and the leading edge of stimulated cells . VASP functions as a binding intermediate between profilin and focal adhesion proteins . Intracellular pathogens, including Listeria monocytogenes, have coat proteins which bind VASP . This is one way in which these pathogens use VASP, and other proteins from the host cell, to assemble the actin filaments they require to move around the cytoplasm of infected cells and enter neighbouring cells . Understanding the role of VASP and other proteins in cell and bacterial motility is likely to lead to development of new therapeutic strategies for diseases including atherosclerosis and tumour growth, and for limiting the spread of intracellular pathogens.

Rev Soc Bras Med Trop, 1998 Mar-Apr, 31(2), 173 - 7
{Listeria monocytogenes meningitis . Case reports in patients from the Federal District}; Hofer E et al.; It has been shown the role of Listeria monocytogenes as a etiological agent identified by bacteriological analysis among cases of human meningitis in Distrito Federal, Brazil . Laboratorial characteristics and some clinical and epidemiological aspects are reported.

Appl Environ Microbiol, 1998 Jun, 64(6), 2065 - 71
Effects of above-optimum growth temperature and cell morphology on thermotolerance of Listeria monocytogenes cells suspended in bovine milk; Rowan NJ et al.; The thermotolerances of two different cell forms of Listeria monocytogenes (serotype 4b) grown at 37 and 42.8 degrees C in commercially pasteurized and laboratory-tyndallized whole milk (WM) were investigated . Test strains, after growth at 37 or 42.8 degreesC, were suspended in WM at concentrations of approximately 1.5 x 10(8) to 3.0 x 10(8) cells/ml and were then heated at 56, 60, and 63 degrees C for various exposure times . Survival was determined by enumeration on tryptone-soya-yeast extract agar and Listeria selective agar, and D values (decimal reduction times) and Z values (numbers of degrees Celsius required to cause a 10-fold change in the D value) were calculated . Higher average recovery and higher D values (i.e., seen as a 2.5- to 3-fold increase in thermotolerance) were obtained when cells were grown at 42.8 degrees C prior to heat treatment . A relationship was observed between thermotolerance and cell morphology of L . monocytogenes . Atypical Listeria cell types (consisting predominantly of long cell chains measuring up to 60 micron in length) associated with rough (R) culture variants were shown to be 1.2-fold more thermotolerant than the typical dispersed cell form associated with normal smooth (S) cultures (P </= 0.001) . The thermal death-time (TDT) curves of R-cell forms contained a tail section in addition to the shoulder section characteristic of TDT curves of normal single to paired cells (i.e., S form) . The factors shown to influence the thermoresistance of suspended Listeria cells (P </= 0.001) were as follows: growth and heating temperatures, type of plating medium, recovery method, and cell morphology . Regression analysis of nonlinear data can underestimate survival of L . monocytogenes; the end point recovery method was shown to be a better method for determining thermotolerance because it takes both shoulders and tails into consideration . Despite their enhanced heat resistance, atypical R-cell forms of L . monocytogenes were unable to survive the low-temperature, long-time pasteurization process when freely suspended and heated in WM.

Med Pediatr Oncol, 1998 Jul, 31(1), 19 - 21
Brain abscesses in children with cancer; Antunes NL et al.; BACKGROUND: Brain abscesses in pediatric patients are rare events, and the causative organism and prognosis vary with the population under study . Children with cancer seem to be particularly susceptible to the development of brain abscesses because of the immunological changes induced by cancer and its treatment . We reviewed the records of children who developed a brain abscess during treatment of a malignancy to define the clinical characteristics, prognosis, and management of these patients . PROCEDURE: We performed a retrospective review of the clinical and laboratory characteristics of all cancer patients younger than age 20 years who were admitted to our institution between 1980 and 1996 for a brain abscess . RESULTS: Twelve children were identified . Cancer diagnoses were brain tumor in two, systemic PNET in two, and leukemia in eight . Six patients had multiple abscesses . Eleven received prior chemotherapy . Abscesses were surgically excised or aspirated in seven, and empiric antibiotics were given to the other five . At surgery, Listeria monocytogenes, Aspergillus fumigatus (3), Fusarium, and Candida lusitanea were cultured . Aspergillus was identified in other locations in four patients . Abscesses were successfully treated in seven patients, two of whom received antibiotics only; five patients (42%) died from infection . CONCLUSIONS: Mortality is high in this immunosuppressed population, in part due to the preponderance of fungal infection . The finding of very rare organisms suggests that drainage and culture should be performed whenever possible; empiric antibiotics that include an antifungal agent may, on occasion, be successful.

