A Comprehensive Study of the Immunophenotype and its Clinicopathologic Significance in Adult T-Cell Leukemia/Lymphoma.

Adult T-cell leukemia/lymphoma (ATLL) is a mature T-cell tumor caused by human T-lymphotropic virus type 1 (HTLV-1). The typical ATLL immunophenotypes are described in the 2017 World Health Organization Classification of Tumours of Haematopoietic and Lymphoid Tissues (positive: CD2, CD3, CD5, CD4, and CD25; negative: CD7, CD8, and cytotoxic markers; and partially positive: CD30, CCR4, and FOXP3). However, limited studies are available on the expression of these markers, and their mutual relationship remains unknown. Furthermore, the expression status of novel markers associated with T-cell lymphomas, including Th1 markers (T-bet and CXCR3), Th2 markers (GATA3 and CCR4), T follicular helper markers (BCL6, PD1, and ICOS), and T-cell receptor (TCR) markers, and their clinicopathologic significance is unclear. In this study, we performed >20 immunohistochemical stains in 117 ATLL cases to determine the comprehensive immunophenotypic profile of ATLL, which were compared on the basis of clinicopathologic factors, including morphologic variants (pleomorphic vs anaplastic), biopsy locations, treatments, Shimoyama classification-based clinical subtype, and overall survival. CD3+/CD4+/CD25+/CCR4+ was considered a typical immunophenotype of ATLL, but approximately 20% of cases did not conform to this pattern. Simultaneously, the following new findings were obtained: (1) most cases were negative for TCR-ß and TCR-δ (104 cases, 88.9%), indicating the usefulness of negative conversion of TCR expression to provide differentiation from other T-cell tumors; (2) the positivity of CD30 and CD15 and the negativity of FOXP3 and CD3 were significantly associated with anaplastic morphology; and (3) atypical cases, such as T follicular helper marker-positive (12 cases, 10.3%) and cytotoxic molecule-positive cases (3 cases, 2.6%), were identified. No single markers could predict the overall survival among patients with acute/lymphoma subtypes of ATLL. The results of this study illustrate the diversity of ATLL phenotypes. In T-cell tumors occurring in HTLV-1 carriers, the possibility of ATLL should not be eliminated even when the tumor exhibits an atypical phenotype, and the confirmation of HTLV-1 in the tissue is recommended.

Recurrent malignant peripheral nerve sheath tumor presenting as an asymptomatic intravenous thrombus extending to the heart: a case report.

BACKGROUND: Malignant peripheral nerve sheath tumor (MPNST) is a rare soft tissue sarcoma mainly treated via surgical resection. Herein, we report a case of MPNST wherein a massive tumor thrombus extended to the major veins and heart. CASE PRESENTATION: A 39-year-old female with a history of neurofibromatosis type 1 developed MPNST from the right radial nerve. In addition to adjuvant chemotherapy, she underwent wide tumor resection and concomitant radial nerve resection, followed by postoperative radiotherapy. Histological evaluation revealed marked venous invasion. The 2-year follow-up CT revealed an asymptomatic recurrent tumor thrombus extending from the right subclavian vein to the heart. An urgent life-saving operation was performed to ligate the base of the right subclavian vein and remove the entire intravenous thrombus that extended to the right ventricle. The remaining tumor in the right subclavian vein increased in size 3 months after thrombectomy. After confirming the absence of any metastatic lesions, the patient underwent extended forequarter amputation to achieve surgical remission. One year later, a new metastasis to the right diaphragm was safely resected. The patient remains alive without any evidence of disease 2 years after the extended forequarter amputation. CONCLUSIONS: In cases of a previous history of microscopic venous invasion, recurrence can occur as a massive tumor thrombus that extends to the great vessels.

Effects of collaborative consultation using iPad application in school-based occupational therapy: A single-arm pre-post pilot study.

