Reiko Tanaka

Trichosporon species infection in bone marrow transplanted patients.

Trichosporon species are emerging as opportunistic agents that cause systemic diseases in immunocompromised patients. Patients undergoing bone marrow transplant are submitted to intense and prolonged periods of neutropenia and consequently to several risk factors to fungal infections as the use of broad spectrum antibiotics and invasive devices. Two cases of fungal infections caused by Trichosporon asahii var. asahii and T. inkin in patients with bone marrow transplant are described T. asahii var. asahii was responsible for fungemia and the identification of this microorganism was later performed. T. inkin caused vascular accesses infection and was recovered from an implanted Hickman-Broviac catheter. Both patients were under oral fluconazole prophylaxis. The patient with systemic infection died despite the therapy with amphotericin B and the patient with catheter-related infection recovered from the fungal infection after catheter removal. Difficulties in the identification of this microorganism lead to delays in treatment and post-mortem diagnosis.

Fungi in bathwater and sludge of bathroom drainpipes

Samples of bathwater from 14 homes and 22 public bathhouses and sludge in drainpipes from 19 house-hold bathrooms were plated out onto potato dextrose agar supplemented with chloramphenicol. Several media were used to study colony morphology of the isolates and the thermotolerance and alkaline tolerance of each isolate were examined.Eleven sludge samples produced 12 isolates of Exophiala jeanselmei, 2 of E. dermatitidis and 1 of E. moniliae. Five household bathwater samples produced 2 isolates of E. jeanselmei, 4 of E. dermatitidis and 1 of E. alcalophila. One isolate of E. jeanselmei, 2 of E. dermatitidis, 3 of E. moniliae and 2 of unidentified Exophiala species were recovered from 6 samples of the bathwater dissolving ‘Chinese medicine’ in the bathtubs of public bathhouses. One isolate of E. jeanselmei was recovered from the 15 samples of bathwater from public bathhouses. Bathwater and sludge in bathroom drainpipes may be an important habitat of Exophiala species.

Pulmonary cysts of Birt-Hogg-Dubé syndrome: a clinicopathologic and immunohistochemical study of 9 families.

Birt-Hogg-Dubé (BHD) syndrome is an autosomal dominant disorder characterized by fibrofolliculomas, renal tumors, and pulmonary cysts with recurrent pneumothorax. Multiple pulmonary cysts and pneumothorax are the key signs for diagnosing BHD syndrome. The pathologic features of BHD pulmonary cysts, however, are poorly understood. This disorder is caused by mutations in the gene that encodes folliculin (FLCN). FLCN is regarded as a tumor suppressor; it mediates cellular activities by interacting with the mammalian target of rapamycin (mTOR). In this study, we investigated the lungs of 11 patients from 9 BHD families. The majority of patients consulting doctors were women between 30 and 60 years of age who had pulmonary cysts and repeated pneumothoraces. Genomic DNA testing revealed 5 different mutation patterns. Histopathologic examination found that the inner surface of cysts was lined by epithelial cells, sometimes with a predominance of type II pneumocyte-like cuboidal cells. The cysts occasionally contained internal septa consisting of alveolar walls or showed an “alveoli within an alveolus” pattern. The cells constituting the cysts stained positive for phospho-S6 ribosomal protein expression, suggesting activation of the mTOR pathway. Although BHD pulmonary cysts are frequently misdiagnosed as nonspecific cystic diseases, they are distinctly different in histopathology from other bullous changes. Mechanical stress such as rupture and postrupture remodeling allows mesothelial invagination and fibrosis. Such modified BHD pulmonary cysts are virtually indistinguishable from nonspecific blebs and bullae. We propose a new insight, namely, that the BHD syndrome-associated pulmonary cyst may be considered a hamartoma-like cystic alveolar formation associated with deranged mTOR signaling.

Antifungal Susceptibilities, Varieties, and Electrophoretic Karyotypes of Clinical Isolates of Cryptococcus neoformans from Brazil, Chile, and Venezuela

ABSTRACT One hundred clinical isolates of Cryptococcus neoformans from human immunodeficiency virus (HIV)-infected and non-HIV-infected patients from Brazil, Chile, and Venezuela were separated according to varieties and tested for antifungal susceptibility. A high susceptibility to antifungal agents was observed among all the isolates. The electrophoretic karyotyping of 51 strains revealed good discrimination among Cryptococcus neoformansvar. neoformans strains.

Serotyping of Cryptococcus neoformans strains isolated from clinical specimens in Thailand and their susceptibility to various antifungal agents

One hundred and thirty-nine strains of Cryptococcus neoformans were isolated in Thailand from clinical specimens including 97 AIDS patients: 67 from Northern, 48 from Central, 17 from Northeastern and 7 from Southern regional hospitals. Six out of the 139 strains were serotype B and the remaining 133 were A. There was no correlation between serotypes and regional distribution. To our knowledge, this is the first report of serotyping studies on C. neoformans in Thailand. Studies on random amplified polymorphic DNA (RAPD) analysis showed that this method is useful for the differentiation of C. neoformans var. gattii (serotypes B and C) and C. neoformans var. neoformans (serotypes A and D). They also indicated that Thai isolates of C. neoformans var. gattii (serotype B) were a homogeneous group on the basis of their genotypes. Antifungal susceptibility tests using 5 antifungal agents including amphotericin B, fluconazole, flucytosine, itraconazole and micronazole against 50 selected strains of C. neoformans showed that they were sensitive to all of the antifungal agents tested except for one strain that was resistant to flucytosine.

