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Kozo Fujisaki - One of the best experts on this subject based on the ideXlab platform.
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glutathione s transferases play a role in the detoxification of flumethrin and chlorpyrifos in Haemaphysalis longicornis
Parasites & Vectors, 2018Co-Authors: Kodai Kusakisako, Remil Linggatong Galay, Emmanuel Pacia Hernandez, Melbourne Rio Talactac, Kozo Fujisaki, Takeshi Hatta, Naotoshi TsujiAbstract:Haemaphysalis longicornis is a tick of importance to health, as it serves as a vector of several pathogens, including Theileria orientalis, Babesia ovata, Rickettsia japonica and the severe fever with thrombocytopenia syndrome virus (SFTSV). Presently, the major method of control for this tick is the use of chemical acaricides. The glutathione S-transferase (GST) system is one mechanism through which the tick metabolizes these acaricides. Two GSTs from H. longicornis (HlGST and HlGST2) have been previously identified. Enzyme kinetic studies were performed to determine the interaction of acaricides with recombinant H. longicornis GSTs. Recombinant HlGST activity was inhibited by flumethrin and cypermethrin, while recombinant HlGST2 activity was inhibited by chlorpyrifos and cypermethrin. Using real-time RT-PCR, the upregulation of the HlGST gene was observed upon exposure to sublethal doses of flumethrin, while the HlGST2 gene was upregulated when exposed to sublethal doses of chlorpyrifos. Sex and strain dependencies in the induction of GST gene expression by flumethrin were also observed. Knockdown of the HlGST gene resulted in the increased susceptibility of larvae and adult male ticks to sublethal doses of flumethrin and the susceptibility of larvae against sublethal doses of chlorpyrifos was increased upon knockdown of HlGST2. HlGST could be vital for the metabolism of flumethrin in larvae and adult male ticks, while HlGST2 is important in the detoxification of chlorpyrifos in larval ticks.
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Glutathione S-transferases play a role in the detoxification of flumethrin and chlorpyrifos in Haemaphysalis longicornis
Parasites & Vectors, 2018Co-Authors: Emmanuel Pacia Hernandez, Kodai Kusakisako, Remil Linggatong Galay, Melbourne Rio Talactac, Naotoshi Tsuji, Kozo Fujisaki, Takeshi Hatta, Tetsuya TanakaAbstract:Background Haemaphysalis longicornis is a tick of importance to health, as it serves as a vector of several pathogens, including Theileria orientalis , Babesia ovata , Rickettsia japonica and the severe fever with thrombocytopenia syndrome virus (SFTSV). Presently, the major method of control for this tick is the use of chemical acaricides. The glutathione S-transferase (GST) system is one mechanism through which the tick metabolizes these acaricides. Two GSTs from H. longicornis (HlGST and HlGST2) have been previously identified. Results Enzyme kinetic studies were performed to determine the interaction of acaricides with recombinant H. longicornis GSTs. Recombinant HlGST activity was inhibited by flumethrin and cypermethrin, while recombinant HlGST2 activity was inhibited by chlorpyrifos and cypermethrin. Using real-time RT-PCR, the upregulation of the HlGST gene was observed upon exposure to sublethal doses of flumethrin, while the HlGST2 gene was upregulated when exposed to sublethal doses of chlorpyrifos. Sex and strain dependencies in the induction of GST gene expression by flumethrin were also observed. Knockdown of the HlGST gene resulted in the increased susceptibility of larvae and adult male ticks to sublethal doses of flumethrin and the susceptibility of larvae against sublethal doses of chlorpyrifos was increased upon knockdown of HlGST2 . Conclusions HlGST could be vital for the metabolism of flumethrin in larvae and adult male ticks, while HlGST2 is important in the detoxification of chlorpyrifos in larval ticks.
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Vector competence of Haemaphysalis longicornis ticks for a Japanese isolate of the Thogoto virus
Scientific Reports, 2018Co-Authors: Melbourne Rio Talactac, Kentaro Yoshii, Kodai Kusakisako, Remil Linggatong Galay, Emmanuel Pacia Hernandez, Masami Mochizuki, Kozo Fujisaki, Tetsuya TanakaAbstract:Thogoto virus (THOV), a tick-borne arbovirus not previously reported in East Asia, was recently isolated from Haemaphysalis longicornis in Kyoto, Japan. In this study, we investigated the vector competence of H. longicornis ticks for a Japanese isolate of the Thogoto virus using anal pore microinjection and experimental virus acquisition. Our results showed that anal pore microinjection can readily infect adult ticks, and THOV-infected ticks can successfully transmit the virus to mice. Blood feeding was also critical in the distribution of the virus in tick organs, most especially in the salivary glands. Furthermore, co-feeding between an infected adult and naïve nymphs can also produce infected molted adults that can horizontally transmit THOV to mice. Altogether, our results suggest that H. longicornis is a competent vector for the Japanese THOV isolate and could be the primary tick vector of the virus in Japan.
