In vitro antibody response to the tetrapeptide repeats of Plasmodium falciparum circumsporozoite protein by splenocytes from mice infected with P. yoelii yoelii

The antibody response to the recombinant protein, R32tet32, which contained the repetitive sequence (NANP)n of Plasmodium falciparum CSP was determined in C57BL/6 mice during the course of nonlethal infection with Plasmodium yoelii 17X. Marked suppression of the IgG antibody response to R32tet32 occurred when mice were immunized at peak parasitemia (on day 16). In vitro antibody responses of spleen cells from acutely infected mice to R32tet32 were similarly suppressed. Stimulation of normal spleen cells cultured for 5 days with 100 ng/ml of R32tet32 gave an optimal IgG antibody response, but spleen cells from infected mice obtained at peak parasitemia failed to respond to a broad range of antigen concentrations. Cocultivation studies employing enriched lymphocyte populations from infected and uninfected C57BL/6 mice indicated that both T and B cells from infected mice were defective in their response to R32tet32. The response to the repetitive region was restored by the addition of recombinant mouse interleukin-2 (IL-2) at a dose of 50 U/ml to cultures of spleen cells from infected mice.     

The role of CD4(+) or CD8(+) T cells in the protective immune response of BALB/c mice to Neospora caninum infection

The role of CD4(+) or CD8(+) T cells in the immune response of BALB/c mice against Neospora caninum infection was examined by using anti-CD4 and/or anti-CD8 monoclonal antibodies (mAbs). Mice were intraperitoneally inoculated with anti-CD4 and/or anti-CD8 mAbs before and after infection with N. caninum and observed for 30 days after infection. Most of the anti-CD4 mAb-treated mice and all of the anti-CD4 and anti-CD8 mAbs-treated mice died within 30 days post-infection (p.i.). In contrast, 100% of PBS-treated mice and 70% of anti-CD8 mAb-treated mice survived more than 30 days. When compared with phosphate-buffered saline (PBS)-treated mice, the weight of mice treated with mAbs tended to decrease. From these results CD4(+) T cells, but not CD8(+) T cells, have an important role for protection of mice against N. caninum infection. Serum antibody levels to N. caninum in infected-mice treated with anti-CD4 mAb or a mixture of anti-CD4 and anti-CD8 mAbs were lower than those in the infected mice treated with anti-CD8 mAb or PBS. The mice treated with anti-interferon-gamma (IFN-gamma) mAb produced high antibody levels to N. caninum, but all mice died within 18 days p.i. These results indicated that IFN-gamma is an important cytokine for protection against N. caninum infection at the early stage of infection. However, since CD4(+) T cells against N. caninum were essential to the production of specific antibody, these antibodies might have important roles in host protection at the later stage of infection.     

Evaluation of the suitability of monoclonal antibodies for flow cytometric intracellular cytokine detection in porcine peripheral blood lymphocytes

Panels of monoclonal antibodies (mAbs) specific for porcine interleukin (IL)-2, IL-4, IL-6, IL-12, interferon (IFN)-gamma and tumour necrosis factor (TNF)-alpha were evaluated for their applicability in flow cytometric intracellular cytokine detection. Peripheral blood mononuclear cells were short-time stimulated in the presence of brefeldin-A, ionomycin and phorbol-12-myristate-13-acetate, fixed and incubated with the respective mAbs as well as phycoerythrin-conjugated second-step antibodies. Suitability of mAbs was judged by use of statistical data and by visual control of scattergrams, considering the capability of mAbs to discriminate between cytokine-positive and cytokine-negative cell populations. The number of suitable clones differed to a high degree between the investigated cytokines, but at least one mAb fitting for flow cytometric intracellular cytokine detection could be identified within each panel. Monoclonal Abs producing scattergrams with a clear demarcation between positive and negative cell populations were found within the anti-IL-2, IL-6 and IFN-gamma panels, whereas less well defined positive and negative cell populations could be generated by use of mAbs within the anti-IL-4 and TNF-alpha panels. Only one moderately fitting mAb was identified within the anti-IL-12 panel. After having evaluated the best fitting mAbs, these were used to obtain reference levels for the physiological range of porcine lymphocytic cytokine production in a second set of experiments. For that reason, 13 clinically healthy pigs aged between 6 weeks and 6 months were investigated. Data presented are given as mean +/- SD of the percentage of positive-staining lymphocytes: IL-2, 45.5 +/- 27.6; IL-4, 34.1 +/- 21.3; IL-6, 45.4 +/- 23.8; IL-12, 13.9 +/- 8.6; TNF-alpha, 43.4 +/- 11.3; IFN-gamma, 65.5 +/- 14.8.     

