-
[show abstract]
[hide abstract]
ABSTRACT: Classical serine proteases use the conserved Ser/His/Asp catalytic triad to hydrolyze substrates. Here, we show that longistatin, a salivary gland protein with two EF-hand domains from the vector tick Haemaphysalis longicornis, does not have the conserved catalytic triad, but still functions as a serine protease. Longistatin was synthesized in and secreted from the salivary glands of ticks, and is injected into host tissues during the acquisition of blood-meals. Longistatin hydrolyzed fibrinogen, an essential plasma protein in the coagulation cascade, and activated plasminogen, into its active form plasmin, a serine protease that dissolves fibrin clots. Longistatin efficiently hydrolyzed several serine protease-specific substrates showing its specificity to the amide bond of Arg. Longistatin did not hydrolyze synthetic substrates specific for other groups of proteases. The enzyme was active at a wide range of temperatures and pHs, with the optimum at 37°C and pH 7. Its activity was efficiently inhibited by various serine protease inhibitors such as phenylmethanesulfonyl fluoride (PMSF), aprotinin, antipain, and leupeptin with the estimated IC(50) of 278.57 μM, 0.35 μM, 41.56 μM and 198.86 μM, respectively. In addition, longistatin was also potently inhibited by Zinc (Zn(2+)) in a concentration-dependent manner with an IC(50) value of 275 μM, and the inhibitory effect of Zn(2+) was revived by ethylenediaminetetra acetic acid (EDTA). Immunization studies revealed that longistatin sharply induced high levels of protective IgG antibodies against ticks. Immunization with longistatin reduced repletion of ticks by about 54%, post engorgement body weight by >11% and molting of nymphs by approximately 34%; thus, the vaccination trial was approximately 73% effective against tick infestation. Taken together, our results suggest that longistatin is a new potent atypical serine protease, and may be an interesting candidate for the development of anti-tick vaccines.
Molecular and Biochemical Parasitology 12/2011; 182(1-2):45-53. · 2.55 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Thrombo-occlusive diseases are major causes of morbidity and mortality, and tissue-type plasminogen activator (t-PA) is recommended for the treatment of the maladies. However, both t-PA and u-PA are rapidly inactivated by plasminogen activator inhibitor-1 (PAI-1). Here, we show that longistatin, a novel plasminogen activator isolated from the ixodid tick, Haemaphysalis longicornis is resistant to PAI-1. Longistatin was relatively less susceptible to the inhibitory effect of SDS-treated platelet lysate than physiologic PAs. Platelet lysate inhibited t-PA and tcu-PA with the IC(50) of 7.7 and 9.1 μg/ml, respectively, whereas for longistatin inhibition IC(50) was 20.1 μg/ml (p<0.01). Similarly, activated PAI-1 (20 nM) inhibited only 21.47% activity of longistatin but almost completely inhibited t-PA (99.17%) and tcu-PA (96.84%). Interestingly, longistatin retained 76.73% initial activity even after 3h of incubation with 20 nM of PAI-1. IC(50) of PAI-1 during longistatin inhibition was 88.3 nM while it was 3.9 and 3.2 nM in t-PA and tcu-PA inhibition, respectively. Longistatin completely hydrolyzed fibrin clot by activating plasminogen efficiently in the presence of 20 nM of PAI-1. Importantly, unlike t-PA, longistatin did not form complex with PAI-1. Collectively, our results suggest that longistatin is resistant to PAI-1 and maybe an interesting tool for the development of a PAI-1 resistant effective thrombolytic agent.
