Hygienic Shortcomings of Frozen Dessert Freezing Equipment and Fate of Listeria monocytogenes on Ice Cream-Soiled Stainless Steel.

Although frozen dairy desserts have a strong record of safety, recent outbreaks of foodborne disease linked to ice creams have brought new attention to this industry. There is concern that small-scale frozen dessert equipment may not comply with or be reviewed against published comprehensive design and construction sanitation specifications (National Sanitation Foundation or 3-A sanitary standards). Equipment sanitary design issues may result in reduced efficacy of cleaning and sanitation, thus increasing the likelihood of postprocess contamination with pathogenic bacteria. In this context, and given that Listeria monocytogenes outbreaks are of great concern for the frozen dessert industry, a complementary study was conducted to evaluate the fate of L. monocytogenes in ice cream mix on a stainless steel surface. Our results showed that L. monocytogenes survived for up to 6 weeks at room temperature and 9 weeks at 4°C in contaminated ice cream on a stainless steel surface. Furthermore, chlorine- and acid-based surface sanitizers had no detrimental effect on the L. monocytogenes when used at a concentration and contact time (1 min) recommended by the manufacturer; significant reduction in CFU required 5 to 20 min of contact time.

Bone marrow lymphoid and myeloid progenitor cells are suppressed in 7,12-dimethylbenz(a)anthracene (DMBA) treated mice.

In this study we used colony forming unit (CFU) assays to demonstrate rapid suppression (within 6h) of lymphoid (CFU-preB) and myeloid (CFU-GM) progenitor cells in DMBA-treated mice. The duration of these changes were consistent with the blood levels of DMBA and its metabolites that were achieved by either IP or oral DMBA administration. CFU-GM and CFU-preB activities returned to control levels by 2 and 7 days after oral DMBA exposure, respectively, but remained suppressed through 7 days after IP DMBA administration. The continued presence of low levels of DMBA in the bloodstream following IP administration was associated with sustained suppression of CFU-preB, total bone marrow lymphoid cells and peripheral blood lymphocytes. The changes noted above were not observed in Cyp1b1 null mice, demonstrating the need for local DMBA metabolism in the bone marrow by Cyp1b1 to impair bone marrow CFU-preB and CFU-GM. Furthermore, these data provide evidence that myeloid-lineage cells are restored more quickly than lymphoid-lineage cells after DMBA exposure.

Viable "Haemophilus somnus" induces myosin light-chain kinase-dependent decrease in brain endothelial cell monolayer resistance.

"Haemophilus somnus" causes thrombotic meningoencephalitis in cattle. Our laboratory has previously reported that H. somnus has the ability to adhere to, but not invade, bovine brain endothelial cells (BBEC) in vitro. The goal of this study was to determine if H. somnus alters brain endothelial cell monolayer integrity in vitro, in a manner that would be expected to contribute to inflammation of the central nervous system (CNS). Monolayer integrity was monitored by measuring transendothelial electrical resistance (TEER) and albumin flux. BBEC incubated with H. somnus underwent rapid cytoskeletal rearrangement, significant increases in albumin flux, and reductions in TEER. Decreased monolayer TEER was preceded by phosphorylation of the myosin regulatory light chain and was partially dependent on tumor necrosis factor alpha and myosin light-chain kinase but not interleukin-1beta. Neither heat-killed H. somnus, formalin-fixed H. somnus, nor purified lipooligosaccharide altered monolayer integrity within a 2-h incubation period, whereas conditioned medium from H. somnus-treated BBEC caused a modest reduction in TEER. The data from this study support the hypothesis that viable H. somnus alters integrity of the blood-brain barrier by promoting contraction of BBEC and increasing paracellular permeability of the CNS vasculature.

Roles of cellular activation and sulfated glycans in Haemophilus somnus adherence to bovine brain microvascular endothelial cells.

