Analysis of complete genome sequence and major surface antigens of Neorickettsia helminthoeca, causative agent of salmon poisoning disease

Abstract

Summary Neorickettsia helminthoeca, a type species of the genus Neorickettsia, is an endosymbiont of digenetic trematodes of veterinary importance. Upon ingestion of salmonid fish parasitized with infected trematodes, canids develop salmon poisoning disease (SPD), an acute febrile illness that is particularly severe and often fatal in dogs without adequate treatment. We determined and analysed the complete genome sequence of N. helminthoeca: a single small circular chromosome of 884 232 bp encoding 774 potential proteins. N. helminthoeca is unable to synthesize lipopolysaccharides and most amino acids, but is capable of synthesizing vitamins, cofactors, nucleotides and bacterioferritin. N. helminthoeca is, however, distinct from majority of the family Anaplasmataceae to which it belongs, as it encodes nearly all enzymes required for peptidoglycan biosynthesis, suggesting its structural hardiness and inflammatory potential. Using sera from dogs that were experimentally infected by feeding with parasitized fish or naturally infected in southern California, Western blot analysis revealed that among five predicted N. helminthoeca outer membrane proteins, P51 and strain‐variable surface antigen were uniformly recognized. Our finding will help understanding pathogenesis, prevalence of N. helminthoeca infection among trematodes, canids and potentially other animals in nature to develop effective SPD diagnostic and preventive measures. Recent progresses in large‐scale genome sequencing have been uncovering broad distribution of Neorickettsia spp., the comparative genomics will facilitate understanding of biology and the natural history of these elusive environmental bacteria.

Isolation and Molecular Analysis of a NovelNeorickettsiaSpecies That Causes Potomac Horse Fever

“…Phylogenetic analysis revealed that only 10 of the isolates were N. risticii strains, whereas the remaining two isolates (Fin17 and Tom16) were a previously uncharacterized Neorickettsia sp. The results were corroborated by whole-genome sequencing of Fin17 and genomic comparison with N. risticii, N. sennetsu, and N. helminthoeca (21,45,46,48). Experimental inoculation of Fin17 clearly showed that it can infect horses and cause PHF or subclinical infection.…”

“…These findings suggest that the multiple ssa genes in Fin17, N. risticii, and N. sennetsu arose as gene duplication events after the two species diverged from N. helminthoeca. Ssas have extensive intramolecular sequence repeats (34,45,46,48). Notably, the Fin17/Tom16 16S rRNA and P51 gene sequences were phylogenetically separated from those of other PHF-related strains, whereas the presence of Ssa3 repeat structures was similar to those of the N. risticii strains and distinct from those of N. sennetsu and the SF agents, suggesting that Ssa3 may be involved in PHF infection and pathogenesis.…”

“…Native P51 of N. sennetsu has porin activity, which is an important means for nutrient acquisition among Gram-negative bacteria (50). The p51 gene is found in N. risticii, N. sennetsu, N. helminthoeca, the SF agent, and Neorickettsia species from F. hepatica (21,45,46,48). P51 is the major surface-exposed protein of N. risticii strains and is highly immunogenic in animals infected with N. risticii or N. helminthoeca (21,34,45,51).…”

“…The p51 gene is found in N. risticii, N. sennetsu, N. helminthoeca, the SF agent, and Neorickettsia species from F. hepatica (21,45,46,48). P51 is the major surface-exposed protein of N. risticii strains and is highly immunogenic in animals infected with N. risticii or N. helminthoeca (21,34,45,51). The bacterial Ssa protein was initially thought to be encoded by a single gene and may be an immunologically reactive surface protein that could be developed as a vaccine (32,52,53).…”

“…Because a signal peptide sequence is not predicted for Ssa3 (46), if it is a bacterial surface protein, it is likely to be secreted in a Sec-independent manner (32,52,53). Our previous whole-genome sequence analyses of N. risticii and N. sennetsu revealed that they harbored three and two (ssa2 has degenerated in N. sennetsu) tandemly repeated ssa genes, respectively (46,48), whereas N. helminthoeca carries only a single ssa gene (45). The current genome analysis showed that Fin17 carries 3 ssa genes (see Table S3 in the supplemental material).…”