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Toxoplasma gondii is a global protozoan pathogen. Clonal lineages predominate in Europe, North America, Africa, and China, whereas highly recombinant parasites are endemic in South/Central America. Far East Asian T. gondii isolates are not included in current global population genetic structure analyses at WGS resolution. Here we report a genome-wide population study that compared eight Japanese and two Chinese isolates against representative worldwide T. gondii genomes using POPSICLE, a novel population structure analyzing software. Also included were 7 genomes resurrected from non-viable isolates by target enrichment sequencing. Visualization of the genome structure by POPSICLE shows a mixture of Chinese haplogroup (HG) 13 haploblocks introgressed within the genomes of Japanese HG2 and North American HG12. Furthermore, two ancestral lineages were identified in the Japanese strains; one lineage shares a common ancestor with HG11 found in both Japanese strains and North American HG12. The other ancestral lineage, found in T. gondii isolates from a small island in Japan, is admixed with genetically diversified South/Central American strains. Taken together, this study suggests multiple ancestral links between Far East Asian and American T. gondii strains and provides insight into the transmission history of this cosmopolitan organism., (© 2024. The Author(s).)

The one-humped camel (Camelus dromedarius) or dromedary is an economically important domestic animal. However, infectious diseases, including those caused by vector-borne hemopathogens, frequently compromise the health and production of camels. In this study, we examined infections caused by Babesia, Theileria, Trypanosoma, and Anaplasma species in camels in Egypt. We analyzed blood DNA samples from 148 camels reared in six Egyptian governorates (Giza, Asyut, Sohag, Qena, Luxor, and the Red Sea) using pathogen-specific Polymerase Chain Reaction (PCR) assays. Our results indicated that 29 (19.6%), 22 (14.9%), 1 (0.7%), 2 (1.4%), 1 (0.7%), 2 (1.4%), and 28 (18.9%) of the surveyed animals were infected with Babesia bovis, B. bigemina, Babesia sp. Mymensingh, Theileria sp. Yokoyama, Theileria equi, Trypanosoma evansi, and Anaplasma marginale, respectively. We found that a total of 68 (45.9%) animals were infected with at least one of the detected hemopathogens. Sequencing analyses of PCR amplicons confirmed our diagnostic results. This study is the first to report Theileria sp. Yokoyama and Babesia sp. Mymensingh in Egypt. This is also the first report of infection with these two species in one-humped camel. In conclusion, this study found that camels in Egypt are infected with several vector-borne hemopathogens, including novel parasite species., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Image-enhanced endoscopy (IEE) allows real-time high-contrast imaging of a targeted lesion without any special equipment. Among various IEE technologies, narrow-band imaging, in which a light of shorter wavelength is used, emphasizes the surface and blood vessel patterns on the mucosal surface. This technology has been widely used in endoscopic diagnosis in the gastrointestinal tract. Red dichromatic imaging (RDI) was recently developed; it utilizes lights of longer wavelengths (520-550, 595-610, and 620-640 nm), which have weak light scattering characteristics in contrast to narrow-band imaging. RDI was designed to enhance the visibility of deep-lying blood vessels and areas of bleeding, and it has been installed in the latest Olympus endoscopy system, EVIS X1, as an advanced version of the optical-digital method that was originally developed. Improving the visibility of deep blood vessels allows more accurate evaluation of esophageal varices and the degree of inflammation in ulcerative colitis. Easier identification of a bleeding source makes hemostasis quicker and easier to accomplish during endoscopic resection procedures such as endoscopic submucosal dissection and peroral endoscopic myotomy as well as during treatment of gastrointestinal bleeding from a peptide ulcer or colon diverticulum. The authors herein review the technological development and principles, review the existing literature on RDI, and discuss the utility and effectiveness of this unique IEE technology in gastrointestinal endoscopy., (© The Author(s), 2022.)

Toxoplasmosis is a cosmopolitan protozoan zoonosis caused by Toxoplasma gondii infamous for inducing severe clinical manifestations in humans. Although the disease affects at least one billion people worldwide, it is neglected in many countries including developed ones. In literature, the epidemiological data documenting the actual incidence of the disease in humans and domestic animals from Japan are limited and importantly many earlier papers on T. gondii infections were published in Japanese and a considerable part is not available online. Herein, we review the current summary about the epidemiological situation of T. gondii infection in Japan and the potential associated risk factors in humans and animals as well as the different T. gondii genotypes isolated in Japan. Several T. gondii isolates have been identified among cats (TgCatJpTy1/k-3, TgCatJpGi1/TaJ, TgCatJpObi1 and TgCatJpOk1-4) and goats (TgGoatJpOk1-13). This literature review underscores the need for a nationwide investigation of T. gondii infection in Japanese people and assessment of the socioeconomic impact of the disease burden. Furthermore, epidemiological studies in domestic and wild animals and estimation of degree of contamination of soil or water with T. gondii oocysts are needed, for a better understanding of the scope of this public health concern., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Gastrointestinal nematodes cause massive economic losses as an important impediment to the development of animals around the world. This study aimed to investigate the microscopical diagnosis of nematode parasites of buffalo from Minoufiya, Egypt and molecular characterization of Bunostomum phlebotomum. We examined 390 fecal samples with floatation and fecal culture techniques to recognize different genera of nematodes. The results revealed B. phlebotomum (2.56%), Strongyloides papillosus (3.85%), Toxocara vitulorum (7.69%), Haemonchus sp. (1.28%), and Dictyocaulus viviparus (1.28%). The recovered eggs and larvae of nematodes were identified as well as the adults of B. phlebotomum. Age-wise, sex-wise, and seasonal prevalences of the recovered nematodes were recorded. Sequence analysis of the ITS-2 gene of B. phlebotomum was highly identical (99-100%) to sequences from Australia and China and occurred in the same clade with B. trigoncephalum. In conclusion, the study presented the coprological survey of gastrointestinal nematodes, and the genetic characterization of B. phlebotomum from Minoufiya Governorate, Egypt for the first time., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Apicomplexan parasite Toxoplasma gondii has three distinct clonal lineages: high, medium and low virulent strains, type I, II and III, respectively. T. gondii avoids the immune response by transforming from fast multiplying tachyzoite to slow multiplying bradyzoite, and establishing a chronic infection. In the present study, we isolated a new strain of T. gondii from cat feces in the Tokachi subprefecture, Hokkaido, Japan and named it as TgCatJpObi1 (Obi1) strain. Genotyping analysis of 12 loci revealed atypical characters close to type II, genotype 4 according to ToxoDB classification. Phenotypically, Obi1 strain shows slow growth rate and the ability of spontaneous cyst formation in both human foreskin fibroblast (HFFs) and mouse peritoneal macrophages in vitro without bradyzoite induction. Parasite virulence was assessed by means of mouse survival upon infection with either Obi1 or ME49 strains. Obi1 strain displayed no mortalities in comparison to type II clonal lineage, ME49 at LD 50 to LD 100 range (1 × 10 3 -10 6 tachyzoites). Although virulence of Obi1 strain is significantly lower than that of ME49, nucleotide sequences analyses revealed that genes of virulence factors such as Gra15, Rop5, 16, 17, and 18 in Obi1 strain were 100% identical to those in the type II strain. Thus, characterization of a newly isolated strain, Obi1, is crucial to clarify the development of toxoplasmosis in both humans and animals., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Cryptosporidium, a waterborne protozoan parasite, has a substantial veterinary and medical impact worldwide. This parasite is more often recognized during waterborne outbreaks because of its resistance to chlorine disinfection, small size making it difficult to inactivate/eliminate through filtration, and presence in many animal species including humans. Migratory waterfowl, in addition to acting as mechanical carriers of Cryptosporidium oocysts, can also serve as natural reservoirs of infection by host-specific Cryptosporidium species. For better understanding of the extent of genetic diversity and inter-relationships among avian isolates of Cryptosporidium, 200 fecal samples of migratory ducks from the Tokachi subprefecture, Hokkaido, Japan were collected and analyzed by nested PCR (N-PCR) at the 18S rRNA gene. N-PCR revealed that 11.5% (23/200) were positive for Cryptosporidium spp. Among all samples, sequence analysis identified that 10% (20/200) were 98-100% identical to Cryptosporidium avian genotype III. On the other hand, 1.5% (3/200) were 99-100% identical to C. baileyi. This is the first molecular study reporting the prevalence of Cryptosporidium in migratory ducks in Japan. Genetic diversity among Cryptosporidium isolates from humans and birds has been reported worldwide. Nevertheless, further studies are important to assess genetic variety and to elucidate the transmission dynamics of Cryptosporidium parasites.

