Listeriosis

J Microbiol Methods, 2005 Feb, 60(2), 259 - 68
Specific detection of cytopathogenic Listeria monocytogenes using a two-step method of immunoseparation and cytotoxicity analysis; Gray KM et al.; The development of rapid methods for detection of viable Listeria monocytogenes is crucial to prevent listeriosis and product recalls . While immunomagnetic separation has been used for isolating Listeria spp., lack of specificity and pathogenicity determination render this method unsatisfactory . A two-step method using Protein A agarose beads (Immunobeads) coated with a more specific antibody, monoclonal antibody (MAb)-C11E9 for L . monocytogenes was developed . Immunobeads were allowed to capture Listeria cells from a variety of samples and tested for cytopathogenic action on a murine hybridoma B-lymphocyte, Ped-2E9 cell line by Trypan blue staining, and by an alkaline phosphatase (AP)-based cytotoxicity assay . The two-step method was used to test uninoculated hotdogs, bologna, and raw beef, chicken, and pork samples, following selective enrichment in half-Fraser broth . Pure culture studies proved the assay to be specific for L . monocytogenes, while a similar assay with Dynal(R) Anti-Listeria immunomagnetic beads was positive for L . monocytogenes, L . ivanovii, and L . seeligeri . Detection and confirmation of cytopathogenicity of Listeria cells from food samples after 24-h selective enrichment were completed in 2-4 h . Isolates were further analyzed by the CAMP test for hemolytic activity and RiboPrinter(R) for genomic patterns . Using immunoseparation and cytotoxicity as a two-step rapid method, viable L . monocytogenes could be isolated, detected, and confirmed as cytopathogenic in 28 h or less.

J Immunol, 2004 Dec 15, 173(12), 7416 - 25
IFN regulatory factor 3-dependent induction of type I IFNs by intracellular bacteria is mediated by a TLR- and Nod2-independent mechanism; Stockinger S et al.; Like viruses, intracellular bacteria stimulate their host cells to produce type I IFNs (IFN-alpha and IFN-beta) . In our study, we investigated the signals and molecules relevant for the synthesis of and response to IFN by mouse macrophages infected with Listeria monocytogenes . We report that IFN-beta is the critical immediate-early IFN made during infection, because the synthesis of all other type I IFN, expression of a subset of infection-induced genes, and the biological response to type I IFN was lost upon IFN-beta deficiency . The induction of IFN-beta mRNA and the IFN-beta-dependent sensitization of macrophages to bacteria-induced death, in turn, was absolutely dependent upon the presence of the transcription factor IFN regulatory factor 3 (IRF3) . IFN-beta synthesis and signal transduction occurred in macrophages deficient for TLR or their adaptors MyD88, TRIF, or TRAM . Expression of Nod2, a candidate receptor for intracellular bacteria, increased during infection, but the protein was not required for Listeria-induced signal transduction to the Ifn-beta gene . Based on our data, we propose that IRF3 is a convergence point for signals derived from structurally unrelated intracellular pathogens, and that L . monocytogenes stimulates a novel TLR- and Nod2-independent pathway to target IRF3 and the type I IFN genes.

Appl Environ Microbiol, 2004 Dec, 70(12), 7555 - 7
Role for compatible solutes glycine betaine and L-carnitine in listerial barotolerance; Smiddy M et al.; Increased listerial barotolerance at elevated osmolarity is attributed, in part, to the presence of accumulated betaine and L-carnitine . The percentage of listerial survival following exposure to 400 MPa for 5 min increased from 0.008 to 0.02% with added L-carnitine (5 mM) and to 0.05% with added betaine (5 mM) . Furthermore, listerial cells incapable of transporting compatible solutes fail to adapt to high pressure at elevated osmolarity.

J Morphol . 2004 Nov 29; {Epub ahead of print}
Ultrastructural observations of spermatozoa of several tetragnathid spiders with phylogenetic implications (Araneae, Tetragnathidae); Michalik P et al.; The present study reports on the ultrastructure of spermatozoa and spermatogenesis of 12 tetragnathid spiders (10 Tetragnatha species {T . boydi, T . dearmata, T . extensa, T . montana, T . nigrita, T . obtusa, T . pinicola, T . reimoseri, T . shoshone, T . striata}; Pachygnatha listeri and Metellina segmentata) . All species develop typical cleistospermia with a coiled nucleus in the center and a coiled axoneme in the periphery of the cell . Remarkable differences in the sperm ultrastructure of the investigated species comprise the shape of the main sperm cell components (nucleus, acrosomal complex, implantation fossa, and centriolar complex) . Within the observed Tetragnatha species, three types of sperms were characterized: T . montana-type, T . boydi-type, and T . striata-type . The highly derivative T . montana-type is characterized by the following remarkable features: an extremely elongated nucleus, shaped like a corkscrew and twisted around the axoneme (before coiling); a deep implantation fossa; a corkscrew-shaped acrosomal vacuole; after the coiling process, the nucleus is coiled five to six times in the center of the spermatozoon and the axoneme is coiled five to six times peripheral to the nucleus . The T . boydi-type hardly differs from the T . montana-type, but is remarkable due to the triangular-shaped nucleus (in cross section) . The T . striata-type differs especially by a peculiar acrosomal vacuole with a long, slightly curved process and a short appendix, as well as a nucleus that describes only three loose coils around the axoneme (before coiling) . The spermatozoa of Pachygnatha listeri and especially Metellina segmentata differ strikingly from the described Tetragnatha-types and are similar to more primitive araneomorph spermatozoa, such as Hypochilus pococki . The described Tetragnatha-types completely correspond with Okuma's (1988a,b, J Fac Agr Kyushu U 32:165-181, 32:183-213) classification of Tetragnatha species . Furthermore, our results suggest an early derivative systematic position of Pachygnatha within Tetragnathinae and the position of Metellina within the Tetragnathidae . (c) 2004 Wiley-Liss, Inc.

Vet Res Commun, 2004 Oct, 28(7), 569 - 79
Humoral and delayed-type hypersensitive responses against listeria monocytogenes phosphatidylinositol-specific phospholipase C in experimentally infected buffaloes; Chaudhari SP et al.; The kinetics of antibody production against phosphatidylinositol-specific phospholipase C (PI-PLC) and the isolation pattern of Listeria monocytogenes from bacteriological samples were studied following oral infection of buffalo calves with 3 x 10(9) cells each of pathogenic L . monocytogenes . Antibodies to PI-PLC appeared by 4-8 days post infection (PI), with a peak between days 7 and 16 PI, when tested by indirect plate-ELISA . Subsequently, antibody titres in all the animals declined and became undetectable on days 26-35 PI onwards until the study concluded on day 211 PI . Dot-ELISA could detect the antibodies to PI-PLC 1-2 days earlier and at higher titres as compared to plate-ELISA . L . monocytogenes could be recovered from faeces, nasal swabs and haemocultures from days 2 to 33, days 2 to 21 and days 11 to 17 PI, respectively . Antibodies to PI-PLC were detected during the course of active infection but their titres declined sharply once animals became culturally negative . Sonicated antigen elicited the highest delayed-type hypersensitivity response, followed by PI-PLC and listeriolysin O.

Vet Res Commun, 2004 Oct, 28(7), 561 - 7
Listeria monocytogenes in products of animal origin in Turkey; Akpolat NO et al.; A study was carried out on 430 samples of different foodstuffs (soft cheese, raw chicken, minced beef, sausage, fish) and 400 carcase samples (sheep, young and adult cattle) for screening of Listeria monocytogenes . It was found that only one of the samples contained L . monocytogenes at > 10(3) cfu/ml in the initial examination, but another 42 samples contained L . monocytogenes following an enrichment process . L . monocytogenes was isolated most frequently from raw chicken samples (18%), but was not isolated from sausage samples . Forty-three isolates were defined as serotypes by using Bacto-Listeria-O-antisera Type 1 (Difco 2300-50-2) and Type 4 (Difco 2301-50-1) except that Type poly was not used . For these reasons, all isolates were classified as type 1 or type 4 and the other was termed untypeable . Twenty-one samples were type 1, 17 were untypeable, and 5 were both serotype 4 and untypeable.

PLoS Biol . 2004 Dec;2(12):e412 . Epub 2004 Nov 30.
In silico reconstitution of Listeria propulsion exhibits nano-saltation; Alberts JB et al.; To understand how the actin-polymerization-mediated movements in cells emerge from myriad individual protein-protein interactions, we developed a computational model of Listeria monocytogenes propulsion that explicitly simulates a large number of monomer-scale biochemical and mechanical interactions . The literature on actin networks and L . monocytogenes motility provides the foundation for a realistic mathematical/computer simulation, because most of the key rate constants governing actin network dynamics have been measured . We use a cluster of 80 Linux processors and our own suite of simulation and analysis software to characterize salient features of bacterial motion . Our "in silico reconstitution" produces qualitatively realistic bacterial motion with regard to speed and persistence of motion and actin tail morphology . The model also produces smaller scale emergent behavior; we demonstrate how the observed nano-saltatory motion of L . monocytogenes,in which runs punctuate pauses, can emerge from a cooperative binding and breaking of attachments between actin filaments and the bacterium . We describe our modeling methodology in detail, as it is likely to be useful for understanding any subcellular system in which the dynamics of many simple interactions lead to complex emergent behavior, e.g., lamellipodia and filopodia extension, cellular organization, and cytokinesis.

Infect Immun, 2004 Dec, 72(12), 7374 - 8
Listeria monocytogenes sigmaB contributes to invasion of human intestinal epithelial cells; Kim H et al.; The role of sigma(B) in Listeria monocytogenes infection of human intestinal epithelial cells was investigated . Invasion defects associated with loss of sigma(B) paralleled those of a DeltainlA strain independently of the sigma(B)-dependent P2(prfA) promoter . Concomitantly, amounts of inlA transcript and InlA protein were significantly decreased in the DeltasigB strain.

Infect Immun, 2004 Dec, 72(12), 7005 - 11
Immunostimulating properties of intragastrically administered Acetobacter-derived soluble branched (1,4)-beta-D-glucans decrease murine susceptibility to Listeria monocytogenes; Li W et al.; We previously found that AC-1, an extracellular polysaccharide, produced by Acetobacter xylinum and composed of (1,4)-beta-D-glucan with branches of glucosyl residues, showed a strong activity to induce production of interleukin-12 (IL-12) p40 and tumor necrosis factor alpha by macrophages in vitro via Toll-like receptor 4 (TLR-4) signaling . In the present study, we examined the effect of oral administration of AC-1 on protective immunity against Listeria monocytogenes . Mice were given AC-1 or phosphate-buffered saline (PBS) intragastrically 2 days before, on the day of, and 2 days after an intraperitoneal inoculation of L . monocytogenes . The survival rate of AC-1-treated mice was significantly improved and bacterial growth in AC-1-treated mice was severely retarded compared to those of PBS-treated mice after infection with L . monocytogenes . IL-12 p40 levels in serum and magnitudes of CD4+ Th1 and CD8+ Tc1 responses against Listeria antigen were significantly higher in AC-1-treated mice than in PBS-treated mice . The effect of AC-1 on antilisterial activity was diminished in C3H/HeJ mice carrying mutated TLR-4 . Thus, AC-1, a potent IL-12 inducer through TLR-4, enhanced protective immunity against L . monocytogenes via augmentation of Th1 responses . These results suggest that infectious processes driven by intracellular microorganisms could be prevented to develop by the (1,4)-beta-D-glucan.

J Immunol, 2004 Dec 1, 173(11), 6694 - 702
Shortening the infectious period does not alter expansion of CD8 T cells but diminishes their capacity to differentiate into memory cells; Williams MA et al.; Following a primary immune response, a portion of effector T cells gives rise to long-lived memory cells . Although primary expansion and differentiation of effector CD8 T cells is dictated by a brief exposure to Ag, it is unclear whether full memory differentiation is also programmed within the same short window . By carefully modulating the kinetics of Listeria monocytogenes infection, we analyzed the requirements for the programming of effector and memory T cell development in vivo . We find that although limiting the infectious period to the first 24-48 h does not impact the size of the primary CD8 response, the ensuing memory population is significantly diminished . This effect is particularly pronounced in the development of tissue-homing memory cells and is inversely proportional to the initial infectious dose . In contrast to CD8 responses, the differentiation of primary CD4 responses was highly dependent on the continued presence of the infection . Shortening the duration of the infection greatly reduced the development of CD4 effector responses in the spleen and prevented their trafficking to peripheral sites of infection . We propose that the stimulus received by CD8 T cells during the early stages of infection largely contribute to the differentiation of CD8 effector cells, whereas continued or distinct signals received at later stages influence their ability to differentiate into memory cells.

J Food Prot, 2004 Nov, 67(11), 2544 - 9
Efficacy of acidic electrolyzed water ice for pathogen control on lettuce; Koseki S et al.; Acidic electrolyzed water (AcEW) was used as frozen AcEW (AcEW-ice) for inactivation of Listeria monocytogenes and Escherichia coli O157:H7 on lettuce . AcEW-ice was prepared from AcEW with 20, 50, 100, and 200 ppm of available chlorine by freezing at -40 degrees C and generated 30, 70, 150, and 240 ppm of chlorine gas (Cl2), respectively . The AcEW-ice was placed into styrene-foam containers with lettuce samples at 20 degrees C for 24 h . Although AcEW-ice generating 30 ppm Cl2 had no effect on L . monocytogenes cell counts, AcEW-ice generating 70 to 240 ppm of Cl2 significantly (P < 0.05) reduced L . monocytogenes by ca . 1.5 log CFU/g . E . coli O157:H7 cell counts were reduced by 1.0 log CFU/g with AcEW-ice generating 30 ppm of Cl2 . AcEW-ice generating 70 and 150 ppm of Cl2 reduced E . coli O157:H7 by 2.0 log CFU/g . Further significant reduction of E . coli O157:H7 (2.5 log CFU/g) was demonstrated by treatment with AcEW-ice generating 240 ppm of Cl2 . However, treatment with AcEW-ice generating 240 ppm of Cl2 resulted in a physiological disorder resembling leaf burn . AcEW-ice that generated less than 150 ppm of Cl2 had no effect on the surface color of the lettuce . AcEW-ice, regardless of the concentration of the emission of Cl2, had no effect on the ascorbic acid content in the lettuce . The weight ratio of lettuce to AcEW-ice required was determined to be over 1:10 . The bactericidal effect of AcEW-ice appeared within the first 2 h . The use of AcEW-ice provides simultaneously for low temperature storage and inactivation of bacteria.

J Food Prot, 2004 Nov, 67(11), 2500 - 14
Longitudinal studies on Listeria in smoked fish plants: impact of intervention strategies on contamination patterns; Lappi VR et al.; Four ready-to-eat smoked fish plants were monitored for 2 years to study Listeria contamination patterns and the impact of plant-specific Listeria control strategies, including employee training and targeted sanitation procedures, on Listeria contamination patterns . Samples from the processing plant environment and from raw and finished product were collected monthly and tested for Listeria spp . and Listeria monocytogenes . Before implementation of intervention strategies, 19.2% of raw product samples (n = 276), 8.7% of finished product samples (n = 275), and 26.1% of environmental samples (n = 617) tested positive for Listeria spp . During and after implementation of Listeria control strategies, 19.0% of raw product samples (n = 242), 7.0% of finished product samples (n = 244), and 19.5% of environmental samples (n = 527) were positive for Listeria spp . In one of the four fish plants (plant 4), no environmental samples were positive for L . monocytogenes, and this plant was thus excluded from statistical analyses . Based on data pooled from plants 1, 2, and 3, environmental Listeria spp . prevalence was significantly lower (P < 0.05) for nonfood contact surfaces and the finished product area and for the overall core environmental samples after implementation of control strategies . Listeria prevalence for floor drains was similar before and after implementation of controls (49.6 and 54.2%, respectively) . Regression analysis revealed a significant positive relationship (P < 0.05) between L . monocytogenes prevalence in the environment and in finished products before implementation of control strategies; however, this relationship was absolved by implementation of Listeria control strategies . Molecular subtyping (EcoRI ribotyping) revealed that specific L . monocytogenes ribotypes persisted in three processing plants over time . These persistent ribotypes were responsible for all six finished product contamination events detected in plant 1 . Ribotype data also indicated that incoming raw material is only rarely a direct source of finished product contamination . While these data indicate that plant-specific Listeria control strategies can reduce cross-contamination and prevalence of Listeria spp . and L . monocytogenes in the plant environment, elimination of persistent L . monocytogenes strains remains a considerable challenge.

J Food Prot, 2004 Nov, 67(11), 2496 - 9
Dairy farm reservoir of Listeria monocytogenes sporadic and epidemic strains; Borucki MK et al.; Identifying the reservoirs of a pathogen is vital for control of sporadic disease and epidemics . Listeria monocytogenes is a zoonotic foodborne pathogen that is responsible for 28% of food-related deaths in the United States annually, as well as a major cause of massive product recalls worldwide . To examine the role of the dairy farm as a potential source or reservoir for L . monocytogenes subtypes shown to cause human listeriosis, we compared the pulsed-field gel electrophoresis (PFGE) restriction enzyme digestion profiles of L . monocytogenes dairy farm-associated strains (milk, environmental, and bovine) to human sporadic and epidemic disease strains . Twenty-three percent of human sporadic strains had PFGE patterns identical to that of farm isolate(s) . Additionally, three farm environmental strains and one human sporadic strain had a PFGE pattern identical to a strain of L . monocytogenes responsible for the 1985 California epidemic . These data indicate that this epidemic strain continues to cause sporadic human illness and has a potential dairy farm as a reservoir.

J Food Prot, 2004 Nov, 67(11), 2488 - 95
Strain-specific differences in the attachment of Listeria monocytogenes to alfalfa sprouts; Gorski L et al.; Contamination of fresh produce with Listeria monocytogenes has resulted in outbreaks of systemic listeriosis and febrile gastroenteritis . Recalls of alfalfa sprouts have occurred due to contamination with L . monocytogenes . Alfalfa sprouts were used as a preharvest model to study the interaction with this human pathogen . Seventeen strains were assessed for their capacity to colonize alfalfa sprouts, and strain-specific differences (not related to source, serotype, or lineage) were revealed when the sprout irrigation water was changed daily . Two of the strains colonized and attached to the sprouts very well, reaching levels of more than 5 log CFU per sprout . The remaining strains varied in their final levels on sprouts between less than 1 to 4.7 log CFU per sprout . All of the L . monocytogenes strains grew to equivalent levels on the sprouts when the irrigation water was not changed, suggesting the differences observed with regular changing of the water resulted from differences in attachment . Further analysis of the best colonizing strains indicated that only between 0.3 and 1 log CFU per sprout could be removed by additional washing of the sprout, and the presence of normal sprout bacteria did not compete with the L . monocytogenes strains on the sprouts . The poorest colonizing strain was able to grow in the irrigation water during the experiment but could not attach to the sprouts . Microscopic examination of the sprouts with L . monocytogenes expressing the green fluorescent protein indicated that L . monocytogenes was associated with the root hairs of the sprouting alfalfa, with few to no cells visible elsewhere on the sprout.

J Food Prot, 2004 Nov, 67(11), 2480 - 7
Prevalence and typing of Listeria monocytogenes in ready-to-eat food products on the Belgian market; Van Coillie E et al.; Listeria monocytogenes is a major concern to producers of ready-to-eat foods because of the high mortality rate associated with listeriosis and the widespread nature of the organism . To investigate the prevalence of this pathogen in different ready-to-eat food products on the Belgian market, a variety of 252 ready-to-eat food products, mainly fish and meat products, were analyzed . Overall, L . monocytogenes was detected in 23.4% of the samples . The highest prevalence of L . monocytogenes was found in prepared minced meat (42.1%) and smoked halibut (33.3%) . Contamination levels were in most cases low (<10 CFU/g); however, levels higher than 100 CFU/g were detected in some samples of smoked salmon, smoked halibut, and prepared minced meat . A high prevalence of Listeria innocua (15.8%) and Listeria welshimeri (36.8%) was detected in prepared minced meat . L . monocytogenes strains isolated from different contaminated products were subjected to repetitive element sequence-based PCR (REP-PCR) typing to determine possible associations with product type, producer, or market . REP-PCR patterns were analyzed using BioNumerics software, and seven different groups with at least 90% similarity were identified . The cluster analysis indicates that cross-contamination occurred at the producer and retail level . Serotype identification of the strains by PCR revealed that most belonged to the 1/2a(3a) serotype group.

J Food Prot, 2004 Nov, 67(11), 2472 - 9
Effect of prepackage and postpackage pasteurization on postprocess elimination of Listeria monocytogenes on deli turkey products; Muriana P et al.; Surface pasteurization for inactivation of Listeria monocytogenes was evaluated for radiant heat prepackage pasteurization, submersed water postpackage pasteurization, and combinations of the two techniques on various types of ready-to-eat deli turkey products obtained from at least four different manufacturers . Products were inoculated either by in-package liquid inoculum or surface sponge-contact with approximately 10(9) CFU of L . monocytogenes . Additional testing of radiant heat pasteurization was performed with low-level inoculation of product undersides with approximately 100 CFU of L . monocytogenes followed by enrichment recovery after pasteurization . Prepackage pasteurization provided 2.0 to 2.8 log reductions when processed for 60 s and 2.8 to 3.8 log reductions when processed for 75 s . An improved radiant oven provided 3.53 (60 s) and 4.76 (75 s) log reductions of L . monocytogenes . No positive samples were detected after enrichment when 40 samples of deli turkey (4 to 4.5 kg) undersides were inoculated at low levels and processed for 75 s . Submersed water postpackage pasteurization provided 1.95 to 3.0 log reductions when processed for 2, 3, 4, or 5 min, and combinations of the two processes gave 3.0 to 4.0 log inactivation of L . monocytogenes using either 60 + 60 s or 60 + 90 s for the prepackage and postpackage pasteurization processes, respectively . These processes, either individually or in combination, can provide postprocess elimination of bacteria for the manufacture of safe ready-to-eat deli meats.

J Food Prot, 2004 Nov, 67(11), 2465 - 71
Control of growth and survival of Listeria monocytogenes on smoked salmon by combined potassium lactate and sodium diacetate and freezing stress during refrigeration and frozen storage; Yoon KS et al.; In this study, we evaluated the antimicrobial effects of different levels of a potassium lactate (PL) plus sodium diacetate (SDA) mixture against the growth and survival of Listeria monocytogenes Scott A inoculated onto smoked salmon stored at 4, 10, and -20 degrees C . The effect of freezing stress on the growth kinetics of L . monocytogenes Scott A on smoked salmon at 4 and 10 degrees C was also investigated . The use of PL+SDA at all tested levels (1.5, 3.3, and 5% of a 60% commercial solution of PURASAL P Opti.Form 4) completely inhibited the growth of L . monocytogenes Scott A on smoked salmon stored at 4 degrees C during 32 days of storage . It also delayed the growth of L . monocytogenes Scott A on smoked salmon stored at 10 degrees C for up to 11 days, but a listeriostatic effect was observed only with 5% PURASAL P Opti.Form 4 at 10 degrees C after 11 days . Addition of PL+SDA at all tested levels decreased the surviving populations of L monocytogenes Scott A on smoked salmon during 10 months of frozen storage at -20 degrees C . Freezing stress significantly (P < 0.001) extended the lag time and delayed the growth of L . monocytogenes Scott A at both 4 and 10 degrees C . However, the effect of freezing stress was more significant at 4 degrees C than at 10 degrees C, indicating the importance of temperature control of smoked salmon during the retail storage period.

J Food Prot, 2004 Nov, 67(11), 2450 - 5
Decontamination of strawberries using batch and continuous chlorine dioxide gas treatments; Han Y et al.; Efficacy of chlorine dioxide (ClO2) gas in reducing Escherichia coli O157:H7 and Listeria monocytogenes on strawberries was determined using batch and continuous flow ClO2 gas treatment systems . Effects of continuous ClO2 gas treatment on total aerobic plate count, color, and residual ClO2 and chlorite on strawberries were also evaluated . Strawberries were spot inoculated with 7 to 8 log CFU per strawberry of each pathogen (E . coli O157:H7 and L . monocytogenes), stored for 1 day at 4 degrees C, and treated at 22 degrees C and 90 to 95% relative humidity with 0.2 to 4.0 mg/liter ClO2 gas for 15 or 30 min using a batch treatment system or with 0.6, 1.8, and 3.0 mg/liter for 10 min using a continuous treatment system . Surviving microbial populations were determined using a membrane-transfer plating recovery method . Increased ClO2 gas concentrations resulted in increased log reductions of each pathogen for both the batch and continuous systems . A batch treatment of strawberries with 4 mg/liter ClO2 for 30 min and continuous treatment with 3 mg/liter ClO2 for 10 min achieved greater than a 5-log reduction for both E . coli O157:H7 and L . monocytogenes . After continuous exposure to 3.0 mg/liter ClO2 gas for 10 min followed by 1 week of storage at 4 degrees C, no aerobic microorganisms were detected and the color of the strawberry surface did not change significantly (P > 0.05) . Residues of ClO2 and chlorite on strawberries after the treatment were 0.19 +/- 0.33 mg ClO2 per kg and 1.17 +/- 2.02 mg Cl2 per kg, respectively, whereas after 1 week of storage no ClO2 residues were detected and residual chlorite levels were down to 0.07 +/- 0.12 mg Cl2 per kg . These results suggest that ClO2 gas treatment is an effective decontamination technique for improving the safety of strawberries while extending shelf life.

J Food Prot, 2004 Nov, 67(11), 2443 - 9
Fate of Escherichia coli O157:H7 and Listeria monocytogenes in strawberry juice and acidified media at different pH values and temperatures; Han Y et al.; Survival and growth of Escherichia coli O157:H7 and Listeria monocytogenes in strawberry juice and acidified media at different pH levels (pH 3.4 to 6.8) and temperatures were studied . Sterile strawberry juice (pH 3.6) and acidified trypticase soy broth (TSB) media (pH 3.4 to 6.8) were inoculated with approximately 6.7 log CFU/ml E . coli O157:H7 or 7.3 log CFU/ ml L . monocytogenes, incubated for 3 days at 4 and 37 degrees C . Bacterial levels were determined after 2 h, 1 day, and 3 days using surface plating nonselectively on tryptic soy agar and selectively on sorbitol MacConkey agar for E . coli O157:H7 or modified Oxford agar for L . monocytogenes . A spectrophotometer (660 nm) was also used to study growth inhibition of L . monocytogenes in different TSB and strawberry juice media (pH 3.4 to 7.3) . E . coli O157:H7 survived well at pH values of 3.4 to 6.8 at 4 degrees C, but the number of injured cells increased as pH decreased and incubation time increased . At 37 degrees C, E . coli O157:H7 was inactivated at pH of < or = 3.6 but could grow at pH 4.7 . L . monocytogenes was quickly injured at pH of < or = 4.7 within 2 h of storage at 4 degrees C and then was slightly and gradually inactivated as storage time increased . L . monocytogenes survived well at pH 6.8 at 4 degrees C and grew well at 37 degrees C . Growth of L . monocytogenes at 37 degrees C was inhibited in TSB by 1% citric acid and 0.5% malic acids at pH 3.4 or by 50% strawberry juice at pH 4.7 . Bacterial injury and inactivation appeared to be induced by the acids in strawberry juice . The acids, pH value, temperature, and time were important factors for bacterial survival, inactivation, and growth in the media tested.

J Food Prot, 2004 Nov, 67(11), 2436 - 42
Effects of recovery, plating, and inoculation methods on quantification of Escherichia coli O157:H7 and Listeria monocytogenes from strawberries; Han Y et al.; Effects of different recovery and inoculation methods on quantification of Escherichia coli O157:H7 and Listeria monocytogenes from strawberries were studied . Strawberries were spot or dip inoculated with 7 to 8 log CFU per strawberry of each pathogen, air dried for 2 h, and stored for 1, 3, and 7 days at 4 degrees C . The inoculated samples were stomached or washed with phosphate-buffered saline (PBS; pH 7.2) or with modified PBS (pH 8.4) . Bacterial levels were determined using a direct selective plating, thin agar layer plating, or membrane-transferring plating (MTP) with tryptic soy agar and sorbital MacConkey agar (E . coli O157:H7) or modified Oxford agar (L . monocytogenes) . Under most test conditions, washing with PBS followed by MTP had significantly higher (P < 0.05) recovery for both bacteria compared with other tested methods . Within a 7-day storage period for spot-inoculated strawberries, a stomaching step resulted in an injury of 0.9 to 1.4 log CFU for E . coli O157: H7 and 1.4 to 1.7 log CFU for L . monocytogenes . When a washing step was used instead, this resulted in an injury of only 0.2 to 0.6 log CFU for E . coli O157:H7 and 0.2 to 0.7 log CFU for L . monocytogenes . Both bacteria could survive on strawberry surfaces, but their recovered levels decreased with the increase of storage time at 4 degrees C for both spot and dip inoculation methods . Dip inoculation generally had a lower recovery than spot inoculation . An ideal protocol to recover and enumerate E . coli O157:H7 and L . monocytogenes from strawberries involved shaking and washing samples with 100 ml of PBS for 15 min at 22 degrees C coupled with a MTP enumeration method.

Mol Genet Genomics . 2004 Nov 10; {Epub ahead of print}
Listeria monocytogenes infection-dependent transfer of exogenously added DNA to fibroblast COS-1 cells; Stritzker J et al.; The addition of double-stranded circular or linear DNA encoding EGFP (the Enhanced Green Fluorescent Protein) to a Listeria -containing infection medium resulted in up to 8.6% COS-1 cells expressing the reporter protein . The transfer of naked DNA into host cells upon infection by Listeria was found to be dependent on the ability of the bacteria to synthesize internalins and listeriolysin . Since no binding of DNA to the bacterial cells was detected, DNA uptake seems to be the consequence of the simultaneous entry of infection medium, and thus of naked DNA, via the phagosomes induced by the bacterium to facilitate its own entry into the host cells.

Lett Appl Microbiol, 2004, 39(6), 528 - 32
Differential inactivation of Listeria monocytogenes by D- and L-lactic acid; Gravesen A et al.; AIMS: To determine inactivation of Listeria monocytogenes by the two lactic acid isomers . METHODS AND RESULTS: The survival of four strains with varying sensitivity to acid was determined following treatment with L- or D-lactic acid at 100 mmol l(-1) (pH 3.7) or HCl at pH 3.37 . There was some, but not complete, similarity in the relative sensitivity of the four strains to the two types of acid . All strains were most sensitive to D-lactic acid, which gave 0.6-2.2 log units greater reduction than L-lactic acid midway in the inactivation curves . Even very low concentrations of the two isomers had an immediate effect on pH(i) which was identical for the two isomers . CONCLUSIONS: The results show that L . monocytogenes is more sensitive to D- than to L-lactic acid; however, this difference is less than the strain variation in L-lactic acid sensitivity . SIGNIFICANCE AND IMPACT OF THE STUDY: This work has implications for the application of lactic acid for food preservation as well as for the understanding of the antibacterial mechanisms of weak organic acids.

Lett Appl Microbiol, 2004, 39(6), 483 - 9
Glycopeptide-resistance transferability from vancomycin-resistant enterococci of human and animal source to Listeria spp; de Niederhausern S et al.; AIMS: The glycopeptide-resistance transferability from vancomycin-resistant enterococci (VRE) of clinical and animal origin to different species of Listeria was investigated . METHODS AND RESULTS: Of 36 matings, performed on membrane filter, the glycopeptide resistance was successfully transferred in six attempts, five with donors of animal origin and only one with donors from clinical source . The acquired glycopeptide resistance in Listeria transconjugants was confirmed by the presence of the conjugative plasmid band and by the amplification of the 732-bp fragment of vanA gene in transferred plasmids . CONCLUSIONS: Despite the lower number of bacteria used in this study, the source of enterococci influenced the outcome of mating . Moreover transferred VanA plasmid induced a different expression in Listeria transconjugants, suggesting that gene expression might be influenced by species affiliation of recipients . SIGNIFICANCE AND IMPACT OF THE STUDY: Our data strengthen the opinion that enterococci are an important source of resistance genes for Listeria via the transfer of movable genetic elements . As these strains are commonly found in the same habitats, a horizontal spread of glycopeptide resistance in Listeria spp . could be possible.

J Appl Microbiol, 2004, 97(6), 1281 - 8
Relationship between inactivation kinetics of a Listeria monocytogenes suspension by chlorine and its chlorine demand; Virto R et al.; AIMS: Chlorine demand by Listeria monocytogenes cells and inactivation of L . monocytogenes by chlorine (0.6-1.0 mg l(-1)) at different temperatures (4, 20 and 30 degrees C) have been investigated in a batch reactor . METHODS AND RESULTS: Chlorine demand depended on the microbial concentration and was independent on the initial chlorine concentration and temperature . Chlorine decay was modelled by the addition of two first-order decay equations . Inactivation of L . monocytogenes by chlorine depended on the initial microbial concentration, initial chlorine concentration and temperature . A mathematical model based on a biphasic inactivation properly described survival curves of L . monocytogenes and a tertiary model was developed that satisfactorily predicted the inactivation of L . monocytogenes by different concentrations of initial chlorine at different temperatures . CONCLUSIONS: Both available chlorine decay and inactivation of L . monocytogenes by chlorine were biphasic and can be modelled by a two-term exponential model . SIGNIFICANCE AND IMPACT OF THE STUDY: The biphasic nature of survival curves of L . monocytogenes did not reflect the effect of a change of available chlorine concentration during the treatment . The microbial inactivation was caused by successive reactions that occur after the consumption of the chlorine by the bacterial cell components.

Int J Food Microbiol, 2004 Dec 15, 97(2), 215 - 9
Factors affecting the antilisterial effects of nisin in milk; Bhatti M et al.; The ability of Listeria monocytogenes to proliferate in milk and the antilisterial activities of nisin are well documented . Although milk fat was reported to reduce the antimicrobial activities of nisin, there is little information on the influence of milk fat on the antilisterial activities of nisin in refrigerated milk, and whether pasteurization and homogenization influence these activities . Fresh, pasteurized, and homogenized milk samples (0.1%, 2.0%, and 3.5% fat) were treated with nisin (0-500 IU/ml) and challenged with 10(4) CFU/ml L . monocytogenes strain Scott A . The organism was most sensitive to nisin in skim milk, showing rapid decline in cell numbers to <10 CFU/ml after 12 days at 5 degrees C following treatment with 250 IU/ml . An initial decline in cell numbers in 2% and whole milk was followed by regrowth of the organism . Loss of the antilisterial effects of nisin was confirmed in homogenized whole milk, whether raw or pasteurized, but not in raw or pasteurized whole milk that was not homogenized . Tween 80, a nonionic emulsifier, partially counteracted the loss of the antilisterial activity of nisin, whereas lecithin, an anionic emulsifier, had no effect . These results demonstrate that the chemical composition and treatment of foods may play an important role in the antilisterial effects of nisin.

Scand J Immunol, 2004 Nov, 60(5), 437 - 48
Interferon-gamma mediates neuronal killing of intracellular bacteria; Jin Y et al.; Neurons can be targets for microbes, which could kill the neurons . Just in reverse, we, in this study, report that bacteria can be killed when entering a neuron . Primary cultures of foetal mouse hippocampal neurons and a neuronal cell line derived from mouse hypothalamus were infected by Listeria monocytogenes . Treatment with interferon-gamma (IFN-gamma) did not affect bacterial uptake, but resulted in increased killing of intracellular bacteria, whereas the neuronal cell remained intact . The IFN-gamma-mediated bacterial killing was mapped to the neuronal cytosol, before listerial actin tail formation . Treatment with IFN-gamma induced phosphorylation of the transcription factor STAT-1 in neurons and IFN-gamma-mediated listerial killing was not observed in STAT-1(-/-) neurons or neurons treated with IFN regulatory factor-1 antisense oligonucleotides . IFN-gamma-treated neuronal cells showed increased levels of inducible nitric oxide synthase (iNOS) mRNA, and antisense iNOS oligonucleotides hampered the bacterial killing by neurons upon IFN-gamma treatment . This novel neuronal function - i.e., that of a microbe killer - could play a crucial role in the control of infections in the immuno-privileged nervous system.

J Clin Microbiol, 2004 Nov, 42(11), 5270 - 6
Selective discrimination of Listeria monocytogenes epidemic strains by a mixed-genome DNA microarray compared to discrimination by pulsed-field gel electrophoresis, ribotyping, and multilocus sequence typing; Borucki MK et al.; Listeria monocytogenes can cause serious illness in humans, and subsequent epidemiological investigation requires molecular characterization to allow the identification of specific isolates . L . monocytogenes is usually characterized by serotyping and is subtyped by using pulsed-field gel electrophoresis (PFGE) or ribotyping . DNA microarrays provide an alternative means to resolve genetic differences among isolates, and unlike PFGE and ribotyping, microarrays can be used to identify specific genes associated with strains of interest . Twenty strains of L . monocytogenes representing six serovars were used to generate a shotgun library, and subsequently a 629-probe microarray was constructed by using features that included only potentially polymorphic gene probe sequences . Fifty-two strains of L . monocytogenes were genotyped by using the condensed array, including strains associated with five major listeriosis epidemics . Cluster analysis of the microarray data grouped strains according to phylogenetic lineage and serotype . Most epidemiologically linked strains were grouped together, and subtyping resolution was the same as that with PFGE (using AscI and ApaI) and better than that with multilocus sequence typing (using six housekeeping genes) and ribotyping . Additionally, a majority of epidemic strains were grouped together within phylogenetic Division I . This epidemic cluster was clearly distinct from the two other Division I clusters, which encompassed primarily sporadic and environmental strains . Discriminant function analysis allowed identification of 22 probes from the mixed-genome array that distinguish serotypes and subtypes, including several potential markers that were distinct for the epidemic cluster . Many of the subtype-specific genes encode proteins that likely confer survival advantages in the environment and/or host.

