近平滑念珠菌形态多样性及毒力因子的分泌活性研究

目的比较近平滑念珠菌不同临床分离株的菌落和细胞形态及其分泌型天冬氨酸蛋白酶(SAP)酶活性,阐明该病原真菌的形态多样性及毒力差异。方法分别用Lee’s Glc NAc、Lee’s glucose、Spider 3种培养基于37℃培养7株近平滑念珠菌临床分离株,观察各菌株的菌落及细胞形态;用牛血清白蛋白培养基(BSA)于30℃培养7株近平滑念珠菌,通过观察形成晕圈能力的强弱确定其毒力因子(SAP)的分泌活性。结果 7株近平滑念珠菌临床分离株在不同条件下菌落表型有较明显的不同,菌落有光滑型、皱褶型及火山口样型,且染色程度不同,各分离株细胞在形成酵母相或菌丝相的能力方面呈现出不同程度的差异;SAP酶活性观察实验中检测到各菌株水解晕圈大小从2.55 cm到1.55 cm各不相等。结论近平滑念珠菌临床分离株具有形态多样性,各菌株SAP酶活性明显不同,提示菌株间的毒力有明显差异。     

硼酸抗白念珠菌作用的试验研究

目的探讨硼酸对白念珠菌临床分离株菌株的生长状态和细胞形态的影响。方法用加入0%、0.1%、0.15%硼酸(w/v)的Lee’s glucose培养基于25℃和37℃分别培养8株临床分离的白念珠菌,观察各菌株在不同培养条件下生长状况。结果 25℃培养条件下,0.1%的硼酸明显抑制白念珠菌生长,其中对HJ065的抑制效果最明显,0.15%的硼酸进一步抑制白念珠菌生长,并且SC5314、HJ058菌株的菌丝生长能力随硼酸浓度增高而减弱。37℃培养条件下,硼酸对白念珠菌生长状态和菌丝生长能力的抑制效果较25℃更明显。相同培养条件下,硼酸对不同CAI重复数的白念珠菌的抑制效应存在差异。结论硼酸对白念珠菌临床分离菌株的抑制作用表现出明显菌株的差异性。其抑制作用与培养温度有关:表现为生理温度下抑制作用更明显。硼酸对8株白念珠菌抑制作用与CAI的重复数无明显相关性。     

血培养分离出212株念珠菌的菌种分布及耐药性分析

目的分析血培养分离出的念珠菌的菌种分布及耐药分析,为临床念珠菌血症的诊治提供理论依据。方法回顾性分析2017年1月—2020年12月期间,本院血培养分离出的念珠菌的菌种分布、药敏试验结果及患者血培养分离出念珠菌前后96 h内的G试验结果。结果血培养中分离出念珠菌314例,阳性率2.1%,其中非重复分离株212例。检出率最高的是近平滑念珠菌(72株,34.0%),其次是白念珠菌(55株,25.9%)和光滑念珠菌(28株,13.2%)。念珠菌检出率最高的科室为ICU(62株,29.2%),其次是新生儿科(39株,18.4%)和血液科(20株,9.4%)。检出的212株念珠菌除一株近平滑念珠菌为两性霉素B的非野生株,其余均为两性霉素B的野生株。白念珠菌对唑类药物的敏感率超过90%。但光滑念珠菌和热带念珠菌对唑类药物的敏感性较低。血培养分离出念珠菌的前后96 h内,G试验的阳性率为73.7%。结论本院念珠菌检出率前3位分别是近平滑念珠菌、白念珠菌和光滑念珠菌。光滑念珠菌、热带念珠菌对唑类药物敏感性比较低,在经验用药时需要合理选择抗真菌药物。G试验对念珠菌血症有较高的价值,需要对结果进行动态监测。     