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the development of oocytes in the ovary of a parthenogenetic tick Haemaphysalis longicornis
Parasitology International, 2018Co-Authors: Ryo Mihara, Kozo Fujisaki, Rika Umemiyashirafuji, Yasuyuki Abe, Tomohide Matsuo, Noriyuki Horiuchi, Suguru Kawano, Hiroshi SuzukiAbstract:Haemaphysalis longicornis is an important vector of various pathogens in domestic animals and humans. The tick is a unique species with bisexual and parthenogenetic races. Although mating induces oocyte development, it is possible in the parthenogenetic race to complete oogenesis without copulation. Here we examined the developmental process of oocytes from unfed to the oviposition period in parthenogenetic H. longicornis. We classified the developmental stages of oocytes into five stages: stage I, germinal vesicle occupies more than half of the cytoplasm; stage II, germinal vesicle occupies less than half of the cytoplasm; stage III, germinal vesicle migrates from the center in the oocyte to the vicinity of the pedicel cells; stage IV, the cytoplasm is filled with yolk granules of various sizes; stage V, the cytoplasm is occupied by large yolk granules. Oocytes at the unfed period were undeveloped and classified as stage I. Stage I and II oocytes were observed at the rapid feeding period, indicating that oocyte development began after the initiation of blood feeding. All developmental stages of oocytes were observed at the pre-oviposition period. At 10 days after the beginning of the oviposition period, the ratios of stage I and II oocytes were higher than those of the previous period, suggesting that the ovarian development and activity may be continuing. Based on these findings, we propose classification criteria for the oocyte development in the parthenogenetic H. longicornis. The criteria will be useful for understanding the mechanisms of tick reproduction and transovarial transmission of pathogens.
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Additional file 2: Figure S2. of Characterization and expression analysis of a newly identified glutathione S-transferase of the hard tick Haemaphysalis longicornis during blood-feeding
2018Co-Authors: Emmanuel Hernandez, Kodai Kusakisako, Remil Linggatong Galay, Melbourne Rio Talactac, Naotoshi Tsuji, Kozo Fujisaki, Takeshi Hatta, Tomohide Matsuo, Tetsuya TanakaAbstract:Nucleotide and deduced amino acid sequences of HlGST (a) and HlGST2 (b) of Haemaphysalis longicornis. Start and stop codons are underlined. Predicted glutathione and substrate binding sites are shaded in black and gray, respectively. The putative polyadenylation signal, AATAAA, is double underlined. (PDF 53 kb
Naotoshi Tsuji - One of the best experts on this subject based on the ideXlab platform.
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glutathione s transferases play a role in the detoxification of flumethrin and chlorpyrifos in Haemaphysalis longicornis
Parasites & Vectors, 2018Co-Authors: Kodai Kusakisako, Remil Linggatong Galay, Emmanuel Pacia Hernandez, Melbourne Rio Talactac, Kozo Fujisaki, Takeshi Hatta, Naotoshi TsujiAbstract:Haemaphysalis longicornis is a tick of importance to health, as it serves as a vector of several pathogens, including Theileria orientalis, Babesia ovata, Rickettsia japonica and the severe fever with thrombocytopenia syndrome virus (SFTSV). Presently, the major method of control for this tick is the use of chemical acaricides. The glutathione S-transferase (GST) system is one mechanism through which the tick metabolizes these acaricides. Two GSTs from H. longicornis (HlGST and HlGST2) have been previously identified. Enzyme kinetic studies were performed to determine the interaction of acaricides with recombinant H. longicornis GSTs. Recombinant HlGST activity was inhibited by flumethrin and cypermethrin, while recombinant HlGST2 activity was inhibited by chlorpyrifos and cypermethrin. Using real-time RT-PCR, the upregulation of the HlGST gene was observed upon exposure to sublethal doses of flumethrin, while the HlGST2 gene was upregulated when exposed to sublethal doses of chlorpyrifos. Sex and strain dependencies in the induction of GST gene expression by flumethrin were also observed. Knockdown of the HlGST gene resulted in the increased susceptibility of larvae and adult male ticks to sublethal doses of flumethrin and the susceptibility of larvae against sublethal doses of chlorpyrifos was increased upon knockdown of HlGST2. HlGST could be vital for the metabolism of flumethrin in larvae and adult male ticks, while HlGST2 is important in the detoxification of chlorpyrifos in larval ticks.
