Genome analysis of an entomopathogenic nematode belonging to the genus Oscheius and its insect pathogenic bacterial endosymbiont

Abstract

The use of synthetic chemical pesticides has several negative implications for the Agricultural industry, which include the development of resistance to the insecticides, crop contamination and the killing of non-target insects. This has brought many scientists in the field of nematology and entomology to investigate biological control agents which can help solve identified challenges and these biocontrol agents have also included entomopathogenic nematodes. The majority of entomopathogenic nematodes species that have been isolated belong to Heterorhabditids and Steinernematids which act as vectors for insect pathogenic bacteria species belonging to the genera, Photorhabdus and Xenorhabdus, respectively. However, other species of nematodes, one of which includes a strain of Caenorhabditis briggsae, have also been shown to act as a vector for an insect pathogenic strain of Serratia marcescens. Oscheius sp. TEL-2014 EPNs have been observed to act as vectors for insect pathogenic bacteria belonging to the genus Serratia. In this study a novel insect pathogenic Serratia sp. strain TEL was isolated from the gut of infective juveniles belonging to a species of Oscheius sp. TEL-2014. Next generation sequencing of the bacteria was conducted by generating genomic DNA paired-end libraries with the Nextera DNA sample preparation kit (Illumina) and indexed using the Nextera index kit (Illumina). Paired-end (2 × 300 bp) sequencing was performed on a MiSeq Illumina using the MiSeq reagent kit v3 at the Agricultural Research Council Biotechnology Platform. Quality control and adapter trimming was performed and the genome was assembled using SPADES. 19 contigs were generated with an average length of 301767 bp and N50 of 200,110 bp. The genome of the Serratia sp. TEL was found to be 5,000,541 bp in size, with a G+C content of 59.1%, which was similar to that of other Serratia species previously identified. Furthermore, the contigs were annotated using NCBI Prokaryotic Genome Automatic Annotation Pipeline. Features of the annotated genome included protein encoding sequence or genes, rRNA encoding genes, tRNA encoding genes, ncRNA sequences and repeat regions. 4,647 genes were found and 4,495 were protein-coding sequences (CDS). The genome contains 36 pseudo genes, 2 CRISPR arrays, 13 rRNA genes with five operons (5S, 16S, 23S), 88 tRNAs genes, 15 ncRNA sequences and 9 frameshifted genes. Several genes involved in virulence, disease, defense, stress response, cell division, motility and chemotaxis were identified. This genome sequence will allow for the investigation of identified genes and that will be critical in furthering the understanding of the insect pathogenicity of Serratia sp. strain TEL. Furthermore, it will provide additional genomic insights about the insect-nematode interactions and thus help us improve their ability to be used as biological control agents in agricultural industries. Oscheius sp. TEL-2014 was tested for its entomopathogenicity and it was found that this species was able to infect and kill two model insects Galleria mellonella and Tenebrio molitor. This new nematode species brought 100% mortality within 72 h post-exposure in G. mellonella and whereas, within 96 hours in T. molitor. Following morphometrics analysis of Oscheius sp. TEL-2014 it was concluded that this nematode is described as a novel entomopathogenic nematode species based on its morphometrics and 18S rRNA gene sequence originality. Whole genome sequencing of Oscheius sp. TEL-2014 inbred lines (7 and 13) was performed using Illumina Hiseq sequencing system and paired ends library preparation protocol. Sequencing reads assembled on Velvet resulted in generation of 75965 contigs (line 7) and 53190 contigs (line 13). Gene prediction tools showed that proteins involved in gene expression and DNA replication are present in Oscheius sp. TEL-2014. The draft genome of Oscheius nematodes will support the improvement and initiation of further studies intended to help us understand the molecular and metabolic processes in this genus.