A rodent model for allergic dermatitis induced by flea antigens

There have been very few reports of experimentally induced animal models of allergic dermatitis, an immunologic disorder. This report describes the induction of histopathology confirmed allergic dermatitis in C57BL/6 mice along with the consistent clinical sign of alopecia following the administration of flea antigens emulsified in complete Freund's adjuvant (CFA). By comparing different strains of mice, routes of injection, types of adjuvants and different dosages of flea antigens, C57BL/6 mice were found to be most susceptible to flea antigens administered intramuscularly (i.m.) and subsequently developed dermatologic excoriations and local alopecia. The level of specific IgE reactive to flea antigens in C57BL/6 mice after the onset of clinical signs was significantly higher than such levels in mice without clinical signs, suggesting that flea antigen-specific IgE level can be correlated to the severity of allergic hyper-reaction. CD4(+) T lymphocytes and IL-4 rather than IL-10, or IFN-gamma were found to be the predominant cytokines associated with the clinical onset of allergic symptoms in C57BL/6 mice. Further, histopathologic analysis indicated that not only mast cells had infiltrated into the area of the skin lesion, but the damage was found to be at a stage where mast cells were degranulating causing considerable exacerbation of the local injury. In conclusion, this murine allergic dermatitis model induced by flea antigens may provide a useful means to evaluate vaccines or immunodulatory drugs; thus providing researchers with a tool to study allergy-related disorders and other parameters needed in the area of allergic investigations.     

Relationship between type 1/type 2 immune responses and occurrence of vertical transmission in BALB/c mice infected with Neospora caninum

To examine the relationship between occurrence of vertical transmission and type 1/type 2 immune responses induced by Neospora caninum infection in BALB/c mice, pregnant (group 1 p) and non-pregnant mice (group 1 np) were inoculated with 2 x 10(6) of the N. caninum parasites and then we examined the vertical transmission rate and production of IFN-gamma and IL-4. We also studied chronically infected mice, which were bred at 4 weeks or more after infection (group 2), and mice inoculated during pregnancy and re-bred at 4 weeks or more after delivery (group 3). In groups 1p, 2 and 3, vertical transmission was observed in 27.4, 41.4, and 50% of the offspring, respectively. The serum IFN-gamma level increased on days 1 and 5 post-inoculation (p.i.) in groups 1 p and 1 np, while no increase level was observed in groups 2 and 3 during pregnancy or after delivery. When the mice in groups 2 and 3 were re-inoculated, all mice showed a transient increase in serum IFN-gamma on day 1 post-re-inoculation. The serum IL-4 level in both of groups 1p and np increased in a similar manner following infection. In group 3, the serum IL-4 level was somewhat higher than that in group 2 after re-inoculation. The anti-N. caninum antibody IgG1 titer in group 3 increased on day 10 post-re-inoculation. These results suggest that the mice infected during pregnancy may acquire a weaker immune response to the parasite than mice infected when they are not pregnant, and that mice infected during pregnancy may show an enhanced type 2 immune response in the recrudescence of the infection.     