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Glutathione S-transferases play a role in the detoxification of flumethrin and chlorpyrifos in Haemaphysalis longicornis
Parasites & Vectors, 2018Co-Authors: Emmanuel Pacia Hernandez, Kodai Kusakisako, Remil Linggatong Galay, Melbourne Rio Talactac, Naotoshi Tsuji, Kozo Fujisaki, Takeshi Hatta, Tetsuya TanakaAbstract:Background Haemaphysalis longicornis is a tick of importance to health, as it serves as a vector of several pathogens, including Theileria orientalis , Babesia ovata , Rickettsia japonica and the severe fever with thrombocytopenia syndrome virus (SFTSV). Presently, the major method of control for this tick is the use of chemical acaricides. The glutathione S-transferase (GST) system is one mechanism through which the tick metabolizes these acaricides. Two GSTs from H. longicornis (HlGST and HlGST2) have been previously identified. Results Enzyme kinetic studies were performed to determine the interaction of acaricides with recombinant H. longicornis GSTs. Recombinant HlGST activity was inhibited by flumethrin and cypermethrin, while recombinant HlGST2 activity was inhibited by chlorpyrifos and cypermethrin. Using real-time RT-PCR, the upregulation of the HlGST gene was observed upon exposure to sublethal doses of flumethrin, while the HlGST2 gene was upregulated when exposed to sublethal doses of chlorpyrifos. Sex and strain dependencies in the induction of GST gene expression by flumethrin were also observed. Knockdown of the HlGST gene resulted in the increased susceptibility of larvae and adult male ticks to sublethal doses of flumethrin and the susceptibility of larvae against sublethal doses of chlorpyrifos was increased upon knockdown of HlGST2 . Conclusions HlGST could be vital for the metabolism of flumethrin in larvae and adult male ticks, while HlGST2 is important in the detoxification of chlorpyrifos in larval ticks.
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Additional file 2: Figure S2. of Characterization and expression analysis of a newly identified glutathione S-transferase of the hard tick Haemaphysalis longicornis during blood-feeding
2018Co-Authors: Emmanuel Hernandez, Kodai Kusakisako, Remil Linggatong Galay, Melbourne Rio Talactac, Naotoshi Tsuji, Kozo Fujisaki, Takeshi Hatta, Tomohide Matsuo, Tetsuya TanakaAbstract:Nucleotide and deduced amino acid sequences of HlGST (a) and HlGST2 (b) of Haemaphysalis longicornis. Start and stop codons are underlined. Predicted glutathione and substrate binding sites are shaded in black and gray, respectively. The putative polyadenylation signal, AATAAA, is double underlined. (PDF 53 kb
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Virucidal activity of Haemaphysalis longicornis longicin P4 peptide against tick-borne encephalitis virus surrogate Langat virus
Parasites & Vectors, 2016Co-Authors: Melbourne Rio Talactac, Kentaro Yoshii, Kodai Kusakisako, Remil Linggatong Galay, Emmanuel Pacia Hernandez, Naotoshi Tsuji, Kozo Fujisaki, Hiroki Maeda, Tetsuya Tanaka, Masami MochizukiAbstract:Background Longicin is a defensin-like peptide, identified from the midgut epithelium of hard tick Haemaphysalis longicornis. Several studies have already shown the antimicrobial and parasiticidal activities of longicin peptide and one of its synthetic partial analogs, longicin P4. In this study, longicin peptides were tested for potential antiviral activity against Langat virus (LGTV), a tick-borne flavivirus.
