Publications by authors named "Kenji Yagita"

29 Publications

  • Page 1 of 1

A Japanese case of amoebic meningoencephalitis initially diagnosed by cerebrospinal fluid cytology.

Clin Case Rep 2020 Sep 15;8(9):1728-1734. Epub 2020 Jul 15.

Department of Diagnostic Pathology (DDP) & Research Center of Diagnostic Pathology (RC-DiP) Gifu Municipal Hospital Gifu Japan.

Microscopy can detect the presence of amoebic trophozoites in cerebrospinal fluid and tissue. The infection was confirmed in the present case by polymerase chain reaction and immunohistochemistry, but we were unable to achieve a cure. Our case rapidly progressed without any skin lesions.
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http://dx.doi.org/10.1002/ccr3.2953DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7495867PMC
September 2020

Utility of the Rapid Antigen Detection Test E. histolytica Quik Chek for the Diagnosis of Entamoeba histolytica Infection in Nonendemic Situations.

J Clin Microbiol 2020 10 21;58(11). Epub 2020 Oct 21.

Department of Parasitology, National Institutes of Infectious Diseases, Tokyo, Japan.

infection is an increasingly common sexually transmitted infection in Japan. Currently, stool ova and parasite examination (O&P) is the only approved diagnostic method. Here, we assessed the utility of the commercially available rapid antigen detection test (Quik Chek) for A multicenter cross-sectional study was conducted. Stool samples that had been submitted for O&P were included. The samples were subjected to both Quik Chek and PCR, and the Quik Chek results were assessed in comparison with PCR as the reference standard. infection was confirmed in 5.8% (38/657) of the samples and comprised 20 diarrheal and 18 nondiarrheal cases. The overall sensitivity and specificity of Quik Chek were 44.7% (95% confidence interval, 30.1 to 60.3) and 99.8% (99.1 to 100), respectively. The sensitivity of Quik Chek was higher for diarrheal cases (60.0%) than for nondiarrheal cases (27.8%). Furthermore, the combined use of Quik Chek with O&P increased the sensitivity (78.9%), especially for diarrheal cases (up to 90%). The burden assessed by quantitative PCR was similar between Quik Chek-positive and -negative samples. The Quik Chek assay sensitivity was lower for cyst-containing stools than for trophozoite-containing stools, although it was shown that cultured clinical strains from Quik Chek-negative cyst-containing stools exhibited antigenicity The present study confirmed the high specificity of Quik Chek for infection. Combined use with O&P increased the sensitivity of detection, facilitating the use of Quik Chek in point-of-care settings in nonendemic situations.
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http://dx.doi.org/10.1128/JCM.01991-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7587111PMC
October 2020

Clinical features and gut microbiome of asymptomatic Entamoeba histolytica infection.

Clin Infect Dis 2020 Jun 21. Epub 2020 Jun 21.

AIDS Clinical Center, National Center of Global Health and Medicine, Tokyo, Japan.

Background: Entamoeba histolytica infection is a sexually transmitted disease in some developed countries. Asymptomatic infection often occurs and can be a source of transmission; however, limited data are available regarding the pathogenesis of E. histolytica.

Methods: This is a single center cross-sectional study. Specimens were prospectively collected from patients with clinically suspected cases. "E. histolytica infection" was defined as a case in which the identification of E. histolytica was confirmed by PCR of a clinical specimen. Data from asymptomatic cases were compared with those from symptomatic invasive cases.

Results: Sixty-four E. histolytica-infected cases, including 13 asymptomatic cases, were identified during the study period. Microbiological diagnosis was made by endoscopic sampling in 26.6% of these cases (17/64). Endoscopy identified macroscopically visible lesions in all cases; however, the sensitivity of histopathology on biopsy samples was low (45.5%) compared with PCR (94.7%). In asymptomatic cases, infection sites were limited around the proximal colon; moreover, trophozoites were frequently identified at infection sites whereas cystic forms were commonly detected in stools. Gut microbiome analyses showed more uniform composition in asymptomatic cases than in symptomatic invasive cases, which were represented by a relatively high abundance of Ruminococcaceae, Coriobacteriaceae, and Clostridiaceae, and a low abundance of Streptococcaceae.

Conclusion: These results indicate that the encystation and attenuation of E. histolytica are highly affected by the intestinal contents, including the gut microbiome.
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http://dx.doi.org/10.1093/cid/ciaa820DOI Listing
June 2020

Pathogenic free-living amoebic encephalitis in Japan.

Neuropathology 2019 Aug 26;39(4):251-258. Epub 2019 Jun 26.

Department of Pathology, Kurume University School of Medicine, Kurume, Japan.