Haemophilus somnus can cause a devastating fibrinopurulent meningitis with thrombotic vasculitis and encephalitis in cattle. The mechanisms used by H. somnus to migrate from the bloodstream into the central nervous system (CNS) are unknown. In this study, we demonstrate that H. somnus adheres to, but does not invade, bovine brain endothelial cells (BBEC) in vitro. The number of adherent H. somnus was significantly increased by prior activation of the BBEC with tumor necrosis factor alpha (TNF-alpha). Addition of exogenous glycosaminoglycans significantly reduced H. somnus adherence to resting and TNF-alpha-activated BBEC. Heparinase digestion of the endothelial cell's glycocalyx or sodium chlorate inhibition of endothelial cell sulfated glycan synthesis significantly reduced the number of adherent H. somnus. In contrast, addition of hyaluronic acid, a nonsulfated glycosaminoglycan, had no inhibitory effect. These findings suggest a critical role for both cellular activation and sulfated glycosaminoglycans in adherence of H. somnus to BBEC. Using heparin-labeled agarose beads, we demonstrated a high-molecular-weight heparin-binding protein expressed by H. somnus. Heparin was also shown to bind H. somnus in a 4 degrees C binding assay. These data suggest that heparin-binding proteins on H. somnus could serve as initial adhesins to sulfated proteoglycans on the endothelial cell surface, thus contributing to the ability of H. somnus to infect the bovine CNS.

Inflammatory cytokines enhance the interaction of Mannheimia haemolytica leukotoxin with bovine peripheral blood neutrophils in vitro.

Mannheimia (Pasteurella) haemolytica A1 produces several virulence factors that play an important role in the pathogenesis of bovine pneumonic pasteurellosis. Foremost among these is a leukotoxin (LKT) that specifically kills ruminant leukocytes. Recent evidence suggests that M. haemolytica LKT binding to bovine leukocytes is mediated by the beta(2)-integrin CD11a/CD18 (lymphocyte function-associated antigen 1 [LFA-1]), which subsequently induces activation and cytolysis of these cells. Inflammatory cytokines, which are released during viral and bacterial infection, are reported to increase LFA-1 expression and conformational activation. We investigated the effects of the inflammatory cytokines interleukin-1beta (IL-1beta), tumor necrosis factor alpha (TNF-alpha), and gamma interferon (IFN-gamma) on the interaction of M. haemolytica LKT with bovine peripheral blood neutrophils (PMNs). In this study we demonstrated, by flow cytometry, that bovine PMNs increased their binding to an anti-bovine LFA-1 monoclonal antibody (BAT75A) following in vitro incubation with IL-1beta, TNF-alpha, or IFN-gamma. Incubation with cytokines also increased CD18 expression, as assessed by real-time PCR and by Western blotting. Increased LFA-1 expression by PMNs exposed to cytokines was associated with increased LKT binding and cytotoxicity. The latter represented, at least in part, enhanced PMN apoptosis, as assessed by propidium iodine staining and caspase-3 activation. The results of this study suggest that inflammatory cytokines may play an important role in enhancing the biological response of bovine PMNs to M. haemolytica LKT.

Use of Hoechst 33342 staining to detect apoptotic changes in bovine mononuclear phagocytes infected with Mycobacterium avium subsp. paratuberculosis.

Mycobacterium avium subsp. paratuberculosis is an intracellular pathogen of macrophages that causes a chronic enteritis (Johne's disease) in ruminants. The purpose of this study was to determine whether M. avium subsp. paratuberculosis infection causes apoptosis in bovine monocytes. Using Hoechst 33342 staining, we observed increased numbers of apoptotic monocytes within 6 h of infection with M. avium subsp. paratuberculosis, and these numbers increased further at 24 and 48 h. This effect appeared to require viable bacilli, because monocytes infected with heat-killed M. avium subsp. paratuberculosis did not exhibit a significant increase in apoptosis. Preincubation of monocytes with bovine growth hormone prior to infection with M. avium subsp. paratuberculosis did not significantly alter the number of apoptotic cells.