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parasiticidal activity of Haemaphysalis longicornis longicin p4 peptide against toxoplasma gondii
Peptides, 2012Co-Authors: Tetsuya Tanaka, Naotoshi Tsuji, Hiroki Maeda, Xuenan Xuan, Rika Umemiyashirafuji, Tomohide Matsuo, Hiroshi Suzuki, Damdinsuren Boldbattar, Aiko Kume, Kozo FujisakiAbstract:The Haemaphysalis longicornis longicin P4 peptide is an active part peptide produced by longicin which displays bactericidal activity against both Gram-negative and Gram-positive bacteria and other microorganisms. In the present study, the effect of the longicin P4 peptide on the infectivity of Toxoplasma gondii parasites was examined in vitro. Tachyzoites of T. gondii incubated with longicin P4 had induced aggregation and lost the trypan blue dye exclusion activity and the invasion ability into the mouse embryonal cell line (NIH/3T3). Longicin P4 bound to T. gondii tachyzoites, as demonstrated by fluoresce microscopic analysis. An electron microscopic analysis and a fluorescence propidium iodide exclusion assay of tachyzoites exposed to longicin P4 revealed pore formation in the cellular membrane, membrane disorganization, and hollowing as well as cytoplasmic vacuolization. The number of tachyzoites proliferated in mouse macrophage cell line (J774A.1) was significantly decreased by incubation with longicin P4. These findings suggested that longicin P4 conceivably impaired parasite membranes, leading to the destruction of Toxoplasma parasites in J774A.1 cells. Thus, longicin P4 is an interesting candidate for antitoxoplasmosis drug design that causes severe toxicity to T. gondii and plays an important role in reducing cellular infection. This is the first report showing that longicin P4 causes aggregation and membrane injury of parasites, leading to Toxoplasma tachyzoite destruction.
Tetsuya Tanaka - One of the best experts on this subject based on the ideXlab platform.
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Glutathione S-transferases play a role in the detoxification of flumethrin and chlorpyrifos in Haemaphysalis longicornis
Parasites & Vectors, 2018Co-Authors: Emmanuel Pacia Hernandez, Kodai Kusakisako, Remil Linggatong Galay, Melbourne Rio Talactac, Naotoshi Tsuji, Kozo Fujisaki, Takeshi Hatta, Tetsuya TanakaAbstract:Background Haemaphysalis longicornis is a tick of importance to health, as it serves as a vector of several pathogens, including Theileria orientalis , Babesia ovata , Rickettsia japonica and the severe fever with thrombocytopenia syndrome virus (SFTSV). Presently, the major method of control for this tick is the use of chemical acaricides. The glutathione S-transferase (GST) system is one mechanism through which the tick metabolizes these acaricides. Two GSTs from H. longicornis (HlGST and HlGST2) have been previously identified. Results Enzyme kinetic studies were performed to determine the interaction of acaricides with recombinant H. longicornis GSTs. Recombinant HlGST activity was inhibited by flumethrin and cypermethrin, while recombinant HlGST2 activity was inhibited by chlorpyrifos and cypermethrin. Using real-time RT-PCR, the upregulation of the HlGST gene was observed upon exposure to sublethal doses of flumethrin, while the HlGST2 gene was upregulated when exposed to sublethal doses of chlorpyrifos. Sex and strain dependencies in the induction of GST gene expression by flumethrin were also observed. Knockdown of the HlGST gene resulted in the increased susceptibility of larvae and adult male ticks to sublethal doses of flumethrin and the susceptibility of larvae against sublethal doses of chlorpyrifos was increased upon knockdown of HlGST2 . Conclusions HlGST could be vital for the metabolism of flumethrin in larvae and adult male ticks, while HlGST2 is important in the detoxification of chlorpyrifos in larval ticks.
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Vector competence of Haemaphysalis longicornis ticks for a Japanese isolate of the Thogoto virus
Scientific Reports, 2018Co-Authors: Melbourne Rio Talactac, Kentaro Yoshii, Kodai Kusakisako, Remil Linggatong Galay, Emmanuel Pacia Hernandez, Masami Mochizuki, Kozo Fujisaki, Tetsuya TanakaAbstract:Thogoto virus (THOV), a tick-borne arbovirus not previously reported in East Asia, was recently isolated from Haemaphysalis longicornis in Kyoto, Japan. In this study, we investigated the vector competence of H. longicornis ticks for a Japanese isolate of the Thogoto virus using anal pore microinjection and experimental virus acquisition. Our results showed that anal pore microinjection can readily infect adult ticks, and THOV-infected ticks can successfully transmit the virus to mice. Blood feeding was also critical in the distribution of the virus in tick organs, most especially in the salivary glands. Furthermore, co-feeding between an infected adult and naïve nymphs can also produce infected molted adults that can horizontally transmit THOV to mice. Altogether, our results suggest that H. longicornis is a competent vector for the Japanese THOV isolate and could be the primary tick vector of the virus in Japan.