INTRODUCTION: This pilot study examined the effect of collaborative consultation with teachers in school-based occupational therapy (SBOT) using iPad application for goal-setting-Aid for Decision-Making in Occupation Choice for School (ADOC-S). METHOD: A single arm pre-post intervention design was conducted at four preschools in Japan. The participants were 10 preschool teachers (10.5 ± 0.4 years of teaching), and 10 children (3.8 ± 0.4 years old; five boys). The teachers received three collaborative consultations (1 hr for each) from one occupational therapist. Firstly, the occupational therapist conducted an observational assessment of the child's school life, then the teacher and occupational therapist identified occupation-based goals for each child using the ADOC-S. Secondly, the teacher and the occupational therapist developed the graded goals using the Goal Attainment Scale (GAS) and discussed the daily intervention plan for the teachers to conduct. Thirdly, the occupational therapist followed up on the intervention process using GAS. The subjective outcome was measured at 10 point-satisfaction and performance score in each goal using Canadian Occupational Performance Measure (COPM). The observed outcome was measured using GAS and Strengths and Difficulties Questionnaire (SDQ). RESULTS: All the participants including teachers and children successfully underwent the intervention process. Significant improvement was found pre-post and effect size (ES) was large on the COPM (ES; r = .85-.94), GAS (ES; r = .98-.99) and SDQ (ES; r = .64). All goals (30/30) exceeded expectations of GAS from zero to +2. CONCLUSION: Although there are some limitations and considerations which need to be resolved for future studies, our results suggest that ADOC-S would help build a rapport with a teacher in SBOT settings, and this collaborative consultation would facilitate a positive outcome.

Dermatopathic reaction of lymph nodes in HTLV-1 carriers: a spectrum of reactive and neoplastic lesions.

AIMS: Dermatopathic reaction is a histopathological finding of lymph nodes that usually occurs in patients with inflammatory pruritic cutaneous lesions. However, it is sometimes seen in patients with cutaneous T cell lymphoma. Adult T cell leukaemia/lymphoma (ATLL) is a T cell malignancy caused by infection with human T cell leukaemia virus type I (HTLV-1), which is frequently accompanied by cutaneous lesions. However, the detailed clinicopathological characteristics of the dermatopathic reaction of lymph nodes in ATLL patients and HTLV-1 carriers, addressed in this study, remains to be clarified. METHODS AND RESULTS: We retrospectively analysed 18 nodal lesions with dermatopathic reaction in HTLV-1 carriers. Axillary and inguinal lymph nodes were the primary affected tissues. Three cases with atypical lymphoid cell infiltration were defined as ATLL with dermatopathic reaction (ATLL-D), showing an abnormal T cell immunophenotype and T cell monoclonality. Two of the three ATLL-D patients died 14 and 7 months after diagnosis (the third case had a very short follow-up). The other 15 patients were indistinguishable from reactive lesions and were defined as HTLV-1-associated lymphadenitis with dermatopathic reaction (HAL-D). They showed an indolent clinical course, with only one case eventually transforming to aggressive disease. CONCLUSIONS: Lymph node lesions accompanied by dermatopathic reaction in HTLV1 carriers represent a spectrum that includes reactive and neoplastic conditions. HAL-D should be distinguished from ATLL-D, especially to avoid overtreatment.

Seroprevalence of Antibodies Against Paracoccidioides Spp. in Captive Dolphins from Three Aquaria in Japan.

The skin disease paracoccidioidomycosis ceti occurs in several dolphin species globally. Infection by the unculturable fungi Paracoccidioides brasilensis or other Paracoccidioides spp. results in chronic cutaneous and granulomatous lesions. In this study we used immunohistochemistry to investigate the seroprevalence of antibodies to Paracoccidioides spp. in captive dolphins from three aquaria in Japan. We had previously reported that there were serological cross-reactions for Paracoccidioides spp. with related species in the order Onygenales. We hypothesized that the degree of serological cross-reactions for Paracoccidioides spp. might be lower in areas, such as Japan, where the fungal diseases coccidiodomycosis and paracoccidiodomycosis are not endemic. Sera from 41 apparently healthy dolphins, including 20 Atlantic bottlenose dolphins (BD: Tursiops truncatus), 6 Indo-Pacific bottlenose dolphins (IPBD: Tursiops aduncus), 2 F1 generation of a cross between BD and IPBD (F1), 3 Pacific white-sided dolphins (PWD: Lagenorhynchus obliquidens), 2 pantropical spotted dolphins (PSD: Stenella attenuata), 6 false killer whales (FKW: Pseudorca crassidens), and 2 rough-toothed dolphins (RTD: Steno bredanensis) were investigated. Sera from three dolphins with paracoccidioidomycosis ceti were used as a positive control. The yeast-form cells of Paracoccidioides spp. in the cutaneous tissue sample derived from the first Japanese paracoccidioidomycosis ceti case were used as the antigen for the immunohistochemistry. Of the 41 dolphins tested, 61.0% had antibodies against Paracoccidioides spp. This indicates that dolphins of several species in Japanese aquaria have likely been exposed to the pathogen Paracoccidioides spp.

