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1.
Primary culture of bovine brain cells was examined for its susceptibility to Neospora caninum infections, and this model was used to investigate the effects of bovine interferon gamma (IFN-gamma) and tumor necrosis factors alpha (TNF-alpha) on tachyzoite growth. Tachyzoites of N. caninum grew well in this culture, and tachyzoite growth in astroglia and microglia were confirmed by immunocytochemical staining. IFN-gamma inhibited the tachyzoite growth, and this inhibition was not reversed by the addition of nitric oxide antagonist. TNF-alpha, to a lesser extent, also inhibited the tachyzoite growth. Th-1 type cytokines may play an important role in host defense mechanisms in N. caninum infection.  相似文献   

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3.
Neospora caninum is an intracellular apicomplexan parasite that infects a wide range of mammals and has been associated with abortion in cattle worldwide. Artemisinin is an effective antimalarial compound derived from a traditional Chinese herbal remedy, qinghao or Artemisia annua L. In the study reported, the cultured host cells (vero cells or mouse peritoneal macrophages) infected with N. caninum tachyzoites were incubated with alpha-MEM (minimal essential medium) 10%HS supplemented with various concentration or artemisinin (20, 10, 1, 0.1 and 0.01 microg/ml) to examine the efficacy of artemisinin against N. caninum tachyzoites intracellular multiplication. In long-term studies, at 20 or 10 microg/ml for 11 days, artemisinin reduced N. caninum and completely eliminated all microscopic foci of N. caninum. At 1 microg/ml for 14 days, artemisinin reduced N. caninum and completely achieved elimination of all microscopic foci of N. caninum. There was no apparent toxicity to host cells in long-term studies. In short-term studies, at > or = 0.1microg/ml, artemisinin reduced N. caninum tachyzoites intracellular multiplication, significantly (P < 0.05) and appeared to depend on the artemisinin concentrations. Pretreatment of host cells or N. caninum tachyzoites with artemisinin had no effect on N. caninum tachyzoites intracellular multiplication. These results demonstrate that artemisinin inhibited N. caninum tachyzoites intracellular multiplication.  相似文献   

4.
A suitable balance in the production of Th1/Th2-type cytokines has a crucial role in the control of microbial infections. We investigated cytokine production patterns and effects during Neospora caninum infection, based on two mouse models and an in vitro system. In the acute infection of N. caninum, BALB/c-background IFN-gamma-deficient mice that were sensitive to the N. caninum infection showed high levels of IL-10 production, whereas significant levels of interferon-gamma (IFN-gamma) and interleukin-4 (IL-4) production were observed in resistant wild type mice. BALB/c mice vaccinated with recombinant vaccinia virus expressing N. caninum surface protein NcSRS2 resisted parasite spread throughout the body, low levels of IFN-gamma production and high levels of IL-4 production were observed compared to unvaccinated animals. The treatment of N. caninum-infected cells with IFN-gamma or IL-10 decreased the host-cell viability in an in vitro system using mouse macrophage J774A.1 cells. On the other hand, IL-4, but not IL-10 administration, increased the viability of N. caninum-infected and IFN-gamma-treated cells. In the light of the balance of Th1/Th2-type cytokine production, an IFN-gamma/IL-4 balance may have a crucial role for the control of cellular responses against the parasite invasion.  相似文献   

5.
Neospora caninum is an apicomplexan parasite associated with abortion in cattle worldwide. Apicidin, histone deacetylase (HDAC) inhibitor, has shown a broad spectrum of anti-protozoal activity against apicomplexan parasites. Cultured vero cells infected with N. caninum tachyzoites were treated with 0.5 microg/ml of depudecin, another known natural product isolated from Altermaria brassiciicola with anti-histone deacetylase activity, to examine the efficacy of depudecin against intracellular multiplication of N. caninum tachyzoite, using 15 ng/ml of apicidin as the control. Depudecin significantly inhibited the intracellular multiplication of N. caninum at a level similar to that of apicidin without exerting any cytotoxicity on the host vero cells. Acid/urea/Triton gel electrophoresis analysis show anti-parasitic activity of depudecin appears to be due to the inhibition of protozoal histone deacetylase, which induces hyperacetylation of histones in N. caninum.  相似文献   

