资料来源: NaturWissenschaften (Springer-Verlag 2001) PDF档(有图)请洽维中 Javier Garcia-Guinea · Victor Cardenes Angel T. Martinez · Maria Jesus Martinez Received: 31 May 2001 Accepted in revised form: 23 June 2001 Published online: 17 July 2001 Fungal bioturbation paths in a compact disk Abstract We report here on bioturbation traces, with micro-dendrite textures, composed of a mixture of altered aluminum and polycarbonate, which have been developed in a common compact disk (CD), destroying information pits. Fungal hyphae proliferated in these deteriorated zones, and Geotrichum-type fungus was isolated from surface-sterilized CD fragments. The severe biodeterioration described is attributed to the slow growth of this arthroconidial fungus on the CD material in the tropical indoor environment of Belize, Central America (~30°C, ~90% humidity). 亚佛‧贾西-朱尼亚, 维克多‧卡登尼斯 安琪拉‧T‧马丁尼兹, 玛利亚‧耶稣‧马丁尼兹 (这名字也太... Orz) 接稿日: 2001年5月31日 最终修正版收稿日: 2001年6月23日 上线发行日: 2001年7月17日 CD片上的真菌生物扰动轨迹 摘要 在此我们要报告的,是一种在普通CD上发展,由变质的铝和聚碳酸酯组成,且呈现微 树突状组织的生物扰动作用痕迹,及其对CD资讯槽所造成的破坏. 真菌菌丝在变质的区域 增生 -- 消毒过的CD碎片表面,分离出了某种地丝菌型的真菌. 这严峻的生物腐蚀,正是 贝里斯的热带环境中(室内,~30°C,~90%相对湿度),该节-分生胞子型的真菌於CD上缓慢 生长所致. Introduction The fungus Geotrichum was described more than 175 years ago as a white hyphomycete with hyphae disarticulating into cylindrical spores. It is commonly found in close proximity to human life. Geotrichum species are present in foodstuffs such as cereals, cheese, yeast and vegetables (Rodionova et al. 1995), in water (Baudoin and Davis 1986; Molimard et al. 1995), in insects (Fischer 1999), as airborne spores (Cosentino and Palmas 1991) and even in the human body (Hrdy et al.1995). 简介 地丝菌型的真菌早在175年以前,就被形容为一种"白色的丝胞属真菌,菌丝会脱离出 圆柱状的胞子". 日常生活环境中很容易可以找到它. 地丝菌会在吃的里头出现,像是 谷物,起司,酵母,和蔬菜,也可能在空气中(胞子),水里,昆虫,甚至跑到人的身上. Compact disks are used worldwide to store sound, music, images, software, text files, and that they are a secure persistent store is taken for granted. A CD consists of a pregrooved polycarbonate substrate, a metal reflector, an overcoat and often a dye (the latter in CD-R). CD producers (i.e., Philips, Sony) advertise the excellent properties of CDs, describing two threshold temperatures involved with the softening of polycarbonate at 170°C and with a transition of the dye at 265°C (Holtslag et al.1992). There have been few reports describing biodegradation of polycarbonates because they present great chemical stability; in addition, their biocompatibility allows polycarbonates to be used in medical implants (Mathur et al. 1997). Japanese research groups have studied the biodegradability of aliphatic polyestercarbonate (Imada al. 1999) and have isolated proteobacteria (Pseudomonas sp. and Variovorax sp.) from degraded polycarbonates in the river Ibaraki, Japan (Suyama et al.1998). Additional research topics on biodegradability of polycarbonates are their cytotoxicity, water resistance and the ability of fungi to penetrate poly- carbonate micropores, as has been reported by Bardage and Daniel(1997). CD广泛的用来储存声音,音乐,影像,软体和文字,被视为是安全持久的储存媒体. CD片是由有沟纹的聚碳酸酯作基底,上面是金属的反射体,再上层是外壳且通常有染色 (近期的CD-R). CD制造商(如新力,飞利浦)宣传CD片的优异特性,并说明其两个门槛 温度,分别为聚碳酸酯软化温度170°C,染色层转化温度265°C(注1). 