Inosine 5'-monophosphate dehydrogenase (IMPDH) is an essential enzyme for the production of guanine nucleotides. Disruption of IMPDH activity has been explored as a therapeutic strategy for numerous purposes, such as for anticancer, immunosuppression, antiviral, and antimicrobial therapy. In the present study, we established a luciferase-based high-throughput screening system to identify IMPDH inhibitors from our chemical library of known bioactive small molecules. The screening of 1400 compounds resulted in the discovery of three irreversible inhibitors: disulfiram, bronopol, and ebselen. Each compound has a distinct chemical moiety that differs from other reported IMPDH inhibitors. Further evaluation revealed that these compounds are potent inhibitors of IMPDHs with k on values of 0.7 × 10 4 to 9.3 × 10 4  M -1 ·s -1 . Both disulfiram and bronopol exerted similar degree of inhibition to protozoan and mammalian IMPDHs. Ebselen showed an intriguing difference in mode of inhibition for different IMPDHs, with reversible and irreversible inhibition to each Cryptosporidium parvum IMPDH and human IMPDH type II, respectively. In the preliminary efficacy experiment against cryptosporidiosis in severe combined immunodeficiency (SCID) mouse, a decrease in the number of oocyst shed was observed upon the oral administration of disulfiram and bronopol, providing an early clinical proof-of-concept for further utilization of these compounds as IMPDH inhibitors.

Toxoplasma gondii and Neospora caninum are protozoan parasites that cause huge economic losses in animal industries worldwide. N. caninum can cause abortion storms and high culling rates in cattle, whereas T. gondii infection is a significant concern in both human and animals because it can induce abortion and clinical symptoms in immunocompromised hosts. The aim of this study was to determine the seroprevalence of T. gondii and N. caninum in cattle in Mongolia. Specific antibodies to T. gondii and N. caninum were detected by using an indirect enzyme-linked immunosorbent assay (iELISA) based on recombinant antigens of dense granule protein 7 of Toxoplasma gondii and surface antigen 1 of Neospora caninum, respectively. A total of 1438 cattle sera from 20 of 21 provinces of Mongolia and the capital city of Ulaanbaatar were tested. Overall, 18.7% and 26.2% of cattle were positive for specific antibodies to T. gondii and N. caninum, respectively. Prevalence rates were higher (T. gondii infection: P < .0001, N. caninum infection: P = .002) in the central region of Mongolia (T. gondii infection: 27.1%, N. caninum infection: 30.8%) compared with western region, suggesting that prevalence rates might be influenced by geographical condition, particularly warmer temperatures around this area in Mongolia. The lowest prevalence rates were observed in the western region of Mongolia (T. gondii: 9%, N. caninum: 20.8%). In addition, the seroprevalence of N. caninum in female animals (27.5%) was significantly higher than that in male animals (20.4%) (P = .018), suggesting an important risk factor of abortion and stillbirth in cattle. The present results showed that T. gondii and N. caninum infections might be a risk for public health and economy of the livestock industry in Mongolia. In conclusion, this study demonstrates high seroprevalences of T. gondii and N. caninum in Mongolia and provides valuable new data for development of control measures against these infections in Mongolia., (Copyright © 2018 Elsevier B.V. All rights reserved.)

The present study investigated the prevalence of Toxoplasma gondii and other intestinal parasites in cats in the Tokachi subprefecture in Japan. A total of 365 household cats were included in the study, and 353 serum and 351 fecal samples were collected and analyzed. T. gondii IgG antibodies were detected in the sera of 16.14% of cats based on Latex agglutination test and ELISA. For ELISA, T. gondii RH strain tachyzoites lysate and T. gondii SAG2 recombinant protein were used as antigens. Low seropositivity was detected in cats younger than one year and older than 11 years; outdoor and hunter cats showed significantly high seropositivities. Neutering either in male or female cats, but not gender, had a considerable effect on seroprevalence. Toxoplasma gondii oocysts were detected in one fecal sample. The overall parasitic infestation in cats was 12.5%. Other detected parasites included Toxocara species, which showed the highest prevalence of 7.7%, followed by Isospora spp. (2%), Taenia spp. (1.7%), and Ancylostoma spp. (0.9%). Spirometra spp. was detected in only one sample. Outdoor cats comprised 50% of all 44 parasite-infested cats. Although T. gondii oocysts were detected in only one sample, the relatively high seroprevalence of T. gondii indicated that it can pose significant risks to the environment. Our findings highlighted the potential of outdoor cats as a source of T. gondii and other parasites.

