Immune responses of immunocompetent and immunocompromised mice experimentally infected with Bartonella henselae

The aim of this study is to understand host immune responses in immunocompetent and immunocompromised mice against Bartonella henselae infection. BALB/c and nude (BALB/c nu/nu) mice were inoculated intraperitoneally with 10(8) colony forming units of B. henselae (Houston-1 strain). Blood, brain, liver, spleen, kidney and bone marrow samples were collected 0, 3, 7, 14, 21 and 28 days after infection and submitted to bacteriological, serological and genetical examinations. B. henselae was isolated only from the liver 3 days after infection. DNA of the inoculums was detected by polymerase chain reaction from blood, liver, and spleen samples collected from BALB/c and blood from nude mice 3 and 7 days after infection. No bacterial DNA was detected from both BALB/c and nude mice thereafter during 4 weeks observation periods. These results indicate that the T-cell may not participate in the effective elimination of the organisms from mice. In addition, western blot analysis revealed that the antigens of 27.3- and 31.5-kDa reacted with IgM antibodies from the blood of BALB/c and nude mice after 3 days of infection, suggesting that these antigens were recognized by thymus-independent mechanism. Furthermore the antigens were detected from the culture-supernatants of B. henselae, indicating that these antigens were secreted from the organisms.     

A pregnant mouse model for bovine Tritrichomonas foetus infection

The economically important effects of Tritrichomonas foetus infection in cattle are abortion and infertility, yet there has not been an animal model to examine the parasite-host interactions during gestation. In this study, 5- and 7- to 8-week-old BALB/cAnNCr, BALB/cJ, and SCID/NCr mice on a BALB/c background were intravaginally infected with T. foetus. All BALB/cAnNCr and BALB/cJ mice, and 89% of SCID/NCr mice sustained infections for 13 weeks, if inoculated before 5 weeks of age. Infection rates were lower in all mouse strains inoculated at 7 weeks of age, although BALB/cAnNCr mice were significantly more susceptible than BALB/cJ or SCID/NCr mice. Vaginal bacterial flora did not account for the variation in mouse-strain susceptibility, although coagulase-negative staphylococci in vaginal flora were associated with failure of T. foetus to infect. As with infected cattle, T. foetus-specific vaginal immunoglobulin (Ig) G and IgA antibodies were elevated after infection. The number and viability of day-10 fetuses were reduced in mice infected at 5 weeks of age and bred 12 weeks after infection. Lesions in pregnant and nonpregnant infected mice, including suppurative and eosinophilic vaginitis; cervicitis; endometritis with distension of the uterine lumen; endometrial ulceration; and glandular ectasia, with neutrophils in the glandular lumen and loss of gland epithelium, were similar to those in cattle. The decidua and placenta were multifocally necrotic. Immunohistochemistry demonstrated trichomonads in vaginal folds and uterine glands, and adjacent to fetal tissues. In summary, experimentally infected BALB/cAnNCr mice showed many pathologic similarities to cattle and may serve as a model to study host-trichomonad interactions.     

Reduced formation of granulomata in gamma(delta) T cell knockout BALB/c mice inoculated with Mycobacterium avium subsp. paratuberculosis

The role of gamma(delta) T cells in the bovine immune response to Mycobacterium avium subsp. paratuberculosis (M. paratuberculosis) infection is poorly understood. Accordingly, using BALB/c mice that are innately susceptible to M. paratuberculosis, we compared wild-type and gamma(delta) T cell knockout BALB/c mice to study the protective roles of gamma(delta) T cells in M. paratuberculosis infection. Ten-week-old mice were inoculated intraperitoneally with either a low dose (4 x 10(6) colony-forming units [CFU]/mouse) or a high dose (4 x 10(9) CFU/mouse) of M. paratuberculosis strain ATCC 19698. Histopathologic and morphometric examinations showed reductions in the number and area of granulomatous lesions in the liver of the knockout mice at 18 weeks after inoculation with either the low or the high dose of the mycobacteria. Furthermore, at 18 weeks after inoculation, the bacterial load in the spleens of the knockout mice inoculated with the high dose was significantly lower than that of wild-type mice. No differences were found in bacterial load between the knockout and the wild-type mice in the low-dose groups. In contrast, in the livers of wild-type mice inoculated with either the low or high mycobacterial dose, increased areas of epithelioid granulomata were observed and the granulomata became disseminated widely during the experimental period. These findings in model mice suggest that gamma(delta) T cells, rather than restricting mycobacterial growth, may play a crucial role in development of epithelioid granulomata similar to those seen consistently in bovine paratuberculosis. The results of this study may have relevance to our understanding of the pathogenesis of paratuberculosis in ruminants, in which a prominent number of gamma(delta) T cells exist in the lymphoid system.     