Microbiology, 2004 Nov, 150(Pt 11), 3843 - 55
sigmaB-dependent gene induction and expression in Listeria monocytogenes during osmotic and acid stress conditions simulating the intestinal environment; Sue D et al.; Listeria monocytogenes must overcome a variety of stress conditions in the host digestive tract to cause foodborne infections . The alternative sigma factor sigma(B), encoded by sigB, is responsible for regulating transcription of several L . monocytogenes virulence and stress-response genes, including genes that contribute to establishment of gastrointestinal infections . A quantitative RT-PCR assay was used to measure mRNA transcript accumulation for the virulence genes inlA and bsh, the stress-response genes opuCA and lmo0669 (encoding a carnitine transporter and an oxidoreductase, respectively) and the housekeeping gene rpoB . Assays were conducted on mid-exponential phase L . monocytogenes cells exposed to conditions reflecting osmotic (0.3 M NaCl) or acid (pH 4.5) conditions typical for the human intestinal lumen . In exponential-phase cells, as well as under osmotic and acid stress, inlA, opuCA and bsh showed significantly lower absolute expression levels in a L . monocytogenes DeltasigB null mutant compared to wild-type . A statistical model that normalized target gene expression relative to rpoB showed that accumulation of inlA, opuCA and bsh transcripts was significantly increased in the wild-type strain within 5 min of acid and osmotic stress exposure; lmo0669 transcript accumulation increased significantly only after acid exposure . It was concluded that sigma(B) is essential for rapid induction of the tested stress-response and virulence genes under conditions typically encountered during gastrointestinal passage . As inlA, bsh and opuCA are critical for gastrointestinal infections in animal models, the data also suggest that sigma(B) contributes to the ability of L . monocytogenes to cause foodborne infections.

J Immunol, 2004 Nov 15, 173(10), 5918 - 22
Cutting edge: immunity and IFN-gamma production during Listeria monocytogenes infection in the absence of T-bet; Way SS et al.; The T-box transcription factor T-bet is an important regulator of IFN-gamma production in all cell types and is considered to be essential for the generation of CD4 Th1 T cells . IFN-gamma in turn plays a critical role in immunity to many infectious agents . In this study, we demonstrate that T-bet is not required for host resistance to primary Listeria monocytogenes (LM) infection . In the innate immune phase, control of LM replication, serum IFN-gamma, and numbers of IFN-gamma-producing NK cells were similar in T-bet-deficient and control mice . In the adaptive immune phase, there was no defect in bacterial clearance or in the numbers of LM-specific IFN-gamma-producing CD8 T cells in T-bet-deficient mice and only a modest, although significant, reduction in the numbers of Th1 CD4 T cells and IFN-gamma secretion by CD4 T cells . Thus, host resistance and the generation of IFN-gamma-producing cells in response to LM infection are not substantially compromised in the absence of T-bet.

Langmuir, 2004 Nov 9, 20(23), 10283 - 7
Iron and cobalt oxide and metallic nanoparticles prepared from ferritin; Hosein HA et al.; Metallic Fe and Co and Fe- and Co-based oxide nanoparticles were prepared by a novel method utilizing the biologically relevant protein ferritin . In particular, iron and cobalt oxyhydroxide nanoparticles were assembled within horse spleen and Listeria innocua derived ferritin, respectively, in the aqueous phase . Ferritin containing either Fe or Co oxide was transferred and dried on a SiO2 support where the protein shell was removed during exposure to a highly oxidizing environment . It was also shown that the metal oxide particles could be reduced to the respective metal by heating in hydrogen . X-ray photoelectron spectroscopy was used to characterize the composition of the particles and atomic force microscopy was used to characterize the size of the nanoparticles . Depending on the Fe or Co loading and/or type of ferritin used, metallic and oxide nanoparticles could be produced within a range of 20-60 A.

Vet Rec, 2004 Oct 9, 155(15), 456 - 9
Listeria monocytogenes in horses in Iceland; Gudmundsdottir KB et al.; Twenty isolates of Listeria monocytogenes associated with five confirmed and four suspected incidents of listeriosis in horses in Iceland were characterised by serotyping, pulsed-field gel electrophoresis and ribotyping . Semiquantitative estimates of the numbers of L monocytogenes were made on faeces from horses with clinical signs of listeriosis and on grass silage fed to them . Large numbers of L monocytogenes were often found in the faeces of horses with severe signs of disease . The 20 isolates could be divided into six genotypes, each incident involving only one genotype . One serovar 1/2a genotype was associated with three confirmed incidents of listeriosis in 1991, 1993 and 1997 . In one incident, the same genotype was isolated from the organs of a horse with listeriosis and from the spoiled grass silage fed to it.

Scand J Infect Dis, 2004, 36(10), 709 - 11
Outcome of Listeria monocytogenes prosthetic valve endocarditis: as bad as it looks?
Miguel-Yanes JM, Gonzalez-Ramallo VJ, Pastor L.
In 1998 we presented 1 successfully treated case of Listeria monocytogenes prosthetic valve endocarditis and made a review of all the cases that had been published to date . We carry out an up-to-date review through Pub-Med of every case of Listeria monocytogenes prosthetic valve endocarditis; mortality rate is calculated and data from several clinical and therapeutical variables are collected; Fisher's exact test is used to identify those variables significantly associated with mortality . Four out of 23 patients died in hospital (17%); among all the variables included, only peripheral embolism (p=0.024), onset on a mechanical prosthesis (p=0.035) and having used only 1 antibiotic instead of a combination of drugs (p=0.026) were independently associated with mortality . Although the overall number of cases is too low to draw definite conclusions (n=23), mortality rate is lower than previously believed . Some variables that have traditionally been associated with a poor prognosis for endocarditides are not for the case of Listeria monocytogenes on valvular prostheses . It seems prudent to treat affected patients with a combination of ampicillin -- or vancomycin, if there is a history of beta-lactam allergy and ampicillin desensitization is not possible -- plus an aminoglycoside.

J Food Prot, 2004 Oct, 67(10), 2296 - 301
Control of Listeria monocytogenes with combined antimicrobials on beef franks stored at 4 degrees C; Uhart M et al.; Contamination of ready-to-eat meat products such as beef franks with Listeria monocytogenes has become a major concern for the meat processing industry and an important food safety issue . The objective of this study was to determine the effectiveness of combinations of antimicrobials as aqueous dipping solutions to control L . monocytogenes on vacuum-packaged beef franks stored at 4 degrees C for 3 weeks . Commercial beef franks were dipped for 5 min in three antimicrobial solutions: pediocin (6,000 AU), 3% sodium diacetate and 6% sodium lactate combined, and a combination of the three antimicrobials . Samples were then inoculated with 10(7) CFU/g of either four L . monocytogenes strains individually or a cocktail of the four strains, vacuum packaged, and stored at 4 degrees C for 3 weeks . Sampling was carried out at day 0 and after 2 and 3 weeks of storage . Individual strains, as well as the cocktail, exhibited different responses to the antimicrobial treatments . After 2 and 3 weeks of storage at 4 degrees C, pediocin-treated beef franks showed a less than 1-log reduction for all bacterial strains . Samples treated with the sodium diacetate-sodium lactate combination showed about a 1-log reduction after 2 weeks of storage for all strains and between a 1- and 2-log reduction after 3 weeks of storage, depending on the bacterial strain . When the three antimicrobials were combined, reductions ranged between 1 and 1.5 log units and 1.5 to 2.5 log units after 2 and 3 weeks of storage, respectively, at 4 degrees C . These results indicate that the use of combined antimicrobial solutions for dipping treatments is more effective at inhibiting L . monocytogenes than treatments using antimicrobials such as pediocin separately.

J Food Prot, 2004 Oct, 67(10), 2212 - 7
Improved quantitative recovery of Listeria monocytogenes from stainless steel surfaces using a one-ply composite tissue; Vorst KL et al.; Four sampling devices, a sterile environmental sponge (ES), a sterile cotton-tipped swab (CS), a sterile calcium alginate fiber-tipped swab (CAS), and a one-ply composite tissue (CT), were evaluated for quantitative recovery of Listeria monocytogenes from a food-grade stainless steel surface . Sterile 304-grade stainless steel plates (6 by 6 cm) were inoculated with approximately 106 CFU/cm2 L . monocytogenes strain Scott A and dried for 1 h . The ES and CT sampling devices were rehydrated in phosphate buffer solution . After plate swabbing, ES and CT were placed in 40 ml of phosphate buffer solution, stomached for 1 min and hand massaged for 30 s . Each CS and CAS device was rehydrated in 0.1% peptone before swabbing . After swabbing, CS and CAS were vortexed in 0.1% peptone for 1 min . Samples were spiral plated on modified Oxford agar with modified Oxford agar Rodac Contact plates used to recover any remaining cells from the stainless steel surface . Potential inhibition from CT was examined in both phosphate buffer solution and in a modified disc-diffusion assay . Recovery was 2.70, 1.34, and 0.62 log greater using CT compared with ES, CS, and CAS, respectively, with these differences statistically significant (P < 0.001) for ES and CT and for CAS, CS, and CT (P < 0.05) . Rodac plates were typically overgrown following ES, positive after CS and CAS, and negative after CT sampling . CT was noninhibitory in both phosphate buffer solution and the modified disc-diffusion assay . Using scanning electron microscopy, Listeria cells were observed on stainless steel plates sampled with each sampling device except CT . The CT device, which is inexpensive and easy to use, represents a major improvement over other methods in quantifying L . monocytogenes on stainless steel surfaces and is likely applicable to enrichment of environmental samples.

J Food Prot, 2004 Oct, 67(10), 2205 - 11
Survival, growth, and thermal resistance of Listeria monocytogenes in products containing peanut and chocolate; Kenney SJ et al.; Outbreaks of listeriosis associated with the consumption of ready-to-eat foods have raised interest in determining growth, survival, and inactivation characteristics of Listeria monocytogenes in a wide range of products . A study was undertaken to determine the thermal tolerance of L . monocytogenes in a peanut-based beverage (3.1% fat), whole-fat (3.5%) milk, wholefat (4.0%) and reduced-fat (1.0%) chocolate milk, a chocolate-peanut spread (39% fat), and peanut butter (53% fat) . The D60 degrees C value (decimal reduction time at 60 degrees C) in peanut beverage (3.2 min) was not significantly different (P > 0.05) than the D60 degrees C value in whole-fat milk (3.3 min) or whole-fat chocolate milk (4.5 min) but significantly lower (P < or = 0.05) than the D60 degrees C value in reduced-fat chocolate milk (5.9 min) . The pathogen was significantly more resistant to heat when enmeshed in chocolate-peanut spread (water activity {aw} of 0.46; D60 degrees C = 37.5 min) and peanut butter (aw of 0.32; D60 degrees C = 26.0 min) than in liquid products . At 10 degrees C, the pathogen grew most rapidly in whole-fat chocolate milk and slowest in peanut beverage . At 22 degrees C, populations increased significantly within 12 and 16 h in whole-fat milk and reduced-fat chocolate milk, respectively, and within 8 h in whole-fat chocolate milk and peanut beverage . Initial populations (3.37 to 4.42 log CFU/g) of L . monocytogenes in chocolate-peanut spread and peanut butter adjusted to an aw of 0.33 and 0.65 declined, but the pathogen was not eliminated during a 24-week period at 20 degrees C . Survival was enhanced at reduced aw . Results indicate that a pasteurization process similar to that used for full-fat milk would be adequate to ensure the destruction of L . monocytogenes in peanut beverage . The pathogen survives for at least 24 weeks in chocolate-peanut spread and peanut butter at an aw range that encompasses that found in these products.

J Food Prot, 2004 Oct, 67(10), 2195 - 204
Modeling the growth boundary of Listeria monocytogenes in ready-to-eat cooked meat products as a function of the product salt, moisture, potassium lactate, and sodium diacetate concentrations; Legan JD et al.; A central composite response surface design was used to determine the time to growth of Listeria monocytogenes as a function of four continuous variables: added sodium chloride (0.8 to 3.6%), sodium diacetate (0 to 0.2%), potassium lactate syrup (60% {wt/wt}; 0.25 to 9.25%), and finished-product moisture (45.5 to 83.5%) in ready-to-eat cured meat products . The design was repeated for ready-to-eat uncured meat products giving a fifth categorical variable for cure status . Products were stored at 4 degrees C . The results were modeled using a generalized regression approach . All five main effects, six two-factor interactions, and two quadratic terms were statistically significant . The model was used to show the boundary between growth and no-growth conditions at 4 degrees C using contour plots of time to growth . It was validated using independent challenge studies of cured and uncured products . Generally, the model predicted well, particularly for cured products, where it will be useful for establishing conditions that prevent the growth of L . monocytogenes . For uncured products, there was good agreement overall between predicted and observed times to growth, but the model is less thoroughly validated than for cured products . The model should initially only be used for screening of formulations likely to prevent growth of Listeria monocytogenes in uncured products, with recommendations subject to confirmation by challenge studies.

Toxicol Appl Pharmacol, 2004 Nov 1, 200(3), 206 - 18
Suppression in lung defense responses after bacterial infection in rats pretreated with different welding fumes; Antonini JM et al.; Epidemiology suggests that inhalation of welding fumes increases the susceptibility to lung infection . The effects of chemically distinct welding fumes on lung defense responses after bacterial infection were compared . Fume was collected during gas metal arc (GMA) or flux-covered manual metal arc (MMA) welding using two consumable electrodes: stainless steel (SS) or mild steel (MS) . The fumes were separated into water-soluble and -insoluble fractions . The GMA-SS and GMA-MS fumes were found to be relatively insoluble, whereas the MMA-SS was highly water soluble, with the soluble fraction comprised of 87% Cr and 11% Mn . On day 0, male Sprague-Dawley rats were intratracheally instilled with saline (vehicle control) or the different welding fumes (0.1 or 2 mg/rat) . At day 3, the rats were intratracheally inoculated with 5 x 10(3) Listeria monocytogenes . On days 6, 8, and 10, left lungs were removed, homogenized, cultured overnight, and colony-forming units were counted to assess pulmonary bacterial clearance . Bronchoalveolar lavage (BAL) was performed on right lungs to recover phagocytes and BAL fluid to measure the production of nitric oxide (NO) and immunomodulatory cytokines, including tumor necrosis factor-alpha (TNF-alpha), interleukin (IL)-2, IL-6, and IL-10 . In contrast to the GMA-SS, GMA-MS, and saline groups, pretreatment with the highly water soluble MMA-SS fume caused significant body weight loss, extensive lung damage, and a dramatic reduction in pulmonary clearance of L . monocytogenes after infection . NO concentrations in BAL fluid and lung immunostaining of inducible NO synthase were dramatically increased in rats pretreated with MMA-SS before and after infection . MMA-SS treatment caused a significant decrease in IL-2 and significant increases in TNF-alpha, IL-6, and IL-10 after infection . In conclusion, pretreatment with MMA-SS increased production of NO and proinflammatory cytokines (TNF-alpha and IL-6) after infection, which are likely responsible for the elevation in lung inflammation and injury . In addition, MMA-SS treatment reduced IL-2 (involved in T cell proliferation) and enhanced IL-10 (involved in inhibiting macrophage function) after bacterial infection, which might result in a possible suppression in immune response and an increase in susceptibility to infection.

Infect Immun, 2004 Nov, 72(11), 6418 - 25
Listeria monocytogenes-based antibiotic resistance gene-free antigen delivery system applicable to other bacterial vectors and DNA vaccines; Verch T et al.; Plasmids represent a powerful tool to rapidly introduce genes into bacteria and help them reach high expression levels . In vaccine development, with live vaccine vectors, this allows greater flexibility and the ability to induce larger antigen amounts through multiple gene copies . However, plasmid retention often requires antibiotic resistance markers, the presence of which has been discouraged in clinical applications by the Food and Drug Administration . Therefore, we developed a Listeria monocytogenes-Escherichia coli shuttle plasmid that is retained by complementation of D-alanine racemase-deficient mutant strains both in vitro and in vivo . Our technology potentially allows the production of antibiotic resistance marker-free DNA vaccines as well as bacterial vaccine vectors devoid of engineered antibiotic resistances . As a proof of concept, we applied the D-alanine racemase complementation system to our Listeria cancer vaccine platform . With a transplantable tumor model, we compared the efficacy of the new Listeria vector to that of an established vector containing a conventional plasmid carrying a tumor-specific antigen . Both vaccine vector systems resulted in long-term regression of established tumors, with no significant difference between them . Thus, the Listeria vaccine vector presented here potentially complies with Food and Drug Administration regulations and could be developed further for clinical use.

Res Microbiol, 2004 Nov, 155(9), 741 - 6
Species-specific PCR determination of Listeria seeligeri; Liu D et al.; Listeria seeligeri is a non-pathogenic bacterium coming under the genus Listeria . As this bacterium resembles other Listeria species such as L . monocytogenes and L . ivanovii that are pathogenic to man and animals, it is important that rapid and precise identification techniques be available for L . seeligeri in cases where such determination is desirable . A specific molecular test on the basis of a uniquely present gene region in L . seeligeri will be of particular value under the circumstances . In this report, after comparative screening of genomic DNA from six Listeria species by dot blot hybridization, we isolated one L . seeligeri-specific clone (lse24-315) that contains an insert of 1538 bp . Using primers (lse24-315F and lse24-315R) derived from this clone, we showed that a specific PCR product of 375 bp was generated from genomic DNA of L . seeligeri strains only, but not of other Listeria species or common bacteria . Therefore, the PCR employing primers lse24-315F and lse24-315R provides a rapid, sensitive and specific method for distinguishing L . seeligeri from other Listeria and common bacteria.

J Immunol, 2004 Nov 1, 173(9), 5679 - 87
Duration of infection and antigen display have minimal influence on the kinetics of the CD4+ T cell response to Listeria monocytogenes infection; Corbin GA et al.; The T cell response to infection consists of clonal expansion of effector cells, followed by contraction to memory levels . It was previously thought that the duration of infection determines the magnitude and kinetics of the T cell response . However, recent analysis revealed that transition between the expansion and contraction phases of the Ag-specific CD8+ T cell response is not affected by experimental manipulation in the duration of infection or Ag display . We studied whether the duration of infection and Ag display influenced the kinetics of the Ag-specific CD4+ T cell response to Listeria monocytogenes (LM) infection . We found that truncating infection and Ag display with antibiotic treatment as early as 24 h postinfection had minimal impact on the expansion or contraction of CD4+ T cells; however, the magnitudes of the Ag-specific CD4+ and CD8+ T cell responses were differentially affected by the timing of antibiotic treatment . Treatment of LM-infected mice with antibiotics at 24 h postinfection did not prevent generation of detectable CD4+ and CD8+ memory T cells at 28 days after infection, vigorous secondary expansion of these memory T cells, or protection against a subsequent LM challenge . These results demonstrate that events within the first few days of infection stimulate CD4+ and CD8+ T cell responses that are capable of carrying out the full program of expansion and contraction to functional memory, independently of prolonged infection or Ag display.

J Immunol, 2004 Nov 1, 173(9), 5652 - 8
Reduced apoptosis and ameliorated listeriosis in TRAIL-null mice; Zheng SJ et al.; Listeriosis is an infectious disease caused by the bacterium Listeria monocytogenes . Although it is well recognized that apoptosis plays a critical role in the pathogenesis of the disease, the molecular mechanisms of cell death in listeriosis remain to be established . We report in this study that mice deficient in TRAIL were partially resistant to primary listeriosis, and blocking TRAIL with a soluble death receptor 5 markedly ameliorated the disease . The numbers of Listeria in the liver and spleen of TRAIL+/+ mice were 10-100 times greater than those in TRAIL-/- mice following primary Listeria infection . This was accompanied by a significant increase in the survival rate of TRAIL-/- mice . Lymphoid and myeloid cell death was significantly inhibited in TRAIL-/- mice, which led to marked enlargement of the spleen . These results establish a critical role for TRAIL in apoptosis during listeriosis.

J Immunol, 2004 Nov 1, 173(9), 5644 - 51
Cross-presentation of Listeria-derived CD8 T cell epitopes requires unstable bacterial translation products; Janda J et al.; Presentation of bacteria-derived CD8 T cell epitopes by dendritic cells (DC) requires either their direct infection or that DC acquire and cross-present Ags from other infected cells . We found that cross-presentation of Listeria monocytogenes-derived CD8 T cell epitopes was much stronger than direct Ag presentation by infected murine DC . Cross-presentation of Listeria-derived CD8 T cell epitopes showed unique physiological requirements . It was dependent upon the delivery of unstable bacterial translation products by infected, but still viable, Ag donor cells . Cross-presentation was enhanced both when unstable translation products in infected Ag donor cells were protected from proteasomal degradation and when the production of misfolded bacterial proteins was increased . The requirement of unstable translation products for cross-presentation may represent a novel pathway that functions to focus the CD8 T cell response toward epitopes derived from newly synthesized proteins.

Biochem Soc Trans, 2004 Nov, 32(Pt 5), 712 - 4
Lipid rafts clustering and signalling by listeriolysin O; Gekara NO et al.; Listeriolysin O, the major virulent determinant of Listeria monocytogenes, is known for forming pores on cholesterol-rich membranes . In the present study, we reveal its other facet, rafts clustering . By immunofluorescence microscopy, we show that the glycosylphosphatidylinositol-anchored proteins CD14 and CD24, which normally exhibit uniform distribution on J774 cells, undergo clustering upon treatment with LLO . The non-raft marker transferrin receptor is unaffected by such treatment . Rafts clustering might explain the induction of tyrosine phosphorylation observed on LLO-treated cells.

J AOAC Int, 2004 Sep-Oct, 87(5), 1123 - 32
Evaluation of VIDAS listeria monocytogenes II (LMO2) immunoassay method for the detection of Listeria monocytogenes in foods: collaborative study; Silberiagel KM et al.; A multilaboratory study was conducted to compare the VIDAS Listeria monocytogenes II (LMO2) immunoassay and the standard cultural methods for the detection of Listeria monocytogenes in foods . Five food types-vanilla ice cream, brie cheese, cooked roast beef, frozen green beans, and frozen tilapia fish-at 3 levels were analyzed by each method . A total of 26 laboratories representing government and industry participated . In this study, 1404 test portions were analyzed of which 1152 were used in the statistical analysis . There were 448 positive by the VIDAS LMO2 assay and 457 positive by the standard culture methods . A chi2 analysis of each of the 5 food types, at the 3 inoculation levels tested, was performed . The resulting chi2 value, 0.36, indicates that overall, there are no statistical differences between the VIDAS LMO2 assay and the standard methods at the 5% level of significance.

Proc Natl Acad Sci U S A, 2004 Oct 26, 101(43), 15440 - 5 Epub 2004 Oct 15.
A totally synthetic vaccine of generic structure that targets Toll-like receptor 2 on dendritic cells and promotes antibody or cytotoxic T cell responses; Jackson DC et al.; A simple generic peptide-based vaccine structure that targets Toll-like receptor 2-expressing dendritic cells and causes their activation is described . The vaccines are totally synthetic, serve as their own adjuvant, and are composed of (i) a single helper T cell epitope, (ii) a target epitope that is either recognized by CD8+ T cells or B cells, and (iii) a Toll-like receptor 2-targeting lipid moiety, S-{2,3-bis(palmitoyloxy)propyl}cysteine, that is situated between the peptide epitopes to form a branched configuration . The different CD8+ T cell epitopes examined were from (i) influenza virus, (ii) the intracellular bacterium Listeria monocytogenes, and (iii) ovalbumin as a model tumor antigen . Vaccines containing a B cell epitope from gastrin or luteinizing hormone-releasing hormone as a B cell epitope were also examined for their ability to elicit antibody against the parent hormones . Each of the vaccines was capable of inducing either CD8+ T cell or antibody-mediated immune responses . The lipidated vaccines, but not the nonlipidated vaccines, were able to mediate protection against viral or bacterial infection and mediate prophylactic and therapeutic anticancer activity . The two hormone-based vaccines induced high antibody titers, which in the case of luteinizing hormone-releasing hormone resulted in abrogation of reproductive function . These results highlight the utility of simple, totally synthetic, epitope-based vaccines.

Annu Rev Microbiol, 2004, 58, 587 - 610
Molecular determinants of Listeria monocytogenes virulence; Dussurget O et al.; Listeria monocytogenes is the etiological agent of listeriosis, a severe human foodborne infection characterized by gastroenteritis, meningitis, encephalitis, abortions, and perinatal infections . This gram-positive bacterium is a facultative intracellular pathogen that induces its own uptake into nonphagocytic cells and spreads from cell to cell using an actin-based motility process . This review covers both well-established and recent advances in the characterization of L . monocytogenes virulence determinants and their role in the pathophysiology of listeriosis.

J Cancer Res Clin Oncol, 2005 Jan, 131(1), 49 - 59 Epub 2005 Jan.
Listeria monocytogenes produces a pro-invasive factor that signals via ErbB2/ErbB3 heterodimers; Oliveira MJ et al.; PURPOSE . We have previously demonstrated that conditioned medium from bacteria, some of which were isolated from the colon of cancer patients, stimulate cancer cell invasion in vitro through a 13-mer beta-casein-derived peptide . Since invasion signalling pathways are coordinated by the balance between protein kinases and phosphatases, we investigated the effect of conditioned medium from bacteria on the overall cellular tyrosine phosphorylation . METHODS . The tyrosine phosphorylation level of HCT-8/E11 human colon cancer cells treated with the pro-invasive conditioned medium of Listeria, prepared on top of collagen type I gels (CM(Coll) Listeria/TSB), were analysed by means of immunoprecipitation and Western blot, with specific anti-phosphotyrosine antibodies . RESULTS . We demonstrated that CM(Coll) Listeria/TSB increases the tyrosine phosphorylation level of ErbB2 and ErbB3, members of the epidermal growth factor receptor (EGFR) family, and the association between ErbB3 and the phosphatidylinositol 3-kinase (PI3K) regulatory subunit (p85alpha) . CM(Coll) Listeria/TSB-stimulated ErbB3 tyrosine phosphorylation and cancer cell invasion were independent from EGFR expression and activity but dependent on ErbB2 activity . CONCLUSIONS . The interaction between Listeria and collagen type I produces, next to the 13-mer peptide, at least another pro-invasive factor that signals via ErbB2/ErbB3 heterodimers.

Ophthalmologe . 2004 Oct 7; {Epub ahead of print}
{Listeria endophthalmitis.}; Berger E et al.; Listeria monocytogenes is a rare cause of endogenous endophthalmitis . During the last 20 years about 30 cases have been published, all of which showed similar clinical features and a profound visual loss mainly owing to delayed diagnosis . This case report is about an otherwise healthy 41-year-old woman whose diagnosis was established 17 days after the onset of symptoms by microbiological cultures . Under sufficient therapy signs of local inflammation disappeared and intraocular pressure decreased . Pars plana vitrectomy was necessary; although post-surgery complications developed, the result was complete recovery of visual acuity.

Cancer Immunol Immunother . 2004 Oct 6; {Epub ahead of print}
Tumor sensitivity to IFN-gamma is required for successful antigen-specific immunotherapy of a transplantable mouse tumor model for HPV-transformed tumors; Dominiecki ME et al.; Purpose: Many human tumors lose responsiveness to IFN-gamma, providing a possible mechanism for the tumor to avoid immune recognition and destruction . Here we investigate the importance of tumor responsiveness to IFN-gamma in the successful immunotherapy of TC1 tumors that were immortalized with human papillomavirus proteins E6 and E7 . Methods: To investigate the role of IFN-gamma in vivo, we constructed a variant of TC1, TC1.mugR, that is unresponsive to IFN-gamma due to overexpression of a dominant negative IFN-gamma receptor . Results: Using recombinant Listeria monocytogenes that express HPV-16 E7 (Lm-LLO-E7) to stimulate an antitumor response, we demonstrate that sensitivity to IFN-gamma is required for therapeutic efficacy in that Lm-LLO-E7 induces regression of TC1 tumors but not TC1.mugR . In addition, we show that tumor sensitivity to IFN-gamma is not required for inhibition of tumor angiogenesis by Lm-LLO-E7 or for trafficking of CD4(+) and CD8(+) T cells to the tumor . However, it is required for penetration of lymphocytes into the tumor mass in vivo . Conclusions: Our findings identify a role for IFN-gamma in immunity to TC1 tumors and show that loss of tumor responsiveness to IFN-gamma poses a challenge to antigen-based immunotherapy.

Cell, 2004 Oct 15, 119(2), 299 - 309
LXR-dependent gene expression is important for macrophage survival and the innate immune response; Joseph SB et al.; The liver X receptors (LXRs) are nuclear receptors with established roles in the regulation of lipid metabolism . We now show that LXR signaling not only regulates macrophage cholesterol metabolism but also impacts antimicrobial responses . Mice lacking LXRs are highly susceptible to infection with the intracellular bacteria Listeria monocytogenes (LM) . Bone marrow transplant studies point to altered macrophage function as the major determinant of susceptibility . LXR-null macrophages undergo accelerated apoptosis when challenged with LM and exhibit defective bacterial clearance in vivo . These defects result, at least in part, from loss of regulation of the antiapoptotic factor SPalpha, a direct target for regulation by LXRalpha . Expression of LXRalpha or SPalpha in macrophages inhibits apoptosis in the setting of LM infection . Our results demonstrate that LXR-dependent gene expression plays an unexpected role in innate immunity and suggest that common nuclear receptor pathways mediate macrophage responses to modified lipoproteins and intracellular pathogens.

Cell, 2004 Oct 15, 119(2), 149 - 51
A nuclear strike against Listeria--the evolving life of LXR; Barish GD et al.; LXRs are members of the nuclear receptor superfamily and function as master regulators of cholesterol metabolism . In the macrophage, they control cholesterol efflux and inhibit the transcription factor NF-kappaB-mediated proinflammatory responses . In this issue of Cell, discover surprising, protective functions for LXRalpha in innate immunity.

J Agric Food Chem, 2004 Oct 20, 52(21), 6585 - 91
Antimicrobial and physicochemical properties of chitosan-HPMC-based films; Moller H et al.; To prepare composite films from biopolymers with anti-listerial activity and moisture barrier properties, the antimicrobial efficiency of chitosan-hydroxy propyl methyl cellulose (HPMC) films, chitosan-HPMC films associated with lipid, and chitosan-HPMC films chemically modified by cross-linking were evaluated . In addition, the physicochemical properties of composite films were evaluated to determine their potential for food applications . The incorporation of stearic acid into the composite chitosan-HPMC film formulation decreased water sensitivity such as initial solubility in water and water drop angle . Thus, cross-linking of composite chitosan-HPMC, using citric acid as the cross-linking agent, led to a 40% reduction in solubility in water . The water vapor transfer rate of HPMC film, approximately 270 g x m(-2) x day(-1) x atm(-1), was improved by incorporating chitosan and was further reduced 40% by the addition of stearic acid and/or cross-linking . Anti-listerial activity of films was determined on solid medium by a numeration technique . Chitosan-HPMC-based films, with and without stearic acid, inhibited the growth of Listeria monocytogenes completely . On the other hand, a loss of antimicrobial activity after chemical cross-linking modification was observed . FTIR and 13C NMR analyses were then conducted in order to study a potential chemical modification of biopolymers such as a chemical reaction with the amino group of chitosan . To complete the study, the mechanical properties of composite films were determined from tensile strength assays .

Pol J Microbiol, 2004, 53(2), 75 - 88
Classes and functions of Listeria monocytogenes surface proteins; Popowska M et al.; Listeria monocytogenes is an opportunistic pathogen that causes infections collectively termed listeriosis, which are related to the ingestion of food contaminated with these gram-positive rods . The pathogenicity of L . monocytogenes is determined by the following virulence factors: listeriolysin O, protein ActA, two phospholipases C, internalins (In1A and In1B), protein CwhA and a metalloprotease . The bacterium is a model organism in studies on the pathogenesis of intracellular parasites . It is able to penetrate, multiply and propagate in various types of eukaryotic cells and is also able to overcome the three main barriers encountered in the host: the intestinal barrier, the blood-brain barrier and the placenta . Based on L . monocytogenes genome sequence analysis 133 surface proteins have been identified . In particular, the large number of proteins covalently bound to murein sets L . monocytogenes apart from other gram-positive bacteria . The ability of this pathogen to multiply in various environments as well as the possibility of its interaction with many kinds of eukaryotic cells is, in fact, made possible by the large number of surface proteins.

J Immunol, 2004 Oct 15, 173(8), 5112 - 20
The host resistance locus sst1 controls innate immunity to Listeria monocytogenes infection in immunodeficient mice; Boyartchuk V et al.; Epidemiological, clinical, and experimental approaches have convincingly demonstrated that host resistance to infection with intracellular pathogens is significantly influenced by genetic polymorphisms . Using a mouse model of infection with virulent Mycobacterium tuberculosis (MTB), we have previously identified the sst1 locus as a genetic determinant of host resistance to tuberculosis . In this study we demonstrate that susceptibility to another intracellular pathogen, Listeria monocytogenes, is also influenced by the sst1 locus . The contribution of sst1 to anti-listerial immunity is much greater in immunodeficient scid mice, indicating that this locus controls innate immunity and becomes particularly important when adaptive immunity is significantly depressed . Similar to our previous observations using infection with MTB, the resistant allele of sst1 prevents formation of necrotic infectious lesions in vivo . We have shown that macrophages obtained from sst1-resistant congenic mice possess superior ability to kill L . monocytogenes in vitro . The bactericidal effect of sst1 is dependent on IFN-gamma activation and reactive oxygen radical production by activated macrophages after infection, but is independent of NO production . It is possible that there is a single gene that controls common IFN-dependent macrophage function, which is important in the pathogenesis of infections caused by both MTB and L . monocytogenes . However, host resistance to the two pathogens may be controlled by two different polymorphic genes encoded within the sst1 locus . The polymorphic gene(s) encoded within the sst1 locus that controls macrophage interactions with the two intracellular pathogens remains to be elucidated.

Appl Environ Microbiol, 2004 Oct, 70(10), 6299 - 301
Rapid quantitative detection of Listeria monocytogenes in meat products by real-time PCR; Rodriguez-Lazaro D et al.; We describe a quick and simple method for the quantitative detection of Listeria monocytogenes in meat products . This method is based on filtration, Chelex-100-based DNA purification, and real-time PCR . It can detect as few as 100 CFU/g and quantify as few as 1,000 CFU/g, with excellent accuracy compared to that of the plate count method . Therefore, it is a promising alternative for the detection of L . monocytogenes in meat products.

Appl Environ Microbiol, 2004 Oct, 70(10), 5833 - 41
Listeria monocytogenes isolates from foods and humans form distinct but overlapping populations; Gray MJ et al.; A total of 502 Listeria monocytogenes isolates from food and 492 from humans were subtyped by EcoRI ribotyping and PCR-restriction fragment length polymorphism analysis of the virulence gene hly . Isolates were further classified into genetic lineages based on subtyping results . Food isolates were obtained through a survey of selected ready-to-eat food products in Maryland and California in 2000 and 2001 . Human isolates comprised 42 isolates from invasive listeriosis cases reported in Maryland and California during 2000 and 2001 as well as an additional 450 isolates from cases that had occurred throughout the United States, predominantly from 1997 to 2001 . Assignment of isolates to lineages and to the majority of L . monocytogenes subtypes was significantly associated with the isolate source (food or human), although most subtypes and lineages included both human and food isolates . Some subtypes were also significantly associated with isolation from specific food types . Tissue culture plaque assay characterization of the 42 human isolates from Maryland and California and of 91 representative food isolates revealed significantly higher average infectivity and cell-to-cell spread for the human isolates, further supporting the hypothesis that food and human isolates form distinct populations . Combined analysis of subtype and cytopathogenicity data showed that strains classified into specific ribotypes previously linked to multiple human listeriosis outbreaks, as well as those classified into lineage I, are more common among human cases and generate larger plaques than other subtypes, suggesting that these subtypes may represent particularly virulent clonal groups . These data will provide a framework for prediction of the public health risk associated with specific L . monocytogenes subtypes.

J Bacteriol, 2004 Oct, 186(20), 6721 - 7
Flagellin from Listeria monocytogenes is glycosylated with beta-O-linked N-acetylglucosamine; Schirm M et al.; Glycan staining of purified flagellin from Listeria monocytogenes serotypes 1/2a, 1/2b, 1/2c, and 4b suggested that the flagellin protein from this organism is glycosylated . Mass spectrometry analysis demonstrated that the flagellin protein of L . monocytogenes is posttranslationally modified with O-linked N-acetylglucosamine (GlcNAc) at up to six sites/monomer . The sites of glycosylation are all located in the central, surface-exposed region of the protein monomer . Immunoblotting with a monoclonal antibody specific for beta-O-linked GlcNAc confirmed that the linkage was in the beta configuration, this residue being a posttranslational modification commonly observed in eukaryote nuclear and cytoplasmic proteins.