Over 600 cases of amoebic encephalitis caused by pathogenic free-living amoebas (Balamuthia mandrillaris, Acanthamoeba spp., and Naegleria fowleri) have been reported worldwide, and in Japan, 24 cases have been reported from the first case in 1976 up to 2018. Among these cases, 18 were caused by B. mandrillaris, four by Acanthamoeba spp., one by N. fowleri, and one was of unknown etiology. Additionally, eight cases were diagnosed with encephalitis due to pathogenic free-living amoebas before death, but only three cases were successfully treated. Unfortunately, all other cases were diagnosed by autopsy. These facts indicate that an adequate diagnosis is difficult, because encephalitis due to pathogenic free-living amoebas does not show typical symptoms or laboratory findings. Moreover, because the number of cases is limited, other cases might have been missed outside of those diagnosed by autopsy. Cases of encephalitis caused by B. mandrillaris have been reported from all over Japan, with B. mandrillaris recently isolated from soil in Aomori prefecture. Therefore, encephalitis caused by pathogenic free-living amoebas should be added to the differential diagnosis of encephalitis patients.
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http://dx.doi.org/10.1111/neup.12582DOI Listing
August 2019

Microsporidial keratitis retrospectively diagnosed by ultrastructural study of formalin-fixed paraffin-embedded corneal tissue: a case report.

Ann Clin Microbiol Antimicrob 2019 Jun 10;18(1):17. Epub 2019 Jun 10.

Department of Ophthalmology, Kindai University Faculty of Medicine, 377-2, Ohnohigashi, Osakasayama, Osaka, 589-8511, Japan.

Background: The utility of formalin-fixed paraffin-embedded (FFPE) corneal tissue specimens for retrospective diagnosis of microsporidial keratitis was evaluated by transmission electron microscopy (TEM) analysis and the possible second case of microsporidial keratitis after Descemet stripping automated endothelial keratoplasty (DSAEK) was described.

Case Presentation: A 68-year-old man presented with multiple crystalline opacities in the corneal stroma that progressed extremely slowly after DSAEK. Fungiflora Y staining of corneal scrapings from the affected regions revealed an oval microorganism. Topical voriconazole administration was ineffective and penetrating keratoplasty was performed. Histological and molecular analyses were carried out on the excised cornea. Ziehl-Neelsen staining revealed an acid-fast, oval organism that was visible by ultraviolet illumination after Fungiflora Y and Uvitex 2B staining, whereas periodic acid-Schiff and Grocott's staining did not yield any significant findings. Microsporidium was detected by TEM of FFPE tissue. Nosema or Vittaforma sp. was suspected as the causative microorganism by PCR of FFPE tissue and by the fact that those species are known to cause eye infection. The corneal graft has maintained transparency at 1 year and half postoperatively.

Conclusions: This is the first known case of microsporidial keratitis diagnosed retrospectively by molecular and ultrastructural study of FFPE tissue, and the possible second case of microsporidial keratitis after DSAEK. Microsporidial keratitis should be considered when corneal opacity refractory to conventionally known therapy would occur after DSAEK. Our findings suggest that more microsporidial keratitis cases than have been reported to date can be identified by TEM or PCR examination of FFPE corneal specimens.
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http://dx.doi.org/10.1186/s12941-019-0316-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6558824PMC
June 2019

The Molecular Detection of and in Sika Deer () 
in Japan.

Food Saf (Tokyo) 2018 Jun 29;6(2):88-95. Epub 2018 Jun 29.

Department of Veterinary Public Health, Iwate University, Ueda 3-18-8, Morioka, Iwate 020-8550, Japan.

Fecal specimens (271 samples) from wild deer, , were collected from nine different areas in Japan; these samples were subjected to a real-time reverse transcription PCR for -and -specific 18S ribosomal RNA to investigate the prevalence of and infection. The incidence of and in the nine areas ranged from 0% to 20.0% and 0% to 3.4%, respectively. The prevalence of among male and female deer was 8.1% and 3.9%, respectively, while that of was 0.7% and 0.8%. Sequence analysis identified the deer genotype, , and from the sequence of specific partial 18S ribosomal RNA and assemblage A from the partial sequence of -specific 18S rRNA. The variation in regional prevalence indicates that infection depends on environmental factors, and that bovine was detected more frequently than cervine These data suggest that wild deer might be a healthy carrier of bovine .
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http://dx.doi.org/10.14252/foodsafetyfscj.2017029DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6989202PMC
June 2018

A case report of granulomatous amoebic encephalitis by Group 1 Acanthamoeba genotype T18 diagnosed by the combination of morphological examination and genetic analysis.

Diagn Pathol 2018 May 10;13(1):27. Epub 2018 May 10.

Department of Hematology and Oncology, Osaka University Graduate School of Medicine, 2-2 Yamada-Oka, Suita, Osaka, 565-0871, Japan.

Background: The diagnosis of granulomatous amoebic encephalitis is challenging for clinicians because it is a rare and lethal disease. Previous reports have indicated that Acanthamoeba with some specific genotypes tend to cause the majority of human infections. We report a case of granulomatous amoebic encephalitis caused by Acanthamoeba spp. with genotype T18 in an immunodeficient patient in Japan after allogenic bone marrow transplantation, along with the morphological characteristics and genetic analysis.

Case Presentation: A 52-year old man, who had undergone allogenic bone marrow transplantation, suffered from rapid-growing brain masses in addition to pneumonia and died within 1 month from the onset of the symptoms including fever, headache and disorientation. Infection with Acanthamoeba in the brain and lung was confirmed by histological evaluation; immunohistochemical staining and polymerase chain reaction analysis using autopsy samples also indicated the growth of Acanthamoeba in the brain. Gene sequence analysis indicated that this is the second documented case of infection with Acanthamoeba spp. with genotype T18 in a human host. Postmortem retrospective evaluation of cerebrospinal fluid sample in our case, as well as literature review, indicated that some cases of granulomatous amoebic encephalitis caused by Acanthamoeba may be diagnosable by cerebrospinal fluid examination.