Cytochrome P4501B1 mediates induction of bone marrow cytotoxicity and preleukemia cells in mice treated with 7,12-dimethylbenz[a]anthracene.

Humans are exposed to polycyclic aromatic hydrocarbons (PAHs) through many environmental pollutants, especially cigarette smoke. These chemicals cause a variety of tumors and immunotoxic effects, as a consequence of bioactivation by P-450 cytochromes to dihydrodiol epoxides. The recently identified cytochrome P4501B1 (CYP1B1) bioactivates PAHs but is also a physiological regulator, as evidenced by linkage of CYP1B1 deficiency to congenital human glaucoma. This investigation demonstrates that CYP1B1 null mice are almost completely protected from the acute bone marrow cytotoxic and preleukemic effects of the prototypic PAH 7,12-dimethylbenz[a]anthracene (DMBA). CYP1B1 null mice did not produce the appreciable amounts of bone marrow DMBA dihydrodiol epoxide DNA adducts present in wild-type mice, despite comparable hepatic inductions of the prominent PAH-metabolizing P-450 cytochrome, CYP1A1. Wild-type mice constitutively expressed low levels of bone marrow CYP1B1. These findings suggest that CYP1B1 is responsible for the formation of DMBA dihydrodiol epoxides in the bone marrow. Furthermore, this study substantiates the importance of DMBA dihydrodiol epoxide generation at the site of cancer initiation and suggests that tissue-specific constitutive CYP1B1 expression may contribute to cancer susceptibility in the human population.

Bone marrow stromal cells constitutively express high levels of cytochrome P4501B1 that metabolize 7,12-dimethylbenz[a]anthracene.

The polycyclic aromatic hydrocarbon 7,12-dimethylbenz[a]anthracene (DMBA) is a potent carcinogen that produces immunotoxic effects in bone marrow. Here, we show that bone marrow stromal cells metabolize DMBA to such products as 3,4-dihydrodiol, the precursor to the most mutagenic DMBA metabolite. The BMS2 bone marrow stromal cell line constitutively expressed higher levels of CYP1B1 protein and mRNA than C3H10T1/2 mouse embryo fibroblasts. BMS2 cells also produced a DMBA metabolite profile that was consistent with CYP1B1 activity. Treatment with the potent aryl hydrocarbon receptor (AhR) ligand 2,3, 7,8-tetrachlorodibenzo-p-dioxin (TCDD) induced a approximately 2-fold increase in CYP1B1 mRNA, protein, and activity in BMS2 cells. Two forms of the AhR (97 and 104 kDa) and the AhR nuclear translocator were detected in BMS2 cells. The AhR translocated to the nucleus after treatment with TCDD or DMBA but was approximately 5 times slower with DMBA. Primary bone marrow stromal (BMS) cell cultures established from AhR-/- mice showed similar basal CYP1B1 expression and activity as cell cultures established from heterozygous littermates or C57BL/6 mice. However, primary BMS cells from AhR-/- mice did not exhibit increased CYP1B1 protein expression after incubation with TCDD. BMS cells therefore constitutively express functional CYP1B1 that is not dependent on the AhR. This contrasts with embryo fibroblasts from the same mouse strain, in which basal CYP1B1 expression is AhR dependent. We therefore conclude that bone marrow toxicity may be mediated by CYP1B1-dependent DMBA metabolism, which is regulated by factors other than the AhR.

Polymerase chain reaction analysis of TNF-alpha and IL-6 mRNA levels in whole blood from cattle naturally or experimentally infected with Mycobacterium paratuberculosis.