Biochemical and Biophysical Research Communications 09/2011; 413(4):599-604. · 2.48 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Ixodid ticks are notorious blood-sucking ectoparasites and are completely dependent on blood-meals from hosts. In addition to the direct severe effects on health and productivity, ixodid ticks transmit various deadly diseases to humans and animals. Unlike rapidly feeding vessel-feeder hematophagous insects, the hard ticks feed on hosts for a long time (5-10 days or more), making a large blood pool beneath the skin. Tick's salivary glands produce a vast array of bio-molecules that modulate their complex and persistent feeding processes. However, the specific molecule that functions in the development and maintenance of a blood pool is yet to be identified. Recently, we have reported on longistatin, a 17.8-kDa protein with two functional EF-hand Ca(++)-binding domains, from the salivary glands of the disease vector, Haemaphysalis longicornis, that has been shown to be linked to blood-feeding processes. Here, we show that longistatin plays vital roles in the formation of a blood pool and in the acquisition of blood-meals. Data clearly revealed that post-transcriptional silencing of the longistatin-specific gene disrupted ticks' unique ability to create a blood pool, and they consequently failed to feed and replete on blood-meals from hosts. Longistatin completely hydrolyzed α, β and γ chains of fibrinogen and delayed fibrin clot formation. Longistatin was able to bind with fibrin meshwork, and activated fibrin clot-bound plasminogen into its active form plasmin, as comparable to that of tissue-type plasminogen activator (t-PA), and induced lysis of fibrin clot and platelet-rich thrombi. Plasminogen activation potentiality of longistatin was increased up to 4 times by soluble fibrin. Taken together, our results suggest that longistatin may exert potent functions both as a plasminogen activator and as an anticoagulant in the complex scenario of blood pool formation; the latter is critical to the feeding success and survival of ixodid ticks.
PLoS Pathogens 03/2011; 7(3):e1001312. · 9.13 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Human indigenous cutaneous leishmaniasis caused by Leishmania donovani complex is endemic in Sri Lanka. We performed an entomological survey to determine the distribution of probable vector species. Sand flies were collected in districts in the dry zone, in the wet zone highlands, and in the wet zone coastal belt of Sri Lanka using CDC light traps, sticky traps and cattle-baited net traps during July, 2005. The survey was reconducted in February, 2006. Overall, 584 sand flies belonging to Phlebotomus (266 specimens, 2 species) and Sergentomyia (318 specimens, 8 species) genera were collected. A total of 266 Phlebotomus was identified as P. argentipes (258/266; 97%) and P. stantoni (8/266; 3%) . The identification studies of Sergentomyia specimens showed that there are at least 8 species in Sri Lanka. Higher number of Phlebotomus sand flies (76/266) were caught in the southern part of the country compared to the other parts probably due to different ecological aspects. P. argentipes were widely distributed throughout the island whereas P. stantoni were collected only in four districts. Since P. argentipes is known to be the vector of L. donovani responsible of visceral leishmaniasis in India, this species may be incriminated as the most possible vector of human cutaneous leishmaniasis in Sri Lanka.
Journal of Vector Ecology 03/2011; 36 Suppl 1:S77-86. · 0.88 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Human indigenous cutaneous leishmaniasis caused by Leishmania donovani complex is endemic in Sri Lanka. We performed an entomological survey to determine the distribution of probable vector species. Sand flies were collected in districts in the dry zone, in the wet zone highlands, and in the wet zone coastal belt of Sri Lanka using CDC light traps, sticky traps and cattle-baited net traps during July, 2005. The survey was reconducted in February, 2006. Overall, 584 sand flies belonging to Phlebotomus (266 specimens, 2 species) and Sergentomyia (318 specimens, 8 species) genera were collected. A total of 266 Phlebotomus was identified as P. argentipes (258/266; 97%) and P. stantoni (8/266; 3%) . The identification studies of Sergentomyia specimens showed that there are at least 8 species in Sri Lanka. Higher number of Phlebotomus sand flies (76/266) were caught in the southern part of the country compared to the other parts probably due to different ecological aspects. P. argentipes were widely distributed throughout the island whereas P. stantoni were collected only in four districts. Since P. argentipes is known to be the vector of L. donovani responsible of visceral leishmaniasis in India, this species may be incriminated as the most possible vector of human cutaneous leishmaniasis in Sri Lanka.
Journal of Vector Ecology. 02/2011; 36(s1):S77 - S86.
