Polkinghorne, Adam

AbstractAdvances in culture-independent methods have meant that we can more readily detect and diagnose emerging infectious disease threats in humans and animals. Metagenomics is fast becoming a popular tool for detection and characterisation of novel bacterial pathogens in their environment, and is particularly useful for obligate intracellular bacteria such as Chlamydiae that require labour-intensive culturing. We have used this tool to investigate the microbial metagenomes of Chlamydia-positive cloaca and choana samples from snakes. The microbial complexity within these anatomical sites meant that despite previous detection of chlamydial 16S rRNA sequences by single-gene broad-range PCR, only a chlamydial plasmid could be detected in all samples, and a chlamydial chromosome in one sample. Comparative genomic analysis of the latter revealed it represented a novel taxon, Ca. Chlamydia corallus, with genetic differences in regards to purine and pyrimidine metabolism. Utilising statistical methods to relate plasmid phylogeny to the phylogeny of chromosomal sequences showed that the samples also contain additional novel strains of Ca. C. corallus and two putative novel species in the genus Chlamydia. This study highlights the value of metagenomics methods for rapid novel bacterial discovery and the insights it can provide into the biology of uncultivable intracellular bacteria such as Chlamydiae.

Summary Several Chlamydiales families are associated with epitheliocystis, a common condition of the fish gill epithelium. These families share common ancestors with the Chlamydiaceae and environmental Chlamydiae . Due to the lack of culture systems, little is known about the biology of these chlamydial fish pathogens. We investigated epitheliocystis in cultured Orange‐spotted grouper ( Epinephelus coioides ) from North Queensland, Australia. Basophilic inclusions were present in the gills of 22/31 fish and the presence of the chlamydial pathogen in the cysts was confirmed by in situ hybridization. Giant grouper ( Epinephelus lanceolatus ) cultured in the same systems were epitheliocystis free. 16S rRNA gene sequencing revealed a novel member of the Candidatus Parilichlamydiaceae: Ca . Similichlamydia epinephelii. Using metagenomic approaches, we obtained an estimated 68% of the chlamydial genome, revealing that this novel chlamydial pathogen shares a number of key pathogenic hallmarks with the Chlamydiaceae , including an intact Type III Secretion system and several chlamydial virulence factors. This provides additional evidence that these pathogenic mechanisms were acquired early in the evolution of this unique bacterial phylum. The identification and genomic characterization of Ca . S. epinephelii provides new opportunities to study the biology of distantly‐related chlamydial pathogens while shining a new light on the evolution of pathogenicity of the Chlamydiaceae .

Summary Gill disease in salmonids is characterized by a multifactorial aetiology. Epitheliocystis of the gill lamellae caused by obligate intracellular bacteria of the order Chlamydiales is one known factor; however, their diversity has greatly complicated analyses to establish a causal relationship. In addition, tracing infections to a potential environmental source is currently impossible. In this study, we address these questions by investigating a wild brown trout ( Salmo trutta ) population from seven different sites within a Swiss river system. One age class of fish was followed over 18 months. Epitheliocystis occurred in a site‐specific pattern, associated with peak water temperatures during summer months. No evidence of a persistent infection was found within the brown trout population, implying an as yet unknown environmental source. For the first time, we detected ‘ Candidatus Piscichlamydia salmonis’ and ‘ Candidatus Clavochlamydia salmonicola’ infections in the same salmonid population, including dual infections within the same fish. These organisms are strongly implicated in gill disease of caged Atlantic salmon in Norway and Ireland. The absence of aquaculture production within this river system and the distance from the sea, suggests a freshwater origin for both these bacteria and offers new possibilities to explore their ecology free from aquaculture influences.