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Theronts from 2 different strains of Ichthyophthirius multifiliis (AR1 and AR5) were exposed to copper sulfate (CuSO 4 ) in waters of different total alkalinities and observed for 4 h to determine relative toxicity and kinetics of parasite mortality. Consistent with the known solubility properties of the metal, Cu was significantly more toxic to cells maintained under low (48 mg l ) total alkalinity conditions. This was reflected in both the median lethal concentration (LC 50 ) values and rates of mortality for both parasite strains; strain differences were also observed. The AR1 strain was significantly more resistant to copper toxicity than the AR5 strain in both high and low alkalinity waters. In general, these strain differences were more evident under conditions of low stress (i.e. low CuSO 4 concentration and high alkalinity), and suggest that genetic factors are overridden under high stress conditions. The present study establishes a role for alkalinity in the effectiveness of CuSO 4 treatment of ichthyophthiriasis and reveals differences in the susceptibility of parasite populations that are clearly important for control programs. KEY WORDS: Strain differences · Ichthyophthirius multifiliis · Copper toxicity Resale or republication not permitted without written consent of the publisherDis Aquat Org 83: [31][32][33][34][35][36] 2009 pounds at high alkalinities (Chakoumakos et al. 1979, Laurén & McDonald 1986. To account for this reduced toxicity or efficacy, the current practice for therapeutic use of CuSO 4 in culture ponds is to increase application rates in direct proportion to the total alkalinity of the water (MacMillan 1985, Tucker & Robinson 1990.It has become clear over the past decade that natural isolates of Ichthyophthirius multifiliis can be distinguished based on a number of criteria, most notably, the expression of surface immobilization antigens or i-antigens (Clark & Forney 2003). These antigens vary among natural isolates, and either monoclonal antibodies or reference polyclonal antisera that bind specific i-antigens can be used to define serotypes in simple immobilization assays (Dickerson et al. 1993, Swennes et al. 2006. Previous work has shown that serotype D strains are the most common in nature and express an abundant 55 kDa i-antigen on their surface (Wang et al. 2002). At least 5 additional serotypes of I. multifiliis have been identified based on antibodyspecific immobilization (Wang et al. 2002, Swennes et al. 2007, Xu et al. 2006 Swennes et al. (2007) has suggested that different strains of I. multifiliis vary with respect to virulence, and it would be reasonable to assume they differ in other phenotypic character traits as well.Previous research has investigated the effectiveness of CuSO 4 in controlling Ichthyophthirius multifiliis in several species of fish (Ling et al. 1993, Straus 1993, Schlenk et al. 1998, Tieman & Goodwin 2001, Goodwin & Straus 2006, Straus 2008, Rowland et al. 2009). The objective of the present study was to evaluate the acute toxicity of...

We are developing Tetrahymena thermophila as a delivery system for recombinant vaccines against parasitic protozoa, including the common fish parasite, Ichthyophthirius multifiliis. T. thermophila cell lines expressing I. multifiliis genes under the control of a cadmium-inducible metallothionein gene promoter conferred strong protection against a lethal parasite challenge when administered parenterally to naive fish. Nevertheless, given that heavy metals can be toxic to parasites, a question arose as to whether protection resulted from Cd residues carried over with the vaccine, rather than acquired immunity per se. To address this issue, we examined the sensitivity of I. multifiliis to Cd in vitro and determined Cd concentrations in different host tissues following i.p. injection of juvenile channel catfish with the recombinant vaccine. We found that CdCl2 at concentrations > or = 50 ppb were lethal to I. multifiliis theronts in vitro. Furthermore, Cd concentrations were clearly elevated in fish tissues and reached levels equivalent to 74 ng/g wet weight (74 ppb) in the skin within 14 days of injection with recombinant T. thermophila. Nevertheless, fish injected with non-transformed Tetrahymena grown in the presence or absence of CdCl2 showed no significant difference in either relative survival or parasite load following direct challenge with I. multifiliis.

Mitochondrial cox1 689 bp barcodes are routinely used for identification of Tetrahymena species. Here, we examine whether two shorter nuclear sequences, the 5.8S rRNA gene region and the intergenic region between H3 and H4 histone genes, might also be useful either singly or in combination with each other or cox1. We obtained sequences from ~300 wild isolates deposited at the Tetrahymena Stock Center and analyzed additional sequences obtained from GenBank. The 5.8S rRNA gene and portions of its transcribed flanks identify isolates as to their major clade and uniquely identify some, but not all, species. The ~330 bp H3/H4 intergenic region possesses low intraspecific variability and is unique for most species. However, it fails to distinguish between two pairs of common species and their rarer counterparts, and its use is complicated by the presence of duplicate genes in some species. The results show that while the cox1 sequence is the best single marker for Tetrahymena species identification, 5.8S rRNA, and the H3/H4 intergenic regions sequences are useful, singly or in combination, to confirm cox1 species assignments or as part of a preliminary survey of newly collected Tetrahymena. From our newly collected isolates, the results extend the biogeographical range of T. shanghaiensis and T. malaccensis and identify a new species, Tetrahymena arleneae n. sp. herein described.