Castellanos-Moguel Judith, González-Barajas Margarita, Mier Teresa, Reyes-Montes María Del Rocío, Aranda Eduardo, Toriello Conchita
Departamento el Hombre y su Ambiente, Universidad Autónoma Metropolitana-Xochimilco, México DF, México.
Rev Iberoam Micol. 2007 Mar;24(1):62-8. doi: 10.1016/s1130-1406(07)70016-5.
To properly characterize several isolates of Paecilomyces fumosoroseus, a fungal entomopathogen of whiteflies (Homoptera: Aleyrodidae) and other insect pests for biocontrol purposes, virulence towards Trialeurodes vaporariorum, and subtilisin-like (Pr1) and trypsin-like (Pr2) protease activity during propagule production were investigated in monospore cultures (MCs). The virulence of three MCs towards second instar whiteflies was measured and expressed as lethal median concentration (LC50). Number and widthlength ratio of propagules (blastospores, hyphal bodies, short hyphae, submerged conidia) and extracellular proteolytic activity was determined simultaneously in liquid medium. Total protease activity was assayed with azocasein, Pr1 and Pr2 activity was determined with the substrates N-Succinyl-Ala-Ala-Pro-Phe-p-nitroanilide and N-Benzoyl-Phe-Val-Arg-pnitroanilide, respectively. Natural variability in virulence, propagule production and cuticle-degrading proteases among isolates was observed. Bioassays showed a LC50 of 1.1 x 1,000, 2.5 x 10,000 and 7.6 x 10,000 conidia/ml for MCs EH-506/3, EH-503/3 and EH-520/3, respectively, EH-506/3 being the most virulent isolate. Isolate EH-503/3 produced the highest yield of propagules (7.7 x 10000000 propagules/ml), followed by EH-520/3 with 6.4 x 10000000 and EH-506/3 with 1.0 x 10000000 propagules/ml. Subtilisin-like (Pr1) and trypsin-like (Pr2) activity was present in the three MCs. Subtilisin-like (Pr1) activity was highest (745.7 UPr1/ml at 120 h) in the most virulent isolate, EH-506/3, pointing at Pr1 as a phenotypic marker of virulence for P. fumosoroseus. EH-506/3 appears to be a good candidate for whitefly biocontrol due to its high virulence, Pr1 concentration and rapid transition to blastospores in submerged liquid medium.
为了准确鉴定玫烟色拟青霉的多个分离株，玫烟色拟青霉是一种用于生物防治目的的粉虱（同翅目：粉虱科）及其他害虫的真菌性昆虫病原，在单孢子培养物（MCs）中研究了其对温室白粉虱的毒力，以及在繁殖体产生过程中类枯草杆菌蛋白酶（Pr1）和类胰蛋白酶（Pr2）的活性。测定了三种MCs对二龄粉虱的毒力，并以致死中浓度（LC50）表示。同时在液体培养基中测定了繁殖体（芽生孢子、菌丝体、短菌丝、浸没分生孢子）的数量和长宽比以及细胞外蛋白水解活性。用偶氮酪蛋白测定总蛋白酶活性，分别用底物N-琥珀酰-丙氨酸-丙氨酸-脯氨酸-苯丙氨酸-对硝基苯胺和N-苯甲酰-苯丙氨酸-缬氨酸-精氨酸-对硝基苯胺测定Pr1和Pr2活性。观察到分离株之间在毒力、繁殖体产生和角质层降解蛋白酶方面存在自然变异。生物测定表明，MCs EH-506/3、EH-503/3和EH-520/3对二龄粉虱的LC50分别为1.1×1000、2.5×10000和7.6×10000个分生孢子/毫升，EH-506/3是毒力最强的分离株。分离株EH-503/3产生的繁殖体产量最高（7.7×10000000个繁殖体/毫升），其次是EH-520/3，为6.4×10000000个，EH-506/3为1.0×10000000个繁殖体/毫升。三种MCs中均存在类枯草杆菌蛋白酶（Pr1）和类胰蛋白酶（Pr2）活性。在毒力最强的分离株EH-506/3中，类枯草杆菌蛋白酶（Pr1）活性最高（120小时时为745.7 UPr1/毫升），表明Pr1是玫烟色拟青霉毒力的表型标记。由于EH-506/3具有高毒力、Pr1浓度高以及在浸没液体培养基中能快速转变为芽生孢子，它似乎是粉虱生物防治的一个良好候选菌株。
