The modern antimycotic era began with the introduction of griseofulvin in the 1950's, which had a tremendous impact on the management of dermatophytoses. Subsequent advances in therapy have lowered our threshold to treat recalcitrant cutaneous mycoses, such as onychomycosls. Newer generation antifungals including the triazoles, itraconazole and fluconazole, as well as the allylamine, terbinafine, may significantly reduce the prevalence of onychomycosis, as fungal nails are no longer incurable. The AIDS epidemic has been associated with an increase in the number of cutaneous and systemic mycotic infections. In spite of recent advances, organisms recalcitrant or resistant to therapy are common. Newer antifungal agents, improved diagnostic techniques, and standardization of fungal susceptibility testing are required to adequately treat patients with systemic and cutaneous mycoses.

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Candida albicansの二形性変換(酵母形から菌糸形への形態変換)は病原性発現の一因子と考えられている.二形性のメカニズムおよびこの現象がどの程度病原性発現に寄与しているかについて以前から膨大な研究がなされてきたが,詳細はいまだ不明である.近年,Candidaの研究分野にも急速に分子遺伝学の技術が導入され,二形性変換の研究分野でも遺伝学的な背景を明らかにすることを目的とした新たな展開が始まっている.この小論は二形性変換における遺伝子発現研究に焦点を当てて最近の知見を紹介し,この現象を分子生物学的に解き明かすためのアプローチについて考察した.

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Adaptation of Aspergillus niger to amphotericin B (AMPH) and two imidazoles (miconazole (MCZ) and ketoconazole (KCZ)) was observed at a single hypha level with a continuous measurement system. It was found that a test hypha adapted to MCZ or KCZ was also adapted to AMPH but that a hypha adapted to AMPH was not adapted either to MCZ or KCZ. These adaptation phenomena to respective agents did not occur after incubation in the medium supplemented with ergosterol. The cross adaptation phenomena were suspected to be due to modification of the synthesis pathway of ergosterol and related derivatives.

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The non-dialyzable fraction of culture filtrate antigen of Aspergillus fumigatus was fractionated by anion exchange chromatography and gel filtration, connecting each column of Mono Q and Superose 6 sequentially. Fractions which had a high protein and reacted strongly with an immunoglobulin-M monoclonal antibody (MAb) 8-3C F5 G9, as determined by enzyme-linked immunosorbent assay (ELISA), were pooled to give four antigenic fractions (F1-F4). Gas-liquid chromatography analysis of the 4 antigenic fractions showed the presence of mannose and galactose. An immunoelectron microscopic experiment with MAb revealed the distribution of antigen on the surface of the hyphae. The fractions had no proteolytic activity. Fraction 4, relatively small in its molecular size but with the highest recovery of the reactivity with MAb among F1 to F4, was selected for further characterization. Antigenic reactivity of F4 with MAb was heat-stable but fully retained after sodium metaperiodate treatment. Pronase E almost destroyed the antigenic reactivity of F4 with MAb. MAb affinity chromatography of F4 gave the specific antigen, F4-AF. In both sodium dodecyl sulfate-polyacrylamide gel electrophoresis by Coomassie blue staining and Western blotting with MAb, F4-AF showed a single band with molecular mass of 66 kDa.

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BioCell-Tracer装置は単一菌糸の先端を追跡し,その成長速度を測定可能にした装置である.本装置を用いAspergillus fumigatus単一菌糸の成長速度を測定しmiconazole(MCZ)の薬剤効果を検討した.その結果,薬剤濃度0.8(1/4MIC)から6.25μg/ml(2MIC)では約1時間で菌糸成長抑制を認めた.また,12.5(4MIC)から100μg/mlでは20分以内に各々成長抑制を認めた.更に,菌糸先端の形態変化でも各薬剤濃度で異なり,0.8-1.6μg/mlでは徐々に成長を停止して菌糸先端は棍棒状になり,3.2-25μg/mlでは先端で異常な短分岐が観察された.次にMCZを7.6μg/ml含む患者血清で効果検討をした結果,菌糸は約90分後に成長抑制を示し,その後の菌糸成長の回復は認められなかった.即ち,本試験法は菌糸に対するMCZの直接的な薬剤併用効果を経時的および形態学的に検討することが可能であることを示唆すると共に,MCZの投与を受けている患者血清もまたA.fumigatusの菌糸の成長を十分抑制することが認められた.

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Aspergillus fumigatus 15株,A. neoellipticus 19株およびNeosartorya fischeri 13株についてミトコンドリアDNA(mtDNA)のrestriction fragment length polymorphism分析を行った.切断パターンによりA.fumigatusは3タイプ,A. neoellipticus 6タイプ,N. fischeri 2タイプに分けられ,切断パターンは種の同定,菌株の識別に有用と考えられた.これら各タイプについてmtDNAの塩基置換率の推計値を基にして系統樹を作成した.その結果,遺伝的にはA.fumigatusとA.neoellipticusは別種,A.neoellipticusとN.fischeri(12株)は極めて近く種内変異としてまとめられ,A.neoellipticusはN.fischeriのアナモルフである可能性が示唆された.

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漢方医薬である十全大補湯製剤TJ-48の深在性真菌症に対する有効性の有無を明らかにするため,マウス感染モデルを用いて,本剤の経口投与による予防及び治療効果を検討した.正常マウスおよび,シクロホスファミド(CY)前処置によって,免疫抑制状態にしたマウスにCandida albicansを静脈内接種し,1日1回5日間,TJ-48を連日投与した.その結果CY前処置の有無にかかわらず,2g/kgまで用量依存的に,生存期間が延長した.TJ-48投与マウスにおいては,腎臓内生菌数の減少が認められた.この治療効果は,TJ-48を菌接種前に投与した場合の予防的効果より,明確に観察された.又,フルコナゾールとTJ-48を併用することにより,より強力な治療効果が得られた.TJ-48の治療効果は,Aspergillus fumigatus感染モデルでも認められたが,Cryptococcus neoformans感染モデルでは明確でなかった.以上より,実験的真菌症に対して十全大補湯が治療効果を有すると結論した.

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The major membrane phospholipids of Candida albicans (C. albicans) are phosphatidylcholine (PC), phosphatidylethanolamine (PE), phosphatidylinositol (PI), and phosphatidylserine (PS). In Saccharomyces cerevisiae (S. cerevisiae), the most abundant component PC is considered to be synthesized mainly by two pathways; stepwise methylation pathway of PE produced from PS by decarboxylation (de novo pathway) and diacylglycerol (DG)/CDP-choline pathway. However, relatively little information is available regarding the phospholipid biosynthesis in C. albicans. In order to clarify the phospholipid metabolism of this organism, we have investigated the ³²Pi incorporation into different phospholipids by continuous and pulse-labeling in cells grown in the synthetic medium (SM) and yeast extractproteose peptone-dextrose (YPD) medium.
The continuous labeling with ³²Pi in YPD medium showed that the decreases in radioactivity in acidic phospholipids (PA, PI and PS) were followed by the great increase in PC, suggesting the existence of de novo pathway. In SM cultured cells showing no growth, PS and PI were heavily labeled, while PC was much less labeled. By the switch from SM to YPD, the rapid increase in ³²P-labeled PC occurred at the expense of PI and PS, suggesting that the principal PC synthesis for membrane biogenesis is via the de novo pathway in C. albicans.

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