6.
Neospora caninum, a strictly intracellular protozoan, is a major leading cause of parasite-induced abortion in cattle. A widely held view of N. caninum infection is that both cellular proliferation and stage interconversion (tachyzoite-bradyzoite transformation) are triggered, perhaps even modulated by, changes in cultural conditions. This study tested the hypothesis that exposure of N. caninum tachyzoites to different pH culture media affects the parasite's entry, proliferation and cyst formation in cultured cells. The endocytic pathway for N. caninum entry into the K562 cell line was found to be mediated by low pH of culture medium. Internalization of N. caninum by host cells was significantly increased in acidic and alkaline culture medium compared to cells maintained in neutral medium as revealed by transmission electron microscopy. Parasite proliferation within Vero cells was assessed by plaque formation assay and was found to be highest when pH level was optimum, paralleled by a decrease in the number of cysts. In contrast, parasite encystation increased when the pH level was alkaline or acidic, as evaluated by indirect immunofluorescence and immunocytochemical analyses. Acidic pH regardless of state of host cell infection suppressed the rate of host cell division. These findings suggest that culture medium pH has a determinable effect on the host cell-N. caninum interaction and support the hypothesis that pH of culture medium influence the entry, growth, and phenotypic plasticity of N. caninum in mammalian cells.  相似文献   

7.
Bats are considered to be natural reservoirs for several viruses of clinical importance, including rabies virus, Nipah virus, and Hendra virus. Type I interferons (IFNs) is an important part of the immune system in the defense against viral infection. To investigate the function of type I IFNs upon viral infection in bats, the nucleic acid, and amino acid sequences of Egyptian Rousette (Rousettus aegyptiacus) IFN-alpha and -beta were characterized. Sequence data indicated that bat IFN-alpha consists of 562-bp encoded 187-aa, and IFN-beta consisted of 558-bp encoded 186-aa. Phylogenetic analysis of the overall identity of IFN-beta shared the highest sequence homology with pig IFN-beta in both nucleotide and amino acid level. Stimulation of bat primary kidney cells (BPKCs) and bat lung cell lines, Tb-1 Lu, with polyinosinic-polycytidylic acid (poly(I:C)) or exogenous bat type I IFNs resulted in increased type I IFNs mRNA expression in BPKCs, but not in Tb-1 Lu. Characterization of the bat IFN-alpha and -beta genes allows understanding of the immune responses upon stimulation in different tissues, thus providing practical strategies for control and treatment of clinically important diseases. These results are important especially for the virus infection, and suggest that future molecular studies on virus infection experiment of bats in vitro will require careful consideration of the differences of type I IFN expression patterns in different cell types.  相似文献   

8.
The direct effects of three steroid hormones (progesterone, estradiol-17beta and corticosterone) on the growth of Neospora caninum (N. caninum) tachyzoite were examined in Vero cells. Subsequently, ovariectomized BALB/c mice infected with N. caninum were treated with physiological concentrations of the steroid hormones for 1 or 2 weeks. These hormones had no direct effect on the parasite growth in vitro. In the infected mice, there was no significant difference in the parasite distribution and histopathological changes between the hormone-injected and control groups. No mice showed parasitemia at the time of autopsy. These results suggest that physiological levels of steroid hormones (progesterone, estradiol-17beta and corticosterone) do not reactivate N. caninum in mice.  相似文献   