Biochemistry (Mosc), 1998 Feb, 63(2), 191 - 4
Teichoic acids of the cell wall of Nocardiopsis listeri, Nocardiopsis lucentensis, and Nocardiopsis tregalosei; Streshinskaya GM et al.; The structures of teichoic acids of three Nocardiopsis species were established . The cell wall of Nocardiopsis listeri VKM Ac-1881T contains two teichoic acids (TA) . TA1 is 1,3-poly(glycerol phosphate) with 50% glycerol phosphate residues substituted by alpha-N-acetylglucosamine in the C-2 position . TA2 is 1, 5-poly(ribitol phosphate) with each ribitol phosphate unit carrying pyruvate acetal groups in the 2 and 4 positions . The cell wall of Nocardiopsis lucentensis VKM Ac-1963T contains only one teichoic acid of the same structure as TA1 of N . listeri . The teichoic acid of the Nocardiopsis tregalosei VKM Ac-942 is a 1,3-poly(glycerol phosphate) substituted (60%) by beta-glucopyranosyl residues . Structures of polymers were studied by chemical and NMR spectroscopy methods . The presented results confirm the species-specificity of teichoic acids from Nocardiopsis genus.

Scand J Gastroenterol, 1998 Apr, 33(4), 430 - 4
Listeria monocytogenes in Crohn's disease; Chiba M et al.; BACKGROUND: In an immunohistochemical study a higher rate of reactivity of intestinal tissues to the antibody against Listeria monocytogenes was reported in Crohn's disease as compared with controls . METHODS: Seventy-six intestinal tissues, either therapeutically resected or biopsied, from 31 patients with Crohn's disease, 20 with ulcerative colitis, and 21 with non-inflammatory bowel disease were studied . DNA extracted from intestinal tissues by proteinase K treatment was used for nested polymerase chain reaction (PCR), using two sets of primers . PCR products were analyzed with agarose gel electrophoresis and subsequent Southern blot analysis . RESULTS: Our amplification system could detect 9 pg of L . monocytogenes DNA . L . monocytogenes was detected in only one sample, that from a patient with ulcerative colitis . CONCLUSIONS: Our study does not support the etiologic significance of L . monocytogenes in Crohn's disease.

Infect Immun, 1998 Jun, 66(6), 2814 - 7
Listeria monocytogenes-infected hepatocytes are targets of major histocompatibility complex class Ib-restricted antilisterial cytotoxic T lymphocytes; Bouwer HG et al.; Subclinical infection of BALB/c mice with the intracellular bacterial pathogen Listeria monocytogenes results in the development of protective antilisterial immunity . L . monocytogenes can infect hepatocytes, and antilisterial cytotoxic T lymphocytes (CTL) lyse Listeria-infected hepatocytes in a major histocompatibility complex (MHC) class Ia-restricted manner . It remained to be determined whether L . monocytogenes-infected hepatocytes are susceptible to MHC class Ib-restricted cytolysis . In this study, we showed that hepatocytes express MHC class Ib molecule Qa-1(b) mRNA and protein . We further showed that Listeria-infected hepatocytes are susceptible to MHC class Ib-restricted cytolysis, since C57BL/6-derived Listeria-infected hepatocytes were lysed by BALB/c-derived antilisterial CTL . These results establish that Listeria-infected hepatocytes are susceptible to cytolysis by MHC class Ib restricted Listeria-specific CTL.

Int J Food Microbiol, 1998 Mar 3, 40(1-2), 133 - 7
Production of bacteriocin-like-substance by Listeria innocua against Listeria monocytogenes; Yokoyama E et al.; Cultures and culture filtrates of 129 Listeria innocua strains were examined for inhibitory activity against 18 strains of Listeria monocytogenes . Of the strains examined, 114 (88.4%) cultures and 126 (97.7%) culture filtrates had an inhibitory activity against strains of L . monocytogenes and most filtrates were sensitive to trypsin treatment . The authors concluded that most L . innocua strains produce a trypsin sensitive bacteriocin-like substance against L . monocytogenes.