外阴阴道念珠菌病的病原检测及体外药敏试验

目的了解STD门诊患者外阴阴道念珠菌病的菌种分布及对常用抗真菌药物的敏感状况,为临床治疗提供参考。方法取STD门诊疑似念珠菌感染患者的阴道分泌物标本,用沙氏琼脂培养基分离纯化菌株,然后用ATB ID 32C酵母菌鉴定系统和ATB Fungus3药敏系统进行菌种鉴定与药物敏感试验。结果950份标本中检出306株念珠菌,占32.21%,其中白色念珠菌226株,阳性率为73.86%;光滑念珠菌37株,阳性率为12.09%;热带念珠菌24株,阳性率为7.84%;近平滑念珠菌13株,占4.25%。药敏结果显示：念珠菌对5-氟胞嘧啶（5-FC）;两性霉素B（AMB）,伏立康唑（VRC）耐药率〈6%;近平滑念珠菌对5种抗真菌药物均敏感;白色念珠菌对氟康唑（FCZ）、伊曲康唑（ICZ）耐药率依次：9.29%、11.06%,光滑念珠菌药耐药率依次为37.84%,67.57%,热带念珠菌药耐药率依次为8.33%、12.5%。结论STD门诊患者阴道念珠菌检出率明显高于普通妇科门诊患者,检出菌种仍以白色念珠菌为主,但光滑念珠菌和热带念珠菌生殖道感染有明显上升趋势;念珠菌对两性霉素B、5-氟胞嘧啶敏感性较高,对唑类药物有耐药菌株。因此,念珠菌鉴定和药敏监测是非常必要的,临床医生应根据药敏结果合理使用抗真菌药物。     

727株真菌的分布与药敏分析

目的对727株真菌的分布及药敏试验进行分析。方法用沙保罗培养基分离培养真菌,用API20C AUX生化鉴定及ATB FUNGUS药敏板。结果 727株真菌中白色念珠菌438株,热带念珠菌155株,光滑念珠菌73株,近平滑念珠菌23株,新型隐球菌15株。5-H氟胞嘧啶、两性霉素B、制霉菌素、咪康唑、益康唑和酮康唑的敏感率分别为92.0%、97.6%、94.6%、94.7%、56.3%、62.9%和70.4%,结论真菌感染仍然以白色念珠菌为主。两性霉素B、制需菌素和一氧胞密啶敏感率较高,而咪康唑、益康唑和酮康唑均出现不同程度的耐药,且呈上升趋势,这需要临床慎选针对性强且副作用低的药物,减少耐药菌株出现。     

念珠菌血流感染临床与实验室特点分析

目的根据血培养真菌阳性标本,进行病原学与药敏试验分析,探讨念珠菌在血流感染中的临床和实验室特点。方法收集某院2012年6月至2018年9月的血培养结果为真菌的临床资料,对真菌种类、阳性报警时间(TTP)、标本送检科室、病原菌来源和氟康唑药敏试验结果进行分析。结果血培养分离出真菌共86株,非重复分离株为50株,44株为念珠菌属。排在前3位的念珠菌分别为白念珠菌18株(40.91%),近平滑念珠菌和光滑念珠菌各10株各占(22.73%);近平滑念珠菌TTP[(56.14±7.22)h]最长,热带念珠菌TTP[(19.36±1.24)h]最短,白念珠菌[(34.67±2.98)h]与光滑念珠菌[(38.16±4.12)h]TTP接近居中;念珠菌血症前3位科室为神经外科10株(22.73%)、肿瘤血液科6株(13.64%)、重症监护病房5株(11.36%);泌尿系统为最常见感染来源,其次为腹腔引流感染和导管相关感染;近平滑念珠菌、热带念珠菌对氟康唑敏感率均为100%,白念珠菌和光滑念珠菌分别为88.89%和70%,克柔酵母菌和酿酒酵母菌各1株为耐药和敏感。结论本院念珠菌血流感染最常见为白念珠菌,其次为近平滑念珠菌和光滑念珠菌。如果血培养需氧瓶阳性且TTP大于40 h,发生念珠菌血流感染的可能较大,临床或实验室应高度重视。泌尿系感染、导管相关血流感染、手术部位感染是念珠菌血症的重要感染源。大部分念珠菌对氟康唑仍敏感。     

