Efficiency of entomopathogenic nematodes and chemical insecticides against Sphenophorus levis and Leucothyreus sp. on sugarcane

The sugarcane billbug, Sphenophorus levis (Coleoptera: Dryophthoridae), is an important pest of sugarcane crops in the São Paulo State, Brazil. Their larvae damage plants rhizome, causing losses of up to 30 t ha-1 year-1 of sugarcane. In order to evaluate the effectiveness of entomopathogenic nematodes and chemical insecticides against the sugarcane billbug and the scarab beetle Leucothyreus sp., three field trials were carried out in Brazil. For the first one, 10 treatments were considered: 1) thiametoxan (Actara 250WG) 500 g ha-1 of commercial product (c.p.); 2) thiametoxan 1.0 kg ha-1 c.p.; 3) thiametoxan 1.5 kg ha-1 c.p.; 4) Steinernema brazilense 108 infective juveniles (IJ) ha-1 + thiametoxan 250 g ha-1 c.p.; 5) S. brazilense (108 IJ ha-1) + thiametoxan 500 g ha-1 c.p.; 6) Heterorhabditis indica (108 IJ ha-1) + thiametoxan 250 g ha-1 c.p; 7) H. indica (108 IJ ha-1) + thiametoxan 500 g ha-1 c.p.; 8) S. brazilense (108 IJ ha-1); 9) H. indica (108 IJ ha-1); and 10) Control. The second one consisted of five treatments: 1) thiametoxan (Actara 250WG) 800 g ha-1 c.p.; 2) fipronil (Regente 800WG) 250 g ha-1 c.p.; 3) S. brazilense (108 IJ ha-1); 4) S. brazilense (108 IJ ha-1) + thiametoxan 200 g ha-1 c.p.; and 5) Control. In the third one, all treatments already mentioned for the second trial plus the mixture S. brazilense (108 IJ ha-1) + fipronil (Regente 800WG) 62.5 g ha-1 c.p. were considered. All treatments with the insecticides and nematodes provided higher sugarcane yield, when compared to their respective controls, which were most damaged by S. levis, but without statistical significance. S. brazilense was more efficient on controlling this pest, if compared to H. indica, but without statistical significance. S. brazilense provided a 50% control of Leucothyreus sp. larvae, meanwhile the best treatments against this scarab were the mixtures of this nematode with fipronil (78%) and thiametoxan (83%)

Occurrence, production and dry mycelium preservation of Batkoa sp. and Furia sp., pathogens of spittlebugs pests of pastures.