Conclusion: This case indicates that Acanthamoeba spp. with genotype T18 can also be an important opportunistic pathogen. For pathologists as well as physicians, increased awareness of granulomatous amoebic encephalitis is important for improving the poor prognosis along with the attempt to early diagnosis with cerebrospinal fluid.
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http://dx.doi.org/10.1186/s13000-018-0706-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5943995PMC
May 2018

An Acute Case of Granulomatous Amoebic Encephalitis-Balamuthia mandrillaris Infection.

Intern Med 2018 May 11;57(9):1313-1316. Epub 2018 Jan 11.

Department of Parasitology, National Institute of Infectious Diseases, Japan.

A 74-year-old woman who exhibited drowsiness was referred to our hospital. Enhanced head magnetic resonance imaging (MRI) revealed multiple ring-enhancing lesions and lesions showing partial mild hemorrhaging. The patient gradually progressed to a comatose condition with notable brain deterioration of unknown cause on follow-up MRI. On day nine, the patient inexplicably died, although brain herniation was suspected. Autopsy and histopathology revealed numerous amoebic trophozoites in the perivascular spaces and within the necrotic tissue. Brain immunostaining tested positive for Balamuthia mandrillaris. Infection due to free-living amoeba is rare in Japan; however, it may increase in the near future due to unknown reasons.
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http://dx.doi.org/10.2169/internalmedicine.0011-17DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5980817PMC
May 2018

Underestimated Amoebic Appendicitis among HIV-1-Infected Individuals in Japan.

J Clin Microbiol 2017 01 28;55(1):313-320. Epub 2016 Dec 28.

AIDS Clinical Center, National Center for Global Health and Medicine, Shinjuku, Tokyo, Japan.

Entamoeba histolytica is not a common causative agent of acute appendicitis. However, amoebic appendicitis can sometimes be severe and life threatening, mainly due to a lack of awareness. Also, its frequency, clinical features, and pathogenesis remain unclear. The study subjects were HIV-1-infected individuals who presented with acute appendicitis and later underwent appendectomy at our hospital between 1996 and 2014. Formalin-fixed paraffin-embedded preserved appendix specimens were reexamined by periodic acid-Schiff (PAS) staining and PCR to identify undiagnosed amoebic appendicitis. Appendectomies were performed in 57 patients with acute appendicitis. The seroprevalence of E. histolytica was 33% (14/43) from the available stored sera. Based on the medical records, only 3 cases were clinically diagnosed as amoebic appendicitis, including 2 diagnosed at the time of appendectomy and 1 case diagnosed by rereview of the appendix after the development of postoperative complications. Retrospective analyses using PAS staining and PCR identified 3 and 3 more cases, respectively. Thus, E. histolytica infection was confirmed in 9 cases (15.8%) in the present study. Apart from a significantly higher leukocyte count in E. histolytica-positive patients than in negative patients (median, 13,760 versus 10,385 cells/μl, respectively, P = 0.02), there were no other differences in the clinical features of the PCR-positive and -negative groups. In conclusion, E. histolytica infection was confirmed in 9 (15.8%) of the appendicitis cases. However, only 3, including one diagnosed after intestinal perforation, were diagnosed before the present analyses. These results strongly suggest there is frequently a failure to detect trophozoites in routine examination, resulting in an underestimation of the incidence of amoebic appendicitis.
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http://dx.doi.org/10.1128/JCM.01757-16DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5228245PMC
January 2017

The Development of a Novel, Validated, Rapid and Simple Method for the Detection of Sarcocystis fayeri in Horse Meat in the Sanitary Control Setting.

Biocontrol Sci 2016 ;21(2):131-4

Kumamoto Prefectual Institute of Public-Health and Environmental Science.

Sarcocystis fayeri (S. fayeri) is a newly identified causative agent of foodborne disease that is associated with the consumption of raw horse meat. The testing methods prescribed by the Ministry of Health, Labour and Welfare of Japan are time consuming and require the use of expensive equipment and a high level of technical expertise. Accordingly, these methods are not suitable for use in the routine sanitary control setting to prevent outbreaks of foodborne disease. In order to solve these problems, we have developed a new, rapid and simple testing method using LAMP, which takes only 1 hour to perform and which does not involve the use of any expensive equipment or expert techniques. For the validation of this method, an inter-laboratory study was performed among 5 institutes using 10 samples infected with various concentrations of S. fayeri. The results of the inter-laboratory study demonstrated that our LAMP method could detect S. fayeri at concentrations greater than 10(4) copies/g. Thus, this new method could be useful in screening for S. fayeri as a routine sanitary control procedure.
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http://dx.doi.org/10.4265/bio.21.131DOI Listing
January 2017

Copper Homeostasis for the Developmental Progression of Intraerythrocytic Malarial Parasite.

Curr Top Med Chem 2016 ;16(27):3048-3057

Division of Tropical Diseases and Parasitology, Department of Infectious Diseases, Kyorin University School of Medicine, Tokyo 181 8611, Japan.