Int J Food Microbiol, 1998 Mar 3, 40(1-2), 105 - 15
Predicted and observed growth of Listeria monocytogenes in seafood challenge tests and in naturally contaminated cold-smoked salmon; Dalgaard P et al.; The performance of the Pathogen Modelling Program, the Food MicroModel, the Murphy-model and the Ross-model for growth of L . monocytogenes was evaluated by comparison with data from 100 seafood challenge tests and data from 13 storage trials with naturally contaminated sliced vacuum-packed cold-smoked salmon . Challenge tests with both cured and noncured products were studied, and graphs as well as the bias- and the accuracy factors were used for comparison of the observed and predicted growth . The Pathogen Modelling Program could not be successfully validated in seafood challenge tests . Growth rates were markedly overestimated and the mu(max)-bias factor was as high as 3.9 in challenge tests with cured products . On the basis of the effect of temperature, NaCl/a(w) and pH, the mu(max)-bias factor of the other three models studied, varied between 1.0 and 2.3 in the challenge tests with cured and noncured seafoods . None of the models accurately predicted the growth in both cured and noncured seafoods . However, the results indicated that a new expanded model, including the additional effect of lactate and phenol, may provide accurate predictions of the growth of L . monocytogenes in challenge tests with various types of seafoods . Storage trials clearly showed the growth of L . monocytogenes in naturally contaminated cold-smoked salmon to be markedly slower than growth in inoculated challenge tests . Consequently, all four models substantially overestimated growth in the naturally contaminated products . Temperature, pH, NaCl/a(w) and lactate were measured in the storage trials and on the basis of these parameters, the Food MicroModel mu(max)-bias factor was 5.2 . Clearly, the model could not be successfully validated with naturally contaminated cold-smoked salmon . To improve the applicability of predictive models to fish products, it is suggested to include studies with naturally contaminated products in the development and validation of models with seafood pathogens.

Int J Food Microbiol, 1998 Mar 3, 40(1-2), 77 - 85
Enhanced detection and enumeration of Listeria monocytogenes from foodstuffs and food-processing environments; Johansson T; Listeria monocytogenes blood agar (LMBA) was compared to Listeria selective agar based on lithium chloride and ceftazidime (LA) and to the Oxford and Palcam media recommended by ISO and IDF for the detection and enumeration of L . monocytogenes from foodstuffs and food-processing environments . LMBA is based on trypticase soy agar with the following additions: sheep blood (5%) and as selective agents lithium chloride (10 g/l), polymyxin B sulphate (10 mg/l) and ceftazidime (20 mg/l), whereas the selectivity of LA is based on lithium chloride (15 g/l) and ceftazidime (15 g/l) . The media were compared in the detection of L . monocytogenes after enrichment from naturally contaminated foodstuffs (n = 423) and from food-processing environments (n = 93), and in the enumeration of the species from naturally contaminated foodstuffs (n = 287) . LMBA was superior both to the standard media and to LA in detection after enrichment and also in enumeration, except in the case of fresh broiler cut samples . The overall sensitivities of the Palcam, Oxford, LA and LMBA media were 68%, 67%, 74% and 96% in detection after enrichment and 64%, 73%, 76% and 80% in the enumeration of the species from ready to eat foods . The superiority of LMBA is based on distinguishing L . monocytogenes from other Listeria species by detection of beta-hemolysis, whereas the other media gave false-negative results because of the overgrowth of Listeria spp . other than L . monocytogenes, especially in detection after enrichment . A more selective medium than LMBA would have been required for the enumeration of the species from samples with high levels of competitive bacteria other than Listeria spp . The results indicate the need for a more specific isolation medium for L . monocytogenes in addition to those recommended by ISO and IDF for both detection and enumeration . LMBA offers an alternative to be used in combination with either Palcam or Oxford as well as with LA.

Int J Food Microbiol, 1998 Mar 3, 40(1-2), 65 - 75
The combined effect of nisin, leucocin F10, pH, NaCl and EDTA on the survival of Listeria monocytogenes in broth; Parente E et al.; The combined effect of the bacteriocins nisin (1-2100 IU/ml) and leucocin F10 (1-2100 AU/ml), pH (4.7-6.5), NaCl (0.7-4.5% w/l), ethylene diaminetetraacetic acid disodium salt (EDTA, 0.08-4.72 mmol/l) and inoculum level (10(3)-10(8) cfu/ml) on the survival of a pool of three strains of Listeria monocytogenes in broth was evaluated in three factorial experiments . Several factor combinations were found to prevent growth . Logistic regression analysis of the categorical data (survival/no survival) was used to generate predictive models for the probability of survival in 0.01 ml (P0.01) or 1 ml (P1) . Predicted and observed probabilities of survival were not significantly different in 72% and 68.9% of treatments for P0.01 and P1, respectively . Unsafe predictions were obtained in 9.4% and 14.8% of treatments for P0.01 and P1, respectively . Nisin had a major effect on the probability of survival but the addition of leucocin F10 was necessary to prevent the survival of L . monocytogenes . Lower pH values significantly decreased the probability of survival, while NaCl and EDTA had only a minor effect . Doses of bacteriocins > 250 AU/ml, pH < 5.6 and EDTA > 0.2 mmol/l (0.074 g/l) were needed to reliably prevent survival of Listeria monocytogenes.

