First Report of Guinea Grass Smut Caused by Conidiosporomyces ayresii in Japan.

Guinea grass (Panicum maximum Jacq.) is an important C-4 perennial plant that grows in southern Japan. In March 2010, a smut disease was found in grass that is cultivated in the Ishigaki Islands, Okinawa, in southernmost Japan. Spikelets of susceptible cultivars were swollen and filled with gray spore masses and seed production was substantially reduced. Two single-spore isolates of a smut fungus were obtained from infected spikelets and deposited at the NIAS Genebank, Japan as MAFF511519 and 511520. The 28S rDNA sequences of the isolates were analyzed as described by Boekhout et al. (1). The 28S rDNA sequence (GenBank Accession No. AB647346) of isolate MAFF511519 matched that of Conidiosporomyces ayresii (GenBank Accession No. AY819017) isolated from P. maximum with 99.8% similarity. Spores were pale brown to brown, globose to subglobose, verrucose, and 14 to 16 × 15 to 18 µm in diameter with relatively thick walls of 2 to 3 µm. With scanning electron microscopy, warts appeared dense and short with pointed tips. Spores germinated under wet conditions and produced masses of basidiospores. Basidiospores were aseptate, long, cylindrical, straight to slightly curved, 20 to 37 × 2 to 3 µm, and often germinated into Y-shaped conidia. This description matches previous descriptions (3) of C. ayresii (Berk.) Vánky (Tilletia ayresii Berk.) of the smut pathogen of guinea grass (2). The smut fungus was identified as C. ayresii on the basis of morphology and molecular phylogenetic analysis. To produce inoculum, the isolates were grown on potato dextrose agar at 25°C in the dark for 7 days. Two plants of cv. Ryukyu 5-gou with half-flowering heads were grown in a greenhouse for approximately 1 month and then inoculated by atomizing them with conidial suspensions of each isolate (106 conidia/ml). A plant sprayed with sterilized distilled water served as the control. Inoculated heads were covered with plastic bags for 48 h at 25°C. After 30 days, all inoculated plants were symptomatic with swollen spikelets releasing gray spores. Controls remained asymptomatic. The smut fungus was reisolated from released gray spores, confirming Koch's postulates. To our knowledge, this is the first report of smut caused by C. ayresii on guinea grass in Japan. References: (1) T. Boekhout et al. Stud. Mycol. 38:175, 1995. (2) J. M. Lenné and P. Trutmann. Diseases of Tropical Pasture Plants. CAB International, Wallingford, UK, 1994. (3) K. Vánky and R. Bauer. Mycotaxon 43:427, 1992.

First Report of Pestalotiopsis microspora Causing Leaf Spot of Hidcote (Hypericum patulum) in Japan.

Hidcote, Hypericum patulum Thunb. ex Murray, is a deciduous shrub that is cultivated as an ornamental in landscape gardens and courtyards in Japan. In early August 2008, severe leaf spotting was observed on plants growing in a courtyard in Nasushiobara, Tochigi, Japan. More than 30% of the leaves on five shrubs exhibited leaf spot symptoms. Small, round, pale brown lesions were initially observed. Later, they expanded to 5 to 12 mm in diameter, round to irregular-shaped with pale brown centers and dark brown margins. Under continuously wet or humid conditions, black acervuli developed on the leaf lesions. Conidia were straight or slightly curved, fusiform to clavate, and five-celled with constrictions at the septa. Conidia ranged from 17 to 21 × 5 to 8 µm with hyaline apical and basal cells. Fifteen percent of apical cells had two and the rest had three appendages (setulae) ranging from 10 to 21 µm long. The basal hyaline cell tapered into a 2 to 4 µm pedicel. The three median cells ranged from light or dark brown to olive green. These morphological characteristics matched those of Pestalotiopsis microspora (Speg.) G.C. Zhao & N. Li (1,2). The identity of the fungus was confirmed by DNA sequencing of the internal transcribed spacer (ITS) region (GenBank Accession No. GU908473) from single-spore isolates, which revealed 100% homology with those of other P. microspora isolates (e.g., GenBank Accession Nos. FJ459950 and DQ456865). Koch's postulates were confirmed using leaves of three detached branches of a field-grown asymptomatic plant of H. patulum. Thirty leaves of each branch were inoculated by placing mycelial plugs obtained from the periphery of 7-day-old single-spore cultures on the leaf surface. Potato dextrose agar plugs without mycelium served as controls. Leaves on branches were covered with plastic bags for 24 h to maintain high relative humidity in a greenhouse (approximately 24 to 28°C). After 5 days, all inoculated leaves showed symptoms identical to those described above, whereas control leaves remained symptom free. Reisolation of the fungus from lesions on inoculated leaves confirmed that the causal agent was P. microspora. To our knowledge, this is the first report of leaf spots on H. patulum caused by P. microspora in Japan. Management options may have to be developed and implemented to protect Hidcote plants in areas where leaf spot cannot be tolerated. References: (1) P. A. Saccardo. Sylloge Fungorum III:789, 1884. (2) G. C. Zhao and N. Li. J. Northeast For. Univ. 23(4):21, 1995.