Fundamentals of host immune response against Brucella abortus: what the mouse model has revealed about control of infection

The studies reviewed here evaluated the role cellular immune system components play in control of brucellosis by conducting comparative studies with brucella-resistant C57BL/10 or C57BL/6 mice and susceptible BALB/c mice. We have shown by both in vitro and in vivo studies that activation of macrophages with interferon-gamma (IFN-γ) is an important factor for control of infection with B. abortus in the mouse model and that the mechanism of anti-brucella activity largely involved reactive oxygen intermediates. Differences in control of the organism by resistant and susceptible mice was not related to inherent differences in the ability of their macrophages to control infection either with or without IFN-γ activation nor was it attributable to NK cells since we found no role for them in control of brucellosis in either mouse strain. However, relative resistance to brucellosis did correlate with increased production of IFN-γ by CD4 T cells during the first weeks after infection while IL-10 contributed to susceptibility in BALB/c mice. Moreover, by 3 weeks post-infection splenocytes from the susceptible BALB/c mice failed to produce IFN-γ and relied on TNF- as well as CD8 T cells to control infection until the end of the plateau phase around 6 weeks post-infection when IFN-γ production resumed and clearance began. In contrast, IFN-γ was crucial for control throughout the infection in the more resistant C57BL/6 mice and the mice died in its absence by 6 weeks post-infection compared to 12 weeks for the more susceptible mice that relied on additional mechanisms of control. In contrast to the IFN-γ knock-out mice, both β2 microglobulin knock-out C57BL/6 mice, which do not express conventional MHC class I molecules and thus cannot present antigen to CD8 T cells, or perforin knock-out C57BL/6 mice, which have no T cell cytotoxic activity, controlled and cleared the infection as well as normal C57BL/6 mice. The hiatus of IFN-γ production in BALB/c mice correlated with very high levels of total IL-12 and it was postulated that the lack of IFN-γ was a consequence of p40 homodimer blocking activity. However, reduction of p40 IL-12 in vivo through administration of indomethacin reduced the infection without a concomitant measurable increase in IFN-γ. Current studies are aimed at elucidating the mechanism of the IFN-γ hiatus.     

Comparative behaviour of Brucella abortus strains 19 and RB51 in the pregnant mouse.

Pregnant BALB/c mice received various doses of either Brucella abortus strain 19, a smooth vaccine strain, or B abortus strain RB51, a stable rough organism, intraperitoneally on day 9 of gestation to compare the relative pathogenicity of the two attenuated strains. Nine days after inoculation, spleens and placentas were collected for bacteriological and histopathological examination. A dose of 10(7.5) and strain 19 organisms produced a severe necrosuppurative placentitis occasionally accompanied by fetal death. This dose resulted in a 10-fold higher level of splenic infection than did a dose of 10(9.5) strain RB51 organisms, which produced only mild to minimal placentitis not associated with fetal death. Strain 19 infected mice showed seroconversion in the standard tube agglutination test in contrast to the seronegative titre of strain RB51 infected mice. The results of this study corroborate previous investigations on the relative pathogenicity and the serological response of the non-pregnant mouse to strain RB51.     

