Entomopathogenic nematode-associated microbiota: from monoxenic paradigm to pathobiome

Abstract

Background: The holistic view of bacterial symbiosis, incorporating both host and microbial environment, constitutes a major conceptual shift in studies deciphering host-microbe interactions. Interactions between Steinernema entomopathogenic nematodes and their bacterial symbionts, Xenorhabdus, have long been considered monoxenic two partner associations responsible for the killing of the insects and therefore widely used in insect pest biocontrol. We investigated this "monoxenic paradigm" by profiling the microbiota of infective juveniles (IJs), the soil-dwelling form responsible for transmitting Steinernema-Xenorhabdus between insect hosts in the parasitic lifecycle. Results: Multigenic metabarcoding (16S and rpoB markers) showed that the bacterial community associated with laboratory-reared IJs from Steinernema carpocapsae, S. feltiae, S. glaseri and S. weiseri species consisted of several Proteobacteria. The association with Xenorhabdus was never monoxenic. We showed that the laboratory-reared IJs of S. carpocapsae bore a bacterial community composed of the core symbiont (Xenorhabdus nematophila) together with a frequently associated microbiota (FAM) consisting of about a dozen of Proteobacteria (Pseudomonas, Stenotrophomonas, Alcaligenes, Achromobacter, Pseudochrobactrum, Ochrobactrum, Brevundimonas, Deftia, etc.). We validated this set of bacteria by metabarcoding analysis on freshly sampled IJs from natural conditions. We isolated diverse bacterial taxa, validating the profile of the Steinernema FAM. We explored the functions of the FAM members potentially involved in the parasitic lifecycle of Steinernema. Two species, Pseudomonas protegens and P. chlororaphis, displayed entomopathogenic properties suggestive of a role in Steinernema virulence and membership of the Steinernema pathobiome. Conclusions: Our study validates a shift from monoxenic paradigm to pathobiome view in the case of the Steinernema ecology. The microbial communities of low complexity associated with EPNs will permit future microbiota manipulation experiments to decipher overall microbiota functioning in the infectious process triggered by EPN in insects and, more generally, in EPN ecology.

Characterization of symbiotic and associated bacteria from entomopathogenic nematode Heterorhabditis sp. (nematode: Heterorhabditidae) isolated from India

“…For a long time, the discovery of EPN-associated bacteria more popularly referred to as "frequently associated microbiota (FAM)" has often been dismissed as laboratory/handling contaminants; but as they became frequently encountered (Walsh and Webster 2003;Gouge andSnyder 2006, andSingh et al 2014) in the hemolymph of infected insects, researchers started paying attention to them. FAM consists predominantly of members of Proteobacteria, while other genera are also present, except for a few of them like P. entomophila, their role in killing the insect host has not been established (Ogier et al 2020). However, their co-existence with the core symbionts even in nematodes that have been maintained for generations in laboratory conditions indicates that they may have some import roles to play, which may be discovered in future.…”

“…However, their co-existence with the core symbionts even in nematodes that have been maintained for generations in laboratory conditions indicates that they may have some import roles to play, which may be discovered in future. In fact, it is now widely debated whether the pathogenicity of IJs is exclusively because of the core symbionts (Xenorhabdus and Photorhabdus) or it is a combined effect of core symbiont and FAM (Ogier et al 2020); such studies point towards the involvement of a rather "pathobiome" and the involvement of more than one pathogen in establishing a disease as against the widely believed concept of one pathogen one disease.…”

Draft Genome Sequence of a Noncognate Bacterium, Achromobacter sp. Strain Bel, Associated with a Rhabditid Entomopathogenic Nematode

“…Entomopathogenic nematodes of the genera Steinernema , Heterorhabditis , and Oscheius are symbiotically associated with cognate symbiotic Enterobacteriaceae bacteria of the Xenorhabdus , Photorhabdus , and Serratia genera, respectively ( 3 ). Other noncognate bacterial species from genera such as Pseudomonas , Acinetobacter , Staphylococcus , Stenotrophomonas , Ochrobactrum , and Achromobacter have been isolated from EPNs ( 4 – 7 ). Although the function of noncognate bacteria is not fully delineated, some noncognate species have been reported to contribute toward the entomopathogenicity of EPNs ( 7 ).…”

“…Other noncognate bacterial species from genera such as Pseudomonas , Acinetobacter , Staphylococcus , Stenotrophomonas , Ochrobactrum , and Achromobacter have been isolated from EPNs ( 4 – 7 ). Although the function of noncognate bacteria is not fully delineated, some noncognate species have been reported to contribute toward the entomopathogenicity of EPNs ( 7 ). Thus, the genome sequences of noncognate bacteria will contribute toward (i) understanding their role toward EPN pathogenicity, (ii) knowing the impact they have on nematode fitness and reproduction, and (iii) knowing how their genomic profiles differ from cognate symbiotic bacteria with regard to infectivity against insects.…”

Host-associated microbe PCR (hamPCR): accessing new biology through convenient measurement of both microbial load and community composition

“…For many host-associated microbiome samples, in particular those from plants 12 , nematodes 13 , insects [14][15][16] , and other organisms in which it is difficult or impossible to physically separate microbes from host tissues, a thorough DNA extraction yields both host and microbial DNA. For such samples, the amount of DNA from host and microbe is directly proportional to the number of cells sampled 17,18 , and therefore the ratio of microbial DNA to host DNA is an intrinsic measure of the microbial load of the sample 6,12,[19][20][21] .…”