Helicobacter monodelphidis sp. nov. and Helicobacter didelphidarum sp. nov., isolated from grey short-tailed opossums (Monodelphis domestica) with endemic cloacal prolapses

Abstract

In a search for potential causes of increased prolapse incidence in grey short-tailed opossum colonies, samples from the gastrointestinal tracts of 94 clinically normal opossums with rectal prolapses were screened for Helicobacter species by culture and PCR. Forty strains of two novel Helicobacter species which differed from the established Helicobacter taxa were isolated from opossums with and without prolapses. One of the Helicobacter species was spiral-shaped and urease-negative whereas the other Helicobacter strain had fusiform morphology with periplasmic fibres and was urease-positive. 16S rRNA gene sequence analysis revealed that all the isolates had over 99 % sequence identity with each other, and were most closely related to Helicobacter canadensis . Strains from the two novel Helicobacter species were subjected to gyrB and hsp60 gene and whole genome sequence analyses. These two novel Helicobacter species formed separate phylogenetic clades, divergent from other known Helicobacter species. The bacteria were confirmed as novel Helicobacter species based on digital DNA–DNA hybridization and average nucleotide identity analysis of their genomes, for which we propose the names Helicobacter monodelphidis sp. nov. with the type strain MIT 15-1451T (=LMG 29780T=NCTC 14189T) and Helicobacter didelphidarum sp. nov with type strain MIT 17-337T (=LMG 31024T=NCTC 14188T)

Helicobacter cyclurae sp. Nov., Isolated From Endangered Blue Iguanas (Cyclura lewisi)

“…A whole-genome phylogenetic tree constructed from the multi-sequence alignment of concatenated core genes confirms the iguana helicobacter is distinct from other known Helicobacter species, including the species isolated from reptiles described by Gilbert et al (2017). Instead, H. cyclurae was most closely placed with Helicobacter monodelphidis MIT 15-1451 isolated from captive opossums with cloacal prolapse (Shen et al, 2020). ANI and dDDH comparisons between the iguana isolate genome and other Helicobacter species were below 95 and 70%, respectively, supporting the finding that the iguana isolate is a novel Helicobacter species (Table 3).…”

“…The novel helicobacter was characterized by biochemical and whole-genome sequencing; 16S rRNA, gyrB, and hsp60 gene sequencing; and ultrastructural characterization, following the guidelines described by On et al (2017) pertaining to the minimal standards for describing new Helicobacter spp. (Shen et al, 2020). According to these guidelines, these phenotypic and genotypic characterizations are necessary to effectively and unambiguously distinguish a new taxon from extant species and subspecies and determine phylogeny in the genus.…”

“…This was particularly important for supporting that H. cyclurae is a novel species, considering helicobacters have been previously isolated from reptilian hosts (Gilbert et al, 2014). A single phenotypic and/or genotypic method is insufficient to make accurate taxonomic determinations, and as demonstrated in our recent publication describing two novel Helicobacter species isolated from opossums, the aggregate of phenotypic and genotypic data is appropriate instead (Shen et al, 2020).…”

“…Detailed biochemical characterization analysis was performed on five individual isolates using a RapID TM NH System (Remel Laboratories, Lenexus, KS). Biochemical characterization of urease, catalase, and oxidase production; sensitivity to nalidixic acid and cephalothin; and growth in the presence of 1% glycine were analyzed as previously described by our laboratory (Shen et al, 2005(Shen et al, , 2017 and following the guideline of minimal standards for describing new species belonging to the families Campylobacteraceae and Helicobacteraceae (On et al, 2017;Shen et al, 2020).…”

“…Number of flagella 1 2--5 2 2 7-10 3-4 2 2 2 2 2 1-2 6-12 1 14-20 2 2 2 1-2 7-14 1 2 2 2 7-14 10-14 4-8 2 1 4-8 2 6-12 4-10 5-7 1-2 1 DNA G+C content (mol%) (On et al, 2017;Shen et al, 2020). concatenated core gene sequences were determined followed by MAFFT for multi-sequence alignment and FastTree to infer approximately maximum-likelihood phylogenetic trees.…”