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The density and regulation of microbial populations are important factors in the success of symbiotic associations. High bacterial density may improve transmission to the next generation, but excessive replication could turn out to be costly to the host and result in higher virulence. Moreover, differences in virulence may also depend on the diversity of symbionts. Using the maternally transmitted symbiont Wolbachia, we investigated how bacterial density and diversity are regulated and influence virulence in host insects subject to multiple infection. The model we used was the wasp Asobara tabida that naturally harbors three different Wolbachia strains, of which two are facultative and induce cytoplasmic incompatibility, whereas the third is necessary for the host to achieve oogenesis. Using insect lines infected with different subsets of Wolbachia strains, we show that: (i) some traits of A. tabida are negatively affected by Wolbachia: (ii) the physiological cost increases with the number of co-infecting strains, which also corresponds to an increase in the total bacterial density; and (iii) the densities of the two facultative Wolbachia strains are independent of one another, whereas the obligatory strain is less abundant when it is alone, suggesting that there is some positive interaction with the other strains.

Several different isolates of different organisms are analyzed to investigate the impact of the multiple infections on the expression and evolution of virulence in a Daphina-endoparasite system. High rates of multiple infections are shown to lead to replacement of less virulent parasite lines by more virulent strains.

The phylogenetic position of Spirobacillus cienkowskii is established through the analysis of genes encoding the 16S rRNA and [beta]-subunit of topoisomerase and has confirmed the presence of the pathogen in multiple species of Daphnia. The geographic distribution, host range and virulence of Spirobacillus cienkowskii have shown that it plays a vital role in the ecology and evolution of daphniids.

Selected sporulation genes and an epitope associated with the spore envelope of Pasteuria ramose and Pasteuria penetrans are compared. The presence of a shared adhesion in these two species with ecologically distant hosts has shown that there is an ancient and ecologically significant recognition process in these endospore-forming bacilli that has contributed to the virulence of both species in their respective hosts.

The study signifies the presence of variable-number tandem repeats (VNTRs) in the populations of Pasteuria ramosa, a castracting endobacterium of Daphnia species. The analysis reveals that VNTRs may act as the biomakers for the genetic characterization of these Pasteuria ramosa isolates.