白念珠菌对特比萘芬耐药性的体外梯级诱导和回复研究

目的以浓度梯级倍增的特比萘芬在体外诱导白念珠菌标准株获得耐药子代菌株,并观察其耐药稳定性,从细胞水平研究白念珠菌对特比萘芬耐药前后生物学特性的变化,为进一步采用基因芯片在基因表达水平上研究特比萘芬对白念珠菌的药理作用及其诱导耐药机制提供理想的实验模型。方法将白念珠菌ATCC90028株在特比萘芬浓度逐渐梯级倍增的YPD液体培养基中分别转种传代,直到最后转种至含1024μg／ml特比萘芬的YPD液体培养基中培养,分别测定诱导后形成的各子代菌株的MIC值;选用以1024μg／ml特比萘芬诱导形成的耐药菌株,在不含特比萘芬的YPD液体培养基中连续传代10次后,测定其MIC值,观察其耐药表型的稳定性;并分别用肉眼、光镜和电镜观察白念珠菌耐药性产生前后的形态学特征。结果特比萘芬MIC值为8μg／ml的白念珠菌母本菌株（白念珠菌ATCC90028）成功地被诱导成特比萘芬MIC值为≥512μg/ml的子代菌株,进一步的耐药稳定性实验说明诱导后形成的子代菌株的表型是相对稳定的,诱导后的子代耐药菌株与其母本相比,生长繁殖速度减慢,细胞形态不规则,部分细胞胞膜不完整。结论通过在药物浓度梯级倍增的培养基中连续传代培养的方法可成功建立相同基因型的对特比萘芬敏感的白念珠菌母本和对特比萘芬耐药的子代模型,为获取有亲本的耐药白念珠菌菌株提供了一个有效的实验方法,是在基因水平研究白念珠菌对特比萘芬耐药机制的理想实验模型。     

福建地区94例女性阴道念珠菌感染分离株分型研究

目的 对福建地区临床阴道分泌物分离出的（94株）念珠菌菌株进行分类鉴定,并通过分子生物学及API试验分析传统科玛嘉分类方法的准确性.方法 ①通过科玛嘉显色培养基鉴定菌种,并观察菌株显微镜下形态学表现.②通过ITS区段分子生物学序列分析进行菌种鉴定.③将科玛嘉试验与ITS区段序列分析结果与化验室检验报告结果对比,将鉴定差别菌株进一步行API试验及LSU区段序列分析.结果 ①94株念珠菌经鉴定结果为白念珠菌78株、光滑念珠菌10株、近平滑念珠菌3株、热带念珠菌1株、酿酒假丝酵母菌1株及其中1株为光滑念珠菌及近平滑念珠菌混合感染.②科玛嘉试验可良好的鉴定念珠菌,但对于少见菌种（如酿酒假丝酵母菌）仍缺乏特异性.③一般化验室通过简单菌落形态学及简易科玛嘉检测鉴定仍存在一定误差率（10/94）.④分子生物学方法鉴定菌种准确性高,且可从基因序列分析中鉴定包含C.parapsilosis sensu strico,Candida metapsilosis以及Candida orthopsilosis的近平滑念珠菌复合体菌种.结论 福建地区女性外阴感染的菌种仍以白念珠菌为主,但非白念珠菌的感染也占据相当的比例（16/94）,而在检测方法上,分子生物学技术较科玛嘉试验更能准确的鉴定念珠菌菌种.     