-
[show abstract]
[hide abstract]
ABSTRACT: To investigate the usefulness of the immunopotentiator from Pantoea agglomerans 1 (IP-PA1) as a supportive drug in melanoma therapy, we analyzed the immunological effects of IP-PA1 on melanoma-inoculated model mice. Oral administration of IP-PA1 increased the serum levels of tumor necrosis factor (TNF)-α at 2 h after the administration and interferon (IFN)-γ and IL-12 at 12 h after the administration in naïve BALB/cCrSlc mice as evaluated by ELISA. IP-PA1 did not affect the proliferation of melanoma cells directly determined by the 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) assay. Combinatory treatment of IP-PA1 with doxorubicin for 9 days increased the serum levels of IFN-γ and IL-12 by 71.0 and 15.3%, respectively, compared to the treatment of doxorubicin alone in melanoma-bearing C57BL/6NCrSlc mice as evaluated by ELISA. It also increased the proportion of natural killer (NK) cells and the ratio of CD4(+) to CD8(+) T cells in the spleen from 6.1 ± 0.3 to 7.4 ± 0.5% and from 1.25 ± 0.03 to 1.38 ± 0.04, respectively, compared to the treatment of doxorubicin alone as analyzed by flow cytometry. The mean survival period of melanoma-bearing, doxorubicin treated mice was prolonged from 31.4 ± 7.1 to 35.3 ± 8.4, 51.1 ± 5.4, and 45.0 ± 8.4 days by combinatory treatment of IP-PA1 at the daily doses of 0.1, 0.5, and 1 mg/kg, respectively. In conclusion, the results of the present study suggest the usefulness of IP-PA1 as a supportive drug in melanoma therapy.
Experimental Animals 01/2011; 60(2):101-9. · 0.92 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Visceral leishmaniasis (VL), also called kala-azar (KA), is the most severe form of leishmaniasis and often fatal if not treated.
The causative species of VL are members of Leishmania donovani sensu lato (s.l.), which includes L. donovani, L. infantum, L. archibaldi, and L. chagasi. Its principal victims are children and young adults. Control of KA is urgently needed worldwide. However, due to the diversity
of the epidemiological features in each endemic focus, it has not been possible to devise a universal control strategy applicable
to all foci. In order to construct a framework of epidemiological, pathological, and taxonomic understanding, and explain
how Leishmania species develop different clinical and epidemiological features, further studies on the pathogenesis of different forms of
KA and comparative studies of the parasites from different foci are clearly necessary. This chapter summarizes the recent
advances in understanding the polymorphism of leishmaniasis as caused by Leishmania donovani s.l. in Asia.
KeywordsCutaneous leishmaniasis-Immunopathology-Kala-azar-
Leishmania donovani
-Post-kala-azar dermal leishmaniasis-Visceral leishmaniasis
12/2010: pages 101-110;
-
[show abstract]
[hide abstract]
ABSTRACT: Immunopotentiator from Pantoea agglomerans 1 (IP-PA1), an edible lipopolysaccharide (LPS) derived from symbiotic bacteria in crops, is a promising immunomodulator. It activates macrophages and protects from chemotherapeutic agent-induced growth inhibition in macrophages in vitro. We showed the immune-recovery effects of IP-PA1 in a chicken model of dexamethasone-induced stress in which IP-PA1 inhibited thymic and bursal atrophy and improved antibody production in response to vaccination. Furthermore, we showed IP-PA1 improved survival of melanoma-bearing, doxorubicin-treated mice, although not directly affecting the proliferation of melanoma cells, dominantly through the improvement of host antitumor immunity. These results suggest that IP-PA1 could have other possible applications in the treatment of various immunosuppression-related disorders in humans and animals.
Anticancer research 08/2010; 30(8):3113-8. · 1.73 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: The immunopotentiator from Pantoea agglomerans 1 (IP-PA1) is an edible lipopolysaccharide (LPS) derived from symbiotic bacteria found in crops. IP-PA1 is known to ameliorate chemotherapy-induced immunosuppression; therefore, its macrophage-activating effect in the presence of chemotherapeutic agents was evaluated.