The δD temperature proxy in Antarctic ice cores varies in parallel with CO 2 through glacial cycles. However, these variables display a puzzling asynchrony. Well-dated records of Southern Ocean temperature will provide crucial information because the Southern Ocean is likely key in regulating CO 2 variations. Here, we perform multiple isotopic analyses on an Antarctic ice core and estimate temperature variations at this site and in the oceanic moisture source over the past 720,000 years, which extend the longest records by 300,000 years. Antarctic temperature is affected by large variations in local insolation that are induced by obliquity. At the obliquity periodicity, the Antarctic and ocean temperatures lag annual mean insolation. Further, the magnitude of the phase lag is minimal during low eccentricity periods, suggesting that secular changes in the global carbon cycle and the ocean circulation modulate the phase relationship among temperatures, CO 2 and insolation in the obliquity frequency band.

Considering the long lifespan of the liver fluke Opisthorchis viverrini, human mobility from prevalent regions to other neighboring areas has the possibility to disperse carriers and complicate the opisthorchiasis problem. To evaluate this, mass screening of the fluke infection was conducted in nine communities of lower Northern Thailand, combined with a questionnaire survey to distinguish the participant's origin. The liver fluke infection was found in 70 individuals (7.2%) of the examined 971 stool samples from seven communities, with light intensity providing small numbers of eggs in the examined stool. Prevalence in the positive communities varied from 2.1% to 28.7%. As a result of generalized linear mixed models fitting, regional origin and raw-fish eating habits were stably selected as variables affecting the parasite infection while occupation and educational background were secondary ones. Majority of the infected cases (64.3%) were found from the immigrants of northeastern Thailand (the fluke prevalent region), providing 2.28-2.42 times higher infectious risk on average against the local residents. Daily consumption of raw fish averaged a 3.12-3.60 times higher risk compared to those with no raw-fish eating habit. Our findings suggest that people's origin and moving history deserve further attentions in health promotion programs including education for safe eating., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Oocyst stage of Toxoplasma gondii is characterized by a durable wall that confers a strong protection to this protozoan parasite in face of harsh environmental conditions. Thus, it is considered the key for transmission of T. gondii. Analysis of oocyst wall composition is mandatory therefore; the aim of this study was to identify novel T. gondii oocyst wall proteins and test their use in detection of these oocysts in environmental samples. Five candidates of novel T. gondii oocyst wall proteins (TgOWPs) were identified and named TgOWP8 through TgOWP12. Recombinant protein of TgOWP8 was expressed in E. coli using glutathione S-transferase as fusion protein. Polyclonal antibody was produced and validated by indirect immunofluorescence antibody assay (IFA). For detection by IFA, we used different methods for fixation and permeabilization of oocysts to improve the antigen-antibody detection. Specificity to wall of T. gondii oocyst was confirmed and revealed absence of cross reactivity with bradyzoite cyst wall and tachyzoites. Although some TgOWPs were identified previously, our study represents a continuation of molecular investigations of oocyst wall proteins as an essential structure for the longevity and infectivity of this stage and also provided new trial to improve T. gondii oocysts detection., (Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.)

In the asexual stages, Toxoplasma gondii stage converts between acute phase rapidly replicating tachyzoites and chronic phase slowly dividing bradyzoites. Correspondingly, T. gondii differentially expresses two distinct genes and isoforms of the lactate dehydrogenase enzyme, expressing LDH1 exclusively in the tachyzoite stage and LDH2 preferentially in the bradyzoite stage. LDH catalyzes the interconversion of pyruvate and lactate in anaerobic growth conditions and is utilized for energy supply, however, the precise role of LDH1 and LDH2 in parasite biology in the asexual stages is still unclear. Here, we investigated the biological role of LDH1 and LDH2 in the asexual stages, and the vaccine strain potential of deletion mutants lacking LDH1, LDH2, or both genes (Δldh1, Δldh2 and Δldh1/2). Deletion of LDH1 reduced acute parasite virulence, impaired bradyzoite differentiation in vitro, and markedly reduced chronic stage cyst burdens in vivo. In contrast, deletion of LDH2 impaired chronic stage cyst burdens without affecting virulence or bradyzoite differentiation. Deletion of both LDH1 and LDH2 induced a more severe defect in chronic stage cyst burdens. These LDH mutant phenotypes were not associated with any growth defect. Vaccination of mice with a low dose of mutants deleted for LDH elicited effective protective immunity to lethal challenge infection, demonstrating the vaccine potential of LDH deletion mutants. These results suggest that lactate dehydrogenase in T. gondii controls virulence, bradyzoite differentiation, and chronic infection and reveals the potential of LDH mutants as vaccine strains.

Climatic variabilities on millennial and longer time scales with a bipolar seesaw pattern have been documented in paleoclimatic records, but their frequencies, relationships with mean climatic state, and mechanisms remain unclear. Understanding the processes and sensitivities that underlie these changes will underpin better understanding of the climate system and projections of its future change. We investigate the long-term characteristics of climatic variability using a new ice-core record from Dome Fuji, East Antarctica, combined with an existing long record from the Dome C ice core. Antarctic warming events over the past 720,000 years are most frequent when the Antarctic temperature is slightly below average on orbital time scales, equivalent to an intermediate climate during glacial periods, whereas interglacial and fully glaciated climates are unfavourable for a millennial-scale bipolar seesaw. Numerical experiments using a fully coupled atmosphere-ocean general circulation model with freshwater hosing in the northern North Atlantic showed that climate becomes most unstable in intermediate glacial conditions associated with large changes in sea ice and the Atlantic Meridional Overturning Circulation. Model sensitivity experiments suggest that the prerequisite for the most frequent climate instability with bipolar seesaw pattern during the late Pleistocene era is associated with reduced atmospheric CO 2 concentration via global cooling and sea ice formation in the North Atlantic, in addition to extended Northern Hemisphere ice sheets.

Gastrointestinal zoonotic helminths of dogs and cats have a public health concern worldwide. We investigated the prevalence of gastrointestinal helminths of zoonotic significance in dogs and cats in lower Northern Thailand and utilized molecular tools for species identification of hookworms and Opisthorchis viverrini. Fecal samples of 197 dogs and 180 cats were collected. Overall prevalence of infection using microscopy was 40.1% in dogs and 33.9% in cats. Helminth infection found in both dogs and cats included hookworms, Spirometra spp., Taenia spp., Toxocara spp., O. viverrini, Strongyloides spp. and Trichuris spp. Hookworms were the most common helminth in dogs, while Spirometra spp. were the most prevalent in cats. Among hookworm infection in dogs and cats, Ancylostoma ceylanicum was the most prevalent hookworm, being 82.1% in hookworm infected dogs and 95.8% in hookworm infected cats. Mixed-infection due to hookworms and Spirometra spp. was the most dominant in both dogs and cats. Our finding showed that zoonotic helminth infection is highly prevalent in dogs and cats in the lower Northern area of Thailand.