Malaria is one of the world's most devastating diseases, particularly in the tropics. In humans, Plasmodium falciparum lives mainly within red blood cells, and malaria pathogenesis depends on the red blood cells being infected with the parasite. Nonesterified fatty acids (NEFAs), including cis-9-octadecenoic acid, and phospholipids have been critical for complete parasite growth in serum-free culture, although the efficacy of NEFAs in sustaining the growth of P. falciparum has varied markedly. Hexadecanoic acid and trans-9-octadecenoic acid have arrested development of the parasite, in association with down-regulation of genes encoding copper-binding proteins. Selective removal of Cu+ ions has blockaded completely the ring-trophozoite-schizont progression of the parasite. The importance of copper homeostasis for the developmental progression of P. falciparum has been confirmed by inhibition of copper-binding proteins that regulate copper physiology and function by associating with copper ions. These data have provided strong evidence for a link between healthy copper homeostasis and successive developmental progression of P. falciparum. Perturbation of copper homeostasis may be, thus, instrumental in drug and vaccine development for the malaria medication. We review the importance of copper homeostasis in the asexual growth of P. falciparum in relation to NEFAs, copperbinding proteins, apoptosis, mitochondria, and gene expression.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5068492PMC
http://dx.doi.org/10.2174/1568026616999160215151704DOI Listing
February 2017

Draft Genome Sequence of High-Temperature-Adapted Protochlamydia sp. HS-T3, an Amoebal Endosymbiotic Bacterium Found in Acanthamoeba Isolated from a Hot Spring in Japan.

Genome Announc 2015 Feb 5;3(1). Epub 2015 Feb 5.

National Institute of Infectious Diseases, Shinjuku-ku, Toyama, Tokyo, Japan.

Here, we report the draft genome sequence of high-temperature-adapted Protochlamydia sp. strain HS-T3, an environmental chlamydia. This bacterium is an amoebal endosymbiont, found in Acanthamoeba isolated from a hot spring in Japan. Strain HS-T3 readily grew in mammalian cells at 37°C, a characteristic not previously reported for environmental chlamydiae.
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http://dx.doi.org/10.1128/genomeA.01507-14DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4319609PMC
February 2015

An autopsy case of Balamuthia mandrillaris amoebic encephalitis, a rare emerging infectious disease, with a brief review of the cases reported in Japan.

Neuropathology 2015 Feb 3;35(1):64-9. Epub 2014 Sep 3.

Department of Pathology & Applied Neurobiology, Graduate School of Medical Science, Kyoto Prefectural University of Medicine, Kyoto, Japan.

Balamuthia mandrillaris is an amoeba found in fresh water and soil that causes granulomatous amoebic encephalitis. We report herein an autopsy case of B. mandrillaris amoebic encephalitis, which was definitely diagnosed by PCR. An 81-year-old man, who had Sjögren's syndrome, manifested drowsiness 2 months before his death with progressive deterioration. Neuroimaging demonstrated foci of T2- and fluid-attenuated inversion recovery high and T1 low-intensity with irregular post-contrast ring enhancement in the cerebral hemisphere, thalamus and midbrain. Pathologically, multiple hemorrhagic and necrotic lesions were found in the cerebrum, thalamus, midbrain, pons, medulla and cerebellum, which were characterized by liquefactive necrosis, marked edema, hemorrhage and necrotizing vasculitis associated with the perivascular accumulation of amoebic trophozoites, a few cysts, and the infiltration of numerous neutrophils and microglia/macrophages. The trophozoites were ovoid or round, 10-60 μm in diameter, and they showed foamy cytoplasm and a round nucleus with small karyosome in the center. The PCR and immunohistochemistry from paraffin-embedded brain specimens revealed angioinvasive encephalitis due to B. mandrillaris. Human cases of B. mandrillaris brain infection are rare in Japan, with only a few brief reports in the literature.
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http://dx.doi.org/10.1111/neup.12151DOI Listing
February 2015

High-temperature adapted primitive Protochlamydia found in Acanthamoeba isolated from a hot spring can grow in immortalized human epithelial HEp-2 cells.

Environ Microbiol 2014 Feb 6;16(2):486-97. Epub 2013 Oct 6.

Department of Medical Laboratory Science, Faculty of Health Sciences, Hokkaido University, North-12, West-5, Kita-ku, Sapporo, 060-0812, Japan.

To elucidate how ancient pathogenic chlamydiae could overcome temperature barriers to adapt to human cells, we characterized a primitive chlamydia found in HS-T3 amoebae (Acanthamoeba) isolated from a hot spring. Phylogenetic analysis revealed the primitive species to be Protochlamydia. In situ hybridization staining showed broad distribution into the amoebal cytoplasm, which was supported by transmission electron microscopic analysis showing typical chlamydial features, with inclusion bodies including both elementary and reticular bodies. Interestingly, although most amoebae isolated from natural environments show reduced growth at 37°C, the HS-T3 amoebae harbouring the Protochlamydia grew well at body temperature. Although infection with Protochlamydia did not confer temperature tolerance to the C3 amoebae, the number of infectious progenies rapidly increased at 37°C with amoebal lysis. In immortalized human epithelial HEp-2 cells, fluorescence microscopic study revealed atypical inclusion of the Protochlamydia, and quantitative real-time polymerase chain reaction analyses also showed an increase in 16S ribosomal RNA DNA amounts. Together, these results showed that the Protochlamydia found in HS-T3 amoebae isolated from a hot spring successfully adapted to immortalized human HEp-2 cells at 37°C, providing further information on the evolution of ancient Protochlamydia to the present pathogenic chlamydiae.
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http://dx.doi.org/10.1111/1462-2920.12266DOI Listing
February 2014

Identification of photoactivated adenylyl cyclases in Naegleria australiensis and BLUF-containing protein in Naegleria fowleri.