IL-4受体阻断对小鼠急性感染肝片吸虫后肝脏单核细胞悬液中细胞因子的影响

本研究旨在探索肝片吸虫感染小鼠急性期肝脏单核细胞悬液中细胞因子表达模式及其原因。试验将小鼠分为4组,分别对感染抗体处置组、感染非抗体处置组、抗体处置非感染组和非抗体处置非感染组的肝脏单核细胞培养上清液中IL-4、IL-5、IL-13及IFN-γ进行测定;同时也对肝脏中IL-4及IL-12P40 mRNA水平进行荧光定量PCR检测。结果显示,IL-4和IL-5的表达模式相似。在非感染状态下,无论是否进行抗体处置,两者都处于较低表达水平。在感染状态下,两者在非抗体处置小鼠中的表达显著高于抗体处置组(P<0.05)。IL-13与IL-4及IL-5的表达差异在于感染并用抗体处置后,小鼠肝脏单核细胞中的IL-13仍有较高水平表达。IFN-γ的表达量在感染后有所增加,但在有无抗体处理间无显著差异(P>0.05)。mRNA检测结果显示,在非感染状态下,抗体处置后的IL-4 mRNA水平显著低于非抗体处置组(P<0.05)。感染后非抗体处置组小鼠中IL-4 mRNA水平极显著高于抗体处置组(P<0.01),同时也极显著高于非感染非抗体处置组(P<0.01)。IL-12P40 mRNA的水平在感染状态下,抗体处置组的IL-12P40 mRNA虽较非抗体处置组低,但无显著差异(P>0.05)。但感染后非抗体处置组的IL-12P40 mRNA水平均极显著高于感染组抗体处置和非抗体处置组(P<0.01)。本试验结果表明,IL-4Rα对肝片吸虫感染急性期IL-5和IL-4的表达极其重要,但对IL-13及IFN-γ的影响较小。     

Vitamin E supplementation does not mitigate the acute morbidity effects of porcine reproductive and respiratory syndrome virus in nursery pigs

The objective of this study was to determine whether feeding a vitamin E-rich diet would benefit nursery pigs infected with porcine reproductive and respiratory syndrome virus (PRRSV). Sixty-four pigs were subjected to one of four treatment combinations (2 x 2 factorial) of dietary vitamin E (adequate or excess) and PRRSV (medium or inoculation with VR-2385 isolate P-129). Pigs were fed experimental diets during a 3-wk period before inoculation as well as during a 12-d period after inoculation. Growth performance was determined throughout the study, and lipid peroxidation in liver, glutathione peroxidase (GPX) activity in serum, circulating white blood cells, and serum interleukin-1beta (IL-1beta) and interferon-gamma (IFN-gamma) were determined in samples collected from pigs killed 4 or 12 d after inoculation. Infection by PRRSV (P < 0.001) induced a marked decrease in both ADFI and ADG, but neither the main effect of diet nor the diet x PRRSV interaction was significant. Neither diet nor PRRSV affected feed efficiency. At 12 d after inoculation, lipid peroxidation in liver and GPX activity in serum were lower in pigs fed excess vitamin E than in those fed adequate vitamin E (P < 0.01), suggesting that the diet high in vitamin E bolstered the antioxidant status of the pigs. However, PRRSV did not affect lipid peroxidation in liver or serum GPX activity, and the diet x PRRSV interaction was not significant. White blood cell counts were decreased and IFN-gamma, and IL-1beta were increased (P < 0.05) 4 and 12 d after inoculation in PRRSV-infected pigs, but neither diet nor the diet x PRRSV interaction was significant. Collectively, these results indicate that increasing antioxidant defenses by feeding high levels of vitamin E did not ameliorate the effects of PRRSV on decreased growth, leukopenia, and increased serum IL-1beta and IFN-gamma. Thus, feeding nursery pigs a diet high in vitamin E may not be useful for mitigating the acute morbidity effects of PRRSV infection.     

Immunostimulation in mice infected with Sendai virus

The effect of Sendai virus infection on the splenic primary plaque-forming cell (PFC) response to sheep RBC in 2 strains of mice, with contrasting susceptibility to Sendai viral pneumonia, was examined. Mice were given single inoculations of sheep RBC, which varied relative to time of inoculation with Sendai virus, PFC were counted 6 days later, and were compared with PFC responses from noninfected mice. The IgM- and IgG-PFC responses were augmented in resistant C57BL/6J mice 7 and 9 days after inoculation with Sendai virus (sheep RBC given 1 and 3 days after inoculation with Sendai virus, respectively) and in susceptible DBA/2J mice 7, 9, 10, and 13 days after inoculation with Sendai virus. Augmentation was restricted mainly to IgM-, IgG3-, and IgG2b-PFC. The number of splenic background antitrinitrophenyl sheep RBC PFC in mice of both strains was examined during the course of Sendai virus infection. Only a marginal increase in background PFC was seen in C57BL/6J mice on or after viral inoculation day 11 and no change was seen in DBA/2J mice. Serum of infected mice also was examined sequentially for alpha/beta interferon (IFN). Despite vigorous lung IFN production, infected mice rarely had detectable circulating IFN. Seemingly, Sendai virus infection can induce transient hyperresponsiveness to a nonviral antigen.     

