Thirty samples of roasted ground coffee beans from 10 different commercial brands were analyzed to investigate the occurrence and levels of Bacillus cereus and Bacillus thuringiensis strains. Strains were evaluated for their genetic diversity by repetitive element sequence polymorphism PCR (Rep-PCR) and for their toxigenic profiles, i.e., the presence of hblA, hblC, hblD, nheA, nheB, nheC, cytK, ces, and entFM. Survival and multiplication of B. cereus sensu lato in the ready-to-drink coffee was determined to evaluate this beverage as a possible vehicle for B. cereus infection. B. cereus was detected in 17 (56.7%) of the 30 samples, and B. thuringiensis was detected in 8 (26.7%) of the 30 samples. Five samples did not produce any characteristic growth. The most common gene, entFM, was detected in 23 strains (92%). The NHE complex (nheA, nheB, and nheC genes) was found in 19 strains (76%). The HBL complex (hblA, hblC, and hblD) was found in 16 strains (64%). All strains were negative for ces. The cytK gene was found in 16 strains (64%). The computer-assisted cluster analysis of Rep-PCR profiles using a clustering criterion of 80% similarity revealed four main clusters. Cluster 1 was the predominant and comprised three B. thuringiensis strains with 100% similarity, cluster 2 comprised two B. cereus strains (100% similarity), cluster 3 comprised two B. thuringiensis strains (90% similarity), and cluster 4 comprised one B. thuringiensis strain and one B. cereus strain (85% similarity). The cluster analysis of fingerprints generated by Rep-PCR revealed a high genetic diversity among the B. cereus strains, suggesting that the contamination could have originated from different sources. In our experiments, when sugar was added and the beverage was kept in thermic bottles there was a significant increase in B. cereus sensu lato levels, which may increase the risk of food poisoning. These results highlight the need for additional studies on this subject to better evaluate coffee as a food poisoning vehicle.

Some strains of the Gram-positive spore-forming bacterium Bacillus thuringiensis are very toxic to various insect larvae. The insecticidal activity is determined by different cytoplasmic insoluble proteins produced during sporulation which aggregate to form parasporal crystals (E Schnepf et al. 1998 Microbiol Mol Biol Rev 62: 775-806). A few strains of B. thuringiensis are used as bioinsecticides to control pests and vector insects in the orders Diptera, Lepidoptera and Coleoptera (PAW Martin & RS Travers 1989 Appl Environ Microbiol 55: 2437-2442) and more recently new isolates of B. thuringiensis have been shown to be active against Hymenoptera, Homoptera, Orthoptera and Mallophaga, as well as nematodes, mites and pro-Partially supported by Conselho Nacional de Desenvolvimento Científico e Tecnológico.

IntroductionLysinibacillus sphaericus is a highly effective and specific bioinsecticide used for the control of Culicidae larvae.ObjectiveThis study aimed to identify and characterize L. sphaericus strains isolated from Culex quinquefasciatus larvae in Brazil.MethodsC. quinquefasciatus larvae were collected from streams in the urban area of São Paulo state. L. sphaericus strains were identified through cytomorphology, biochemical, and physiological analyses. Qualitative bioassays were performed to evaluate the toxicity of the strains against C. quinquefasciatus. The crystal compound protein pattern of L. sphaericus strains was analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis. Five reference strains were used as standards in all tests performed. Repetitive extragenic palindromic elements-polymerase chain reaction (REP-PCR) was utilized in an attempt to differentiate pathogenic and nonpathogenic isolates.ResultsTwenty-one strains were isolated. Only one presented toxic activity against C. quinquefasciatus. REP-PCR results identified 23 patterns among the 26 strains used in the study, and the fragment analysis showed low similarity (16%) between L. sphaericus isolates and the five reference strains.ConclusionComparison of strains isolated in this study using REP-PCR showed a low similarity to other strains, demonstrating the high intraspecific variability for L. sphaericus.