Nuclear factor-kappaB (NF-kappaB) activation in IP-PA1-treated RAW264 and J774.1 cells was examined using Western blot analyses; Griess assay and ELISA were used to examine the production of nitric oxide and tumour necrosis factor alpha, respectively. The expression of apoptosis-related proteins was also assessed using Western blot analyses. The effect of IP-PA1 on doxorubicin-induced apoptosis was analyzed by flow cytometry after annexin-V staining. The growth of macrophages treated with chemotherapeutic agents and IP-PA1 was analyzed using an MTT assay.
IP-PA1 activated NF-kappaB and ameliorated chemotherapy induced growth inhibition in the cells.
IP-PA1 is an edible drug that can potentially support chemotherapy by ameliorating chemotherapy-induced immunosuppression.
Anticancer research 06/2010; 30(6):2033-40. · 1.73 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Considering the usefulness of the immunopotentiator from Pantoea agglomerans 1 (IP-PA1), which is a purified lipopolysaccharide (LPS) derived from symbiotic gram-negative bacteria of food crops, in controlling immunosuppression in poultry husbandry, in this study, we examined its immune-recovery effects in dexamethasone-treated stressed chicken models. Three-week-old chickens daily administered 10 microg/kg of dexamethasone for 35 days to induce stress showed more whole body weight loss; relative thymic, bursal, and splenic weight losses; and decrease in the number of peripheral blood lymphocytes, as compared with the control chickens on day 35; the IP-PA1-pretreated, dexamethasone-treated chickens showed reduced weight losses. Five- to eight-week-old chickens administered 5 mg/kg of dexamethasone showed excessive apoptosis of thymic and bursal lymphocytes 24 hr after a single dexamethasone treatment; apoptosis was inhibited in the IP-PA1-pretreated, dexamethasone-treated chickens. Chickens daily administered 10 microg/kg of dexamethasone for 35 days and injected with commercial Salmonella Enteritidis (SE) vaccine or sheep red blood cells (SRBC) on days 7 and 21 showed about 8- or 2-fold lower antibody production in response to SE or SRBC, respectively, as compared with the control chickens on day 35; the antibody production in response to SE or SRBC was increased in the IP-PA1-pretreated, dexamethasone-treated chickens. These results indicate that IP-PA1 exerts inhibitory effects on dexamethasone-induced immunosuppression and that it may be useful in controlling immunosuppression in poultry husbandry.
Journal of Veterinary Medical Science 04/2010; 72(4):435-42. · 0.85 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: The mucosal vaccination is a non-invasive alternative approach for not only mucosal pathogens but also parenteral pathogens, since it induces both mucosal and systemic immunoreactions. The purpose of this study was to evaluate the application of intranasal (i.n.) immunization with a recombinant leishmanial protein against Leishmania infection. BALB/c mice were i.n. administered 1-3 times with Leish-111f plus cholera toxin (CT) adjuvant (Leish-111f/CT). Splenocytes from i.n. immunized mice produced high level of IFN-gamma but not IL-4 in response to Leish-111f. When infected with 1x10(6) of Leishmania major promastigotes 2 weeks after the final administration, lesion development was completely controlled in all mice i.n. administered with Leish-111f/CT. Mice i.n. administered with Leish-111f alone showed neither cytokine productions nor lesion control even after 6 administrations, suggesting the importance of CT adjuvant. This report demonstrated for the first time that i.n. administration of a recombinant leishmanial protein induces Th1 type immunity and protects mice from Leishmania infection.
Vaccine 03/2010; 28(10):2207-13. · 3.77 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Calcium and the EF-hand Ca(++)-binding proteins have been undisputedly recognised as the key players in almost all aspect of cell functions, starting from the cell's birth, during mitosis to its end with apoptosis. But in a few exceptional cases the EF-hand proteins are secreted from the cells and play their crucial roles extracellularly. Here, to our knowledge for the first time, we have identified and characterised an EF-hand Ca(++)-binding protein from the salivary glands of the ixodid tick, Haemaphysalis longicornis, herein called longistatin. Longistatin possesses two EF-hand domains which conserve canonical structure and bind with Ca(++). Both the recombinant and endogenous proteins were stained with Rutheninum red. Reverse-transcription PCR data showed that longistatin-specific transcript was expressed in all life-cycle stages of H. longicornis and was up-regulated only in blood-fed ticks. Organ-specific transcription analysis revealed a salivary gland-specific expression of the gene which peaked at 96-120 h of feeding when ticks acquired full blood-meals and become engorged but its expression declined sharply as they detached and dropped off the host. Consistently, endogenous protein was localised in the salivary glands of adult ticks and in the lumen of the functional acini of the salivary glands. Furthermore, longistatin was detected in feeding lesions at the site of attachment of ticks on the host. These results suggest that longistatin is synthesised in, and is secreted from, the salivary glands and may have functional roles in the feeding process of ixodid ticks.
