Yayoi Nishiyama

Candida auris sp. nov., a novel ascomycetous yeast isolated from the external ear canal of an inpatient in a Japanese hospital

A single strain of a novel ascomycetous yeast species belonging to the genus Candida was isolated from the external ear canal of an inpatient in a Japanese hospital. Analyses of the 26S rDNA D1/D2 domain, nuclear ribosomal DNA ITS region sequences, and chemotaxonomic studies indicated that this strain represents a new species with a close phylogenetic relationship to Candida ruelliae and Candida haemulonii in the Metschnikowiaceae clade. This strain grew well at 40 °C, but showed slow and weak growth at 42 °C. The taxonomic description of Candida auris sp. nov. is proposed (type strain JCM15448T= CBS10913T= DSM21092T).

Inhibitory effect of essential oils on apical growth of Aspergillus fumigatus by vapour contact

The inhibitory effect of seven essential oils on the apical growth of hyphae of Aspergillus fumigatus was studied using a bio cell tracer by vapour contact in a sealed vessel. Based on the inhibitory pattern, these essential oils were classified into three groups. The first group, composed of citron, lavender and tea tree oils, stopped the apical growth in a loading dose of 63 μg ml−1 air, but allowed the regrowth of the hyphae after removal of the vapour, indicating fungistatic action. The second group, consisting of perilla and lemongrass oils, stopped the apical growth in a loading dose of 6.3 μg ml−1 air, and did not allow the regrowth after gaseous contact at 63 μg ml−1 air, indicative of fungicidal action. The third group, consisting of cinnamon bark and thyme oils, retarded the growth in a dose of 6.3 μg ml−1 air, stopped it in a dose of 63 μg ml−1 air, and incompletely suppressed regrowth of the hyphae. Gas chromatographic analysis revealed that vapours of essential oils were absorbed on fungal mycelia and agar medium most abundantly by the first group, followed by the second and third groups, reflecting the volatility of the respective groups. Suppression of the apical growth by vapour contact was ascribed to the direct deposition of essential oils on fungal mycelia, together with an indirect effect via the agar medium absorbed.

Antisporulating and respiration‐inhibitory effects of essential oils on filamentous fungi

Summary. Sporulation of four species of filamentous fungi, namely Aspergillus fumigatus, Fusarium solani, Penicillium expansum and Rhizopus oryzae, was suppressed by gaseous contact with citron, lavender and thyme oils and, to a lesser extent, by that of perilla and tea tree oils. Lemongrass and cinnamon bark oils were scarcely active. The antisporulating effect of the essential oils was not observed when they were applied as a solution, indicating that their vapours were the active form. Moreover, exposure of fungal cultures to vapours of the active essential oils caused curling of the tips of aerial hyphae (R. cryzae) or incomplete development of conidiophores (A. fumigatus). These antisporulating effects of the vapourizing essential oils seemed to be correlated with their respirationinhibitory activity, rather than with their growthinhibitory activity.

Protective effects of farnesol against oral candidiasis in mice

Farnesol is known as a quorum‐sensing molecule for Candida albicans and is recognized to play pathogenic roles in Candida infection. To assess the possible role of farnesol in mucosal C. albicans infection, the effects of farnesol treatment against experimental oral candidiasis in mice were examined. Prednisolone‐pretreated ICR mice were orally infected with C. albicans and 3, 24 and 30 hr later the animals were orally given farnesol. Forty‐eight hr later they were killed for observation. Farnesol treatment in a dose ranging between 1.125 and 9 μmol/mouse showed a protective effect against oral candidiasis in a dose‐dependent manner, at least as estimated by symptom scores of tongues. At 9 μmol/mouse it decreased bodyweight loss. Histological studies of 2.25 μmol/mouse farnesol‐treated animals indicated that farnesol suppressed mycelial growth of C. albicans on the surface of tongues, but microbiological study did not prevent the change of CFU of C. albicans cells not only on tongues but also in feces, kidneys and livers. These results suggest that farnesol has very characteristic roles in protection against mucosal candidiasis.

Mechanism of Action of Efinaconazole, a Novel Triazole Antifungal Agent

ABSTRACT The mechanism of action of efinaconazole, a new triazole antifungal, was investigated with Trichophyton mentagrophytes and Candida albicans. Efinaconazole dose-dependently decreased ergosterol production and accumulated 4,4-dimethylsterols and 4α-methylsterols at concentrations below its MICs. Efinaconazole induced morphological and ultrastructural changes in T. mentagrophytes hyphae that became more prominent with increasing drug concentrations. In conclusion, the primary mechanism of action of efinaconazole is blockage of ergosterol biosynthesis, presumably through sterol 14α-demethylase inhibition, leading to secondary degenerative changes.

In vitro activity of novel imidazole antifungal agent NND-502 against Malassezia species.

The in vitro activity of NND-502, a novel antifungal imidazole compound, was tested against the three major Malassezia species by an agar dilution method with modified Dixon medium and compared with the activities of three reference antifungal drugs of topical use, lanoconazole (LCZ), bifonazole (BFZ) and terbinafine (TBF). The geometric mean (GM)-MICs of NND-502 for 25 strains of M. furfur, 15 strains of M. sympodialis and ten strains of M. slooffiae were approximately 1.4, 0.1 and 1.0 mg/l, respectively, showing the greatest activity against M. sympodialis and the least against M. slooffiae. These values were similar to that of LCZ, but four to 69 times lower than that of BFZ and two to three times lower than that of TBF. The results suggest that NND-502 might be beneficial in the treatment of Malassezia-associated skin diseases.