Int J Food Microbiol, 1998 Mar 3, 40(1-2), 35 - 42
Synergistic effect of nisin and the lactoperoxidase system on Listeria monocytogenes in skim milk; Zapico P et al.; Nisin added at 10 or 100 IU/ml to ultra-high temperature processed (UHT) skim milk had no effect on counts of Listeria monocytogenes after 24 h at 30 degrees C, whereas addition of the lactoperoxidase system (LPS) resulted in counts of viable cells three log units lower than those of control milk after 24 h at 30 degrees C . Addition of nisin and LPS showed a synergistic effect and resulted in counts up to 5.6 log units lower than the control milk . When the two preservatives were added to actively growing cells of L . monocytogenes in two steps with a 2 h interval, their synergistic effect was enhanced . Counts of L . monocytogenes Ohio after 24 h at 30 degrees C in milk with nisin and LPS added together after 3 h of growth were 5.7 log units lower than the control milk . The difference in counts increased to 7.4 log units if LPS was added after 3 h and nisin after 5 h of growth . Similar but less pronounced effects were observed for the more resistant strain L . monocytogenes Scott A.

Scand J Immunol, 1998 Apr, 47(4), 369 - 74
Endogenous interleukin-4 does not suppress the resistance against a primary or a secondary Listeria monocytogenes infection in mice; Samsom JN et al.; Interleukin-4 (IL-4), a cytokine produced by T-helper 2 (Th2) cells, can inhibit the development of T-helper 1 (Th1) cells, which results in a decreased release of cytokines by the latter . As interferon-gamma (IFN-gamma), produced by Th1 cells, is involved in the resistance against a Listeria monocytogenes infection, the role of endogenously formed IL-4 during a Listeria infection in mice was investigated . Neutralization of endogenously formed cytokines by subcutaneously injected alginate-encapsulated monoclonal antibody (MoAb)-forming cells results in high antibody titres in the circulation over a long time period . The aim of the present study was to reevaluate the effect of neutralization of IL-4 during a primary Listeria infection and to investigate the role of IL-4 during a secondary infection in mice using encapsulated MoAb-forming cells . During the course of a primary infection in mice given anti-IL-4 antibody-forming cells (anti-IL-4-FC), the number of Listeria found in the liver and spleen was comparable to that found in control mice given anti-beta-galactosidase antibody-forming cells (anti-beta-gal-FC) . Activation of macrophages measured by inhibition of Toxoplasma gondii proliferation and the release of reactive nitrogen intermediates (RNI) was not affected by anti-IL-4-FC treatment during infection . Furthermore, during a secondary L . monocytogenes infection the number of bacteria in the liver and spleen of anti-IL-4-treated immune mice was comparable to anti-beta-gal-FC-treated, control, immune mice . The concentration of IFN-gamma in plasma of anti-IL-4-treated immune mice was similar to that of control immune mice . Taken together, these findings demonstrate that neutralization of endogenously formed IL-4 does not affect resistance to a primary or a secondary L . monocytogenes infection in mice.

FEMS Microbiol Lett, 1998 May 1, 162(1), 169 - 76
Host cell protein tyrosine kinases are activated during the entry of Listeria monocytogenes . Possible role of pp60c-src family protein kinases; Van Langendonck N et al.; Listeria monocytogenes is able to invade a wide range of cell types by inducing its own internalization . Little is known, however, about the host cell proteins affecting the entry process which involves triggering the host cell signal transduction mechanism . We report here that entry of L . monocytogenes strains (serotypes 4b and 1/2a) into Caco-2 cells induced tyrosine phosphorylation of several host cell proteins including pp60c-src substrates . Using specific synthetic peptide substrates, we showed that L . monocytogenes activates, as early as 5 min after bacteria-cell contact, the pp60c-src family-related (srcFR) proteins by an inlAB-dependent pathway . The activation of srcFR proteins seems to be crucial in the entry of L . monocytogenes into Caco-2 cells . Indeed, specific inhibition of the srcFR signal by herbimycin A blocked the entry of L . monocytogenes strains . Taken together, our data show that L . monocytogenes enhances cell tyrosine phosphorylations and activates the pp60c-src family-related proteins by an inlAB-dependent pathway.