International journal for parasitology 12/2009; 40(6):721-9. · 3.39 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: This study was performed to isolate a velogenic Newcastle disease virus (NDV) strain currently found in Indonesia for establishing a domestic reference virus for future pathological and molecular epidemiological studies. A chicken suspected to have contracted Newcastle disease (ND) in a local outbreak in Bali was selected for NDV isolation. Atrophy of lymphoid tissues such as the bursa of Fabricius, thymus, and spleen; intestinal haemorrhage; and oedema of the brain were observed in the chicken. Histopathological examination revealed severe non-suppurative meningoencephalomyelitis characterised by neuronal necrosis, multifocal to diffuse gliosis, and perivascular cuffing of mononuclear cells, hemorrhagic necrosis of the trachea, intestines and bursa of Fabricius, and various degree of lymphoid depletion and necrosis of the lymphoid tissues. After ND was confirmed immunohistochemically, the NDV was propagated by inoculating tissue homogenate of the diseased chicken in embryonated eggs. Phylogenetic analysis based on the F gene nucleotide sequence revealed that this isolate belonged to genotype VII. The deduced amino acid sequence of the isolated NDV F protein at the cleavage site was (112)RRQKRF(117), which is typically found in virulent NDV isolates. Pathogenicity indexes such as the mean death time (MDT) and intracerebral pathogenicity index (ICPI) were 54 hr and 1.77, respectively. Pathological findings, phylogenetic analysis, amino acid sequence of the F protein cleavage site, and pathogenicity index test results revealed the NDV isolate, designated as NDV/Bali-1/07, to be a novel Indonesian velogenic NDV strain belonging to group VII.
Journal of Veterinary Medical Science 12/2009; 72(3):313-9. · 0.85 Impact Factor
-
Takeshi Arakawa,
Mayumi Tachibana,
Takeshi Miyata,
Tetsuya Harakuni,
Hideyasu Kohama, Yasunobu Matsumoto,
Naotoshi Tsuji,
Hajime Hisaeda,
Anthony Stowers,
Motomi Torii,
Takafumi Tsuboi
[show abstract]
[hide abstract]
ABSTRACT: Malaria vaccines based on ookinete surface proteins (OSPs) of the malaria parasites block oocyst development in feeding mosquitoes and hence disrupt the parasite life cycle and prevent the disease from being transmitted to other individuals. To investigate whether a noninvasive mucosal vaccination regimen effectively blocks parasite transmission in vivo, Plasmodium yoelii Pys25, a homolog of the Pfs25 and Pvs25 OSPs of Plasmodium falciparum and Plasmodium vivax, respectively, was intranasally (i.n.) administered using a complement-deficient DBA/2 mouse malaria infection model, in which a highly elevated level of oocysts develops in feeding mosquitoes. Vaccinated mice developed a robust antibody response when the vaccine antigen was given together with cholera toxin adjuvant. The induced immune serum was passively transferred to DBA/2 mice 3 days after infection with P. yoelii 17XL, and Anopheles stephensi mosquitoes were allowed to feed on the infected mice before or after serum transfusion. This passive immunization completely blocked oocyst development; however, immune serum induced by the antigen or adjuvant alone did not have such a profound antiparasite effect. Further, when i.n. vaccinated mice were infected with the parasite and then mosquitoes were allowed to directly feed on the infected mice, complete blockage of transmission was again observed. To our knowledge, this is the first time that mucosal vaccination has been demonstrated to be efficacious for directly preventing parasite transmission from vaccinated animals to mosquitoes, and the results may provide important insight into rational design of nonparenteral vaccines for use against human malaria.
