Genome analysis of salivary gland hypertrophy virus (SGHV) reveals a novel large double- stranded circular DNA virus from Glossina pallidipes

We report the first complete genome sequence of tsetse fly Glossina pallidipes salivary gland hypertrophy virus (GpSGHV). This virus infects several species of tsetse fly, causes salivary gland hypertrophy symptom and reduces significantly the fertility of the infected flies. The SGHV has a doublestranded circular DNA genome of 190, 9074 bp encoding 150 open reading frames (ORFs). The G + C content of the SGHV virus is 28% with a gene density of one gene per 1.7 gene/ kb. Although rod-shaped SGHV is morphologically similar to insect baculoviruses there is very little homology at the amino acid level. In fact, some SGHV genes are more homologous to eukarytotic genes than to viral genes. Extensive sequence analysis indicates that SGHV differs from all known viruses, although a few genes do display a weak homology to genes of other viruses, i.e. baculovirus, entomopoxvirus, poxvirus, whispovirus, ascovirus and invertebrate iridescent virus. In addition, most of the ORFs encode proteins that bear no homology to any known proteins suggesting that SGHV represents a novel class of insect viruses. Determination of the genome sequence of SGHV will facilitate a better understanding of the molecular mechanisms underlying the pathogenesis of this virus. Microbial approaches to insect biocontrol Diversity and ecology of Bacillus thuringiensis Jacques Mahillon, Géraldine Van der Auwera, Sophie Timmery, Florence Hoton, Izabela Swiecicka Laboratory of Food and Environmental Microbiology, Université catholique de Louvain, Croix du Sud 2/12, B-1348 Louvain-la-Neuve, Belgium, Department of Microbiology, Institute of Biology, University of Bialystok, 15-950 Bialystok, 20B Swierkowa Street, Poland Bacillus thuringiensis belongs to the group of Bacillus cereus sensu lato, which also includes Bacillus mycoides, Bacillus pseudomycoides, Bacillus weihenstephanensis and Bacillus anthracis. Although these bacteria share similar genetic background, they differ significantly by their spectrum of virulence and lifestyles (Jensen et al., 2003). Whereas B. thuringiensis is an entomopathogen (due to the production of crystal δ-endotoxins), B. anthracis displays virulence against mammals, including humans. Similarly, B. cereus sensu stricto strains comprise both “pathotypes”, such as food pathogens or eye and tooth opportunists (Jensen et al., 2003), and “ecotypes”, known to stimulate plant growth through their antimicrobial activity on phytopathogens (Silo-Suh et al., 1998). In this context, it is important to better understand the diversity and variability of these bacteria, including their capacity to adapt through lateral gene transfers. Corollary, it is equally relevant to get insights into their ecology and their potential of mutual adaptation. The present communication focuses on the ecological aspects of B. thuringiensis, studied in both environmental and laboratory conditions. Although B. thuringiensis isolates have been recovered from extremely diverse niches including soil, plants, animals and food, our investigations were mainly dedicated on natural microflora associated with the digestive system of arthropods and mammals, The diversity of B. thuringiensis strains isolated from the intestines of small rodents and insectivores has been investigated using both their δ-endotoxin (crystal) gene content and their general genome organization (Swiecicka and Mahillon, 2005). Interestingly, among ca. 100 bacterial isolates, only 9 distinct genotypes could be observed, indicating a clonal structure for these B. thuringiensis populations. The data also indicated no correlation between the B. thuringiensis strain genotypes and their crystal contents, suggesting the occurrence of horizontal gene transfer among these natural B. thuringiensis isolates (Swiecicka and Mahillon, 2005). These strains isolated from wild mammals were also tested for the presence of genetic determinants potentially associated with enterotoxins. In fact, all the B. thuringiensis isolates displayed various combinations of putative haemolytic and nonhemolytic enterotoxin genes (Swiecicka and Mahillon, 2006b). It is therefore important to recognize the existence of this potential virulence arsenal in natural B. thuringiensis strains and the role of small mammals in its dissemination. However, further detailed investigations will be required to better interpret these observations with regard to possible environmental and health issues. In order to characterize the behaviour of B. thuringiensis and B. cereus in terrestrial soil animals, several strains were isolated from the digestive tract of sow bugs and were genetically marked by selecting mutants resistant to antibiotic or by obtaining recombinant clones expressing the Green Fluorescent proteins. These tagged bacteria were then reintroduced in sow bugs and monitored over time using their new phenotypes. Both spores and vegetative cells of B. thuringiensis and B. cereus were recovered from the arthropods over a 30-day period, strongly suggesting that these bacteria are genuine residents of terrestrial isopods (Swiecicka and Mahillon, 2006a). Do B. thuringiensis strains exchange their plasmids (including the cry-bearing molecules) in natural settings? Although several reports have indicated the potential transfer of relevant B. thuringiensis plasmids outside laboratory conditions, i.e. soil (Vilas-Boas et al., 2000), river water (Thomas et al., 2001) or insect larvae (Thomas et al., 2000), the observed frequencies were always rather low and the transfers were mostly dependent on the growth potential of the strains. These results contrasted with those obtained recently in food matrices. Indeed, plasmid conjugation (and mobilization) has been observed to occur in several foodstuffs, at frequencies higher than those obtained in standard laboratory conditions (Van der Auwera et al., 2007; Entomological Research 37 (Suppl. 1) (2007) A11–A73 A33 © 2007 The Entomological Society of Korea and Blackwell Publishing Asia Pty Ltd ICIBI 2007, Daegu, Republic of Korea A34 Entomological Research 37 (Suppl. 1) (2007) A11–A73 © 2007 The Entomological Society of Korea and Blackwell Publishing Asia Pty Ltd Mahillon et al., unpublished results). It would now be interesting to further investigate the actual contribution of these phenomena to the genetic and virulence diversity observed among the B. thuringiensis strains. The expected results should shed new light on the genetic and genomic tools used to shuffle and re-assort the entomopathogenic properties of B. thuringiensis.