Cats are the only definitive hosts of Toxoplasma gondii and constitute an essential source of infection to all warm blooded animals and humans. Diagnosis of T. gondii infection in cats is fundamental for proper management and control of infection in humans and animals. In the current study, we have evaluated the diagnostic performance of tachyzoite lysate antigen (TLA) and different T. gondii recombinant antigens including surface antigen 2 (SAG2), dense granule proteins 2, 6, 7, 15 (GRA2, GRA6, GRA7, GRA15) and microneme 10 protein (MIC10) in immunoglobulin G enzyme linked-immunosorbent assay (IgG ELISA) using cat serum samples, with reference to latex agglutination test (LAT). Remarkably, TLA showed better performance than other recombinant antigens in IgG ELISAs as compared to LAT, with concordance and Kappa values of 94.27% and 0.93, respectively. Furthermore, to improve the reactivity of the recombinant antigens, we have developed IgG ELISAs using different combinations with these recombinant antigens. Strikingly, a combination of SAG2 and GRAs has relatively similar performance as TLA evidenced by concordance and Kappa values of 94.27% and 0.81, respectively. The developed ELISA with a combination of recombinant antigens can be used as a promising diagnostic tool for routine testing of T. gondii infection and mass screening in cats. The major advantages of this assay are the high sensitivity and specificity, lower cost, safer production and easiness of standardization in various laboratories worldwide., (Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.)

Goat farming is important for the livelihood of millions of rural people because it contributes to food security and creation of assets. However, infection of goats with Toxoplasma gondii could be a source of parasite transmission to humans. The information on T. gondii infection of goat was not reported yet in Myanmar. A total of 119 goat serum samples were collected from three cities in the central region of Myanmar for T. gondii antibody survey. With the occurrence value obtained in this first study, a second one, more complete, with larger number (162) of animals and properties, was carried out and the risk factors and prevalence were determined. In both studies the samples were analyzed by the LAT. Of these, 32 (11.4%) samples were showed to be positive. The infection was associated with the presence of cats at the farm (odds ratio [OR] = 4.66, 95% confidential interval [CI] = 1.03-21.06), farming with different animal species (sheep, cattle, and pigs) (OR = 4.33, 95% CI = 1.57-11.94), and farming without good management practices (OR = 0.23, 95% CI = 0.06-0.83). This is the first T. gondii prevalence study in goats in the country.

Background: The glyoxalase pathway, which includes two enzymes, glyoxalase 1 and 2 (Glo1 and Glo2), is a ubiquitous cellular system responsible for the removal of cytotoxic methylglyoxal produced during glycolysis. Protozoan parasites, including Toxoplasma gondii (T. gondii) tachyzoites, produce methylglyoxal because of increased glycolytic fluxes. A Glo1 inhibitor such as curcumin could be considered a drug candidate for anti-protozoan, anti-inflammatory, and anti-cancer therapy., Methods: The T. gondii Glo1 gene (TgGlo1) was cloned and the recombinant protein was produced. Enzyme kinetics of TgGlo1 and five mutants were evaluated by adding methylglyoxal and glutathione to a reaction mixture. Finally, the inhibitory effects of various concentrations of curcumin on recombinant TgGlo1 were evaluated using in vitro cultures of T. gondii., Results: Active recombinant TgGlo1 was successfully produced and the active sites (E166 and E251) of TgGlo1 were verified by point mutagenesis. Curcumin at the tested doses inhibited the enzymatic activity of recombinant TgGlo1 as well as the parasitic propagation of in vitro-cultured T. gondii. The Ki and IC50 were 12.9 ± 0.5 μM and 38.3 ± 0.9 μM, respectively., Conclusion: The inhibitory effect of curcumin on the enzymatic activity of TgGlo1 and parasitic propagation of T. gondii could be explored in the potential development of a potent drug for the treatment of toxoplasmosis. However, considering the fact that curcumin is known to have many effects on other molecules in the micromolar range, further elucidation of curcumin's direct inhibition of the glyoxalase system of T. gondii will be needed.

Parasites of the phylum Apicomplexa are highly successful pathogens of humans and animals worldwide. As obligate intracellular parasites, they have significant energy requirements for invasion and gliding motility that are supplied by various metabolic pathways. Aldolases have emerged as key enzymes involved in these pathways, and all apicomplexans express one or both of fructose 1,6-bisphosphate (F16BP) aldolase and 2-deoxyribose 5-phosphate (dR5P) aldolase (DERA). Intriguingly, Toxoplasma gondii, a highly successful apicomplexan parasite, expresses F16BP aldolase (TgALD1), d5RP aldolase (TgDERA), and a divergent dR5P aldolase-like protein (TgDPA) exclusively in the latent bradyzoite stage. While the importance of TgALD1 in glycolysis is well established and TgDERA is also likely to be involved in parasite metabolism, the detailed function of TgDPA remains elusive. To gain mechanistic insight into the function of different T. gondii aldolases, we first determined the crystal structures of TgALD1 and TgDPA. Structural analysis revealed that both aldolases adopt a TIM barrel fold accessorized with divergent secondary structure elements. Structural comparison of TgALD1 and TgDPA with members of their respective enzyme families revealed that, while the active-site residues are conserved in TgALD1, key catalytic residues are absent in TgDPA. Consistent with this observation, biochemical assays showed that, while TgALD1 was active on F16BP, TgDPA was inactive on dR5P. Intriguingly, both aldolases are competent to bind polymerized actin in vitro. Altogether, structural and biochemical analyses of T. gondii aldolase and aldolase-like proteins reveal diverse functionalization of the classic TIM barrel aldolase fold., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

The potential contamination of Toxoplasma gondii and Neospora caninum oocysts in the human environment is a concern from the public health viewpoint. However, estimation of their seroprevalences in humans cannot be performed in a manner that distinguishes between oocysts and tissue cysts as a source of infection. Rabbits are considered popular pet animals in Japan that can acquire natural infections by the aforementioned parasites only through the ingestion of oocysts. Therefore, this study was conducted to estimate the seroprevalences of T. gondii and N. caninum in pet rabbits in Japan as an indicator of the possible oocyst contamination in the environment surrounding human beings. Serum samples of 337 rabbits were examined by different serological methods. Enzyme-linked immunosorbent assays were performed to measure the titer of IgG and IgM antibodies. Samples revealed to be seropositive by ELISA were further analyzed by a latex agglutination test, Western blotting and an indirect immunofluorescence assay. The rates of seropositivity for T. gondii were 0.89% (3/337) and 0.29% (1/337) in IgG and IgM ELISA, respectively. SAG1 and SAG2 were detected as major antigens by the positive rabbit sera in Western blotting associated with strong staining observed by IFA in T. gondii tachyzoites. Regarding N. caninum, none of the serum samples showed a specific reaction in both Western blotting and the IFA. The results of this study indicate low seroprevalences of toxoplasmosis and neosporosis in pet rabbits in Japan, suggesting low oocyst contamination in the human environment.

