Cooperative endurance: A remarkable strategy adopted by symbiotic bacteria to persist in their nematode vector

Symbioses between microbes and animals are ubiquitous, yet little is known about the intricate mechanisms maintaining such associations. In an emerging animal-microbe symbiosis model system represented by the partnership between insect-pathogenic bacteria Photorhabdus temperata and insect-parasitic nematodes Heterorhabditis bacteriophora, we investigated molecular mechanisms adopted by the bacteria to persist in the enduring nematode vector in search of their insect host. Using selective capture of transcribed sequences approach, 50 transcripts were identified to be up-regulated and 56 were down-regulated by the bacteria during persistence in the nematode compared with growth in culture medium. Real-time PCR analysis of 14 representative transcripts displayed 6-12 fold change in expression, reflecting a significant shift in bacterial gene expression in the nematode. The identified transcripts included but not limited to genes involved in proton transport, metabolic pathways, biofilm formation and cell motility, suggesting that the bacteria undergo major transcriptional reshaping in the nematode vector. Besides general starvation mechanisms, the bacteria induce cellular acidification to slow down growth, switch to pentose phosphate pathway to overcome oxidative stress and nutrition limitation, and shed motility but develop biofilm to persist in the nematode intestine until being released into the insect hemolymph. Our mutation data further confirm that such transcriptional reshaping is critical for bacteria to persist in the nematode infective juvenile. These findings demonstrate how the symbiotic bacteria reduce their nutritional dependence on the enduring nematode partner to ensure successful transmission of the couple to the next insect host

Differences in susceptibility of white grub species to entomopathogenic nematodes: the relative contribution of symbiotic bacteria and nematodes

As susceptibility of white grub species to entomopathogenic nematodes differs, we compared the virulence of Photorhabdus temperata and Xenorhabdus koppenhoferi, the symbionts of nematodes Heterorhabditis bacteriophora and Steinernema scarabaei, respectively, to three white grub species Popillia japonica, Rhizotrogus majalis, and Cyclocephala borealis. Both bacteria were virulent to all three grub species even at as low as 2 cells /grub. However, the median lethal dose at 48 h post injection and median lethal time at 20 cells /grub showed that P. temperata was more virulent than X. koppenhoferi to C. borealis. There were no differences in virulence of two bacteria against P. japonica and R. majalis. Although H. bacteriophora carrying P. temperata is less pathogenic than S. scarabaei carrying X. koppenhoferi to R. majalis, P. temperata grew faster than X. koppenhoferi both in vitro and in vivo. We then tested the pathogenicity of oral and hemolymph introduced H. bacteriophora to R. majalis to determine whether nematodes are able to successfully vector the bacteria into the hemolymph. Hemolymph injected H. bacteriophora were pathogenic to R. majalis indicating successful bacterial release, but orally introduced H. bacteriophora were not. Dissection of grubs confirmed that orally introduced H. bacteriophora were unable to penetrate into the hemolymph through the gut wall. Therefore, we conclude that the low susceptibility of R. majalis to H. bacteriophora is not due to the symbiotic bacteria, but is due to the nematode’s poor ability to penetrate through the gut wall to vector the bacteria into the hemolymph.This research was supported by an interdisciplinary grant and a graduate research competitive grant from the Ohio Agricultural Research and Development Center, Wooster, Ohio