Molecular Characterization of Novel Cryptosporidium Fish Genotypes in Edible Marine Fish

Abstract

Current knowledge of Cryptosporidium species/genotypes in marine fish is limited. Following phylogenetic analysis at the 18S rDNA locus, a recent study identified six new genotypes of Cryptosporidium colonizing edible fish found in European seas. Of these, five grouped in a clade together (#Cryptofish 1-5) and one grouped separately (#Cryptofish 7). In the present study, after phylogenetic analyses of #Cryptofish1, #Cryptofish2, #Cryptofish4, #Cryptofish5 and #Cryptofish7 at the actin locus, the presence of two major clades was confirmed. In addition, when possible, longer 18S amplicons were generated. In conclusion, the small genetic distances between these genotypes designated as a novel marine genotype I (#Cryptofish 1-5) suggest that they may be genetic variants of the same species, while the designated novel marine genotype 2 (#Cryptofish 7) is clearly representative of a separate species.

Keywords: 18S rDNA gene; actin gene; edible marine fish; genetic characterization; molecular phylogeny; piscine Cryptosporidium.

Figure 1

Phylogenetic relationships between #Cryptofish1, 2, 4, 5 and 7 genotypes (identified in the present study) and other piscine Cryptosporidium species inferred by ML analysis of actin gene. Percentage support (>50%) from 1000 pseudoreplicates from ML analyses is indicated at the left of the supported node. Scale bars indicate the number of substitutions per nucleotide position.

Figure 2

Stained sections of the intestinal tract of one Scomber scombrus (Ss VII 4BO) infected by the genotype Cryptofish7 (A) Presence of round bodies suggestive of the developmental stages of Cryptosporidium spp. observed in the apical position of a (stained with H&E) (B) Detail of A (delimited area). (C) Structure labelled in red with the Sporoglo (Waterborne) antibody and blue with DAPI (4′,6-diamidino-2-phenylindole) suggestive of an intracellular stage of Cryptosporidium spp.