The histopathogenesis of paralytic rabies in six-week-old C57BL/6J mice following inoculation of the CVS-11 strain into the right triceps surae muscle

A fatal encephalomyelitis was developed after intracerebral and hind limb inoculation of in 6-week-old C57BL/6J mice by the inoculation of fixed rabies virus (CVS-11 strain), intracerebrally and into hind. After the intracerebral inoculation, virus antigens were detected in the cerebral cortex and hippocampus at 2 days postinoculation (PI), and later spread centrifugally to thalamus, brain stem, cerebellum, spinal cord and spinal ganglia. At 4 days PI, severe apoptosis and DNA fragmentation were observed in the hippocampus and cerebral cortex. All mice infected intracerebrally were dead without limb paralysis at from 10 to 11 days PI. In contrast, mice infected with virus intramuscularly were persistently observed virus antigens in the myocytes at the site of inoculation from 2 days PI. At 4 days PI, the antigens were demonstrated in the spinal dorsal root ganglia, spinal cord and muscle spindles without their detection in the cerebrum and hippocampus. There were no apoptosis in the spinal cord and dorsal root ganglia, however hind limb paralysis was found in all infected mice. Hind limb paralysis was progressed to quadriparalysis, and mice were dead from 11 to 13 days PI. From 4 days PI, necrosis of neuron was observed in the the spinal and dorsal ganglia with infiltration of lymphocyte. This study suggested that the necrosis of spinal neurons was more important to cause the paralysis of hind limb rather than the severe cerebral infection and apoptosis in C57BL/6J mice infected with CVS-11 strain. The virus primarily replicated in the muscles was ascended the spinal cord via afferent fibers and retrogradely invaded the cerebrum, and with subsequent spread to muscle spindles.     

坏死梭杆菌FN(A)p2001株小鼠感染模型的建立

将鹿源坏死梭杆菌FN(A)p2001分离株以3×108、3×107、3×106、3×105、3×104、3×103个/只等不同菌量,分别接种于6组小鼠,每组10只,逐日观察小鼠感染情况.结果表明,3×106、3×107、3×108个/只菌量感染组小鼠,在接种后3 d～8 d先后死亡,5 d后死亡小鼠的病理变化明显,并从死亡小鼠内脏及脓汁中均检到长丝状及小杆状等多形态典型的坏死梭杆菌;3×104个/只和3×105个/只菌量感染仅表现体重减轻,3×103个/只菌量感染无明显临床表现.由此表明,坏死梭杆菌FN(A)P2001分离株具有很强的感染毒力;对小鼠的最小致死量为106个/只菌量;腹腔接种是适宜的感染途径.从而建立了坏死梭杆菌分离株小鼠感染模型.     

Characterization of monoclonal antibodies to equine interleukin-10 and detection of T regulatory 1 cells in horses

Interleukin-10 (IL-10) terminates inflammatory immune responses and inhibits activation and effector functions of T-cells, monocytes, macrophages and dendritic cells. IL-10 has also been found to be a key cytokine expressed by subpopulations of regulatory T-cells. In this report, we describe the generation and characterization of three monoclonal antibodies (mAbs) to equine IL-10. The antibodies were found to be specific for equine IL-10 using different recombinant equine cytokine/IgG fusion proteins. Two of the anti-equine IL-10 mAbs were selected for ELISA to detect secreted IL-10 in supernatants of mitogen stimulated equine peripheral blood mononuclear cells (PBMC). The sensitivity of the ELISA for detecting secreted IL-10 was found to be around 200pg/ml. The production of intracellular IL-10 was measured in equine PBMC by flow cytometry. PBMC were stimulated with phorbol 12-myristate 13-acetate (PMA) and ionomycin in the presence of the secretion blocker Brefeldin A. All three anti-IL-10 mAbs detected a positive population in PMA stimulated lymphocytes which was absent in the medium controls. Around 80% of the IL-10(+) cells were CD4(+). Another 15% were CD8(+) cells. Double staining with IL-4 or interferon-gamma (IFN-gamma) indicated that PMA and ionomycin stimulation induced 80% IL-10(+)/IFN-gamma(+) lymphocytes, while only 5% IL-10(+)/IL-4(+) cells were observed. By calculation, at least 60% of the IL-10(+)/IFN-gamma(+) cells were CD4(+) lymphocytes. This expression profile corresponds to the recently described T regulatory 1 (T(R)1) cell phenotype. In summary, the new mAbs to equine IL-10 detected native equine IL-10 by ELISA and flow cytometry and can be used for further characterization of this important regulatory cytokine in horses.     

