Quorum Sensing Influences Vibrio harveyi Growth Rates in a Manner Not Fully Accounted For by the Marker Effect of Bioluminescence

The light-emitting Vibrios provide excellent material for studying the interaction of cellular communication with growth rate because bioluminescence is a convenient marker for quorum sensing. However, the use of bioluminescence as a marker is complicated because bioluminescence itself may affect growth rate, e.g. by diverting energy. quorum mutants. growth rate can be either positive or negative and includes both bioluminescence-dependent and independent components. Bioluminescence tends to slow growth rate but not enough to account for the effects of quorum sensing on growth rate

Peptidoglycan Induces Loss of a Nuclear PGRP During Host Tissue Development in a Beneficial Animal–Bacterial Symbiosis

Peptidoglycan recognition proteins (PGRPs) are mediators of innate immunity and recently have been implicated in developmental regulation. To explore the interplay between these two roles, we characterized a PGRP in the host squid Euprymna scolopes (EsPGRP1) during colonization by the mutualistic bacterium Vibrio fischeri. Previous research on the squid-vibrio symbiosis had shown that, upon colonization of deep epithelium-lined crypts of the host light organ, symbiont-derived peptidoglycan monomers induce apoptosis-mediated regression of remote epithelial fields involved in the inoculation process. In this study, immunofluorescence microscopy revealed that EsPGRP1 localizes to the nuclei of epithelial cells, and symbiont colonization induces the loss of EsPGRP1 from apoptotic nuclei. The loss of nuclear EsPGRP1 occurred prior to DNA cleavage and breakdown of the nuclear membrane, but followed chromatin condensation, suggesting that it occurs during late stage apoptosis. Experiments with purified peptidoglycan monomers and with V. fischeri mutants defective in peptidoglycan-monomer release provided evidence that these molecules trigger nuclear loss of EsPGRP1 and apoptosis. The demonstration of a nuclear PGRP is unprecedented, and the dynamics of EsPGRP1 during apoptosis provide a striking example of a connection between microbial recognition and developmental responses in the establishment of symbiosis

Phylogenetic Evidence for Ancient and Persistent Environmental Symbiont Reacquisition in Largidae (Hemiptera: Heteroptera)

The insect order Hemiptera, one of the best-studied insect lineages with respect to bacterial symbioses, still contains major branches that lack comprehensive characterization of associated bacterial symbionts. The Pyrrhocoroidea (Largidae [220 species] and Pyrrhocoridae [∼300 species]) is a clade of the hemipteran infraorder Pentatomomorpha. Studies on bacterial symbionts of this group have focused on members of Pyrrhocoridae, but recent examination of species of two genera of Largidae demonstrated divergent symbiotic complexes in these putative sister families. We surveyed the associated bacterial diversity of this group using paired-end Illumina sequencing and targeted Sanger sequencing of bacterial 16S rRNA amplicons of 30 pyrrhocoroid taxa, including 17 species of Largidae, in order to determine bacterial associates and the similarity of associated microbial communities among species. We also used molecular data (4,800 bp in 5 loci, for 57 ingroup and 12 outgroup taxa) to infer a phylogeny of the host superfamily, in order to trace the evolution of symbiotic complexes among Pentatomomorpha species. We undertook multiple lines of investigation (i.e., experimental rearing, fluorescence in situ hybridization microscopy, and phylogenetic and coevolutionary analyses) to elucidate potential transmission routes for largid symbionts. We found a prevalent and specific association of Largidae with Burkholderia strains of the plant-associated beneficial and environmental clade, housed in midgut tubules. As in other distantly related Heteroptera, symbiotic bacteria seem to be acquired from the environment every generation. We review the current understanding of symbiotic complexes within Pentatomomorpha and discuss means to further investigate the evolution and function of these symbioses. IMPORTANCE Obligate symbioses with bacteria are common in insects, particularly Hemiptera, in which various forms of symbiosis occur. However, knowledge regarding symbionts remains incomplete for major hemipteran lineages. Thus, an accurate understanding of how these partnerships evolved and changed over millions of years is not yet achievable. We contribute to our understanding of the evolution of symbiotic complexes in Hemiptera by characterizing bacterial associates of Pyrrhocoroidea, focusing on the family Largidae. Members of Largidae are associated with specific symbiotic Burkholderia strains from a different clade than Burkholderia symbionts in other Burkholderia-associated Hemiptera. Evidence suggests that species of Largidae reacquire specific symbiotic bacteria from the environment every generation, which is a rare strategy for insects, with potentially volatile evolutionary ramifications, but one that must have persisted in Largidae and related lineages since their origin in the Cretaceous Period

Mutations in ampG and Lytic Transglycosylase Genes Affect the Net Release of Peptidoglycan Monomers from Vibrio fischeri▿ †

The light-organ symbiont Vibrio fischeri releases N-acetylglucosaminyl-1,6-anhydro-N-acetylmuramylalanyl-γ-glutamyldiaminopimelylalanine, a disaccharide-tetrapeptide component of peptidoglycan that is referred to here as “PG monomer.” In contrast, most gram-negative bacteria recycle PG monomer efficiently, and it does not accumulate extracellularly. PG monomer can stimulate normal light-organ morphogenesis in the host squid Euprymna scolopes, resulting in regression of ciliated appendages similar to that triggered by infection with V. fischeri. We examined whether the net release of PG monomers by V. fischeri resulted from lytic transglycosylase activity or from defects in AmpG, the permease through which PG monomers enter the cytoplasm for recycling. An ampG mutant displayed a 100-fold increase in net PG monomer release, indicating that AmpG is functional. The ampG mutation also conferred the uncharacteristic ability to induce light-organ morphogenesis even when placed in a nonmotile flaJ mutant that cannot infect the light-organ crypts. We targeted five potential lytic transglycosylase genes singly and in specific combinations to assess their role in PG monomer release. Combinations of mutations in ltgA, ltgD, and ltgY decreased net PG monomer release, and a triple mutant lacking all three of these genes had little to no accumulation of PG monomers in culture supernatants. This mutant colonized the host as well as the wild type did; however, the mutant-infected squid were more prone to later superinfection by a second V. fischeri strain. We propose that the lack of PG monomer release by this mutant results in less regression of the infection-promoting ciliated appendages, leading to this propensity for superinfection

Complete genome sequence of Vibrio fischeri: A symbiotic bacterium with pathogenic congeners

Vibrio fischeri belongs to the Vibrionaceae, a large family of marine γ-proteobacteria that includes several dozen species known to engage in a diversity of beneficial or pathogenic interactions with animal tissue. Among the small number of pathogenic Vibrio species that cause human diseases are Vibrio cholerae, Vibrio parahaemolyticus, and Vibrio vulnificus, the only members of the Vibrionaceae that have had their genome sequences reported. Nonpathogenic members of the genus Vibrio, including a number of beneficial symbionts, make up the majority of the Vibrionaceae, but none of these species has been similarly examined. Here we report the genome sequence of V. fischeri ES114, which enters into a mutualistic symbiosis in the light organ of the bobtail squid, Euprymna scolopes. Analysis of this sequence has revealed surprising parallels with V. cholerae and other pathogens