Johne's disease is characterized by a chronic enteritis that results in granulomatous inflammation, cachexia, and eventual death of cattle infected with Mycobacterium paratuberculosis. The cytokines tumor necrosis factor-alpha (TNF-alpha) and interleukin-6 (IL-6) have been associated with granuloma formation and wasting in other disease syndromes. The potential role of these cytokines in the development and progression of Johne's disease has not been investigated. Using the polymerase chain reaction (PCR) and specific bovine oligonucleotide cytokine primers and probes, we examined the expression of messenger RNA for these cytokines in whole blood from M. paratuberculosis infected and uninfected cattle. Cytokine mRNA levels were examined before and after in vitro incubation with E.coli lipopolysaccharide (LPS) and lipoarabinomannan (LAM) purified from M. paratuberculosis. Uninfected calves, experimentally infected calves, and naturally infected cattle all displayed similar cytokine mRNA expression patterns. However, individual animals demonstrated variability in the levels of IL-6 and TNF-alpha mRNA expression as determined by a semiquantitative PCR method using 32P-labelled oligonucleotide probes.

Inhibition of the multiplication of Listeria monocytogenes in a murine hepatocyte cell line (ATCC TIB73) by IFN-gamma and TNF-alpha.

The effects of IFN-gamma and TNF-alpha on intracellular multiplication of Listeria monocytogenes in a murine enbryonic hepatocyte cell line (ATCC TIB 73) were investigated. Neither IFN-gamma nor TNF-alpha alone (10, 25, or 100 ng/ml) significantly inhibited intracellular multiplication of L. monocytogenes. In contrast, addition of both IFN-gamma and TNF-alpha (10, 25, or 100 ng/ml) significantly inhibited intracellular listerial multiplication. The anti-listerial effects of IFN-gamma and TNF-alpha were not blocked by adding aminoguanidine or catalase. Nor did IFN-gamma and TNF-alpha treated hepatocyte monolayers produce detectable amounts of nitric oxide or hydrogen peroxide. These findings suggest that the effects of IFN-gamma and TNF-alpha may be independent of reactive oxygen (ROI) and nitrogen (RNI) intermediates.

Ex vivo induction of TNF-alpha and IL-6 mRNA in bovine whole blood by Mycobacterium paratuberculosis and mycobacterial cell wall components.

Johne's disease is a chronic enteritis of cattle and other ruminant species that is of worldwide economic importance. The cytokines tumor necrosis factor-alpha (TNF-alpha) and interleukin-6 (IL-6) have been associated with granuloma formation and wasting in other disease syndromes. The potential role of these cytokines in the development and progression of Johne's disease has not been investigated. Using reverse transcriptase polymerase chain reaction (RT-PCR) and specific bovine oligonucleotide cytokine primers and probes for bovine TNF-alpha and IL-6, we examined the ex vivo expression of mRNA for these inflammatory cytokines in whole blood from healthy cattle. Cytokine mRNA levels increased after a brief incubation of bovine whole blood with Mycobacterium paratuberculosis or its lipoarabinomannan (LAM). Muramyl dipeptide (MDP) and Escherichia coli LPS also stimulated TNF-alpha and IL-6 mRNA expression. Several strains of M. paratuberculosis were tested and found to have similar abilities to stimulate TNF-alpha and IL-6 mRNA expression. Several strains of the closely related Mycobacterium avium, and the unrelated saprophyte, Mycobacterium phlei, had somewhat less ability to stimulate TNF-alpha and IL-6 mRNA expression.

Dissociation of cytolysis and monokine release by bovine mononuclear phagocytes incubated with Pasteurella haemolytica partially purified leukotoxin and lipopolysaccharide.