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Additional file 2: Figure S2. of Characterization and expression analysis of a newly identified glutathione S-transferase of the hard tick Haemaphysalis longicornis during blood-feeding
2018Co-Authors: Emmanuel Hernandez, Kodai Kusakisako, Remil Linggatong Galay, Melbourne Rio Talactac, Naotoshi Tsuji, Kozo Fujisaki, Takeshi Hatta, Tomohide Matsuo, Tetsuya TanakaAbstract:Nucleotide and deduced amino acid sequences of HlGST (a) and HlGST2 (b) of Haemaphysalis longicornis. Start and stop codons are underlined. Predicted glutathione and substrate binding sites are shaded in black and gray, respectively. The putative polyadenylation signal, AATAAA, is double underlined. (PDF 53 kb
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Synchronous Langat Virus Infection of Haemaphysalis longicornis Using Anal Pore Microinjection
Viruses, 2017Co-Authors: Melbourne Rio Talactac, Kentaro Yoshii, Kodai Kusakisako, Remil Linggatong Galay, Emmanuel Pacia Hernandez, Masami Mochizuki, Kozo Fujisaki, Tetsuya TanakaAbstract:The tick-borne encephalitis virus (TBEV) serocomplex of flaviviruses consists of arboviruses that cause important diseases in animals and humans. The transmission of this group of viruses is commonly associated with tick species such as Ixodes spp., Dermacentor spp., and Hyalomma spp. In the case of Haemaphysalis longicornis, the detection and isolation of flaviviruses have been previously reported. However, studies showing survival dynamics of any tick-borne flavivirus in H. longicornis are still lacking. In this study, an anal pore microinjection method was used to infect adult H. longicornis with Langat virus (LGTV), a naturally attenuated member of the TBEV serocomplex. LGTV detection in ticks was done by real-time PCR, virus isolation, and indirect immunofluorescent antibody test. The maximum viral titer was recorded at 28 days post-inoculation, and midgut cells were shown to be the primary replication site. The tick can also harbor the virus for at least 120 days and can successfully transmit LGTV to susceptible mice as confirmed by detection of LGTV antibodies. However, no transovarial transmission was observed from the egg and larval samples. Taken together, our results highly suggest that anal pore microinjection can be an effective method in infecting adult H. longicornis, which can greatly assist in our efforts to study tick and virus interactions.
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Characterization and antiviral activity of a newly identified defensin-like peptide, HEdefensin, in the hard tick Haemaphysalis longicornis.
Developmental and comparative immunology, 2016Co-Authors: Melbourne Rio Talactac, Kentaro Yoshii, Kodai Kusakisako, Remil Linggatong Galay, Emmanuel Pacia Hernandez, Masami Mochizuki, Kozo Fujisaki, Hiroki Maeda, Yurika Yada, Tetsuya TanakaAbstract:Tick defensins are antimicrobial peptides that play a major role in the innate immunity of ticks by providing a direct antimicrobial defense. In this study, we identified and characterized a defensin-like encoding gene, HEdefensin, from the expressed sequence tags (EST) database of hemolymph from the hard tick Haemaphysalis longicornis. Expression of the gene in whole adult ticks and in different organs was upregulated during blood feeding, though not after Langat virus (LGTV) challenge. A synthetic HEdefensin peptide demonstrated significant virucidal activity against LGTV but not against an adenovirus in co-incubation virucidal assays. Moreover, the RNAi-mediated gene silencing of HEdefensin did not significantly affect the virus titer as compared to the control group. The data reported here have established the in vitro virucidal activity of the peptide against LGTV. However, its role in the innate antiviral immunity of H. longicornis remains to be explored, and further studies are needed to fully evaluate the potential biological activities of the peptide against bacteria, fungi or parasites.
Misao Onuma - One of the best experts on this subject based on the ideXlab platform.
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cross immunity with Haemaphysalis longicornis glutathione s transferase reduces an experimental rhipicephalus boophilus microplus infestation
Experimental Parasitology, 2011Co-Authors: Luis Fernando Parizi, Saiki Imamura, Misao Onuma, Kazuhiko Ohashi, Aoi Masuda, Kiyoko Uemura Utiumi, Itabajara Da Silva VazAbstract:Abstract Recombinant Glutathione S-transferase of Haemaphysalis longicornis (rGST-Hl) was expressed in Escherichia coli , purified by affinity chromatography and used in the immunization of cattle. Western blot analysis showed positive antibody response in cattle immunized with rGST-Hl. The tests also showed that immunized bovine sera recognize native Rhipicephalus microplus proteins in different tissue extracts. Furthermore, the vaccine potential of rGST-Hl was investigated against infestation of Hereford cattle by R. microplus . Vaccination of cattle with rGST-Hl conferred partial cross-protection immunity against R. microplus . Considering the effect on number of engorged ticks, egg laying capacity and egg fertility, the overall efficacy of vaccination was of 57%, as compared with control group.