The interparticle distance between nanoparticles (NPs) dispersed on on SiO2 was shown to be controlled by PEGylation. Ferritins with nanoparticle cores were prepared and PEGylated with poly(ethylene glycol)s (PEGs) of two different molecular weights. It was shown that the thickness of the PEG layer on the ferritin surface determines the interparticle distance under short Debye lengths. Under conditions where the Debye length was greater than the PEG layer thickness, distance between ferritins increased due to the electrostatic repulsive force. Results suggest that the PEG layer accommodated a small amount of counterions insufficient to cancel the ferritin outer surface charges. Simulation showed that ferritins adsorbed randomly and interparticle distance can be predicted theoretically. We demonstrate that PEGylated ferritins, that is, NP cores, can be dispersed on a surface with interval distances between particles determined by the combination of the ionic strength of the solution and the molecular weight of the PEG.

Host cell invasion by apicomplexan parasites driven by gliding motility and empowered by actin-based movement is essential for parasite survival and pathogenicity. The parasites share a conserved invasion process: actin-based motility led by the coordination of adhesin-cytoskeleton via aldolase. A number of studies of host cell invasion in the Plasmodium species and Toxoplasma gondii have been performed. However, the mechanisms of host cell invasion by Babesia species have not yet been studied. Here, we show that Babesia gibsoni aldolase (BgALD) forms a complex with B. gibsoni thrombospondin-related anonymous protein (BgTRAP) and B. gibsoni actin (BgACT), depending on tryptophan-734 (W-734) in BgTRAP. In addition, actin polymerization is mediated by BgALD. Moreover, cytochalasin D, which disrupts actin polymerization, suppressed B. gibsoni parasite growth and inhibited the host cell invasion by parasites, indicating that actin dynamics are essential for erythrocyte invasion by B. gibsoni. This study is the first molecular approach to determine the invasion mechanisms of Babesia species., (Copyright © 2013 Elsevier Inc. All rights reserved.)

We successfully fabricated defect-free, distributed and sub-20-nm GaAs quantum dots (named GaAs nanodisks (NDs)) by using a novel top-down technique that combines a new bio-template (PEGylated ferritin) and defect-free neutral beam etching (NBE). Greater flexibility was achieved when engineering the quantum levels of ND structures resulted in greater flexibility than that for a conventional quantum dot structure because structures enabled independent control of thickness and diameter parameters. The ND height was controlled by adjusting the deposition thickness, while the ND diameter was controlled by adjusting the hydrogen-radical treatment conditions prior to NBE. Photoluminescence emission due to carrier recombination between the ground states of GaAs NDs was observed, which showed that the emission energy shift depended on the ND diameters. Quantum level engineering due to both diameter and thickness was verified from the good agreement between the PL emission energy and the calculated quantum confinement energy.

A highly periodical Si nanodisk superlattice has been fabricated by our top-down process. Based on the realistic structure, a 3D simulation program using the finite element method is developed to calculate energy band structure, optical and electrical properties, as well as the intermediate band solar cell operation. Both the experiments and simulations reveal that miniband formation enhances the optical and electrical collections. Consequently, detailed electronic structure and conversion efficiency are examined to guide the optimal design of minibands. A theoretically predicted maximal efficiency of the explored Si nanodisk superlattice is 50.3%, which is promising, compared with well-known complicated Si tandem solar cells.

We fabricated a three-dimensional (3D) stacked Si nanodisk (Si-ND) array with a high aspect ratio and uniform size by using our advanced top-down technology consisting of bio-template and neutral beam etching processes. We found from conductive atomic microscope measurements that conductivity became higher as the arrangement was changed from a single Si-ND to two-dimensional (2D) and 3D arrays with the same matrix of SiC, i.e., the coupling of wave functions was changed. Moreover, our theoretical calculations suggested that the formation of minibands enhanced tunneling current, which well supported our experimental results. Further analysis indicated that four or more Si-NDs basically maximized the advantage of minibands in our structure. However, it appeared that differences in miniband widths between 2D and 3D Si-ND arrays did not affect the enhancement of the optical absorption coefficient. Hence, high photocurrent could be observed in our Si-ND array with high photoabsorption and carrier conductivity due to the formation of 3D minibands.

We study the temperature dependence of time-resolved photoluminescence (PL) in closely packed alignment of Si nanodisks (NDs) with SiC barriers, fabricated by neutral beam etching using bio-nano-templates. The PL time profile indicates three decaying components with different decay times. The PL intensities in the two slower decaying components depend strongly on temperature. These temperature dependences of the PL intensity can be quantitatively explained by a three-level model with thermal activation energies of 410 and 490 meV, depending on the PL components. The activation energies explain PL quenching due to thermal escape of electrons from individual NDs. This thermal escape affects the PL decay times above 250 K. Dark states of photo-excited carriers originating from the separate localization of electron and hole into different NDs are elucidated with the localization energies of 70 and 90 meV. In contrast, the dynamics of the fastest PL decaying component is dominated by electron tunneling among NDs, where the PL intensity and decay time are constant for temperature.

The intracellular protozoan parasite Toxoplasma gondii is capable of invading any nucleated cell and replicates within a parasitophorous vacuole (PV). This microenvironment is modified by secretory proteins from organelles named rhoptries and dense granules. In this report, we identify a novel dense granule protein, which we refer to as GRA22. GRA22 has no significant homology to any other known proteins. GRA22 possesses a signal peptide at the N-terminal end which is responsible for dense granule and PV localization. The RH strain GRA22 contains 12 copies of tandem repeats consisting each of 21 amino acids located between the 42nd and 293rd amino acid residues from a full length of 624 amino acids. On the other hand, ME49 strain GRA22 has 10 copies of tandem repeats. The Neospora caninum GRA22 ortholog completely lacks this repetitive sequence. GRA22 knock out parasites show a similar growth rate as the parental strain. However, the timing of egress is earlier than that of the parental strain. These results suggest that GRA22 is involved in regulating parasite egress in T. gondii., (Copyright © 2013 Elsevier B.V. All rights reserved.)

A sub-10 nm, high-density, periodic silicon nanodisk (Si-ND) array with a SiC interlayer has been fabricated using a new top-down process that involves a 2D array of a bio-template etching mask and damage-free neutral beam etching. Optical and electrical measurements were carried out to clarify the formation of mini-bands due to wavefunction coupling. We found that the SiC interlayer could enhance the optical absorption coefficient in the layer of Si-NDs due to the stronger coupling of wavefunctions. Theoretical simulation also indicated that wavefunction coupling was effectively enhanced in Si-NDs with a SiC interlayer, which precisely matched the experimental results. Furthermore, the I-V properties of a 2D array of Si-NDs with a SiC interlayer were studied through conductive AFM measurements, which indicated conductivity in the structure was enhanced by strong lateral electronic coupling between neighboring Si-NDs. We confirmed carrier generation and less current degradation in the structure due to high photon absorption and conductivity by inserting the Si-NDs into p-i-n solar cells.