Mol Microbiol, 1998 Apr, 28(1), 81 - 93
Internalin B is essential for adhesion and mediates the invasion of Listeria monocytogenes into human endothelial cells; Parida SK et al.; Listeria monocytogenes causes rhombencephalitis in humans and animals and also affects the fetus in utero, causing disseminated sepsis . In both instances, the infection occurs by the crossing of endothelial cells lining a physiological barrier, the blood-brain barrier or the transplacental barrier . In this study, the ability of L . monocytogenes wild-type EGD to invade human umbilical vein endothelial cells (HUVECs) was evaluated using wild-type bacteria and isogenic Listeria mutants . Here, we show that invasion of HUVECs by L . monocytogenes is dependent on the expression of the internalin B gene product . This was demonstrated in several ways . First, L . monocytogenes strains lacking the inlB gene did not invade HUVECs . Secondly, avid invasion was obtained when a strain deleted for inlAB was complemented with a plasmid harbouring inlB only, whereas strains expressing inlA did not enter HUVECs . Thirdly, entry of wild-type EGD could be blocked effectively with antibodies to InlB . Fourthly, cell binding assays and flow cytometry with HUVECs showed binding of purified InlB, but not InlA, suggesting a tropism of InlB for this cell type . Finally, physical association of purified native InlB with the surface of non-invasive mutants dramatically increased their ability to invade HUVECs . In laser-scanning confocal microscopy, binding of InlB was observed as focal and localized patches on the cell surface of HUVECs . Qualitative examination of the entry process by scanning electron microscopy revealed that both wild-type EGD and a recombinant strain overexpressing only InlB enter HUVECs in a similar fashion . The entry process was polarized, involved single bacteria and occurred over the entire surface of endothelial cells.

Otolaryngol Head Neck Surg, 1998 May, 118(5), 625 - 9
Effects of topical oral antiseptic rinses on bacterial counts of saliva in healthy human subjects; Balbuena L et al.; Wound infections remain a significant source of morbidity in patients undergoing major head and neck operations that invade the aerodigestive tract . Infection rates have been significantly reduced by the administration of perioperative intravenous antibiotics; however, the incidence of infection remains unacceptably high . This study was undertaken to help identify an oral antiseptic that could significantly reduce the bacterial colony count of human saliva . A randomized, prospective clinical trial was conducted to analyze and compare the effects of Listerine antiseptic and Peridex oral rinse on the aerobic and anaerobic bacterial counts in healthy human subjects . Thirty healthy adult volunteers between the ages of 18 and 61 participated in the study . The patients were randomized to receive normal saline solution, Listerine antiseptic, or Peridex oral rinse . Aerobic and anaerobic bacterial colony counts of saliva were measured before treatment and at 1 and 4 hours after treatment . Both Listerine antiseptic and Peridex oral rinse significantly reduced bacterial counts at 1 hour after treatment in our volunteers . At 4 hours after treatment, Peridex oral rinse showed a further reduction in the bacterial colony count whereas Listerine antiseptic showed no difference compared with normal saline solution . At 4 hours after treatment, Peridex oral rinse reduced the total bacterial colony count by 85%.