Parasitol Today, 1988 Dec, 4(12), 340 - 7
Oxidative killing of intracellular parasites mediated by macrophages; Hughes HP; An important function of macrophages is to eliminate invading pathogens, and one of their main weapons involves the generation of lethal oxygen radicals . Yet some parasites and pathogens - notably Leishmania, Toxoplasma, and Listeria and Mycobacterium - make use of macrophages as their primary cellular hosts displaying a capacity to survive the oxidative killing mechanisms of these host cells . It is now clear that more than one pathway is involved in the activation of macrophages to kill intracellular pathogens . Here, Huw Hughes discusses the biochemistry of the oxidative metabolism of macrophages, and the steps taken by parasites to survive within this hostile environment.

Nat Rev Immunol, 2004 Oct, 4(10), 812 - 23
Immune responses to Listeria monocytogenes; Pamer EG; Listeria monocytogenes is a Gram-positive bacterium that is often used to study the mammalian immune response to infection because it is easy to culture, is relatively safe to work with and causes a highly predictable infection in laboratory mice . The broad application of this mouse model has resulted in a torrent of studies characterizing the contributions of different cytokines, receptors, adaptors and effector molecules to resistance against infection with Listeria monocytogenes . These studies, which are yielding one of the most comprehensive pictures of the 'battle' between host and microorganism, are reviewed here.

J Food Prot, 2004 Sep, 67(9), 1866 - 75
Combining pediocin with postpackaging irradiation for control of Listeria monocytogenes on frankfurters; Chen CM et al.; Frankfurters, in 1-link, 5-link, or 10-link packages, were surface inoculated with a five-strain mixture of Listeria monocytogenes (3.40 or 5.20 log CFU/g) after treatment with 3,000 arbitrary units (AU) or 6,000 AU of pediocin (in ALTA 2341) per link . The frankfurters were vacuum packaged, after which the 1-link and 5-link packages were irradiated at 1.2 or 2.3 kGy and the 10-link packages were irradiated at 1.4 or 3.5 kGy . L . monocytogenes was enumerated following the treatments . Selected treatments were subsequently evaluated during storage at 4, 10, and 25 degrees C for up to 12 weeks . Combination of pediocin with postpackaging irradiation at 1.2 kGy or more was necessary to achieve a 50% reduction of L . monocytogenes on frankfurters in 1-link or 5-link packages . The combination of 6,000 AU of pediocin and irradiation at 2.3 kGy or more was effective in all package sizes for inhibition of the pathogen for 12 weeks at 4 or 10 degrees C . There was a synergistic effect between pediocin and irradiation for inhibition of L . monocytogenes . Storage at 4 degrees C enhanced the antilisterial effects of the treatment combinations, with little or no growth of the pathogen in 1-link or 5-link packages during 12 weeks of storage . In general, these treatments did not affect the sensory quality of frankfurters.

J Food Prot, 2004 Sep, 67(9), 1855 - 65
Combining pediocin (ALTA 2341) with postpackaging thermal pasteurization for control of Listeria monocytogenes on frankfurters; Chen CM et al.; Frankfurters packaged in 1-link, 5-link, or 10-link packages were surface-inoculated with a five-strain mixture of Listeria monocytogenes (3.40 or 5.20 log CFU/g) after treatments with 3,000 arbitrary units (AU) or 6,000 AU pediocin (in ALTA 2341) per link . The frankfurters were vacuum packaged, after which the packages were heated in hot water at 71, 81, or 96 degrees C for 30, 60, or 120 s . L . monocytogenes was enumerated following the treatments . Selected treatments were subsequently evaluated during storage at 4, 10, and 25 degrees C for up to 12 weeks . L . monocytogenes was reduced by all treatments, but 81 degrees C or more for at least 60 s in combination with pediocin (Pdn-6000) was necessary to achieve a 50% reduction of initial inoculations . Heat treatments were most effective for 1-link packages and least effective for 10-link packages . Little or no growth of L . monocytogenes occurred on frankfurters for 12 weeks at 4 or 10 degrees C, and for 12 days at 25 degrees C . Generally, the treatments mentioned above did not significantly (P > 0.05) affect the sensory qualities of frankfurters . Therefore, pediocin (in ALTA 2341) in combination with postpackaging thermal treatment offers an effective treatment combination for improved control of L . monocytogenes on frankfurters.

J Food Prot, 2004 Sep, 67(9), 1848 - 54
Inactivation of Listeria innocua in nisin-treated salmon (Oncorhynchus keta) and sturgeon (Acipenser transmontanus) caviar heated by radio frequency; Al-Holy M et al.; Recent regulatory concerns about the presence of the pathogen Listeria monocytogenes in ready-to-eat aquatic foods such as caviar has prompted the development of postpackaging pasteurization processes . However, caviar is heat labile, and conventional pasteurization processes affect the texture, color, and flavor of these foods negatively . In this study, chum salmon (Oncorhynchus keta, 2.5% total salt) caviar or ikura and sturgeon (Acipenser transmontanus, 3.5% total salt) caviar were inoculated with three strains of Listeria innocua in stationary phase at a level of more than 10(7) CFU/g . L innocua strains were used because they exhibit an equivalent response to L monocytogenes for many physicochemical processing treatments, including heat treatment . The products were treated by immersion in 500 IU/ml nisin solution and heat processed (an 8-D process without nisin or a 4-D process with 500 IU/ml nisin) in a newly developed radio frequency (RF; 27 MHz) heating method at 60, 63, and 65 degrees C . RF heating along with nisin acted synergistically to inactivate L . innocua cells and total mesophilic microorganisms . In the RF-nisin treatment at 65 degrees C, no surviving L . innocua microbes were recovered in sturgeon caviar or ikura . The come-up times in the RF-heated product were significantly lower compared with the water bath-heated caviar at all treatment temperatures . The visual quality of the caviar products treated by RF with or without nisin was comparable to the untreated control.

FEMS Microbiol Lett, 2004 Oct 1, 239(1), 157 - 61
Resistance of Gram-positive bacteria to nisin is not determined by lipid II levels; Kramer NE et al.; Lipid II is essential for nisin-mediated pore formation at nano-molar concentrations . We tested whether nisin resistance could result from different Lipid II levels, by comparing the maximal Lipid II pool in Micrococcus flavus (sensitive) and Listeria monocytogenes (relatively insensitive) and their nisin-resistant variants, with a newly developed method . No correlation was observed between the maximal Lipid II pool and nisin sensitivity, as was further corroborated by using spheroplasts of nisin-resistant and wild-type strains of M . flavus, which were equally sensitive to nisin.

FEMS Microbiol Lett, 2004 Oct 1, 239(1), 63 - 70
Influence of environmental conditions on the expression of virulence factors by Listeria monocytogenes and their use in species identification; Lemes-Marques EG et al.; The hemolytic, lecithinase or phosphatidylinositol-specific phospholipase C activities of Listeria monocytogenes can be used to differentiate this pathogenic bacteria from L . innocua, apathogenic, frequently isolated from environmental sources and food . However, the interpretation of these characteristics is problematic because of the variation in the expression of virulence factors by L . monocytogenes, which can be influenced by environmental conditions . We used a cheap, simple plate assay to monitor this expression in strains obtained from various sources and grown under different culture conditions . The results were increasingly significant and were obtained adding activated charcoal and different salts to the culture media, and in some cases changing the culture temperature, all with a rigorous control on the process of media sterilization.

Infect Immun, 2004 Oct, 72(10), 5622 - 9
Growth, virulence, and immunogenicity of Listeria monocytogenes aro mutants; Stritzker J et al.; Mutants of Listeria monocytogenes with deletions in genes of the common branch of the biosynthesis pathway leading to aromatic compounds were constructed as possible virulence-attenuated carrier strains for protein antigens or vaccine DNA . aroA, aroB, and in particular aroE mutants showed strongly reduced growth rates in epithelial cells and even in rich culture media . The metabolism of the aro mutants under these conditions was predominantly anaerobic . Aerobic metabolism and a wild-type growth rate were, however, regained upon the addition of vitamin K2, suggesting that the aro mutants are deficient in oxidative respiration due to the lack of menaquinone . Replication of the aro mutants in the host cell's cytosol and cell-to-cell spread were drastically slowed down, and all aro mutants showed high virulence attenuation in mice, i.e., the 50% lethal dose in BALB/c mice was increased at least 10(4)-fold for the aroA, aroB, and aroA/B mutants and >10(5)-fold for the aroE mutant compared to the parent strain . Nevertheless, mice preimmunized with aro mutant bacteria elicited good T-cell response and full protection against a subsequent challenge with the virulent wild-type strain . A total of 5 x 10(6) aroA, aroB, and aroA/B mutant bacteria were sufficient to obtain a protective T-cell response, while 5 x 10(8) aroE or aroA/E mutants were necessary to achieve comparable numbers of antigen-specific T cells . These numbers were well tolerated without causing any signs of disease, indicating that Listeria strains with deletions in genes of the basic branch of the aromatic amino acid pathway could be useful vaccine carriers for inducing T-cell immunity.

Eur J Immunol, 2004 Nov, 34(11), 3070 - 81
Expression of selectin ligands on murine effector and IL-10-producing CD4+ T cells from non-infected and infected tissues; Kretschmer U et al.; Endothelial selectins are crucial for the recruitment of leukocytes into sites of inflammation . On T cells, ligands for selectins become induced upon differentiation into the effector/memory stage . Initial in vitro studies suggested a correlation between the Th1 phenotype and ligand expression, but whether this also holds true in vivo remained uncertain . We here analyzed selectin ligands on CD4+ T cells producing IFN-gamma, IL-4 or IL-10, prototypic cytokines of the Th1, Th2 and Tr1 subset, respectively . We analyzed mice infected with influenza virus, the bacterium Listeria, and the parasites Toxoplasma (all Th1 models) or Nippostrongylus (Th2 model) . A link between the Th1 phenotype and ligand expression was not found in vivo . Surprisingly, the potentially regulatory IL-10-producing T cells displayed the highest frequency of ligand-positive cells in general . Within the inflamed tissues, the frequencies of P-selectin-binding cells increased in the dominant subset, either Th1 or Th2 . Up-regulation was also found for E-selectin ligands during influenza, but not Nippostrongylus infection . In conclusion, conditions driving T cell polarization into either Th1 or Th2 in vivo do not affect the expression of selectin ligands, but acquisition of P-selectin binding and hence migration into inflamed tissues is boosted by an inflammatory milieu.

J Immunol, 2004 Oct 1, 173(7), 4324 - 30
Intestinal epithelial antigen induces mucosal CD8 T cell tolerance, activation, and inflammatory response; Liu Z et al.; Intestinal autoimmune diseases are thought to be associated with a breakdown in tolerance, leading to mucosal lymphocyte activation perhaps as a result of encounter with bacterium-derived Ag . To study mucosal CD8(+) T cell activation, tolerance, and polarization of autoimmune reactivity to self-Ag, we developed a novel (Fabpl(4x at -132)-OVA) transgenic mouse model expressing a truncated form of OVA in intestinal epithelia of the terminal ileum and colon . We found that OVA-specific CD8(+) T cells were partially tolerant to intestinal epithelium-derived OVA, because oral infection with Listeria monocytogenes-encoding OVA did not elicit an endogenous OVA-specific MHC class I tetramer(+)CD8(+) T cell response and IFN-gamma-, IL-4-, and IL-5-secreting T cells were decreased in the Peyer's patches, mesenteric lymph nodes, and intestinal mucosa of transgenic mice . Adoptive transfer of OVA-specific CD8(+) (OT-I) T cells resulted in their preferential expansion in the Peyer's patches and mesenteric lymph nodes and subsequently in the epithelia and lamina propria but failed to cause mucosal inflammation . Thus, CFSE-labeled OT-I cells greatly proliferated in these tissues by 5 days posttransfer . Strikingly, OT-I cell-transferred Fabpl(4x at -132)-OVA transgenic mice underwent a transient weight loss and developed a CD8(+) T cell-mediated acute enterocolitis 5 days after oral L . monocytogenes-encoding OVA infection . These findings indicate that intestinal epithelium-derived "self-Ag" gains access to the mucosal immune system, leading to Ag-specific T cell activation and clonal deletion . However, when Ag is presented in the context of bacterial infection, the associated inflammatory signals drive Ag-specific CD8(+) T cells to mediate intestinal immunopathology.

Mil Med, 2004 Aug, 169(8), 600 - 3
A survey of preprocedural antiseptic mouth rinse use in Army dental clinics; Hennessy B et al.; OBJECTIVE: The objective of this project was to evaluate the use of preprocedural mouth rinses in Army dental clinics . MATERIALS AND METHODS: Three hundred six-question surveys were distributed to 10 Army dental organizations throughout the United States and Germany during the period from March 2001 to March 2002 . Two hundred fifty-four surveys were completed and returned . Simple mathematics were used to evaluate answers to the questionnaires . RESULTS: The 254 respondents included military dentists (n = 190), civilian dentists used by the military (n = 27), registered dental hygienists (n = 20), and military-trained dental hygiene technicians (n = 17) . Eighty-four and one-tenth percent of respondents (n = 216) use preprocedural rinses in their practices to prevent possible disease transmission (n = 85) or to decrease chances of postoperative infection (n = 167) . Chlorhexidine gluconate (n = 170) and phenol-based essential oil preparations (n = 84) are the most commonly used products . The perceived greatest benefits of preprocedural rinsing are to decrease oral bacterial load (38%), to decrease incidence of postoperative infection (21%), and to decrease aerosolization of bacteria (8.66%) . CONCLUSIONS: Army dental clinics make extensive use of antimicrobial preprocedural rinses . Chlorhexidine and Listerine (Warner-Lambert Consumer Healthcare, Morris Plains, NJ) are the most commonly used products . Currently available literature appears to support the use of these products in preventing or diminishing the chances of postoperative infection.

Proteomics, 2004 Oct, 4(10), 2991 - 3006
The cell wall subproteome of Listeria monocytogenes; Schaumburg J et al.; The surface subproteome of Listeria monocytogenes that includes many proteins already known to be involved in virulence and interaction with host cells has been characterized . A new method for the isolation of a defined surface proteome of low complexity has been established based on serial extraction of proteins by different salts at high concentration, and in all 55 proteins were identified by N-terminal sequencing and mass spectrometry . About 16% of these proteins are of unknown function and three proteins have no orthologue in the nonpathogenic L . innocua and might be involved in virulence mechanisms . Remarkably, a relatively high number of proteins with a function in the cytoplasmic compartment was identified in this surface proteome . These proteins had neither predicted or detectable signal peptides nor could any modification be observed except removal of the N-terminal methionine . Enolase (Lmo2455) is one of these proteins . It was shown to be present in the cell wall of the pathogen by immunoelectron microscopy and, along with heat shock factor DnaK (Lmo1473), elongation factor TU (Lmo2653), and glyceraldehyde-3-phosphate dehydrogenase (Lmo2459), it was found to be able to bind human plasminogen in overlay blots and surface plasmon resonance (SPR) experiments . The KD values of these interactions were determined by SPR measurements . The data indicate a possible role of these proteins as receptors for human plasminogen on the bacterial cell surface . The potential role of this recruitment of a host protease for extracellular invasion mechanisms is discussed.

Proteomics, 2004 Oct, 4(10), 3187 - 201
Two-dimensional electrophoresis database of Listeria monocytogenes EGDe proteome and proteomic analysis of mid-log and stationary growth phase cells; Folio P et al.; Listeria monocytogenes is the causative agent of listeriosis, one of the most significant foodborne diseases in industrialized countries . The complete genome of the L . monocytogenes EGDe strain, belonging to the serogroup 1/2a, has been sequenced and is comprised of 2853 open reading frames . The objective of the current study was to construct a two-dimensional (2-D) database of the proteome of this strain . The soluble protein fractions of the microorganism were recovered either in the mid-log or in the stationary phase of growth at 37 degrees C . These fractions were analyzed by 2-D electrophoresis (2-DE), using immobilized pH gradient strips of various pH values (3-10, 3-6, and 5-8) for the first-dimensional separations and 12.5% acrylamide gels for sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) . 201 protein spots corresponding to 126 different proteins were identified by matrix assisted laser desorption/ionization-time of flight-mass spectrometry (MALDI-TOF-MS) . The 2-DE maps presented here provide a first basis for further investigations of protein expression in L . monocytogenes . In this way, the comparison of proteome between cells in the exponential or stationary phase of growth at 37 degrees C allowed us to characterize 161 variations in protein spot intensity, of which 38 were identified . Among the differentially expressed proteins were ribosomal proteins (RpsF, RplJ, and RpmE), proteins involved in cellular metabolism (GlpD, PdhD, Pgm, Lmo1372, Lmo2696, and Lmo2743) or in stress adaptation (GroES and ferritin), a fructose-specific phosphotransferase enzyme IIB (Lmo0399) and different post-translational modified forms of listeriolysin (LLO).

J Dairy Sci, 2004 Sep, 87(9), 2803 - 12
Molecular Subtyping and Tracking of Listeria monocytogenes in Latin-Style Fresh-Cheese Processing Plants; Kabuki DY et al.; Latin-style fresh cheeses, which have been linked to at least 2 human listeriosis outbreaks in the United States, are considered to be high-risk foods for Listeria monocytogenes contamination . We evaluated L . monocytogenes contamination patterns in 3 Latin-style fresh-cheese processing plants to gain a better understanding of L . monocytogenes contamination sources in the manufacture of these cheeses . Over a 6-mo period, 246 environmental samples were collected and analyzed for L . monocytogenes using both the Food and Drug Administration (FDA) method and the Biosynth L . monocytogenes detection system (LMDS) . Finished cheese samples from the same plants (n = 111) were also analyzed by the FDA method, which was modified to include L . monocytogenes plating medium (LMPM) and the L . monocytogenes confirmatory plating medium (LMCM) used in the LMDS method . Listeria monocytogenes was detected in 6.3% of cheese and 11.0% of environmental samples . Crates, drains, and floor samples showed the highest contamination rates, with 55.6, 30.0, and 20.6% L . monocytogenes positive samples, respectively . Finished products and food contact surfaces were positive in only one plant . The FDA method showed a higher sensitivity than the LMDS method for detection of L . monocytogenes from environmental samples . The addition of LMPM and LMCM media did not further enhance the performance of the FDA method for L . monocytogenes detection from finished products . Molecular subtyping (PCR-based allelic analysis of the virulence genes actA and hly and automated ribotyping) was used to track contamination patterns . Ribotype DUP-1044A, which had previously been linked to a 1998 multistate human listeriosis outbreak in the United States, was the most commonly identified subtype (20/36 isolates) and was isolated from 2 plants . This ribotype was persistent and widespread in one factory, where it was also responsible for the contamination of finished products . We hypothesize that this ribotype may represent a clonal group with a specific ability to persist in food processing environments . While previous listeriosis outbreaks were linked to Latin-style fresh cheeses made from unpasteurized milk, the presence of this organism in pasteurized cheese products illustrates that persistent environmental contamination also represents an important source of finished product contamination.

Inhal Toxicol, 2004 May, 16(5), 311 - 7
Sensitivity to ozone, diesel exhaust particles, and standardized ambient particulate matter in rats with a listeria monocytogenes-induced respiratory infection; Steerenberg P et al.; Ambient particulate matter may increase respiratory allergic skewing of the T-cell-mediated immune response toward a T-helper-2 (Th2) response, with the consequence that the Th1 response develops less well . Successful clearing of a respiratory bacterial infection depends on an adequate Th1 immune response; therefore, the subject would not control the infection as well if exposed to particulate matter . To substantiate this hypothesis, we examined the effect of exposure to diesel exhaust particles (DEP) and urban particulate matter (EHC-93, Ottawa dust) on rats with a Listeria monocytogenes respiratory infection . Since this hypothesis has been confirmed for ozone, we used it as a positive control . Wistar rats were exposed to ozone (2 mg/m3 for 24 h/day for 7 days) and to DEP or to EHC-93 (50 microg/rat intranasally daily for 7 consecutive days) . Twenty-four hours after the last exposure, the rats were infected intratracheally with 1 x 10(6) L . monocytogenes bacteria . The number of L . monocytogenes was determined after 3, 4 and 5 days . Statistically significant increases of the number of L . monocytogenes in rats exposed to ozone were observed in the lungs and spleen at all three times . However, we found no significant differences in the numbers of bacteria that were found in rats exposed to DEP or EHC-93 compared to the saline-treated group at any of the three times . In conclusion, the results of this study do not support the hypothesis that exposure to DEP or EHC-93 reduces subsequent resistance to a respiratory infection in rats . Copyright Taylor & Francis Inc.

Syst Appl Microbiol, 2004 Aug, 27(4), 454 - 61
Genetic characterization of Listeria monocytogenes food isolates and pathogenic potential within serovars 1/2a and 1/2b; Cabrita P et al.; A total of 39 Listeria monocytogenes strains isolated from raw milk, smoked meat, chicken carcass and reference strains, belonging to serovars 1/2a, 4a, 1/2b, 3b and 4b, were analysed by RAPD and by polymorphisms of the virulent genes inlAB and iap . Ten isolates, belonging to serovars 1/2a and 1/2b and, collected from raw milk and smoked meat, were further tested for pathogenicity by IP injection into mice . The clustering of the 39 L . monocytogenes strains in 3 groups at 0.45 similarity level, based on molecular typing, was observed . Distribution of serovars in these clusters was in agreement with the proposed three Listeria monocytogenes lineages . Within serovar 1/2b, the 50% lethal dose (LD50) ranged from 8.4 x 10(4) to 1.7 x 10(6) cfu.ml(-1) . One of the serovar 1/2b strains, isolated from smoked meat, exhibited the lowest virulence potential evaluated by LD50 and by mean time to death (MTD) and, from this point of view, was completely different from the other strains . Our results suggest the existence of heterogeneity in virulence levels within serovars 1/2a and 1/2b . However, when comparing the isolates based on genotyping, virulence indicators and food origin, no relation could be assessed.

Eur J Immunol, 2004 Oct, 34(10), 2681 - 9
Expression of housekeeping and immunoproteasome subunit genes is differentially regulated in positively and negatively selecting thymic stroma subsets; Nil A et al.; The expression of housekeeping and/or immunoproteasomes in isolated thymic stroma subsets has so far not been analyzed but may have important consequences for self peptide repertoires presented by MHC class I molecules during positive and negative thymic selection . Here we determined the expression of housekeeping and immunoproteasome beta subunits and of PA28 in positively and negatively selecting stroma subsets . Positively selecting cortical thymic epithelial cells (cTEC) expressed only housekeeping but no immunoproteasome beta subunit mRNA and proteins . However, immunoproteasome beta subunits could be induced in cTEC by infection with Listeria monocytogenes or injection of IFN-gamma . In negatively selecting stroma including medullary epithelial cells and dendritic cells, incomplete and low representation of housekeeping beta subunit proteins but high and complete expression of immunoproteasome beta subunit proteins suggests absence of proper housekeeping proteasomes and predominance of immunoproteasomes . Expression of immunoproteasome beta subunits in negatively selecting stroma was independent of IFN-gamma receptor as shown in knockout (KO) mice . Absence of LMP2 altered thymic selection of the MHC class I-restricted transgenic P14 TCR in KO mice . The data suggest that negative selection may primarily involve immunoproteasome peptide repertoires and that peripheral infection may influence peptide repertoires involved in positive selection.

J Clin Periodontol, 2004 Oct, 31(10), 878 - 84
Comparative antiplaque and antigingivitis effectiveness of a chlorhexidine and an essential oil mouthrinse: 6-month clinical trial; Charles CH et al.; OBJECTIVES: The objective of this study was to compare the antiplaque and antigingivitis effectiveness and the side-effect profiles of an essential oil-containing mouthrinse and a chlorhexidine-containing mouthrinse . MATERIAL AND METHODS: One hundred and eight qualifying subjects, aged 20-57 years, were randomized into three groups: essential oil mouthrinse (ListerineAntiseptic); 0.12% chlorhexidine mouthrinse (Peridex); or 5% hydroalcohol negative control . At baseline, subjects received a complete oral soft tissue examination and scoring of the Loe-Silness gingival index (GI), Quigley-Hein plaque index (PI), Volpe-Manhold calculus index (CI), and Lobene extrinsic tooth stain index (SI) . Following a complete dental prophylaxis, subjects started rinsing twice daily with their respective mouthrinse as an adjunct to their usual mechanical oral hygiene procedures . One of the rinses on each weekday was supervised . Subjects were reexamined at 3 and 6 months . The treatment groups were compared with respect to baseline demographic and clinical variables . The primary efficacy variables were GI and PI . Intergroup differences for all clinical variables were tested at 3 and 6 months using appropriate statistical procedures . RESULTS: All of the 108 randomized subjects were evaluable at 3 months, and 107 subjects were evaluable at 6 months . There were no statistically significant differences among the three groups at baseline, with the exception that the control group PI was significantly lower than that of the essential oil group (p<0.05) and the chlorhexidine group (p<0.001), and the essential oil mouthrinse group had a significantly greater number of subjects than the control group with body region SI scores > or =1.0 (p=0.021) . At 6 months, the essential oil and chlorhexidine mouthrinses produced statistically significant (p<0.001) GI reductions of 14.0% and 18.2%, respectively, and statistically significant (p<0.001) PI reductions of 18.8% and 21.6%, respectively, compared with the control and were not statistically significantly different from each other with respect to plaque and gingivitis reduction . The chlorhexidine mouthrinse group had significantly more calculus and extrinsic tooth stain than either the essential oil mouthrinse group or the control group . CONCLUSION: This 6-month controlled clinical study demonstrated that the essential oil mouthrinse and the chlorhexidine mouthrinse had comparable antiplaque and antigingivitis activity . Insofar as side effects associated with the chlorhexidine mouthrinse may limit patient compliance, it is suggested that each product can have a distinct role in the management of patients with periodontal diseases . Copyright Blackwell Munksgaard, 2004

J Agric Food Chem, 2004 Sep 22, 52(19), 5769 - 72
Discrimination of intact and injured Listeria monocytogenes by Fourier transform infrared spectroscopy and principal component analysis; Lin M et al.; Fourier transform infrared spectroscopy (FT-IR, 4000-600 cm(-)(1)) was used to discriminate between intact and sonication-injured Listeria monocytogenes ATCC 19114 and to distinguish this strain from other selected Listeria strains (L . innocua ATCC 51742, L . innocua ATCC 33090, and L . monocytogenes ATCC 7644) . FT-IR vibrational overtone and combination bands from mid-IR active components of intact and injured bacterial cells produced distinctive "fingerprints" at wavenumbers between 1500 and 800 cm(-)(1) . Spectral data were analyzed by principal component analysis . Clear segregations of different intact and injured strains of Listeria were observed, suggesting that FT-IR can detect biochemical differences between intact and injured bacterial cells . This technique may provide a tool for the rapid assessment of cell viability and thereby the control of foodborne pathogens.

J Infect Chemother, 2004 Aug, 10(4), 242 - 4
A case of listerial meningitis treated with a regimen containing panipenem-betamipron; Hitomi S et al.; Although panipenem-betamipron, which is commercially available only in Japan, is recommended for treatment of pediatric bacterial meningitis by some experts, only a limited number of clinical studies have been reported . In the present report, we describe a 2-year-old boy with meningitis caused by Listeria monocytogenes who was treated with a regimen containing panipenem-betamipron and recovered without any apparent neurological sequelae . On the basis of our experience and previous reports, panipenem-betamipron appears to be effective for the treatment of listerial meningitis.

Proc Natl Acad Sci U S A, 2004 Sep 21, 101(38), 13832 - 7 Epub 2004 Sep 13.
Listeria-based cancer vaccines that segregate immunogenicity from toxicity; Brockstedt DG et al.; The facultative intracellular bacterium Listeria monocytogenes is being developed as a cancer vaccine platform because of its ability to induce potent innate and adaptive immunity . For successful clinical application, it is essential to develop a Listeria platform strain that is safe yet retains the potency of vaccines based on wild-type bacteria . Here, we report the development of a recombinant live-attenuated vaccine platform strain that retains the potency of the fully virulent pathogen, combined with a >1,000-fold reduction in toxicity, as compared with wild-type Listeria . By selectively deleting two virulence factors, ActA (DeltaactA) and Internalin B (DeltainlB), the immunopotency of Listeria was maintained and its toxicity was diminished in vivo, largely by blocking the direct internalin B-mediated infection of nonphagocytic cells, such as hepatocytes, and the indirect ActA-mediated infection by cell-to-cell spread from adjacent phagocytic cells . In contrast, infection of phagocytic cells was not affected, leaving intact the ability of Listeria to stimulate innate immunity and to induce antigenspecific cellular responses . Listeria DeltaactA/DeltainlB-based vaccines were rapidly cleared from mice after immunization and induced potent and durable effector and memory T-cell responses with no measurable liver toxicity . Therapeutic vaccination of BALB/c mice bearing murine CT26 colon tumor lung metastases or palpable s.c . tumors (>100 mm(3)) with recombinant Listeria DeltaactA/DeltainlB expressing an endogenous tumor antigen resulted in breaking of self-tolerance and long-term survival . We propose that recombinant Listeria DeltaactA/DeltainlB expressing human tumor-associated antigens represents an attractive therapeutic strategy for further development and testing in human clinical trials.

Int J Food Microbiol, 2004 Nov 1, 96(2), 181 - 7
Growth of Listeria monocytogenes as influenced by viscosity and water activity; Stecchini ML et al.; The effects of osmotic stress on Listeria monocytogenes growth parameters was examined in relation to the viscosity of the growth media . In low-viscosity systems, growth of L . monocytogenes in glucose-supplemented media was comparable to growth in sucrose-supplemented media . The relative lag time (RLT: the lag time divided by the generation time) responses were found to increase in the more restrictive water activity conditions . In high-viscosity systems containing polyvinylpyrrolidone (PVP), growth rate was reduced, whereas lag time showed no discernible modification . Osmotic stress in medium- and high-viscosity media supplemented with glucose resulted in approximately exponential increasing of the RLT values . Thus, the biological effects of osmotic stress on L . monocytogenes could be affected by the physical properties of the system, such as viscosity and diffusivity.

Int J Food Microbiol, 2004 Oct 1, 96(1), 85 - 96
Molecular epidemiology and disinfectant susceptibility of Listeria monocytogenes from meat processing plants and human infections; Heir E et al.; We have investigated the molecular epidemiology of Listeria monocytogenes from the meat processing industry producing cold cuts and from cases of human listeriosis by discriminative pulsed-field gel electrophoresis (PFGE) . A subset of the isolates was also investigated for susceptibility to a disinfectant based on quaternary ammonium compounds (QAC) frequently used in the meat processing industry . The purpose of this investigation was to obtain knowledge of sources, routes of contamination and genetic types of L . monocytogenes present along the production line in the meat processing industry, and to compare meat industry isolates and human isolates . Of the 222 isolates from four meat-processing plants, 200 were from two plants responsible for nearly 50% of the production of cold cuts in the Norwegian market . The strain collection included historical routinely sampled isolates (1989-2002) and isolates systematically sampled through a one year period (November 2001 to November 2002) from fresh meat and production environments in three plants . No isolates were obtained in samples from employees (throat, faeces) . Human strains included all available reported isolates from Norwegian patients in selected time periods . The L . monocytogenes PFGE data showed a large genetic heterogeneity, with isolates separated into two genetic lineages and further subdivided into 56 different PFGE profiles . Certain profiles were observed on both sides of production (before and after heat treatment) indicating contamination of end products by fresh meat or fresh meat environments . While fresh meat isolates almost exclusively grouped within lineage I, isolates from end products showed a more balanced distribution between lineages I and II . Ten profiles were common among isolates from human and meat industry . Typing of human isolates identified a previously unrecognised outbreak . Generally, a higher QAC resistance incidence was observed among isolates from the meat processing industry than among human isolates although large plant to plant differences were indicated . No correlation between resistance and PFGE profile or resistance and persistence was observed .

Int J Food Microbiol, 2004 Oct 1, 96(1), 49 - 59
Time-temperature profiles of chilled ready-to-eat foods in school catering and probabilistic analysis of Listeria monocytogenes growth; Rosset P et al.; The purpose of this study was to evaluate the chill chain in school catering by monitoring time-temperature profiles . Chilled ready-to-eat foods have been chosen as subject of this study because of their high risk due to their production, storage and distribution steps, separated in time, followed by consumption without any further thermal treatment . In order to integrate the effects of storage duration and storage temperature, a quantitative criterion, namely "TTE" or "Time-Temperature Equivalent", was proposed . To illustrate the sanitary consequences of the recorded thermal history, Listeria monocytogenes growth was predicted based on reference growth curves in chilled ready-to-eat food products . The study of five centralised kitchens and 11 school-lunch canteens demonstrated in general a satisfactory maintenance of the chill chain . However, the coincidence of extended storage duration (due to weekends) and temperature abuse was observed and could lead to a significant microbial development .

J Immunol, 2004 Sep 15, 173(6), 4084 - 90
The development of functional CD8 T cell memory after Listeria monocytogenes infection is not dependent on CD40; Montfort MJ et al.; The immunologic requirements for generating long-lived protective CD8 T cell memory remain unclear . Memory CD8 populations generated in the absence of CD4 Th cells reportedly have functional defects, and at least a subset of CD8 T cells transiently express CD40 after activation, suggesting that direct CD4-CD8 T cell interactions through CD40 may influence the magnitude and functional quality of memory CD8 populations . To ascertain the role of CD40 in such direct T cell interactions, we investigated CD8 T cell responses in CD40-/- mice after infection with Listeria monocytogenes, an intracellular bacterium that induces APC activation and thus priming of CD8 T cells independently of CD4 Th cell help through CD40 . In this study we show that memory CD8 T cells generated in CD40-deficient mice show in vivo cytotoxicity and cytokine production equivalent to CD8 memory T cells from wild-type mice . Upon secondary Listeria infection, CD40-/- memory CD8 T cells expand to greater numbers than seen in wild-type mice . These results indicate that CD40 ligation on CD8 T cells, although reportedly a part of CD8 T cell memory development in an H-Y-directed response, is not needed for the development of functional memory CD8 T cell populations after Listeria infection .

J Immunol, 2004 Sep 15, 173(6), 3660 - 7
Fas-Fas ligand interactions are essential for the binding to and killing of activated macrophages by gamma delta T cells; Dalton JE et al.; Gammadelta T cells have a direct role in resolving the host immune response to infection by eliminating populations of activated macrophages . Macrophage reactivity resides within the Vgamma1/Vdelta6.3 subset of gammadelta T cells, which have the ability to kill activated macrophages following infection with Listeria monocytogenes (Lm) . However, it is not known how gammadelta T cell macrophage cytocidal activity is regulated, or what effector mechanisms gammadelta T cells use to kill activated macrophages . Using a macrophage-T cell coculture system in which peritoneal macrophages from naive or Lm-infected TCRdelta-/- mice were incubated with splenocytes from wild-type and Fas ligand (FasL)-deficient mice (gld), the ability of Vgamma1 T cells to bind macrophages was shown to be dependent upon Fas-FasL interactions . Combinations of anti-TCR and FasL Abs completely abolished binding to and killing of activated macrophages by Vgamma1 T cells . In addition, confocal microscopy showed that Fas and the TCR colocalized on Vgamma1 T cells at points of contact with macrophages . Collectively, these studies identify an accessory or coreceptor-like function for Fas-FasL that is essential for the interaction of Vgamma1 T cells with activated macrophages and their elimination during the resolution stage of pathogen-induced immune responses .

Annu Rev Microbiol . 2004 Jun 16; {Epub ahead of print}
Molecular Determinants of Listeria Monocytogenes Virulence; Dussurget O et al.; Listeria monocytogenes is the etiological agent of listeriosis, a severe human foodborne infection characterized by gastroenteritis, meningitis and encephalitis, abortions and perinatal infections . This gram-positive bacteria is a facultative intracellular pathogen that induces its own uptake into nonphagocytic cells and spreads from cell to cell by using an actin-based motility process . This review covers both well-established as well as recent advances in the characterization of L . monocytogenes virulence determinants and their role in the pathophysiology of listeriosis . Expected online publication date for the Annual Review of Microbiology Volume 58 is September 8, 2004 . Please see for revised estimates.