The aim of this study was the preparation and characterization of a bioluminescence bacteria sheet in view of the fabrication of an oxygen marker inside a biofilm. Photobacterium kishitanii sheets were prepared using sodium alginate, and a biofilm of Pseudomonas aeruginosa was dropped over it. The relationship between the dissolved oxygen (DO) concentration in the biofilm and luminescence from the sheet was examined. As a conclusion, the alginate-gel film method was effective in fabricating a DO-monitoring thin film.

The authors study plasmonic enhancements of photoluminescence (PL) in Si nanodisk (ND) arrays hybridized with nanostructures such as nanoplates of Au, where these hybrid nanostructures are fabricated by fully top-down lithography: neutral-beam etching using bio-nano-templates and high-resolution electron-beam lithography. The separation distance between the Si ND and Au nanostructure surfaces is precisely controlled by inserting a thin SiO2 layer with a thickness of 3 nm. We observe that PL intensities in the Si NDs are enhanced by factors up to 5 depending on the wavelength by integrating with the Au nanoplates. These enhancements also depend on the size and shape of the Au nanoplates.

The picosecond carrier dynamics in a closely packed Si-nanodisk (Si-ND) array with ultrathin potential barrier fabricated by neutral beam etching using bio-nano-templates was investigated by time-resolved photoluminescence (PL). The PL decay curves were analyzed as a function of photon energy by the global fitting method. We show three spectral components with different decay times, where the systematic energy differences of the spectral peaks are clarified: 2.03 eV for the fastest decaying component with a decay time τ = 40 ps, 2.02 eV for τ = 300 ps, and 2.00 eV for τ = 1.6 ns. These energy separations ranging from 10 to 30 meV among the emissive states can be attributed to the coupling of wavefunctions of carriers between neighboring NDs.

A sub-10 nm, high-density, periodic silicon-nanodisc (Si-ND) array has been fabricated using a new top-down process, which involves a 2D array bio-template etching mask made of Listeria-Dps with a 4.5 nm diameter iron oxide core and damage-free neutral-beam etching (Si-ND diameter: 6.4 nm). An Si-ND array with an SiO(2) matrix demonstrated more controllable optical bandgap energy due to the fine tunability of the Si-ND thickness and diameter. Unlike the case of shrinking Si-ND thickness, the case of shrinking Si-ND diameter simultaneously increased the optical absorption coefficient and the optical bandgap energy. The optical absorption coefficient became higher due to the decrease in the center-to-center distance of NDs to enhance wavefunction coupling. This means that our 6 nm diameter Si-ND structure can satisfy the strict requirements of optical bandgap energy control and high absorption coefficient for achieving realistic Si quantum dot solar cells.

The effect of intra-gastric infusion of skipjack tuna (Katsuwonus pelamis) muscle soluble extract and of carnosine, anserine and histidine, on pepsin-like protease activity was studied in the isolated, externally batch-cultured stomach of the rockfish (Sebastes schlegelii). Stomach was isolated from the fish, then intragastrically infused with the above solutions or with a balanced saline solution (control solution) as the stimulant at 0.1 mL/min for 6h. Intra-gastric efflux was collected for measurement of pepsin-like protease activity. Skipjack tuna soluble extract, but not carnosine, anserine or histidine alone caused significant enhancement of pepsin-like protease activity during the infusion. Pepsin-like protease activity from skipjack tuna soluble extract responded immediately after infusion and was kept for 150 min after infusion. Stomach motility at the end of infusion was also observed. These results suggest that isolated stomach can receive mucosal side stimulants in vitro. It is likely that some effective components for stomach digestion in fish exist in skipjack tuna soluble extract., (Copyright © 2010 Elsevier Inc. All rights reserved.)

The DnaK-tetratricopeptide repeat (DnaK-TPR) gene (ToxoDB ID, TGME49&#95;002020) is expressed predominantly at the bradyzoite stage. DnaK-TPR protein has a heat shock protein (DnaK) and tetratricopeptide repeat (TPR) domains with amino acid sequence similarity to the counterparts of other organisms (40.2-43.7% to DnaK domain and 41.1-66.0% to TPR domain). These findings allowed us to infer that DnaK-TPR protein is important in the tachyzoite-to-bradyzoite development or maintenance of cyst structure although the function of this gene is still unknown. An immunofluorescence assay (IFA) revealed that DnaK-TPR protein was expressed in Toxoplasma gondii-encysted and in vitro-induced bradyzoites and distributed in the whole part of parasite cells. We conducted yeast two-hybrid screening to identify proteins interacting with DnaK-TPR protein, and demonstrated that DnaK-TPR protein interacts with p23 co-chaperone protein (Tgp23). It was expected that DnaK-TPR protein would have a function as a molecular chaperon in bradyzoite cells associated with Tgp23. Possible mechanisms for this gene are discussed., (Copyright © 2011 Elsevier Inc. All rights reserved.)

We created a two-dimensional array of sub-10 nm Si-nanodiscs (Si-NDs), i.e. a 2D array of Si-NDs, with a highly ordered arrangement and dense NDs by using a new top-down technique comprising advanced damage-free neutral-beam (NB) etching and a bio-template (iron oxide core) as a uniform sub-10 nm etching mask. The bandgap energy (E(g)) of the fabricated 2D array of Si-NDs can be simply controlled from 2.2 to 1.3 eV by changing the ND thickness from 2 to 12 nm. Due to weak quantum confinement existing in the diameter direction resulting from the sub-10 nm Si-ND diameter, even though the thickness of the Si-ND is much larger than the Bohr radius of Si, E(g) is still larger than the 1.1 eV E(g) of bulk Si. Si-ND not only has wide controllable E(g) but also a high absorption coefficient due to quantum confinement in three dimensions. This new technique is a promising candidate for developing new nanostructures and could be integrated into the fabrication of nanoelectronic devices.

The Toxoplasma gondii deoxyribose phosphate aldolase-like (TgDPA) gene is expressed predominantly in bradyzoites. This finding allowed us to infer that TgDPA is important in the tachyzoite-to-bradyzoite development or maintenance of cyst structure although the function of this gene is still unknown. We conducted yeast two-hybrid screening to identify proteins interacting with TgDPA, and the actin depolymerizing factor (TgADF) gene was obtained. Co-immunoprecipitation and a GST pull-down assay demonstrated that TgDPA interacts with TgADF. To reveal the significance of the protein-protein interaction between TgDPA and TgADF, actin polymerization and disassembly kinetics were examined. Addition of GST-TgDPA to TgADF lowered the extent of actin polymerization and enhanced the filamentous actin disassembly. These results demonstrated that this is the novel protein-protein interaction in T. gondii, and that TgDPA can enhance the activity of TgADF. This phenomenon might play an important role in T. gondii bradyzoites by affecting the actin turnover., (2010 Elsevier B.V. All rights reserved.)