First Report of Alternaria panax Causing Leaf Spot of Aralia cordata in Japan.

Udo, Aralia cordata Thumb, Araliaceae, is a traditional Japanese perennial vegetable and used in Chinese herbal medicines. During the last 10 days of July 2008, before the period of flower, leaf spots were observed on udo growing under pine trees in Nasushiobara, Tochigi, Japan. Leaf spots affected more than 40% of the plants. Early symptoms appeared as small, round or irregular, water-soaked, dark brown lesions on the leaves. These areas expanded to 15 to 30 mm in diameter, were irregular and pale brown in the central area and the margin of the lesions were water soaked and dark brown. Later, some lesions coalesced. In continuously wet or humid conditions, conidiophores with conidia appeared on the surface of leaf spots. Conidiophores were medium brown and simple (approximately 70 to 160 × 6 to 8 µm). Well-developed conidia were long-obclavate, base obtuse, straight, yellowish brown, smooth walled, with six to nine transverse septa and three to five longitudinal or oblique septa, constricted at some main septa, some cells easily swelled, conidium body was 72 to 100 × 19 to 34 µm, and the rostra extension was 40 to 90 × 4 to 5 µm. The pathogen was identified as Alternaria panax on the basis of the morphology and sequence of ITS1-5.8s-ITS2 of rDNA. The sequence (GenBank Accession No. FJ607183) exactly matched the sequences of two A. panax (e.g., GenBank Accession Nos. AY898639 and AY898640) (2-4). The fungus was isolated on V8 agar from a single conidium found on symptomatic leaf tissue. Colonies of A. panax were gray-to-black and did not easily produce conidia on the agar. Koch's postulates were performed with the leaves of three branches on a field plant of Aralia cordata. Leaves were inoculated with a mycelial plug harvested from the periphery of a 7-day-old colony; an equal number of leaves on the same plant inoculated with plugs of V8 medium served as the control. All test leaves were covered with plastic bags for 24 h to maintain high relative humidity and incubated at a natural temperature (approximately 24 to 28°C). After 7 days, all inoculated leaves showed symptoms identical to those observed in natural conditions, whereas the controls remained symptom free. Reisolation of the fungus from lesions on inoculated leaves confirmed that the causal agent was A. panax. This species has been previously reported on Aralia cordata in Korea (1). To our knowledge, this is the first report of leaf spots caused by A. panax on Aralia cordata in Japan. References: (1) W. D. Cho and H. D. Shin. List of Plant Diseases in Korea. 4th ed. Korean Society of Plant Pathology, 2004. (2) E. G. Simmons. Mycotaxon 14:17, 1982. (3) E. G. Simmons. Alternaria: An Identification Manual. CBS Fungal Biodiversity Centre, Utrecht, the Netherlands, 2007. (4) T. Y. Zhang et al. Flora Fungorum Sinicorum: Alternaria. Vol. 16. Science Press (in Chinese), Beijing, 2003.