The bovine respiratory pathogen Pasteurella haemolytica secretes an exotoxin that is specific for ruminant leukocytes (leukotoxin). Previous studies have shown that subcytolytic concentrations of the leukotoxin stimulate bovine neutrophils to undergo a respiratory burst and degranulate. Relatively little is known about the stimulatory effects of the leukotoxin on bovine mononuclear phagocytes. In this study, we compared the relative cytolytic effects of partially purified leukotoxin on bovine peripheral blood monocytes and alveolar macrophages. We found monocytes to be approximately 8- to 10-fold more sensitive than alveolar macrophages to the cytolytic effect of leukotoxin. In addition, incubation of monocytes and alveolar macrophages with sublethal doses of leukotoxin stimulated release of IL-1 and TNF activities in a dose-dependent manner. Addition of an antileukotoxin MAb neutralized the cytolytic effects of leukotoxin, but potentiated TNF release. Heat inactivation also blocked the cytolytic activity of LKT, but only slightly reduced its ability to stimulate TNF release. Although the leukotoxin preparations were estimated to have only small amounts of lipopolysaccharide (LPS) contamination, as determined by a standard Limulus amebocyte lysate coagulation assay, a chromogenic Limulus assay indicated much greater amounts of LPS were present. Adding equivalent doses of P. haemolytica LPS largely duplicated the monokine release stimulated by leukotoxin. These results suggest that the stimulatory effects of the P. haemolytica leukotoxin on bovine mononuclear phagocytes may principally involve LPS, perhaps complexed with leukotoxin.

Recombinant interleukin-12 enhances resistance of mice to Listeria monocytogenes infection.

The effect of recombinant murine IL-12 (rIL-12) or anti-IL-12 antibody administration on resistance to murine listeriosis was investigated. Mice given a single 0.5 micrograms dose of rIL-12 had 1.5 log10 fewer listeriae in their spleens and livers as compared with control infected mice 3 days after L. monocytogenes challenge. Conversely, administration of anti-IL-12 IgG caused an equivalent increase in the cfu of L. monocytogenes recovered from the spleens and livers as compared to control mice. This is the first report of such a protective effect from a single dose of rIL-12. Treatment of uninfected mice with rIL-12 induced IFN-gamma mRNA production in their livers. Infection of mice with L. monocytogenes caused a similar increase in IFN-gamma mRNA levels that was not increased further by concurrent treatment with rIL-12. Treatment of mice with an anti-IFN-gamma MAb eliminated the protective effect of IL-12 on Listeria infection. Expression of TNF-alpha, IL-10 and IL-12p40 mRNA in L. monocytogenes-infected mice were not significantly altered by administration of either anti-IL-12 IgG or rIL-12. rIL-12 administration was associated with increased serum AST levels, a measure of liver damage, 1 day after treatment in L. monocytogenes-infected mice. In addition, rIL-12 administration was associated with the increased presence of small inflammatory foci and necrotic hepatocytes in both infected and uninfected mice, suggesting a proinflammatory role for IL-12 in the liver.

Mycobacterial cell wall components induce the production of TNF-alpha, IL-1, and IL-6 by bovine monocytes and the murine macrophage cell line RAW 264.7.

Johne's disease is characterized by a chronic enteritis that results in granulomatous inflammation, cachexia, and eventual death of cattle infected with Mycobacterium paratuberculosis. The cytokines tumor necrosis factor-alpha (TNF-alpha), interleukin-1 (IL-1), and interleukin-6 (IL-6) have been associated with granuloma formation and wasting in other disease syndromes. The potential role of these cytokines in the development and progression of Johne's disease has not been investigated. Freshly isolated bovine peripheral blood monocytes and the murine macrophage cell line RAW 264.7 were examined for their ability to release inflammatory cytokines in response to mycobacterial cell wall components. Bovine monocytes and RAW 264.7 cells incubated with M. paratuberculosis lipoarabinomannan (LAM), muramyl dipeptide (MDP), or lipopolysaccharide (LPS) released TNF-alpha, IL-1 beta, and IL-6 as detected by appropriate bioassays. Using the RAW 264.7 cells, cytokine mRNA levels were elevated after in vitro incubation with live M. paratuberculosis or LPS as determined using a reverse-transcriptase polymerase chain reaction procedure.

In vivo administration of a monoclonal antibody against the type I IL-1 receptor inhibits the ability of mice to eliminate Mycobacterium paratuberculosis.