727株真菌失发布与药敏分析

目的 对727株真菌的分布及药敏试验进行分析。方法 用沙保罗培养基分离培养真菌,用API20CAUX生化鉴定及SATBFUNGUS药敏板。结果 727株真菌中白色念珠菌438株,热带念珠菌155株,光滑念珠菌73株,近平滑念珠菌23株,新型隐球菌15株。5－H氟胞嘧啶、两性毒素B、制霉菌素、咪康唑、益康唑和酮康唑的敏感率分别为92．0％、97．6％、94．6％、56．3％、62．9％和70．4％,结论真菌感染仍然以白色念珠菌为主。两性霉素B、制需菌素和一氧胞密啶敏感率较高,而咪康唑、益康唑和酮康唑均出现不同程度的耐药,且呈上升趋势,这需要临床慎选针对性强且副作用低的药物,减少耐药菌株出现。     

1557株酵母菌的鉴定及其药敏试验分析

目的 研究临床分离的酵母菌种类及其对药物的敏感性。方法 采用API 20C、显色培养基对2004-2005年北京协和医院细菌室分离到的1557株念珠菌进行菌株鉴定,并对2005年收集的酵母菌采用ATB FUNGUS 2对氟康唑,伊曲康唑,两性霉素B,5-氟胞嘧啶进行药物敏感性试验;并以whonet 5．3软件进行结果分析。结果 在1557株酵母菌中自念珠菌为57．5％,热带念珠菌为17．0％,光滑念珠菌为14．8％,克柔念珠菌为2．8％,近平滑念珠菌为2．3％,葡萄牙念珠菌为0．9％,新生隐球菌为0．7％,季也蒙念珠菌0．4％,其他菌株共为3．6％。药敏结果显示：近平滑念珠菌、白念珠菌、热带念珠菌、光滑念珠菌、克柔念珠菌和其他念珠菌对氟康唑的敏感率分别为100％、97．6％、94．5％、81．1％、9．1％和75．0％;对5-氟胞嘧啶的敏感率分别为100％、98．0％、98．0％、99．0％、9．5％和88．0％。伊曲康唑对克柔念珠菌和未鉴定到种的念珠菌的MIC90为1μg／ml,光滑念珠菌为4μg／ml,热带念珠菌为0．5μg／ml,白念珠菌和近平滑念珠菌均为0．125μg／ml;两性霉素B对所有念珠菌的MIC50和MIC90均为0．5μg／ml。结论 白念珠菌比例最高,其次为热带念珠菌及光滑念珠菌。5-氟胞嘧啶和氟康唑对除克柔念珠菌之外的所有念珠菌敏感度都很高,两性霉素B对所有念珠菌的MIC90都很低,伊曲康唑对除克柔念珠菌和光滑念珠菌之外的其他菌株的MIC90较低。     

Wall mannoproteins in cells from colonial phenotypic variants of Candida albicans

Candida albicans ATCC 26555 switched at high frequency (10(-1) to 10(-3)) between several phenotypes identified by colony morphology on a defined mineral amino-acid-containing agar medium supplemented with arginine and zinc (LAZ medium). When cells taken from colonies exhibiting distinct morphologies were plated directly onto LAZ agar, spontaneous conversion to all the variant phenotypes occurred at combined frequencies of 2.1 x 10(-1) to 9.5 x 10(-3). However, when cells taken from the different colonial phenotypes were plated directly onto an undefined medium (yeast extract/peptone/dextrose; YPD medium), or first incubated in liquid YPD medium and then cloned on YPD agar, all colonies observed exhibited the same phenotype (smooth-white). When cells from the smooth-white colonies were plated as clones on LAZ agar, the original switch phenotype reappeared. These results suggest that environmental conditions such as the growth medium (and possibly the temperature) influence switching by suppressing phenotype expression, but have no effect on genotype. The variant colony morphologies also appeared to be associated with differences in the relative proportions of yeast and mycelial cells. Zymolyase digests of wall preparations obtained from cells belonging to different colonial phenotypes were analysed by SDS-PAGE. After blotting to nitrocellulose paper, the mannoproteins were stained with Concanavalin A, with a polyclonal antiserum enriched in antibodies against mycelium-specific wall components, and with a monoclonal antibody raised against a high-molecular-mass mannoprotein band (260 kDa) specific to the walls of mycelial cells. The results suggest that phenotypic switching might be associated with changes in the degree of glycosylation in high-molecular-mass mannoproteins, or in the way these mannoproteins are bound to other cell wall components.     