9.
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.  相似文献   

10.
Neospora caninum infection provokes neurological disorders, recurrent abortion and death in dogs and cattle. Dogs are both intermediate and definitive host of N. caninum. Thus, the development of sensitive and specific immunoassays to diagnose canine neosporosis is essential to control this disease. This work investigated serum anti-neosporal IgG and IgE antibodies in 140 dogs represented by 30 healthy animals (group I), 11 dogs showing acute N. caninum infection (group II), 50 urban dogs with serological evidence of canine neosporosis in indirect fluorescent antibody test (IFAT) (group III) and 49 urban dogs without clinical and laboratory evidences of neosporosis (group IV). Enzyme-linked immunosorbent assay (ELISA) and western immunoblotting, both using a soluble N. caninum tachyzoite antigen (SNA), investigated these two isotypes of antibodies, while a Urea-ELISA measured the avidity of the IgG antibodies. Anti-Toxoplasma gondii IgG antibodies were also investigated in the animals. Anti-neosporal IgG was found in all animals from groups II and III, whereas 32.7% (16/49) of dogs from group IV were reactive. IgG antibodies of low avidity were demonstrated in dogs from group II (median 35.3%), while animals from groups III and IV had IgG antibodies of high avidity (medians of 61.5% and 61.7% respectively). IgE antibodies were found in four (13.3%) and five (16.6%) dogs from groups III and IV respectively. Dogs presenting acute infection (group II) or chronic infection (group III) had IgG antibodies to several neosporal antigens, mainly of 29-30 and 35 kDa, while 13 of 16 dogs from group IV recognized antigens from 14 to 170 kDa. Antibodies to T. gondii were detected in 36 of 50 (72%) sera from group III and 25 of 49 (51%) sera from group IV. We concluded that IgG-ELISA and Urea-ELISA with SNA may substitute for IFAT in both laboratory routine and epidemiological studies of canine neosporosis.  相似文献   

11.
Apart from their veterinary importance, the hookworms Ancylostoma caninum, Ancylostoma braziliense and Ancylostoma caninum are also capable of causing zoonotic disease in humans. A highly sensitive and species-specific PCR-RFLP technique was utilised to detect and differentiate the various canine Ancylostoma spp directly from eggs in faeces. This technique was utilised to screen 101 canine faecal samples from parasite endemic tea growing communities in Assam, India, as part as an ongoing epidemiological investigation into canine parasitic zoonoses. The prevalence of hookworms in dogs was found to be 98% using a combination of PCR and conventional microscopy. Overall, 36% of dogs were found positive for single hookworm infections with A. caninum, 24% positive for single infections with A. braziliense and 38% had mixed infections with both A. caninum and A. braziliense. No dogs were found positive for A. ceylanicum in the community under study. The high prevalence of A. caninum and A. braziliense in dogs in this community may account for the high incidence of cutaneous larva migrans (CLM) observed among the human population residing at the tea estates. The PCR-RFLP technique described herein allows epidemiological screening of canine hookworms to be conducted rapidly, with ease and accuracy, and has the potential to be applied to a number of different clinical, pharmacological and epidemiological situations.  相似文献   

12.
Here, we studied the potential of Neospora caninum tachyzoites to infect heifers when administered in utero by artificial insemination via contaminated semen. Eighteen primiparous cyclic heifers were hormonally synchronized and artificially inseminated. Nine of them, which were inseminated with semen containing 10(7) live N. caninum NC-1 isolate-tachyzoites, reacted with seroconversion and a specific IFN-gamma response. Moreover, N. caninum DNA was demonstrated by a nested-PCR in the blood of all nine heifers and in brain, lungs, liver and uterine horn of several of them. In contrast, nine heifers inseminated with tachyzoite-free semen developed no antibody or IFN-gamma responses, and no parasite DNA was detected in blood or organs. At necropsy, viable embryos were detected in one and six of the infected and non-infected heifers, respectively. No specific Neospora DNA was detected in any of the embryos. This study provides evidence that intrauterine inoculation via contaminated semen cause N. caninum infection in cattle.  相似文献   

13.
We have previously shown that resistance to Leishmania infantum in dogs is associated with a Th1 type of immune response. In this study, we use a canine macrophage cell line (030-D) that can readily be infected with this protozoan parasite. Our aim is to further characterize the effector mechanisms involved in killing of Leishmania parasite in dogs. We observed that activation of 030-D cells by incubation with a supernatant derived from a Leishmania-specific T cell line containing IFN-gamma, TNF-alpha and interleukin-2 (IL-2) resulted in enhanced nitric oxide (NO) production by these cells. In addition, we observed enhanced anti-leishmanial activity of infected 030-cells after activation. Both, NO production and anti-leishmanial activity were abrogated by addition of L-N(G)-nitroargininemethyl ester (L-NAME), an analogue of L-arginine. Thus, NO play an important role in the anti-leishmanial activity of these canine macrophages. We propose the infection of the 030-D cell line as a good in vitro model to further investigate parasite-host cell interactions in dogs, a natural host of Leishmania parasites.  相似文献   