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cross immunity with Haemaphysalis longicornis glutathione s transferase reduces an experimental rhipicephalus boophilus microplus infestation
Experimental Parasitology, 2011Co-Authors: Luis Fernando Parizi, Saiki Imamura, Misao Onuma, Kazuhiko Ohashi, Aoi Masuda, Kiyoko Uemura Utiumi, Itabajara Da Silva VazAbstract:Recombinant Glutathione S-transferase of Haemaphysalis longicornis (rGST-Hl) was expressed in Escherichia coli, purified by affinity chromatography and used in the immunization of cattle. Western blot analysis showed positive antibody response in cattle immunized with rGST-Hl. The tests also showed that immunized bovine sera recognize native Rhipicephalus microplus proteins in different tissue extracts. Furthermore, the vaccine potential of rGST-Hl was investigated against infestation of Hereford cattle by R. microplus. Vaccination of cattle with rGST-Hl conferred partial cross-protection immunity against R. microplus. Considering the effect on number of engorged ticks, egg laying capacity and egg fertility, the overall efficacy of vaccination was of 57%, as compared with control group.
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a serine protease inhibitor serpin from Haemaphysalis longicornis as an anti tick vaccine
Vaccine, 2005Co-Authors: Saiki Imamura, Kazuhiko Ohashi, Itabajara Da Silva Vaz, Maiko Sugino, Misao OnumaAbstract:The application of anti-tick vaccine has been shown to be the most promising alternative tick control strategy compared to the current use of acaricides that suffer from a number of limitations. The success of this strategy is dependent on the cloning, and characterization of tick molecules involved in the mediation of tick central physiological roles. Rapid amplification of the cDNA ends (RACE) and primers designed based on a conserved serpin amino acid motif (NAVYFKG) were used to clone a cDNA with high similarity in the reactive center loop (RCL) to representative serpin, heparin cofactor II. We have named this novel gene as Haemaphysalis longicornis serpin-2 (HLS2). RT-PCR analysis showed that HLS2 mRNA transcripts are not expressed in salivary glands but in hemolymph by feeding ticks. HLS2 was not introduced into the bite site as measured by Western blot analysis. The activated partial thromboplastin time (APTT) and the thrombin inhibitory assay using recombinant HLS2 (rHLS2) demonstrated prolonged coagulation time and inhibition of thrombin activity. These results suggested that HLS2 is present only in hemolymph of the feeding ticks and the function of HLS2 is homeostasis in tick physiological compartment. Vaccination of rabbits with rHLS2 conferred protective immunity against ticks, resulting in 44.6 and 43.0% mortality in nymphal and adult ticks, respectively. These results show that rHLS2 could be an important candidate as a component of a cocktail anti-tick vaccine.
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molecular cloning and sequence analysis of cdnas encoding for boophilus microplus Haemaphysalis longicornis and rhipicephalus appendiculatus actins
Veterinary Parasitology, 2005Co-Authors: Itabajara Da Silva Vaz, Saiki Imamura, Kazuhiko Ohashi, Chie Nakajima, F C Cardoso, Carlos A Ferreira, Gaby Renard, Aoi Masuda, Misao OnumaAbstract:The nucleotide and deduced amino acid sequences of the actins from ticks, Boophilus microplus, Haemaphysalis longicornis and Rhipicephalus appendiculatus, have been determined. Nucleotide sequence analysis showed open reading frames of 1128-nucleotide-long encoding proteins of 376 amino acids with a predicted molecular weight of 41.82 kDa each. Comparison between the nucleic acid and deduced amino acid sequences as well as structural and phylogenetic analyses of these genes confirmed the high similarity among actins from ticks in comparison to other species.
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cloning expression and partial characterization of a Haemaphysalis longicornis and a rhipicephalus appendiculatus glutathione s transferase
Insect Molecular Biology, 2004Co-Authors: I Da Silva Vaz Jnr, Saiki Imamura, Kazuhiko Ohashi, Misao OnumaAbstract:The ticks Haemaphysalis longicornis and Rhipicephalus appendiculatus are important parasites worldwide. The current method for control of cattle ticks involves the use of chemicals. Nevertheless, parasite resistance is an ever increasing global problem. Glutathione S-transferases (GSTs) play a central role in detoxication of xenobiotic and endogenous compounds. Several authors have noted that an increase in GST activity is associated with resistance to insecticides and acaricides. In the present study, we report the cloning and expression of GST cDNAs from H. longicornis and R. appendiculatus. In addition, we determine the effect of three acaricides (ethion, deltamethrin and diazinon) on the enzymatic activity of rGSTs.
