Inactivation of a Novel Gene Produces a Phenotypic Variant Cell and Affects the Symbiotic Behavior of Xenorhabdus nematophilus

Abstract

Xenorhabdus nematophilus is an insect pathogen that lives in a symbiotic association with a specific entomopathogenic nematode. During prolonged culturing, variant cells arise that are deficient in numerous properties. To understand the genetic mechanism underlying variant cell formation, a transposon mutagenesis approach was taken. Three phenotypically similar variant strains of X. nematophilus, each of which contained a single transposon insertion, were isolated. The insertions occurred at different locations in the chromosome. The variant strain, ANV2, was further characterized. It was deficient in several properties, including the ability to produce antibiotics and the stationary-phase-induced outer membrane protein, OpnB. Unlike wild-type cells, ANV2 produced lecithinase. The emergence of ANV2 from the nematode host was delayed relative to the emergence of the parental strain. The transposon in ANV2 had inserted in a gene designated var1, which encodes a novel protein composed of 121 amino acid residues. Complementation analysis confirmed that the pleiotropic phenotype of the ANV2 strain was produced by inactivation of var1. Other variant strains were not complemented by var1. These results indicate that inactivation of a single gene was sufficient to promote variant cell formation in X. nematophilus and that disruption of genetic loci other than var1 can result in the same pleiotropic phenotype.

The global regulator Lrp contributes to mutualism, pathogenesis and phenotypic variation in the bacterium Xenorhabdus nematophila

“…In Escherichia coli , early appearing GASP strains contain mutant alleles of the stationary phase sigma factor, RpoS (Zambrano et al ., 1993), but an X. nematophila rpoS null mutant has no effects on phenotypic variation (Vivas and Goodrich‐Blair, 2001). A mutant hunt for genes that convert X. nematophila from primary to secondary form revealed the involvement of var1 (named for its involvement in variation) (Volgyi et al ., 2000). The function of this gene, however, is not known, and thus, the regulatory mechanisms controlling X. nematophila phenotypic variation remain obscure.…”

Analysis of the PixA Inclusion Body Protein of Xenorhabdus nematophila

“…and Photorhabdus spp. (4,6,20,21). The X. nematophila variant cells are able to support growth of S. carpocapsae, while Heterorhabditis bacteriophora nematodes are unable to grow on the secondary variant strains of P. luminescens.…”

The ner Gene of Photorhabdus : Effects on Primary-Form-Specific Phenotypes and Outer Membrane Protein Composition

“…undergo a similar range of phenotypic switching whose mechanism is equally poorly understood. Recently, a gene was inactivated in primary cells of X. nematophilus which generated a strain that was secondary-like for most phenotypes tested (39). In addition, the mutant was defective in its packaging or survival in or release from the nematode.…”

“…Whether or not the secondary form is a spontaneous mutant, it is clear that the primary-phase-specific phenotypes are coordinately regulated, and the induction of phase shift by multiple independent transposon insertions in X. nematophilus (39) implies that a complex pathway regulates this kind of phenotypic variation. In the work described here, we hypothesize that phenotypic variation in Photorhabdus is controlled by a cascade of regulatory genes and ask whether a repressor inhibits expression of the phase-specific phenotypes in the secondary form.…”