Effect of phenotypic switching on the biological properties and susceptibility to chlorhexidine in Candida krusei ATCC 14243

Phenotypic switching is characterized as a virulence factor of Candida spp. This study was carried out to evaluate the phenotypic switching ability of C.?krusei ATCC 14243 and to determine its effect on the biological properties, adherence capacity and susceptibility towards chlorhexidine digluconate (CHX). To induce switched generations C.?krusei was cultured under nitrogen-depleted growth conditions by adding phloxine B. These phenotypically switched colonies were designated as the 1st generation. Subsequent sub-culturing was performed to produce the 2nd, 3rd and 4th switched generations. The recovery of the 3rd generation was the highest at 85.7% while that of the 4th generation was lower at 70.8%, and the recovery of the 1st and 2nd generations gradually reduced to 46.6% and 36.4%, respectively. All generations of C.?krusei were susceptible towards CHX. The unswitched C.?krusei was the most susceptible but the least adherent to coated hard surfaces. The 2nd generation was the least susceptible, but with the highest adherent ability. The minimum inhibition concentration and minimal fungicidal concentration of C.?krusei of all generations were determined at 0.4?mg?mL(-1) . These observations suggest that the switching activity of C.?krusei induces changes to its biological properties and susceptibility towards CHX.     

Ploidy evolution in the yeast Saccharomyces cerevisiae: a test of the nutrient limitation hypothesis

The nutrient limitation hypothesis provides a nongenetic explanation for the evolution of life cycles that retain both haploid and diploid phases: differences in nutrient requirements and uptake allow haploids to override the potential genetic advantages provided by diploidy under certain nutrient limiting conditions. The relative fitness of an isogenic series of haploid, diploid and tetraploid yeast cells (Saccharomyces cerevisiae), which were also equivalent at the mating type locus, was measured. Fitness was measured both by growth rate against a common competitor and by intrinsic growth rate in isolated cultures, under four environmental conditions: (1) rich medium (YPD) at the preferred growth temperature (30 °C); (2) nutrient poor medium (MM) at 30 °C; (3) YPD at a nonpreferred temperature (37 °C); and (4) MM at 37 °C. In contrast to the predictions of the nutrient limitation hypothesis, haploids grew significantly faster than diploids under nutrient rich conditions, but there were no apparent differences between them when fitness was determined by relative competitive ability. In addition, temperature affected the relative growth of haploids and diploids, with haploids growing proportionately faster at higher temperatures. Tetraploids performed very poorly under all conditions compared. Cell geometric parameters were not consistent predictors of fitness under the conditions measured.     

Characterization of Switch Phenotypes in <Emphasis Type="Italic">Candida albicans</Emphasis> Biofilms

The aim of this study was to characterize switch phenotypes in Candida albicans biofilms. Cells of Candida albicans 192887g biofilms (24 h) were resuspended and these together with their planktonic counterparts were separately inoculated on Lee’s medium agar supplemented with arginine and zinc, at 25 °C for 9 days, for colony formation. The different switch phenotypes, as reflected by varying colony morphologies, were then examined for their (i) stability under various growth conditions, (ii) carbohydrate assimilation profiles, (iii) susceptibility to the polyene antifungal, nystatin, (iv) adhering and biofilm-forming ability, (v) filamentation, and (vi) growth rate in yeast nitrogen base medium supplemented with 100 mM glucose. Our data showed that the frequency of phenotypic switching in C. albicans biofilms was approximately 1%. Compared with the planktonic yeasts, cells derived from candidal biofilms generated one of the phenotypes less frequently (Chi-square-tests: P = 0.017). The five phenotypes derived from the biofilm growth demonstrated differing profiles for carbohydrate assimilation, adhesion, biofilm formation, filamentation, and growth rate. These findings reported here, for the first time, imply that phenotypic switching in the candidal biofilms differs from that in the planktonic growth, and affects multiple biological attributes.     