Xuenan Xuan - One of the best experts on this subject based on the ideXlab platform.
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porin expression profiles in Haemaphysalis longicornis infected with babesia microti
Frontiers in Physiology, 2020Co-Authors: Rika Umemiyashirafuji, Hiroshi Suzuki, Mingming Liu, Weiqing Zheng, Kiyoshi Okado, Paul Franck Adjou Moumouni, Qian Zhang, Haiying Chen, Xuenan XuanAbstract:The porin gene is widely disseminated in various organisms and has a pivotal role in the regulation of pathogen infection in blood-sucking arthropods. However, to date, information on the porin gene from the Haemaphysalis longicornis tick, an important vector of human and animal diseases, remains unknown. In this study, we identified the porin gene from H. longicornis and evaluated its expression levels in Babesia microti-infected and -uninfected H. longicornis ticks at developmental stages. We also analyzed porin functions in relation to both tick blood feeding and Babesia infection and the relationship between porin and porin-related apoptosis genes such as B-cell lymphoma (Bcl), cytochrome complex (Cytc), caspase 2 (Cas2), and caspase 8 (Cas8). The coding nucleotide sequence of H. longicornis porin cDNA was found to be 849 bp in length and encoded 282 amino acids. Domain analysis showed the protein to contain six determinants of voltage gating and two polypeptide binding sites. Porin mRNA levels were not significantly different between 1-day-laid and 7-day-laid eggs. In the nymphal stage, higher porin expression levels were found in unfed, 12-h-partially-fed (12 hPF), 1-day-partially-fed (1 dPF), 2 dPF nymphs and nymphs at 0 day post-engorgement (0 dAE) vs. nymphs at 2 dAE. Cytc and Cas2 mRNA levels were higher in 2 dPF nymphs in contrast to nymphs at 2 dAE. Porin expression levels appeared to be higher in the infected vs. uninfected nymphs during blood feeding except at 1 dPF and 0-1 dAE. Especially, the highest B. microti burden negatively affected porin mRNA levels in both nymphs and female adults. Porin knockdown affected body weight and Babesia infection levels and significantly downregulated the expression levels of Cytc and Bcl in H. longicornis female ticks. In addition, this study showed that infection levels of the B. microti Gray strain in nymphal and female H. longicornis peaked at or around engorgement from blood feeding to post engorgement. Taken together, the research conducted in this study suggests that H. longicornis porin might interfere with blood feeding and B. microti infection.
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identification of Haemaphysalis longicornis genes differentially expressed in response to babesia microti infection
Pathogenetics, 2020Co-Authors: Rika Umemiyashirafuji, Hiroshi Suzuki, Mingming Liu, Weiqing Zheng, Shengen Chen, Kiyoshi Okado, Paul Franck Adjou Moumouni, Shu Yang, Xuenan XuanAbstract:Haemaphysalis longicornis is a tick and a vector of various pathogens, including the human pathogenetic Babesia microti. The objective of this study was to identify female H. longicornis genes differentially expressed in response to infection with B. microti Gray strain by using a suppression subtractive hybridization (SSH) procedure. A total of 302 randomly selected clones were sequenced and analyzed in the forward subtracted SSH cDNA library related to Babesia infection, and 110 clones in the reverse cDNA library. Gene ontology assignments and sequence analyses of tick sequences in the forward cDNA library showed that 14 genes were related to response to stimulus or/and immune system process, and 7 genes had the higher number of standardized sequences per kilobase (SPK). Subsequent real-time PCR detection showed that eight genes including those encoding for Obg-like ATPase 1 (ola1), Calreticulin (crt), vitellogenin 1 (Vg1) and Vg2 were up-regulated in fed ticks. Compared to uninfected ticks, infected ticks had six up-regulated genes, including ola1, crt and Vg2. Functional analysis of up-regulated genes in fed or Babesia-infected ticks by RNA interference showed that knockdown of crt and Vg2 in infected ticks and knockdown of ola1 in uninfected ticks accelerated engorgement. In contrast, Vg1 knockdown in infected ticks had delayed engorgement. Knockdown of crt and Vg1 in infected ticks decreased engorged female weight. Vg2 knockdown reduced B. microti infection levels by 51% when compared with controls. The results reported here increase our understanding of roles of H. longicornis genes in blood feeding and B. microti infection.
