Conservation in the first internal transcribed spacer region (ITS1) in Hematodinium species infecting crustacean hosts found in the UK and Newfoundland

Abstract

Parasitic dinoflagellates in the genus Hematodinium infect a number of decapod crustaceans in waters off the UK, including the Norway lobster Nephrops norvegicus and the edible crab Cancer pagurus. This study investigated sequence variability in the first internal transcribed spacer (ITS1) region of the ribosomal RNA complex of Hematodinium spp. infecting N. norvegicus, C. pagurus, and Pagurus bernhardus from 4 locations in the UK and from the Hematodinium sp. infecting Chionoecetes opilio from the province of Newfoundland and Labrador, Canada. Phylogenetic analysis of the Hematodinium ITS1 sequences from N. norvegicus, C. pagurus, P. bernhardus and C. opilio suggest that these crustaceans are infected with the same species of Hematodinium. Length variability of the ITS1 region was observed (324 to 345 bp) and attributed to 4 variable microsatellite regions (CATG) n , (GCC) n TCCGC(TG) n , (TA) n , and (GAA) n (GGA) n within the sequenced ITS1 fragment. The observed variation may be due to co-infection of the host crustacean with several different strains of Hematodinium or differences among copies of ITS1 region within the genome of a single parasite cell. The Hematodinium ITS1 sequence from N. norvegicus, C. pagurus, P. bernhardus and C. opilio isolates was sufficiently conserved in primer binding regions targeted by previous molecular diagnostic assays; therefore, we suggest that this assay could be used to screen for Hematodinium infections in these crustacean hosts. KEY WORDS: Hematodinium · Conservation of ITS1 · Crustacean hosts · Parasite Resale or republication not permitted without written consent of the publisherDis Aquat Org 75: [251][252][253][254][255][256][257][258] 2007 In waters surrounding the UK Hematodinium-like infections have been reported in Nephrops norvegicus from the Clyde and Irish seas (Field et al. 1992, Briggs & McAliskey 2002 and in Cancer pagurus from the English Channel (Stentiford et al. 2002) and from the Atlantic Ocean (Anonymous 2004). Both N. norvegicus and C. pagurus support large and valuable fisheries in UK waters; however, it is unknown whether different species of Hematodinium parasites infect each of these different hosts or whether one species infects multiple hosts.PCR-based diagnostics to detect Hematodinium spp. have been developed for detecting infections in several host species (Hudson & Adlard 1994, Gruebl et al. 2002, Small et al. 2006, 2007. In the first 2 studies the primer binding regions were in the 18S and 5.8S ribosomal RNA genes and are almost certainly genus specific, not species specific, due to the conserved nature of these genes. Small et al. (2006) developed a potentially species-specific set of PCR primers to target the Hematodinium sp. infecting Nephrops norvegicus using the conserved 18S rRNA gene and variable first internal transcribed spacer region (ITS) of the rRNA gene complex. This PCR assay also detected infections in Cancer pagurus from the English Channel tentatively suggesting that this is either the same species or a closel...

Molecular detection of Hematodinium sp. in Northeast Pacific Chionoecetes spp. and evidence of two species in the Northern Hemisphere

“…In parasitic species like Hematodinium, intra-and inter-individual variation may be confounded if hosts are infected by several genetically differentiated genotypes. Small et al (2007b) found variation only within repetitive regions in ITS1 within Hematodinium isolates from several host species. Because the amplified PCR products in this study were not cloned, it is uncertain whether one or several clones were present in each crab.…”

“…Dis Aquat Org 89: [155][156][157][158][159][160][161][162][163][164][165][166] 2010 including Portunus pelagicus, Norway lobsters Nephrops norvegicus and blue crabs Callinectes sapidus (reviewed in Stentiford & Shields 2005, Small et al 2006, 2007b. Despite the potential economic and ecological impact of Hematodinium-related mortality on affected populations, little is known about the method of infection, the life history of the parasite outside the host and the underlying causes for the recent worldwide increase in frequency of Hematodiniumassociated disease.…”

“…ex N. norvegicus is low, with only 9 differences in ITS1, each confined to single sequences, within 34 sequences (3 in the present study, and 31 in GenBank). As noted by Small et al (2007b), the sequence reported by Hudson & Adlard (1996) should be regarded with caution.The extreme difference in level of variability between similar 18S rDNA and non-alignable ITS1 sequences could be explained by either conservation of 18S rDNA or fast evolution of ITS1. There is little evidence to suggest that 18S is more conserved in dinoflagellates than in other groups; in fact, 18S is often used to define morphologically similar species of parasitic, symbiotic and free-living dinoflagellates (e.g.…”

Hematodinium sp. (Alveolata, Syndinea) detected in marine decapod crustaceans from waters of Denmark and Greenland

“…Different Hematodinium genotypes were related to different host species (Small et al 2007b, Hamilton et al 2010, Jensen et al 2010) and geographical regions (Hamilton et al 2010). It is also clear, however, that a single Hematodinium genotype may infect more than one host species (Hamilton et al 2007, Small et al 2007a, Jensen et al 2010. However, so far only one other species of Hematodinium has been formally described, namely H. australis, which was described on the basis of morphology, host species, and austral location (Hudson & Shields 1994).…”

Laser-assisted microdissection: a new tool for aquatic molecular parasitology

“…ITS1 has previously been used to differentiate between closely related Hematodinium spp. infecting a number of hosts (using primers HITS1F and HITS1R also used in the present study, see Small et al 2007a) or confirm a single infective species (Small et al 2007b); if required, it would be possible to use LMD and targeted PCR to amplify further phylogenetically informative regions. This application could be repeated for numerous other aquatic parasites where the diagnostic PCR primers amplify a small fragment.…”