First Report of Blast of Guinea Grass Caused by Pyricularia sp. LS-Group in Japan.

Guinea grass (Panicum maximum Jacq.) is an important C-4 perennial herbage in the southern part of Japan. In February 2002, a blast disease was found on the grass cultivated on the Okinawa Islands, the southern most region of Japan. Early symptoms appeared as small, round or ellipsoid lesions on leaves. Lesions later expanded to 2 to 5 × 1 to 2 mm and were spindle shaped and grayish white in the central area with dark brown margins. We obtained three single-conidia isolates of a Pyricularia-like fungus from the lesions and deposited them in the NIAS Genebank, Japan as MAFF306662, 306671, and 306672. The isolates were grown under near-UV light on V8 juice agar for 7 days to produce conidia, and guinea grass plants of the seven- to eight-leaf stage grown from seeds in a green house, five plants for each isolate, were inoculated by atomizing them with the conidial suspension of 105 conidia/ml. The same number of plants sprayed with sterilized distilled water served as the control. The experiments were repeated twice. All plants were covered with plastic bags for 24 h at 25°C to maintain high relative humidity. After 7 days, all inoculated plants showed symptoms identical to those observed in the field. Controls remained symptom free. The Pyricularia-like fungus was reisolated from lesions on inoculated leaves. The morphologies of the isolates were observed and described from the colonies grown under the condition described above. Conidiophores were pale brown, emerging singly or in small groups, straight or flexuous, geniculate toward the apex, and 36 to 197 × 2 to 5 µm. Conidia were obpyriform, straight, colorless to pale brown, smooth, and 19 to 30 × 5 to 10 µm with two to three septa. The morphologies were the same as those of the description of the genus Pyricularia. Previously, all Pyricularia isolates from Gramineae had been identified as P. grisea, except for those from rice (3,4). However, a new taxonomy of Pyricularia spp. based on DNA analyses was proposed by Couch and Kohn (1). Only the isolates from Digitaria were classified as P. grisea and those from C-3 grasses classified as P. oryzae. However, the species names for the isolates from the other C-4 grasses were not described. We analyzed the sequences of the rDNA-ITS region (ITS1-5.8s-ITS2) and ß-tubulin gene of the isolates from guinea grass following Couch and Kohn (1). The sequences of rDNA-ITS (GenBank Accession No. AB512785) and ß-tubulin (AB512786) of the isolate MAFF306672 matched the sequences of those of the Pyricularia sp. LS-group (AB274426 and AB274458, respectively) isolated from Leersia oryzoides. Hirata et al. (2) reclassified Pyricularia isolates from Gramineae by multilocus phylogenetic analysis and showed that non-P. oryzae and non-P. grisea isolates could be classified into two groups of the Pyricularia sp., a LS- and a CE-group, corresponding to those isolated from Leersia spp. and Setaria spp. or Cenchrus spp. of grasses, respectively. Since no Magnaporthe teleomorph was produced by the crossing tests using the isolates, we identified the isolates from guinea grass as the Pyricularia sp. LS-group on the basis of their morphology and the molecular phylogenetic analysis. To our knowledge, this is the first report of blast on guinea grass in Japan. References: (1) B. C. Couch and L. M. Kohn. Mycologia 94:683, 2002. (2) K. Hirata et al. Mycol. Res. 111:799, 2007. (3) K. D. Hyde. Australas. Plant Pathol. 22:73, 1993. (4) R. Sprague. Diseases of Cereals and Grasses in North America. Ronald Press Company, New York, 1950.

First Report of Botrytis cinerea Causing Leaf Spot of Aquilegia vulgaris in Japan.