Curr Microbiol, 1998 May, 36(5), 309 - 18
Identification and characterization of nucleotide sequence differences in three virulence-associated genes of listeria monocytogenes strains representing clinically important serotypes; Vines A et al.; Listeria monocytogenes is a Gram-positive, facultative intracellular bacterium that causes invasive, often fatal, disease in susceptible hosts . As a foodborne pathogen, the bacterium has emerged as a significant public health problem and has caused several epidemics in the United States and Europe . Three serotypes (1/2a, 1/2b, 4b) of L . monocytogenes are responsible for nearly 95% of all reported cases of human listeriosis . L . monocytogenes serotype 4b has caused all well-characterized foodborne epidemic outbreaks in North America and Europe between 1981 and 1993 . However, most of the genetic studies to characterize virulence factors of L . monocytogenes have been done by using serotypes 1/2a and 1/2c . In this investigation, we examined three virulence-associated genes (hly encoding listeriolysin, plcA encoding phosphotidylinositol-specific phospholipase C, and inlA encoding internalin) of two serotype 4b and two serotype 1/2b strains . We chose these virulence-associated genes on the basis of published sequence differences among strains from Listeria subgroups containing serotypes 1/2a and 1/2c versus 4b, respectively . They correspond to sequence homologies that include very highly conserved (hlyA), highly conserved (plcA) and mostly conserved (inlA) . We found by using nucleotide sequence analysis of the hly, plcA, and inlA genes, the two L . monocytogenes strains (including a strain associated with a foodborne disease outbreak in California in 1985) in this study, two serotype 1/2b strains from a study that we recently reported, and other similar published data for serotypes 1/2a, 1/2c, and 4b, had a high degree of sequence conservation at the gene and protein levels for all three genes . However, the sequences for the hly gene of L . monocytogenes strains of serotypes 1/2b and 4b were more closely related to each other and showed significant divergence from serotypes 1/2a and 1/2c . A unique nonsynonymous mutation was found in the hly gene of L . monocytogenes isolates that were associated with the 1985 California outbreak and were the epidemic phage type . When 158 L . monocytogenes isolates from the collection at the Centers for Disease Control and Prevention were screened, the mutation was found only in one other strain that had been isolated in California 3 years before the epidemic . Although the California epidemic clone was lactose negative, other L . monocytogenes serotype 4b isolates that were lactose negative did not possess the unique mutation observed in that epidemic clone.

Curr Microbiol, 1998 May, 36(5), 266 - 70
Flow cytometry demonstrates bacteriocin-induced injury to Listeria monocytogenes; Swarts AJ et al.; Flow cytometry was used to study the effect of the bacteriocin leucocin B-TA11a on Listeria (L.) monocytogenes . Mixed proportions of dead and live control populations were analyzed by flow cytometry to determine detection limits of the Dead/Live Baclight Bacterial Viability KitTM . High correlations for flow cytometric detection of defined proportions of live or dead cells in mixtures between 10 and 100% of dead (r2 = 0.97) or live (r2 = 0.99) cells were obtained . However, mixtures containing less than 10% of either live or dead control cells gave correlations below 0.72 . The growth of L . monocytogenes in the absence and presence of leucocin B-TA11a was analyzed by flow cytometry with Baclight, plate counts, and optical density measurements . Although leucocin B-TA11a initially inhibited listerial growth, the uptake of both Baclight dyes suggested that cells remained viable but became leaky, possibly indicating bacteriocin-induced pore formation in the target membranes.

W V Med J, 1998 Mar-Apr, 94(2), 80 - 3
Listeria monocytogenes rhomboencephalitis with cranial-nerve palsies: a case report; Shaffer DN et al.; Listeria monocytogenes rhomboencephalitis is an uncommon complication of L . monocytogenes meningitis . It presents in a typical biphasic pattern characterized by a non-specific prodromal period followed by any combination of asymmetrical, cranial-nerve palsies; cerebellar signs; hemiparesis or hypesthesia; and diminished consciousness . The survival rate is greater than 70% when appropriate antibiotic therapy is initiated early . However, approximately 60 percent of the survivors develop neurological sequelae . We present the case of a 33-year-old woman who developed L . monocytogenes meningitis with subsequent rhomboencephalitis and cranial-nerve palsie, and review the literature on this syndrome.

