Identification and taxonomy of some entomopathogenic Paecilomyces spp. ( Ascomycota ) isolates using rDNA-ITS Sequences

A phylogenetic analysis of the 5.8S rDNA and internal transcribed spacer (ITS1 and ITS2) sequences from some entomogenous Paecilomyces species supports the polyphyly of the genus and showed the existence of cryptic species. In the Eurotiales, anamorphs Paecilomyces variotii and Paecilomyces leycettanus were related to the teleomorphs Talaromyces and Thermoascus. In the Hypocreales, three major ITS subgroups were found, one of which included Paecilomyces viridis, Paecilomyces penicillatus, Paecilomyces carneus and isolates identified as Paecilomyces lilacinus and Paecilomyces marquandii. However, the majority of the P. lilacinus and P. marquandii isolates formed a distinct and distantly related subgroup, while the other major subgroup contained Paecilomyces farinosus, Paecilomyces amoeneroseus, Paecilomyces fumosoroseus and Paecilomyces tenuipes.

Genetic diversity of Hirsutella thompsonii isolates from Thailand based on AFLP analysis and partial beta-tubulin gene sequences

Amplified fragment length polymorphism (AFLP) was used to investigate polymorphism among 43 Hirsutella thompsonii isolates (33 from Thailand) obtained from various mite species. The outgroups were an unidentified Hirsutella isolate along with Hirsutella nodulosa and Hirsutella kirchneri. Phylogenetic analyses of the AFLP data showed significant variation among isolates and the existence of three H. thompsonii clades. We also investigated the isolates using PCR with specific primers for the Hirsutella exotoxin gene Hirsutelin A (HtA) and 18 of these isolates were used for sequencing of the partial beta-tubulin gene. Phylogenetic analyses of beta-tubulin sequences showed two distinct H. thompsonii clades, one of which included AFLP clades I and II. For both markers grouping of the H. thompsonii isolates was not related to either host mite species or geographical origin, although for the HtA gene one clade contained only isolates with no detectable HtA band. These results confirm the high intraspecific polymorphism of H. thompsonii, and maximum likelihood analysis showed no monophyletic group within this species. To refine the taxonomy of this genus other studies should be undertaken using additional molecular markers and several other Hirsutella isolates.

Activity of oil-formulated Beauveria bassiana against Triatoma sordida in peridomestic areas in central Brazil

Field tests were carried out during the rainy season of 2001/2002 in São Luís de Montes Belos, Goiás, Brazil, to evaluate the potential of the entomopathogenic fungus, Beauveria bassiana, against peridomestic Triatoma sordida. An oil-water formulation of the isolate CG 14 (Embrapa) was applied in triatomine infested hen houses of four farms at a final concentration of 10(6) conidia/cm². Numbers of T. sordida decreased over the next 25 days, after application of the fungus, and B. bassiana developed on dead insects in one hen house. A high number of B. bassiana colonies was detected in substrates collected in treated hen houses 24 h after application of CG 14. In the following three months the presenceof B. bassiana declined to values found before treatment.

Cinética da esporulação e viabilidade de conídios de Nomuraea rileyi (Farlow) samson sobre cadáveres da lagarta-da-soja, Anticarsia gemmatalis Hübner (Lepidoptera: Noctuidae), em condições de campo / Sporulation kinetics and conidial viability of Nomuraea rileyi (Farlow) samson on cadavers of the velvetbean caterpillar, Anticarsia gemmatalis Hübner (Lep: Noctuidae) in the field

The entomopathogenic fungus Nomuraea rileyi (Farlow) Samson produces epizootics on populations of the velvetbean caterpillar, Anticarsia gemmatalis Hübner, resulting in its natural control. However, sometimes this phenomenon does not occur soon enough to prevent pest population outbreaks and damage to the crop. Observations of N. rileyi conidial production on the surface of A. gemmatalis cadavers in the field showed that high relative humidity and precipitation determine the sporulation dynamics. In fact, initiation of sporulation is delayed and conidia production is reduced when A. gemmatalis cadavers infected with N. rileyi are exposed to water stress (relative humidity 25-30%) in the laboratory. Field observations also demonstrated that N. rileyi conidial viability remain high (>75% germination) until 10 days on A. gemmatalis cadaver surface, acting as an inoculum source for this period.