Os fungos Batkoa sp. e Furia sp.(Entomophthorales), embora ocasionalmente ocorrendo de forma epizoótica em populações das cigarrinhas-das-pastagens e cana-deaçúcar, nunca foram avaliados quanto ao nível de incidência e potencial de controle biológico. Esse estudo teve por objetivo avaliar a ocorrência natural, produção "in vitro" e preservação de micélio seco desses fungos. Nos trabalhos de produção incluiu-se como padrão o fungo Neozygites floridana, representante de outra família de Entomophthorales. O estudo sobre a ocorrência natural foi realizado na região de Pindamonhangaba, SP, no período de 22 de janeiro a 9 de fevereiro de 1998 com o objetivo de avaliar a incidência de Batkoa sp. na população de Mahanarva fimbriolata em pastagem de capim napier e de Furia sp. sobre Deois schach em pastagem de Brachiaria sp. O estudo sobre produção foi realizado em condições de laboratório com o objetivo de desenvolver meios líquidos para a produção de micélio ou corpos hifais de Batkoa sp., Furia sp. e N. floridana, tendo como base o meio líquido para cultura de tecido de inseto Graces suplementado, o qual permite bom crescimento do último patógeno. Finalmente, estudou-se o efeito da combinação de dois dessecantes (sílica e glicerol) com dois redutores de oxigênio (vácuo e Agelessâ ZPT-200) na preservação de micélio seco de Batkoa sp. e Furia sp. sob temperaturas de 3°C e 23°C. Concluiu-se que: Batkoa sp. e Furia sp. causam doença na população de M. fimbriolata e D. schach, respectivamente; Furia sp. possui maior potencial epizoótico que Batkoa sp.; Batkoa sp., Furia sp. e N. floridana se assemelham quanto ao crescimento em meios adicionados de sais, vitaminas e aminoácido, e em meio completo; A adição de sais ao meio básico proporciona um aumento significativo na produção das três espécies de fungos, e maior do que a adição de vitaminas e aminoácidos; A mistura dos sais CaCl2.2H2O (0,06%), KCl (0,28%), MgCl2.6H2O (0,16%), MgSO4.7H2O (0,2%), NaHCO3 (0,03%) e NaH2PO4.H2O (0,1%) proporciona maior produção dos fungos do que os sais avaliados isoladamente na concentração de 0,83%; O sulfato de magnésio (MgSO4) pode ser utilizado como único sal, na concentração de 0,83%, visando a produção de Batkoa sp., Furia sp. e N. floridana; Esses fungos se assemelham quanto ao crescimento em meios contendo diferentes fontes de carbono, mas se diferenciam em meios com diferentes fontes de nitrogênio; O meio contendo 2,66% de glucose proporciona uma produção das três espécies de fungos significativamente maior do que o meio com 2,66% de sacarose; A adição de 0,1% de monossacarídeos no meio com 2,66% de sacarose não aumenta significativamente a produção dos fungos; O extrato de levedura proporciona maior produção de Batkoa sp.; A concentração de 0,5% de extrato de levedura é a mais adequada para a produção vegetativa de Batkoa sp.; A combinação de extrato de levedura + extrato de carne + leite desnatado proporciona a maior produção de Furia sp.; A combinação de extrato de levedura + leite desnatado proporciona o segundo maior rendimento de Furia sp., sendo mais adequada para a produção do fungo devido ao menor custo; A concentração de 1% da mistura dessas fontes de nitrogênio é adequada para a produção vegetativa de Furia sp.; A combinação de extrato de levedura + peptona + leite desnatado proporciona a maior produção de N. floridana; A concentração de 3% da mistura dessas fontes de nitrogênio é adequada para a produção vegetativa de N. floridana; A combinação de sílica + Ageless® prolonga a sobrevivência desses dois fungos formulados como micélio seco até 90 dias, armazenados a 3°C e 23°C; O uso de vácuo como redutor de oxigênio não prolonga a sobrevivência desses dois fungos, mesmo em associação com dessecantes; A temperatura de 3°C provoca uma ligeira queda no potencial de crescimento de Batkoa sp., porém prolonga a sobrevivência do fungo até 90 dias, independentemente do tratamento.The fungi Batkoa sp. and Furia sp. although occasionally occurring at epizootic levels in populations of spittlebug pests of pasture and sugarcane, have never been evaluated as to natural incidence level nor biological control potential. This study was designed to evaluate the natural occurrence, in vitro production and dry mycelium preservation of these fungi. Neozygites floridana, a representative of another family of Entomophthorales, was included as a standard in the production studies. The study on natural occurrence was done in Pindamonhangaba county, São Paulo State, from January 22 to February 9, 1998, to evaluate the incidence of Batkoa sp. in the Mahanarva fimbriolata population on pasture of napier grass, and Furia sp. in Deois schach, on Brachiaria sp. pasture. The study on production was done in laboratory conditions with the purpose to develop liquid media for the production of mycelium or hyphal bodies of Batkoa sp. Furia sp. and N. floridana, using as a base a liquid medium for insect tissue culture [Graces supplemented (HyQ® TNM-FH, #SH30280.02)], which allowed good growth of the latter pathogen. Finally, the effect of combinations of two desiccants (silica and glycerol) with two oxygen reducers (vacuum and Ageless® ZPT-200) on the preservation of dry mycelium of Batkoa sp. and Furia sp. under temperatures of 3º C and 23º C was studied. It is concluded that: Batkoa sp. and Furia sp. cause disease in M. fimbriolata and D. schach sp. populations, respectively. Furia sp. has higher epizootic potential than Batkoa sp. Batkoa sp., Furia sp. and N. floridana are similar concerning their growth patterns in media with added salts, vitamins and amino acids, as well as in the complete one. The addition of salts to the basic medium causes a significant increase in biomass production of the three fungal species, and higher than that from the addition of vitamins and amino acids. The mixture of the salts CaCl2.2H2O (0,06%), KCl (0,28%), MgCl2.6H2O (0,16%), MgSO4.7H2O (0,2%), NaHCO3 (0,03%) and NaH2PO4.H2O (0,1%) afforded higher production of the three fungi than the salts solely evaluated, at the concentration of 0,83%. MgSO4 can be used as sole salt for the production of Batkoa sp., Furia sp. and N. floridana. These fungi are similar concerning their growth patterns in media with different sources of carbone, but different in media with different sources of nitrogen. The production of the three fungal species is significantly higher in medium containing 2.66% glucose than in 2.66% sucrose. The addition of 0.1% monossacarides to media containing 2.66% sucrose does not increase significantly biomass production. Yeast extract allows the highest production of Batkoa sp. The concentration of 0.5% yeast extract is the most suitable for vegetative growth of Batkoa sp. The combination of yeast extract + beef extract + skim milk allows the highest production of Furia sp. The combination of yeast extract + skim milk allows the second highest production of Furia sp., and is the most suitable for mass production due to the lower cost. The 1% concentration of the mixture of these nitrogen sources is suitable for vegetative growth of Furia sp. The combination of yeast extract + peptone + skim milk allows the highest production of N. floridana. The 3% concentration of the mixture of these nitrogen sources is suitable for the vegetative growth of N. floridana. The combination of silica + Ageless® extends the survival of these two fungi formulated as dry mycelium for 90 days, stored at 3º C and 23º C. The use of vacuum to reduce oxygen does not extend the survival of these fungi, even in association with desiccants. Storage at 3º C causes a slight decrease in growth potential of Batkoa sp., but it extends fungus survival for 90 days, independent of other treatments