14.
Interferon-alpha (IFN-alpha) is a type I interferon that is secreted during the early stages of the innate immune response and is often induced upon infection with viral pathogens. IFN-alpha production affects multiple downstream events influencing both innate and adaptive immune responses. Here, we describe the expression of an equine rIFN-alpha/IgG4 fusion protein in mammalian cells. The anti-viral activity of rIFN-alpha/IgG4 was found to be 70-fold higher than that of a previously described IFN-gamma/IgG1 as tested by bioassay. The purified rIFN-alpha was subsequently used for the generation of six monoclonal antibodies (mAbs) to equine IFN-alpha. Four of these mAbs inhibited the protective anti-viral effect of equine leukocyte IFN in bioassays. One mAb (clone 240-2) showed a high-neutralizing capacity. An ELISA was established using two anti-equine IFN-alpha mAbs (clones 29B and 240-2) and its analytical sensitivity for was found to be around 800 pg/ml and 3 U/ml for rIFN-alpha and equine leukocyte IFN, respectively. When analyzing samples with a likely dominance of IFN-alpha among type I IFNs, such as supernatants from equine peripheral blood mononuclear cells stimulated with CpG-oligodeoxyribonucleotides, the results obtained by ELISA and IFN bioassay showed a high agreement (r(2)(sp)=0.98). When analyzing samples likely containing a mixture of type I IFNs, such as serum and nasal secretions from virally infected horses, the ELISA only detected some of the IFN-activity recorded in the bioassay. Overall, the data showed that the new anti-equine IFN-alpha mAbs are valuable tools to detect native IFN-alpha for further characterization of the early innate immune response and anti-viral immunity in horses.  相似文献   

15.
INTRODUCTION: Neospora caninum is the most frequently diagnosed cause of cattle abortion in New Zealand and is an important pathogen worldwide. The parasite has been found in aborted bovine foetuses and in puppies with neurological disease. Recent developments have provided new insights into the epidemiology of the disease that are reviewed in this article. TRANSMISSION: Transplacental infection is of major importance in the spread of N. caninum overseas. Most congenital infections result in the birth of apparently healthy calves. Seroepidemiological studies indicate introduced point-source infections are also likely, as do investigations of abortion outbreaks in cattle herds attributed to N. caninum. Horizontal transmission is an important determinant of the stability of infection prevalence in a herd. Two potential sources of horizontal transmission that have recently been reported are: via a canine definitive host and; cow-to-calf transmission through pooled colostrum or milk. Transmission parameters for the organism have been estimated using mathematical modeling, to explore the epidemiology and options for control of N. caninum infection in dairy cattle in the absence of precise epidemiological data. LIFE-CYCLE: Seroepidemiologic studies, combined with the knowledge that dogs can be a definitive host, provide evidence supporting a dog-cattle life-cycle. The importance of dogs in the epidemiology of bovine neosporosis is not yet clear. It is likely that N. caninum oocysts in dog faeces could serve as a source of infection for cattle and recent studies have shown that the canine and bovine isolatesflare the same organism. Infection of cattle orally exposed to oocysts has been demonstrated experimentally but not in nature. It is not known if other wild carnivores may serve as definitive hosts of N. caninum. CONCLUSION: Despite active research for over a decade, very little is known about the mechanisms of transmission of N. caninum. Vertical transmission is well documented but horizontal spread and the pathogenesis of abortion from this disease need further investigation. It is evident that, even with a very high probability of vertical transmission, some form of horizontal transmission is required for the disease to persist in a herd.  相似文献   

