Identification of 'Candidatus Piscichlamydia salmonis' in Arctic charr Salvelinus alpinus during a survey of charr production facilities in North America

Authors: DRAGHI, ANDREW - UNIVERSITY OF CONNECTICUT; Bebak, Julie; DANIELS, STEPHEN - UNIVERSITY OF CONNECTICUT; TULMAN, EDAN - University Of Connecticut; STEVEN, GEARY - University Of Connecticut; WEST, BRIAN - University Of Connecticut; POPOV, VSEVOLD - University Of Connecticut; FRASCA, SALVATORE - University Of Connecticut

Submitted to: Diseases of Aquatic Organisms
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 10/27/2009
Publication Date: 2/24/2010
Citation: Draghi, A., Bebak, J.A., Daniels, S., Tulman, E.R., Steven, G.J., West, B.A., Popov, V.L., Frasca, S. 2010. Identification of 'Candidatus Piscichlamydia salmonis' in Arctic charr Salvelinus alpinus during a survey of charr production facilities in North America. Diseases of Aquatic Organisms. 89:39-49.

Interpretive Summary: The gill disease epitheliocystis can limit production of Arctic charr, Salvelinus alpinus, and is associated with bacteria from the order Chlamydiales. Arctic charr production facilities and non-production water sources in the United States and Canada were sampled to determine whether chlamydia-like bacteria were present in water, and whether those chlamydiae were associated with epitheliocystis. Gill samples from 607 fish obtained from 13 sites were processed for histopathologic examination and DNA extraction. Water was collected from 21 locations for DNA isolation. Eighteen fish, from one location, had inclusions and pathology consistent with epitheliocystis. DNA sequencing and in situ hybridization confirmed the presence of Piscichlamydia salmonis in 12 out of 18 of these fish. Members of the family Chlamydiaceae were isolated from the water samples. This study demonstrates the feasibility of using DNA isolation, PCR and ISH to identify chlamydiae from production systems and resident fish population for comparison purposes. This is the first identification of Piscichlamydia salmonis associated with epitheliocystis in charr.

Technical Abstract: Artic charr Salvelinus alpinus production facilities, nonproduction water sources and effluents in the United States and Canada were sampled to determine if chlamydiae associated with epitheliocystis were present in water and were associated with inclusions of epitheliocystis in gill tissue. Gills from 607 fish from 13 sites were processed for histopathologic examination and DNA extraction. Water was collected from 21 locations for DNA testing. Eighteen fish from one location had inclusions of epitheliocystis with proliferative and inflammatory gill lesions. Inclusions were stained using the Gimenez technique and, at the ultrastructural level, consisted on intracytoplasmic membrane-bound vacuoles containing reticulate and intermediate bodies in a fibrillar matrix. PCR using Order Chlamydiales-specific primers performed on DNA extracts from 12 of 13 infected fish yielded amplicons that were identical to (GQ302988) or differed at one base from (GQ302987) the 16S ribosomal RNA gene signature sequence of 'Candidatus Piscichlamydia salmonis', which is the Chlamydia that was previously identified in epitheliocystis inclusions of farmed Atlantic salmon. In situ hybridization using a ~1.5 kb riboprobe corresponding to the 'Candidatus Piscichlamydia salmonis' 16S rRNA genetic sequence (AY462244) confirmed its presence within Artic charr gill inclusions. DNA isolated from water samples was tested by Chlamydiales-specific PCR and yielded 54 partial 16S rRNA genetic sequences spanning the signature region; however, no 16S rRNA genetic sequences associated with epitheliocystis were identified. This is the first report of 'Candidatus Piscichlamydia salmonis' associated with epitheliocystis in Artic charr, the first identification of 'Candidatus Piscichlamydia salmonis' from a freshwater production location, and the first reported occurrence in North America.