Identification of entomopathogenic Bacillus isolated from Simulium (Diptera, Simuliidae) larvae and adults

Abstract

Entomopathogenic bacteria isolated fromSimuliids are hematophagus insects naturally occurring in waterways throughout Brazil. Although some species have been involved in the transmission of human onchocerciasis in other regions (Gerais & Ribeiro 1986, Maia-Herzog et al. 1999) the main importance of these insects in the southeast of Brazil is socio-economic. The frequent and intensive attacks by simuliids on the transitory population in this area can reduce tourism in summer months, a fact that could negatively impact the economies of affected municipalities. To decrease the presence of these insects in these areas, a full-scale programme, based on Bacillus thuringiensis serovar israelensis, has been carried out in the north littoral zone of the State of São Paulo since 1990 (Araújo-Coutinho 1995).Recent findings of B. thuringiensis serovar oswaldocruzi and B. thuringiensis serovar braziliensis from unusual habitats (Rabinovitch et al. 1995) encourage the continued search for other endemic entomopathogens. For that purpose, Simulium larvae and adults from the north littoral zone of São Paulo and some other rivers in the State of Rio de Janeiro were evaluated for the presence of entomopathogenic bacteria. MATERIALS AND METHODSCollection of simuliids -Black fly larvae and adults were collected from breeding sites in Pau d'Alho river, State of São Paulo. This river was divided into two regions, one downstream of B. thuringiensis serovar israelensis applications and another, upstream. Another collection point was the Soberbo river, Guapimirim, State of Rio de Janeiro (Fonseca et al. 1998). This river has never been exposed to commercial formulations based on B. thuringiensis serovar israelensis. Only larvae were collected in this waterway. The third point where larvae were collected was in Rio das Pedras, Mangaratiba, State of Rio de Janeiro.Isolation of Bacillus -To eliminate external contamination, the insects were sterilized following the methodology described by Alves (1986) with a slight modification; "Superbonder" glue (cyanoacrilate ester) was used to close both oral and anal cavities. This modification was adopted because Simulium larvae are very small and the dental floss used in the original technique was not applicable. After this step, the original method was performed passing the larvae through three solutions, first in 70% alcohol for 2 sec, second in 5% sodium hypoclorite for 3 min and finally in sterile 10% sodium thiossulfate for 5 min. The specimens were then washed three times in sterile distilled water.Two different methods were used to isolate B. thuringiensis samples. In both, Simulium larvae were transferred aseptically into a sterile mortar and macerated with a sterile pestle. In the first method, the macerate was placed in tubes containing distilled water (10 ml). The suspension was heated (65ºC) for 12 min, and immediately diluted (1x10 -2

Bacillus thuringiensis isolates entomopathogenic for Culex quinquefasciatus (Diptera: Culicidae) and Anticarsia gemmatalis (Lepidoptera: Noctuidae)

“…Some of our new B. thuringiensis isolates from both soil and insects produced high mean percentage mortality in C. quinquefasciatus, A. gemmatalis or both, confirming the reports of various authors that both soil and insects are good sources for new entomopathogenic B. thuringiensis strains (Hansen et al, 1997;Schnepf et al, 1998;Kim, 2000;Cavados et al, 2001;Quesada-Moraga et al, 2004;Hajaij et al, 2005;Hernandez et al, 2005).…”

“…The relationship between the protein profile of isolates ID-MD1 and ID-MD4 and their toxicity to C. quinquefasciatus may have been due to the 27 kDa band in the protein profile of these isolates (Figure 1), since the Cyt protein, known to trigger cytolytic action against diptera, also resolves at 27 kDa Tokcaer et al, 2006). Isolate SR-VV2 more virulent for C. quinquefasciatus than A. gemmatalis, displayed a band at approximately 100 kDa (Figure 1b), similar results have been reported for two isolates toxic against A. aegypti (Cavados et al, 2001). The isolates showing…”

Histopathological and ultrastructural effects of delta-endotoxins of Bacillus thuringiensis serovar israelensis in the midgut of Simulium pertinax larvae (Diptera, Simuliidae)

“…Simulium pertinax is the most important species in Southeast region inducing a negative socio-economic impact. The frequent and intensive attacks by S. pertinax on the transitory populations in this area helps to reduce tourism during summer months (Gerais & Ribeiro 1986, Araújo-Coutinho 1995, MaiaHerzog et al 1999, Cavados et al 2001, Araújo-Coutinho et al 2003.…”

“…After isolation of the B. thuringiensis serovar israelensis (Bti) by Goldberg and Margalit (1977) and its characterization by De Barjac (1978), several studies were realized confirming the action of Bti as a larvicide mainly for controlling Simulium and Aedes species (De Barjac 1978, Lacey et al 1982, Charles & De Barjac 1983, Becker 1990, Araújo-Coutinho 1995, Mardini et al 1999, Cavados et al 2001, Regis et al 2001.…”

Occurrence of Bacillus thuringiensis and their phages in Yemen soil

“…More than 60 cry genes, with toxicity spectra extending over several orders of insects and other pests, have been reported from natural Bt strains [17]. Bt strains have been isolated from diverse habitats such as, soils and aquatic environments [24], plants [28] insects [14], animal faeces [25] and arid environments [10]. The natural occurrence of Bt in the environment and its insecticidal activity has been reported from various countries in Europe [8,36], North America [29,35], South America [9,23,29,35], Asia [13,20,21,29,34,49], and Africa [2,29,33,38].…”