目前已有少数 报告叙述了聚碳酸酯的生物降解,毕竟它们的化学性质实在稳定. 另外,生物上的兼容 性让聚碳酸酯被拿来制作医疗用植入管. 日本的团队研究了脂肪族聚酯碳酸酯的生物 可降解性,且从取自日本茨城县河流中降解的聚碳酸酯上,分离出变形菌. 其他关於 聚碳酸酯生物可降解性的研究,尚有它的细胞毒性,防水性以及真菌对微孔的穿透性 (注2). 注1: 是指熔点Tc跟玻璃转化温度Tg?? 似乎不怎麽重要,至於想看CD加热後会怎样的, 请点 http://pslc.ws/fire/polyburn/polyburn.htm 注2: ()中的期刊文章就省略不翻,以下同. Compact disk sample Last summer, one of the authors, Victor Cardenes, made a social visit to Belmopan, Cayo District, Belize, Central America, where he stayed with his friends Miguel Cano and Zayda Villar. While talking about life in Belize, Victor's hosts showed him a CD (Fig.1a-c), which contained clearly visible biodeterioration paths on its surface and joked, "Look, life is so strong here, we even have something that eats CDs". The paths on the CD caught Victor's (a geologist) interest because of their similarity with the trails left by burrowing worms, a subject he had recently been studying. Miguel and Zayda gave him the affected CD (by the Kronos Quartet, issued by Elektra Entertainment of Warner Communications, Baltimore,USA) and he brought it back to Spain with him at the end of his trip. At the Spanish Natural Science Museum of the CSIC, Victor decided, with author Dr.Garcia-Guinea, to observe the CD with polarizing and scanning electron microscopes. Together they observed the paths, which formed fractal structures and destruction of the pits and halls in the aluminum on the disk. In search of fungi or bacteria, the disk was taken to the authors, Dr M.J. Martinez and Dr A.T. Martinez, biologists at the Centro de Investigaciones Biologicas (CIB) of the CSIC. Here, under a confocal microscope, the hyphae of a fungus became clearly visible. The CD was cut into two pieces – one for the Museum and the other placed in the CIB in a Petri glass at room tempera- ture and 100% humidity, where the fungus continues to grow. CD试片 去年暑假,作者之一的维克多‧卡登尼斯,到位於中美洲贝里斯,卡右区的贝尔墨邦 访问,和同行的两位友人米高‧卡诺和捷达‧维拉在那儿作了停留. 当聊到贝里斯的生活 时,饭店老板拿了张CD给他看(图1a–c),并开玩笑说: "瞧,我们这里东西的生命力特强, 连CD都被当成食物给啃了." 该CD片上有着明显的生物腐蚀痕迹. 那轨迹兴起了卡登尼斯 (身为地质学家)的好奇心,因为看起来很像他才刚刚研究过的,会钻洞的蠕虫留下来的 足迹. 米高和捷达把那张受到侵袭的CD(注1)交给卡登尼斯,好让他在旅途结束时带回 西班牙. 在西班牙CSIC(注2)所辖的自然科学博物馆中,维克多和另一位作者贾西-朱尼亚博士 决定用偏光电子扫描显微镜去观察这张CD. 他们发现那些轨迹呈现不规则碎片形,而碟片 铝层的资讯槽也遭到破坏(注3). 为了研究到底是真菌还是细菌,片子又转交给另外两位 作者安琪拉‧T‧马丁尼兹和玛利亚‧耶稣‧马丁尼兹. 他们是CSIC旗下CIB(注4)的研究 人员. 在雷射扫描共轭焦显微镜之下,真菌菌丝清晰可见. 後来CD被切成两半,一半拿回 博物馆,另一半则留在CIB,且置於培养皿中,以室温/100%相对湿度让真菌继续生长. 注1: 文中CD是Kronos弦乐四重奏在Elektra的录音,但现在厂牌好像变成nonesuch? 因原文语焉不详,只能推算是2000年以前发行的,无法确定是哪张. 请参考连结 http://kronosquartet.org/records/index.php 注2: Consejo Superior de Investigaciones Cientificas 相当於我们的国科会. 注3: CD上的0跟1好像分别叫作pit跟land,我不晓得文中的"pits and halls"是在说啥. 注4: Centro de Investigaciones Biologicas Results and discussion The study of these paths in the transparent zones of the CD, by transmi- tted and reflected light microscopy, X-ray diffraction and scanning electron microscopy, revealed dendrite microstructures of aluminum–polycarbonate mixtures and the complete loss of the original information pits. Fungal growth and partial degradation of metal and polycarbonate were detected by direct observation of the original CD with a Zeiss photomicroscope coupled to a CCD