Pyrimidine Nucleoside Salvage Confers an Advantage to Xenorhabdus nematophila in Its Host Interactions

Abstract

Xenorhabdus nematophila is a mutualist of entomopathogenic nematodes and a pathogen of insects. To begin to examine the role of pyrimidine salvage in nutrient exchange between X. nematophila and its hosts, we identified and mutated an X. nematophila tdk homologue. X. nematophila tdk mutant strains had reduced virulence toward Manduca sexta insects and a competitive defect for nematode colonization in plate-based assays. Provision of a wild-type tdk allele in trans corrected the defects of the mutant strain. As in Escherichia coli, X. nematophila tdk encodes a deoxythymidine kinase, which converts salvaged deoxythymidine and deoxyuridine nucleosides to their respective nucleotide forms. Thus, nucleoside salvage may confer a competitive advantage to X. nematophila in the nematode intestine and be important for normal entomopathogenicity.

Influence of nematode age and culture conditions on morphological and physiological parameters in the bacterial vesicle of Steinernema carpocapsae (Nematoda: Steinernematidae)

“…The data presented here expand the understanding of which metabolic pathways are important for the establishment of colonization. Earlier analyses Orchard and Goodrich-Blair, 2005) had indicated that X. nematophila folate biosynthesis (defective in pabA and aroA mutants) is necessary for normal colonization. There are numerous folate-dependent biosynthetic pathways in bacteria, but it was suggested that folate-dependent methionine biosynthesis in particular might be necessary for eYcient colonization.…”

An encoded N-terminal extension results in low levels of heterologous protein production in Escherichia coli

“…Cultures were grown in a tube roller at 30°C in Luria-Bertani (LB) broth [11] except for the FUdR growth assays, which were performed in semi-defined medium containing 5-fluorodeoxyuridine ([12]; FUdR obtained from Fisher Scientific, Pittsburgh, PA) at 37°C with shaking in a microplate reader (Molecular Devices, Sunnyvale, CA) as described previously [3]. LB agar (20 g l -1 ) plates and all liquid media were supplemented when appropriate with kanamycin (kan; 20 μg ml -1 ), chloramphenicol (cam; 20 μg ml -1 ), isopropyl-β-D-thiogalactopyranoside (IPTG; 0.2 mM) or arabinose (0.2%).…”

“…The predicted Xenorhabdus nematophila Tdk protein is 70% identical to E. coli Tdk and has been shown to have deoxythymidine kinase activity [3], converting salvaged deoxythymidine to deoxythymidine monophosphate [4]. A translational start site was predicted for X. nematophila tdk based on alignment with tdk sequences from other organisms.…”

Microaerobic Conversion of Glycerol to Ethanol in Escherichia coli

“…Unless otherwise indicated, all DNA procedures followed standard protocols and specific recommendations from manufacturers. E. coli knockout mutants were constructed by the suicide vector method (16). E. coli genes were amplified from genomic DNA by using AccuPrime Pfx DNA polymerase (Life Technologies, Invitrogen) and cloned into vectors by using the Quick Ligation kit (New England BioLabs Inc., Ipswich, MA) or In-Fusion HD cloning kit (Clontech Laboratories Inc., Mountain View, CA).…”