Fungal flora on board the Mir-Space Station, identification by morphological features and ribosomal DNA sequences.

This report is on the morphological and molecular biological identification, using 18S‐ and ITS1‐rDNA sequences, of the “space fungi” isolated on board the Russian Mir‐Space Station as the major constituents of the fungal flora. The six fungal strains were isolated from air by using an air sampler or from condensation. Strains were identified as Penicillium chrysogenum, Aspergillus versicolor, or Penicillium sp. by both methods. The species of space fungi were common saprophytic fungi in our living environment, potential pathogens, and allergens. This study concluded that the environment on board the space station Mir allows the growth of potentially pathogenic fungi as true in residential areas on the earth. Therefore, to prevent infection or other health disorders caused by these fungi, easy and reliable methods should be established to survey the fungal flora in a space station.

A novel mechanism of fluconazole resistance associated with fluconazole sequestration in Candida albicans isolates from a myelofibrosis patient.

A series of 10 strains of Candida albicans, from TIMM 3309 to TIMM 3318, were repeatedly isolated in one myelofibrosis‐complicated patient with recurrent candidemia. The latter five isolates, from TIMM 3314 to TIMM 3318, became suddenly resistant to fluconazole during the 10 to 16 weeks after antimycotic therapy. We investigated the resistant mechanism of fluconazole using one susceptible isolate and two of the five resistant isolates in the series. The ergosterol synthesis by cell‐free extracts from the two resistant isolates was less susceptible to fluconazole partly as a result of a decreased affinity of cytochrome P‐450. Unexpectedly, these two resistant isolates showed higher levels of an intracellular accumulation of [3H]fluconazole than the susceptible isolate and the control strain of C. albicans ATCC 10231. In the resistant isolate, TIMM 3318, most intracellular incorporated fluconazole was distributed in the 12,000 × g pellet (P‐120) fraction by centrifugation unlike the two susceptible strains. An observation of the ultrastructure of TIMM 3318 showed the most notable alteration to be the characteristic appearance of numerous vesicular vacuoles (diameter, 150 to 400 nm); these vacuoles were not observed, however, in either of the susceptible strains. A direct observation of the subcellular fraction prepared from TIMM 3318 by the electron microscopy negative‐staining method suggests that most of the vesicular vacuoles were recovered in the P‐120 fraction. These results suggest that fluconazole sequestration caused by vesicular vacuoles of the resistant isolate might act as a novel mechanism of fluconazole resistance besides the decreased affinity of cytochrome P‐450.

Microbe-I: fungal biota analyses of the Japanese experimental module KIBO of the International Space Station before launch and after being in orbit for about 460 days

In addition to the crew, microbes also find their way aboard the International Space Station (ISS). Therefore, microbial monitoring is necessary for the health and safety of the crew and for general maintenance of the facilities of this station. Samples were collected from three sites in the Japanese experimental module KIBO on the ISS (air diffuser, handrail, and surfaces) for analysis of fungal biota approximately 1 year after this module had docked with the ISS. Samples taken from KIBO before launch and from our laboratory were used as controls. In the case of KIBO, both microbe detection sheet (MDS) and swab culture tests of orbital samples were negative. The MDS were also examined by field emission‐scanning electron microscopy; no microbial structures were detected. However, fungal DNAs were detected by real‐time PCR and analyzed by the clone library method; Alternaria sp. and Malassezia spp. were the dominant species before launch and in space, respectively. The dominant species found in specimens from the air conditioner diffuser, lab bench, door push panel, and facility surfaces on our laboratory (ground controls) were Inonotus sp., Cladosporium sp., Malassezia spp., and Pezicula sp., respectively. The fungi in the KIBO were probably derived from contamination due to humans, while those in our laboratory came from the environment (e.g., the soil). In conclusion, the cleanliness in KIBO was equivalent to that in a clean room environment on the ground.

Invasion process of Candida albicans to tongue surface in early stages of experimental murine oral candidiasis

We analyzed the morphologic and microbiologic aspects of the process of adhesion and invasion in the early stages of Candida albicans oral infection in a murine system. ICR mice were anesthetized by intramuscular injection with chlorpromazine chloride and then orally inoculated by swabbing with the C. albicans yeast cells. Their tongues were resected 1-3h after inoculation, washed sequentially with a physiological saline and 0.25% trypsin-solution and then homogenized. The number of viable C. albicans cells on the tongue surface was counted and fround to increase from 1-3h after inoculation. Most of the Candida cells attached to the tongue surface were present in clusters, mainly located in the gaps between lingual papillae and were covered with a mucoidal substance. By 3h after inoculation, these clusters frequently formed mycelia and could not be easily detached from the tongue surface by trypsin treatment. Observation of SEM and histological sections stained by Fungiflora Y revealed that the Candida hyphae at 3h stretched out of the cluster and entered the tongues through the surface. These results indicate that Candida hyphae begin to invade the tongue surface within 3h after inoculation and suggest that the mucus-like substance covering these cells may have an important early role in the interaction between the Candida cells and the tongue mucosal epithelium.