For the last couple of years, a method that permits the collection of precise positional information of transcriptional start sites (TSSs) together with digital information of the gene-expression levels in a high-throughput manner was established. We applied this novel method, 'tss-seq', to elucidate the transcriptome of tachyzoites of the Toxoplasma gondii, which resulted in the identification of 124,000 TSSs, and they were clustered into 10,000 transcription regions (TRs) with a statistics-based analysis. The TRs and annotated ORFs were paired, resulting in the identification of 30% of the TRs and 40% of the ORFs without their counterparts, which predicted undiscovered genes and stage-specific transcriptions, respectively. The massive data for TSSs make it possible to execute the first systematic analysis of the T. gondii core promoter structure, and the information showed that T. gondii utilized an initiator-like motif for their transcription in the major and novel motif, the downstream thymidine cluster, which was similar to the Y patch observed in plants. This encyclopaedic analysis also suggested that the TATA box, and the other well-known core promoter elements were hardly utilized.

Toxoplasma gondii is an obligate intracellular protozoan parasite that invades a wide range of host cells. The parasite releases a large variety of proteins from a secretory organelle, microneme, and the secretion is essential for the parasite invasion. We cloned a secreted protein with an altered thrombospondin repeat of Toxoplasma gondii (TgSPATR), which was the homologue of Plasmodium SPATRs. Immunofluorescence double staining experiment revealed that TgSPATR was co-localized with a microneme protein, MIC2, and immuno-electron microscopic (IEM) analysis detected TgSPATR in the microneme-like structure. TgSPATR secretion was induced by ethanol, while an intracellular Ca(2+) chelator, 1,2-bis(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid, tetraacetoxymethyl ester (BAPTA-AM), suppressed the ethanol-induced secretion, suggesting the secretion was Ca(2+)-dependent, similarly to known microneme proteins. Furthermore, TgSPATR, existed on outer surface of the parasites, was detected by incomplete membrane permeabilization by saponin and immunofluorescent antibody test (IFAT). Both TgSPATR and MIC2 were detected on outer surface of extracellular parasites, but not of intracellular single parasites, suggesting they were similarly secreted during early stages of parasite invasion. Therefore, TgSPATR is probably new member of microneme protein and maybe involved in parasite invasion.

The aim of the present study using near-infrared spectroscopy (NIRS) is to evaluate the correlation of cerebral (parieto-temporal lobe) hemoglobin changes and vertiginous sensation during unilateral caloric stimulation. During the hot water (44 degrees C) stimulus, cerebral hemoglobin was increased bilaterally, but it was dominant ipsilaterally. During the unilateral cold water (30 degrees C) stimulus, cerebral hemoglobin was decreased on both sides, especially on the ipsilateral side. Vertigious sensation was strong in cases with marked left-right difference of cerebral hemoglobin changes. The usefulness of NIRS to investigate the relationship between peripheral vestibular organ and vestibular cortex was verified.

Toxoplasma gondii undergoes stage conversion from tachyzoites to bradyzoites in intermediate hosts. There have been many reports on bradyzoite-specific genes which are thought to be involved in stage conversion. Here, we described a novel T. gondii deoxyribose phosphate aldolase-like gene (TgDPA) expressing predominantly in bradyzoites. The TgDPA gene encodes 286 amino acids having a predicted molecular weight of 31kDa. Sequence analysis revealed that TgDPA had a deoxyribose phosphate aldolase (DeoC) domain with about 30% homology with its Escherichia coli counterpart. RT- and quantitative PCR analyses showed that the TgDPA gene was more expressed in bradyzoites and that its expression gradually increased during in vitro tachyzoite-to-bradyzoite stage conversion. A polyclonal antibody against recombinant TgDPA protein was raised in rabbits, and immunofluorescent analysis demonstrated that TgDPA was expressed in bradyzoites in vivo and in vitro. These findings indicate that the TgDPA gene is a new bradyzoite-specific marker and might play a role in bradyzoites.

Infection with Encephalitozoon cuniculi in rabbits frequently exists as a chronic, latent infection, and only a percentage of infected animals develop clinical disease. This study presents a seroepidemiological study of E. cunicucli infection in 337 pet rabbits collected from 20 prefectures in Japan in 2006 and 2007, using enzyme-linked immunosorbent assay (ELISA) capable of measuring IgG and IgM antibodies. These rabbits were divided into the following four groups: healthy and isolated rabbits (n=74, group I), healthy and companioned rabbits (n=121, group II), neurologically diseased rabbits (n=105, group III), and other diseased rabbits (n=37, group IV). Using ELISA for IgG antibodies, the highest detection rate, 81%, was seen in group III, the second highest, 75.2%, in group II, and the lowest, 29.7%, in group I, which was significantly different to the other groups except for group IV (43.2%). On the other hand, when ELISA was used for IgM antibody detection, 14-40% of rabbits in the four groups were also observed to have anti-E. cuniculi IgM. This study demonstrated high seroprevalence of E. cuniculi in not only neurologically diseased rabbits but also healthy and other diseased rabbits.

Conclusions: A lengthy alteration of gravity direction produced different effects on the intrinsic horizontal and vertical optokinetic oculomotor systems., Objective: To examine both optokinetic nystagmus (OKN) and optokinetic after-nystagmus (OKAN) in a 6 h 6 degrees head-down bedrest study, in which the subjects were kept lying under simulated micro-gravity conditions., Subjects and Methods: In six normal healthy adults, we repeatedly (five times) and comparatively studied OKN and OKAN evoked by horizontal and vertical stimuli. Stage 1 was an upright sitting position. During the 6 h bedrest condition, we studied OKN and OKAN in 90 degrees recumbent lateral positions (stages 2, 3, and 4). In stage 5 the subject returned to an upright position., Results: We confirmed that the change in gravity direction had various effects on the condition of OKN and OKAN. Also, we found that it took more than 3 h to reach a desirable level of systemic adaptive modification to the unique environmental condition. We considered that the early change was basically due to the changes in sensory inputs through the otolith organs, and the latter changes represented the adaptive process of the spatial orientation system. During the tilt, the occurrence rates of both horizontal and vertical OKANs were decreased; however, the conditions of these changes were different.