Culture media to detect and criteria to evaluate and report the activity of extracellular enzymes produced by phytopathogenic fungi

ABSTRACT: Extracellular enzymes are involved in the fungal pathogenesis in plants. Currently, culture media, data analyses, and data report related to extracellular enzymes produced in vitro conditions are different and therefore, lack standardization. This work aimed to compare the culture media cited on the literature (normal) with the potato-dextrose-agar (PDA) medium combined with a specific compound to produce extracellular enzymes through three soilborne phytopathogenic fungi (F. solani f. sp. passiflorae, S. rolfsii, and R. solani AG-4 HGI), as well as to analyze and report enzyme data based on five different criteria. The assay was randomized, with three factors (culture media, isolates, and enzymes) and six repetitions. The studied enzymes were amylase (AM), carboxymethylcellulase (CMCase), lipase (LP), laccase (LC), catalase (CT), and gelatinase (GT). The normal media detected more enzymes and was more precise compared to the PDA medium plus specific compound. The criteria that calculated the area of the circular crown of AM, CMCase, LP, and LC and measured the intensity (0 = absence, up to 4 = intense) of CT and GT adopting note scale were the best to evaluate and report the results of the enzymes. We suggest the normal media culture to study enzyme production, as well as the criteria mentioned to assess and report the data related to enzyme activities

Colonization of vines by Petri disease fungi, susceptibility of rootstocks to Phaeomoniella chlamydospora and their disinfection

ABSTRACT: Petri disease is complex, attacks young vine plants and it is difficult to be controlled. The fungus Phaeomoniella chlamydospora (Phc) has been identified as the main causative agent of this disease. This study aimed to evaluate the prevalent colonization of the Petri disease fungi in different portions of vine plants; to assess the susceptibility of grapevine rootstocks to the fungus P. chlamydospora; to assess the effect of solarization and biofumigation, followed by hot-water treatment (HWT), on the disinfection of cuttings of the rootstock IAC 766 infected with P. chlamydospora, and assess the effect of solarization and biofumigation, followed by HWT, on the rooting of cuttings of the rootstock IAC 766. For the prevalent colonization test, the fungus species detected and identified in ‘Niagara Rosada’ grafted on two rootstocks different were Phc and Phialemoniopsis ocularis. This is the first report of P. ocularis in a young vineyard in Brazil. Both fungi, in particular Phc, colonized only the plant’s basal part, drawing attention to the rootstock as target for control measures. Measurement of the dark streaks in the vascular system revealed that Golia was the least susceptible rootstock, and IAC 572 was the most susceptible to Phc. Moreover, biofumigation or temperature of 37°C applied for 7 and 14 days, both followed by HWT, suppressed Phc in cuttings of the rootstock IAC 766 without hampering their rooting. Meanwhile, new studies are needed to validate the efficiency of these disinfection techniques

Pathogenicity and Virulence of Different Concentrations of Brazilian Isolates of Entomopathogenic Nematodes Against Drosophila suzukii