Int Immunol, 2004 Oct, 16(10), 1535 - 48 Epub 2004 Sep 06.
Modulation of T cell development and activation by novel members of the Schlafen (slfn) gene family harbouring an RNA helicase-like motif; Geserick P et al.; The regulatory networks governing development and differentiation of hematopoietic cells are incompletely understood . Members of the Schlafen (Slfn) protein family have been implicated in the regulation of cell growth and T cell development . We have identified and chromosomally mapped four new members, slfn5, slfn8, slfn9 and slfn10, which belong to a distinct subgroup within this gene family . The characteristic feature of these proteins is the presence of sequence motifs identifying them as distinct members of the superfamily I of DNA/RNA helicases . A significant role of these newly identified members in hematopoietic cell differentiation is suggested based on their differential regulation (i) in developing and activated T cells, (ii) in LPS or IFNgamma activated macrophages, (iii) upon IL6 or LIF driven terminal differentiation of myeloblastic M1 cells into macrophage-like cells, and (iv) in splenocytes of mice infected with Listeria monocytogenes . In contrast to wild-type cells, IRF-1 and IFNalpha/betaR deficient macrophages, although undergoing growth arrest, fail to upregulate slfn gene expression upon IFNgamma or LPS stimulation, respectively . Therefore, an essential participation in IFNgamma or LPS induced growth arrest appears unlikely . Likewise, ectopic expression of the newly identified slfn family members in fibroblasts did not reveal a general impact on growth control . In contrast, transgenic T-cell specific expression of a representative member of this new subfamily, slfn8, resulted in profoundly impaired T cell development and peripheral T cells showed a reduced proliferative potential . Thus, functional participation of slfn8 in the regulatory networks governing T cell development and growth appears to be cell type specific.

Microbiology, 2004 Sep, 150(Pt 9), 3025 - 33
Cell-surface alterations in class IIa bacteriocin-resistant Listeria monocytogenes strains; Vadyvaloo V et al.; Strains of the food-borne pathogen Listeria monocytogenes, showing either intermediate or high-level resistance to class IIa bacteriocins, were investigated to determine characteristics that correlated with their sensitivity levels . Two intermediate and one highly resistant spontaneous mutant of L . monocytogenes B73, a highly resistant mutant of L . monocytogenes 412, and a highly resistant, defined (mptA) mutant of L . monocytogenes EGDe were compared with their respective wild-type strains in order to investigate the contribution of different factors to resistance . Decreased mannose-specific phosphotransferase system gene expression (mptA, EIIAB(Man) component) was implicated in all levels of resistance, confirming previous studies by the authors' group . However, a clear correlation between d-alanine content in teichoic acid (TA), in particular the alanine : phosphorus ratio, and a more positive cell surface, as determined by cytochrome c binding, were found for the highly resistant strains . Furthermore, two of the three highly resistant strains showed a significant increase in sensitivity towards d-cycloserine (DCS) . However, real-time PCR of the dltA (d-alanine esterification), and dal and ddlA genes (peptidoglycan biosynthesis) showed no change in transcriptional levels . The link between DCS sensitivity and increased d-alanine esterification of TA may be that DCS competes with alanine for transport via the alanine transporter . A possible tendency towards increased lysinylation of membrane phospholipid in the highly resistant strains was also found . A previous study reported that cell membranes of all the resistant strains, including the intermediate resistant strains, contained more unsaturated phosphatidylglycerol, which is an indication of a more fluid cell membrane . The results of that study correlate with the possible lysinylation, decreased mptA expression, d-alanine esterification of TA and more positive cell surface charge found in this study for resistant strains . The authors' findings strongly indicate that all these factors could contribute to class IIa bacteriocin resistance and that the combination and contribution of each of these factors determine the level of bacteriocin resistance.

Appl Environ Microbiol, 2004 Sep, 70(9), 5672 - 8
Inhibition of Listeria monocytogenes in fish and meat systems by use of oregano and cranberry phytochemical synergies; Lin YT et al.; Optimized phenolics from oregano and cranberry extracts were evaluated for antimicrobial activity against Listeria monocytogenes in laboratory media and in beef and fish . The antimicrobial activity increased when oregano and cranberry extracts were mixed at a ratio of 75% oregano and 25% cranberry (wt/wt) with 0.1 mg of phenolic per disk or ml, and the efficacy was further enhanced by lactic acid . The inhibition by phytochemical and lactic acid synergies was most effective when beef and fish slices were stored at 4 degrees C.

Appl Environ Microbiol, 2004 Sep, 70(9), 5644 - 50
Raw cow milk bacterial population shifts attributable to refrigeration; Lafarge V et al.; We monitored the dynamic changes in the bacterial population in milk associated with refrigeration . Direct analyses of DNA by using temporal temperature gel electrophoresis (TTGE) and denaturing gradient gel electrophoresis (DGGE) allowed us to make accurate species assignments for bacteria with low-GC-content (low-GC%) (<55%) and medium- or high-GC% (>55%) genomes, respectively . We examined raw milk samples before and after 24-h conservation at 4 degrees C . Bacterial identification was facilitated by comparison with an extensive bacterial reference database ( approximately 150 species) that we established with DNA fragments of pure bacterial strains . Cloning and sequencing of fragments missing from the database were used to achieve complete species identification . Considerable evolution of bacterial populations occurred during conservation at 4 degrees C . TTGE and DGGE are shown to be a powerful tool for identifying the main bacterial species of the raw milk samples and for monitoring changes in bacterial populations during conservation at 4 degrees C . The emergence of psychrotrophic bacteria such as Listeria spp . or Aeromonas hydrophila is demonstrated.

J Bacteriol, 2004 Sep, 186(18), 6265 - 76
A novel mutation within the central Listeria monocytogenes regulator PrfA that results in constitutive expression of virulence gene products; Wong KK et al.; The PrfA protein of Listeria monocytogenes functions as a key regulatory factor for the coordinated expression of many virulence genes during bacterial infection of host cells . PrfA activity is controlled by multiple regulatory mechanisms, including an apparent requirement for either the presence of a cofactor or some form of posttranslational modification that regulates the activation of PrfA . In this study, we describe the identification and characterization of a novel PrfA mutation that results in constitutive activation of the PrfA protein . The PrfA L140F mutation was found to confer high-level expression of PrfA-regulated genes and to be functionally dominant over the wild-type allele . The presence of the PrfA L140F mutation resulted in the aggregation of L . monocytogenes in broth culture and, unlike previously described prfA mutations, appeared to be slightly toxic to the bacteria . High-level PrfA-dependent gene expression showed no additional increase in L . monocytogenes strains containing an additional copy of prfA L140F despite a >4-fold increase in PrfA protein levels . In contrast, the introduction of multiple copies of the wild-type prfA allele to L . monocytogenes resulted in a corresponding increase in PrfA-dependent gene expression, although overall expression levels remained far below those observed for PrfA L140F strains . These results suggest a hierarchy of PrfA regulation, such that the relative levels of PrfA protein present within the cell correlate with the levels of PrfA-dependent gene expression when the protein is not in its fully activated state; however, saturating levels of the protein are then quickly reached when PrfA is converted to its active form . Regulation of the PrfA activation status must be an important facet of L . monocytogenes survival, as mutations that result in constitutive PrfA activation may have deleterious consequences for bacterial physiology.

Bacteriol Virusol Parazitol Epidemiol, 2003 Apr-Sep, 48(2-3), 149 - 52
{Effect of transferrin on the expression of virulence factors in Listeria monocytogenes strains}; Mihaescu G et al.; Iron is an essential element for the great majority of microorganisms, which developed transport systems for iron acquisition, because during the colonization and invasion processes the microorganisms encounter iron-limiting conditions . The bacterial genes implicated in the iron transport and those codifying for other virulence factors are simultaneously depressed . The purpose of this study was to investigate the role of transferrin on the virulence factors expression in food isolated Listeria monocytogenes strains . Our results showed that transferrin stimulates the bacterial growth rates, as well as the adherence and invasion capacity (27 x 10(4) CFU/ml vs 13 x 10(4) CFU/ml) to cellular substrate and inert one (as shown by slime test) . Transferrin also induced the secretion of haemolysins, amylases and lecithinases, demonstrating the role of iron ions in the virulence genes derepression and the simulation of bacterial growth rate.

J Cell Biol, 2004 Aug 30, 166(5), 743 - 53
Role of lipid rafts in E-cadherin-- and HGF-R/Met--mediated entry of Listeria monocytogenes into host cells; Seveau S et al.; Listeria monocytogenes uptake by nonphagocytic cells is promoted by the bacterial invasion proteins internalin and InlB, which bind to their host receptors E-cadherin and hepatocyte growth factor receptor (HGF-R)/Met, respectively . Here, we present evidence that plasma membrane organization in lipid domains is critical for Listeria uptake . Cholesterol depletion by methyl-beta-cyclodextrin reversibly inhibited Listeria entry . Lipid raft markers, such as glycosylphosphatidylinositol-linked proteins, a myristoylated and palmitoylated peptide and the ganglioside GM1 were recruited at the bacterial entry site . We analyzed which molecular events require membrane cholesterol and found that the presence of E-cadherin in lipid domains was necessary for initial interaction with internalin to promote bacterial entry . In contrast, the initial interaction of InlB with HGF-R did not require membrane cholesterol, whereas downstream signaling leading to F-actin polymerization was cholesterol dependent . Our work, in addition to documenting for the first time the role of lipid rafts in Listeria entry, provides the first evidence that E-cadherin and HGF-R require lipid domain integrity for their full activity.

FEMS Microbiol Lett, 2004 Sep 1, 238(1), 37 - 41
Involvement of the mpo operon in resistance to class IIa bacteriocins in Listeria monocytogenes; Arous S et al.; High resistance to class IIa bacteriocins in Listeria monocytogenes has been clearly linked to lack of expression of the mptACD operon, encoding the EIIt Man mannose PTS permease . Also, intermediate resistance has been associated with membrane phospholipid modifications in the spontaneous mutants L . monocytogenes B73-V1 and B73-V2 . We constructed a new mutant of L . monocytogenes that was interrupted in mpoA, and which also exhibited an intermediate resistance phenotype . The mpoABCD operon putatively encodes a PTS permease of the mannose family similar to that encoded by the mpt operon . In silico analysis indicated that mpo transcription might be dependent on sigma54 . Our study demonstrated that the three intermediate resistant mutants have a slight decrease in mptACD expression, showing that the level of sensitivity is correlated to the level of mpt expression . We show a cross-regulation between mpo and mpt . In particular, the mpo mutant has a defect in mpt expression that possibly could explain its intermediate resistance phenotype.

FEMS Microbiol Lett, 2004 Sep 1, 238(1), 29 - 36
Modeling the synergistic effect of high pressure and heat on inactivation kinetics of Listeria innocua: a preliminary study; Buzrul S et al.; The survival curves of Listeria innocua CDW47 by high hydrostatic pressure were obtained at four pressure levels (138, 207, 276, 345 MPa) and four temperatures (25, 35, 45, 50 degrees C) in peptone solution . Tailing was observed in the survival curves . Elevated temperatures and pressures substantially promoted the inactivation of L . innocua . A linear and two non-linear (Weibull and log-logistic) models were fitted to these data and the goodness of fit of these models were compared . Regression coefficients (R2), root mean square (RMSE), accuracy factor (Af) values and residual plots suggested that linear model, although it produced good fits for some pressure-temperature combinations, was not as appropriate as non-linear models to represent the data . The residual and correlation plots strongly suggested that among the non linear models studied the log-logistic model produced better fit to the data than the Weibull model . Such pressure-temperature inactivation models form the engineering basis for design, evaluation and optimization of high hydrostatic pressure processes as a new preservation technique.

Biotechniques, 2004 Aug, 37(2), 246 - 8, 250-3
Potential influence of the first PCR cycles in real-time comparative gene quantifications; Nogva HK et al.; There is an underlying assumption in real-time PCR that the amplification efficiency is equal from the first cycles until a signal can be detected . In this study, we evaluated this assumption by analyzing genes with known gene copy number using real-time PCR comparative gene quantifications . Listeria monocytogenes has six 23S rRNA gene copies and one copy of the hlyA gene . We determined 23S rRNA gene copy numbers between 0.9 and 1.6 relative to hlyA when applying the comparative gene quantification approach . This paper focuses on the first cycles of PCR to explain the difference between known and determined gene copy numbers . Both theoretical and experimental evaluations were done . There are three different products (types 1-3) dominating in the first cycles . Type 1 is the original target, type 2 are undefined long products, while type 3 are products that accumulate during PCR . We evaluated the effects of type 1 and 2 products during the first cycles by cutting the target DNA with a restriction enzyme that cuts outside the boundaries of the PCR products . The digestion resulted in a presumed increased amplification efficiency for type 1 and 2 products . Differences in the amplification efficiencies between type 1, 2, and 3 products may explain part of the error in the gene copy number determinations using real-time PCR comparative gene quantifications . Future applications of real-time PCR quantifications should account for the effect of the first few PCR cycles on the conclusions drawn.

Arch Latinoam Nutr, 2004 Mar, 54(1), 72 - 80
{Design of a Hazard Analysis and Critical Control Points (HACCP) plan to assure the safety of a bologna product produced by a meat processing plant}; Bou Rached L et al.; The Hazard Analysis and Critical Control Point (HACCP) is a systematic integral program used to identify and estimate the hazards (microbiological, chemical and physical) and the risks generated during the primary production, processing, storage, distribution, expense and consumption of foods . To establish a program of HACCP has advantages, being some of them: to emphasize more in the prevention than in the detection, to diminish the costs, to minimize the risk of manufacturing faulty products, to allow bigger trust to the management, to strengthen the national and international competitiveness, among others . The present work is a proposal based on the design of an HACCP program to guarantee the safety of the Bologna Special Type elaborated by a meat products industry, through the determination of hazards (microbiological, chemical or physical), the identification of critical control points (CCP), the establishment of critical limits, plan corrective actions and the establishment of documentation and verification procedures . The used methodology was based in the application of the seven basic principles settled down by the Codex Alimentarius, obtaining the design of this program . In view of the fact that recently the meat products are linked with pathogens like E . coli O157:H7 and Listeria monocytogenes, these were contemplated as microbiological hazard for the establishment of the HACCP plan whose application will guarantee the obtaining of a safe product.

C R Biol, 2004 Jun, 327(6), 523 - 31
Exploitation of host cell cytoskeleton and signalling during Listeria monocytogenes entry into mammalian cells; Pizarro-Cerda J et al.; Deciphering how Listeria monocytogenes exploits the host cell machinery to invade mammalian cells during infection isa key issue for the understanding how this food-borne pathogen causes a pleiotropic disease ranging from gastro-enteritis to meningitis and abortions . Using multidisciplinary approaches, essentially combining bacterial genetics and cell biology, we have identified two bacterial proteins critical for entry into target cells, InlA and InlB . Their cellular ligands have been also identified: InlA interacts with the adhesion molecule E-cadherin, while InlB interacts with the receptor for the globular head of the complement factor Clq (gClq-R), with the hepatocyte growth factor receptor (c-Met) and with glycosaminoglycans(including heparan sulphate) . The dynamic interaction between these cellular receptors and the actin cytoskeleton is currently under investigation . Several intracellular molecules have been recognized as key effectors for Listeria entry into target cells,including catenins (implicated in the connection of E-cadherin to actin) and the actin depolymerising factor/cofilin (involved in the rearrangement of the cytoskeleton in the InlB-dependent internalisation pathway) . At the organism level, species specificity has been discovered concerning the interaction between InlA and E-cadherin, leading to the generation of transgenic mice expressing the human E-cadherin, in which the critical role of InlA in the crossing of the intestinal barrier has been clearly determined . Listeria appears as an instrumental model for addressing critical questions concerning both the complex process of bacterial pathogenesis and also fundamental molecular processes, such as phagocytosis.

J Vet Med B Infect Dis Vet Public Health, 2004 Jun, 51(5), 209 - 15
Neuropathological findings in brains of Bavarian cattle clinically suspected of bovine spongiform encephalopathy; Miyashita M et al.; The brains of 26 Bavarian bovines clinically suspected of bovine spongiform encephalopathy (BSE) were the subject of a neuropathological evaluation containing histopathology and immunohistochemistry . Six animals tested positive for BSE . In these six brains severe histological lesions that correlated with previous reports from the United Kingdom were observed . Immunohistochemistry with prion protein (PrP(Sc)), glial fibrillary acidic protein (GFAP) and synaptophysin were conducted on the mid-brain containing the red nucleus . All BSE-positive brains stained positively for PrP(Sc), and no plaques were observed . The BSE-affected brains showed a substantially more intense staining pattern for GFAP in comparison with the control groups, some of which were diagnosed with severe neuropathological disorders . Synaptophysin staining on BSE-positive brains was substantially reduced in the neuropil of the mid-brain, especially in the red nucleus . Twenty animals tested negative for BSE . The most common diagnoses were listeriosis, viral infections of unknown aetiology, brain oedema and hypomagnesaemia . These disorders may represent the most important clinical differential diagnoses for BSE in Bavaria.

J Food Prot, 2004 Aug, 67(8), 1682 - 6
Optimizing concentration and timing of a phage spray application to reduce Listeria monocytogenes on honeydew melon tissue; Leverentz B et al.; A phage cocktail was applied to honeydew melon pieces 1, 0.5, and 0 h before contamination with Listeria monocytogenes strain LCDC 81-861 and 0.5, 1, 2, and 4 h after contamination . The phage application was most effective when applied 1, 0.5, or 0 h before contamination with L . monocytogenes, reducing pathogen populations by up to 6.8 log units after 7 days of storage . This indicates that under commercial conditions, if contamination occurs at the time of cutting, phage would have to be applied as soon as possible after cutting the produce . However, all phage applications from 1 h before to 4 h after contamination and all phage concentrations ranging from 10(4) to 10(8) PFU/ml reduced bacterial populations on honeydew melon pieces . Higher phage concentrations were more effective in reducing pathogen populations . A phage concentration of approximately 10(8) PFU/ml was necessary to reduce the pathogen populations to nondetectable levels immediately after treatment, and pathogen growth was suppressed by phage concentrations of 10(6) through 10(8) throughout the storage period of 7 days at 10 degrees C . In an attempt to enhance the effectiveness of the phage cocktail on low pH fruit, such as apples, the phage was applied in combination with MnCl2 . This combination, however, did not enhance the effectiveness of the phage on apple tissue . The results from this study indicate that the effectiveness of the phage application on honeydew melon pieces can be optimized by using a phage concentration of at least 10(8) PFU/ml applied up to 1 h after processing of the honeydew melons.

J Food Prot, 2004 Aug, 67(8), 1676 - 81
Fate of Listeria monocytogenes in bovine manure-amended soil; Jiang X et al.; The survival and growth of Listeria monocytogenes in soil amended with bovine manure was studied under different environmental conditions of temperature, nutrients, and soil microflora . Autoclaved soil was compared with unautoclaved soil for assessing the influence of competitive soil microflora on the survival of L . monocytogenes . Initial L . monocytogenes cell numbers of 5 to 6 log CFU/g survived for up to 43, 43, and 14 days in manure-amended autoclaved soil at 5, 15, and 21 degrees C, respectively . In manure-amended unautoclaved soil, the pathogen was detectable for up to 43, 21, and 21 days at 5, 15, and 21 degrees C, respectively . L . monocytogenes was inactivated more rapidly in autoclaved soil amended with manure at a manure/soil ratio of 1:10 than in the more dilute (1:100) manure in soil samples at both 15 and 21 degrees C . However, in manure-amended unautoclaved soil, L . monocytogenes survived longer in samples with ratios of 1:10 than in the more dilute (1:100) manure-amended soil . The persistence of L . monocytogenes for several weeks in manure-amended soil suggests listeriae could be transmitted through soil to fresh produce or to shoes, clothing, and hands of field workers, especially during the cold months.

J Food Prot, 2004 Aug, 67(8), 1671 - 5
High-pressure processing of Gorgonzola cheese: influence on Listeria monocytogenes inactivation and on sensory characteristics; Carminati D et al.; The presence of Listeria monocytogenes on the rind of Gorgonzola cheese is difficult to avoid . This contamination can easily occur as a consequence of handling during ripening . The aims of this study were to determine the efficiency of high-pressure processing (HPP) for inactivation of L . monocytogenes on cheese rind and to evaluate the influence of HPP treatments on sensory characteristics . Gorgonzola cheese rinds, after removal, were inoculated (about 7.0 log CFU/g) with L . monocytogenes strains previously isolated from other Gorgonzola cheeses . The inoculated cheese rinds were processed with an HPP apparatus under conditions of pressure and time ranging from 400 to 700 MPa for 1 to 15 min . Pressures higher than 600 MPa for 10 min or 700 MPa for 5 min reduced L . monocytogenes more than 99% . A reduction higher than 99.999% was achieved pressurizing cheese rinds at 700 MPa for 15 min . Lower pressure or time treatments were less effective and varied in effectiveness with the cheese sample . Changes in sensory properties possibly induced by the HPP were evaluated on four different Gorgonzola cheeses . A panel of 18 members judged the treated and untreated cheeses in a triangle test . Only one of the four pressurized cheeses was evaluated as different from the untreated sample . HPP was effective in the reduction of L . monocytogenes on Gorgonzola cheese rinds without significantly changing its sensory properties . High-pressure technology is a useful tool to improve the safety of this type of cheese.

J Food Prot, 2004 Aug, 67(8), 1656 - 65
High-resolution genotyping of Listeria monocytogenes by fluorescent amplified fragment length polymorphism analysis compared to pulsed-field gel electrophoresis, random amplified polymorphic DNA analysis, ribotyping, and PCR-restriction fragment length polymorphism analysis; Fonnesbech Vogel B et al.; The purpose of this study was to evaluate fluorescent amplified fragment length polymorphism (AFLP) analysis for the inter- and intraspecies differentiation of a collection of 96 strains of Listeria monocytogenes and 10 non-L . monocytogenes strains representing six other Listeria species of different origin . The AFLP technique was compared with three other molecular typing methods--ribotyping, random amplified polymorphic DNA analysis (RAPD), and pulsed-field gel electrophoresis (PFGE)--in terms of discriminatory ability . PCR-restriction fragment length polymorphism was included for virulence gene allele characterization . The 96 L . monocytogenes strains were divided into two major clusters by AFLP fingerprinting at a similarity level of 82% in concordance with the results of PFGE, RAPD, and ribotyping . One main cluster consisted of all of the 24 L . monocytogenes hly allele 1 strains, while another main cluster consisted of all of the 72 L . monocytogenes hly allele 2 strains . This indicates the existence of two distinct phylogenetic divisions . Isolates of the remaining Listeria species were not included in the clusters . AFLP, PFGE, and RAPD typing were highly discriminatory methods, with discrimination (D) indices of 0.974, 0.969, and 0.954, respectively, whereas ribotyping had a lower D index of 0.874 . AFLP, PFGE, and RAPD typing showed some level of agreement in terms of strain grouping and differentiation . However, all three methods subdivided types of strains grouped by the other methods . Isolates with identical DNA profiles were distributed across the spectrum of origin . It was not possible to associate certain types with specific food sectors or clinical cases, which is indicative of the spread of L . monocytogenes clones across species . Overall, AFLP fingerprinting was suitable for the high-resolution genotyping of L . monocytogenes and had an equally high or higher differentiation power compared to PFGE or RAPD typing.

J Food Prot, 2004 Aug, 67(8), 1646 - 55
A validated PCR-based method to detect Listeria monocytogenes using raw milk as a food model--towards an international standard; D'Agostino M et al.; A PCR assay with an internal amplification control was developed for Listeria monocytogenes . The assay has a 99% detection probability of seven cells per reaction . When tested against 38 L . monocytogenes strains and 52 nontarget strains, the PCR assay was 100% inclusive (positive signal from target) and 100% exclusive (no positive signal from nontarget) . The assay was then evaluated in a collaborative trial involving 12 European laboratories, where it was tested against an additional 14 target and 14 nontarget strains . In that trial, the inclusivity was 100% and the exclusivity was 99.4%, and both the accordance (repeatability) and the concordance (reproducibility) were 99.4% . The assay was incorporated within a method for the detection of L . monocytogenes in raw milk, which involves 24 h of enrichment in half-Fraser broth followed by 16 h of enrichment in a medium that can be added directly into the PCR . The performance characteristics of the PCR-based method were evaluated in a collaborative trial involving 13 European laboratories . In that trial, a specificity value (percentage of correct identification of blank samples) of 81.8% was obtained; the accordance was 87.9%, and the concordance was 68.1% . The sensitivity (correct identification of milk samples inoculated with 20 to 200 L . monocytogenes cells per 25 ml) was 89.4%, the accordance was 81.2%, and the concordance was 80.7% . This method provides a basis for the application of routine PCR-based analysis to dairy products and other foodstuffs and should be appropriate for international standardization.

J Food Prot, 2004 Aug, 67(8), 1641 - 5
Survival and recovery of viable but nonculturable Listeria monocytogenes cells in a nutritionally depleted medium; Foong SC et al.; Survival of a desiccated five-strain Listeria monocytogenes mixture during storage in sand at 4 degrees C for 2 months was determined using the acridine orange direct count method with novobiocin and plate counts . Samples of inoculated sand were taken every 2 weeks, incubated at 37 degrees C for 6 h, stained with acridine orange, and then examined with a fluorescence microscope . Elongated viable but nonculturable cells were most frequently observed during weeks 2 and 4 . At weeks 6 and 8, most of the cells either remained viable or were dead . In each microscopic field, only one or two viable but nonculturable cells were observed among hundreds of other viable culturable cells, indicating that L . monocytogenes does not generally become viable but nonculturable . Therefore, viable but nonculturable cells are not a concern when plating environmental samples or desiccated L . monocytogenes cells on nonselective media . Tryptic soy agar with 0.6% (wt/vol) yeast extract (TSAYE) and Columbia agar were used as nonselective plate count media . Modified Oxford agar and TSAYE + 5% (wt/vol) sodium chloride were used as the selective plate count media . The effects of aerobic or anaerobic incubation and media supplementation with 0.1% or 1% (wt/vol) sodium pyruvate were tested to optimize recovery of desiccated cells . Nonselective media showed better recovery when TSAYE and Columbia agar contained 0.1% (wt/vol) pyruvate and were incubated aerobically . These two culture methods were equally effective (P > 0.05) for recovering desiccated L . monocytogenes cells.

J Food Prot, 2004 Aug, 67(8), 1634 - 40
Optimization of rapid detection of Escherichia coli O157:H7 and Listeria monocytogenes by PCR and application to field test; Moon GS et al.; For rapid detection of Escherichia coli O157:H7 and Listeria monocytogenes, simple methods for sample preparation and PCR were established and applied to a field test . To improve specificity, primer sets LP43-LP44 and C(+)-D(-) were selected for E . coli O157:H7 and L . monocytogenes, respectively . Through centrifugation and partial heat treatment after enrichment, E . coli O157:H7 and L . monocytogenes were detected at 1 initial CFU without genomic DNA extraction in the culture and with artificially inoculated food samples including milk, chicken, ham, and pork . Based on the optimized PCR method, a feasibility test was carried out using randomly collected field samples . To remove false positives and false negatives, a PCR method using several primer sets, including the optimized primer set, and a standard culture method were used . With the PCR detection and standard culture methods, two pork samples were positive for L . monocytogenes after enrichment, indications that the PCR assay could be effectively used for rapid, sensitive, and species-specific detection of foodborne pathogens.

Med Sci Monit, 2004 Sep, 10(9), BR317 - 24 Epub 2004 Aug 20.
Immunosuppressive treatment inhibits the development of amebic liver abscesses in hamsters; Quintanar-Quintanar ME et al.; BACKGROUND: The aim of the present study was to determine if the inflammation and/or immunosuppression induced by Entamoeba histolytica may contribute to amebic invasion . MATERIAL/METHODS: Dexamethasone was administered three days before and three days after inoculation of hamsters with E . histolytica . Seven days alter inoculation the animals were sacrificed and the sizes of their amebic liver abscesses were determined . The number of neutrophils, macrophages, T and B cells in the peritoneum as well as the production of nitric oxide and the susceptibility to Listeria monocytogenes infection was also determined . RESULTS: Dexamethasone treatment significantly reduced the number of T lymphocytes in thymus and spleen . The number of neutrophils, macrophages and T lymphocytes in the peritoneal exudate was also reduced as well as the production of nitric oxide and the microbicidal activity against Listeria monocytogenes . However, in the animals treated with a high dose of dexamethasone the size of the liver abscesses was significantly smaller than in the untreated animals . CONCLUSIONS: The results suggest that macrophage and T cell-mediated immunity is not relevant as a protective mechanism because tissue invasion by E . histolytica was reduced in immunosuppresed animals . On the contrary, the inflammatory process may contribute to the invasion and liver damage.

Ginecol Obstet Mex, 2001 May, 69, 206 - 8
{Hematometra & Listeria monocytogenes}; Gomez Arzapalo E et al.; The hematometra is a nosological entity that may not always be attributed to an embryonic defect of the paramesonefros; cervical-vaginal infections such as etiological possibilities due to Listeria monocytogenes (Lm), cervix malignant neoplasias, iatrogenias due to endometrial ablation with Lasser, traumatic bloody uterine curetage and because of cervical cryocoagulation or electrocoagulation are also mentioned . The case to be reported is from a woman in reproductive stage, who is 32 years old, and had menarca at the age of 13, starting her sexual life at 31, not using any method to control her fertility . When having an eight-week amenorrhea after 8 months of marriage, she visited the doctor for assumed pregnancy, within the prenatal analysis a pelvic echographic study was requested, finding out images that we concluded as hematometra, having been drained and demonstrated the presence of LM by anti-Lm antibodies, being administered Azitromicina and Espiramicina.

Indian J Exp Biol, 2003 Dec, 41(12), 1466 - 8
Prevalence of Listeria in soil; Moshtaghi H et al.; One hundred thirty soil samples from agricultural fields and animal-inhabited areas were examined for the presence of Listeria . The microorganism was identified in 23 (17.7%) samples . L . monocytogenes was detected in 7 samples (5.4%), L . ivanovii in 2 (1.5%), L . innocua in 10 (7.7%) and L . welshimeri in 4 samples (3.1%) . Prevalence of Listeria in soil from agricultural fields (17.5%) did not differ considerably from that in the soil and animal-inhabited area (18.0%), but L . ivanovii was isolated only from the latter source . Frequency of occurrence of different species of Listeria differed from place to place.

Toxicol Sci, 2004 Nov, 82(1), 143 - 53 Epub 2004 Aug 19.
Roles of reactive oxygen species and heme oxygenase-1 in modulation of alveolar macrophage-mediated pulmonary immune responses to Listeria monocytogenes by diesel exhaust particles; Yin XJ et al.; Diesel exhaust particles (DEP) have been shown to suppress alveolar macrophage (AM)-mediated pulmonary immune responses to Listeria monocytogenes in vivo . In this study, effects of DEP-derived reactive oxygen species (ROS) and heme oxygenase (HO)-1 on AM-mediated immune responses to L . monocytogenes were investigated . Brown Norway rats were intratracheally inoculated with 100,000 L . monocytogenes, and AM were isolated at 7 days post-infection . Exposure to DEP or their organic extract (eDEP), but not the washed DEP (wDEP) or carbon black, increased intracellular ROS and HO-1 expression in AM . Induction of ROS and HO-1 by eDEP was partially reversed by alpha-naphthoflavone, a cytochrome P450 1A1 inhibitor, and totally blocked by N-acetylcysteine . In addition, exposure to eDEP, but not wDEP, inhibited lipopolysacchride-stimulated secretion of tumor necrosis factor-alpha (TNF-alpha) and interleukin-12 (IL-12), but augmented production of IL-10 by AM . Kinetic studies showed that modulation of cytokines by eDEP was preceded by ROS and HO-1 induction . Furthermore, pretreatment of AM with superoxide dismutase (SOD) or zinc protoporphrin IX (Znpp), which attenuated eDEP-induced HO-1 expression/activity, substantially inhibited eDEP effect on IL-10 . Finally, direct stimulation with pyrogallol (PYR), a superoxide donor, upregulated HO-1 and IL-10 but decreased secretion of IL-12 in L . monocytogenes-infected AM . These results show that DEP, through eDEP-mediated ROS, induce HO-1 expression and IL-10 production and at the same time inhibit AM production of TNF-alpha and IL-12 to dampen the host immune responses . The results also suggest that HO-1 may play an important role in regulating production of IL-10 by DEP-exposed and L . monocytogenes-infected AM.

Neuroimmunomodulation, 2004, 11(5), 323 - 31
Splenic norepinephrine and serum corticosterone level fluctuations associated with bacteria-induced stress; Kim D et al.; Corticosterone (CORT) and norepinephrine (NE), two effector molecules of the hypothalamic-pituitary-adrenal (HPA) and the sympathetic-lymphoid (SL) axes, respectively, differentially influence murine host resistance to Listeria monocytogenes (LM) . Serum CORT and splenic NE levels early (< or =24 h) after infection correlated positively with host resistance, as long as the LM burden did not exceed approximately 10(6) cfu LM per spleen . As previously reported, mice with right-circling preference (R-mice) have significantly greater host resistance to LM than those with left-circling preference (L-mice) and early after infection, R-mice had significantly higher serum CORT levels than L-mice . However, rapid pathogenesis with a high bacterial burden induced high activation of the HPA and SL axes, which prevented observable differences in the defense against LM, especially later in infection . With the high bacterial inoculum (10(5) LM), the splenic NE levels significantly increased, but no differences among R- and L-mice were discernible . We suggest that endogenous asymmetry of neuroimmune circuits contributes to differential host resistance, but the level of stress (bacterial inoculum) is critical . With regard to the neuroendocrine factors assessed, CORT, but not NE, levels significantly correlated with the enhanced defenses of R-mice in comparison to L-mice . The differential host resistance based on brain laterality seems to be more a function of the HPA axis and possibly other CNS effects on peripheral immunity than neurotransmitter release by the sympathetic innervation of the spleen.

J Immunother, 2004 Sep-Oct, 27(5), 339 - 46
CD4+CD25+ regulatory T cells that secrete TGFbeta and IL-10 are preferentially induced by a vaccine vector; Hussain SF et al.; CD4CD25 T cells generated in a vaccine scenario can play a critical role in limiting antitumor therapy, thus having widespread implications for the immunotherapy-based treatment of cancer . The authors previously used Listeria monocytogenes to develop two vaccine constructs that express HPV-16 E7 protein and induce strong cellular immunity to HPV-E7-expressing tumors . Immunization of mice bearing established E7-expressing tumors with Lm-LLO-E7 induced regression of the tumors, whereas Lm-E7 showed little or no tumor regression . To investigate the possibility that regulatory CD4CD25 T-cell populations may be responsible for the differences in tumor regression, the authors characterized the role of these cells generated by the two vaccine systems . The authors compared the prevalence of CD4CD25 T cells in tumor-bearing vaccinated mice and demonstrate that Lm-E7-vaccinated mice have significantly increased numbers of CD4CD25 T cells in both the spleen and tumor-infiltrating lymphocytes compared with Lm-LLO-E7-vaccinated mice . The authors confirm that these increased numbers of CD4CD25 T cells are indeed suppressor in function by in vitro suppression assays and that the mechanism of action of the tumor-infiltrating cells involves the production of suppressor cytokines interleukin-10 and transforming growth factor beta . These results show that it is possible for a tumor vaccine system to generate tumor-infiltrating CD4CD25 regulatory T cells that critically affect tumor regression and the overall success of vaccine therapy .

Microb Pathog, 2004 Aug, 37(2), 87 - 94
Characteristics of the gastritis induced by Listeria monocytogenes in mice: microbiology, histopathology, and mRNA expression of inflammatory mediators with time course of infection; Park JH et al.; Listeria monocytogenes induces the suppurative gastritis in some mice strains . In this study, characteristics of the gastritis caused by L . monocytogenes infection in mice were examined with time course of infection . Mice were administered intragastrically with 1.8 x 10(8) CFU of L . monocytogenes . Each three mice were sacrificed by cervical dislocation at 1, 3, 5, 7, 10, 14, 17, 21, and 28 days postinoculation (pi), respectively . Bacterial colonization in the stomachs reached the peak at 3 days pi, maintained over 4.3 log10 CFU/g tissue until 14 days pi, and was cleared by 28 days pi . However, in the spleens and livers, the bacteria could not be detected after 7 days pi . The gastric lesions were the most prominent at between 3 and 7 days pi . The lesions consisted of marked neutrophilic infiltration, edema, vacuolar degeneration and necrosis of muscle cells and were more severe in the nonglandular region and fundus than in the pylorus, and were in submucosa, lamina muscularis, and serosa than in mucosa . mRNA expression of several cytokines (INF-gamma, IL-1beta, IL-5, IL-6, IL-12, and TNF-alpha) and chemokines (KC, MCP-1) increased in the gastric tissue of infected mice at 1-7 days pi and slightly decreased at 14 days pi . These findings would be useful for studying the pathological mechanism of human febrile gastroenteritis due to L . monocytogenes infection .

Epidemiol Infect, 2004 Aug, 132(4), 769 - 72
Nationwide survey of human Listeria monocytogenes infection in Japan; Okutani A et al.; Listeriosis, caused by Listeria monocytogenes, is a significant public-health concern as a result of its clinical severity and high mortality . Large foodborne outbreaks of listeriosis have occurred during the last two decades in Europe and the United States, but to date there have been no food-mediated epidemics of the disease and very little information is available on the number of cases of listeriosis in Japan . We performed a nationwide surveillance study of listeriosis . The data were collected between 1980 and 2002, and 95 case reports were identified from 1996 to 2002 . We divided 13.6 (cases per year between 1996 and 2002) by the ratio of the number of beds in hospitals that replied to the questionnaire, to that of all the hospitals in Japan and estimated that there is an average of 83 cases of listeriosis per year and an incidence of 0.65 cases per million of the population in Japan.