Bacteriologic and histologic features in mice after intranasal inoculation of Brucella melitensis

OBJECTIVE: To characterize effects of intranasal inoculation of virulent Brucella melitensis strain 16M in mice. ANIMALS: Female Balb/c mice, 6 to 8 weeks old. PROCEDURE: Studies were designed to elucidate gross morphologic lesions, bacterial burden in target organs, and histologic changes in tissues following experimental intranasal inoculation of mice with B melitensis 16M, which could be used to characterize a model for testing vaccine efficacy. RESULTS: Measurable splenomegaly was evident at 3 and 7 weeks after inoculation. A demonstrable increase in splenic colony-forming units (CFU) from infected mice increased over time with increasing dose when comparing inocula of 10(3), 10(4), and 10(5) CFU. Recovery of brucellae from the lungs was possible early in infection with 10(1), 10(3), and 10(5) CFU, but only the group inoculated with 10(5) CFU consistently yielded quantifiable bacteria. At a dose of 10 CFU, few organisms were located in the spleen. Bacteria were recovered up to 140 days after inoculation in mice given 10(3) CFU. At an inoculum of 10(5) CFU, bacterial counts were highest early in infection. Histologic examination of tissues revealed an increase in white pulp and marginal zone in the spleen and lymphohistiocytic hepatitis. CONCLUSION AND CLINICAL RELEVANCE: Changes in the spleen and liver increased with increases in dose and with increased time following intranasal inoculation with B melitensis 16M. Surprisingly, histologic changes were not observed in the lungs of inoculated mice.     

Viral replication and lesions in BALB/c mice experimentally inoculated with porcine circovirus isolated from a pig with postweaning multisystemic wasting disease

Eight-week-old BALB/c mice were either sham inoculated (control mice) or were inoculated intraperitoneally (IP) and intranasally (IN) with a single (sPCV mice) or multiple (mPCV mice) doses of porcine circovirus 2 (PCV2). Four control mice and 4 sPCV mice were sacrificed 7, 14, 28, and 42 days postinoculation (PI). All 4 mPCV mice were sacrificed 42 days PI. In addition, 7-day and 14-day pregnant BALB/c mice were either sham inoculated (control mice) or were inoculated IP and IN with a single dose of PCV2. Newborn mice were euthanatized 1, 8, and 15 days after birth. Necropsies were performed on all euthanatized mice and tissues were collected for histopathology, electron microscopy, in situ hybridization, and polymerase chain reaction (PCR). PCV2 replicated in 8-week-old BALB/c mice that were inoculated with PCV2 and caused fetal infection when inoculated into pregnant BALB/c mice at 7 days and 14 days of gestation. PCV was detected by in situ hybridization and PCR in sPCV mice on days 7, 14, 28, and 42 PI; in mPCV mice on day 42 PI; and in newborn mice from mothers inoculated with PCV at 7 days and 14 days of gestation at 1, 8, and 15 days after birth, but not in control mice. No clinical signs or gross lesions were found in sPCV or mPCV mice during the study. Microscopic lesions in sPCV mice and mPCV mice were characterized by expansion of germinal centers in lymphoid organs with large numbers of histiocytic cells and lymphoblasts, apoptosis of histiocytic cells in germinal centers, and mild lymphoid depletion of the paracortex. PCV nucleic acid was detected in the nuclei and cytoplasm of histiocytes and apoptotic cells in germinal centers in lymphoid tissues as well as in the nuclei of hepatocytes in the liver, in the nuclei of renal tubular epithelial cells, and in the cytoplasm of single lymphocytes in the thymus. Congenitally infected mice only had PCV nucleic acid detected in putative Kupffer cells in livers.     

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.     

Factors affecting the efficacy of ivermectin against Heligmosomoides polygyrus (Nematospiroides dubius) in mice

The efficacy of ivermectin against Heligmosomoides polygyrus was investigated in 5 mouse strains (CFLP, NIH, C57Bl10, BALB/c and CBA) using a variety of dose levels, and subcutaneous and oral administration. Heligmosomoides polygyrus were not completely eliminated when 5 mg kg-1 of ivermectin was given 6 days after infection, but 10 mg kg-1 was totally effective. Significant mouse-strain variation in drug efficacy was detected, NIH mice requiring treatment with higher doses than CFLP mice in order to bring about a comparable level of larvicidal activity. Ivermectin was more effective when given subcutaneously than when given orally, regardless of the dose administered or the strain of mouse tested. The anthelmintic effect of the treatment was more persistent in CFLP mice given 20 mg kg-1 subcutaneously than in NIH mice or in mice treated orally, and a 20-day interval between administration and infection was insufficient to prevent inhibition of parasite survival. Ivermectin was shown to be totally effective at 20 mg kg-1 given subcutaneously in killing immune arrested larvae of H. polygyrus.     