Modulation of cytokine gene expression and secretion during the periparturient period in dairy cows naturally infected with Mycobacterium avium subsp. paratuberculosis

Johne's disease (JD), caused by Mycobacterium avium subsp. paratuberculosis (MAP), is estimated to infect more than 22% of US dairy herds. Periods of immunosuppression may contribute to the transition from the subclinical to the clinical stage of infection. Understanding the effects of stressors such as parturition on the escalation of disease may provide information that will help to manage JD. The objective of this study was to characterize cytokine gene expression and secretion in periparturient dairy cows naturally infected with MAP. Blood was collected from the jugular vein of healthy noninfected, and subclinically and clinically infected dairy cows for 3 weeks pre-calving to 4 weeks post-calving. Real-time PCR was performed to evaluate the expression of the following cytokine genes by peripheral blood mononuclear cells: IFN-gamma, TNF-alpha, IL-12p35, IL-10, TGF-beta, and IL-4. To assess the effects of parturient immunosuppression on cytokine gene expression, RT-PCR data were analyzed by using 2(-ddCt) values calibrated to dCt value at +1 day relative to calving for each animal. Overall, cytokine gene expression was not influenced by infection status of the cows in this study. However, significant effects in cytokine gene expression were noted across sampling days within the periparturient period. Expression of IFN-gamma by NS and ConA-stimulated PBMCs declined at calving compared with prepartum values in both control and infected cows. Similarly, a decline in expression of IL-4 and IL-10 was observed for cells isolated from subclinically infected cows after stimulation with ConA. ConA-stimulated PBMCs isolated from infected cows secreted higher concentrations of IFN-gamma compared with the controls. A significant decline in IFN-gamma secretion was noted for MPS-stimulated cells for clinical cows from -21 days to +1 day. Stimulating cells with MPS resulted in greater secretion of IL-10 by infected cows during the postpartum period. A trend was also observed for higher TGF-beta secretion by NS PBMCs isolated from clinical cows in the postpartum period. Cells isolated from clinically infected cows and stimulated with MPS secreted higher levels of nitric oxide throughout the periparturient period when compared to control or subclinically infected cows. These data suggest that parturition is a very dynamic time period for host immunity, with potential for altered immunity to hinder the ability of dairy cows to thwart infectious diseases.     

Dehydroepiandrosterone increases resistance to experimental infection by Trypanosoma cruzi

Dehydroepiandrosterone (DHEA) enhances immune responses against a wide range of viral, bacterial, and parasitic pathogens. In a previous study, we reported that administration of DHEA significantly decreased the numbers of blood parasites in Trypanosoma cruzi experimental infection. The present study was undertaken to determine the effectiveness of DHEA in reducing the severity of acute phase T. cruzi infection of male and female Wistar rats. Animals were treated subcutaneously with 40 mg/kg body weight/day of DHEA. The concentration of nitric oxide (NO) was determined in spleen peritoneal cavity. Interleukin-2 (IL-2) and interferon-gamma (IFN-gamma) were determined in the sera of uninfected and infected animals. DHEA treatment augments NO production for both sexes after in vitro LPS treatment for uninfected animals. Infection triggered enhanced NO levels although not significant. IL-2 and IFN-gamma were detectable in higher concentrations in treated and infected rats of both genders when compared to untreated controls. These data suggest that DHEA may have a potent immunoregulatory function that can affect the course of T. cruzi infection.     