Wide-spread ignorance on the treatment of subcutaneous malignant tumors; a questionnaire-based study.

BACKGROUND: Subcutaneous malignant tumors are often treated by non-specialized clinicians in musculoskeletal oncology. While the resection of subcutaneous tumors appears technically feasible, unplanned resection of malignant tumors can result in a devastating clinical outcome. The aim of this study was to evaluate the potential estrangement in the awareness of and the treatment strategy for the patients with subcutaneous soft tissue tumors between musculoskeletal oncologists and non-specialized clinicians. METHODS: A questionnaire probing the clinical assessment of subcutaneous tumors was sent to orthopedic surgeons, dermatologists, plastic surgeons, and general surgeons. Results of the questionnaire were statistically analyzed. RESULTS: One hundred sixteen clinicians out of 150 responded to the questionnaire; the response rate was 77.3%. Among those, 46 clinicians had treated subcutaneous tumors. Thirty-nine of these 46 clinicians (27 musculoskeletal oncologists and 12 non-specialized clinicians) preoperatively performed enhanced MRI for diagnostic evaluation. Preoperative incisional biopsy to confirm the pathological diagnosis was performed by 36 of the 46 clinicians (29 musculoskeletal oncologists and seven non-specialized clinicians). These results indicate that musculoskeletal oncologists perform preoperative enhanced MRI (P = 0.08) and biopsy (P < 0.01) more frequently than non-specialized clinicians. The recognition rate of 'myxofibrosarcoma' was 60.8% among clinicians with an experience with sarcoma treatment (25 musculoskeletal oncologists and three non-specialized clinicians). The recognition rate of 'myxofibrosarcoma' between musculoskeletal oncologists and non-specialized clinicians was statistically significant (P < 0.01). CONCLUSIONS: Preoperative evaluations for subcutaneous tumors are more often inappropriate in non-specialized clinicians than those who are. Therefore, it will be mandatory to raise the awareness of this condition to improve the clinical outcome of patients with subcutaneous tumors.

Immunohistochemical Cross-Reactivity Between Paracoccidioides sp. from Dolphins and Histoplasma capsulatum.

Paracoccidioidomycosis ceti is a cutaneous disease of cetaceans caused by uncultivated Paracoccidioides brasiliensis or Paracoccidioides spp. Serological cross-reactions between paracoccidioidomycosis ceti and paracoccidioidomycosis, paracoccidioidomycosis and histoplasmosis, and paracoccidioidomycosis and coccidioidomycosis have been reported before. The present study aimed to detect immunohistochemical cross-reaction between antibodies to Paracoccidioides sp. and Histoplasma capsulatum, and vice versa. Thirty murine sera, obtained from experimental infections of 6 isolates of H. capsulatum, were reacted with paraffin-embedded yeast-form cells of Paracoccidioides sp. derived from a case of paracoccidioidomycosis ceti in Japan. The murine sera were also reacted with human isolates of H. capsulatum yeast cells, with P. brasiliensis yeast cells, and with fungal cells of Coccidioides posadasii. Three dolphins' sera from cases of paracoccidioidomycosis ceti, two human sera from patients with paracoccidioidomycosis, and a serum from a healthy person with a history of coccidioidomycosis were used in order to determine that the tested fungal cells reacted properly. Sera derived from mice infected with an isolate of H. capsulatum reacted positively against yeast cells of Paracoccidioides sp., yeast cells of P. brasiliensis, and fungal cells of C. posadasii, while those derived from other strains were negative. The present study recorded for the first time the cross-reaction between the yeast cells of H. capsulatum and antibodies against Paracoccidioides spp., the yeast cells of Paracoccidioides sp. and antibodies against H. capsulatum, the yeast cells of Paracoccidioides sp. and antibodies against Coccidioides sp., and fungal cells of C. posadasii and antibodies against Paracoccidioides spp.

Depletion of Epidermal Langerhans Cells in the Skin Lesions of Pellagra Patients.