Duration of strain 2308 infection and immunogenicity of Brucella abortus lipopolysaccharide in five strains of mice

A study was conducted to compare immunogenicity of a Brucella abortus lipopolysaccharide (LPS) and the duration of infection in 5 strains of mice. Mice of strains CBA/NJ, BALB/c, CD-1, C3H/HeN, and C3H/HeJ were allotted into 2 large groups (vaccinated with proteinase K-treated LPS or nonvaccinated) and 6 subgroups based on the intervals between challenge exposure to B abortus strain 2308 and the week the response data were obtained. Criteria used in comparing responses between the various strains of mice as well as between vaccinated and nonvaccinated mice were splenomegaly, colony-forming units (CFU) from spleens, and antibody titers. Responses were evaluated at 1, 2, 3, 5, 8, and 12 weeks after challenge exposure. Results indicated that all strains of mice became infected and maintained infection throughout the 12-week period, the percentages of mice infected were significantly (P less than 0.05) less in vaccinated mice for the first 5 weeks after challenge exposure, and there were no direct correlations between increased immunoglobulins (IgM and IgG titers) and reduction in CFU. Vaccinated mice of strains BALB/c, CD-1, C3H/HeN, and C3H/HeJ had increased titers when challenge exposed and also had significantly (P less than 0.05) smaller spleens and lower CFU. Vaccinated CBA/NJ mice did not have marked antibody titers. The overall results indicated that vaccination with LPS offers some initial protection against B abortus strain 2308 infection, but this protection disappears gradually and in various degrees in the 5 strains of mice studied.     

Immunity to Babesia in mice. III. The effects of corticosteroids and anti-thymocyte serum on mice immune to Babesia rodhaini

BALB/c mice, immunized against Babesia rodhaini by an amicarbalide controlled infection, were exposed to selective immunosuppressive treatment with corticosteroids and anti-thymocyte serum (ATS) respectively. Hydrocortisone acetate, 100 mg/kg, given i.p. six times during the three weeks after challenge inoculation caused a rising parasitaemia and high mortality (6/7). Dexamethasone in the drinking water at 20 mg/l or 10 mg/l for 22 days had a similar suppressive effect on the protection against B. rodhaini. Mortality, 100% at the dose rate of 20 mg/l and 50% at 10 mg/l, occurred both in challenged and in carrier animals after the reappearance of parasites in the bloodstream. All the ATS-treated immune mice demonstrated parasitaemia after challenge, although at a lower level than did the corticosteroid treated mice. Seven out of 9 animals died. Corticosteroid-sensitive macrophages together with T-lymphocytes are considered to play an important role in protection against B. rodhaini in specifically induced immunity in mice.     

Effect of phytohaemagglutinin-P on apoptosis and necrosis in Trichinella spiralis infected mice

Flow cytometry analyses were used to evaluate the contribution of apoptotic and necrotic lymphocytes in the selected organs of Trichinella spiralis infected mice treated with phytohaemagglutinin-P (PHA-P). The Tunnel method was used to examine apoptosis in a cryostat section from the jejunum and masseter muscle. CFW mice (Groups I and II) were infected with 200 larvae of T. spiralis. PHA-P was administered intravenously at a dose of 10 mg/kg 24 h prior to infection in Group II mice only. Group III mice were treated with PHA-P without T. spiralis infection, and Group IV mice were untreated controls. The lymphocytes obtained from the spleen, mesenteric lymph nodes (MLN) and muscular inflammatory infiltration on 7, 14, 21, 28, 35, 42 and 60 days post infection (DPI) were incubated with the Annexin-V-Fluos Staining Kit (Roche). The cryostat preparation made from the jejunum and masseter muscle was evaluated using a fluorescence microscope. PHA-P administration stimulated apoptosis in the jejunal mucosa and in the muscular inflammatory infiltration. In Group I mice, infected with T. spiralis only, the highest percentage of apoptotic cells was found on 7 DPI in the spleen and in MLN, and on 14 DPI among the cells of the muscular inflammatory infiltration. The peak of the necrotic lymphocytes was found on 7 DPI in the spleen, on 28 DPI in MLN, and on 21 DPI in the cells of muscular inflammatory infiltration. In Group II mice, infected with T. spiralis and treated with PHA-P, the peak in apoptotic cells occurred on 7 DPI in the spleen and in the muscular inflammatory infiltration. The highest level of necrotic lymphocytes was observed only on 7 DPI in the muscular inflammatory infiltration. Percentage of necrotic lymphocytes in the spleen was the same and in MLN it was lower than in Group I (T. spiralis only). Moreover, the number of muscle larvae in mice treated with PHA-P (Group II) was lower than in Group I (T. spiralis only).     