16.
Attempts were made to isolate Neospora caninum from naturally infected white-tailed deer (Odocoileus virginianus). A total of 110 deer killed during the 2003 hunting season in Virginia region were used for the isolation of N. caninum. Of these, brains from 28 deer that had NAT titer of 1:200 were inoculated into interferon-gamma gene knock out (KO) mice. N. caninum was isolated from the tissues of three deer and all three isolates were mildly virulent to KO mice. Only one of the isolates could be adapted to in vitro growth. Protozoa in the tissues of KO mice reacted with N. caninum-specific polyclonal antibodies and N. caninum DNA was demonstrated in infected tissues by PCR assays; sequences of portions of the ITS-1 and gene 5 loci were identical to those in the public database. This is the first record of in vitro isolation of N. caninum from white-tailed deer and lends credence to the white-tailed deer as an intermediate host for this parasite.  相似文献   

17.
Neospora caninum is an intracellular protozoan parasite that was first recognized in dogs in 1988. N. caninum may cause neuromuscular disease in dogs. Later, it was discovered that N. caninum has a wide host range and is an important cause of abortion in cattle. In this article, the literature on N. caninum in the dog is reviewed, with emphasis on clinical signs, pathology, diagnosis, treatment, and prognosis.  相似文献   

18.
Neosporosis by Neospora caninum causes losses to livestock production through abortion in cattle while, in dogs, it induces neuromuscolar disease. This study investigated neosporosis seroprevalence associated risk factors (including the seropositivity to leishmaniosis) in dogs of southern Italy, determined the prevalence of N. caninum oocyst shedding and examined the relationship between seroprevalence of neosporosis in farm dogs and cattle. Using an inhibition ELISA, 20.9% of 306 dogs had percent inhibition values >10 (indicative of exposure) and farm dogs had a significantly (p<0.001) higher seroprevalence than dogs in a rescue kennel. Whilst N. caninum seroprevalence was associated with increasing age in dogs (p< or =0.01) there was no association between seropositivity for N. caninum and for Leishmania infantum. Oocysts of N. caninum were not detected in faecal samples from 230 dogs including 160 farm dogs. The results indicated that neosporosis infection is common in southern Italy both in dogs and in cattle and that dogs at higher risk of exposure are free-ranging ones living in farms. The lack of correlation between canine seroprevalence for N. caninum and L. infantum assumes a particular significance in an endemic area for leishmaniosis.  相似文献   

19.
Neospora caninum is a canine parasite which is considered a significant cause of bovine abortion. Two cattle herd groups were serologically studied with the objective of studying the prevalence of infection by N. caninum associated with BHV1 and BVDV infections. In group I, 15 dairy herds (476 samples) naturally infected by the three infectious agents were analyzed,. In group II, three dairy herds (100 samples) of cows vaccinated for two viruses were analyzed, in order to determine the infection prevalence by N. caninum. In the first group, an infection prevalence of 12.61, 34 and 28.3% was determined for N. caninum BHV1 and BVDV, respectively. In the second group, a seropositive prevalence of 46, 85 and 76%, respectively, was determined for N. caninum, BVH1 and BVDV. In the first group, the virus and N. caninum had shown in the first group 4.41% positive samples in association with BVH1, 3.15% with BVDV, and 8.41% with BVH1 and BVDV.  相似文献   

20.
Neospora caninum was isolated from the brain of an aborted 4-month-old fetus from a dairy cow herd with endemic neosporosis in Porto, Portugal. The fetal brain homogenate was inoculated interperitoneally first into outbred Swiss Webster mice given dexamethasone and then the peritoneal exudates from these mice was co-inoculated with mouse sarcoma cells in the peritoneal cavity of mice given dexamethasone. N. caninum tachyzoites were seen in peritoneal exudate of the second passage. Tachyzoites from the peritoneal exudate reacted positively with anti-N. caninum antibodies and not with anti-Toxoplasma gondii antibodies and contained N. caninum specific DNA. This Portuguese isolate of N. caninum has been successfully maintained in cell culture. The dam of the aborted fetus had an antibody titer of 1:10240 in the Neospora agglutination test (NAT). Antibodies to N. caninum were found in 76 of 106 cows from this herd in titers of 1:40 in 31, 1:80 in 22, > or =1:160 or more in 23 in the Neospora agglutination test. This is the first isolation of a viable N. caninum-like parasite from any host in Portugal.  相似文献   

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