Eur J Immunol, 2004 Sep, 34(9), 2589 - 98
Antibody-mediated activation of the classical pathway of complement may compensate for mannose-binding lectin deficiency; Roos A et al.; Deficiency of mannose-binding lectin (MBL), a recognition molecule of the lectin pathway of complement, is associated with increased susceptibility to infections . The high frequency of MBL deficiency suggests that defective MBL-mediated innate immunity can be compensated by alternative defense strategies . To examine this hypothesis, complement activation by MBL-binding ligands was studied . The results show that the prototypic MBL ligand mannan can induce complement activation via both the lectin pathway and the classical pathway . Furthermore, antibody binding to mannan restored complement activation in MBL-deficient serum in a C1q-dependent manner . Cooperation between the classical pathway and the lectin pathway was also observed for complement activation by protein 60 from Listeria monocytogenes . MBL pathway analysis at the levels of C4 and C5b-9 in the presence of classical pathway inhibition revealed a large variation of MBL pathway activity, depending on mbl2 gene polymorphisms . MBL pathway dysfunction in variant allele carriers is associated with reduced MBL ligand binding and a relative increase of low-molecular-mass MBL . These findings indicate that antibody-mediated classical pathway activation can compensate for impaired target opsonization via the MBL pathway in MBL-deficient individuals, and imply that MBL deficiency may become clinically relevant in absence of a concomitant adaptive immune response .

J Vet Diagn Invest, 2004 Jul, 16(4), 305 - 12
Investigations into shaking mink syndrome: an encephalomyelitis of unknown cause in farmed mink (Mustela vison) kits in Scandinavia; Gavier-Widen D et al.; An apparently novel neurological disease clinically characterized by shaking, tremors, seizures, staggering gait, and ataxia was first observed in farmed mink kits in Denmark in 2000 and subsequently in Sweden, Denmark, and Finland in 2001, and again in Denmark in 2002 . Lymphoplasmacytic encephalomyelitis was found in the affected kits . The lesions were most severe in the brainstem and cerebellum and consisted of neuronal degeneration and necrosis, neuronophagia, focal and diffuse gliosis, perivascular cuffs formed by lymphocytes, plasma cells and macrophages, and segmental loss of Purkinje cells . Testing was conducted to determine the cause of the disease, including general virological investigations (virus culture, negative-staining electron microscopy, immunoelectron microscopy, polymerase chain reaction for herpesviruses, adenoviruses, pestiviruses, and coronaviruses), tests for specific viral diseases (canine distemper, Borna disease, Louping ill, West Nile virus infection, tick-borne encephalitis, Aleutian disease), tests for protozoa (Toxoplasma gondii, Neospora caninum, Encephalitozoon cuniculi), bacteria (general culture, listeria, Clamydophila psittaci), and intracerebral inoculation of neonatal mice . The results of all these investigations were negative . One group of 3 mink kits inoculated intracerebrally with brain homogenate of affected mink developed clinical signs and histological lesions similar to those observed in naturally infected mink . Based on the histopathological features, it is postulated that the disease is caused by a yet unidentified virus.

Proc Natl Acad Sci U S A, 2004 Aug 17, 101(33), 12318 - 23 Epub 2004 Aug 09.
Listeria monocytogenes regulates flagellar motility gene expression through MogR, a transcriptional repressor required for virulence; Grundling A et al.; Previous studies have shown that Listeria monocytogenes flagellar motility genes, including flaA, encoding flagellin, are transcriptionally down-regulated at 37 degrees C . For some L . monocytogenes strains, temperature-dependent motility gene expression is less stringent . By using flaA-lacZ transcriptional fusions, we identified regions upstream of the -35/-10 promoter elements that are necessary for temperature-dependent expression of flaA in L . monocytogenes strain EGDe . Whereas the sequence of the flaA promoter region was identical in L . monocytogenes strain 10403S, transcriptional activity was only partially down-regulated at 37 degrees C in 10403S . This finding suggested that a transacting regulatory protein with differential expression or activity in EGDe might be involved in temperature-dependent transcription of flaA . Indeed, a protein factor capable of specifically binding to the flaA promoter region was identified in cytoplasmic extracts of EGDe by using affinity purification and MS . Deletion of the factor-encoding gene (lmo0674) resulted in loss of temperature-dependent flaA expression and an increase in flaA promoter activity . Expression of other motility genes was also deregulated in the lmo0674 deletion . We have designated lmo0674 as mogR, indicating its role as a motility gene repressor . In tissue culture models, MogR repression of flaA during intracellular infection was independent of temperature and a deletion of mogR reduced the capacity for cell-to-cell spread . During in vivo infection, a deletion of mogR resulted in a 250-fold decrease in virulence . These studies indicate that regulation of flagellar motility gene expression and/or other genes controlled by MogR is required for full virulence of L . monocytogenes.

J Exp Med, 2004 Aug 16, 200(4), 437 - 45 Epub 2004 Aug 09.
Type I interferon production enhances susceptibility to Listeria monocytogenes infection; O'Connell RM et al.; Numerous bacterial products such as lipopolysaccharide potently induce type I interferons (IFNs); however, the contribution of this innate response to host defense against bacterial infection remains unclear . Although mice deficient in either IFN regulatory factor (IRF)3 or the type I IFN receptor (IFNAR)1 are highly susceptible to viral infection, we show that these mice exhibit a profound resistance to infection caused by the Gram-positive intracellular bacterium Listeria monocytogenes compared with wild-type controls . Furthermore, this enhanced bacterial clearance is accompanied by a block in L . monocytogenes-induced splenic apoptosis in IRF3- and IFNAR1-deficient mice . Thus, our results highlight the disparate roles of type I IFNs during bacterial versus viral infections and stress the importance of proper IFN modulation in host defense.

J Exp Med, 2004 Aug 16, 200(4), 535 - 40 Epub 2004 Aug 09.
Type I interferon sensitizes lymphocytes to apoptosis and reduces resistance to Listeria infection; Carrero JA et al.; Infection with Listeria monocytogenes causes lymphocyte apoptosis that is mediated by the actions of the pore-forming virulence factor listeriolysin O (LLO) . Previous work showed that activated lymphocytes were highly sensitive to LLO-induced apoptosis, whereas resting lymphocytes were less susceptible . We now show that mice deficient in the type I interferon (IFN) receptor were more resistant to Listeria infection and had less apoptotic lesions than wild-type counterparts . Furthermore, treatment of resting splenic lymphocytes with recombinant IFN-alphaA enhanced their susceptibility to LLO-induced apoptosis . Together, these data suggest that type I IFN signaling is detrimental to handling of a bacterial pathogen and may enhance the susceptibility of lymphocytes undergoing apoptosis in response to bacterial pore-forming toxins.

J Exp Med, 2004 Aug 16, 200(4), 527 - 33 Epub 2004 Aug 09.
Mice lacking the type I interferon receptor are resistant to Listeria monocytogenes; Auerbuch V et al.; Listeria monocytogenes is a facultative intracellular pathogen that induces a cytosolic signaling cascade resulting in expression of interferon (IFN)-beta . Although type I IFNs are critical in viral defense, their role in immunity to bacterial pathogens is much less clear . In this study, we addressed the role of type I IFNs by examining the infection of L . monocytogenes in BALB/c mice lacking the type I IFN receptor (IFN-alpha/betaR-/-) . During the first 24 h of infection in vivo, IFN-alpha/betaR-/- and wild-type mice were similar in terms of L . monocytogenes survival . In addition, the intracellular fate of L . monocytogenes in macrophages cultured from IFN-alpha/betaR-/- and wild-type mice was indistinguishable . However, by 72 h after inoculation in vivo, IFN-alpha/betaR-/- mice were approximately 1,000-fold more resistant to a high dose L . monocytogenes infection . Resistance was correlated with elevated levels of interleukin 12p70 in the blood and increased numbers of CD11b+ macrophages producing tumor necrosis factor alpha in the spleen of IFN-alpha/betaR-/- mice . The results of this study suggest that L . monocytogenes might be exploiting an innate antiviral response to promote its pathogenesis.

Curr Microbiol, 2004 Aug, 49(2), 95 - 8
Catabolite repression and virulence gene expression in Listeria monocytogenes; Gilbreth SE et al.; Previous studies have suggested that carbohydrates may affect expression of virulence genes in Listeria monocytogenes . Which carbohydrates influence virulence gene expression and how carbohydrates mediate expression, however, is not clear . The goal of this work was to examine how carbohydrates affect virulence gene expression in L . monocytogenes 10403S . Growth studies were conducted in medium containing glucose and various sugars . Metabolism of arbutin, arabitol, cellobiose, mannose, maltose, trehalose, and salicin were repressed in the presence of glucose . Only when glucose was consumed were these sugars fermented, indicating that catabolite repression by glucose had occurred . To determine whether virulence gene expression was also influenced by catabolite repression, we performed primer extension experiments, using primers for hly and prfA, which encode for a hemolysin and the regulator protein PrfA, respectively . In the presence of cellobiose and arbutin, transcription of hemolysin was reduced . However, none of the sugars affected transcription of prfA . The results demonstrate that catabolite repression occurs in L . monocytogenes and suggests that, at least in strain 10403S, cellobiose and arbutin repress expression of hemolysin.

J Clin Microbiol, 2004 Aug, 42(8), 3819 - 22
Differentiation of the major Listeria monocytogenes serovars by multiplex PCR; Doumith M et al.; A new multiplex PCR assay was developed to separate the four major Listeria monocytogenes serovars isolated from food and patients (1/2a, 1/2b, 1/2c, and 4b) into distinct groups . The PCR test, which constitutes a rapid and practical alternative to laborious classical serotyping, was successfully evaluated with 222 Listeria strains.

J Immunol, 2004 Aug 15, 173(4), 2641 - 51
Cytoplasmic entry of Listeria monocytogenes enhances dendritic cell maturation and T cell differentiation and function; Brzoza KL et al.; Protective immunity to the intracellular bacterial pathogen, Listeria monocytogenes, is mediated by a vigorous T cell response . In particular, CD8(+) cytolytic T cells provide essential effector function in the clearance of bacterial infection . The cytoplasmic entry of Listeria facilitated by listeriolysin O is an essential feature not only of the bacteria's virulence, but of the ability of the bacteria to elicit protective immunity in the host . To determine how cytoplasmic entry of Listeria regulates the development of protective immunity, we examined the effects of this process on the maturation of murine dendritic cells (DC) and on their ability to prime naive CD8(+) T cell responses . Costimulatory molecules (CD40, CD80, and CD86) were induced by listerial infection only when the bacteria invaded the cytoplasm . In addition, the production of IL-12, IL-10, IL-6, and TNF-alpha was most efficiently triggered by cytosolic Listeria . Naive T cells primed by peptide-loaded DC infected with either wild-type or nonhemolytic mutant Listeria proliferated equivalently, but a much larger proportion of those primed by wild-type Listeria monocytogenes produced IFN-gamma . Costimulatory molecules induced by cytosolic entry regulated T cell proliferation and, as a result, the number of functional T cells generated . DC-produced cytokines (specifically IL-12 and IL-10) were the major factors determining the proportion of T cells producing IFN-gamma . These data highlight the requirement for listerial cytoplasmic invasion for the optimal priming of T cell cytokine production and attest to the importance of this event to the development of protective CTL responses to this pathogen.

J Immunol, 2004 Aug 15, 173(4), 2594 - 606
Mechanisms regulating the positioning of mouse p47 resistance GTPases LRG-47 and IIGP1 on cellular membranes: retargeting to plasma membrane induced by phagocytosis; Martens S et al.; The recently identified p47 GTPases are one of the most effective cell-autonomous resistance systems known against intracellular pathogens in the mouse . One member of the family, LRG-47, has been shown to be essential for immune control in vivo of Listeria monocytogenes, Toxoplasma gondii, Mycobacterium tuberculosis, and Mycobacterium avium, possibly by promoting acidification of the phagosome . However, the intracellular localization of LRG-47, and the nature of its association with the phagosomal or any other membrane system is unknown . In this study, we show that LRG-47 is a Golgi-associated protein in the IFN-stimulated cell, which is rapidly recruited to active plasma membrane upon phagocytosis and remains associated with phagosomes as they mature . We show that the Golgi localization of LRG-47 is dependent on the integrity of an amphipathic helix near the C terminus, whereas the plasma membrane localization depends on an unidentified signal associated with the G domain . Unlike LRG-47, but like the published p47 resistance GTPase, IGTP, a further p47 GTPase, IIGP1, is associated with the endoplasmic reticulum . However, unlike IGTP, IIGP1 is associated with the endoplasmic reticulum by an N-terminal myristoylation modification . Thus, the p47 GTPases are a diverse battery of intracellular defense factors dynamically associated with different membrane systems.

Microbiol Res, 2004, 159(2), 167 - 71
Hypochlorite inactivation kinetics of Listeria monocytogenes in phosphate buffer; Erkmen O; The inactivation kinetics of Listeria monocytogenes in a phosphate buffer (PB) was determined at different hypochlorite concentrations, pH values and temperatures . D-values, using a linear regression, of L . monocytogenes in PB (pH 6.5) were 23.54, 17.40, 14.24 and 12.00s at 5, 10, 50 and 100 mg l(-1) hypochlorite, respectively, at 30 degrees C . The k-values ranged from 0.098 to 0.192s(-1) and 0.007 to 0.018s(-1) for hypochlorite concentrations (from 5 to 100 mg l(-1)) in PB (pH 6.5) and PB containing 0.1% peptone (pH 6.5), respectively, at 30 degrees C . D-values of L . monocytogenes exposed to hypochlorite were decreased with decreasing pH of PB (pH from 8.5 to 4.5) . Hypochlorite showed higher antimicrobial activity at higher temperature . Not only the effect of hypochlorite concentration on the inactivation of L . monocytogenes but also other parameters like temperature, pH and suspending solutions effect the inactivation rates.

Lett Appl Microbiol, 2004, 39(3), 290 - 5
Quantification of Listeria monocytogenes in fermented sausages by MPN-PCR method; Martin B et al.; AIMS: To combine the principles of most-probable-number (MPN) statistics and the conventional PCR technique to enumerate Listeria monocytogenes in fermented sausages . METHODS AND RESULTS: A simple method to enumerate L . monocytogenes in fermented sausages was developed and compared with direct plating in Palcam agar . Species-specific MPN-PCR, but not direct plating, made the enumeration of L . monocytogenes possible in all assayed samples . CONCLUSIONS: MPN-PCR proved to be a rapid and reliable method for enumerating L . monocytogenes in fermented sausages, including low contaminated samples . SIGNIFICANCE AND IMPACT OF THE STUDY: This MPN-PCR technique may facilitate the enumeration of L . monocytogenes for routine analyses in fermented sausages without excessive work.

Expert Rev Vaccines, 2004 Aug, 3(4 Suppl), S119 - 34
Progress towards the use of Listeria monocytogenes as a live bacterial vaccine vector for the delivery of HIV antigens; Paterson Y et al.; Listeria monocytogenes is a facultative intracellular bacterium that enters the cell by phagocytosis after which it colonizes the cytosol of the host cell . It is thus a potent vaccine vector for the presentation of passenger antigens to the major histocompatability complex class II and class I pathways of antigen processing and presentation . This article shall review the progress made in developing this unusual bacterium as a vaccine vector . In mouse models, recombinant Listeria carrying a number of different antigens have been shown to provide protective immunity against infectious organisms and therapeutic immunity directed towards tumor-associated antigens . Listeria has been engineered to express a number of HIV/SIV antigens . Measurements of immune responses using these recombinant strains in the mouse, after oral and parenteral immunization, and in the rhesus macaque after oral immunization indicate that strong cell-mediated immunity can be induced against these antigens . This review also discusses safety issues associated with live bacterial vaccine vectors and problems to be overcome in developing Listeria as a HIV vaccine for human use.

Immunol Cell Biol, 2004 Aug, 82(4), 370 - 6
Changes in the immune functions and susceptibility to Listeria monocytogenes infection in mice fed dietary lipids; Puertollano MA et al.; The direct examination of the effects that fish oil diets (composed of long-chain n-3 polyunsaturated fatty acids) exert on immune system function indicates a reduction of host natural resistance to infectious diseases mainly because of a suppression of immune function generated by the fatty acids contained in this diet . Here, we evaluated the concentration of IL-12, IL-4, prostaglandin E2 and leukotriene B4 in the serum from BALB/c mice receiving four different diets . Each group was fed a diet that differed only in the source of fat: a low-fat diet (2.5% by weight), an olive oil diet (20% by weight), a fish oil diet (20% by weight) or a hydrogenated coconut oil diet (20% by weight) . Mice were fed for 4 weeks and then infected with the intracellular pathogen Listeria monocytogenes . An initial reduction in the Th1-type response as a result of a decrease in IL-12p70 secretion, an inefficient action of IL-4 (Th2-type response) and no modification of pro-inflammatory lipid-mediator production could be, at least in part, the key events responsible for the inadequate elimination of L . monocytogenes from the spleens of mice fed a fish oil diet . Furthermore, our results suggest that the type of dietary lipids may affect the circulating concentration of IL-12p70 and IL-4, leading to a modulation in the protective cellular immune response to L . monocytogenes infection.

Infect Immun, 2004 Aug, 72(8), 4401 - 9
Sequence and binding activity of the autolysin-adhesin Ami from epidemic Listeria monocytogenes 4b; Milohanic E et al.; Ami is an autolytic amidase from Listeria monocytogenes that is targeted to the bacterial surface via its C-terminal cell wall anchoring (CWA) domain . We recently showed that the CWA domain from Ami of L . monocytogenes EGD (serovar 1/2a) (Ami 1/2a) mediated bacterial binding to mammalian cells . Here we studied the sequence and binding properties of Ami from CHUT 82337 (serovar 4b) (Ami 4b) . The Ami 4b polypeptide is predicted to be 770 amino acids long (compared with the 917 amino acids of Ami 1/2a from EGD) . Ami 1/2a and Ami 4b are almost identical in the N-terminal enzymatic domain (approximately 98% amino acid identity), but the sequence is poorly conserved in the C-terminal CWA domain, with only approximately 54% amino acid identity and eight GW modules in Ami 1/2a compared with six GW modules in Ami 4b . The purified Ami 4b CWA domain efficiently bound serovar 4b bacterial cells and only poorly bound serovar 1/2a bacterial cells . The Ami 4b CWA domain was also significantly less able to bind Hep-G2 human hepatocytic cells than the Ami 1/2a CWA domain . We sequenced the ami regions encoding CWA domains of reference strains belonging to the 12 L . monocytogenes serovars . The phylogenic tree constructed from the sequences yielded a binary division into group I (serovars 1/2a, 1/2b, 1/2c, 3a, 3b, 3c, and 7) and group II (serovars 4a, 4b, 4c, 4d, and 4e) . This is the first direct evidence of divergence between serovars 1/2a and 4b in a gene involved in the adhesion of L . monocytogenes to mammalian cells, as well as the first demonstration of allelic polymorphism correlated with the somatic antigen in this species.

J Food Prot, 2004 Jul, 67(7), 1507 - 11
The BAX PCR assay for screening Listeria monocytogenes targets a partial putative gene lmo2234; Zhang W et al.; The BAX PCR for screening Listeria monocytogenes is a commercial PCR assay for specifically targeting L . monocytogenes, a foodborne pathogen that can contaminate a variety of foods and cause a potentially fatal disease, listeriosis, among high-risk populations . The high specificity (> 98%) of this PCR assay is achieved by targeting a species-specific genomic region (approximately 400 bp) presumably found only in L . monocytogenes . In this study, the identity of the BAX PCR-targeted genomic region was determined by using PCR cloning, DNA sequencing, and basic local alignment search tool (BLAST) analysis of the amplicon sequences of an L . monocytogenes serotype 1/2a strain . BLAST analysis identified the BAX PCR amplicon (GenBank accession no . AY364605) as a 423-bp genomic region between nucleotides 224,409 and 224,831 in the genome of L . monocytogenes (serotype 1/2a strain EGD-e), including a 145-bp noncoding region and a 278-bp partial coding sequence of a putative gene, lmo2234 . The translated amino acid sequence (92 amino acids) of this partial coding region is highly conserved between L . monocytogenes and Listeria innocua (93% homology) . Reverse-position-specific BLAST analysis identified a conserved domain in Lmo2234 that was similar (95.3% aligned, E value = 9E-18) to the consensus amino acid sequence of sugar phosphate isomerases/epimerases (National Center for Biotechnology Information conserved domain database accession no . COG 1082.1, IolE), indicating that Lmo2234 might be involved in bacterial carbohydrate transport and metabolism.

J Food Prot, 2004 Jul, 67(7), 1417 - 28
Distribution of Listeria monocytogenes molecular subtypes among human and food isolates from New York State shows persistence of human disease--associated Listeria monocytogenes strains in retail environments; Sauders BD et al.; While there is considerable information available regarding Listeria monocytogenes contamination patterns in food processing plants, our understanding of L . monocytogenes contamination and transmission in retail operations is limited . We characterized 125 food, 40 environmental, and 342 human clinical L . monocytogenes isolates collected in New York State from 1997 to 2002 using automated ribotyping and hly allelic variation . All environmental isolates were obtained from retail establishments and the majority of food isolates (98 isolates) were obtained from foods that were prepared or handled at retail . Overall, food and/or environmental isolates from 50 different retail establishments were characterized . The 125 food and 40 environmental isolates were differentiated into 29 and 10 ribotypes, respectively . For 16 retail establishments, we found evidence for persistence of one or more specific L . monocytogenes strains as indicated by isolation of the same EcoRI ribotype from food or environmental samples collected in a given establishment on different days . The human isolates were differentiated into 48 ribotypes . Statistical analyses showed that two ribotypes were significantly (P < 0.0001) more common among food isolates as compared with human isolates . However, a total of 17 ribotypes found among the human clinical isolates were also found among the food and environmental isolates . We conclude that L . monocytogenes, including subtypes that have been linked to human disease, can persist in retail environments . Implementation of Listeria control procedures in retail operations, which process and handle products that permit the growth of L . monocytogenes, are thus a critical component of a farm-to-table L . monocytogenes control program.

J Food Prot, 2004 Jul, 67(7), 1408 - 16
Use of carvacrol and cymene to control growth and viability of Listeria monocytogenes cells and predictions of survivors using frequency distribution functions; Periago PM et al.; The antibacterial action of carvacrol and cymene on two Listeria monocytogenes strains (STCC4031 and NCTN4032) was studied . Carvacrol or cymene showed inhibitory effect on the growth of L . monocytogenes during lag and exponential growth phases and was more evident with increasing concentrations in brain heart infusion broth at 30 degrees C . Carvacrol or cymene also decreased the survival of mid-exponential-growth-phase L . monocytogenes STCC4031 cells in potassium-N-2-hydroxy-ethylpiperazine-N-ethanesulfo nic acid, at 30 degrees C . The combination of carvacrol and cymene resulted in an increased antibacterial effect on the growth and a synergistic effect on the viability of L . monocytogenes compared with the natural compounds applied separately . The analysis of survival curves by the Weibull frequency distribution function allowed an accurate prediction of the level of inactivation achieved . Interestingly, an important bactericidal effect (4.7-log reduction) of low concentrations of both antimicrobials combined (0.75 mM) was observed on L . monocytogenes in carrot juice . This study indicates the potential use of carvacrol and cymene applied simultaneously for preservation of minimally processed foods.

Cochrane Database Syst Rev . 2004;(3):CD003069.
Interventions for replacing missing teeth: maintaining health around dental implants; Esposito M et al.; BACKGROUND: To maintain healthy tissues around dental implants it is important to institute an effective preventive regimen (supportive therapy) . Different maintenance regimens have been suggested, however it is unclear which are the most effective . OBJECTIVES: To test the null hypothesis of no difference between different interventions for maintaining healthy tissues around dental implants . SEARCH STRATEGY: We searched the Cochrane Oral Health Group's Trials Register, the Cochrane Central Register of Controlled Trials (CENTRAL), MEDLINE and EMBASE . Handsearching included several dental journals . We checked the bibliographies of the identified randomised controlled trials (RCTs) and relevant review articles for studies outside the handsearched journals . We wrote to authors of all identified RCTs, to more than 55 oral implant manufacturers and an internet discussion group to find unpublished or ongoing RCTs . No language restrictions were applied . The last electronic search was conducted on 2 February 2004 . SELECTION CRITERIA: All randomised controlled trials of oral implants comparing agents or interventions for maintaining or recovering healthy tissues around dental implants . DATA COLLECTION AND ANALYSIS: We carried out a quality assessment of the included RCTs in duplicate and contacted the authors for missing information . We independently extracted the data in duplicate . We followed the Cochrane Oral Health Group's statistical guidelines . MAIN RESULTS: Fourteen RCTs were identified . Five of these trials, which reported results from a total of 127 patients, were suitable for inclusion in the review . Two trials evaluated the efficacy of powered and sonic toothbrushes, respectively, when compared to manual toothbrushing and showed no statistically significant differences . One RCT compared Listerine versus placebo mouthwashes showing a reduction of 54% in plaque and 34% in marginal bleeding compared with the placebo . One trial compared self administered subgingival chlorhexidine irrigation versus chlorhexidine mouthwash . The group using chlorhexidine irrigation resulted in statistically significantly lower mean plaque scores and a marginal bleeding index than the group using chlorhexidine mouthwash, however the mouthwash was given at a suboptimal dosage . One study compared etching gel with mechanical debridement showing no statistical differences . Follow ups ranged between 6 weeks and 5 months . It was not possible to make any meta-analysis as each trial assessed different interventions . REVIEWERS' CONCLUSIONS: There is only little reliable evidence for which are the most effective interventions for maintaining health around peri-implant tissues . There was no evidence that the use of powered or sonic toothbrushes was superior to manual toothbrushing . There is weak evidence that Listerine mouthwash, used twice a day for 30 seconds, as adjunct to routine oral hygiene is effective in reducing plaque formation and marginal bleeding around implants . There was no evidence that phosphoric etching gel offered any clinical advantage over mechanical debridement . These findings are based on RCTs having short follow-up periods and few subjects . There is not any reliable evidence for the most effective regimens for long term maintenance . More RCTs should be conducted in this area . In particular, there is a definite need for trials powered to find possible differences, using primary outcome measures and with much longer follow up . Such trials should be reported according the CONSORT guidelines .

J Immunol, 2004 Aug 1, 173(3), 1994 - 2002
Neutrophil involvement in cross-priming CD8+ T cell responses to bacterial antigens; Tvinnereim AR et al.; Substantial CD8(+) T cell responses are generated after infection of mice with recombinant Listeria monocytogenes strains expressing a model epitope (lymphocytic choriomeningitis virus NP(118-126)) in secreted and nonsecreted forms . L . monocytogenes gains access to the cytosol of infected cells, where secreted Ags can be accessed by the endogenous MHC class I presentation pathway . However, the route of presentation of the nonsecreted Ag in vivo remains undefined . In this study we show that neutrophil-enriched peritoneal exudate cells from L . monocytogenes-infected mice can serve as substrates for in vitro cross-presentation of both nonsecreted and secreted Ag by dendritic cells as well as for in vivo cross-priming of CD8(+) T cells . In addition, specific neutrophil depletion in vivo by low dose treatment with either of two Ly6G-specific mAb substantially decreased the relative CD8(+) T cell response against the nonsecreted, but not the secreted, Ag compared with control Ab-treated mice . Thus, neutrophils not only provide rapid innate defense against infection, but also contribute to shaping the specificity and breadth of the CD8(+) T cell response . In addition, cross-presentation of bacterial Ags from neutrophils may explain how CD8(+) T cell responses are generated against Ags from extracellular bacterial pathogens.

Expert Opin Pharmacother, 2004 Aug, 5(8), 1727 - 35
An update on the medical management of listeriosis; Hof H; It is still not quite well understood why there is no optimal or even a satisfactory antibiotic therapy for listeriosis . Although almost all Listeria strains that induce sepsis, meningitis and encephalitis, as well as many other manifestations--in particular, in immunocompromised individuals--are susceptible to most of the common antibiotics, the cure rate is only approximately 70% . The most effective regimen still consists of a combination of an aminopenicillin (amoxicillin or ampicillin) plus an aminoglycoside . In vitro, this combination is bactericidal, whereas aminopenicillin alone only exerts a weak bactericidal activity against Listeriae . These antibiotics only poorly penetrate the cerebrospinal fluid and thus, only high doses given over a prolonged period of 2-3 weeks are curative . Furthermore, Listeria monocytogenes belongs to the group of facultative intracellular bacteria, which means that a certain population is inaccessible for antibiotics . Theoretically, a drug which is endowed with bactericidal activity superior to that of ampicillin would be preferable . Furthermore, the candidate drug should easily cross the blood-brain barrier into the CNS, be able to accumulate within host cells, reach the cytoplasm and be active under these unusual conditions . Because of all these arguments, the new quinolones are of particular interest; but broad clinical data are still lacking . It is unclear as to whether antibiotics alone will be sufficient to increase the prognosis . Adjunctive therapy with immunomodulators, which are able to reconstitute the defective defence capacities, would presumably create the conditions necessary to finally resolve listeriosis.

J Bacteriol, 2004 Aug, 186(15), 4994 - 5002
Intraspecific phylogeny and lineage group identification based on the prfA virulence gene cluster of Listeria monocytogenes; Ward TJ et al.; Listeria monocytogenes is a serious food-borne pathogen that can cause invasive disease in humans and other animals and has been the leading cause of food recalls due to microbiological concerns in recent years . In order to test hypotheses regarding L . monocytogenes lineage composition, evolution, ecology, and taxonomy, a robust intraspecific phylogeny was developed based on prfA virulence gene cluster sequences from 113 L . monocytogenes isolates . The results of the multigene phylogenetic analyses confirm that L . monocytogenes comprises at least three evolutionary lineages, demonstrate that lineages most frequently (lineage 1) and least frequently (lineage 3) associated with human listeriosis are sister-groups, and reveal for the first time that the human epidemic associated serotype 4b is prevalent among strains from lineage 1 and lineage 3 . In addition, a PCR-based test for lineage identification was developed and used in a survey of food products demonstrating that the low frequency of association between lineage 3 isolates and human listeriosis cases likely reflects rarity of exposure and not reduced virulence for humans as has been previously suggested . However, prevalence data do suggest lineage 3 isolates may be better adapted to the animal production environment than the food-processing environment . Finally, analyses of haplotype diversity indicate that lineage 1 has experienced a purge of genetic variation that was not observed in the other lineages, suggesting that the three L . monocytogenes lineages may represent distinct species within the framework of the cohesion species concept.

J Virol, 2004 Aug, 78(15), 8210 - 8
Oral immunization with recombinant listeria monocytogenes controls virus load after vaginal challenge with feline immunodeficiency virus; Stevens R et al.; Recombinant Listeria monocytogenes has many attractive characteristics as a vaccine vector against human immunodeficiency virus (HIV) . Wild-type and attenuated Listeria strains expressing HIV Gag have been shown to induce long-lived mucosal and systemic T-cell responses in mice . Using the feline immunodeficiency virus (FIV) model of HIV we evaluated recombinant L . monocytogenes in a challenge system . Five cats were immunized with recombinant L . monocytogenes that expresses the FIV Gag and delivers an FIV Env-expressing DNA vaccine (LMgag/pND14-Lc-env) . Control cats were either sham immunized or immunized with wild-type L . monocytogenes (LM-wt) . At 1 year after vaginal challenge, provirus could not be detected in any of the nine tissues evaluated from cats immunized with the recombinant bacteria but was detected in at least one tissue in 8 of 10 control animals . Virus was isolated from bone marrow of four of five LMgag/pND14-Lc-env-immunized cats by use of a stringent coculture system but required CD8(+) T-cell depletion, indicating CD8(+) T-cell suppression of virus replication . Control animals had an inverted CD4:CD8 ratio in mesenteric lymph node and were depleted of both CD4(+) and CD8(+) intestinal epithelial T cells, while LMgag/pND14-Lc-env-immunized animals showed no such abnormalities . Vaginal FIV-specific immunoglobulin A was present at high titer in three LMgag/pND14-Lc-env-immunized cats before challenge and in all five at 1 year postchallenge . This study demonstrates that recombinant L . monocytogenes conferred some control of viral load after vaginal challenge with FIV.

J Immunol Methods, 2004 Jun, 289(1-2), 147 - 55
Single-chain Fv antibody with specificity for Listeria monocytogenes; Paoli GC et al.; Single chain antibodies (scFv) exhibiting specific binding to Listeria monocytogenes strains were isolated from a pool of random scFvs expressed on the surface of filamentous bacteriophages . Positive selection (panning) using L . monocytogenes was used to enrich for phage clones with the desired binding affinity, and negative selection using L . innocua and L . ivanovii was used to remove phages expressing cross-reactive antibody fragments . A single phage clone, P4:A8, was selected using two independent panning schemes . A rapid assay was devised to determine phage antibody binding specificity and was used to develop a selectivity profile for individual phage clones . The P4:A8 clone was screened against a panel of bacteria consisting of eight strains of L . monocytogenes, one each of the other six species of Listeria and nine other relevant bacterial species . A collection of individual clones from the penultimate panning was also screened against a subset of the panel of bacteria . The selectivity profiles indicate that multiple clones, including P4:A8, exhibit binding to one or more strains of L . monocytogenes without cross-reactivity toward any other species in the panel . This is the first report of a species-specific antibody for viable cells of L . monocytogenes (i.e., the ability to bind to L . monocytogenes without cross-reactivity toward any other species of Listeria).

Nat Immunol, 2004 Aug, 5(8), 809 - 17 Epub 2004 Jul 11.
CD8+ T cell contraction is controlled by early inflammation; Badovinac VP et al.; Pathogen-specific CD8(+) T cells expand in number after infection and then their numbers invariably contract by 90-95%, leaving a stable memory cell pool . The chief features of this response are programmed early after infection; however, the factors regulating contraction are mostly undefined . Here we show that antibiotic treatment before Listeria monocytogenes infection induced numbers of protective memory CD8(+) T cells similar to those in control infected mice, by a pathway without contraction . The absence of contraction correlated with decreased early inflammation and interferon-gamma production and an increased fraction of CD8(+) T cells expressing the interleukin 7 receptor at the peak of the response . Thus, contraction is controlled by early inflammation but is not essential for the generation of protective memory CD8(+) T cells after infection.

Int J Food Microbiol, 2004 Aug 1, 94(3), 323 - 8
Listeria monocytogenes isolated from cold-smoked fish products in Osaka City, Japan; Nakamura H et al.; Listeria monocytogenes contamination of ready-to-eat seafood products commercially available in Osaka was examined between 1999 and 2000 . L . monocytogenes was isolated from 12 (13%) of the 95 products tested . All positive samples were from cold-smoked fish with 9 being obtained during the summer . Thirteen isolates of L . monocytogenes were typed by pulsed-field gel electrophoresis (PFGE) and polymerase chain reaction (PCR)-based typing methods . Isolates of the same serotype originating from the same manufacturer gave similar DNA profiles, irrespective of the type of sample or date of isolation . The finding suggest that persistent strains in each manufacturing facility proliferate during the summer and contaminate products during manufacturing processes.

Phys Rev E Stat Nonlin Soft Matter Phys . 2004 Jun;69(6 Pt 1):061906 . Epub 2004 Jun 02.
Soft Listeria: actin-based propulsion of liquid drops; Boukellal H et al.; We study the motion of oil drops propelled by actin polymerization in cell extracts . Drops deform and acquire a pearlike shape under the action of the elastic stresses exerted by the actin comet, a tail of cross-linked actin filaments . We solve this free boundary problem and calculate the drop shape taking into account the elasticity of the actin gel and the variation of the polymerization velocity with normal stress . The pressure balance on the liquid drop imposes a zero propulsive force if gradients in surface tension or internal pressure are not taken into account . Quantitative parameters of actin polymerization are obtained by fitting theory to experiment.

J Appl Microbiol, 1998 Jan, 84(1), 18 - 24
Acetoin production as an indicator of growth and metabolic inhibition of Listeria monocytogenes; Romick TL et al.; It has been shown that Listeria monocytogenes produces acetoin from glucose under aerobic conditions . A defined medium with glucose as the sole carbon source was used in an aerobic shake flask culture to reliably produce acetoin . Acetoin, the reactive compound in the Voges-Proskauer test, was assayable in the medium and was used to quantify the metabolic response when inhibitors were added to the medium . Inhibitors such as lactic, acetic, propionic and benzoic acids were used to demonstrate the utility of acetoin production as an indicator of metabolic disruption . With increasing levels of inhibitor, the metabolic and growth responses were measured by acetoin production and optical density change, respectively . Both measurements decreased in a similar manner with increasing inhibitor concentrations . The data also showed the apparent mode of action of the inhibitors . A bacteriostatic effect was observed for the protonated organic acids, acetic (4 mmol l(-1)) and propionic (4 mmol l(-1)), whereas protonated lactic (4 mmol l(-1)) and benzoic (0.16 mmol l(-1)) acids gave an irreversible (apparent bacteriocidal) effect . Lactic, acetic, and propionic acids showed stimulation of metabolic activity at low concentrations, but benzoic did not . Acetoin production is a novel method for quantifying and assessing the mode of action of inhibitors against L . monocytogenes . This system can be used to screen inhibitors for applications in food safety.