J Gen Appl Microbiol 2013 ;59(5):361-9

Academic Group of Applied Life Sciences, Iwate University.

Complete genome sequencing of Naegleria gruberi has revealed that the organism encodes polypeptides similar to photoactivated adenylyl cyclases (PACs). Screening in the N. australiensis genome showed that the organism also encodes polypeptides similar to PACs. Each of the Naegleria proteins consists of a "sensors of blue-light using FAD" domain (BLUF domain) and an adenylyl cyclase domain (AC domain). PAC activity of the Naegleria proteins was assayed by comparing sensitivities of Escherichia coli cells heterologously expressing the proteins to antibiotics in a dark condition and a blue light-irradiated condition. Antibiotics used in the assays were fosfomycin and fosmidomycin. E. coli cells expressing the Naegleria proteins showed increased fosfomycin sensitivity and fosmidomycin sensitivity when incubated under blue light, indicating that the proteins functioned as PACs in the bacterial cells. Analysis of the N. fowleri genome revealed that the organism encodes a protein bearing an amino acid sequence similar to that of BLUF. A plasmid expressing a chimeric protein consisting of the BLUF-like sequence found in N. fowleri and the adenylyl cyclase domain of N. gruberi PAC was constructed to determine whether the BLUF-like sequence functioned as a sensor of blue light. E. coli cells expressing a chimeric protein showed increased fosfomycin sensitivity and fosmidomycin sensitivity when incubated under blue light. These experimental results indicated that the sequence similar to the BLUF domain found in N. fowleri functioned as a sensor of blue light.
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http://dx.doi.org/10.2323/jgam.59.361DOI Listing
April 2014

Genetic characterization of clinical acanthamoeba isolates from Japan using nuclear and mitochondrial small subunit ribosomal RNA.

Korean J Parasitol 2013 Aug 30;51(4):401-11. Epub 2013 Aug 30.

Department of Parasitology, Graduate School of Medical Science, Kanazawa University, Kanazawa, Ishikawa, Japan.

Because of an increased number of Acanthamoeba keratitis (AK) along with associated disease burdens, medical professionals have become more aware of this pathogen in recent years. In this study, by analyzing both the nuclear 18S small subunit ribosomal RNA (18S rRNA) and mitochondrial 16S rRNA gene loci, 27 clinical Acanthamoeba strains that caused AK in Japan were classified into 3 genotypes, T3 (3 strains), T4 (23 strains), and T5 (one strain). Most haplotypes were identical to the reference haplotypes reported from all over the world, and thus no specificity of the haplotype distribution in Japan was found. The T4 sub-genotype analysis using the 16S rRNA gene locus also revealed a clear sub-conformation within the T4 cluster, and lead to the recognition of a new sub-genotype T4i, in addition to the previously reported sub-genotypes T4a-T4h. Furthermore, 9 out of 23 strains in the T4 genotype were identified to a specific haplotype (AF479533), which seems to be a causal haplotype of AK. While heterozygous nuclear haplotypes were observed from 2 strains, the mitochondrial haplotypes were homozygous as T4 genotype in the both strains, and suggested a possibility of nuclear hybridization (mating reproduction) between different strains in Acanthamoeba. The nuclear 18S rRNA gene and mitochondrial 16S rRNA gene loci of Acanthamoeba spp. possess different unique characteristics usable for the genotyping analyses, and those specific features could contribute to the establishment of molecular taxonomy for the species complex of Acanthamoeba.
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http://dx.doi.org/10.3347/kjp.2013.51.4.401DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3770870PMC
August 2013

Detection and genotype of Encephalitozoon cuniculi DNA from urine and feces of pet rabbits in Japan.

J Vet Med Sci 2013 29;75(8):1017-20. Epub 2013 Mar 29.

Laboratory of Veterinary Parasitology, Faculty of Agriculture, Iwate University, 3-18-8 Ueda, Morioka 020-8550, Japan.

A newly developed nested polymerase chain reaction (PCR) assay targeting the internal transcribed spacer (ITS) region of the ribosomal DNA was applied to detect and characterize Encephalitozoon cuniculi DNA from pet rabbits in Japan. The analysis was carried out using 257 urinary samples and 314 fecal samples collected from 307 pet rabbits in the age group of 1 month to 12 years from 30 different prefectures of Japan and 107 fecal samples and 3 urinary samples collected from 1-month-old rabbits from 3 breeding facilities in Japan. We detected 840-bp amplicons in 20 urinary samples (7.78%) from the pet rabbits of the 13 prefectures and in 1 urinary (33.3%) and 6 fecal (5.6%) samples from the rabbits of the 2 breeding facilities. The sequences (803 bp) of the 27 amplicons had no variations and completely coincided with the sequence of E. cuniculi genotype I. To the best of our knowledge, this is the first report on the detection and genotype characterization of E. cuniculi DNA from pet rabbits in Japan.
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http://dx.doi.org/10.1292/jvms.12-0567DOI Listing
April 2014

Cytopathic effect of Acanthamoeba on human corneal fibroblasts.

