Diversity of Xenorhabdus and Photorhabdus spp. and their symbiotic entomopathogenic nematodes from Thailand

Abstract

Xenorhabdus and Photorhabdus spp. are bacterial symbionts of entomopathogenic nematodes (EPNs). In this study, we isolated and characterized Xenorhabdus and Photorhabdus spp. from across Thailand together with their associated nematode symbionts, and characterized their phylogenetic diversity. EPNs were isolated from soil samples using a Galleria-baiting technique. Bacteria from EPNs were cultured and genotyped based on recA sequence. The nematodes were identified based on sequences of 28S rDNA and internal transcribed spacer regions. A total of 795 soil samples were collected from 159 sites in 13 provinces across Thailand. A total of 126 EPNs isolated from samples taken from 10 provinces were positive for Xenorhabdus (n = 69) or Photorhabdus spp. (n = 57). Phylogenetic analysis separated the 69 Xenorhabdus isolates into 4 groups. Groups 1, 2 and 3 consisting of 52, 13 and 1 isolates related to X. stockiae, and group 4 consisting of 3 isolates related to X. miraniensis. The EPN host for isolates related to X. stockiae was S. websteri, and for X. miraniensis was S. khoisanae. The Photorhabdus species were identified as P. luminescens (n = 56) and P. asymbiotica (n = 1). Phylogenenic analysis divided P. luminescens into five groups. Groups 1 and 2 consisted of 45 and 8 isolates defined as subspecies hainanensis and akhurstii, respectively. One isolate was related to hainanensis and akhurstii, two isolates were related to laumondii, and one isolate was the pathogenic species P. asymbiotica subsp. australis. H. indica was the major EPN host for Photorhabdus. This study reveals the genetic diversity of Xenorhabdus and Photorhabdus spp. and describes new associations between EPNs and their bacterial symbionts in Thailand.

Figure 1. Geographical distribution of sample collection from 159 sites in Thailand and the distribution of sampling sites that were positive or negative for Steinernema and Heterorhabditis.

Figure 2. Maximum likelihood tree based on a 646 bp region of recA for 69 Xenorhabdus isolates from Thailand (codes ending with TH), together with Xenorhabdus sequences downloaded from the GenBank database (shown in red).

Bootstrap values are based on 1,000 replicates. Numbers shown above branches are bootstrap percentages for clades supported above the 50% level. The bar indicates 1% sequence divergence. The EPN species from which they were isolated are shown.

Figure 3. Maximum likelihood tree based on a 646 bp region of recA for 57 Photorhabdus isolates from Thailand (codes ending with TH), together with Photorhabdus sequences downloaded from GenBank (shown in red).

Bootstrap values are based on is 1,000 replicates. Numbers shown above branches are bootstrap percentages for clades supported above the 50% level. The bar indicates 1% sequence divergence. The EPN species from which they were isolated are shown.