Am J Dent, 2004 Feb, 17(1), 23 - 6
Use of essential oil-containing mouthrinses by xerostomic individuals: determination of potential for oral mucosal irritation; Fischman SL et al.; PURPOSE: To assess the irritation potential of an essential oil-containing mouthrinse (Listerine Antiseptic) in a population with objectively documented xerostomia (hyposalivation) using an exaggerated-exposure clinical model . METHODS: Following a baseline oral soft tissue examination, 19 qualifying female subjects with a mean age of 61 years and a mean unstimulated baseline salivary flow of 0.06 mL/min were randomly assigned either the essential oil mouthrinse or a negative control rinse . They rinsed under supervision with 20 ml of their assigned rinse for 30 seconds and 5 minutes later a second salivary flow rate was determined . They then rinsed unsupervised with 20 ml for 30 seconds three times daily for the next 14 days, and received soft tissue examinations on days 7 and 14 . After a 1-week interim period, subjects switched to the alternate rinse and the examination and rinsing regimens were repeated during the subsequent 2 weeks . RESULTS: The oral irritation potential of the essential oil mouthrinse was minimal . Oral mucosal abnormalities attributable to the test rinses were seen in only 2 subjects, both at the 7-day examination . These subjects were both using the essential oil mouthrinse . The abnormalities consisted of an asymptomatic "whitish slough" which was readily wiped off leaving a normal appearing, non-erythematous mucosa . In both subjects, the oral mucosa appeared normal at the 14-day examination.

J Immunol, 2004 Jul 15, 173(2), 969 - 75
Fully functional memory CD8 T cells in the absence of CD4 T cells; Marzo AL et al.; The role of CD4 T cells in providing help to CD8 T cells in primary and secondary responses to infection remains controversial . Using recombinant strains of virus and bacteria expressing the same Ag, we determined the requirement for CD4 T cells in endogenous CD8 T cell responses to infection with vesicular stomatitis virus and Listeria monocytogenes (LM) . Depletion of CD4 T cells had no effect on the frequency of primary or secondary vesicular stomatitis virus-specific CD8 T cells in either lymphoid or nonlymphoid tissues . In contrast, the primary LM-specific CD8 T cell response was CD4 T cell dependent . Surprisingly, the LM-specific CD8 T cell recall response was also CD4 T cell dependent, which correlated with a requirement for CD40/CD40L interactions . However, concomitant inhibition of CD40L and CD4 T cell removal revealed that these pathways may be operating independently . Importantly, despite the absence of CD4 T cells during the recall response or throughout the entire response, CD8 memory T cells were functional effectors and proliferated equivalently to their "helped" counterparts . These data call into question the contention that CD4 T cells condition memory CD8 T cells during the primary response and indicate that the principal role of CD4 T cells in generating CD8 memory cells after infection is augmentation of proliferation or survival through costimulatory signals.

Appl Environ Microbiol, 2004 Jul, 70(7), 4256 - 66
Natural atypical Listeria innocua strains with Listeria monocytogenes pathogenicity island 1 genes; Johnson J et al.; Identification of bona fide Listeria isolates into the six species of the genus normally requires only a few tests . Aberrant isolates do occur, but even then only one or two extra confirmatory tests are generally needed for identification to species level . We have discovered a hemolytic-positive, rhamnose and xylose fermentation-negative Listeria strain with surprising recalcitrance to identification to the species level due to contradictory results in standard confirmatory tests . The issue had to be resolved by using total DNA-DNA hybridization testing and then confirmed by further specific PCR-based tests including a Listeria microarray assay . The results show that this isolate is indeed a novel one . Its discovery provides the first fully documented instance of a hemolytic Listeria innocua strain . This species, by definition, is typically nonhemolytic . The L . innocua isolate contains all the members of the PrfA-regulated virulence gene cluster (Listeria pathogenicity island 1) of L . monocytogenes . It is avirulent in the mouse pathogenicity test . Avirulence is likely at least partly due to the absence of the L . monocytogenes-specific allele of iap, as well as the absence of inlA, inlB, inlC, and daaA . At least two of the virulence cluster genes, hly and plcA, which encode the L . monocytogenes hemolysin (listeriolysin O) and inositol-specific phospholipase C, respectively, are phenotypically expressed in this L . innocua strain . The detection by PCR assays of specific L . innocua genes (lin0198, lin0372, lin0419, lin0558, lin1068, lin1073, lin1074, lin2454, and lin2693) and noncoding intergenic regions (lin0454-lin0455 and nadA-lin2134) in the strain is consistent with its L . innocua DNA-DNA hybridization identity . Additional distinctly different hemolytic L . innocua strains were also studied.

Appl Environ Microbiol, 2004 Jul, 70(7), 4158 - 64
Epidemic clone I-specific genetic markers in strains of Listeria monocytogenes serotype 4b from foods; Yildirim S et al.; Listeria monocytogenes contamination of ready-to-eat foods has been implicated in numerous outbreaks of food-borne listeriosis . However, the health hazards posed by L . monocytogenes detected in foods may vary, and speculations exist that strains actually implicated in illness may constitute only a fraction of those that contaminate foods . In this study, examination of 34 serogroup 4 (putative or confirmed serotype 4b) isolates of L . monocytogenes obtained from various foods and food-processing environments, without known implication in illness, revealed that many of these strains had methylation of cytosines at GATC sites in the genome, rendering their DNA resistant to digestion by the restriction endonuclease Sau3AI . These strains also harbored a gene cassette with putative restriction-modification system genes as well as other, genomically unlinked genetic markers characteristic of the major epidemic-associated lineage of L . monocytogenes (epidemic clone I), implicated in numerous outbreaks in Europe and North America . This may reflect a relatively high fitness of strains with these genetic markers in foods and food-related environments relative to other serotype 4b strains and may partially account for the repeated involvement of such strains in human food-borne listeriosis.

Appl Environ Microbiol, 2004 Jul, 70(7), 4111 - 7
CO2- and anaerobiosis-induced changes in physiology and gene expression of different Listeria monocytogenes strains; Jydegaard-Axelsen AM et al.; Although carbon dioxide (CO(2)) is known to inhibit growth of most bacteria, very little is known about the cellular response . The food-borne pathogen Listeria monocytogenes is characterized by its ability to grow in high CO(2) concentrations at refrigeration temperatures . We examined the listerial responses of different strains to growth in air, 100% N(2), and 100% CO(2) . The CO(2)-induced changes in membrane lipid fatty acid composition and expression of selected genes were strain dependent . The acid-tolerant L . monocytogenes LO28 responded in the same manner to CO(2) as to other anaerobic, slightly acidic environments (100% N(2), pH 5.7) . An increase in the expression of the genes encoding glutamate decarboxylase (essential for survival in strong acid) as well as an increased amount of branched-chain fatty acids in the membrane was observed in both atmospheres . In contrast, the acid-sensitive L . monocytogenes strain EGD responded differently to CO(2) and N(2) at the same pH . In a separate experiment with L . monocytogenes 412, an increased isocitrate dehydrogenase activity level was observed for cells grown in CO(2)-containing atmospheres . Together, our findings demonstrate that the CO(2)-response is a partly strain-dependent complex mechanism . The possible links between the CO(2)-dependent changes in isocitrate dehydrogenase activity, glutamate metabolism and branched fatty acid biosynthesis are discussed.

Appl Environ Microbiol, 2004 Jul, 70(7), 4021 - 9
Effect of high-pressure-induced ice I-to-ice III phase transitions on inactivation of Listeria innocua in frozen suspension; Luscher C et al.; The inactivation of Listeria innocua BGA 3532 at subzero temperatures and pressures up to 400 MPa in buffer solution was studied to examine the impact of high-pressure treatments on bacteria in frozen matrices . The state of aggregation of water was taken into account . The inactivation was progressing rapidly during pressure holding under liquid conditions, whereas in the ice phases, extended pressure holding times had comparatively little effect . The transient phase change of ice I to other ice polymorphs (ice II or ice III) during pressure cycles above 200 MPa resulted in an inactivation of about 3 log cycles, probably due to the mechanical stress associated with the phase transition . This effect was independent of the applied pressure holding time . Flow cytometric analyses supported the assumption of different mechanisms of inactivation of L . innocua in the liquid phase and ice I (large fraction of sublethally damaged cells due to pressure inactivation) in contrast to cells subjected to ice I-to-ice III phase transitions (complete inactivation due to cell rupture) . Possible applications of high-pressure-induced phase transitions include cell disintegration for the recovery of intracellular components and inactivation of microorganisms in frozen food.

Int J Food Microbiol, 2004 Aug 15, 95(1), 1 - 9
New chromogenic plating media for detection and enumeration of pathogenic Listeria spp.--an overview; Reissbrodt R; In recent years a number of selective chromogenic plating media for pathogenic Listeria spp . have been developed and marketed . Their advantages are direct detection and enumeration of pathogenic Listeria spp . utilizing cleavage of substrates by the virulence factor phosphatidylinositol-phospholipase C (PI-PLC) and, to a lesser extent, by phosphatidylcholin-phospholipase C (PC-PLC) . There are two groups of such media: the first utilizes cleavage by PI-PLC of L-alpha-phosphatidyl-inositol, forming a white precipitation zone around the colony, combined with the chromogenic substrate 5-bromo-4-chloro-3-indoxyl-beta-D-glucopyranoside for detection of beta-d-glucosidase, which occurs in all Listeria spp . All Listeria spp . produce turquoise colonies on these media which include ALOA , CHROMagar Listeria, BBL CHROMagar Listeria, and OCLA . The second group of media utilizes 5-bromo-4-chloro-3-indoxyl-myoinositol-1-phosphate, forming blue-turquoise colonies of pathogenic Listeria spp . and white colonies of non-pathogenic Listeria spp . BCM trade mark Listeria monocytogenes plating medium, Rapid'L.mono and LIMONO-Ident-Agar belong to this group . Selective chromogenic L . monocytogenes plating media offer the attraction of rapid economic detection and enumeration of pathogenic Listeria spp . within 24 or 48 h of incubation at 36+/-1 degrees C . This overview summarises the characteristics of these chromogenic plating media, reviews important evaluations, and focuses on replacement of conventional by these chromogenic plating media, particularly for applications in the food industry.

Cell Microbiol, 2004 Aug, 6(8), 761 - 9
A Rickettsia WASP-like protein activates the Arp2/3 complex and mediates actin-based motility; Jeng RL et al.; Spotted fever group Rickettsia are obligate intracellular pathogens that exploit the host cell actin cytoskeleton to promote motility and cell-to-cell spread . Although other pathogens such as Listeria monocytogenes use an Arp2/3 complex-dependent nucleation mechanism to generate comet tails consisting of Y-branched filament arrays, Rickettsia polymerize tails consisting of unbranched filaments by a previously unknown mechanism . We identified genes in several Rickettsia species encoding proteins (termed RickA) with similarity to the WASP family of Arp2/3-complex activators . Rickettsia rickettsii RickA activated both the nucleation and Y-branching activities of the Arp2/3 complex like other WASP-family proteins, and was sufficient to direct the motility of microscopic beads in cell extracts . Actin tails generated by RickA-coated beads consisted of Y-branched filament networks . These data suggest that Rickettsia use an Arp2/3 complex-dependent actin-nucleation mechanism similar to that of other pathogens . We propose that additional Rickettsia or host factors reorganize the Y-branched networks into parallel arrays in a manner similar to a recently proposed model of filopodia formation.

J Vet Med B Infect Dis Vet Public Health, 2004 May, 51(4), 176 - 9
Clinical and histopathological aspects of naturally occurring mastitis caused by Listeria monocytogenes in cattle and ewes; Winter P et al.; Listeria monocytogenes was isolated from the milk of two cows and two sheep with mastitis in one quarter and one udder half . The animals were observed over a period of 2-12 months . Clinical examination of the udder, bacteriological examinations and determination of somatic cell counts of milk samples were performed monthly . All four cases suffered from a subclinical mastitis characterized by an elevated somatic cell count (0.8-10.1 x 10(6) cells/ml), a persistent shedding of Listeria and by a normal appearance of the milk . The animals did not show any systemic reaction, but all animals developed an atrophy of the infected mammary gland . Histological examinations revealed a chronic interstitial mastitis with diffuse infiltration of lymphocytes, plasma cells and macrophages . All internal organs showed no abnormalities, no Listeria could be isolated . Listeria could however be isolated from the affected mammary parenchyma and from the mammary lymph node . The results of the bacteriological examination could be confirmed by means of PCR . Using PFGE, all the isolates from the same animal were identical . Immunohistochemical examination of the ovine mammary glands achieved a very strong immunoreactivity for CD5 cells . The mode of infection and the reaction of the immune system's defense of the ovine udders are discussed.

Mol Microbiol, 2004 Jul, 53(2), 639 - 49
FbpA, a novel multifunctional Listeria monocytogenes virulence factor; Dramsi S et al.; Listeria monocytogenes is a Gram-positive intracellular bacterium responsible for severe opportunistic infections in humans and animals . Signature-tagged mutagenesis (STM) was used to identify a gene named fbpA, required for efficient liver colonization of mice inoculated intravenously . FbpA was also shown to be required for intestinal and liver colonization after oral infection of transgenic mice expressing human E-cadherin . fbpA encodes a 570-amino-acid polypeptide that has strong homologies to atypical fibronectin-binding proteins . FbpA binds to immobilized human fibronectin in a dose-dependent and saturable manner and increases adherence of wild-type L . monocytogenes to HEp-2 cells in the presence of exogenous fibronectin . Despite the lack of conventional secretion/anchoring signals, FbpA is detected using an antibody generated against the recombinant FbpA protein on the bacterial surface by immunofluorescence, and in the membrane compartment by Western blot analysis of cell extracts . Strikingly, FbpA expression affects the protein levels of two virulence factors, listeriolysin O (LLO) and InlB, but not that of InlA or ActA . FbpA co-immunoprecipitates with LLO and InlB, but not with InlA or ActA . Thus, FbpA, in addition to being a fibronectin-binding protein, behaves as a chaperone or an escort protein for two important virulence factors and appears as a novel multifunctional virulence factor of L . monocytogenes.

J Food Prot, 2004 Jun, 67(6), 1238 - 42
Efficacy of chlorine and a peroxyacetic acid sanitizer in killing Listeria monocytogenes on iceberg and Romaine lettuce using simulated commercial processing conditions; Beuchat LR et al.; The efficacy of chlorine (100 microg/ml) and a peroxyacetic acid sanitizer (80 microg/ml; Tsunami 100) in killing Listeria monocytogenes inoculated at populations of 1 to 2, 2 to 3, and 4 to 5 log CFU/g of iceberg lettuce pieces, shredded iceberg lettuce, and Romaine lettuce pieces was determined by treatment conditions simulating those used by a commercial fresh-cut lettuce processor . The lettuce/treatment solution ratio was 1:100 (wt/vol), treatment temperature was 4 degrees C, and total treatment time was 30 s . Compared with washing in water, treatment of iceberg lettuce pieces containing all levels of inoculum and shredded iceberg lettuce containing 2 to 3 or 4 to 5 log CFU/g with chlorine or Tsunami resulted in significant reductions (P < or = 0.05) of pathogen populations . Populations recovered from Romaine lettuce pieces treated with chlorine or Tsunami were not significantly different from populations recovered from pieces washed with water, regardless of the inoculum level . Within lettuce type and inoculum level, in no instance was the number of L . monocytogenes recovered from lettuce treated with chlorine or Tsunami significantly different . The rate of decrease in free chlorine concentration in treatment solution as affected by the weight/volume ratio (1:100, 1:10, 2:10, and 4:10) of lettuce and solution was determined . The rate of reduction increased as the ratio decreased . The overall order of magnitude of reduction was shredded iceberg lettuce > iceberg pieces > Romaine pieces . The highest reductions in free chlorine concentration in solutions used to treat shredded lettuce are attributed to the release of tissue juices, which increases the concentration of soluble organic materials available for reaction with chlorine.

J Food Prot, 2004 Jun, 67(6), 1170 - 6
Survival of Listeria monocytogenes strain H7762 and resistance to simulated gastric fluid following exposure to frankfurter exudate; Wonderling LD et al.; Listeria monocytogenes strain H7762, a frankfurter isolate, was tested to determine whether it was able to survive at 4 degrees C in frankfurter pack fluid (exudate) and to determine whether food exposure affects its acid sensitivity . Cultures were sampled and tested for acid sensitivity by challenge with simulated gastric fluid (SGF) . SGF challenges performed immediately after inoculation revealed that between 20 and 26% of the cells survived the full 30 min of SGF challenge regardless of whether the cells were inoculated into brain heart infusion broth (BHI) or exudate . After 2 days of incubation, cells exposed to both exudate and BHI had significantly decreased SGF resistance; however, the cells exposed to exudate were significantly more SGF resistant than cells exposed to BHI (after 15 min of SGF treatment, 33% of the exudate-exposed cells survived and 12% of the BHI-exposed cells survived) . L . monocytogenes exposed to exudate had greater SGF resistance at all challenge times compared with BHI-exposed cells from day 2 through day 4 . From days 8 to 15, exudate-exposed cells continued to have greater SGF resistance than BHI-exposed cells up to 10 min of SGF challenge but were as sensitive as the BHI-exposed cells at 20 to 30 min of challenge . By day 25, cells exposed to exudate were significantly more sensitive to SGF challenge than BHI-exposed cells . The survivor data generated from SGF challenges were modeled by a nonlinear regression analysis to calculate the underlying distribution of SGF resistance found in the challenged populations . These analyses indicated that L . monocytogenes exposed to exudate at 4 degrees C had a broader distribution of resistance to SGF compared with cells exposed to BHI at 4 degrees C . In addition, the mean time of death during SGF treatment was greater after exposure to exudate, indicating that cells exposed to exudate were more resistant to killing by SGF These data suggest that exposure to frankfurter exudate might render L . monocytogenes more able to survive the stomach environment during the initial stages of infection.

J Food Prot, 2004 Jun, 67(6), 1163 - 9
Impact of intervention strategies on Listeria contamination patterns in crawfish processing plants: a longitudinal study; Lappi VR et al.; Two ready-to-eat crawfish processing plants were monitored for 2 years to study the impact of Listeria control strategies, including employee training and targeted sanitation procedures, on Listeria contamination . Environmental, raw material, and finished product samples were collected weekly during the main processing months (April to June) and tested for Listeria spp . and Listeria monocytogenes . Before implementation of control strategies (year 1), the two processing plants showed Listeria spp . prevalences of 29.5% (n = 78) in raw, whole crawfish, 5.2% (n = 155) in the processing plant environment, and 0% (n = 78) in finished products . In year 2, after plant-specific Listeria control strategies were implemented, Listeria spp . prevalence increased in raw crawfish (57.5%, n = 101), in the processing plant environment (10.8%, n = 204), and in the finished product (1.0%, n = 102) . Statistical analysis showed a significant increase in Listeria spp . prevalence (P < 0.0001) and a borderline nonsignificant increase in L . monocytogenes prevalence (P = 0.097) on raw material in year 2 . Borderline nonsignificant increases were also observed for Listeria spp . prevalence in environmental samples (P = 0.082) . Our data showed that Listeria spp . prevalence in raw crawfish can vary significantly among seasons . However, the increased contamination prevalence for raw materials only resulted in a limited Listeria prevalence increase for the processing plant environment with extremely low levels of finished product contamination . Heat treatment of raw materials combined with Listeria control strategies to prevent cross-contamination thus appears to be effective in achieving low levels of finished product contamination, even with Listeria spp . prevalences for raw crawfish of more than 50%.

Biometals, 2004 Jun, 17(3), 197 - 202
Iron and proteins for iron storage and detoxification; Chiancone E et al.; Iron is required by most organisms, but is potentially toxic due to the low solubility of the stable oxidation state, Fe(III), and to the tendency to potentiate the production of reactive oxygen species, ROS . The reactivity of iron is counteracted by bacteria with the same strategies employed by the host, namely by sequestering the metal into ferritin, the ubiquitous iron storage protein . Ferritins are highly conserved, hollow spheres constructed from 24 subunits that are endowed with ferroxidase activity and can harbour up to 4500 iron atoms as oxy-hydroxide micelles . The release of the metal upon reduction can alter the microorganism-host iron balance and hence permit bacteria to overcome iron limitation . In bacteria, the relevance of the Dps (DNA-binding proteins from starved cells) family in iron storage-detoxification has been recognized recently . The seminal studies on the protein from Listeria innocua demonstrated that Dps proteins have ferritin-like activity and most importantly have the capacity to attenuate the production of ROS . This latter function allows bacterial pathogens that lack catalase, e.g . Porphyromonas gingivalis, to survive in an aerobic environment and resist to peroxide stress.

Blood, 2004 Oct 15, 104(8), 2397 - 402 Epub 2004 Jun 24.
Targeted deletion of T-cell clones using alpha-emitting suicide MHC tetramers; Yuan RR et al.; Immunosuppressive agents in current use are nonspecific . The capacity to delete specific CD8 T-cell clones of unique specificity could prove to be a powerful tool for dissecting the precise role of CD8(+) T cells in human disease and could form the basis for a safe, highly selective therapy of autoimmune disorders . Major histocompatibility complex (MHC) tetramers (multimeric complexes capable of binding to specific CD8 T-cell clones) were conjugated to (225)Ac (an alpha-emitting atomic nanogenerator, capable of single-hit killing from the cell surface) to create an agent for CD8 T-cell clonal deletion . The "suicide" tetramers specifically bound to, killed, and reduced the function of their cognate CD8 T cells (either human anti-Epstein-Barr virus (EBV) or mouse anti-Listeria in 2 model systems) while leaving the nonspecific control CD8 T-cell populations unharmed . Such an approach may allow a pathway to selective ablation of pathogenic T-cell clones ex vivo or in vivo without disturbing general immune function.

Infect Immun, 2004 Jul, 72(7), 4318 - 21
Oral inoculation of A/J mice for detection of invasiveness differences between Listeria monocytogenes epidemic and environmental strains; Kim SH et al.; Four-week-old Harlan A/J mice were orally infected with six epidemic and six environmental strains of Listeria monocytogenes . Epidemic strains were significantly more invasive as a group than were environmental strains (P < 0.05), and the intestines of some mice infected with epidemic strains had extensive hemorrhage . Mice inoculated with epidemic strains were significantly more likely to become systemically infected than mice inoculated with environmental strains (P < 0.01).

Infect Immun, 2004 Jul, 72(7), 3849 - 54
Intestinal P glycoprotein acts as a natural defense mechanism against Listeria monocytogenes; Neudeck BL et al.; Mechanisms by which the intestinal epithelium resists invasion by food-borne pathogens such as Listeria monocytogenes are an evolving area of research . Intestinal P glycoprotein is well known to limit the absorption of xenobiotics and is believed to act as a cytotoxic defense mechanism . The aim of this study was to determine if intestinal P glycoprotein is involved in host defense against L . monocytogenes . Caco-2 cells and a P-glycoprotein-overexpressing subclone (Caco-2/MDR) were employed in addition to mdr1a(-/-) mice and wild-type controls . In vitro invasion assays and in vivo experiments were employed to measure bacterial invasion and dissemination . In addition, L . monocytogenes proteins were labeled with {(35)S}methionine, and the transepithelial transport across Caco-2 monolayers was characterized in both directions . Overexpression of P glycoprotein in Caco-2/MDR cells led to increased resistance to L . monocytogenes invasion, whereas P-glycoprotein inhibition led to increased invasion . Flux of {(35)S}methionine-labeled L . monocytogenes proteins was significantly greater in the basolateral-to-apical direction than in the apical-to-basolateral direction, indicating dependence on an apically located efflux transporter . Moreover, inhibiting P glycoprotein reduced the basolateral-to-apical flux of the proteins . Early dissemination of L . monocytogenes from the gastrointestinal tract was significantly greater in the mdr1a(-/-) mice than in wild-type controls . Expression and function of intestinal P glycoprotein is an important determinant in resistance to early invasion of L . monocytogenes.

J Immunol, 2004 Jul 1, 173(1), 420 - 7
Listeria monocytogenes as a vaccine vector: virulence attenuation or existing antivector immunity does not diminish therapeutic efficacy; Starks H et al.; The bacterium L . monocytogenes is a proposed vaccine carrier based upon the observation that this pathogen replicates within the intracytoplasmic environment facilitating delivery of Ag to the endogenous Ag processing and presentation pathway with subsequent stimulation of peptide specific MHC class I-restricted CD8(+) effector cells . In this report, we evaluate virulence-attenuated strains of Listeria monocytogenes as vaccine vectors and examine whether existing antivector (antilisterial) immunity limits or alters its efficacy as a therapeutic cancer vaccine . Following immunization with virulence-attenuated mutants, we found that the effectiveness of L . monocytogenes as a recombinant cancer vaccine remains intact . In addition, we found that antibiotic treatment initiated 24 or 36 h following therapeutic immunization with recombinant L . monocytogenes allows full development of the antitumor response . We also demonstrate that the vaccine vector potential of L . monocytogenes is not limited in animals with existing antilisterial immunity . For these latter studies, mice previously immunized with wild-type L . monocytogenes were infused with melanoma cells and then 5 days later challenged with recombinant tumor Ag expressing L . monocytogenes . Collectively, these results add additional support for the use of L . monocytogenes as a vaccine vector and underscore its potential to be used repeatedly for stimulation of recall responses concomitant with primary cell-mediated responses to newly delivered heterologous tumor-associated epitopes.

Arch Dis Child Fetal Neonatal Ed, 2004 Jul, 89(4), F328 - 30
Preterm meconium staining of the amniotic fluid: associated findings and risk of adverse clinical outcome; Tybulewicz AT et al.; BACKGROUND: The incidence of preterm meconium staining of the amniotic fluid (MSAF) is uncertain . It may be an indicator of possible listeriosis . It is unclear how great this risk is or whether preterm MSAF is a risk factor for adverse neonatal outcome . OBJECTIVE: To investigate the incidence of preterm MSAF, the incidence of associated maternal and neonatal infection, and the outcomes of the infants at discharge . DESIGN: Retrospective case-control study . METHODS: Infants < 33 weeks gestation with preterm MSAF born in the Simpson Memorial Maternity Pavilion, Edinburgh between 1 January 1994 and 2 January 2001 were matched with the next infant of the same sex and gestation with clear liquor . Maternal and infant characteristics, culture results, placental histology, and clinical outcomes were compared . RESULTS: Preterm MSAF was observed in 45/1054 (4.3%) infants below 33 weeks gestation . No maternal or infant listeriosis was identified in cases or controls . There was no significant difference in birth weight, Apgar score, or first pH between cases and controls . Preterm MSAF was associated with prolonged rupture of the membranes (odds ratio (OR) 3.34, 95% confidence interval (CI) 1.07 to 10.49), but not maternal hypertension, sepsis, or chorioamnionitis . Severe (grade 3/4) intraventricular haemorrhage was significantly more common in infants with preterm MSAF (OR 2.03, 95% CI 1.62 to 2.53) . There was no significant difference in mortality . Early onset sepsis was observed in two cases and three controls . CONCLUSIONS: Preterm meconium staining of the amniotic fluid may be associated with increased risk of intraventricular haemorrhage . It does not appear to be a useful indicator of listeriosis.

Int J Clin Pract, 2004 May, 58(5), 536 - 8
Listeria monocytogenes pneumonia in a cirrhotic child; De Sa FR et al.; A 13-year-old girl with cirrhosis and cyanotic heart disease was admitted with a three-day history of pneumonia . The chest roentgenogram revealed left-sided pleural effusion and cultures from the pleural fluid yielded Listeria monocytogenes . The authors discuss the epidemiologic, clinical, and pathophysiological aspects of L . monocytogenes pneumonia and its association with cirrhosis.

J Bacteriol, 2004 Jul, 186(13), 4276 - 84
Heterologous expression and purification of active divercin V41, a class IIa bacteriocin encoded by a synthetic gene in Escherichia coli; Richard C et al.; Divercin V41, a class IIa bacteriocin with strong antilisterial activity, is produced by Carnobacterium divergens V41 . To express a recombinant version of divercin V41, we constructed a synthetic gene that encodes the mature divercin V41 peptide and then overexpressed the gene in pET-32b by using the T7 RNA polymerase promoter in the Escherichia coli Origami (DE3)(pLysS) strain . The DvnRV41 peptide was expressed as a translational fusion protein with thioredoxin and accumulated in the cell cytoplasm in a soluble anti-Listeria active form . The fusion protein was then purified and cleaved to obtain pure, soluble, folded DvnRV41 (462 microg per 20 ml of culture) . This paper describes the first design of a synthetic bacteriocin gene and the first bacteriocin expressed in the E . coli cytoplasm.

Vet Clin North Am Food Anim Pract, 2004 Jul, 20(2), 243 - 73, vi
Brainstem and cranial nerve abnormalities: listeriosis, otitis media/interna, and pituitary abscess syndrome; Morin DE; This article reviews three disorders associated with multiple asymmetric cranial nerve deficits in ruminants: encephalitic listeriosis,otitis media/interna, and pituitary abscess syndrome . Emphasis is placed on encephalitic listeriosis, an infectious disease of the brainstem and cranial nerves caused by Listeria monocytogenes.The epidemiology, pathophysiology, clinical manifestations, diagnosis,and treatment of encephalitic listeriosis are reviewed, and differences between cattle and small ruminants are noted . Physical and neurologic examination findings that distinguish otitis media/interna and pituitary abscess syndrome from encephalitic listeriosis are highlighted.

Curr Biol, 2004 Jun 22, 14(12), R467 - 9
Actin polymerization: forcing flat faces forward; Upadhyaya A et al.; Actin polymerization has been shown to be sufficient to propel curved objects, for example beads and vesicles coated with the Listeria monocytogenes protein ActA . Recent studies suggest that actin polymerization on flat surfaces can also provide the propulsive force to push them forward.

J Nutr Educ Behav, 2004 May-Jun, 36(3), 121 - 7
Pregnant women and listeriosis: preferred educational messages and delivery mechanisms; Cates SC et al.; OBJECTIVE: To characterize pregnant women's food safety practices, to evaluate the impact of existing educational messages on the risks and prevention of listeriosis, and to identify preferred delivery methods for educational initiatives . DESIGN: Eight focus group discussions conducted with pregnant women in 4 locations . SETTING: Focus group discussions led by moderators using a prepared moderator guide . PARTICIPANTS: Purposeful sampling was used to select the 63 pregnant women who participated in this study . The focus groups were segmented by location and education level . PHENOMENON OF INTEREST: Food safety knowledge and food-handling practices, food safety practices during pregnancy, attitudes toward listeriosis brochure, and preferred delivery methods . ANALYSIS: Focus group discussions were videotaped and audiorecorded . Detailed summaries of each discussion were prepared and systematically analyzed to identify common themes within and across groups . RESULTS: Participants were not aware of the risks of listeriosis and recommended practices for listeriosis prevention; thus, they were not taking precautions during their pregnancy to prevent listeriosis . CONCLUSIONS AND IMPLICATIONS: The study identified the need to develop educational materials on listeriosis targeted specifically to pregnant women and to partner with obstetricians and other health care providers to deliver these materials to pregnant women.

Clin Microbiol Infect, 2004 Jun, 10(6), 562 - 8
Diversity of Listeria monocytogenes isolates of human and food origin studied by serotyping, automated ribotyping and pulsed-field gel electrophoresis; Lukinmaa S et al.; Automated ribotyping, pulsed-field gel electrophoresis (PFGE) and serotyping were evaluated for the epidemiological study of isolates of Listeria monocytogenes collected in Finland in 1997-1999 from human blood (n = 116) and the food industry (n = 72) . The isolates divided into six serotypes, 23 EcoRI ribotypes, 54 AscI PFGE types, and 57 final subtypes if all results were combined . The discrimination index of ribotyping was lower (0.873) than that of PFGE (0.946) . Two final subtypes dominated among human isolates, and identical subtypes were also found among food industry isolates . All PFGE types were serotype-specific, whereas two ribotypes included isolates of two serotypes . Isolates of serotype 3a, involved in an outbreak in Finland in 1999, matched one of these ribotypes, which also included some food industry isolates of serotype 1/2a . Ribotyping with EcoRI would not have been sufficient to define the outbreak in Finland caused by serotype 3a isolates . Although ribotyping is applicable as the first method in outbreak situations, human and food isolates with identical ribotypes should be investigated further by PFGE.

Zh Mikrobiol Epidemiol Immunobiol, 2004 Mar-Apr, (2), 29 - 33
{Influence of temperature on the variability of Listeria monocytogenes in the course of its prolonged existence in soil-packed water-circulating columns}; Buzoleva LS et al.; The capacity of L . monocytogenes for prolonged existence in the soil at different temperatures (18-20 degrees C and 6-8 degrees C) has been shown . The viability of these bacteria in the soil depends both on their properties and the temperature factor . Low temperatures (6-8 degrees C) ensure, on the whole, the stable preservation of the biological properties of the bacteria . More pronounced variability of L . monocytogenes at a temperature of 18-20 degrees C is an adaptive property, ensuring high ecological plasticity of the bacteria and their prolonged existence in the soil.

J Appl Microbiol, 2004, 97(1), 68 - 77
The microbial ecology of high-risk, chilled food factories; evidence for persistent Listeria spp . and Escherichia coli strains; Holah JT et al.; AIMS: The intention of this study was to provide evidence of any Listeria spp . or Escherichia coli strain persistence, and to identify whether strains of these organisms adapt to specific environmental or product niches in food factories . METHODS AND RESULTS: A 3-year assessment of the microbial ecology of four, ready-to-eat food-processing factories was undertaken in which approx . 196 000 and 75 000 product and environmental samples were examined for Escherichia coli and Listeria spp . respectively . A total of 152 E . coli isolates (44 environmental and 108 product in 62 ribogroups) and 260 Listeria spp . isolates (174 environmental and 86 product in 30 ribogroups) were identified and ribotyped . The overall prevalence of E . coli (0.08%), all Listeria spp . (0.35%) and L . monocytogenes (0.23%) was very low . Some 10 E . coli ribogroups and 14 Listeria spp . ribogroups showed evidence for persistence, defined as the isolation of the same strain, from the same site, over a prolonged time period . The majority of E . coli strains were product niche oriented whilst the majority of Listeria spp . strains were environmental niche oriented . CONCLUSION: Current UK high-risk food factory designs, personnel hygiene and cleaning and disinfection regimes are sufficient to control Listeria spp . and E . coli to very low levels . SIGNIFICANCE AND IMPACT OF THE STUDY: Persistent strains of these organisms, however, can remain within factory high-risk production areas over considerable time periods, warranting an examination of the strain persistence mechanisms and alternative hygiene controls.

Mol Microbiol, 2004 Jun, 52(6), 1553 - 65
New Listeria monocytogenes prfA* mutants, transcriptional properties of PrfA* proteins and structure-function of the virulence regulator PrfA; Vega Y et al.; PrfA, a transcription factor structurally related to Crp/Fnr, activates Listeria monocytogenes virulence genes during intracellular infection . We report two new PrfA* mutations causing the constitutive overexpression of the PrfA regulon . Leu-140Phe lies in alphaD adjacent to the DNA-binding motif in the C-terminal domain, like a previously characterized PrfA* mutation (Gly-145Ser) . Ile-45Ser, in contrast, maps to the N-terminal beta-roll, a structure similar to that of the Crp cAMP binding site . The in vitro transcriptional properties of recombinant PrfA*(I45S) and PrfA*(G145S) were compared to those of PrfA(WT) at two differentially regulated PrfA-dependent promoters, PplcA and PactA . The two PrfA* mutations increased the affinity for the target DNA to a different extent, and the differences in DNA binding (PrfA*(G145S) > PrfA*(I45S) >>> PrfA(WT)) correlated with proportional differences in transcriptional activity . The use of the PrfA* proteins revealed that PplcA had a greater affinity for, and was more sensitive to, PrfA than PactA . RNA polymerase (RNAP) initiated transcription independently of PrfA at PplcA, but not at PactA, consistent with bandshift experiments suggesting that PplcA has a greater affinity for RNAP than PactA . Thus, differences in affinity for both PrfA and RNAP appear to determine the different expression pattern of PrfA-regulated promoters . Modelling of the PrfA* mutations in the crystal structure of PrfA and comparison with structure-function analyses of Crp, in which similar mutations lead to constitutively active (cAMP-independent) Crp* proteins, suggested that PrfA shares with Crp an analogous mechanism of cofactor-mediated allosteric shift . Our data support a regulatory model in which changes in PrfA-dependent gene expression are primarily accounted for by changes in PrfA activity.