Pellagra is a nutrient deficiency disease caused by insufficient niacin levels. Recent studies have shown that numbers of epidermal Langerhans cells decreased in other diseases caused by nutritional deficiencies, including necrolytic migratory erythema and acrodermatitis enteropathica. Epidermal Langerhans cells are capable of modulating or even halting the inflammatory reaction. The aim of this study was to examine changes in the number of Langerhans cells and other dendritic cells, and maturation of epidermal Langerhans cells in the lesional and adjacent non-lesional skin in pellagra patients. Seven pellagra patients and 10 healthy individuals who served as controls were included. The number and distribution of dendritic cells and other cutaneous cells were examined by immunohistochemistry. Epidermal Langerhans cells decreased considerably in the skin lesions of pellagra patients, whereas other dendritic cells did not change. The decrease in the number of Langerhans cells was positively correlated with the histological severity of skin lesions. As the number of Langerhans cells was not reduced in the undisturbed neighboring skin, the depletion of epidermal Langerhans cells did not precede skin damage but was a cause of prolonged severe inflammation.

Fecal calprotectin is a clinically relevant biomarker of mucosal healing in patients with quiescent ulcerative colitis.

BACKGROUND AND AIM: Calprotectin is an abundant protein in neutrophils, which infiltrate the mucosa during inflammation. Fecal calprotectin (FC) level has shown correlation with disease activity in ulcerative colitis (UC) patients. Additionally, FC level is expected to indicate mucosal healing (MH). This study was to see the significance of FC for predicting MH in patients with quiescent UC. METHODS: A total of 112 patients with quiescent UC were included. After taking blood and stool samples, patients underwent total colonoscopy, and the Mayo endoscopic subscore was recorded. FC was measured by fluorescence enzyme immunoassay. C-reactive protein, hemoglobin, erythrocyte sedimentation rate, and serum albumin were measured as conventional biomarkers. MH was defined as Mayo 0 or 0 and 1, and receiver-operator characteristic analyses were undertaken to determine the significance levels of measurements. RESULTS: Data from 105 patients were available. Eleven patients showed Mayo ≥ 2. The median (interquartile range) of FC level of all patients was 115 µg/g (45.4-420). The area under the curve (AUC) in receiver operator characteristic analysis of FC to predict Mayo 0 and 1 was 0.869 with a cut-off value of 200 µg/g yielding 67% sensitivity and 91% specificity, which were the best among all biomarkers. However, the power of FC to predict Mayo 0 was modest; the AUC was 0.639 and cut-off value 194 µg/g with 71% sensitivity and 58% specificity. CONCLUSIONS: Based on the findings of this study, we believe that FC is a clinically relevant biomarker of MH in patients with quiescent UC. Other favorable features of FC test include feasibility and non-invasiveness.

Butyrate attenuates lipolysis in adipocytes co-cultured with macrophages through non-prostaglandin E2-mediated and prostaglandin E2-mediated pathways.

BACKGROUND: Interactions between adipocytes and macrophages are associated with metabolic disorders. Production of pro-inflammatory mediators and the release of free fatty acids (FFAs) increase when these cells are co-cultured; butyrate significantly diminishes these effects by suppressing both the macrophage inflammatory and adipocyte lipolysis pathways. Butyrate is known to up-regulate the expression of prostaglandin E2 (PGE2). Therefore, we hypothesized that PGE2 is associated with the suppression of lipolysis by butyrate in co-culture. METHODS: Using contact or transwell co-culture methods with differentiated 3T3-L1 adipocytes and RAW264.7 macrophages, we investigated the effects of butyrate on the release of PGE2 into the medium and on lipolysis in adipocytes. To elucidate the underlying mechanism, we examined the effects of butyrate on cyclooxygenase-2 (COX2) and phospholipase A2 (PLA2) in co-cultured cells, and cyclic adenine monophosphate (cAMP) and protein kinase A type 1-α regulatory subunit (PRKAR1A) in co-cultured adipocytes. Silent interfering (si)RNA targeting of G-protein-coupled receptor (GPR)41 and 109A was employed to examine the effect on lipolysis in TNF-α-stimulated adipocytes. RESULTS: Co-culture increased PGE2 release into the medium, compared with cells cultured separately. Butyrate significantly increased PGE2 production. Co-culture elevated COX2 expression in macrophages and adipocytes, and butyrate further enhanced this effect. Co-culture enhanced cytosolic PLA2 activity in macrophages, which was further enhanced by butyrate. As for lipolysis, co-culture increased the release of FFAs and free glycerol into the medium, whereas butyrate (and to a lesser extent, PGE2) suppressed FFAs and free glycerol release. An inhibition study using a prostaglandin E receptor 3-selective antagonist suggested that approximately 40% of the suppressive effect of butyrate depends on the PGE2-mediated pathway, whereas 60% depends on a non-PGE2-mediated pathway. Co-culture increased cAMP and PRKAR1A levels in adipocytes, whereas butyrate restored the levels to those of the control. Similarly, in TNF-α-stimulated adipocytes, butyrate reduced FFAs and free glycerol release. siRNA inhibition of GPR41 and GPR109A suggested that the GPR109A-mediated pathway predominates, but the GPR41-mediated pathway also regulates the effect of butyrate on lipolysis in TNF-α-stimulated 3T3-L1 cells. CONCLUSIONS: Butyrate attenuates lipolysis in adipocytes co-cultured with macrophages via non-PGE2-mediated and PGE2-mediated pathways.