camera. The hyphae growing in the natural deteriorated areas of the CD were easily observed in the peripheral region under the light microscope (Fig.2a-c). To isolate and identify the fungus, small fragments (2×4 mm) from the deteriorated areas of the Belize CD were superficially sterilized by immersion in 70% ethanol (3 min), washed with sterile water and plated on malt extract (2%) agar and malt extract (2%) + glucose (2%) agar containing cloramphenicol (12 mg/l), and incubated at 26°C in darkness. From the steri- lized CD fragments, a unique type of fungal colony with white aerial mycelium was detected in both media at different incubation times. When the fungus was grown on malt extract agar, white colonies with a velvety surface and a growth rate of 8 mm/day were obtained. Under the light microscope, the fungus dis- played septate hyphae with dichotomous branching. The morphological characte- ristics of the strain isolated from the CD did not change with the presence of glucose in the medium (Allermann et al. 1978). The fungus presents two types of hyphae (2.5-3 and 4-6 μm wide), the main hyphae disarticulating into cylindrical cells (arthroconidia) with 4–6×6–13 μm size. The oldest arthroconidia became barrel-shaped. A difference between arthroconidia from lateral and main branches was not observed. 结果与讨论 在研究这片CD透明区域上的痕迹时,共动用了穿透式,反射式显微镜,X光绕射仪, 电子扫描显微镜,结果呈现出由铝-聚碳酸酯混杂成的树突状的微组织,而原本该有的 资讯槽(pits)则一整个消失. CD上真菌的生成,以及金属跟聚碳酸酯有部份被降解的 情形,都被接上数位相机的蔡司可照像式显微镜给直接捕捉到了. 在光学显微镜下可 轻易观察到,菌丝在CD圆周附近自然变质的区域增生(Fig.2a-c). 为了隔离并确认真菌, 先从这张贝里斯CD的变质部份取下小的碎片(2×4 mm),并做表面消毒,依序为浸放在70% 乙醇(3分钟),以无菌水清洗,分置於麦芽抽出物(2%)培养基,麦芽抽出物(2%)加葡萄糖 (2%)培养基,内含氯丝菌素(12 mg/l,注1),最後放在26°C的暗室中培养. 各个不同的 孵育时段中,两份消毒过的CD碎片皆侦测到一种伴随着白色气生菌丝,十分独特的真菌 菌落. 麦芽抽出物培养基这边,表面光滑的白色菌落生长速度为8毫米/天. 用光学显微 镜就可看到有隔膜菌丝,并有岔状的分支. 对CD上分离出的的本菌种而言,其形貌特徵, 不因葡萄糖的的存在与否而有所不同. 该真菌有两种菌丝(2.5-3&4-6微米宽),主干的 菌丝脱出圆柱状的细胞(桶状节生胞子),大小为4-6x6-13微米(注2). 最老的桶状节生 胞子则变成中广型的桶状. 而侧向跟主分支上的节生胞子,并未发现两者有什麽不同. 注1: 原文cloramphenicol应是chloramphenicol误植(?!) 注2: 我看不懂4-6x6-13微米是在说啥,或许6x6是截面积,4~13是长度吧. @@" 事实上後半段翻得还颇语无伦次 +.+ The above characteristics suggested that the fungus isolated from the CD should be classified in the genus Geotrichum. This genus is restricted to ascomycetous species of arthroconidial fungi, whereas the morphologically related genus Trichosporon is reserved for basidiomycetous fungi. The study of carbohydrate composition of fungal cell walls can be used as a taxonomical indicator to differentiate between related fungi with ascomycetous and basi- diomycetous affinities in these and other genera. With this in mind, at the CIB, the mycelium was hydrolyzed (5 M trifluoroacetic acid, 90 min, 120°C) and neutral sugars analyzed as alditol acetates by gas chromatography using a SP-2380 capillary column programmed from 210°C to 240°C, with a 3-min ini- tial hold, a heating rate of 15°C min–1 and a final time of 7 min. The pre- sence of xylose is most often characteristic of basidiomycetous arthroconidial fungi. The absence of this sugar after GC analysis of the cell walls of the fungus isolated from the CD confirmed that it presents ascomicetous characte- ristics. 从上面这些特性看来,CD上的这些真菌应该是归类在地丝菌属. 