IL-4 and IL-10 inhibition of IFN-gamma- and TNF-alpha-dependent nitric oxide production from bovine mononuclear phagocytes exposed to Babesia bovis merozoites.

The requirement for IFN-gamma and/or TNF-alpha as co-stimulants with Babesia bovis merozoites for nitric oxide (NO) production was examined, as well as the regulatory role of IL-4 and IL-10. Purified B. bovis merozoites did not induce the production of NO in undifferentiated monocytes without addition of exogenous IFN-gamma and TNF-alpha unless the monocytes taken ex vivo were producing TNF-alpha endogenously. Under the latter condition, the NO production resulting from merozoite stimulation remained IFN-gamma-dependent. There was no evidence for endogenous synthesis of TNF-alpha in monocyte-derived macrophages (MDM), and merozoites alone were incapable of inducing TNF-alpha mRNA in MDM. However, while merozoites plus IFN-gamma induced TNF-alpha mRNA expression in MDM, NO was not produced. Both IL-4 and IL-10 inhibited expression of iNOS and production of NO in merozoite-stimulated monocytes.     

Regulated upon activation normal T-cell expressed and secreted (RANTES) contributes to abortion caused by Brucella abortus infection in pregnant mice

Brucella abortus (B. abortus) is a facultative intracellular pathogen that can survive inside macrophages and trophoblast giant cells, and the causative agent of brucellosis. In the present study, we found that production of regulated upon activation normal T-cell expressed and secreted (RANTES) due to B. abortus infection contributes to abortion in pregnant mice. B. abortus infected pregnant interferon-gamma (IFN-gamma) knockout mice died within 15 days of infection, but non-pregnant IFN-gamma knockout mice were still alive. With infection by wild type B. abortus, a large amount of RANTES production was observed in pregnant IFN-gamma knockout mice, and induction of RANTES was also observed in normal pregnant mice infected with the wild type, but not in those infected with the intracellular replication-defective mutant. Production of RANTES and IFN-gamma were inhibited in mice inoculated with the respective RANTES or IFN-gamma antibody. Neutralization of RANTES, induced by B. abortus infection, served to prevent abortion. These results indicate that the production and function of RANTES are correlated with IFN-gamma in pregnant mice infected with B. abortus.     

Optimization of the use of C57BL/6 mice as a laboratory animal model for Neospora caninum vaccine studies

The development and testing of vaccines for Neospora caninum in mice require challenge studies to demonstrate a reduction in clinical signs or prevention of vertical transmission of the parasite after vaccination. Genetic susceptibility to N. caninum varies with the strain of mice. In this study, C57BL/6 mice were evaluated as a model for Neospora vaccine studies. A lethal challenge model was developed and the LD(50) was determined to be 1.5 x 10(7)N. caninum tachyzoites/mouse, delivered intraperitoneally. Brain lesions encountered in sections from sub-lethally challenged mice were scored on the basis of severity and total number of lesions to develop a histopathological scoring system for vaccine efficacy. A vertical transmission model for N. caninum vaccine studies was developed by studying mice that were infected either 2 weeks prior to mating or between days 12 and 14 of pregnancy. It was found that infection prior to mating reduced the average number of pups per litter. DNA extracted from fetal tissue was examined by a N. caninum specific polymerase chain reaction (PCR). The rate of vertical transmission was 0, 100 and 90.5% for the uninfected controls, mice infected during pregnancy and mice infected before mating, respectively. This study demonstrates that the C57BL/6 strain of mice is a good model for N. caninum vaccine studies because it is possible to establish a clear-cut lethal challenge model in C57BL/6 mice and they transmit the disease to their offspring efficiently.     

Hepatitogenicity of three plaque purified mutants of hepatotropic mouse hepatitis virus, MHV-2.