The purpose of this study was to determine the influence of endogenous interleukin-1 (IL-1) on resistance to paratuberculosis infection in experimentally infected gnotobiotic mice. Following a 6-month treatment with prednisolone to facilitate bacillary multiplication, control mice substantially reduced the numbers of M. paratuberculosis in the liver and ileum. In contrast, mice injected with a monoclonal antibody against the type I IL-1 receptor failed to reduce the numbers of M. paratuberculosis in the liver and ileum and exhibited more liver granulomas, which contained numerous acid-fast bacilli. These results indicate a significant role for endogenous IL-1 in host defense against experimental M. paratuberculosis infection in mice.

Treatment with anti-interleukin-10 monoclonal antibody enhances early resistance to but impairs complete clearance of Listeria monocytogenes infection in mice.

Mice that received an anti-interleukin-10 (anti-IL-10) neutralizing monoclonal antibody (MAb) (SXC-1) prior to infection with Listeria monocytogenes initially demonstrated resistance to the infection, as indicated by reduced recovery of L. monocytogenes from their spleens and livers during the first 5 days after challenge. Anti-IL-10 MAb-treated mice then demonstrated reduced resistance during the later stage of infection, as indicated by persistent infection with L. monocytogenes in their livers 11 days after challenge. Aspartate aminotransferase (AST) levels (a measure of liver damage) in the sera of control mice increased between 1 and 5 days after challenge, while anti-IL-10 MAb-treated mice maintained lower AST levels. At 7 days after challenge, AST levels in the sera of control mice decreased as the numbers of organisms declined. In contrast, AST levels increased as the infections persisted in anti-IL-10 MAb-treated mice. The AST levels in serum reflected liver histopathology as anti-IL-10 MAb-treated mice exhibited fewer granulomatous lesions and less necrosis of liver tissue than the control mice during the first 5 days after challenge. Anti-IL-10 MAb treatment altered the expression of inflammatory cytokine mRNAs during L. monocytogenes infection. Control MAb-treated mice exhibited increased expression of tumor necrosis factor alpha and granulocyte-macrophage colony-stimulating factor mRNA in their lives during L. monocytogenes infection, but this increase did not occur in anti-IL-10 MAb-treated mice. Gamma interferon mRNA expression in the livers of the control MAb-treated mice was increased between 1 and 5 days after L. monocytogenes challenge and then decreased at 7 days after challenge. In contrast, gamma interferon mRNA expression in the livers of anti-IL-10 MAb-treated mice was not decreased until 7 days after challenge. These results indicate that endogenous IL-10 has both beneficial and detrimental effects on the host response to L. monocytogenes infection in mice.

Administration of anti-granulocyte mAb RB6-8C5 impairs the resistance of mice to Listeria monocytogenes infection.

Mice injected i.p. with RB6-8C5 mAb experienced a profound depletion of neutrophils in the bloodstream and spleen and significant impairment of their resistance to experimental infection with Listeria monocytogenes. Control mice survived i.v. inoculation with 5 x 10(4) L. monocytogenes; whereas, most RB6-8C5 mAb-treated mice inoculated i.v. with as few as 10 L. monocytogenes died within 6 days. RB6-8C5 mAb treatment was particularly deleterious when given within the first 24 h after i.v. inoculation with L. monocytogenes; however, some adverse effect was observed even when administration was delayed until 3 or 5 days after bacterial inoculation. Histopathologic examination of the livers of RB6-8C5 mAb-treated mice revealed necrotic foci that were characterized by few inflammatory cells and massive numbers of Gram-positive bacteria within hepatocytes. Additional evidence that the effects of RB6-8C5 mAb administration were chiefly due to neutrophil depletion include: 1) the effects of RB6-8C5 mAb treatment occurred more rapidly than what is generally seen in mice treated with anti-T cell mAbs, 2) similar results were observed with normal and scid mice, 3) RB6-8C5 mAb administration did not diminish delayed-type hypersensitivity nor the ability of spleen cells from immunized mice to transfer resistance, and 4) natural killer cell activity was unaffected by RB6-8C5 mAb administration. The results of this study provide additional evidence in support of the importance of neutrophils in the early stage of innate resistance to murine listeriosis.