Appl Environ Microbiol, 2004 Jun, 70(6), 3457 - 66
Identification of sigma factor sigma B-controlled genes and their impact on acid stress, high hydrostatic pressure, and freeze survival in Listeria monocytogenes EGD-e; Wemekamp-Kamphuis HH et al.; The gene encoding the alternative sigma factor sigma(B) in Listeria monocytogenes is induced upon exposure of cells to several stresses . In this study, we investigated the impact of a sigB null mutation on the survival of L . monocytogenes EGD-e at low pH, during high-hydrostatic-pressure treatment, and during freezing . The survival of Delta sigB mutant exponential-phase cells at pH 2.5 was 10,000-fold lower than the survival of EGD-e wild-type cells . Moreover, the Delta sigB mutant failed to show an acid tolerance response . Upon preexposure for 1 h to pH 4.5, the survival at pH 2.5 was 100,000-fold lower for the Delta sigB mutant than for the wild type . The glutamate decarboxylase (GAD) acid resistance system is important in survival and adaptation of L . monocytogenes in acidic conditions . The sigma(B) dependence of the gad genes (gadA, gadB, gadC, gadD, and gadE) was analyzed in silico . Putative sigma(B)-dependent promoter sites were found upstream of the gadCB operon (encoding a glutamate/gamma-aminobutyrate antiporter and a glutamate decarboxylase, respectively) and the lmo2434 gene (gadD, encoding a putative glutamate decarboxylase) . Reverse transcriptase PCR revealed that expression of the gadCB operon and expression of gadD are indeed sigma(B) dependent . In addition, a proteomics approach was used to analyze the protein expression profiles upon acid exposure . Although the GAD proteins were not recovered, nine proteins accumulated in the wild type but not in the Delta sigB strain . These proteins included Pfk, GalE, ClpP, and Lmo1580 . Exposure to pH 4.5, in order to preload cells with active sigma(B) and consequently with sigma (B)-dependent general stress proteins, also provided considerable protection against high-hydrostatic-pressure treatment and freezing . The combined data argue that the expression of sigma(B)-dependent genes provides L . monocytogenes with nonspecific multiple-stress resistance that may be relevant for survival in the natural environment as well as during food processing.

Appl Environ Microbiol, 2004 Jun, 70(6), 3292 - 7
Novel expression system for large-scale production and purification of recombinant class IIa bacteriocins and its application to piscicolin 126; Gibbs GM et al.; Piscicolin 126 is a class IIa bacteriocin isolated from Carnobacterium piscicola JG126 that exhibits strong activity against Listeria monocytogenes . The gene encoding mature piscicolin 126 (m-pisA) was cloned into an Escherichia coli expression system and expressed as a thioredoxin-piscicolin 126 fusion protein that was purified by affinity chromatography . Purified recombinant piscicolin 126 was obtained after CNBr cleavage of the fusion protein followed by reversed-phase chromatography . Recombinant piscicolin 126 contained a single disulfide bond and had a mass identical to that of native piscicolin 126 . This novel bacteriocin expression system generated approximately 26 mg of purified bacteriocin from 1 liter of E . coli culture . The purified recombinant piscicolin 126 acted by disruption of the bacterial cell membrane.

Vet Microbiol, 2004 Jun 21, 101(2), 83 - 9
PCR detection of a putative N-acetylmuramidase gene from Listeria ivanovii facilitates its rapid identification; Liu D et al.; Listeria ivanovii is a Gram-positive bacterial pathogen that is capable of causing abortions and stillbirths in farm animals, particularly sheep and cattle . In terms of morphological, biochemical and molecular characteristics, L . ivanovii resembles other Listeria species such as L . monocytogenes, a pathogen of both man and animals . In this study, through comparative analysis of genomic DNA from the six Listeria species, a L . ivanovii specific clone (liv22-228) containing a 946 bp insert was isolated . This clone contained the 5' ends of two divergently transcribed L . ivanovii genes and an intergenic spacer region, similar in organization to homologous regions from the L . innocua and L . monocytogenes genomes . Regions of low homology in the clone were identified by comparing to the L . innocua and L . monocytogenes genomes, and oligonucleotide primers (liv22-228F and liv22-228R) were designed . These primers amplified a 463 bp band from genomic DNA of L . ivanovii strains only, but not from other Listeria species or common bacteria . Thus, PCR employing L . ivanovii specific primers (liv22-228F and liv22-228R) provides a useful and straightforward method for rapid and precise determination of L . ivanovii .

Int J Food Microbiol, 2004 Jul 1, 94(1), 67 - 78
Effect of a(w), controlled by the addition of solutes or by water content, on the growth of Listeria innocua in broth and in a gelatine model; Lebert I et al.; The effect of a(w) on the growth of Listeria innocua was investigated in broth and on the surface of a gelatine food model . In broth, a(w) was controlled from 0.91 to 0.99 by the addition of solutes such as NaCl, KCl, glucose, sucrose and glycerol . In the gelatine food model, a(w) was controlled by removal of water . In the a(w) range, 0.92-0.99, the generation times observed in broth in the presence of NaCl, KCl, sucrose and glucose were similar but were longer than those in glycerol . For lag times, the inhibition of L . innocua growth followed the order: NaCl = KCl = sucrose>glucose>glycerol . When comparing growth at a(w) 0.95 for the three media--broth + NaCl, gelatine gel (a(w) controlled by removal of water) and gelatine gel with NaCl (gel + NaCl, a(w) controlled by NaCl)--the shortest generation time was observed in broth + NaCl, followed by gel + NaCl and, finally, on gel with a larger gap between the last two . The generation time on gel was five times greater than the generation time in broth + NaCl and 2.5 times greater on gel + NaCl . It was concluded that not only the structure of the media (solid or liquid) had an effect on Listeria inhibition but also and mainly the way the a(w) was adjusted . Removal of water was more stressful to Listeria than the addition of NaCl.

Int J Food Microbiol, 2004 Jul 1, 94(1), 15 - 22
Comparison of selective and nonselective primary enrichments for the detection of Listeria monocytogenes in cheese; Rijpens N et al.; A completely selective enrichment procedure was compared with two partially nonselective ones for the detection of Listeria monocytogenes in cheeses . After enrichment for approximately 48 h, the enrichment media were streaked on selective agars and presumptive Listeria colonies were confirmed using PCR . In some cases, PCR was also performed directly on the enrichment broth . The conventional, completely selective enrichment procedure was not always the best choice for the detection of stressed L . monocytogenes in cheeses . Especially in the case of semi-hard cheeses from pasteurized milk and soft cheeses of the blue veined and the red smear types, the methods that incorporated a nonselective enrichment step gave better results than the completely selective method . For mold ripened, soft cheeses, the results were highly dependent on the brand of cheese and time of sampling, but the best results were obtained using the completely selective enrichment procedure.

J Clin Pathol, 2004 Jun, 57(6), 665 - 6
Treatment difficulties of a listerial rhombencephalitis in an adult patient allergic to penicillins; Popescu GA et al.; Rhombencephalitis is not a rare presentation of listerial central nervous system infections in healthy adults . This report describes a case with several management difficulties linked to antibiotic related adverse events, pointing to alternative solutions to aminopenicillins . In addition, the role of dexamethasone in the management of inflammation and neurological symptoms is discussed.

J AOAC Int, 2004 Mar-Apr, 87(2), 395 - 410
Evaluation of the BAX system for detection of Listeria monocytogenes in foods: collaborative study; Silbernagel K et al.; A multilaboratory study was conducted to compare the automated BAX system and the standard cultural methods for detection of Listeria monocytogenes in foods . Six food types (frankfurters, soft cheese, smoked salmon, raw, ground beef, fresh radishes, and frozen peas) were analyzed by each method . For each food type, 3 inoculation levels were tested: high (average of 2 CFU/g), low (average of 0.2 CFU/g) and uninoculated controls . A total of 25 laboratories representing government and industry participated . Of the 2335 samples analyzed, 1109 were positive by the BAX system and 1115 were positive by the standard method . A Chi square analysis of each of the 6 food types, at the 3 inoculation levels tested, was performed . For all foods, except radishes, the BAX system performed as well as or better than the standard reference methods based on the Chi square results.

Int J Clin Pract, 2004 Apr, 58(4), 420 - 1
Delayed presentation of prosthetic joint infection due to Listeria monocytogenes; Chougle A et al.; Infection with Listeria monocytogenes is rare and has been described in prosthetic valves, stent grafts and prosthetic joints . The route of infection appears to be haematogenous . The choice between conservative treatment with antibiotics or surgical treatment with debridement and revision of the components remains controversial . The best antibiotic treatment is not known with ampicillin being the first choice in most cases . Prosthetic infections with Listeria monocytogenes usually occur in patients with malignancy, diabetes mellitus, chronic renal disease, liver disease, elderly patients and patients receiving immunosuppressive therapy . The hip is the commonest prosthetic joint affected followed by the knee . We report the seventh case of Listeria monocytogenes infection in a non-immunocompromised patient involving a prosthetic joint.

Infect Immun, 2004 Jun, 72(6), 3543 - 8
CpG oligodeoxynucleotides improve the survival of pregnant and fetal mice following Listeria monocytogenes infection; Ito S et al.; Listeria infection during pregnancy can cause the death of both mother and fetus . Previous studies established that immunostimulatory CpG oligodeoxynucleotides (ODN) increase the resistance of healthy adult mice to many infectious pathogens, including Listeria monocytogenes . This study examines whether the innate immune response elicited by CpG ODN can reduce the susceptibility of pregnant mice to lethal listeria challenge . The results indicate that CpG ODN treatment significantly improves maternal survival and reduces pathogen transmission to offspring . CpG ODN administered during pregnancy did not induce abortion, birth defects, or reduce the size or health of litters . These findings suggest that CpG ODN may provide a safe and effective means of improving the health of mothers and fetuses during pregnancy.

Infect Immun, 2004 Jun, 72(6), 3237 - 44
Role of flagellin and the two-component CheA/CheY system of Listeria monocytogenes in host cell invasion and virulence; Dons L et al.; The flagellum protein flagellin of Listeria monocytogenes is encoded by the flaA gene . Immediately downstream of flaA, two genes, cheY and cheA, encoding products with homology to chemotaxis proteins of other bacteria, are located . In this study we constructed deletion mutants with mutations in flaA . cheY, and cheA to elucidate their role in the biology of infection with L . monocytogenes . The DeltacheY, DeltacheA, and double-mutant DeltacheYA mutants, but not DeltaflaA mutant, were motile in liquid media . However, the DeltacheA mutant had impaired swarming and the DeltacheY and DeltacheYA mutants were unable to swarm on soft agar plates, suggesting that cheY and cheA genes encode proteins involved in chemotaxis . The DeltaflaA, DeltacheY, DeltacheA, and DeltacheYA mutants (grown at 24 degrees C) showed reduced association with and invasion of Caco-2 cells compared to the wild-type strain . However, spleens from intragastrically infected BALB/c and C57BL/6 mice showed larger and similar numbers of the DeltaflaA and DeltacheYA mutants, respectively, compared to the wild-type controls . Such a discrepancy could be explained by the fact that tumor necrosis factor receptor p55 deficient mice showed dramatically exacerbated susceptibility to the wild-type but unchanged or only slightly increased levels of the DeltaflaA or DeltacheYA mutant . In summary, we show that listerial flaA . cheY, and cheA gene products facilitate the initial contact with epithelial cells and contribute to effective invasion but that flaA could also be involved in the triggering of immune responses.

J Food Prot, 2004 May, 67(5), 1022 - 6
Prevalence and growth of Listeria on naturally contaminated smoked salmon over 28 days of storage at 4 degrees C; Lappi VR et al.; Only limited data are available on the growth characteristics of Listeria in naturally contaminated ready-to-eat foods . To evaluate Listeria contamination patterns and growth in smoked salmon, 72 smoked salmon product samples from two processing plants were tested for Listeria spp . and L . monocytogenes . Samples were divided into four approximately equal portions: one portion was tested on receipt, and the other three were vacuum sealed and stored at 4 degrees C for 7, 14, and 28 days . Listeria testing was performed using both an enrichment procedure and direct plating to enumerate Listeria in samples that contained >2 to 10 CFU/g . Five samples were positive for Listeria spp., including one sample that was positive for L . monocytogenes . Most samples yielded only sporadic positive results among the portions tested on days 0, 7, 14, and 28 . Only one sample contained Listeria spp . in numbers above the detection limit for enumeration . For this sample, the portions tested on days 7 and 28 contained 46 and 52 CFU/g, respectively, whereas the portion tested on day 14 was negative . Overall, our data indicate that there is considerable heterogeneity in Listeria spp . distribution within a single positive smoked fish sample . Even with refrigerated storage for 28 days, none of the naturally contaminated samples reached Listeria spp . numbers >100 CFU/g, which indicates that Listeria growth was limited within a 4-week storage period . However, because of the apparent heterogeneity of Listeria distribution within samples, the interpretation of growth data collected on naturally contaminated samples is difficult.

J Food Prot, 2004 May, 67(5), 1017 - 21
Evaluation of nisin-coated cellulose casings for the control of Listeria monocytogenes inoculated onto the surface of commercially prepared frankfurters; Luchansky JB et al.; Commercially prepared frankfurters were formulated with and without approximately 1.4% potassium lactate and 0.1% sodium diacetate and were subsequently processed in cellulose casings coated with and without nisin (approximately 50,000 IU per square inch of internal surface area) to control the outgrowth of Listeria monocytogenes during refrigerated storage . The frankfurters were inoculated with approximately 5 log CFU per package of a five-strain mixture of L . monocytogenes and then vacuum sealed before being stored at 4 degrees C for 60 to 90 days . Surviving organisms were recovered and enumerated by rinsing each package with 18 ml of sterile 0.1% peptone water and plating onto MOX selective agar . The data for each of two trials were averaged . In packages that contained frankfurters formulated with potassium lactate and sodium diacetate and prepared in nisin-coated casings, L . monocytogenes levels decreased by 1.15 log CFU per package after 90 days of storage . L . monocytogenes levels decreased by 0.95 log CFU per package in frankfurters that were prepared in casings that were not coated with nisin . In packages of frankfurters that were formulated without potassium lactate and sodium diacetate and prepared in nisin-coated casings, L . monocytogenes levels decreased by 0.88 log CFU per package after 15 days of storage but then increased appreciably thereafter over a 60-day period of refrigerated storage . There was also an appreciable increase in pathogen numbers during 60 days of storage in otherwise similar frankfurters formulated without potassium lactate and sodium diacetate prepared in casings that were not coated with nisin . These data confirm that potassium lactate and sodium diacetate display listeriostatic activity as an ingredient of commercial frankfurters . These data also establish that cellulose casings coated with nisin display only moderate antilisterial activity in vacuum-sealed packages of commercially prepared frankfurters during storage at 4 degrees C.

J Food Prot, 2004 May, 67(5), 922 - 7
Encapsulation of nisin and lysozyme in liposomes enhances efficacy against Listeria monocytogenes; Were LM et al.; The efficacy and stability against Listeria monocytogenes of nisin and lysozyme encapsulated in phospholipid liposomes was evaluated . Antimicrobial-containing liposomes were prepared by hydrating dried lipids with buffer containing nisin, nisin plus the fluorescence probe calcein, or calcein and lysozyme . Mixtures were then centrifuged and sonicated, and encapsulated liposomes were collected using size-exclusion chromatography . Antimicrobial concentration in liposomes was determined by bicinchoninic acid assay prior to determination of antimicrobial activity against strains of L . monocytogenes . When nisin was encapsulated in liposomes, protein concentrations of 0.39, 0.27, and 0.23 mg/ml for phosphatidylcholine (PC), PC-cholesterol (7:3), and PC-phosphatidylglycerol (PG)-cholesterol (5:2:3), respectively, were obtained . Encapsulation of nisin with calcein yielded protein concentrations of 0.35, 0.39, and 0.28 mg/ml for PC, PC-cholesterol, and PC-PG-cholesterol, respectively . Encapsulation of calcein with lysozyme resulted in protein concentrations of 0.43, 0.26, and 0.19 mg/ml for PC, PC-cholesterol, and PC-PG-cholesterol, respectively . Encapsulated nisin in 100% PC and PC-cholesterol liposomes inhibited bacterial growth by >2 log CFU/ml compared with free nisin . Growth inhibition with liposomal lysozyme was strain dependent, with greater inhibition observed for strains 310 and Scott A with PC-cholesterol and PC-PG-cholesterol liposomes . Inhibition of L . monocytogenes indicated the potential of liposomes to serve as delivery vehicles for antimicrobials in foods while improving stability of antimicrobials.

J Food Prot, 2004 May, 67(5), 915 - 21
Effectiveness of acidic calcium sulfate with propionic and lactic acid and lactates as postprocessing dipping solutions to control Listeria monocytogenes on frankfurters with or without potassium lactate and stored vacuum packaged at 4.5 degrees C; Nunez de Gonzalez MT et al.; The safety of ready-to-eat meat products such as frankfurters can be enhanced by treating with approved antimicrobial substances to control the growth of Listeria monocytogenes . We evaluated the effectiveness of acidic calcium sulfate with propionic and lactic acid, potassium lactate, or lactic acid postprocessing dipping solutions to control L . monocytogenes inoculated (ca . 10(8) CFU/ml) onto the surface of frankfurters with or without potassium lactate and stored in vacuum packages at 4.5 degrees C for up to 12 weeks . Two frankfurter formulations were manufactured without (control) or with potassium lactate (KL, 3.3% of a 60% {wt/wt} commercially available syrup) . After cooking, chilling, and peeling, each batch was divided into inoculated (four strains of L . monocytogenes mixture) and noninoculated groups . Each group was treated with four different dips: (i) control (saline solution), (ii) acidic calcium sulfate with propionic and lactic acid (ACS, 1:2 water), (iii) KL, or (iv) lactic acid (LA, 3.4% of a 88% {wt/wt} commercially available syrup) for 30 s . Noninoculated frankfurters were periodically analyzed for pH, water activity, residual nitrite, and aerobic plate counts (APCs), and L . monocytogenes counts (modified Oxford medium) were determined on inoculated samples . Surface APC counts remained at or near the lower limit of detection (<2 log CFU per frank) on franks with or without KL and treated with ACS or LA throughout 12 weeks at 4.5 degrees C . L . monoctogenes counts remained at the minimum level of detection on all franks treated with the ACS dip, which indicated a residual bactericidal effect when L . monocytogenes populations were monitored over 12 weeks . L . monocytogenes numbers were also reduced, but not to the same degree in franks made without or with KL and treated with LA . These results revealed the effectiveness of ACS (bactericidal effect) or LA (bacteriostatic effect) as postprocessing dipping solutions to inhibit or control the growth of L . monocytogenes on vacuum-packaged frankfurters stored at 4.5 degrees C for up to 12 weeks.

J Bacteriol, 2004 Jun, 186(11), 3355 - 62
The RNA-binding protein Hfq of Listeria monocytogenes: role in stress tolerance and virulence; Christiansen JK et al.; In gram-negative bacteria, the RNA-binding protein Hfq has emerged as an important regulatory factor in a variety of physiological processes, including stress resistance and virulence . In Escherichia coli, Hfq modulates the stability or the translation of mRNAs and interacts with numerous small regulatory RNAs . Here, we studied the role of Hfq in the stress tolerance and virulence of the gram-positive food-borne human pathogen Listeria monocytogenes . We present evidence that Hfq is involved in the ability of L . monocytogenes to tolerate osmotic and ethanol stress and contributes to long-term survival under amino acid-limiting conditions . However, Hfq is not required for resistance to acid and oxidative stress . Transcription of hfq is induced under various stress conditions, including osmotic and ethanol stress and at the entry into the stationary growth phase, thus supporting the view that Hfq is important for the growth and survival of L . monocytogenes in harsh environments . The stress-inducible transcription of hfq depends on the alternative sigma factor sigmaB, which controls the expression of numerous stress- and virulence-associated genes in L . monocytogenes . Infection studies showed that Hfq contributes to pathogenesis in mice, yet plays no role in the infection of cultured cell lines . This study provides, for the first time, information on the role of Hfq in the stress tolerance and virulence of a gram-positive pathogen.

J Evol Biol, 2004 May, 17(3), 506 - 18
Plant-like mating in an animal: sexual compatibility and allocation trade-offs in a simultaneous hermaphrodite with remote transfer of sperm; Pemberton AJ et al.; The importance of sexual compatibility between mates has only recently been realized in zoological research into sexual selection, yet its study has been central to botanical research for many decades . The reproductive characteristics of remote mating, an absence of precopulatory mate screening, internal fertilization and embryonic brooding are shared between passively pollinated plants and a phylogenetically diverse group of sessile aquatic invertebrates . Here, we further characterize the sexual compatibility system of one such invertebrate, the colonial ascidian Diplosoma listerianum . All 66 reciprocal pairings of 12 genetic individuals were carried out . Fecundities of crosses varied widely and suggested a continuous scale of sexual compatibility . Of the 11 animals from the same population c . 40% of crosses were completely incompatible with a further c . 20% having obvious partial compatibility (reduced fecundity) . We are unaware of other studies documenting such high levels of sexual incompatibility in unrelated individuals . RAPD fingerprinting was used to estimate relatedness among the 12 individuals after a known pedigree was successfully reconstructed to validate the technique . In contrast to previous results, no correlation between genetic similarity and sexual compatibility was detected . The blocking of many genotypes of sperm is expected to severely modify realized paternity away from 'fair raffle' expectations and probably reduce levels of intra-brood genetic diversity in this obligatorily promiscuous mating system . One adaptive benefit may be to reduce the bombardment of the female reproductive system by outcrossed sperm with conflicting evolutionary interests, so as to maintain female control of somatic : gametic investment.

J Infect Dis, 2004 Jun 1, 189(11), 2101 - 9 Epub 2004 May 12.
Impairment of growth of Listeria monocytogenes in THP-1 macrophages by granulocyte macrophage colony-stimulating factor: release of tumor necrosis factor-alpha and nitric oxide; Carryn S et al.; BACKGROUND: Listeria monocytogenes tends to survive in phagocytes . Granulocyte macrophage colony-stimulating factor (GM-CSF) protects mice against L . monocytogenes infection, and mice knocked out for the GM-CSF gene are more susceptible to these infections . METHODS: THP-1 cells were used to characterize the GM-CSF receptor (binding isotherms; STAT5 phosphorylation), measure the intracellular growth of L . monocytogenes (5 h after phagocytosis), examine the influence of a 24-h incubation with GM-CSF before infection, measure the production of tumor necrosis factor (TNF)-alpha and the expression of nitric oxide synthase (iNOS), and evaluate the influence of anti-GM-CSF receptor (GM-CSFR alpha ) and anti-TNF-alpha antibodies and the addition of N(omega)-nitro-L-arginine methyl ester (L-NAME) and catalase . RESULTS: THP-1 cells display functional GM-CSFR alpha . GM-CSF impairs the intracellular growth of L . monocytogenes to approximately 65% of its value in unstimulated cells . This effect is abolished by anti-GM-CSFR alpha, anti-TNF-alpha antibodies, and catalase (and, to a lesser extent, by L-NAME) . GM-CSF stimulates the release of TNF-alpha and the expression of iNOS . TNF-alpha added to unstimulated cells (even in large amounts) does not fully reproduce the impairment in the growth of L . monocytogenes caused by GM-CSF . CONCLUSIONS: GM-CSF impairs the intracellular growth of L . monocytogenes by a synergistic action of the GM-CSF-triggered release of autocrine TNF-alpha and hydrogen peroxide and the production of NO (associated with the stimulation of the expression of iNOS).

J Infect Dis, 2004 Jun 1, 189(11), 2094 - 100 Epub 2004 May 14.
A molecular marker for evaluating the pathogenic potential of foodborne Listeria monocytogenes; Jacquet C et al.; BACKGROUND: Internalin mediates entry of Listeria monocytogenes into some human cultured cell lines and crossing of the intestinal barrier in transgenic mice expressing its receptor, human E-cadherin, in enterocytes . The relevance of these findings for humans is challenged by the observation that some L . monocytogenes isolates express a truncated nonfunctional form of internalin . METHODS: We investigated expression of internalin by use of immunoblot assay in 300 clinical strains obtained in France in a single year and a representative set of 150 strains obtained from food products during the same period . RESULTS: Clinical strains expressed full-length internalin far more frequently (288/300 strains {96%}) than did strains recovered from food products (98/150 strains {65%}; odds ratio, 12.73; 95% confidence interval, 6.27-26.34; P<1 x 10(-7)) . All 61 strains (100%) from pregnancy-related cases, 55 (98%) of 56 strains from patients with central nervous system infections, and 151 (93%) of 162 strains from patients with bacteremia expressed full-length internalin . All 110 strains belonging to serovar 4b, the most frequently implicated serovar in human listeriosis, expressed full-length internalin . CONCLUSIONS: This study demonstrates the critical role of internalin in the pathogenesis of human listeriosis . It provides a molecular explanation for the predominance of serovar 4b among clinical strains and supports the usefulness of studying the expression of internalin as a marker of virulence in humans.

Biosens Bioelectron, 2004 Jul 15, 19(12), 1759 - 61
Studies of Listeria monocytogenes-antibody binding using electro-orientation; Bunin VD et al.; An electro-optical (EO) approach has been used for studies of Listeria monocytogenes-antibody binding . The EO analyzer, which has been developed at the State Research Center for Applied Microbiology, Obolensk, was used as a basic instrument for EO measurements . AC electro-kinetic effects depend on dielectric properties of bioparticles, their composition, morphology, the medium, and the frequency of applied electrical field . Electro-orientational spectra were used for discrimination of bacteria before and after selective binding with antibodies . The measurements were performed using a discrete set of frequencies of the orienting electric field (10, 100, 250, and 500 kHz) . During biospecific interactions an antibody is bound to the microorganism causing a change in the dielectric properties of the microorganism-antibody complex and the EO signal reaches its maximum at 100-200 kHz . It has been shown that the biospecific interactions of L . monocytogenes cells with anti-Listeria antibody in the presence of Escherichia coli K-12, and Azospirillum brasilense Sp7 change the EO signals significantly . Thus, the determination of the presence of particular bacteria within a mixed sample may be achieved by selection and matching of antibodies specific to individual bacterium types and by comparing spectra of bacterium in the presence and in the absence of specific antibody.

Eur J Clin Microbiol Infect Dis, 2004 Jun, 23(6), 484 - 6 Epub 2004 May 13.
Meropenem therapy failure in Listeria monocytogenes infection; Stepanovic S et al.; Listeria monocytogenes is highly susceptible to meropenem in vitro, but data on the efficacy of meropenem in clinical cases of listeriosis are scarce . Described here is the case of a child with aplastic anemia who acquired nosocomial listeriosis and failed to respond to initial meropenem therapy . Resolution of fever was not noted after 5 days of therapy with meropenem and, more importantly, clinical worsening was observed during this period . The patient began to improve after ampicillin was introduced to the therapeutic regimen . In total, meropenem was administered for 15 days and ampicillin for 10 days.

Clin Diagn Lab Immunol, 2004 May, 11(3), 446 - 51
Use of monoclonal antibodies that recognize p60 for identification of Listeria monocytogenes; Yu KY et al.; Listeria monocytogenes causes major food-borne outbreaks of disease worldwide . Specific identification of this microorganism is of utmost importance to public health and industry . Listeria species are known to secrete a 60-kDa protein collectively termed p60, which is encoded by the iap (invasion-associated protein) gene and secreted in large quantities into the growth media . p60 is a highly immunogenic murein hydrolase that is essential for cell division . Due to these properties, p60 is an ideal diagnostic target for the development of immunological detection systems for L . monocytogenes . We report here two independent lines of monoclonal antibody (MAb): p6007, which specifically recognizes L . monocytogenes p60, and p6017, which reacts with a wide range of Listeria p60 proteins . By combining these antibodies with a polyclonal antibody, we developed efficient sandwich enzyme-linked immunosorbent assay (ELISA) systems which can specifically identify L . monocytogenes or generally detect Listeria species . Since an excess amount of the peptide corresponding to PepA or PepD did not interfere with the ELISA, and direct ELISAs were unable to detect both peptides, we concluded that the epitope presumed to be recognized by p6007 or p6017 could be distinguished from PepA and PepD as described by Bubert et al . (Appl . Environ . Microbiol . 60:3120-3127, 1997) . To our best knowledge, this is the first example of an immunological identification system that uses p60-recognizing MAbs.

Br J Nutr, 2004 May, 91(5), 733 - 9
Fatty acid-mediated inhibition of IL-12 production by murine macrophages is independent of PPARgamma; Zhang M et al.; Our laboratory has reported that n-3 PUFA can reduce host resistance to Listeria infection, in part, by impairing in vivo IL-12 biosynthesis . Recently, PUFA were shown to be ligands for PPAR, a novel family of nuclear receptors with three isoforms: PPARalpha, PPARdelta/beta and PPARgamma . PPARgamma is expressed in immune cells, such as T cells and macrophages . Two PPARgamma agonists, 15-deoxy-delta(12,14)-prostaglandin (PG) J2 and rosiglitazone, have been shown to have immunomodulatory activity in vitro, including inhibiting IL-12 biosynthesis . We hypothesized that n-3 PUFA inhibit IL-12 production through activating PPARgamma . We used thioglycolate-elicited mouse peritoneal macrophages to study the effect of various fatty acids and their oxidized metabolites on in vitro IL-12 production . Our present results demonstrate that both n-3 and n-6 PUFA can reduce in vitro IL-12 biosynthesis, though less potently than 15-deoxy-PGJ2 and rosiglitazone . GW9662, a PPARgamma antagonist, reversed the inhibitory effect of rosiglitazone, but not that of PUFA . Our present findings suggest that fatty acid-mediated inhibition of IL-12 production is independent of PPARgamma.

Int J Food Microbiol, 2004 Jun 1, 93(2), 131 - 40
Overview of Listeria monocytogenes contamination in Japan; Okutani A et al.; Listeriosis is a relatively rare foodborne illness but can be life threatening with high fatality rates . In Japan, the incidence of listeriosis has been very low for the past 40 years compared with that of Western Europe and North America . We hypothesized that less Listeria monocytogenes contamination in Japanese foods would be related to the lower incidence in Japan . For this purpose, we collected data of Japanese foods contaminated with L . monocytogenes, mainly from Japanese-written reports, and reviewed them . From this review, we found that the proportion of L . monocytogenes, Listeria spp . isolation from foods in Japan is similar to those reported from other countries and that other factors might be responsible for the lower occurrence of listeriosis .

Int J Food Microbiol, 2004 May 15, 93(1), 87 - 99
Screening of glutamate decarboxylase activity and bile salt resistance of human asymptomatic carriage, clinical, food, and environmental isolates of Listeria monocytogenes; Olier M et al.; Following consumption, stomach acidity is the first major barrier encountered by the food-borne pathogen Listeria monocytogenes . Analysis of low pH sensitivity and glutamate decarboxylase (GAD) acid resistance system of 14 isolates of L . monocytogenes carried asymptomatically by humans showed that levels of GAD activity were subjected to strain variation . Similar variations were observed for strains responsible for 18 cases of listeriosis, whereas in comparison, 13 strains isolated from food and food-processing plant environments showed lower GAD activity . Following survival of the stomach barrier, L . monocytogenes also has to resist bile salts encountered in the small intestines . Analysis revealed that all strains tested were able to grow in the presence of bile salts with concentrations as high as those encountered in the small intestines and had previously identified bile salt hydrolase (BSH) activity . Strain variation was observed but there was no relationship between the origin of the strains and the ability to degrade bile salts .

Cell Immunol, 2004 Feb, 227(2), 109 - 20
The role of CD1d in the immune response against Listeria infection; Arrunategui-Correa V et al.; To address the role of CD1d in mucosal immune regulation in bacterial infection, we infected CD1d KO mice with Listeria monocytogenes (Lm) . A higher systemic bacterial burden associated with inflammatory lymphocytic infiltrations within the intestine was found in CD1d KO compared with wild type (WT) mice . Lm induced strong IFN-gamma mRNA expression in the liver of WT and the intestine of CD1d KO mice, thus demonstrating the dual, opposing immune activities of IFN-gamma in Lm infection that is dependent on CD1d and/or NKT cells . Analysis of hepatic T cell population demonstrated a reduction of NK1.1(+)TCRbeta+ cells in both mice, followed by recovery only in WT mice . Last, the proportion of alpha4beta1 integrin on lung lymphocytes from CD1d KO was dramatically increased compared with WT mice . Thus, the absence of CD1d resulted in increased susceptibility towards Listeria infection, induced changes in NKT cells, and increased trafficking of alpha4beta1 molecule to inflamed lung.

Microbiology, 2004 May, 150(Pt 5), 1581 - 90
Global analysis of gene expression in an rpoN mutant of Listeria monocytogenes; Arous S et al.; The role of the alternative sigma(54) factor, encoded by the rpoN gene, was investigated in Listeria monocytogenes by comparing the global gene expression of the wild-type EGDe strain and an rpoN mutant . Gene expression, using whole-genome macroarrays, and protein content, using two-dimensional gel electrophoresis, were analysed . Seventy-seven genes and nine proteins, whose expression was modulated in the rpoN mutant as compared to the wild-type strain, were identified . Most of the modifications were related to carbohydrate metabolism and in particular to pyruvate metabolism . However, under the conditions studied, only the mptACD operon was shown to be directly controlled by sigma(54) . Therefore, the remaining modifications seem to be due to indirect effects . In parallel, an in silico analysis suggests that sigma(54) may directly control the expression of four different phosphotransferase system (PTS) operons, including mptACD . PTS activity is known to have a direct effect on the pyruvate pool and on catabolite regulation . These results suggest that sigma(54) is mainly involved in the control of carbohydrate metabolism in L . monocytogenes via direct regulation of PTS activity, alteration of the pyruvate pool and modulation of carbon catabolite regulation.

J Immunol, 2004 May 15, 172(10), 6030 - 8
The ability of two Listeria monocytogenes vaccines targeting human papillomavirus-16 E7 to induce an antitumor response correlates with myeloid dendritic cell function; Peng X et al.; Previous work from our laboratory has shown that Lm-LLO-E7 induces complete regression of approximately 75% of established TC-1 tumors, whereas Lm-E7 only slows the growth of such tumors . In this study, we examine the effects of Lm-LLO-E7 vs Lm-E7 on APCs . We hypothesize that the difference in antitumor efficacy of the two vaccines is due to the ability of each of these vectors to render immature dendritic cells (DCs) effective APCs in terms of MHC class II or costimulatory molecule expression . We also examine the ability of these vectors to stimulate cytokine production by DCs . Both vectors induced IL-12 and TNF-alpha, but only Lm-LLO-E7 induced IL-2 production by DCs . Lm-LLO-E7 also induced significantly higher levels of MHC class II molecules, CD40, and B7 costimulatory molecules (CD86, B7-H1, and B7-DC) on DCs than Lm-E7 . Interestingly, a shift of CD11c(+) cells from CD86(low) to CD86(high) is observed post-Lm-LLO-E7 infection . A similar shift is also observed for B7-H1 and B7-DC molecules . Moreover, Lm-LLO-E7, but not Lm-E7-pulsed DCs, stimulate naive T cell proliferation . These results indicate that Lm-LLO-E7 is more effective than Lm-E7 at inducing DC maturation . This effect is independent of the E7 Ag, because Lm-LLO-NP, and a mixture of Lm-LLO-NP and Lm-E7 induce the same changes in DC phenotype as Lm-LLO-E7 . Taken together, the changes in DC expression correlate well with the differences in antitumor efficacy between these two vaccines.

Appl Environ Microbiol, 2004 May, 70(5), 3176 - 9
Osmotic stress leads to decreased intracellular pH of Listeria monocytogenes as determined by fluorescence ratio-imaging microscopy; Fang W et al.; Intracellular pH (pH(i)) of Listeria monocytogenes was determined after exposure to NaCl or sorbitol in liquid and solid media (agar) . Both compounds decreased pH(i), and recovery on solid medium was impaired compared to that in liquid medium . N,N'-dicyclohexylcarbodiimide abolished pH(i) recovery, and lowering a(w) with glycerol showed no effect on pH(i).

Appl Environ Microbiol, 2004 May, 70(5), 2912 - 8
Molecular and physiological analysis of the role of osmolyte transporters BetL, Gbu, and OpuC in growth of Listeria monocytogenes at low temperatures; Wemekamp-Kamphuis HH et al.; Listeria monocytogenes is a ubiquitous food-borne pathogen found widely distributed in nature as well as an undesirable contaminant in a variety of fresh and processed foods . This ubiquity can be at least partly explained by the ability of the organism to grow at high osmolarity and reduced temperatures, a consequence of its ability to accumulate osmo- and cryoprotective compounds termed osmolytes . Single and multiple deletions of the known osmolyte transporters BetL, Gbu, and OpuC significantly reduce growth at low temperatures . During growth in brain heart infusion broth at 7 degrees C, Gbu and OpuC had a more pronounced role in cryoprotection than did BetL . However, upon the addition of betaine to defined medium, the hierarchy of transporter importance shifted to Gbu > BetL > OpuC . Upon the addition of carnitine, only OpuC appeared to play a role in cryoprotection . Measurements of the accumulated osmolytes showed that betaine is preferred over carnitine, while in the absence of a functional Gbu, carnitine was accumulated to higher levels than betaine was at 7 degrees C . Transcriptional analysis of the genes encoding BetL, Gbu, and OpuC revealed that each transporter is induced to different degrees upon cold shock of L . monocytogenes LO28 . Additionally, despite being transcriptionally up-regulated upon cold shock, a putative fourth osmolyte transporter, OpuB (identified by bioinformatic analysis and encoded by lmo1421 and lmo1422), showed no significant contribution to listerial chill tolerance . Growth of the quadruple mutant LO28deltaBCGB (deltabetL deltaopuC deltagbu deltaopuB) was comparable to the that of the triple mutant LO28deltaBCGsoe (deltabetL deltaopuC deltagbu) at low temperatures . Here, we conclude that betaine and carnitine transport upon low-temperature exposure is mediated via three osmolyte transporters, BetL, Gbu, and OpuC.