Synergistic antiproliferative effect of imatinib and adriamycin in platelet-derived growth factor receptor-expressing osteosarcoma cells.

Osteosarcoma (OS) is the most frequent primary solid malignant tumor of bone. Its prognosis remains poor in the substantial proportion of patients who do not respond to chemotherapy and novel therapeutic options are therefore needed. We previously established a mouse model that mimics the aggressive behavior of human OS. Enzyme-linked immunosorbent assay-based screening of such mouse tumor lysates identified platelet-derived growth factor-BB (PDGF-BB) as an abundant soluble factor, the gene for which was expressed dominantly in surrounding non-malignant cells of the tumor, whereas that for the cognate receptor (PDGF receptor ß) was highly expressed in OS cells. Platelet-derived growth factor-BB induced activation of both MEK-ERK and phosphatidylinositol 3-kinase-protein kinase B signaling pathways and promoted survival in OS cells deprived of serum, and these effects were blocked by the PDGF receptor inhibitor imatinib. However, these actions of PDGF-BB and imatinib were mostly masked in the presence of serum. Whereas imatinib alone did not manifest an antitumor effect in mice harboring OS tumors, combined treatment with imatinib and adriamycin exerted a synergistic antiproliferative effect on OS cells in vivo. These results suggest that treatment of OS with imatinib is effective only when cell survival is dependent on PDGF signaling or when imatinib is combined with another therapeutic intervention that renders the tumor cells susceptible to imatinib action, such as by inducing cellular stress.

A case of Degos disease: demonstration of C5b-9-mediated vascular injury.

A 68-year-old Japanese male presented with atrophic erythematous white lesions with peripheral dark reddish rims on his back. Multiple ulcers were detected from his stomach to his large intestine using endoscopy. Although the patient was given high doses of a steroid, aspirin, dipyridamole, and intravenous immunoglobulin therapy, he died of gastrointestinal hemorrhage, perforation and septic shock. An autopsy examination revealed pauci-inflammatory thrombotic microangiopathy with endothelial cell injury, fibrous occlusive arteriopathy, and vascular C5b-9 deposition in the wall of the gastrointestinal tract from the esophagus to the large intestine as well as in the dermis of the skin.

Isolation of dermatophytes and related species from domestic fowl (Gallus gallus domesticus).

We investigated 793 bird combs [645 chickens and 148 fighting cocks (Shamo)] to determine the prevalence of dermatophytes and their related fungal species. The targeted fungal species were recovered from 195 of the 793 examined birds (24.6 %). Prevalence ratios were compared in temperate (the mainland) and subtropical (Nansei Islands) areas, genders, strains, breeding scale (individual and farm), and housing system (in cage and free ranging). The frequency of the fungal species in the mainland, males, fighting cocks, breeding scale by individual nursing, and free-range housing system exhibited significantly higher positive ratios than that in the other groups. A total of 224 dermatophytes and related species were isolated, including 101 Arthroderma (Ar.) multifidum, 83 Aphanoascus (Ap.) terreus, five Uncinocarpus queenslandicus, two U. reesii, two Ap. pinarensis, one Amauroascus kuehnii, one Ar. simii, one Gymnoascus petalosporus, one Microsporum gallinae, and 28 Chrysosporium-like (Chrysosporium spp.) isolates, which were identified using internal transcribed spacer regions of ribosomal RNA gene sequences. The predominant fungal species in the mainland was Ap. terreus and that in the Nansei Islands was Ar. multifidum. Pathogenic fungal species to humans and animals were limited to M. gallinae and Ar. simii, which corresponded to 0.025 % of the isolates in this study.