本属限制为节-分生 胞子的子囊菌种,而其他形态类似的像是丝包菌则分类在担子菌. 真菌纤维细胞壁的碳水 化合物组成,是区别子囊菌和担子菌等近似的属的指标. 基於此理论,她们在CIB先将菌丝 水解(5M三氟醋酸,90分钟/120°C),藉醣醇乙酸酯分析其中性糖组成,方法是使用SP-2380 毛细层析管,从210~240°C间开始,先加热3分钟後,再以每分上升15°C的速度加热到7 分钟,作气相色层分析. 若是节-分生胞子的担子菌,几乎都会出现木糖,但此次针对CD片 真菌细胞壁所做的气相色层分析并未出现,故可确定他呈现出子囊菌的特徵. This agrees with other morphological criteria, such as the absence of blastoconidia. The fungus displays some of the characteristics reported for Geotrichum candidum although the main branches of this fungus are wider. The fact that intense biodeterioration in CDs, destroying the information pits, has been caused by a common fungus (a species close to G. candidum) leads to speculation on the future of compact disks as a secure persistent storage medium for sound, image and computer files. The Belize fungus has been well studied, and thanks to the information explosion and the Internet, we are receiving reports and samples from Guatemala, Mexico, Panama, Costa Rica, Hong Kong and other tropical countries. We now have three pending tasks (1)to determine the geographical and climatical distribution of this bio- deteroration phenomenon, (2) to study the interaction of the fungus with the host materials of the CD and (3) to confirm its systematic relationships by molecular biology techniques. 实验分析所得,也符合仅依型态标准去判断的结果,好比说,它并没有芽生分生胞子. 这真菌呈现出若干已知的白色地丝菌的特点,虽然其主要分支较粗些. 而摆在眼前的,是 CD上由(某种接近白色地丝菌的)真菌引起的严重生物腐蚀,且资讯槽不保,这让人思考CD 是否真为持久可靠的影音/档案储存媒体. 贝里斯的真菌已经过研究,拜资讯爆炸和网际 网路盛行之赐,我们又收到许多来自瓜地马拉,墨西哥,巴拿马,哥斯大黎加,香港和其他 热带国家的样本与相关回报. 现在,我们仍有三个悬而未决的任务: 1. 针对有这种现象的地区,找出其地域性和天候性的分布. 2. 探讨真菌跟其宿主CD材质间的关系. 3. 利用分子生物技术确认其系统性的关连. Acknowledgements We are grateful to A. Prieto for gas chromatography ana- lysis and Angeles Guijarro for technical assistance in Madrid. The work was supported by the project 1FD97–0959-CO3–01 of the Plan Nacional de Inves- tigacion Cientifica y Desarrollo Tecnologico (Spain). Thanks to Josefo Bedoya for his help with the scanning electron microscope. Thanks also to Paul Giblin for critical revision of the manuscript. 我们对A.普利托和安吉莉斯‧古加洛分别在气相色层分析与技术上给予的帮助表达 由衷感激. 本辑为(西班牙)国家科学技术研究部第1FD97–0959-CO3–01号计画. 谢谢 乔瑟夫‧比多雅对电子扫描显微镜的支援,也同样的感谢保罗‧吉卜林在原稿修订上给的 建议. Fig 1 a Deteriorated compact disk from Belize, Central America. b Detail of the bioturbation paths visible to the naked eye. c Scanning-electron microscopy image of a deteriorated fragment of the Belize compact disk showing dendrite textures of the bioturbed aluminum–polycarbonate mixture 图1 a 中美洲贝里斯,毁损的(被腐蚀的)CD b 生物扰动作用的痕迹用肉眼也可以看得清楚 (300μm可以用肉眼看得清楚??) c 腐蚀的碎片於电子扫描显微镜秀出的图中,有着生物扰动下由铝和聚碳酸酯混杂成的 树突状微组织 Fig 2 a Light microscopy image of the bioturbation paths displaying the fungal growth in the peripheral region of the compact disk alteration. b Hyphae and disarticulated conidia (arthroconidia) in pure culture of the Geotrichum-type fungus isolated from the deteriorated compact disk (light microscopy). c Detail of typical conidial chains produced by this fungus (light micro- scopy) 图二 a 光学显微镜拍到的图上,生物扰动纪录痕迹可看出真菌在CD圆周附近变质的区域生长 b 自CD分离出的地丝菌型真菌,做纯培养後所得菌丝和脱出的分生子柄(节-分生胞子) c 该真菌特有的分生胞子链 待续..... -- 音响的莫非定律 : 每当朋友要来家里听音乐时,音响不是莫名的故障,就是衰声连连. --

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