European columbine, Aquilegia vulgaris L., Ranunculaceae, is an herbaceous flower widely used in gardens, parterres, and courtyards and is a traditional herbal plant. During the summer of 2008, leaf spots were observed on a plant cultivated along a roadside area in Nasushiobara, Tochigi, Japan. In some courtyards, the leaf spot affected more than 60% of the plants. Early symptoms appeared as small, round or elliptic, brown lesions on the leaves. Lesions expanded to 5 to 15 × 4 to 10 mm, irregular spots that were dark brown to black in the middle, with pale yellow-brown or purple-brown margins. In continuously wet or humid conditions, thick, gray mycelium and conidia appeared on the surface of leaf spots. Conidiophores were melanized at the base and hyaline near the apex, branched, and septated (approximately 3 mm × 16 to 18 µm). Conidia were hyaline, aseptate, ellipsoidal to obovoid, with a slightly protuberant hilum, and ranged from 9 to 14.5 × 5.5 to 6.5 µm. The pathogen was identified as Botrytis cinerea Pers.:Fr on the basis of morphology and sequence of ITS1-5.8s-ITS2 region of rDNA. The sequence (GenBank Accession No. FJ424510) exactly matched the sequences of two Botryotinia fuckeliana (anamorph Botrytis cinerea), (e.g., GenBank Accession Nos. AY686865 and FJ169666) (2). The fungus was isolated on potato dextrose agar (PDA) from a single conidium found on the symptomatic leaf tissue. Colonies of B. cinerea were first hyaline and later turned gray to black when the spores differentiated. Koch's postulates were performed with three whole plants of potted aquilegia. Leaves were inoculated with mycelia plugs harvested from the periphery of a 7-day-old colony; an equal number of plants were inoculated with the plugs of PDA medium only and served as controls. All plants were covered with plastic bags for 24 h to maintain high relative humidity and incubated at 25°C. After 8 days, all mycelium-inoculated plants showed symptoms identical to those observed on leaves from A. vulgaris infected in the field, whereas controls remained symptom free. Reisolation of the fungus from lesions on inoculated leaves confirmed that the causal agent was B. cinerea. B. cinerea has been previously reported on A. vulgaris in the United States and China (1,3). To our knowledge, this is the first report of leaf spots caused by B. cinerea on A. vulgaris in Japan. References: (1) Anonymous. Index of Plant Diseases in the United States. USDA Agric. Handb. No 165, 1960. (2) M. B. Ellis. Dematiaceous Hyphomycetes. Commonwealth Mycological Institute, Kew, England, 1971. (3) Z. Y. Zhang. Flora Fungorum Sinicorum. Vol. 26. Botrytis, Ramularia. Science Press, Beijing, 2006.

Prenatal diagnosis of Lowe syndrome by OCRL1 messenger RNA analysis.

Prenatal screening of oculo-cerebro-renal syndrome of Lowe (OCRL; McKusick 309000) was performed using cultured amniocytes. Following identification of defective mRNA expression in the OCRL; gene of the proband's fibroblasts, the mRNA size and quantity of the cultured amniocytes were compared. Based on this analysis, the fetus was diagnosed as being normal and was subsequently delivered as a healthy boy. This is the first reported successful prenatal screening of OCRL using a comparison with defective mRNA of OCRL1 from affected subjects.

Proliferative defect and embryonic lethality in mice homozygous for a deletion in the p110alpha subunit of phosphoinositide 3-kinase.

Phosphatidylinositol 3,4,5-trisphosphate is a phospholipid signaling molecule involved in many cellular functions including growth factor receptor signaling, cytoskeletal organization, chemotaxis, apoptosis, and protein trafficking. Phosphorylation at the 3 position of the inositol ring is catalyzed by many different 3-kinases (classified as types IA, IB, II, and III), but the physiological roles played by each of the different 3-kinase isozymes during embryonic development and in homeostasis in animals is incompletely understood. Mammalian type IA kinase isozymes are heterodimers that are active at 37 degrees C when the catalytic 110-kDa subunit interacts through an amino-terminal binding domain with a regulatory 85- or 55-kDa subunit. Using gene targeting in embryonic stem cells, we deleted this binding domain in the gene encoding the alpha isoform of the 110-kDa catalytic subunit (Pik3ca) of the alpha isozyme of the type IA kinases, leading to loss of expression of the p110 catalytic subunit. We show that Pik3cadel/del embryos are developmentally delayed at embryonic day (E) 9.5 and die between E9.5 and E10.5. E9. 5 Pik3cadel/del embryos have a profound proliferative defect but no increase in apoptosis. A proliferative defect is supported by the observation that fibroblasts from Pik3cadel/del embryos fail to replicate in Dulbecco's modified Eagle's medium and fetal calf serum, even with supplemental growth factors.