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multiple ferritins are vital to successful blood feeding and reproduction of the hard tick Haemaphysalis longicornis
The Journal of Experimental Biology, 2013Co-Authors: Remil Linggatong Galay, Hiroki Maeda, Xuenan Xuan, Rika Umemiyashirafuji, Tomohide Matsuo, Hiroshi Suzuki, Kyaw Min Aung, Hiroaki Kawaguchi, Noriaki Miyoshi, Masami MochizukiAbstract:Ticks are obligate hematophagous parasites and important vectors of diseases. The large amount of blood they consume contains great quantities of iron, an essential but also toxic element. The function of ferritin, an iron storage protein, and iron metabolism in ticks need to be further elucidated. Here, we investigated the function a newly identified secreted ferritin from the hard tick Haemaphysalis longicornis (HlFER2), together with the previously identified intracellular ferritin (HlFER1). Recombinant ferritins, expressed in Escherichia coli, were used for anti-serum preparation and were also assayed for iron-binding activity. RT-PCR and western blot analyses of different organs and developmental stages of the tick during blood feeding were performed. The localization of ferritins in different organs was demonstrated through an indirect immunofluorescent antibody test. RNA interference (RNAi) was performed to evaluate the importance of ferritin in blood feeding and reproduction of ticks. The midgut was also examined after RNAi using light and transmission electron microscopy. RT-PCR showed differences in gene expression in some organs and developmental stages. Interestingly, only HlFER2 was detected in the ovary during oviposition and in the egg despite the low mRNA transcript. RNAi induced a reduction in post-blood meal body weight, high mortality and decreased fecundity. The expression of vitellogenin genes was affected by silencing of ferritin. Abnormalities in digestive cells, including disrupted microvilli, and alteration of digestive activity were also observed. Taken altogether, our results show that the iron storage and protective functions of ferritin are crucial to successful blood feeding and reproduction of H. longicornis.
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parasiticidal activity of Haemaphysalis longicornis longicin p4 peptide against toxoplasma gondii
Peptides, 2012Co-Authors: Tetsuya Tanaka, Naotoshi Tsuji, Hiroki Maeda, Xuenan Xuan, Rika Umemiyashirafuji, Tomohide Matsuo, Hiroshi Suzuki, Damdinsuren Boldbattar, Aiko Kume, Kozo FujisakiAbstract:The Haemaphysalis longicornis longicin P4 peptide is an active part peptide produced by longicin which displays bactericidal activity against both Gram-negative and Gram-positive bacteria and other microorganisms. In the present study, the effect of the longicin P4 peptide on the infectivity of Toxoplasma gondii parasites was examined in vitro. Tachyzoites of T. gondii incubated with longicin P4 had induced aggregation and lost the trypan blue dye exclusion activity and the invasion ability into the mouse embryonal cell line (NIH/3T3). Longicin P4 bound to T. gondii tachyzoites, as demonstrated by fluoresce microscopic analysis. An electron microscopic analysis and a fluorescence propidium iodide exclusion assay of tachyzoites exposed to longicin P4 revealed pore formation in the cellular membrane, membrane disorganization, and hollowing as well as cytoplasmic vacuolization. The number of tachyzoites proliferated in mouse macrophage cell line (J774A.1) was significantly decreased by incubation with longicin P4. These findings suggested that longicin P4 conceivably impaired parasite membranes, leading to the destruction of Toxoplasma parasites in J774A.1 cells. Thus, longicin P4 is an interesting candidate for antitoxoplasmosis drug design that causes severe toxicity to T. gondii and plays an important role in reducing cellular infection. This is the first report showing that longicin P4 causes aggregation and membrane injury of parasites, leading to Toxoplasma tachyzoite destruction.
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multiple vitellogenins from the Haemaphysalis longicornis tick are crucial for ovarian development
Journal of Insect Physiology, 2010Co-Authors: Damdinsuren Boldbaatar, Tetsuya Tanaka, Xuenan Xuan, Rika Umemiyashirafuji, Min Liao, Kozo FujisakiAbstract:Ovarian development and egg maturation are crucial processes for the success of reproduction in ticks. Three full-length cDNAs encoding the precursor of major yolk protein, vitellogenin, were obtained from cDNA libraries of the Haemaphysalis longicornis tick and designated as HlVg-1, HlVg-2 and HlVg-3. The HlVg mRNAs were found in fed females with major expression sites in the midgut, fat body and ovary. Native PAGE and Western blot demonstrated that HlVgs in the hemolymph, fat body and ovary of fed females consisted of four major polypeptides. RNAi results showed that HlVg dsRNA-injected ticks obtained lower body weight, egg weight and showed higher mortality of engorged females after blood sucking than control groups. Our results indicate that all HlVgs are essential for egg development and oviposition.