Mol Vis 2012 9;18:2221-8. Epub 2012 Aug 9.

Department of Ophthalmology, Tokyo Women's Medical University Medical Center East, Tokyo Japan.

Purpose: Acanthamoeba keratitis is associated with keratocyte depletion in humans. We investigated how Acanthamoebae isolated from corneas affected by Acanthamoeba keratitis interacted with human corneal stromal cells in vitro.

Methods: Acanthamoebae were isolated from 6 patients with Acanthamoeba keratitis and genotyping was done. Whether the isolated Acanthamoebae could invade the corneal stroma was assessed with denuded corneal stroma ex vivo. The cytopathic effect of Acanthamoeba on cultured corneal fibroblasts from donor corneas was quantitatively evaluated by the MTT assay after culture under various conditions. Terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) and Annexin V staining were employed to detect apoptotic cells among the corneal fibroblasts co-cultured with Acanthamoebae.

Results: All 6 Acanthamoebae isolated from the patients with Acanthamoeba keratitis were shown to have the T4 genotype by 18S rDNA sequence analysis. Acanthamoebae invaded the denuded corneal stroma in the ex vivo experiments and had a cytopathic effect on human corneal fibroblasts after direct adhesion, but not via chemical mediators. A cytopathic effect was detected with all 6 Acanthamoebae and corneal fibroblasts mainly died by apoptosis, as evidenced by Annexin V staining.

Conclusions: Acanthamoebae isolated from patients with Acanthamoeba keratitis had a cytopathic effect on human corneal fibroblasts, mainly via induction of apoptosis after direct adhesion. Our findings may provide some clues to the pathophysiology of corneal keratocyte depletion in patients with Acanthamoeba keratitis.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3429359PMC
December 2012

Depletion of Cryptosporidium parvum oocysts from contaminated sewage by using freshwater benthic pearl clams (Hyriopsis schlegeli).

Appl Environ Microbiol 2012 Oct 17;78(20):7420-8. Epub 2012 Aug 17.

Section of Drinking Water Chemistry, Division of Environmental Hygiene, Hokkaido Institute of Public Health, Sapporo, Japan.

The freshwater benthic pearl clam, Hyriopsis schlegeli, was experimentally exposed to Cryptosporidium parvum oocysts, and it was verified that the oocysts were eliminated predominantly via the fecal route, retaining their ability to infect cultured cells (HCT-8). The total fecal oocyst elimination rate was more than 90% within 5 days after exposure to the oocysts. H. schlegeli was able to survive in the final settling pond of a sewage plant for long periods, as confirmed by its pearl production. In the light of these findings, the clam was placed in the final settling pond in a trial to test its long-term efficacy in depleting oocysts contaminating the pond water. The number of clams placed was set to ensure a theoretical oocyst removal rate of around 50%, and the turbidity and the density of feed microbes in the overflow trough water of the pond were about 35% and 40 to 60% lower, respectively, than in the control water throughout the year. It was found that the clam feces containing oocysts were sufficiently heavy for them to settle to the bottom of the pond, despite the upward water flow. From these results, we concluded that efficient depletion of oocysts in the sewage water of small or midscale sewage treatment plants can be achieved by appropriate placement of H. schlegeli clams.
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http://dx.doi.org/10.1128/AEM.01502-12DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3457110PMC
October 2012

Assessment of real-time polymerase chain reaction detection of Acanthamoeba and prognosis determinants of Acanthamoeba keratitis.

Ophthalmology 2012 Jun 3;119(6):1111-9. Epub 2012 Mar 3.

Division of Ophthalmology and Visual Science, Faculty of Medicine, Tottori University, Tottori, Japan.

Objective: To evaluate the diagnostic value of real-time polymerase chain reaction (PCR) for detecting Acanthamoeba in eyes diagnosed with Acanthamoeba keratitis (AK) by conventional tests. In addition, to determine the preoperative prognosis-determining factors in eyes with AK.

Design: Retrospective, cross-sectional study.

Participants: A total of 104 eyes of 103 patients who were diagnosed with AK or with bacterial or bacteria-associated keratitis (BK) by conventional tests.

Methods: Twenty-nine eyes with AK and 75 eyes with BK were evaluated for Acanthamoeba and bacterial DNA by real-time PCR. The Acanthamoeba copy numbers, bacterial load, and clinical parameters in the patients with AK were assessed for those significantly associated with poor outcome, that is, final visual acuity of <20/50 or requiring keratoplasty, by logistic regression analysis.

Main Outcome Measures: Acanthamoeba DNA copy number, bacterial DNA copy number, and odds ratio (OR) for poor prognosis.