[TNM classification--pediatric tumors].

The clinical and postsurgical TNM classifications (cTNM and pTNM) for neuroblastoma (NB), nephroblastoma (WT) and soft tissue sarcomas were presented in 1982 by the TNM Committee in UICC in collaboration with SIOP. The Japanese TNM Committee proposed new pTNM systems (J-pTNM) for NB and WT, and new cTNM and pTNM system for primary liver carcinoma in infants and children (HT). These pTNMs were based on the staging systems developed by the Malignant Tumor Committee of the Japanese Society of Pediatric Surgeons. The proposal of subdivision of M category in NB was presented for testing the new telescopic ramifications of TNM. The TNM for HT was added as a new classification recommended for testing. The effectiveness of these TNM systems was assessed using NB, WT and hepatoblastoma (HB) cases which were registered in collaborating institutes. The analyses suggested that pTNM, especially the J-pTNM system in NB, WT and HT were effective for the assessment of prognoses, although cTNM systems were not enough to assess the extent of the disease.

Prevention of visual field defect progression with brovincamine in eyes with normal-tension glaucoma.

PURPOSE: A prospective investigation of the effect of brovincamine fumarate, a Ca2+-channel blocker, on visual field changes in normal-tension glaucoma (NTG). METHODS: A total of 28, age- and field-matched, patients with NTG were allocated randomly to either brovincamine fumarate (20 mg 3 times daily) or placebo (3 times daily). The patients were followed at least every 4 months for a minimum of 2.5 years, and visual field examinations were carried out at least every 6 months. The mean follow-up periods (+/-standard deviation) were 39.1 +/- 8.7 months in the brovincamine-treated group and 37.9 +/- 10.1 months in the placebo group. Stepwise discriminant analyses were performed to separate the patients who showed improvement of their visual fields from those who failed to improve in the brovincamine-treated group, and to identify factors that may determine the visual field prognosis of all patients with NTG enrolled in the study. RESULTS: In the brovincamine-treated group, six patients showed visual field improvement, whereas none showed improvement in the placebo group using the Statpac 2 linear regression analysis. Discriminant analyses identified better cold recovery rate and higher initial systolic blood pressure to be significantly contributory to a favorable outcome in the brovincamine-treated group, and the use of brovincamine, better cold recovery rate, and higher initial systolic blood pressure were identified to be significantly contributory to a favorable prognosis in all subjects. CONCLUSION: Brovincamine seems to have a favorable effect on visual field in at least some patients with NTG.

Unique presentation of heterotopic pancreatic tissue arising from small-bowel mesentery.

Heterotopic pancreatic tissue in the small-bowel mesentery is an extremely rare anomaly: no such case has been reported in the English literature. We report a 5-year-old boy in whom the aberrant tissue formed a retroperitoneal cyst. The condition was further complicated by a ventral pancreatic duct defect. The unique clinical manifestations and difficulty in making the diagnosis are discussed.

Identification and chromosomal mapping of the mouse inositol polyphosphate 1-phosphatase gene.

A mouse inositol polyphosphate 1-phosphatase (Inpp1) cDNA fragment (348 bp) was amplified by means of the polymerase chain reaction using a mouse cDNA library as template with primers designed from published human and bovine cDNA sequences. We isolated a 1623-bp full-length Inpp1 cDNA from a mouse brain cDNA library using this amplified cDNA fragment as probe. Amino acid sequences of mouse, human, and bovine inositol polyphosphate 1-phosphatase are highly conserved. Northern blot analysis shows a major transcript of 1.65-kb mRNA and several higher molecular weight mRNAs that are expressed in a variety of mouse tissues. Utilizing the Jackson Lab backcross DNA panel map service, we mapped Inpp1 to chromosome 1, 1.06 cM proximal to Ctla4.