Infection and immunity 09/2009; 77(12):5496-500. · 4.21 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Plants are attractive vaccine production and oral delivery systems. Cereals are excellent candidate for edible vaccines, which can express and store high levels of proteins for extended periods of time without degradation. In this study, we produced a 14-kDa protective surface antigen of Ascaris suum L3 larvae and its fusion chimera with a mucosal carrier molecule cholera toxin B subunit (CTB) in rice (Oryza sativa L.) under the control of the endosperm-specific glutelin-B promoter. We found that the recombinant protein expression levels reached 1.5 microg per seed, a comparably high amount as compared to previously reported transgenic rice expression experiments. Potentials of transgenic rice plants as a source of oral vaccines against swine roundworm are discussed.
Journal of Veterinary Medical Science 08/2009; 71(7):995-1000. · 0.85 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: To investigate the potential applicability of mucosal vaccines against mucosa-unrelated pathogens, a non-parenteral vaccination approach was taken as a prophylactic strategy against mosquito-borne Japanese encephalitis virus (JEV). Intranasal (i.n.) immunization with a mouse brain-derived formalin-inactivated JE vaccine induced a robust virus-neutralizing antibody in mice, and this induction was augmented by co-administration with cholera toxin (CT) and pertussis toxin, but not with killed Bordetella pertussis. The antibody response induced by the i.n. administration of the JE vaccine with bacterial toxins was comparable in intensity to that induced by a parenteral immunization regime, and the former was considerably more effective in terms of delayed-type hypersensitivity and local antibody response. In addition, the adjuvant effects of bacterial toxins were much more prominent for the mucosal than the parenteral route. Two other non-invasive routes, oral and transcutaneous administration, were examined, but the i.n. route was by far the most effective. Finally, the vaccine efficacy of a chimeric fusion protein between the B subunit of CT and the JEV envelope protein showed some promise for the development of non-invasive JE vaccine. Our results suggest that the mucosal vaccination approach is feasible for a non-mucosal pathogen such as JEV, but that the adjuvant, carrier molecule, and administration route must be optimized for construction of an effective vaccine platform.
Japanese journal of infectious diseases 02/2009; 62(1):37-45. · 1.49 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Cereal crops such as maize and rice are considered attractive for vaccine production and oral delivery. Here, we evaluated the rice Oryza sativa for production of As16-an antigen protective against the roundworm Ascaris suum. The antigen was produced as a chimeric protein fused with cholera toxin B subunit (CTB), and its expression level in the endosperm reached 50 microg/g seed. Feeding the transgenic (Tg) rice seeds to mice elicited an As16-specific serum antibody response when administered in combination with cholera toxin (CT) as the mucosal adjuvant. Although omitting the adjuvant from the vaccine formulation resulted in failure to develop the specific immune response, subcutaneous booster immunization with bacterially expressed As16 induced the antibody response, indicating priming capability of the Tg rice. Tg rice/CT-fed mice orally administered A. suum eggs had a lower lung worm burden than control mice. This suggests that the rice-delivered antigen functions as a prophylactic edible vaccine for controlling parasitic infection in animals.
Transgenic Research 10/2008; 18(2):185-92. · 2.75 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Interferon (IFN)-gamma is essential but not sufficient to control leishmaniasis. It is known that IFN-gamma is one of the major macrophage-activating cytokines, and the activated macrophages are a principal source of interleukin (IL)-12, which induces autocrine macrophage activation. In this study, the combined effect of IFN-gamma and IL-12 on the susceptibility of macrophages to Leishmania major infection was evaluated. Macrophages pretreated with IFN-gamma and/or IL-12 were infected with the parasites. Four hr post-infection (p.i.), the levels of infection and parasite load in the macrophages treated with the combination of IFN-gamma and IL-12 (IFN-gamma/IL-12) were significantly lower than those in the nontreated cells. However, the macrophages treated with either IFN-gamma or IL-12 did not show resistance to L. major infection. In addition, 72 hr p.i., the IFN-gamma/IL-12-treated and IFN-gamma-treated macrophages showed significantly lower levels of infection and parasite load than the nontreated cells, and higher levels of resistance was observed in the IFN-gamma/IL-12-treated macrophages than in the IFN-gamma-treated macrophages. Although IFN-gamma/IL-12 treatment of macrophages prior to the infection led to the induction of resistance, as described above, this resistance was not induced when these cytokines and the parasites were added simultaneously to the macrophage culture. These results suggest that IFN-gamma/IL-12 treatment prior to the infection restricts the early phase of the infection.