Results: The detection of amoebic DNA was 50 times more sensitive by real-time PCR than by conventional cyst counting. The Acanthamoeba copy numbers at the first visit (mean: 4.7×10(5)±3.2×10(5) copies) were significantly correlated with the AK stage, and both were significant risk factors for a poor outcome. The Acanthamoeba DNA copy numbers at the first visit and AK stage had a significantly high risk for poor outcome (OR of Acanthamoeba DNA copy per logarithm of copy numbers: 3.48, 95% confidence interval [CI], 1.04-111.63, P<0.05; OR of AK stage: 2.8 per stage increase, 95% CI, 1.07-7.30, P<0.05, after adjustment of age). In the AK cases with poor outcome, the amoebic DNA was not reduced by more than 90% after 1 month of treatment. The weak amoebic reduction was significantly associated with advanced AK stages or previous use of steroids. Bacterial 16S rDNA was detected in 53.6% of the eyes with AK, but it was not associated with any risk for refractoriness.

Conclusions: Real-time PCR was effective in detecting and managing AK. The Acanthamoeba copy number and AK stage at the first visit were significantly associated with poor outcome.

Financial Disclosure(s): The author(s) have no proprietary or commercial interest in any materials discussed in this article.
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http://dx.doi.org/10.1016/j.ophtha.2011.12.023DOI Listing
June 2012

Multisite performance evaluation of an enzyme-linked immunosorbent assay for detection of Giardia, Cryptosporidium, and Entamoeba histolytica antigens in human stool.

J Clin Microbiol 2012 May 29;50(5):1762-3. Epub 2012 Feb 29.

Division of Infectious Diseases and International Health, University of Virginia School of Medicine, Charlottesville, Virginia, USA.

A novel fecal antigen detection assay for fresh and frozen human samples that detects but does not differentiate Giardia spp, Cryptosporidium spp, and Entamoeba histolytica, the Tri-Combo parasite screen, was compared to three established enzyme-linked immunosorbent assays (ELISAs) at three international sites. It exhibited 97.9% sensitivity and 97.0% specificity, with positive and negative predictive values of 93.4% and 99.1%, respectively. The Tri-Combo test proved a reliable means to limit the use of individual parasite ELISAs to positive samples.
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http://dx.doi.org/10.1128/JCM.06483-11DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3347128PMC
May 2012

Silent information regulator 2 proteins encoded by Cryptosporidium parasites.

Parasitol Res 2010 Aug 19;107(3):707-12. Epub 2010 Jun 19.

Graduate School of Science and Engineering, University of Toyama, Toyama, Japan.

Screening in a database has revealed that Cryptosporidium hominis encodes a silent information regulator 2 (Sir2), a nicotinamide adenine dinucleotide (NAD)-dependent protein deacetylase. Cellular localization of the protein, ChSir2, was analyzed by the use of the social amoeba Dictyostelium discoideum as a model system. Fluorescent microscopic analysis showed that ChSir2 fused with green fluorescent protein was localized in the D. discoideum nucleus. D. discoideum expressing ChSir2 grew faster and reached higher cell density than did D. discoideum harboring a control vector. These results suggest that ChSir2 is a nucleus-localizing protein that plays an important role in the growth of C. hominis. We cloned and sequenced the genes for Sir2 orthologs encoded by three isolates of C. hominis, two isolates of Cryptosporidium parvum and one isolate of Cryptosporidium meleagridis. The orthologs conserve critical catalytic or NAD-binding residues but do not have similarity with human Sir2 proteins (SIRTs). Cryptosporidium Sir2 orthologs would therefore be attractive therapeutic targets. The Cryptosporidium orthologs were classified into four variants based on their nucleotide sequences. Each of the four variants produces its own unique restriction fragment length polymorphism pattern by digestion with TfiI.
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http://dx.doi.org/10.1007/s00436-010-1925-8DOI Listing
August 2010

[Legionella contamination risk factors in non-circulating hot spring water].

Kansenshogaku Zasshi 2009 Jan;83(1):36-44

Ehime Prefectural Institute of Public Health and Environmental Science.

We examined water from 182 non-circulating hot spring bathing facilities in Japan for possible Legionella occurrence from June 2005 to December 2006, finding Legionella-positive cultures in 119 (29.5%) of 403 samples. Legionellae occurrence was most prevalent in bathtub water (39.4%), followed by storage tank water (23.8%), water from faucets at the bathtub edge (22.3%), and source-spring water (8.3%), indicating no statistically significant difference, in the number of legionellae, having an overall mean of 66 CFU/100mL. The maximum number of legionellae in water increased as water was sampled downstream:180 CFU/100 mL from source spring, 670 from storage tanks, 4,000 from inlet faucets, and 6,800 from bathtubs. The majority--85.7%--of isolated species were identified as L. pneumophila : L. pneumophila serogroup (SG) 1 in 22%, SG 5 in 21%, and SG 6 in 22% of positive samples. Multivariate logistic regression models used to determine the characteristics of facilities and sanitary management associated with Legionella contamination indicated that legionellae was prevalent in bathtub water under conditions where it was isolated from inlet faucet/pouring gate water (odds ratio [OR] = 6.98, 95% confidence interval [CI] = 2.14 to 22.8). Risk of occurrence was also high when the bathtub volume exceeded 5 m3 (OR = 2.74, 95% CI = 1.28 to 5.89). Legionellae occurrence was significantly reduced when the bathing water pH was lower than 6.0 (OR = 0.12, 95% CI = 0.02 to 0.63). Similarly, occurrence was rare in inlet faucet water or the upper part of the plumbing system for which pH was lower than 6.0 (OR = 0.06, 95% CI = 0.01 to 0.48), and when the water temperature was maintained at 55 degrees C or more (OR = 0.10, 95% CI = 0.01 to 0.77). We also examined the occurrence of amoeba, Mycobacterium spp., Escherichia coli, Pseudomonas aeruginosa, and Staphylococcus aureus in water samples.
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http://dx.doi.org/10.11150/kansenshogakuzasshi.83.36DOI Listing
January 2009