Analysis of cytokine mRNA expression in Listeria-resistant C57BL/6 and Listeria-susceptible A/J mice during Listeria monocytogenes infection.

This laboratory previously reported that mRNA expression for many cytokines, as determined by reverse transcription-polymerase chain reaction analysis, is induced rapidly in the spleen during murine listeriosis. In the present study, the patterns of cytokine mRNA expression in spleens and livers of Listeria-resistant C57BL/6 and Listeria-susceptible A/J mice were compared. In addition, in situ hybridization was performed to evaluate the distributions of cytokine mRNA-expressing cells in these tissues. Listeria-resistant C57BL/6 mice demonstrated greater expression of gamma interferon (IFN-gamma) and granulocyte-macrophage colony-stimulating factor (GM-CSF) mRNAs in the spleen than Listeria-susceptible A/J mice. Greater numbers of cells expressing IFN-gamma and GM-CSF mRNAs were observed by in situ hybridization in the spleens of C57BL/6 mice than in those of A/J mice. C57BL/6 and A/J mice did not differ in their expression of IFN-gamma mRNA in the liver. Nor did C57BL/6 and A/J mice differ in their expression of tumor necrosis factor alpha, interleukin-1 alpha (IL-1 alpha), IL-2, IL-4, or IL-6 mRNA in the liver or spleen, as determined by reverse transcription-polymerase chain reaction and in situ hybridization. These results indicate that the greater resistance of C57BL/6 mice to Listeria monocytogenes infection is associated with greater expression of IFN-gamma and GM-CSF mRNAs in the spleen and GM-CSF mRNA in the liver.

Multiplication of Listeria monocytogenes in a murine hepatocyte cell line.

Listeria monocytogenes was shown to invade and multiply in a murine hepatocyte cell line (ATCC TIB73). Hemolytic and nonhemolytic L. monocytogenes strains exhibited similar abilities to invade hepatocytes, but only hemolytic L. monocytogenes multiplied within this cell line. Microscopic evaluation of monolayers stained with Wright stain demonstrated focal necrosis (plaques) in the hepatocyte monolayers, with large numbers of intracellular listeriae visible within the hepatocytes that lined the margins of these plaques. Murine recombinant interleukin-1 alpha, human recombinant tumor necrosis factor alpha, and murine recombinant gamma interferon did not affect the multiplication of L. monocytogenes in the hepatocytes. These data confirm in vivo observations of the intracellular multiplication of L. monocytogenes in hepatic lesions in infected mice.

Induction of interleukin-1 release by high- and low-passage isolates of Borrelia burgdorferi.

Low-passage isolates of Borrelia burgdorferi induced arthritis when injected into the hind paws of irradiated hamsters, while high-passage isolates did not. To examine a possible mechanism for induction of arthritis, peritoneal exudate cells were coincubated with high- and low-passage isolates of B. burgdorferi, and the resultant conditioned medium was assayed for interleukin-1 (IL-1) activity. Comparable amounts of IL-1 activity were detected in culture supernatants generated by high- and low-passage spirochetes and were dependent on the number of spirochetes added. Live B. burgdorferi stimulated greater release of IL-1 activity than did heat-killed organisms. No evidence of release of IL-1 due to shedding of soluble components from spirochetes was obtained. A recombinant human IL-1 receptor antagonist blocked the proliferative activity of conditioned medium in a murine thymocyte assay for IL-1 activity. The greater ability of low-passage spirochetes to survive in vivo may be more important than the ability to induce IL-1 production in the pathogenesis of Lyme arthritis.