Bilateral Upper Arm Granulomas Induced by Leuprorelin Acetate Injection Mimicking Malignant Soft Tissue Tumors: A Case Report.

Leuprorelin acetate is a common anticancer medication used for prostate cancer treatment. One of the local adverse reactions after leuprorelin injection is the development of reactive granulomas, typically presenting as subcutaneous nodules. In this case report, we describe a 73-year-old patient with prostate cancer who developed unusually large sized intramuscular reactive granulomas, which mimicked malignant soft tissue tumors. The patient, who had been receiving leuprorelin acetate treatment for the past 12 months, noticed painful masses in both upper arms. Based on the findings of magnetic resonance imaging and fluorodeoxyglucose-positron emission tomography/computed tomography, a diagnosis of malignant soft tissue tumor was strongly suggested. However, further investigation through needle biopsy ultimately led us to the final diagnosis of reactive granuloma. The masses spontaneously resolved after discontinuation of leuprorelin injection. While reactive granulomas after leuprorelin injections are not rare, intramuscular cases are relatively uncommon. Despite using imaging studies as a rational initial approach in the diagnostic process, as we did in our case, their results turned out to be indistinguishable from those of malignant soft tissue tumors, thus highlighting the importance of pathological examination in confirming diagnosis, especially when a patient presents with atypical clinical manifestations.

Twist2 functions as a tumor suppressor in murine osteosarcoma cells.

The epithelial-mesenchymal transition (EMT) contributes to the malignant progression of cancer cells including acquisition of the ability to undergo metastasis. However, whereas EMT-related transcription factors (EMT-TF) are known to play an important role in the malignant progression of epithelial tumors, their role in mesenchymal tumors remains largely unknown. We show that expression of the gene for Twist2 is downregulated in human osteosarcoma and correlates inversely with tumorigenic potential in mouse osteosarcoma. Forced expression of Twist2 in highly tumorigenic murine osteosarcoma cells induced a slight inhibition of cell growth in vitro but markedly suppressed tumor formation in vivo. Conversely, knockdown of Twist2 in osteosarcoma cells with a low tumorigenic potential promoted tumor formation in vivo, suggesting that Twist2 functions as a tumor suppressor in osteosarcoma cells. Furthermore, Twist2 induced expression of fibulin-5, which has been reported as a tumor suppressor. Medium conditioned by mouse osteosarcoma cells overexpressing Twist2 inhibited expression of the MMP9 gene as well as invasion in mouse embryonic fibroblasts, and forced expression of Twist2 in osteosarcoma cells suppressed MMP9 gene expression in tumor tissue. Data from the present study suggest that Twist2 inhibits formation of a microenvironment conducive to tumor growth and thereby attenuates tumorigenesis in osteosarcoma.

Isolation of Microsporum gallinae from a fighting cock (Gallus gallus domesticus) in Japan.

A case of tinea corporis caused by Microsporum gallinae was found in 2011 in Okinawa, located in the southern part of Japan. The patient was a 96-year-old, otherwise healthy, Japanese man, who had been working as a breeder of fighting cocks for more than 70 years. He was bitten on his right forearm by one of the cocks and a few weeks later, two erythematous macules appeared on the right forearm, accompanied by a slight itchy sensation. While the first isolate of this dermatophyte was recovered from the region by Miyasato et al. in 2011, it was not obtained from the same fighting cock owned by the patient. However, frequent exchanges of fighting cocks and special domestic breeds of chickens related to fighting, mating, and/or bird fairs are common among the fans and breeders. We investigated 238 chickens and 71 fighting cocks in Okinawa and in the suburbs of Tokyo (Chiba, Tokyo, Ibaraki, and Sizuoka). One isolate of M. gallinae from a fighting cock in Chiba Prefecture in the Tokyo metropolitan area exhibited a different genotype, with a single base difference from the patient isolate based on the internal transcribed spacer 1-5.8s-ITS2 regions (ITS1-5.8S-ITS2) of the ribosomal RNA gene sequence. The isolation of M. gallinae from a fighting cock on the mainland of Japan is the first such finding in animals in our country.