Determination of ploidy in Cryptococcus neoformans by flow cytometry

We determined the ploidy of Cryptococcus neoformans (28 strains) isolated from patients and nature. The cellular DNA content of these strains, which stained with propidium iodide in comparison to that of two authentic haploid strains, was determined by flow cytometry. All the strains exhibited diphasic histograms. In case of the authentic haploid strains, the first peak was centred around channel 9, and the second peak around channel 18. Most strains exhibited this type of histogram. Some strains exhibited another type of histogram: the first peak was centred around channel 18, and the second one around channel 35. In flow cytometry, the channel number is correlated with the intensity of fluorescence, namely, in proportion to the channel number the DNA content in the cells increases. The cellular DNA content of the second type of histograms showed twice that of the authentic haploid strains, and thus, five of 28 isolates were concluded to be diploid, and the others haploid.

Lung cysts in Birt-Hogg-Dube syndrome : Histopathological characteristics and aberrant sequence repeats

Birt‐Hogg‐Dubé (BHD) syndrome is a rare disorder inherited in an autosomal dominant manner. The affected patients are predisposed to cutaneous fibrofolliculomas, renal cell tumors and lung cysts with recurrent pneumothorax. Contrary to neoplastic events in the skin and the kidney, the lung cysts have frequently been confused with non‐neoplastic changes such as blebs or bullae. Herein is reported a case of multiple lung cysts associated with BHD syndrome. Detailed histopathological characteristics of the lesion are also given. The lung cysts were closely associated with the peripheral interlobular septum, visceral pleura or septal‐pleural junctional region. These cysts were partly abutting alveolar structures, and lined by a layer of alveolar epithelium. These unique microscopic features supported the notion that the BHD lung lesions are distinct from other types of bullous changes. Genomic DNA analysis indicated an aberrant sequence repeat that caused frameshift mutation. Immunohistochemistry showed the localization of folliculin, the BHD gene‐encoding protein, in macrophages and epithelial cells in the patients and normal controls lungs. Haploinsufficiency of folliculin may cause deranged alveolar development, leading to the aberrant cystic alveolar formation. The unique mutation patterns of abnormal sequence repeats in patients with BHD syndrome are also reviewed.

The isolation of Cryptococcus neoformans from pigeon droppings and serotyping of naturally and clinically sourced isolates in China.

This is the first report on the isolation ofCryptococcus neoformans from pigeon droppings in China and their serotypes.C. neoformans colonies which produced brown colonies on caffeic acid-cornmeal agar were found in Twenty-five out of thirty-six samples of pigeon droppings. Fifty-one colonies randomly picked from the positive samples were identified asC. neoformans by a commercially available kit for carbon source assimilation test and Christensens urea agar. Forty (78%) out of the 51 strains were serotyped as A and 11 (22%) as AD. At the same time, seventeen out of nineteen clinical isolates were serotyped as A and 2 as B. There are three findings in our results. One is that onlyC. neoformans var.neoformans strains could be isolated from pigeon droppings, although the varietygattii strains were found in the clinical isolates obtained in the same geographic site in China. The second is that serotype A strains were most frequently seen in natural and clinical materials in the southeast part of China, and serotype AD strains were isolated in pigeon droppings but not in clinical materials. The third is that the coexistence of serotype A and AD cells ofC. neoformans strains in same samples of pigeon droppings were observed.

Identification and subtyping of Trichophyton mentagrophytes by random amplified polymorphic DNA.

Summary. Trichophyton mentagrophytes is one of the most common pathogens for human and animal dermatophytoses and known as a complex species with variable morphology, and ecologic and genetic backgrounds. We performed random amplified polymorphic DNA (RAPD) analysis on 43 human and 18 animal isolates of T. mentagrophytes along with other 10 anamorphic species of dermatophytes and three teleomorphic species of T. mentagrophytes. Using RAPD analysis with primer 5′‐ATGGATC(G,C)(G,C)C‐3′ (ATGS), all T. mentagrophytes strains produced identical band patterns with those of Arthroderma vanbreuseghemii, one of the teleomorphs of T. mentagrophytes, regardless of their phenotypes. Therefore, T. mentagrophytes could be identified by RAPD analysis with primer ATGS. Using RAPD analysis with primer 5′‐ATGGATCGGC‐3′ (ATG) on T. mentagrophytes, human isolates yielded two distinct subgroups related by their colony morphologies at the time of primary isolation from patients. Three morphologic types – cottony, powdery and persicolor – revealed identical bands whereas the granular type lacked one minor band (0.74 kbp). Animal isolates of T. mentagrophytes produced five band patterns and some of them were identical with those of human isolates. With primer 5′‐GAAGGCTCCC‐3′ (OPAO‐15), animal isolates of T. mentagrophytes showed diverse band patterns in contrast to the uniform band pattern of human isolates. These results suggest that RAPD analysis may be a useful tool to identify and subtype T. mentagrophytes complex.

Survival rates of some terrestrial microorganisms under simulated space conditions

In connection with planetary quarantine, we have been studying the survival rates of nine species of terrestrial microorganisms (viruses, bacteria, yeasts, fungi, etc.) under simulated interstellar conditions. If common terrestrial microorganisms cannot survive in space even for short periods, we can greatly reduce expenditure for sterilizing space probes. The interstellar environment in the solar system has been simulated by low temperature, high vacuum (77 k, 4 x 10(-6) torr), and protons irradiation from a Van de Graaff generator. After exposure to a barrage of protons corresponding to about 250 years of irradiation in solar space, Tobacco mosaic virus, Bacillus subtilis spores, Aspergillus niger spores and Clostridiun mangenoti spores showed survival rates of 82%, 45%, 28%, and 25%, respectively. Furthermore. pathogenic Candida albicans showed 7% survival after irradiation corresponding to about 60 years in space.