Isolation,Identification and Pathogenicity on Mice of Streptococcus suis in Pig in Guangxi Border Area

16 strains of Streptococcus were isolated and identified from 370 pig tissues of clinically morbidity farms and slaughter houses in Guangxi border area.Morphological observation, PCR identification and pathogenicity test were carried out. The results showed that the morphology,dyeing,biochemical characteristics were coincidence with the characteristics of Streptococcus suis,confirmed that 16 strains were Streptococcus suis,and named as GX1 to GX16. Further by divided type identification (35 serotypes) by GDH,1 strains was type 1,1 strains was type 2,1 strains was type 5,1 strain was type 29,5 strains were type 8 and 7 strains could not differentiate types. The pathogenicity of 16 strains Streptococcus suis were detected by Kunming mice and BALB/c mice. The results showed that the letal rate of GX10 for Kunming mice and BALB/c mice was 100%;GX11 for BALB/c mice was 80%;Other strains were apathogenicity to the Kunming mice and BALB/c mice. The LD₅₀ of GX10 and GX11 was 4.5×10⁵ and 3.6×10⁹ CFU for BALB/c mice,respectively. The results showed that pathogenic Streptococcus suis type 8 occured in Guangxi border area. The sensitivity of BALB/c mice to Streptococcus suis was higher than Kunming mice, BALB/c mice was more suitable for the pathogenic study of Streptococcus suis.     

广西部分地区猪链球菌的分离鉴定及小鼠致病性研究

对来自广西边境地区部分发病猪场及屠宰场的370份猪组织进行细菌分离鉴定,共分离到16株链球菌,对这16株链球菌进行了形态学观察、PCR鉴定及小鼠致病性试验。结果表明,其形态、染色、生化特性均符合猪链球菌的特点,通过GDH鉴定与测序,证实这16株菌株均为猪链球菌,分别命名为GX1~GX16。通过特异性引物PCR分型（35个血清型）,其中有1株为1型,1株为2型,1株为5型,1株为29型,5株为8型,7株未分出型。用这16株猪链球菌对昆明小鼠和BALB/c小鼠进行动物致病性试验,结果发现,菌株GX10对昆明小鼠和BALB/c小鼠致死率均为100%;菌株GX11对BALB/c小鼠致死率为80%;其他菌株对昆明小鼠和BALB/c小鼠均无致病性。菌株GX10、GX11对BALB/c小鼠的LD₅₀分别为4.5×10⁵和3.6×10⁹ CFU。本试验结果表明,广西边境地区存在致病性猪链球菌8型。BALB/c小鼠对猪链球菌的敏感性高于昆明小鼠,更适合作为猪链球菌致病性研究的动物模型。     

A fatty acid binding protein from Fasciola hepatica induced protection in C57/BL mice from challenge infection with Schistosoma bovis.

Three strains of mice (NMRI, C57/BL, BALB/c) were each immunized with a 12 kDa purified, native Fasciola hepatica fatty acid binding protein (Fh12) and challenged percutaneously with Schistosoma bovis cercariae. C57/BL mice immunized with Fh12 had significant reductions in S. bovis worm burden recoveries (96 and 87% reductions over controls in two separate experiments). When using NMRI or BALB/c mice, Fh12 alone or in Freund's adjuvant failed to induce significant protection against S. bovis. In C57/BL mice vaccinated against Fh 12, antibodies to the IgG2a isotype, but not to the IgG1 isotype, increased by 2 weeks after the second immunization and remained high through 8 weeks of S. bovis infection. Antibodies to S. bovis increased after 4 weeks of infection. Regarding cytokine production by spleen mononuclear cells, C57/BL mice vaccinated with Fh12 in adjuvant, and having the highest protective response against challenge infection with S. bovis, had an increase of IFN-gamma production with Concanavalin A but no increase of IL-4 in similarly stimulated cells. These results suggest that the protection obtained in this group of mice is mediated by a Th1 immune response.     