Hepatitogenicity of three plaque purified mutant strains of mouse hepatitis virus, designated as MHV-2S, -2M and -2L, isolated from MHV-2 infected SR-CDF1-DBT cells was studied. After intraperitoneal inoculation with 2 x 10(5) PFU of parental MHV-2 and its mutants to 4-week-old female ICR mice, 40% of mice inoculated with MHV-2S and 20% of mice with -2M died in one week, whereas with -2L all mice survived. All mice inoculated with MHV-2 died in 3 days postinoculation (p.i.). Virus titer of the liver of mice inoculated with MHV-2, -2S and -2M reached peaks (MHV-2:10(7) PFU/0.2 g, -2S: 10(5) PFU/0.2 g and -2M: 10(6) PFU/0.2 g) at 96 hr p.i., while with -2L a peak titer (10(3) PFU/0.2 g) was shown at 48 hr p.i. Immunofluorescence revealed MHV specific antigen in the liver of MHV-2S infected mice in and around necrotic areas though less extensive than that of parental MHV-2 infected mice. With MHV-2M specific fluorescence was restricted in degenerated hepatocytes in the small necrotic foci. In mice inoculated with MHV-2L only faint fluorescence was detected. Histopathologically, in the liver of MHV-2S infected mice zonal necrosis and cell infiltration were observed. There were spotty necrosis and focal cell infiltration in the liver of MHV-2M infected mice and only small inflammatory foci were seen in MHV-2L infected mice. Large number of extracellular virions were detectable in MHV-2S but not in -2M and -2L infected mice by electron microscopy.     

The local immune response of mice after Helicobacter suis infection: strain differences and distinction with Helicobacter pylori

Helicobacter (H.) suis colonizes the stomach of pigs and is the most prevalent gastric non-H. pylori Helicobacter species in humans. Limited information is available on host immune responses after infection with this agent and it is unknown if variation in virulence exists between different H. suis strains. Therefore, BALB/c and C57BL/6 mice were used to compare colonization ability and gene expression of various inflammatory cytokines, as determined by real-time PCR, after experimental infection with 9 different H. suis strains. All strains were able to persist in the stomach of mice, but the number of colonizing bacteria at 59 days post inoculation was higher in stomachs of C57BL/6 mice compared to BALB/c mice. All H. suis strains caused an upregulation of interleukin (IL)-17, which was more pronounced in BALB/c mice. This upregulation was inversely correlated with the number of colonizing bacteria. Most strains also caused an upregulation of regulatory IL-10, positively correlating with colonization in BALB/c mice. Only in C57BL/6 mice, upregulation of IL-1β was observed. Increased levels of IFN-γ mRNA were never detected, whereas most H. suis strains caused an upregulation of the Th2 signature cytokine IL-4, mainly in BALB/c mice. In conclusion, the genetic background of the murine strain has a clear impact on the colonization ability of different H. suis strains and the immune response they evoke. A predominant Th17 response was observed, accompanied by a mild Th2 response, which is different from the Th17/Th1 response evoked by H. pylori infection.     

Infections in immunocompetent and immune-deficient mice with promastigotes of a North American isolate of Leishmania infantum

Leishmania infantum, an etiologic agent of zoonotic visceral leishmaniasis, is widespread among foxhounds in the United States. Experimental infections with a North American isolate of L. infantum were evaluated using two inoculation routes in immunocompetent and immunosuppressed mouse strains. Groups of 2-5 interferon gamma gene knockout (IFN-gamma-KO) (BALB/c-Ifng), inducible nitric oxide synthase (NOS) gene knockout (iNOS-KO) (C57BL/6), B-cell-deficient (microMT) (C57BL/6), and BALB/c mice were intravenously (i.v.) or subcutaneously (s.c.) inoculated with various doses of promastigotes of the LIVT-1 strain of L. infantum. None of the mice developed clinical signs of leishmaniasis during the 8-9 weeks of the study. Promastigotes were cultured from spleens of all i.v.-infected mice by 3 days post culture. Spleens from s.c.-infected mice inoculated with greater than 1 x 10(6) parasites became culture positive 3-24 days post culture, but promastigotes were not cultured from mice infected with 1 x 10(5) or 5 x 10(5) LIVT-1 promastigotes. Histological lesions were prominent in the livers of i.v.-infected mice but were mild to nonexistent in s.c. infection. Serological responses were low and transient determined by indirect fluorescent antibody testing in all groups. These results indicate that the i.v. route of infection is superior to the s.c. route in a mouse model of North American leishmaniasis and that mice lacking INF-gamma, iNOS or mice that are B-cell-deficient are not more susceptible to acute infection.