江西铅山红芽芋胚性愈伤组织的包埋玻璃化超低温保存

为长期安全保存江西铅山红芽芋种质资源,本文以江西铅山红芽芋的胚性愈伤组织为对象,研究了包埋玻璃化冻存过程中各因素对细胞活力和愈伤组织成活率的影响,优化建立了江西铅山红芽芋胚性愈伤组织包埋玻璃化超低温保存体系。将约0.2 g胚性愈伤组织块包埋成海藻酸钙凝胶珠后,在25℃下转入MS+2 mg/L TDZ+1 mg/L NAA+0.75 mol/L蔗糖的培养基中于14 h/d光周期下预培养1 d;预培养后的胚性愈伤组织块用2 mol/L甘油和0.4 mol/L蔗糖的混合物在25℃下装载40 min;采用PVS2在25℃下脱水30 min,更换PVS2后直接投入液氮保存1 d;再将胚性愈伤组织块置于37℃恒温水浴中化冻3 min,然后用MS+2 mg/L TDZ+1 mg/L NAA+1.2 mol/L蔗糖的液体培养基洗涤3次,每次10 min;洗涤后的胚性愈伤组块转入MS+2 mg/L TDZ+1mg/L NAA固体培养基上先暗培养7 d再转到14 h/d光周期中培养。7 d后胚性愈伤组织块开始恢复生长,并且在30 d内分化出胚状体;将胚状体再次转入MS+2 mg/L TDZ+1 mg/L NAA固体培养基上,60 d后形成完整的植株。红芽芋胚性愈伤组织包埋玻璃化超低温保存后的平均成活率约为60%,并且红芽芋胚性愈伤组织冻后再生苗没有发生形态性状和染色体数目的变异,此结果为长期安全保存江西铅山红芽芋种质资源奠定了良好的基础。     

影响枯草芽胞杆菌和荧光假单胞菌原生质体再生的因素

目的：为了提高再生率,对影响革兰阳性菌枯草芽胞杆菌KR株和革兰阴性菌荧光假单胞菌B13株原生质体再生的因素进行研究。方法：研究了酶解时间,再生方式,再生培养基中稳定剂的种类,Ca^2＋、Mg^2＋、琥珀酸钠、L-色氨酸的浓度及培养基的放置时间对KR和B13株原生质体再生的影响。结果：对KR株酶解20min,采用夹层培养,再生培养基中加入0.6mol/L蔗糖、0.03mol/L Ca^2＋、0.02mol/L Mg^2＋、0.3mol/L琥珀酸钠、0.2mol/L L-色氨酸,培养基在37℃放置72h,原生质体再生率可达42.7%;对B13酶解15min,采用夹层培养,培养基中加入0.6mol/L NaCl、0.02mol/L Ca^2＋、0.01mol/L Mg^2＋、0.3mol/L琥珀酸钠、0.1mol/L L-色氨酸,培养基在37℃放置48h,原生质体再生率可达15.3%。结论：影响革兰阳性菌枯草芽胞杆菌KR株和革兰阴性菌荧光假单胞菌B13株原生质体再生的因素是不同的。     

Comparative evaluation of spinosad and phloxine B as toxicants in protein baits for suppression of three fruit fly (Diptera: Tephritidae) species