Effects of social isolation,re‐socialization and age on cognitive and aggressive behaviors of Kunming mice and BALB/c mice

Both Kunming (KM) mice and BALB/c mice have been widely used as rodent models to investigate stress‐associated mental diseases. However, little is known about the different behaviors of KM mice and BALB/c mice after social isolation, particularly cognitive and aggressive behaviors. In this study, the behaviors of KM and BALB/c mice isolated for 2, 4 and 8 weeks and age‐matched controls were evaluated using object recognition, object location and resident‐intruder tests. The recovery of behavioral deficits by re‐socialization was also examined for the isolated mice in adolescence. Our study showed that isolation for 2, 4 and 8 weeks led to cognitive deficits and increased aggressiveness for both KM and BALB/c mice. An important finding is that re‐socialization could completely recover spatial/non‐spatial cognitive deficits resulted from social isolation for both KM and BALB/c mice. In addition, age only impacted aggressiveness of KM mice. Moreover, isolation duration showed different impacts on cognitive and aggressive behaviors for both KM and BALB/c mice. Furthermore, BALB/c mice showed weak spatial/non‐spatial memory and low aggressiveness when they were at the same age and isolation duration, compared to KM mice. In conclusion, KM mice and BALB/c mice behaved characteristically under physiology and isolation conditions.     

牛源犬新孢子虫孕鼠感染模型的建立

为建立牛源犬新孢子虫孕鼠感染模型,深入研究牛源犬新孢子虫对孕鼠的致病作用,本试验以雌性BALB/c小鼠为试验动物,分离Vero细胞中培养的牛源犬新孢子虫速殖子,分不同剂量组腹腔接种雌性BALB/c小鼠后,与雄性BALB/c小鼠合笼,每天观察小鼠临床症状和发病情况,观察主要脏器组织的病理变化,应用PCR方法检测孕鼠脑、肝脏、脾脏等脏器组织及胎盘中犬新孢子虫Nc5基因,并测定孕鼠胎盘湿重和胎盘系数。结果显示,感染模型小鼠的最佳攻虫剂量为10⁵个虫体;感染犬新孢子虫孕鼠先后出现精神不振、共济失调等临床症状,并有不同程度死亡;病理学观察模型小鼠脑、肝脏、脾脏等脏器组织出现充血、出血、肿大等病理变化;在模型小鼠脑、肝脏、脾脏等脏器组织及胎盘中检测到犬新孢子虫Nc5基因;随攻虫天数的增加,模型小鼠胎盘重量和胎鼠重量均不断增加,胎盘系数逐步降低,在第12、14、16天时,模型组与对照组相比,胎盘重量和胎盘系数均差异显著（P < 0.05）。本试验成功建立了牛源犬新孢子虫孕鼠感染模型,为犬新孢子虫致病机制研究奠定了基础。     

Mycobacterium paratuberculosis monoassociated nude mice as a paratuberculosis model

In this study, a paratuberculosis (Johne's disease) model was developed by intragastrically dosing gnotobiotic athymic nude mice with Mycobacterium paratuberculosis. The mice infrequently shed bacilli from their intestinal tracts during the first 4 months after inoculation. Following this time, increasing numbers of M. paratuberculosis (greater than 4.0 log10 bacilli per fecal pellet by 40 weeks) were recovered from the feces of the 12 mice that remained in the isolator. A similar pattern of recovery of M. paratuberculosis was obtained from the ileum, cecum, colon, and liver. Histopathologic lesions and acid-fast bacilli were rare during the first 4 months of infection and then, with time, increased in prevalence and severity. Mice maintained for 7 months or longer exhibited severe granulomatous inflammation and large numbers of acid-fast bacilli in the gastrointestinal tract and liver (up to 10(8) log10 colony forming units per gram wet weight). Five mice maintained for 7 months or more developed clinical signs consistent with those seen in paratuberculosis (weight loss, chronic diarrhea); three of these mice eventually died or became moribund and were euthanatized. M. paratuberculosis monoassociated mice released increased levels of tumor necrosis factor activity into their sera, as compared to uninfected control mice, when they were injected with bacterial lipopolysaccharide. The clinical signs, fecal shedding of M. paratuberculosis, granuloma formation, and progressive bacillary multiplication observed with these mice are consistent with naturally occurring M. paratuberculosis infection of ruminants (Johne's disease). This model will be useful for future studies of immunoregulation and antimicrobial therapy of paratuberculosis.     