O fungo entomopatogênico Nomuraea rileyi (Farlow) Samson pode produzir epizootias em populações de lagarta-da-soja, Anticarsia gemmatalis Hübner, controlando essa praga naturalmente. No entanto, o fenômeno nem sempre ocorre a tempo de evitar que populações de A. gemmatalis atinjam o nível de dano econômico na cultura. Observações sobre a produção de conídios em cadáveres de A. gemmatalis em condições de campo mostraram que alta umidade relativa e precipitação são fatores determinantes na cinética da esporulação do fungo in vivo. O início da esporulação foi retardado e a produção de conídios diminuída quando cadáveres de A. gemmatalis infectados com N. rileyi foram submetidos a períodos longos de estresse hídrico (umidade relativa de 25-30%). Observações no campo também demonstraram que a viabilidade de conídios de N. rileyi pode permanecer elevada (>75% germinação) por períodos de até 10 dias na superfície de cadáveres, servindo como fonte de inóculo durante esse período.

Transformation of the entomopathogenic fungi, paecilomyces fumosoroseus and paecilomyces lilacinus (deuteromycotina: hyphomycetes) to benomyl resistence

The entomopathogenic fungi Paecilomyces fumosoroseus and P. lilacinus have been transformed to resistance to the fungicide benomyl by a polyethylene glycol (PEG)-mediated procedure using a mutant b-tubulin gene from Neurospora crassa carried on plasmid pBT6. Benomyl-resistant transformants of P. lilacinus were obtained that could tolerate greater than 30 µg/ml benomyl and P. fumosoroseus transformants were obtained that could tolerate 20 µg/ml benomyl. Following 5 serial passages of transformants on benomyl-containing media and 5 serial passages on non-selective media, 100 per cent of P. lilacinus transformants were found to be mitotically stable by a conidial germination test. In contrast, only 4 out of 9 transformants of P. fumosoroseus were mitotically stable. Southern blot analysis of genomic DNA from both species suggested that the mechanism of transformation in all transformants was by gene replacement of the b-tubulin allele. Non-homologous vector sequences were not detectable in the genomes of transformants.

Selection of Beauveria bassiana and Metarhizium anisopliae isolates to control Triatoma infestans

Twenty three isolates of Beauveria bassiana and 13 isolates of Metarhizium anisopliae were tested on third instar nymphs of Triatoma infestans, a serious vector of Chagas disease. Pathogenicity tests at saturated humidity showed that this insect is very susceptible to fungal infection. At lower relative humidity (50 per cent), conditions expected in the vector microhabitat, virulence was significantly different among isolates. Cumulative mortality 15 days after treatment varied from 17.5 to 97.5 per cent and estimates of 50 per cent survival time varied from 6 to 11 days. Maintaining lower relative humidity, four B. bassiana and two M. anisopliae isolates were selected for analysis of virulence at different conidial concentrations and temperatures. Lethal concentration sufficient to kill 50 per cent of insects (LC50) varied from 7.1x10 5 to 4.3x10 6 conidia/ml, for a B. bassiana isolate (CG 14) and a M. anisopliae isolate (CG 491) respectively. Most isolates, particularly B. bassiana isolates CG 24 and CG 306, proved to be more virulent at 25 and 30ºC, compared to 15 and 20ºC. The differential virulence at 50 per cent humidity observed among some B. bassiana isolates was not correlated to phenetic groups in cluster analysis of RAPD markers. In fact, the B. bassiana isolates analyzed presented a high homogeneity (>73 per cent similarity).