Toxicol Sci, 1998 Apr, 42(2), 91 - 8
Pyrimethamine impairs host resistance to infection with Listeria monocytogenes in BALB/c mice; Freund YR et al.; Increased mortality has been observed when HIV-infected patients were treated with pyrimethamine (Pyr) as prophylaxis for toxoplasmic encephalitis, suggesting that Pyr might possess immunosuppressive activity . To analyze this in an animal model, immune function was assessed in BALB/c mice using a battery of in vivo and ex vivo assays and an in vivo model of host resistance to Listeria monocytogenes infection . Treatment for 30 days with 60 mg/kg Pyr decreased circulating white blood cell and lymphocyte counts but not neutrophil, red blood cell, or platelet counts or hemoglobin levels . Splenic B cell percentages and lipopolysaccharide-induced B cell proliferation decreased significantly after treatment with 60 mg/kg Pyr, as did levels of anti-keyhole limpet hemocyanin (KLH) IgM in serum 7 days after immunization with KLH . Anti-KLH IgG levels 14 days after immunization were not affected . Percentages of splenic T cells and macrophages and T cell proliferation in the presence of concanavalin A or allogeneic cells were not decreased by Pyr treatment . An ex vivo assay of T-cell-mediated cytotoxicity was also unaffected . When host resistance to L . monocytogenes infection was assessed, dramatic increases in mortality were observed in Pyr-treated compared to control mice . Increased numbers of L . monocytogenes organisms were observed in liver and spleen of Pyr-treated mice, compared to controls . The reduction in Listeria resistance, which is T cell mediated, contrasts with the fact that no significant changes in T-cell-mediated immunity were observed . It is possible that Pyr affects parameters of innate immunity, which were not monitored in this study.

Biol Neonate, 1998, 73(5), 340 - 5
Effects of endotoxemia and sepsis on bilirubin oxidation by rat brain mitochondrial membranes; Allen JW et al.; Sepsis is believed to increase the risk of bilirubin brain toxicity, but the mechanism is not known . Adult male Sprague-Dawley rats were injected intraperitoneally with either 20 mg/kg Escherichia coli lipopolysaccharide, approximately 5 x 10(9)/kg CFU Listeria monocytogenes or vehicle 48 h prior to sacrifice . Rats were killed with an intraperitoneal injection of pentobarbital . Mitochondrial membrane fractions were produced by homogenization of the brains and differential centrifugation in 0.32 M sucrose . The mitochondrial pellet was resuspended in distilled water and sonicated to rupture the mitochondria . The protein concentration of the suspension was standardized to 2.5 mg/ml . Bilirubin oxidation was assayed in a pH 8.2, 0.1 M barbital buffer containing 10 microM bilirubin, 5 mM EDTA, and 500 U/ml catalase . Optical density was measured at 440 nm before and after a 60-min incubation at 37.5 degrees C . There were no differences between the control, endotoxemic, and septic groups as far as the ability of brain mitochondrial membranes to oxidize bilirubin (bilirubin oxidation rate: 289 +/- 11 vs . 295 +/- 9 vs . 296 +/- 12 pmol/min/mg protein, mean +/- SD) . We conclude that endotoxemia or sepsis do not change the ability of brain mitochondrial membranes to oxidize bilirubin . If sepsis truly increases the risk of bilirubin encephalopathy in neonatal jaundice, this is likely to involve other mechanisms.

What Is Molecular Biology?, What Is Water Purification?, What Is Growth Medium?, What Is Activated Sludge?, What Is Biofilm?, a, Bacteria, n, Bacterium, e, Microorganisms, n, Microbes, e, Bacteriology, r, Bacillus, n, Yeasts, n, Microorganism, a, Bioreactor, s, Bacteriophages, c, Bioreactors, a, Antibiotics, i, Microorganism, e, Rhizobacter, s, Enterococci, r, Escherichia coli, o, Micrococci, i, Staphylococcus, s, Bacteroides, c, Yeast, c, Bacillus, i, E coli O157, e, Escherichia coli, i, Lactobacillus, e, Bacillus, i, Growth media

Scientific Publications - Work Done by Microbiology Reader Bioscreen C
Agricultural Microbiology Anaerobic Microbiology Antimicrobial Susceptibility Artificial Atmosphere Bioassay of Antibiotics Biofilm Microbiology Bioreactor Technology Biotechnology Cell Biology Clinical Microbiology Environmental Microbiology Experiments with Yeast	Fermentation Food Microbiology Functional Genomics Gene Technology Growth Media Development Growth Rate and Lag Time Industrial Microbiology Medical/Pharmaceutical Field Microbiological Assay Microbiological Research Microbiology of Cosmetics go to a specific theme...	Military Microbiology Molecular Microbiology Mutagenicity and Genotoxicity Oral Microbiology Patents Postantibiotic Studies Soil Microbiology Spore Microbiology Veterinary Microbiology Waste/Wastewater Treatment Water Microbiology Wine Microbiology

� 2005 Transgalactic Ltd (manufacturer of Bioscreen C software) | Privacy Statement | P.O. Box 1393, 00101 Helsinki, Finland, phone: +358 9 85172920, fax: +358 9 8749481, e-mail: microbiology@bionewsonline.com

Last modified: May 25, 2005