Journal of Veterinary Medical Science 06/2008; 70(6):589-93. · 0.85 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: The mucosal immunization method is a needle-free alternative way of vaccination. This study evaluated the efficacy of mucosal immunization for rabies. Mice were intranasally administered five times with inactivated and concentrated rabies virus antigen (CRV) supplemented with or without cholera toxin (CT). The anti-rabies virus antibody titer of mice intranasally immunized with CRV plus CT (CRV/CT) was comparable to that of mice intraperitoneally immunized twice with the same amount of CRV. Virus neutralizing (VNA) titers of mice immunized intranasally with CRV/CT were slightly lower than those of intraperitoneally immunized mice. Both anti-rabies virus ELISA antibody and VNA titers of mice immunized with CRV without CT were significantly lower than those of mice immunized with CRV/CT. In mice intranasally immunized with CRV/CT, and intraperitoneally immunized mice, high levels of IgG(2a) antibody were detected, suggesting the activation of Th1-driven cellular immunity by the two ways of immunization. All immunized mice were challenged intracerebrally with a lethal dose of virulent rabies virus CVS strain. The survival rates of mice immunized with CRV/CT and CRV without CT were 67% and 17%, respectively, while the rate of intraperitoneally immunized mice was 100%. Antigen-specific whole IgG and IgG(2a), and VNA titers of survived mice were significantly higher than those of dead mice at the challenge day. These data suggest the possibility of intranasal immunization with inactivated antigen as a rabies vaccination strategy and the importance of a mucosal adjuvant such as CT.
Experimental Animals 02/2008; 57(1):1-9. · 0.92 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: A previous study revealed that antibodies to hepatitis E virus (HEV) (anti-HEV) are highly prevalent among healthy individuals and farm pigs in Bali, Indonesia, and suggested that HEV infection may occur via zoonosis among Balinese people. However, there were no reports of acute hepatitis E in Bali. To elucidate whether Balinese HEV strains recovered from infected humans and pigs have significant sequence similarity, serum samples obtained from 57 patients (age, mean +/- standard deviation, 31.1 +/- 11.9 years) with sporadic acute hepatitis and from one hundred and one 2- or 3-month-old farm pigs in Bali were tested for anti-HEV and HEV RNA. Among the 57 patients, 2 (3.5%) had high-titer IgM/IgA class anti-HEV antibodies and one of them had detectable HEV RNA (BaliE03-46). Overall, 58 pigs (57.4%) tested positive for anti-HEV, while 5 pigs (5.0%) had detectable HEV RNA. Based on the 412-nucleotide sequence within open reading frame 2, the BaliE03-46 isolate and the 5 swine HEV isolates recovered from the viremic pigs were phylogenetically classified in genotype 4, but were only 77.3-90.8% identical to the genotype 4 HEV isolates reported thus far in China, India, Japan, Taiwan, and Vietnam. The BaliE03-46 isolate of human origin shared high identities of 97.3-98.3% with 4 of the 5 Balinese swine isolates, but differed by 16.1% from the remaining swine isolate. These results suggest that indigenous HEV strains of genotype 4 with marked heterogeneity are circulating in Bali, Indonesia, and that pigs are reservoirs of HEV for Balinese people who have a habit of ingesting uncooked pigs.
Journal of Medical Virology 09/2007; 79(8):1138-46. · 2.82 Impact Factor