Toxoplasma gondii, unlike its mammalian host, utilizes a type II fatty acid biosynthesis pathway in which the steps of fatty acid biosynthesis are catalyzed by independent enzymes. Due to this difference, the enzymes of this pathway are good targets for the development of new therapeutic drugs directed against toxoplasmosis. In this report, we show by using reverse transcription-polymerase chain reaction analysis that beta-Hydroxyacyl-acyl carrier protein dehydratase (TgFABZ) is expressed both in tachyzoites and bradyzoites. Indirect immunofluorescence antibody test further shows the localization of TgFABZ protein in the apicoplast of both tachyzoites and bradyzoites. Enzyme dynamic analysis shows that the purified recombinant TgFABZ protein is soluble and active. The Km value of the enzyme for its substrate analog crotonoyl-CoA was estimated to be 82.57 +/- 10 microM.

We developed a sandwich ELISA for the detection of circulating Toxoplasma gondii MIC10 antigens. In T. gondii culture supernatant, MIC10 was detected in a growth dependent manner. Mice were infected with a lethal dose of either a virulent RH strain, an avirulent Beverley strain or a sub-lethal dose of a PLK strain of T. gondii. MIC10 appeared 2 days after infection and increased gradually in the sera of RH-infected mice. A detectable but significantly lower amount of MIC10 was observed in the sera of mice infected intraperitoneally with Beverley tachyzoites. In contrast, the MIC10 antigen in mice sera following oral infection with Beverley cysts was below detectable levels during the course of the experiment. In sera of PLK-infected mice, MIC10 was predominantly observed between late acute and early chronic phase. Our data show that the kinetics of circulating MIC10 differs depending on the strain and route of infection.

Recent evidence has implied that disruption of a limited number of defined cellular pathways is necessary and sufficient for neoplastic conversion of a variety of normal human cell types in tissue culture. We show instead that malignancy in such models results from an iterative process of clonal selection in vitro and/or in vivo. Normal human fibroblasts underwent malignant transformation after transduction with telomerase, cyclin-dependent kinase 4, dominant-negative p53, and activated Ras or MEK. Furthermore, culture conditions favoring overgrowth resulted in clonal selection, which with added Ras or MEK oncogenes led to the emergence of tumorigenic clones. Such tumors showed variable degrees of malignancy with some even exhibiting metastasis. SV40 small t antigen (ST) has been reported to be necessary and sufficient to convert human fibroblasts with these pathway aberrations to a polyclonal tumor. However, we observed that clonal tumors emerged even with ST addition. Genomic instability was markedly increased by p53 and Rb pathway abrogation. Under the same conditions, fibroblasts with these alterations failed to induce tumors, implying that genomic instability may be necessary but not sufficient for malignant transformation. These findings indicate that the minimum number of events required for malignant transformation of human fibroblasts is greater than has been enumerated by such oncogene addition strategies and support a stochastic cancer progression model initiated by four defined cellular alterations.

A combination of antigenic regions of microneme proteins have been previously reported as being protective against chronic toxoplasmosis. In this work, we evaluated immune responses induced by immunizing BALB/c and C57BL/6 mice intradermally with plasmid DNA encoding the protein sequences of Toxoplasma gondii AMA1, MIC2, M2AP and BAG1. Mice immunized with the AMA1 gene developed high levels of serum IgG2a and c antibodies as well as cellular immune responses associated with IFN-gamma synthesis suggesting a modulated Th1 type of response. Immunization with the AMA1 gene resulted in a partial but significant protection against the acute phase of toxoplasmosis compared to MIC2, M2AP and BAG1 genes. Therefore, the AMA1 gene appears to generate a strong specific immune response and also provides effective protection against toxoplasmosis more than the MIC2, M2AP and BAG1 genes.

Background: We developed a new enzyme-linked immunosorbent assay (ELISA) for ornithine carbamoyltransferase (OCT) and evaluated its usefulness., Methods: Recombinant human OCT expressed in E. coli was used as an antigen to obtain the monoclonal antibodies for this assay., Results: The reactivity of the antibodies to native OCT as well as recombinant OCT was enhanced at alkaline pH (8.5-10), and the assay's sensitivity was markedly improved. The antibodies react identically with native and recombinant OCT at pH 9.4. The dilution test showed a good linearity between dilution ratios and the concentrations. Different concentrations of OCT added were recovered on average at 90.4%. There was a good correlation between OCT protein levels in the ELISA and OCT enzyme activities (r=0.987, p<0.0001). A significant difference in the serum level of OCT was observed between chronic hepatitis patients (110.7+/-80 ng/ml) and healthy subjects (34.4+/-20.7 ng/ml) (p<0.0001). The serum levels of OCT between sexes differed significantly in the healthy subjects (p<0.0001)., Conclusions: Our newly established ELISA for OCT using monoclonal antibodies is sensitive enough for clinical application.

The Svalbard archipelago in arctic Norway receives considerable semivolatile organic contaminant (SOC) inputs from the atmosphere. To measure the history of net SOC accumulation there, we analyzed the upper 40 m of an ice core from Austfonna, the largest ice cap in Eurasia, for several legacy organochlorine (OC) compounds and current-use pesticides (CUPs) including organophosphorus (OP), triazine, dinitroaniline, and chloroacetamide compounds. Five OP compounds (chlorpyrifos, terbufos, diazinon, methyl parathion, and fenitrothion), two OCs (methoxychlor and dieldrin), and metolachlor--an herbicide--had historical profiles in the core. The highest OC concentration observed was aldrin (69.0 ng L(-1)) in the surface sample (1992-1998). The most concentrated OP was dimethoate (87.0 ng L(-1)) between 1986 and 1992. The surface sample also had highest concentrations of pendimethalin (herbicide, 18.6 ng L(-1)) and flutriafol, the lone observed fungicide (9.6 ng L(-1)). The apparent atmospheric persistence of CUPs likely results from little or no oxidation by OH* during the dark polar winter and in spring. Long-range atmospheric pesticide transport to Svalbard from Eurasia is influenced by the positive state of the North Atlantic Oscillation Index since 1980 and also by occasional fast-moving summer air masses from northern Eurasian croplands.

Brain and serum were collected from 120 and 12 free-ranging sika deer (Cervus nippon yesoensis), respectively, from six regions in eastern Hokkaido during controlled hunts in the autumn of 2003. Brains were tested for Neospora caninum and Toxoplasma gondii DNA by polymerase chain reaction (PCR) assays. Antibodies to Toxoplasma gondii were measured by means of a latex agglutination test. No brain tested positive for either type of DNA, and no antibody to Toxoplasma gondii was detected in serum, suggesting a low prevalence of infection with these organisms in free-ranging sika deer from eastern Hokkaido. Further examination of multiple tissues by PCR and serologic surveys will be necessary to confirm this.