Occurrence of Cryptosporidium sp. in snakes in Japan.

Parasitol Res 2008 Sep 13;103(4):801-5. Epub 2008 Jun 13.

Department of Microbiology, Kanagawa Prefectural Institute of Public Health, 1-3-1 Shimomachiya, Chigasaki, Kanagawa, 253-0087, Japan,

The aim of this study was to determine the prevalence of Cryptosporidium in snakes in Japan. Fecal samples or intestinal contents of 469 snakes, consisting of five species, were analyzed and Cryptosporidium oocysts were detected only from the Japanese grass snake Rhabdophis tigrinus. The mean prevalence of Cryptosporidium sp. in Japanese grass snakes was approximately 26% in the region studied. Histopathological observations revealed that the organism caused proliferative enteritis in the small intestine. Sequence analysis of a fragment of the small subunit rRNA gene has shown that the partial sequence of Cryptosporidium sp. isolated from the snakes was identical to that of the Cryptosporidium snake genotype W11 from New Guinea viper boa.
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http://dx.doi.org/10.1007/s00436-008-1045-xDOI Listing
September 2008

Genotyping of Giardia intestinalis from domestic and wild animals in Japan using glutamete dehydrogenase gene sequencing.

Vet Parasitol 2005 Nov;133(4):283-7

Laboratory of Veterinary Parasitology, Faculty of Agriculture, Iwate University, 3-18-8 Ueda, Morioka 020-0105, Japan.

To determine the genotypes of Giardia intestinalis from domestic and wild animals in Japan, Giardia isolates obtained from feces of 24 dogs kept in households and breeding kennels, three companion cats, five dairy calves and three wild monkeys, Macaca fuscata, were genotyped using the 177 bp sequence of the glutamete dehydrogenase gene (gdh). The genotypes were assemblages A, C, D or A/D for dog isolates, Assemblage F for cat isolates, assemblages A or E for calf isolates and assemblage B for monkey isolates. This is the first report on the genotypes of Giardia isolates from cats, calves and wild monkeys in Japan.
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http://dx.doi.org/10.1016/j.vetpar.2005.05.061DOI Listing
November 2005

[The largest outbreak of legionellosis in Japan associated with spa baths: epidemic curve and environmental investigation].

Kansenshogaku Zasshi 2005 Jun;79(6):365-74

Miyazaki Prefectural Institute for Public Health and Environment.

In July 2002, a large outbreak of legionellosis occurred in a bathhouse with spa facilities in Miyazaki Prefecture. Two hundred-ninety-five patients (including suspected cases) that had pneumonia and/or symptoms of fever, cough and so forth were reported; 37% of them were hospitalized and seven people died. In environmental investigations, Legionella pneumophila serogroups (SGs) land 8, L. dumoffii, L. londiniensis, some other Legionella species and many kinds of amoeba were isolated from 55 samples of bathtub water, tank water, filters and so forth in the spa facilities. The dominant isolates from the bathtab waters belonged to L. londiniensis, L. dumoffii and L. pneumophila SG1, and their maximum concentrations were 1.5 x 10(6), 5.2 x 10(5) and 1.6 x 10(5) cfu/100 mL, respectively. L. pneumophila SG1 strains isolated from bathtub water, tank water, filters and sputa of patients showed a indistinguishable DNA fingerprint pattern by pulsed-field gel electrophoresis (PFGE), confirming that the source of infection was the spa water. Our study indicate that spas may be a significant health hazard if hygienic management fails.
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http://dx.doi.org/10.11150/kansenshogakuzasshi1970.79.365DOI Listing
June 2005

Occurrence and distribution of Naegleria species in thermal waters in Japan.

J Eukaryot Microbiol 2003 ;50 Suppl:514-5

Department of Parasitology, National Institute of Infectious Diseases, Toyama 1-23-1, Shinjyuku-ku, Tokyo 162-8640, Japan.

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http://dx.doi.org/10.1111/j.1550-7408.2003.tb00614.xDOI Listing
March 2004

[Giardiasis].

Nihon Rinsho 2003 Feb;61 Suppl 2:618-22

Department of Parasitology, National Institute of Infectious Diseases.

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February 2003

[Protozoa and food hygiene].

Shokuhin Eiseigaku Zasshi 2002 Dec;43(6):J343-7

National Institute of Infectious Diseases: 1-23-1, Toyama, Shinjuku-ku, Tokyo 162-8640, Japan.

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December 2002