Spinosad and phloxine B are two more environmentally friendly alternative toxicants to malathion for use in bait sprays for tephritid fruit fly suppression or eradication programs. Laboratory tests were conducted to assess the relative toxicity of these two toxicants for melon fly, Bactrocera cucurbitae Coquillett; oriental fruit fly, Bactrocera dorsalis Hendel; and Mediterranean fruit fly, Ceratitis capitata (Wiedemann) females. Field tests also were conducted with all three species to compare these toxicants outdoors under higher light and temperature conditions. In laboratory tests, spinosad was effective at much lower concentrations with LC50 values at 5 h of 9.16, 9.03, and 4.30 compared with 250.0, 562.1, and 658.9 for phloxine B (27, 62, and 153 times higher) for these three species, respectively. At 16 ppm spinosad, LT50 values were lower for all three species (significantly lower for C. capitata and B. dorsalis) than 630 ppm phloxine B LT50 values. At 6.3 ppm spinosad, the LT50 value for C. capitata (3.94) was still significantly less than the 630 ppm phloxine B LT50 value (6.33). For all species, the 100 ppm spinosad concentrations gave LT50 values of < 2 h. In comparison among species, C. capitata was significantly more sensitive to spinosad than were B. cucurbitae or B. dorsalis, whereas B. cucurbitae was significantly more sensitive to phloxine B than were C. capitata or B. dorsalis. LC50 values were reduced for both toxicants in outdoor tests, with greater reductions for phloxine B than for spinosad for B. dorsalis and B. cucurbitae. Fly behavior, though, is likely to keep flies from being exposed to maximum possible outdoor light intensities. Comparable levels of population suppression for any of the three species tested here will require a much higher concentration of phloxine B than spinosad in the bait.     

Standard YPD, even supplemented with extra nutrients, does not always compensate growth defects of Saccharomyces cerevisiae auxotrophic strains

Conventional complex media are routinely used to grow auxotrophic strains under the assumption that they can compensate the latter's nutritional deficiencies. We here demonstrate that this is not always true. This study compares the growth parameters of Saccharomyces cerevisiae (S288C) and its derived auxotrophic strains FY1679-14C and BY4741 in synthetic minimal medium (SD), standard YPD medium from two of the most commonly used suppliers, or modified YPD medium. Maximum specific growth rates of auxotrophic strains were slightly lower than the prototrophic case in all growth conditions tested. Also, the biomass production of auxotrophic strains in synthetic medium was slightly less than the prototrophic case. However in both of the two standard YPD media used, the biomass production of both auxotrophic strains was markedly lower than that of the prototrophic one. The extent of the differences depended on the medium used. Indeed in one of the two YPD media, the lower biomass production of auxotrophic strains was evident even at the diauxic shift. Uracil seems to be the main limiting growth factor for both auxotrophic strains growing in the two standard YPD medium tested. No YPD media or specific supplement was able to compensate for the effect of the auxotrophic mutations in the multiple auxotrophic marker strain BY4741. The fact that auxotrophic strains grew poorly on YPD when compared to their prototrophic counterpart indicates that standard YPD medium is not sufficient to overcome the effect of auxotrophic mutations.     

Toxicity of Xanthene Dyes to Entomopathogenic Fungi

Effects of xanthene dyes on mycelial growth and conidial germination in three species of entomopathogenic fungi, Beauveria bassiana, Metarhizium anispoliae and Paecilomyces fumosoroseus, were evaluated in a variety of assay systems. In a disk-diffusion assay, erythrosin B and phloxine B (but not eosin B) produced zones of inhibition in colonies of all three species under continuous exposure to light at disk-loadings of 100mug. None of the dyes produced zones of inhibition in the absence of light at disk loadings of 100mug. Both erythrosin B and phloxine B inhibited mycelial growth of all three species in the light in a dose-dependent manner. Weaker dose-responses for inhibition of growth in the dark were observed for some fungus/dye combinations. Erythrosin B, tested singly, completely inhibited conidial germination in the light in all eight fungal strains tested at 100mug ml-1 medium, but failed to inhibit conidial germination in any of these strains in the dark at the same concentration of dye. For single strains of each of the three fungi, erythrosin B and phloxine B inhibited conidial germination in a dose-dependent manner in the light with IC50s < 6.2mug dye ml-1 medium for all fungus/dye combinations. Phloxine B was a more potent inhibitor of germination than erythrosin B for all three fungal species. At fixed dosages of erythrosin B and phloxine B, inhibition of conidial germination in all three species increased with time of exposure to light. These results constitute the first quantitative demonstration of photodynamic inhibition of conidial germination in fungi by xanthene dyes.