Appl Environ Microbiol, 2004 May, 70(5), 2717 - 21
Increased ATPase activity is responsible for acid sensitivity of nisin-resistant Listeria monocytogenes ATCC 700302; McEntire JC et al.; The growth of the foodborne pathogen Listeria monocytogenes can be controlled by nisin, an antimicrobial peptide . A spontaneous mutant of L . monocytogenes shows both resistance to nisin and increased acid sensitivity compared to the wild type . Changes in the cell membrane correlated with nisin resistance, but the mechanism for acid sensitivity appears unrelated . When hydrochloric or lactic acid is added to cultures, intracellular ATP levels drop significantly in the mutant (P < 0.01) compared to the results seen with the wild type . Characterization of the F(0)F(1) ATPase, which hydrolyzes ATP to pump protons from the cell cytoplasm, shows that the enzyme is more active in the mutant than in the wild type . These data support a model in which the increased activity of the mutant ATPase upon acid addition depletes the cells' supply of ATP, resulting in cell death.

Clin Infect Dis, 2004 May 1, 38(9), 1261 - 5 Epub 2004 Apr 15.
Granulomatous infectious diseases associated with tumor necrosis factor antagonists; Wallis RS et al.; The relationship between the use of tumor necrosis factor antagonists and onset of granulomatous infection was examined using data collected through the Adverse Event Reporting System of the US Food and Drug Administration for January 1998-September 2002 . Granulomatous infections were reported at rates of approximately 239 per 100,000 patients who received infliximab and approximately 74 per 100,000 patients who received etanercept (P<.001) . Tuberculosis was the most frequently reported disease, occurring in approximately 144 and approximately 35 per 100,000 infliximab-treated and etanercept-treated patients, respectively (P<.001) . Candidiasis, coccidioidomycosis, histoplasmosis, listeriosis, nocardiosis, and infections due to nontuberculous mycobacteria were reported with significantly greater frequency among infliximab-treated patients . Seventy-two percent of these infection occurred < or =90 days after starting infliximab treatment, and 28% occurred after starting etanercept treatment (P<.001) . These data indicate a risk of granulomatous infection that was 3.25-fold greater among patients who received infliximab than among those who received etanercept . The clustering of reports shortly after initiation of treatment with infliximab is consistent with reactivation of latent infection.

J Egypt Soc Parasitol, 2004 Apr, 34(1), 349 - 66
Coinfection with Listeria monocytogenes potentlxtes the response of BALB/c mice against Leishmania major; Tabbara KS et al.; Protection against L . major is dependent on the stimulation of an anti-leishmanial T helper1 (Th1) response and the production of Interferon (IFN)-y . BALB/c mice develop a Th2 response and fatal infection with Leishmania major . Strategies that boost IL-12 production have shown to be protective . The innate response to Listeria monocytogenes is associated with IL-12 production . The co-infection of BALB/c mice with L . monocytogenes attenuates the course of L . major infection . Lesion sizes were smaller, and co-infected mice out-survived controls injected with L . major alone . The parasite load was reduced at site of injection, in draining lymph nodes (LN) and spleen . During the first week of infection, in-vitro Leishmania-restimulated LN cells from co-infected mice produced higher levels of IFN-7 and undetectable levels of IL-4 compared to controls . Significant IL-4 mRNA expression was detected in LN cells of control but not in co-infected mice.

J Cell Biol, 2004 Apr 26, 165(2), 233 - 42
Fascin-mediated propulsion of Listeria monocytogenes independent of frequent nucleation by the Arp2/3 complex; Brieher WM et al.; Actin-dependent propulsion of Listeria monocytogenes is thought to require frequent nucleation of actin polymerization by the Arp2/3 complex . We demonstrate that L . monocytogenes motility can be separated into an Arp2/3-dependent nucleation phase and an Arp2/3-independent elongation phase . Elongation-based propulsion requires a unique set of biochemical factors in addition to those required for Arp2/3-dependent motility . We isolated fascin from brain extracts as the only soluble factor required in addition to actin during the elongation phase for this type of movement . The nucleation reaction assembles a comet tail of branched actin filaments directly behind the bacterium . The elongation-based reaction generates a hollow cylinder of parallel bundles that attach along the sides of the bacterium . Bacteria move faster in the elongation reaction than in the presence of Arp2/3, and the rate is limited by the concentration of G-actin . The biochemical and structural differences between the two motility reactions imply that each operates through distinct biochemical and biophysical mechanisms.

Int J Clin Pract, 2004 Mar, 58(3), 292 - 6
Fatal meningitis due to Listeria monocytogenes in elderly patients with underlying malignancy; Levidiotou S et al.; Adult patients with malignancies are considered to be at a high risk for Listeria monocytogenes meningitis . The Microbiology Laboratory's database of the University Hospital of Ioannina, Greece, was searched for cases of L . monocytogenes during the period from January 1990 to December 2002 . Listerial meningitis occurred in three patients: one with brain tumour, one with chronic lymphocytic leukaemia, and one with non-Hodgkin's lymphoma . All the patients were older than 70 and they were actively receiving therapy for their malignancy . L . monocytogenes type 4b was isolated from blood and cerebrospinal fluid . All were treated with ampicillin and gentamicin, but they died shortly after the initiation of the treatment . Experience with the three present cases indicated the high mortality rate due to listerial meningitis in this immunosuppressed population . So, listeriosis should be suspected in patients with meningitis and underlying malignancy . Since meningitis due to L . monocytogenes is not distinguishable clinically from other types of bacterial meningitis, it is recommended to cover Listeria in the initial empirical therapy of bacterial meningitis in immunosuppressed patients.

Nucleic Acids Res, 2004 Apr 28, 32(8), 2386 - 95 Print 2004.
Whole genome comparisons of serotype 4b and 1/2a strains of the food-borne pathogen Listeria monocytogenes reveal new insights into the core genome components of this species; Nelson KE et al.; The genomes of three strains of Listeria monocytogenes that have been associated with food-borne illness in the USA were subjected to whole genome comparative analysis . A total of 51, 97 and 69 strain-specific genes were identified in L.monocytogenes strains F2365 (serotype 4b, cheese isolate), F6854 (serotype 1/2a, frankfurter isolate) and H7858 (serotype 4b, meat isolate), respectively . Eighty-three genes were restricted to serotype 1/2a and 51 to serotype 4b strains . These strain- and serotype-specific genes probably contribute to observed differences in pathogenicity, and the ability of the organisms to survive and grow in their respective environmental niches . The serotype 1/2a-specific genes include an operon that encodes the rhamnose biosynthetic pathway that is associated with teichoic acid biosynthesis, as well as operons for five glycosyl transferases and an adenine-specific DNA methyltransferase . A total of 8603 and 105 050 high quality single nucleotide polymorphisms (SNPs) were found on the draft genome sequences of strain H7858 and strain F6854, respectively, when compared with strain F2365 . Whole genome comparative analyses revealed that the L.monocytogenes genomes are essentially syntenic, with the majority of genomic differences consisting of phage insertions, transposable elements and SNPs.

Virology, 2004 May 1, 322(2), 337 - 48
Improved protection conferred by vaccination with a recombinant vaccinia virus that incorporates a foreign antigen into the extracellular enveloped virion; Kwak H et al.; Recombinant poxviruses have shown promise as vaccine vectors . We hypothesized that improved cellular immune responses could be developed to a foreign antigen by incorporating it as part of the extracellular enveloped virion (EEV) . We therefore constructed a recombinant vaccinia virus that replaced the cytoplasmic domain of the B5R protein with a test antigen, HIV-1 Gag . Mice immunized with the virus expressing Gag fused to B5R had significantly better primary CD4 T-cell responses than recombinant virus expressing HIV-Gag from the TK-locus . The CD8 T-cell responses were less different between the two groups . Importantly, although we saw differences in the immune response to the test antigen, the vaccinia virus-specific immune responses were similar with both constructs . When groups of vaccinated mice were challenged 30 days later with a recombinant Listeria monocytogenes that expresses HIV-Gag, mice inoculated with the virus that expresses the B5R-Gag fusion protein had lower colony counts of Listeria in the liver and spleen than mice vaccinated with the standard recombinant . Thus, vaccinia virus expressing foreign antigen incorporated into EEV may be a better vaccine strategy than standard recombinant vaccinia virus.

Microbiol Immunol, 2004, 48(4), 329 - 37
Modulation of the immune system by Listeria monocytogenes-mediated gene transfer into mammalian cells; Shen H et al.; In this study, we established a method for Listeria monocytogenes(Lm)-mediated gene transfer into mammalian cells to manipulate the immune response of the host during infection by pathogens . We used the Lm-mediated gene transfer method in an in vivo study to manipulate host immune responses against Leishmania major(L . major )-infection . The injection of Lm modulated the susceptible host into a resistant state against L . major-infection . A more efficient protective effect was obtained with the injection of IL-12-cDNA containing Lm, and the protective effect was stronger than that of the resistant strain . The protective mechanism of Lm-injection against L . major-infection observed here appeared to be a result of the activation of the local immune system by the Lm-mediated gene transfer method . The present study is the first demonstration that a gene introduced into a host by Lm works to modulate the murine host immune response against infections in vivo . Since this system strongly induces Th1 responses and suppresses Th2 responses in infected hosts, the system can be used for controlling infectious diseases and for protection against allergic responses in the future.

J Leukoc Biol, 2004 Jul, 76(1), 104 - 15 Epub 2004 Apr 23.
Changes in peritoneal myeloid populations and their proinflammatory cytokine expression during infection with Listeria monocytogenes are altered in the absence of gamma/delta T cells; Skeen MJ et al.; Evidence that gamma/delta T cells play a broad, immunoregulatory role has been accumulating steadily . We show here that myeloid cells are disregulated after peritoneal infection with Listeria monocytogenes in mice lacking gamma/delta T cells . Inflammatory populations of neutrophils and monocytes recruited to the site of infection remained longer . Intracellular cytokine analysis showed that frequencies of myeloid cells producing interleukin-12 and tumor necrosis factor alpha were higher and remained elevated longer after infection in mice genetically deficient in gamma/delta T cells . In vivo dye-tracking studies indicated that the majority of inflammatory monocytes differentiated into resident tissue macrophages in situ . In vitro experiments confirmed that monocytes harvested from mice lacking gamma/delta T cells were defective in their maturation process . This evidence suggests that gamma/delta T cells promote differentiation in the monocyte/macrophage lineage . These cells are important for bactericidal activity, inflammatory cytokine production, clearance of inflammatory neutrophils, and ultimately, antigen presentation to T cells . Regulation of monocyte/macrophage differentiation may underlie a broad segment of the phenotypic alterations that have been reported in mice lacking gamma/delta T cells.

J Med Microbiol, 2004 May, 53(Pt 5), 399 - 402
Use of PFGE to characterize clonal relationships among Belgian clinical isolates of Listeria monocytogenes; Yde M et al.; The Belgian Listeria Reference Centre receives between 30 and 50 human clinical strains of Listeria monocytogenes per year . In general, epidemiological data are absent or incomplete, preventing recognition of episodes of listeriosis . However, data on a clonal relationship between strains can indirectly give an idea of the occurrence of episodes . Human isolates of L . monocytogenes from 2001 were serotyped, their arsenic-cadmium resistance profiles were determined, and they were pulsotyped with the application of pulsed-field gel electrophoresis using AscI and ApaI restriction endonucleases . On five occasions, two or more strains presented the same serovar, metal-resistance profile and pulsovar, suggesting a clonal relationship . This is the first report to identify accurately potential listeriosis episodes occurring in Belgium.

J Cell Sci, 2004 Apr 15, 117(Pt 10), 2121 - 30
Unconventional myosin VIIa and vezatin, two proteins crucial for Listeria entry into epithelial cells; Sousa S et al.; Listeria monocytogenes is a bacterial pathogen with the capacity to invade non-phagocytic cells . This dynamic process involves coordinated membrane remodelling and actin cytoskeleton rearrangements . Although some of the molecular factors promoting these events have been identified, the driving force allowing internalization is unknown . One of the receptors for L . monocytogenes on epithelial cells is E-cadherin, a transmembrane protein normally involved in homophilic interactions that allow cell-cell contacts at the adherens junctions . E-cadherin has to be connected to the actin cytoskeleton to mediate strong cell-cell adhesion and to trigger Listeria entry; alpha- and beta-catenins play key roles in these processes . We have recently identified an unconventional myosin, myosin VIIa and its ligand vezatin, at the adherens junctions of polarized epithelial cells . Here, we demonstrate by pharmacological and genetic approaches that both myosin VIIa and vezatin are crucial for Listeria internalization . These results provide the first evidence for the role of an unconventional myosin in bacterial internalization and a novel example of the exploitation of mammalian proteins, by a pathogen, to establish a successful infection.

Biol Neonate, 2004, 86(1), 66 - 72 Epub 2004 Apr 06.
Virulence and cord blood mononuclear cells cytokine production induced by perinatal listeria monocytogenes strains from different phylogenetic lineages; Mereghetti L et al.; Some of the phylogenetic lineages of Listeria monocytogenes are more likely to cause invasive disease in humans than are strains from other phylogenetic lineages . This suggests that strains belonging to these lineages display different levels of pathogenicity . To investigate this, we carried out a plaque-forming assay with HT-29 cells to evaluate the virulence of six perinatal strains from the three lineages that compose the species . All of the strains were largely over the 3.34 cutoff (between 4.29 and 5.97 mean log) with the HT-29 model and can therefore be considered to be equally virulent . We also explored part of the immune response of cord blood mononuclear cells by measuring cytokine production . All strains induced the production of similar amounts of TNF-alpha and IL-1beta . High concentrations of IL-6 and IL-8 were produced (between 6,000 and 17,000 pg/ml), whereas little or no IFN-gamma or IL-12 was produced . Thus, there is no difference between the strains from the three genetic lineages in terms of virulence or cytokine response . Given the epidemiological distribution of the serotypes responsible for human listeriosis and the genetic structure of the L . monocytogenes species, our results suggest: (i) that all strains from lineage I (serotypes 1/2b and 4b), a genetically homogeneous subpopulation, have a similar level of pathogenicity, and (ii) that lineage II (serotypes 1/2a and 1/2c), which is genetically more heterogeneous, is composed of strains with different levels of pathogenicity . The ones responsible for invasive diseases, particularly perinatal infections, display a similar level of pathogenicity to lineage-I strains and are not virulence-attenuated strains that can only infect the most immunocompromised hosts, whereas the other lineage-II strains are probably less pathogenic for humans .

J Food Prot, 2004 Apr, 67(4), 833 - 48
Antimicrobial edible films and coatings; Cagri A et al.; Increasing consumer demand for microbiologically safer foods, greater convenience, smaller packages, and longer product shelf life is forcing the industry to develop new food-processing, cooking, handling, and packaging strategies . Nonfluid ready-to-eat foods are frequently exposed to postprocess surface contamination, leading to a reduction in shelf life . The food industry has at its disposal a wide range of nonedible polypropylene- and polyethylene-based packaging materials and various biodegradable protein- and polysaccharide-based edible films that can potentially serve as packaging materials . Research on the use of edible films as packaging materials continues because of the potential for these films to enhance food quality, food safety, and product shelf life . Besides acting as a barrier against mass diffusion (moisture, gases, and volatiles), edible films can serve as carriers for a wide range of food additives, including flavoring agents, antioxidants, vitamins, and colorants . When antimicrobial agents such as benzoic acid, sorbic acid, propionic acid, lactic acid, nisin, and lysozyme have been incorporated into edible films, such films retarded surface growth of bacteria, yeasts, and molds on a wide range of products, including meats and cheeses . Various antimicrobial edible films have been developed to minimize growth of spoilage and pathogenic microorganisms, including Listeria monocytogenes, which may contaminate the surface of cooked ready-to-eat foods after processing . Here, we review the various types of protein-based (wheat gluten, collagen, corn zein, soy, casein, and whey protein), polysaccharide-based (cellulose, chitosan, alginate, starch, pectin, and dextrin), and lipid-based (waxes, acylglycerols, and fatty acids) edible films and a wide range of antimicrobial agents that have been or could potentially be incorporated into such films during manufacture to enhance the safety and shelf life of ready-to-eat foods.

J Food Prot, 2004 Apr, 67(4), 805 - 8
Prevalence and genetic diversity of Listeria monocytogenes in the tonsils of pigs; Autio T et al.; This study was set up to establish the prevalence of Listeria monocytogenes in the tonsils of sows and fattening pigs from five Finnish slaughterhouses and to evaluate the genetic similarity of L . monocytogenes strains isolated from the tonsils . A total of 271 pig tonsils (132 tonsils from fattening pigs and 139 from sows) from five different slaughterhouses in various parts of Finland were studied from June 1999 to March 2000 . Overall, 14 and 4% of pig tonsils harbored L . monocytogenes and Listeria innocua, respectively . The prevalence of L . monocytogenes in tonsils of fattening pigs (22%) was significantly higher than in sows (6%) . The isolates (n = 38) recovered from tonsils showed a wide genetic diversity by means of 24 different pulsed-field gel electrophoresis (PFGE) types presented by the strains . Moreover, in numerical analyses of restriction patterns, no association was found between the clustering of strains and the slaughterhouses, and strains showing a similar PFGE type were recovered from pigs of different slaughterhouses . The high prevalence of L . monocytogenes showing various PFGE types in the tonsils of pigs could indicate a potential source of contamination of pluck sets, carcasses, and the slaughterhouse environment and of subsequent processing steps.

J Food Prot, 2004 Apr, 67(4), 767 - 71
Reduction of microbial pathogens during apple cider production using sodium hypochlorite, copper ion, and sonication; Rodgers SL et al.; Sodium hypochlorite (100 ppm), copper ion water (1 ppm), and sonication (22 to 44 kHz and 44 to 48 kHz) were assessed individually and in combination for their ability to reduce populations of Escherichia coli O157:H7 and Listeria monocytogenes on apples and in apple cider . Commercial unpasteurized cider was inoculated to contain approximately 10(6) CFU/ml of either pathogen and then sonicated at 44 to 48 kHz, with aliquots removed at intervals of 30 to 60 s for up to 5 min and plated to determine numbers of survivors . Subsequently, whole apples were inoculated by dipping to contain approximately 10(6) CFU/g E . coli O157:H7 or L . monocytogenes, held overnight, and then submerged in 1 ppm copper ion water with or without 100 ppm sodium hypochlorite for 3 min with or without sonication at 22 to 44 kHz and examined for survivors . Treated apples were also juiced, with the resulting cider sonicated for 3 min . Populations of both pathogens decreased 1 to 2 log CFU/ml in inoculated cider following 3 min of sonication . Copper ion water alone did not significantly reduce populations of either pathogen on inoculated apples . However, when used in combination with sodium hypochlorite, pathogen levels decreased approximately 2.3 log CFU/g on apples . Sonication of this copper ion-sodium hypochlorite solution at 22 to 44 kHz did not further improve pathogen reduction on apples . Numbers of either pathogen in the juice fraction were approximately 1.2 log CFU/ml lower after being juiced, with sonication (44 to 48 kHz) of the expressed juice decreasing L . monocytogenes and E . coli O157:H7 populations an additional 2 log . Hence, a 5-log reduction was achievable for both pathogens with the use of copper ion water in combination with sodium hypochlorite followed by juicing and sonication at 44 to 48 kHz.

J Food Prot, 2004 Apr, 67(4), 751 - 7
Effect of pH on the survival of Listeria innocua in calcium ascorbate solutions and on quality of fresh-cut apples; Karaibrahimoglu Y et al.; Fresh-cut apple slices were dipped in calcium ascorbate (CaA) solution at pH values ranging from 2.5 to 7.0 to inhibit browning . After treatment, the cut apples were stored at 4 and 10 degrees C for up to 21 days . Color and texture of the apples were determined on days 1, 14, and 21 . In a separate experiement, the pH of CaA solution was adjusted with acetic acid to six different pH levels, and the solution was inoculated with Listeria innocua . The survival of the bacterium and the stability of CaA were determined at 0, 20, and 96 h . The cut apples maintained fresh quality when the pH of the CaA solution was above 4.5, but slight discoloration of apple slices dipped in pH 4.5 solution was observed after 14 days at 10 degrees C . At pH 5.0, the CaA dip maintained the quality of the apples at both temperatures for at least 21 days . The L . innocua population was reduced by 4 to 5 log CFU/ml at pH 4.5 after 96 h . At pH 5, the bacterial population in the CaA solution was reduced by approximately 2 log CFU/ml during the same period . The CaA solution was stable at pH 5 for at least 96 h . Reduction of the pH to between 4.5 and 5.0 might reduce the risk of foodborne illness due to consumption of fresh-cut apples treated with a CaA solution contaminated with Listeria.

J Food Prot, 2004 Apr, 67(4), 742 - 50
Influence of variations in methodology on populations of Listeria monocytogenes recovered from lettuce treated with sanitizers; Burnett AB et al.; The elimination of Listeria monocytogenes inoculated onto a piece of cut iceberg lettuce (3.8 by 3.8 cm) by treatment with chlorinated water (200 micrograms/ml free chlorine) and a 0.5% (wt/vol) solution of FIT Professional Line Antibacterial Cleaner (FIT) was investigated . The efficacy of the two sanitizers was not influenced by the composition of the medium used to culture the L . monocytogenes used in the inocula, the number of strains in the inoculum, or the recovery medium used to enumerate the pathogen on lettuce after treatment . Drying inoculum on lettuce for 45 min at 37 degrees C caused more cells to die or not be retrieved compared with drying inoculum for 30 min at 25 degrees C . However, the percentage of cells in the inoculum recovered from lettuce treated with chlorine or FIT was not significantly different, regardless of the drying method . Stomaching, homogenizing, or stomaching followed by homogenizing lettuce treated with sanitizers resulted in recovery of similar numbers of L . monocytogenes, indicating that stomaching and homogenizing are equivalent in extracting cells; the sequential use of both processing methods did not substantially increase the efficiency of recovery . Washing lettuce with water or treating lettuce with 200 micrograms/ml chlorine or FIT resulted in decreases in populations of 0.60, 1.76, and 1.51 log CFU per lettuce piece, respectively, regardless of variations in test parameters . Reductions caused by sanitizers were significantly greater (alpha = 0.05) than that observed for water but not significantly different from each other . It is concluded that evaluation of sanitizers for their efficacy in killing L . monocytogenes on lettuce can be determined by spot inoculating 50 microliters of a five-strain mixture of cells from 24-h cultures suspended in 5% horse serum albumen, followed by drying the inoculum for 45 min at 37 degrees C, treatment by submerging in 50 ml of sanitizer for 5 min, stomaching samples in 50 ml of Dey-Engley neutralizing broth for 2 min, and enumerating survivors on modified Oxford medium.

J Food Prot, 2004 Apr, 67(4), 721 - 31
A comparison of different chemical sanitizers for inactivating Escherichia coli O157:H7 and Listeria monocytogenes in solution and on apples, lettuce, strawberries, and cantaloupe; Rodgers SL et al.; Ozone (3 ppm), chlorine dioxide (3 and 5 ppm), chlorinated trisodium phosphate (100- and 200-ppm chlorine), and peroxyacetic acid (80 ppm) were assessed for reduction of Escherichia coli O157:H7 and Listeria monocytogenes in an aqueous model system and on inoculated produce . Initially, sanitizer solutions were inoculated to contain approximately 10(6) CFU/ml of either pathogen, after which aliquots were removed at 15-s intervals over a period of 5 min and approximately plated to determine log reduction times . Produce was dip inoculated to contain approximately 10(6) E . coli O157:H7 or L . monocytogenes CFU/g, held overnight, submerged in each sanitizer solution for up to 5 min, and then examined for survivors . In the model system study, both pathogens decreased > 5 log following 2 to 5 min of exposure, with ozone being most effective (15 s), followed by chlorine dioxide (19 to 21 s), chlorinated trisodium phosphate (25 to 27 s), and peroxyacetic acid (70 to 75 s) . On produce, ozone and chlorine dioxide (5 ppm) were most effective, reducing populations approximately 5.6 log, with chlorine dioxide (3 ppm) and chlorinated trisodium phosphate (200 ppm chlorine) resulting in maximum reductions of approximately 4.9 log . Peroxyacetic acid was the least effective sanitizer (approximately 4.4-log reductions) . After treatment, produce samples were stored at 4 degrees C for 9 days and quantitatively examined for E . coli O157:H7, L . monocytogenes, mesophilic aerobic bacteria, yeasts, and molds . Populations of both pathogens remained relatively unchanged, whereas numbers of mesophilic bacteria increased 2 to 3 log during storage . Final mold and yeast populations were significantly higher than initial counts for chlorine dioxide- and ozone-treated produce . Using the nonextended triangle test, whole apples exposed to chlorinated trisodium phosphate (200 ppm chlorine) and shredded lettuce exposed to peroxyacetic acid were statistically different from the other treated samples.

Proc Natl Acad Sci U S A, 2004 Apr 20, 101(16), 5992 - 7 Epub 2004 Apr 12.
Forces generated during actin-based propulsion: a direct measurement by micromanipulation; Marcy Y et al.; Dynamic actin networks generate forces for numerous types of movements such as lamellipodia protrusion or the motion of endocytic vesicles . The actin-based propulsive movement of Listeria monocytogenes or of functionalized microspheres have been extensively used as model systems to identify the biochemical components that are necessary for actin-based motility . However, quantitative force measurements are required to elucidate the mechanism of force generation, which is still under debate . To directly probe the forces generated in the process of actin-based propulsion, we developed a micromanipulation experiment . A comet growing from a coated polystyrene bead is held by a micropipette while the bead is attached to a force probe, by using a specially designed "flexible handle." This system allows us to apply both pulling and pushing external forces up to a few nanonewtons . By pulling the actin tail away from the bead at high speed, we estimate the elastic modulus of the gel and measure the force necessary to detach the tail from the bead . By applying a constant force in the range of -1.7 to 4.3 nN, the force-velocity relation is established . We find that the relation is linear for pulling forces and decays more weakly for pushing forces . This behavior is explained by using a dimensional elastic analysis.

J Appl Microbiol, 2004, 96(5), 931 - 7
Effects of divercin V41 combined to NaCl content, phenol (liquid smoke) concentration and pH on Listeria monocytogenes ScottA growth in BHI broth by an experimental design approach; Lebois M et al.; AIMS: To investigate the main effects and interactions of different factors : divercin V41 (0-4 ng ml(-1)), NaCl content (0.5-5.5% w v(-1)), phenol (liquid smoke) concentration (0-8 ppm), and pH (5.5-7.5) on Listeria monocytogenes ScottA growth . METHODS AND RESULTS: Experiments were carried out in BHI broth using a central composite design . Divercin V41 (div41), NaCl content and pH were found to be the most influential factors whereas phenol concentration in liquid smoke had no effect on L . monocytogenes ScottA growth in our experimental domain . The combined effects of div41, NaCl content and pH decreased L . monocytogenes ScottA maximum specific growth rate (mu(max)) from 0.34 to 0.01 h(-1) and led to a significant increase in lag time (t(lag)) from 5.5 to 25 h . CONCLUSION: In this study, NaCl, pH and phenol conditions were similar to those currently observed in smoked salmon production . This shows that L . monocytogenes ScottA growth could be efficiently delayed by the use of div41 in addition to the usual technological hurdles . SIGNIFICANCE AND IMPACT OF THE STUDY: In conclusion, the technological hurdles of cold smoked salmon production could be further optimized and combined with the use of div41 or the div41 producer strain to improve the food safety of the product.

J Appl Microbiol, 2004, 96(5), 913 - 21
The diversity of Listeria monocytogenes strains from 10 Icelandic sheep farms; Gudmundsdottir KB et al.; AIMS: The purpose of this study was to examine the diversity of Listeria monocytogenes strains from healthy sheep, winter feed and environment of sheep farms in Iceland . METHODS AND RESULTS: A total of 104 L . monocytogenes isolates from animals, winter feed and environment on 10 Icelandic sheep farms were compared by serotyping, ribotyping, and pulsed-field gel electrophoresis with ApaI and AscI . The isolates were divided into 24 genotypes, all identified as serovars 1/2a, 1/2b, or 4b . Nine genotypes were detected on more than one farm . On three of the farms there seemed to be a dominant strain of L . monocytogenes . Isolates from incidents of listeriosis in animals occurring on two of the farms belonged to the genotype most commonly found on the particular farm . Nine of the 24 genotypes found on the sheep farms have been associated with disease in animals and/or humans elsewhere in Iceland . CONCLUSIONS: Certain strains of L . monocytogenes seem to be widely distributed on Icelandic sheep farms . On some farms there appears to be a dominant strain of L . monocytogenes . Incidents of listeriosis in animals may tend to be associated with strains commonly found on the farm . SIGNIFICANCE AND IMPACT OF THE STUDY: This study demonstrates the diversity of L . monocytogenes present in healthy sheep and their environment.

Risk Anal, 2004 Apr, 24(2), 409 - 14
Modeling growth and reduction of microorganisms in foods as functions of temperature and time; McMasters RL et al.; Food safety objectives (FSOs) are established in order to minimize the risk of foodborne illnesses to consumers, but these have not yet been incorporated into regulatory policy . An FSO states the maximum frequency and/or concentration of a microbiological hazard in a food at the time of consumption that provides an acceptable level of protection to the public and leads to a performance criterion for industry . However, in order to be implemented as a regulation, this criterion has to be achievable by the affected industry . In order to determine an FSO, the steps to produce and store that food need to be known, especially where they have an impact on contamination, growth, and destruction . This article uses existing models for growth of Listeria monocytogenes in conjunction with calculations of FSOs to approximate the outcome of more than one introduction of the foodborne organism throughout the food-processing path from the farm to the consumer . Most models for the growth and reduction of foodborne illnesses are logarithmic in nature, which fits the nature of the growth of microorganisms, spanning many orders of magnitude . However, these logarithmic models are normally limited to a single introduction step and a single reduction step . The model presented as part of this research addresses more than one introduction of food contamination, each of which can be separated by a substantial amount of time . The advantage of treating the problem this way is the accommodation of multiple introductions of foodborne pathogens over a range of time durations and conditions.

Risk Anal, 2004 Apr, 24(2), 389 - 99
Risk assessment of listeriosis linked to the consumption of two soft cheeses made from raw milk: Camembert of Normandy and Brie of Meaux; Sanaa M et al.; This article reports a quantitative risk assessment of human listeriosis linked to the consumption of soft cheeses made from raw milk . Risk assessment was based on data purposefully acquired inclusively over the period 2000-2001 for two French cheeses, namely: Camembert of Normandy and Brie of Meaux . Estimated Listeria monocytogenes concentration in raw milk was on average 0.8 and 0.3 cells/L, respectively, in Normandy and Brie regions . A Monte Carlo simulation was used to account for the time-temperature history of the milk and cheeses from farm to table . It was assumed that cell progeny did not spread within the solid cheese matrix (as they would be free to do in liquid broth) . Interaction between pH and temperature was accounted for in the growth model . The simulated proportion of servings with no L . monocytogenes cell was 88% for Brie and 82% for Camembert . The 99th percentile of L . monocytogenes cell numbers in servings of 27 g of cheese was 131 for Brie and 77 for Camembert at the time of consumption, corresponding respectively to three and five cells of L . monocytogenes per gram . The expected number of severe listeriosis cases would be < or =10(-3) and < or =2.5 x 10(-3) per year for 17 million servings of Brie of Meaux and 480 million servings of Camembert of Normandy, respectively.

Vet Rec, 2004 Mar 20, 154(12), 365 - 70
Clinicopathological findings in sheep from Sardinia showing neurological signs of disease; Ligios C et al.; Histopathological and bacteriological examinations were performed on 178 brains from Sardinian sheep which were showing neurological signs . The sheep represented the total number of sheep with neurological syndromes submitted for diagnostic investigations over a three-year period in Sardinia . Scrapie was detected in 57 cases, cerebrocortical necrosis in 25, intoxication by a typical Mediterranean plant (Cistus species) was suspected in 25, coenurosis was detected in 11 cases, Listeria monocytogenes in eight cases and focal symmetrical encephalomalacia in six cases . Non-suppurative inflammatory changes were observed in three of the brains and suppurative changes were noted in two . Lesions restricted to the spinal cord were found in three cases . In the remaining 38 cases there were no significant neuropathological changes.

Proc Natl Acad Sci U S A, 2004 Apr 20, 101(16), 6152 - 7 Epub 2004 Apr 08.
Targeting and crossing of the human maternofetal barrier by Listeria monocytogenes: role of internalin interaction with trophoblast E-cadherin; Lecuit M et al.; Listeria monocytogenes produces severe fetoplacental infections in humans . How it targets and crosses the maternofetal barrier is unknown . We used immunohistochemistry to examine the location of L . monocytogenes in placental and amniotic tissue samples obtained from women with fetoplacental listeriosis . The results raised the possibility that L . monocytogenes crosses the maternofetal barrier through the villous syncytiotrophoblast, with secondary infection occurring via the amniotic epithelium . Because epidemiological studies indicate that the bacterial surface protein, internalin (InlA), may play a role in human fetoplacental listeriosis, we investigated the cellular patterns of expression of its host receptor, E-cadherin, at the maternofetal interface . E-cadherin was found on the basal and apical plasma membranes of syncytiotrophoblasts and in villous cytotrophoblasts . Established trophoblastic cell lines, primary trophoblast cultures, and placental villous explants were each exposed to isogenic InlA+ or InlA- strains of L . monocytogenes, and to L . innocua expressing or not InlA . Quantitative assays of cellular invasion demonstrated that bacterial entry into syncytiotrophoblasts occurs via the apical membrane in an InlA-E-cadherin dependent manner . In human placental villous explants, bacterial invasion of the syncytiotrophoblast barrier and underlying villous tissue and subsequent replication produces histopathological lesions that mimic those seen in placentas of women with listeriosis . Thus, the InlA-E-cadherin interaction that plays a key role in the crossing of the intestinal barrier in humans is also exploited by L . monocytogenes to target and cross the placental barrier . Such a ligand-receptor interaction allowing a pathogen to specifically cross the placental villous trophoblast barrier has not been reported previously.

J Immunol, 2004 Apr 15, 172(8), 4866 - 74
Listeriolysin O from Listeria monocytogenes is a lymphocyte apoptogenic molecule; Carrero JA et al.; Infection of mice with Listeria monocytogenes caused marked lymphocyte apoptosis in the white pulp of the spleen on day 2 postinfection . We prove in this study that listeriolysin O (LLO), a pore-forming molecule and a major virulence factor of Listeria, could directly induce murine lymphocyte apoptosis both in vivo and in vitro at nanomolar and subnanomolar doses . Induction of apoptosis by LLO was rapid, with caspase activation seen as early as 30 min post-treatment . T cells lost their mitochondrial membrane potential and exposed phosphatidylserine within 8 h of treatment . Incubation of lymphocytes with a pan-caspase inhibitor blocked DNA laddering and caspase-3 activation, but did not block phosphatidylserine exposure or loss of mitochondrial membrane potential . We describe a novel function for LLO: induction of lymphocyte apoptosis with rapid kinetics, effected by both caspase-dependent and -independent pathways.

Appl Environ Microbiol, 2004 Apr, 70(4), 2383 - 90
Genetic markers unique to Listeria monocytogenes serotype 4b differentiate epidemic clone II (hot dog outbreak strains) from other lineages; Evans MR et al.; A small number of closely related strains of Listeria monocytogenes serotype 4b, designated epidemic clone I (ECI), have been implicated in numerous outbreaks of food-borne listeriosis described during the past two decades in Europe and North America . In 1998 to 1999, a multistate outbreak traced to contaminated hot dogs involved a different strain type of serotype 4b, with genetic fingerprints rarely encountered before . In spite of the profound economic and public health impact of this outbreak, the implicated bacteria (designated epidemic clone II {ECII}) have remained poorly characterized genetically, and nucleotide sequences specific for these strains have not been reported . Using genome sequence information, PCR, and Southern blots, we identified DNA fragments which appeared to be either absent or markedly divergent in the hot dog outbreak strains but conserved among other serotype 4b strains . PCR with primers derived from these fragments as well as Southern blots with the amplicons as probes readily differentiated ECII from other serotype 4b strains . The serotype 4b-specific region harboring these fragments was adjacent to inlA, which encodes a well-characterized virulence determinant . The findings suggest that ECII strains have undergone divergence in portions of a serotype-specific region that is conserved in other serotype 4b strains . Although the mechanisms that drive this divergence remain to be identified, DNA-based tools from this region can facilitate the detection and further characterization of strains belonging to this lineage.

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