Biological characteristics of neuroblastoma with spontaneous tumor reduction: a case report.

The authors examined the biological characteristics of a neuroblastoma with spontaneous tumor reduction. A 6-month-old boy with a pelvic neuroblastoma underwent surgical extirpation of the tumor 1 month after diagnosis. The size of the tumor reduced spontaneously while he was awaiting operation. The low proliferative activity of the tumor cells and the presence of apoptosis in the tumor tissue were shown by an immunohistochemical method using anti-PCNA antibody and a DNA fragmentation analysis, respectively. These results suggest that the spontaneous tumor reduction seen in this patient may well be caused by the overwhelming apoptosis of tumor cells.

[Hypotonous maculopathy following trabeculectomy with mitomycin C].

We retrospectively studied incidence, prognosis and predictive factors of hypotonous maculopathy following trabeculectomy with adjunctive mitomycin C. The subjects were 208 cases (208 eyes) with various types of glaucoma with clear media, who were followed up for at least three months postoperatively. The follow up periods ranged from 3 to 44 months with a mean of 21.1 months. The incidence of the maculopahy was 9.1% (19/208). The postoperative intraocular pressures (IOPs) varied between 1 and 6 mmHg in the presence of maculopathy. The maculopathy disappeared in 14 eyes (74%) spontaneously or following ocular hypertensive therapy including application of trichloracetic acid and subconjunctival injection of autologous blood. The final visual acuity deteriorated two lines or more in 8 eyes (42%). A discriminant analysis showed that age, history of intraocular surgery, preoperative maximum IOP, and mean deviation are the predictive factors for maculopathy. As compared with the hypotonous eyes without maculopathy, those with maculopathy tended to be younger and to have less severe visual field changes.

Autologous bone marrow transplantation in children with advanced neuroblastoma.

BACKGROUND: Encouraging results have been reported with high dose chemotherapy and total body radiation followed by bone marrow autotransplantation in children with advanced neuroblastoma; however, relapse remains a significant problem. METHODS: The authors treated 22 children with advanced neuroblastoma with high dose chemotherapy, surgery, intraoperative radiation, and a bone marrow autotransplant (treated in vitro to remove tumor cells) followed by 13-cis-retinoic acid. RESULTS: The 3-year relapse rate was 25% (95% confidence interval [CI], 6-44%). The 3-year disease free survival rate was 72% (95% CI, 52-92%). Toxicities included hemolytic uremic syndrome, herpes infection, and hepatic venoocclusive disease. CONCLUSION: These data suggest that this treatment strategy offers an increased rate of 3-year disease free survival. The nonrandomized nature of this study and its use of multiple modalities precludes the analysis of the specific contribution of each treatment component and comparison with conventional therapy.

[Methylmalonic acidemia with bilateral MRI high intensities of the globus pallidus].

We report a patient with methylmalonic acidemia who developed an acute extrapyramidal disorder after severe ketoacidosis. The neurologic findings resulted from bilateral destruction of the globus pallidus. A 10-year-old girl was the term product of an uncomplicated pregnancy and delivery. Poor feeding and vomiting were noted after one month. She was hospitalized at 6 months of age with vomiting, coma and tachypnea. Analysis of urinary organic acids revealed a massive amount methylmalonic acid. She was not vitamin B 12-responsive and was maintained on a low-protein diet. At 33 months of age, she was able to walk and speak, but she developed acute severe ketoacidosis. Involuntary movements and spastic paraplegia became evident two days after admission. Subsequently, the patient has had metabolic ketoacidosis once or twice a year. Her intelligence quotient was 47. Neurologic examination revealed spastic paraplegia and generalized hypotonicity with mild dystonia. Some relief from dystonic symptoms has been obtained through the use of L-dopa. A brain CT scan at 5 years of age disclosed bilaterally symmetric lucencies of the globus pallidus. T2-weighted brain MRI at 8 years of age showed bilateral symmetric high intensities of the globus pallidus.