Elucidation of the Photorhabdus temperata Genome and Generation of a Transposon Mutant Library To Identify Motility Mutants Altered in Pathogenesis.
The entomopathogenic nematode Heterorhabditis bacteriophora forms a specific mutualistic association with its bacterial partner Photorhabdus temperata. The microbial symbiont is required for nematode growth and development, and symbiont recognition is strain specific. The aim of this study was to sequence the genome of P. temperata and identify genes that plays a role in the pathogenesis of the Photorhabdus-Heterorhabditis symbiosis. A draft genome sequence of P. temperata strain NC19 was generated. The 5.2-Mb genome was organized into 17 scaffolds and contained 4,808 coding sequences (CDS). A genetic approach was also pursued to identify mutants with altered motility. A bank of 10,000 P. temperata transposon mutants was generated and screened for altered motility patterns. Five classes of motility mutants were identified: (i) nonmotile mutants, (ii) mutants with defective or aberrant swimming motility, (iii) mutant swimmers that do not require NaCl or KCl, (iv) hyperswimmer mutants that swim at an accelerated rate, and (v) hyperswarmer mutants that are able to swarm on the surface of 1.25% agar. The transposon insertion sites for these mutants were identified and used to investigate other physiological properties, including insect pathogenesis. The motility-defective mutant P13-7 had an insertion in the RNase II gene and showed reduced virulence and production of extracellular factors. Genetic complementation of this mutant restored wild-type activity. These results demonstrate a role for RNA turnover in insect pathogenesis and other physiological functions.


OBJECTIVE
The relationship between Photorhabdus and entomopathogenic nematode Heterorhabditis represents a well-known mutualistic system that has potential as a biological control agent. The elucidation of the genome of the bacterial partner and role that RNase II plays in its life cycle has provided a greater understanding of Photorhabdus as both an insect pathogen and a nematode symbiont.
Department of Cellular, Molecular, and Biomedical Sciences, University of New Hampshire, Durham, New Hampshire, USA.
Sheldon
S
Hurst
Department of Cellular, Molecular, and Biomedical Sciences, University of New Hampshire, Durham, New Hampshire, USA.
Holli
H
Rowedder
Department of Cellular, Molecular, and Biomedical Sciences, University of New Hampshire, Durham, New Hampshire, USA.
Brandye
B
Michaels
Department of Cellular, Molecular, and Biomedical Sciences, University of New Hampshire, Durham, New Hampshire, USA.
Hannah
H
Bullock
Department of Cellular, Molecular, and Biomedical Sciences, University of New Hampshire, Durham, New Hampshire, USA.
Ryan
R
Jackobeck
Department of Cellular, Molecular, and Biomedical Sciences, University of New Hampshire, Durham, New Hampshire, USA.
Feseha
F
Abebe-Akele
Department of Cellular, Molecular, and Biomedical Sciences, University of New Hampshire, Durham, New Hampshire, USA.
Umjia
U
Durakovic
Department of Cellular, Molecular, and Biomedical Sciences, University of New Hampshire, Durham, New Hampshire, USA.
Jon
J
Gately
Department of Cellular, Molecular, and Biomedical Sciences, University of New Hampshire, Durham, New Hampshire, USA.
Erik
E
Janicki
Department of Cellular, Molecular, and Biomedical Sciences, University of New Hampshire, Durham, New Hampshire, USA louis.tisa@unh.edu.
Louis S
LS
Tisa
D000818:Animals; D004251:DNA Transposable Elements / Q000235:genetics*; D004269:DNA, Bacterial; D015964:Gene Expression Regulation, Bacterial; D015723:Gene Library; D016680:Genome, Bacterial; D006790:Host-Parasite Interactions; D009036:Moths / Q000469:parasitology; D009068:Movement; D009154:Mutation; D009348:Nematoda / Q000382:microbiology* / Q000502:physiology; D020637:Photorhabdus / Q000235:genetics*; D013559:Symbiosis
D004251:DNA Transposable Elements; D004269:DNA, Bacterial