The invasive pest Drosophila suzukii (Matsumura) (Diptera: Drosophilidae) was recently recorded in Brazil and constitutes a threat to fruit growing, mainly for small, soft fruits. Recent advances in research on ways of controlling D. suzukii involve the use of entomopathogenic nematodes (EPNs). In this context, the objective of this study was to evaluate the pathogenicity and virulence of four isolates in different concentrations against D. suzukii pupae. The EPN isolates used in trials were Steinernema brazilense IBCBn 06, S. carpocapsae IBCBn 02, Heterorhabditis bacteriophora HB, and H. amazonensis IBCBn 24. Both H. amazonensis IBCBn 24 and H. bacteriophora HB were effective in controlling D. suzukii as they caused a mortality rate of 86.25% and 80.0%, and virulence of 549.75 IJs/pupae and 787.75 IJs/pupae in the concentrations of 1800 IJs/ml and 5400 IJs/ml, respectively. The lowest lethal concentrations (LC50) of juveniles were found in host pupae with 771.63 IJs/ml of H. bacteriophora HB and 1115.49 IJs/ml of H. amazonensis IBCBn 24. Results showed that both EPNs, H. amazonensis IBCBn 24 and H. bacteriophora HB, could be promising eco-friendly biological agents to control D. suzukii.Fil: Dias, Sérgio Costa. Universidade Federal de Pelotas; BrasilFil: de Brida, Andressa Lima. Universidade Federal de Pelotas; BrasilFil: Maguintontz Cedney, Jean-Baptiste. Universidade Federal de Pelotas; BrasilFil: Leite, Luís Garrigós. Instituto Biológico de Campinas; BrasilFil: Ovruski Alderete, Sergio Marcelo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Planta Piloto de Procesos Industriales Microbiológicos; ArgentinaFil: Garcia, Flavio Roberto Mello. Universidade Federal de Pelotas; Brasi

Mortality of Conotrachelus humeropictus in response to combined application of the nematode Steinernema brazilense and the fungus Beauveria bassiana

ABSTRACT: The fruit borer Conotrachelus humeropictus is one of the most important pests of the cupuaçu crop, and can promote yield losses of around 50%. The present study involved the selection of Beauveria bassiana isolates on Conotrachelus psidii larvae, which was used as test insect due to its taxonomic closeness to C. humeropictus. In order to assess their interaction using combined application, we studied the ability of the nematode to carry conidia of B. bassiana and Metarhizium anisopliae to soil layers. This study also aimed to establish a biological control method for cupuaçu borer with the use of the fungus B. bassiana at 10 kg/ha and the entomopathogenic nematode Steinernema brazilense at 1, 3, 9, and 27 IJ/cm2, applied as single and combined treatments. In the selection of B. bassiana strains for C. psidii, the IBCB 276 and IBCB 165 isolates were the most promising ones, causing 86 and 84% mortality, respectively. The IBCB 276 strain was applied in the field to control C. humeropictus. In the study of carriage of conidia in soil, the nematode S. brazilense was found to be capable of carrying conidia of B. bassiana to deep 7 to 10 cm soil layers. This was not observed with M. anisopliae, found only in the topsoil. In field trials against C. humeropictus larvae, B. bassiana promoted 15.6% mortality. The nematode promoted 60.0% mortality at the highest concentration used, and their combined use with B. bassiana (10 kg/ha) increased mortality to 65.6%, with an additive interaction observed between the two entomopathogens

Two methods for isolation of endophytic and edaphic Bacillus spp. from sugarcane fields

ABSTRACT: Bacillus has been widely studied and used for the control of pests and diseases. The adapted protocol proposed by POLANCZYK (2004) proved to be more efficient than the one by the World Health Organization (WHO, 1985) to isolate edaphic strains of Bacillus. However, it has not been assessed for isolation of endophytic strains, which are much less abundant in the nature and more difficult to be isolated. This study aimed to compare two methodological procedures for isolation of Bacillus, established by the WHO (1985) and by POLANCZYK (2004), regarding their efficiency for isolation of endophytics and edaphics Bacillus strains from inside the root tissue of sugarcane, as well as from the associated soil sample, collected from 11 locations; and to compare the density of bacteria in both environments. Endophytic and edaphic strains of Bacillus were isolated by both procedures. However, the isolation protocol performed by POLANCZYK (2004) made more efficient by having a greater number of colony forming units (CFU) per gram of soil and root indicating that this procedure is more useful, especially for isolation of endophytic strains of Bacillus, which are much less abundant in the nature than edaphic strains, being therefore more difficult to be isolated. Using the Polanczyk protocol (2004), Bacillus strains were recovered from all roots (endophytic) and soil (edaphic) samples of all the 11 fields, suggesting that the plant root may be another important source for isolation of Bacillus besides the soil. Higher densities of Bacillus were isolated from the edaphic environment compared with the endophytic environment, with significant differences when isolated by Polanczyk method (2004)