Vertical transmission of Neospora caninum in BALB/c mice in both acute and chronic infection

To examine the frequency of congenital infection by Neospora caninum, BALB/c mice were inoculated intraperitoneally with tachyzoites of N. caninum either during pregnancy (Group 1) or 4 weeks or more before pregnancy (Group 2). Further, the mice inoculated during pregnancy were bred at 4 weeks or more after delivery to form Group 3. Congenital transmission was observed in 76% of the neonates of the mice in Group 1 and in 50% of the neonates of the mice in Group 2. Interestingly, congenital transmission was observed in 86% of the neonates from Group 3. These results suggest that chronically-infected BALB/c mice efficiently transmit N. caninum infection to their offspring.     

Oral vaccination of brushtail possums (Tichosurus vulpecula) with BCG: immune responses, persistence of BCG in lymphoid organs and excretion in faeces

AIMS: To determine immune responses, and the localisation and persistence of Mycobacterium bovis bacille Calmette-Guérin (BCG) in gut-associated lymphoid tissues (GALT) and other organs in possums vaccinated orally with lipid-formulated BCG vaccine. To determine the duration of excretion and longevity of survival of BCG in the faeces of vaccinated animals. METHODS: Possums (n=28) were vaccinated with lipid-formulated BCG (1 x 10(8) colony forming units (cfu) of formulated BCG) by the oral route. Control possums (n=17) were fed oral bait pellets containing formulation medium only. Possums were sacrificed at 3 days and at 1, 3, 6 and 8 weeks after vaccination or ingestion of bait. Proliferation responses to bovine purified protein derivative (PPD) were measured in lymphocytes from blood and mesenteric lymph nodes (MLN) and samples of lung, spleen, liver, MLN and Peyer's patches (PP) were cultured for the presence of BCG. The number of BCG organisms excreted in faeces and the duration of excretion were determined in eight vaccinated possums and eight control possums over a 3-week period. In a separate experiment, a further six possums were vaccinated with oral BCG vaccine (5-10 x 10(8) cfu BCG/possum) and their faeces collected over 48-72 h, for culture of BCG. The longevity of survival of BCG in these faeces was determined by storing faecal samples (n=12) under three different conditions: in an incubator (22.5 degrees C), and conditions which simulated the forest floor and open pasture. A proportion (1-2 g) of these faecal samples was collected after storage for 1, 3, 5, 8 or 20 weeks, and cultured for BCG. RESULTS: Possums vaccinated orally with BCG vaccine showed strong proliferation responses to bovine PPD in peripheral blood lymphocytes at 6-8 weeks post-vaccination (p.v.). Positive lymphocyte proliferation assay (LPA) responses to bovine PPD were first evident in MLN at 3 weeks p.v. BCG was cultured from MLN and PP in a proportion of animals at 3-8 weeks p.v. BCG was not cultured from sections of spleen, lung or liver at any time p.v. BCG was recovered in low to moderate numbers from the faeces of vaccinated possums for up to 7 days, and maximal numbers were cultured in faeces collected 48-72 h p.v. After storage for 1 week, BCG was cultured from all faecal samples placed in the incubator and from a proportion of faeces exposed to conditions similar to those on the forest floor and pasture. With the exception of one faecal sample stored under forest floor conditions which was culture-positive for BCG at 3 and 5 weeks, BCG was not cultured from any other faecal sample stored for more than 1 week. CONCLUSIONS: Ingestion of oral BCG vaccine by possums was associated with the development of strong cell-mediated immunity in both blood and MLN. Following oral vaccination with BCG, the organisms were localised and persisted in GALT but did not spread to the spleen, liver or lungs. BCG was shed in low to moderate numbers in the faeces for up to 7 days p.v. The viability of BCG excreted in faeces decreased rapidly, particularly when faeces were exposed to an open pasture environment. Oral vaccination of possums with formulated BCG is unlikely to result in undue contamination of the environment with BCG.