The genome of a steinernematid-associated Pseudomonas piscis bacterium encodes the biosynthesis of insect toxins

Ryan Musumba Awori; Prasad Hendre; Nelson O Amugune

doi:10.1099/acmi.0.000659.v3

The genome of a steinernematid-associated Pseudomonas piscis bacterium encodes the biosynthesis of insect toxins

Access Microbiol. 2023 Oct 5;5(10):000659.v3. doi: 10.1099/acmi.0.000659.v3. eCollection 2023.

Authors

Ryan Musumba Awori^{1

2}, Prasad Hendre², Nelson O Amugune³

Affiliations

¹ Elakistos Biosciences, P. O. Box 19301-00100, Nairobi, Kenya.
² International Centre for Research on Agroforestry, P. O. Box 30677-00100, Nairobi, Kenya.
³ Department of Biology, University of Nairobi, P. O. Box 30197-00100, Nairobi, Kenya.

Abstract

Several species of soil-dwelling Steinernema nematodes are used in the biocontrol of crop pests, due to their natural capacity to kill diverse lepidopteran species. Although this insect-killing trait is known to be augmented by the nematodes' Xenorhabdus endosymbionts, the role of other steinernematid-associated bacterial genera in the nematode lifecycle remains unclear. This genomic study aimed to determine the potential of Pseudomonas piscis to contribute to the entomopathogenicity of its Steinernema host. Insect larvae were infected with three separate Steinernema cultures. From each of the three treatments, the prevalent bacteria in the haemocoel of cadavers, four days post-infection, were isolated. These three bacterial isolates were morphologically characterised. DNA was extracted from each of the three bacterial isolates and used for long-read genome sequencing and assembly. Assemblies were used to delineate species and identify genes that encode insect toxins, antimicrobials, and confer antibiotic resistance. We assembled three complete genomes. Through digital DNA-DNA hybridisation analyses, we ascertained that the haemocoels of insect cadavers previously infected with Steinernema sp. Kalro, Steinernema sp. 75, and Steinernema sp. 97 were dominated by Xenorhabdus griffiniae Kalro, Pseudomonas piscis 75, and X. griffiniae 97, respectively. X. griffiniae Kalro and X. griffiniae 97 formed a subspecies with other X. griffiniae symbionts of steinernematids from Kenya. P. piscis 75 phylogenetically clustered with pseudomonads that are characterised by high insecticidal activity. The P. piscis 75 genome encoded the production pathway of insect toxins such as orfamides and rhizoxins, antifungals such as pyrrolnitrin and pyoluteorin, and the broad-spectrum antimicrobial 2,4-diacetylphloroglucinol. The P. piscis 75 genome encoded resistance to over ten classes of antibiotics, including cationic lipopeptides. Steinernematid-associated P. piscis bacteria hence have the biosynthetic potential to contribute to nematode entomopathogenicity.

Keywords: Pseudomonas piscis; Xenorhabdus; bacterial insect toxins; biosynthetic gene clusters; digital DNA-DNA hybridisation; nematode microbiota; orfamides.