Béatrice M. Després

The Lobster Parasite Anophryoides haemophila (Scuticociliatida: Orchitophryidae): Nuclear 18S rDNA Sequence, Phylogeny and Detection Using Oligonucleotide Primers

ABSTRACT. We have determined the nucleotide sequence of the nuclear gene encoding small‐subunit ribosomal ribonucleic acid of the ciliate Anophryoides haemophila, a parasite of the American lobster Homarus americanus. The gene is 1763 bp in length, and has a guanosine‐plus‐cytosine content of 43.9%. Inferred phylogenetic frameworks strongly support the monophyly of the scuticociliates, and suggest that order Scuticociliatida should be elevated to at least subclass rank. Oligonucleotide probes based on A. haemophila ssu‐rDNA can discriminate between DNAs of A. haemophila and other investigated hymenostome ciliates, and effectively prime polymerase chain reaction‐based detection of A. haemophila deoxyribonucleic acid against at least a 1600‐fold excess of total deoxyribonucleic acid from H. americanus.

Microheterogeneity and Coevolution: An Examination of rDNA Sequence Characteristics in Neoparamoeba pemaquidensis and Its Prokinetoplastid Endosymbiont

ABSTRACT. Neoparamoeba pemaquidensis, the etiological agent of amoebic gill disease, has shown surprising sequence variability among different copies of the 18S ribosomal RNA gene within an isolate. This intra‐genomic microheterogeneity was confirmed and extended to an analysis of the internal transcribed spacer (ITS) region. High levels of intra‐genomic nucleotide diversity (Pi=0.0201–0.0313) were found among sequenced ITS regions from individual host amoeba isolates. In contrast, the ITS region of its endosymbiont revealed significantly lower levels of intra‐genomic nucleotide diversity (Pi=0.0028–0.0056) compared with the host N. pemaquidensis. Phylogenetic and ParaFit coevolution analyses involving N. pemaquidensis isolates and their respective endosymbionts confirmed a significant coevolutionary relationship between the two protists. The observation of non‐shared microheterogeneity and coevolution emphasizes the complexity of the interactions between N. pemaquidensis and its obligate endosymbiont.

Case Report: Outbreak of Bumper Car Disease Caused by Anophryoides haemophila in a Lobster Holding Facility in Nova Scotia, Canada

Abstract This case report provides pathologic and confirmatory molecular characterization of an outbreak of bumper car disease caused by the scuticociliate Anophryoides haemophila in American lobster Homarus americanus, from a commercial holding facility in Nova Scotia, Canada. Although sporadically and anecdotally reported to be present in Atlantic Canada, this is the first report detailing observations from a natural outbreak of bumper car disease by gross and microscopic pathology and its positive identification by small subunit rRNA gene sequencing. Randomly amplified polymorphic DNA analysis confirmed that the genotype of the present outbreak isolate is identical to the original isolate of A. haemophila reported from Maine lobsters in 1993.

Analysis of expressed sequence tags (ESTs) and gene expression changes under different growth conditions for the ciliate Anophryoides haemophila, the causative agent of bumper car disease in the American lobster (Homarus americanus)

The scuticociliate Anophryoides haemophila, causes bumper car disease in American lobster (Homarus americanus) in commercial holding facilities in Atlantic Canada. While the parasite has been recognized since the 1970s and much has been learned about its biology, minimal molecular characterization exists. With genome consortiums turning to model organisms like the ciliates Tetrahymena and Paramecium, the amount of relevant sequence data available has made sequence surveys more attractive for gene discovery in related ciliates. We sequenced 9984 expressed sequence tags (ESTs) from a non-normalized A. haemophila cDNA library to characterize gene expression patterns, functional gene distribution and to discover novel genes related to the parasitic life history. The A. haemophila ESTs were grouped into 843 clusters and singletons with 658 EST clusters having identifiable homologs, while 159 ESTs were unique and had no similarity to any sequences in the public databases. Not unexpectedly, about 67% of the A. haemophila ESTs have similarity to annotated and hypothetical genes from the related oligohymenophorean ciliate, Tetrahymena. Numerous cysteine proteases, hypothetical proteins and novel sequences possess putative secretory signal peptides suggesting that they may contribute to the pathogenesis of bumper car disease in lobster. Real time RT-qPCR analysis of cathepsin L and two homologs of cathepsin B did not show any changes in gene expression under varying in vitro growth conditions or during a modified-in vivo infection which may be suggestive of the opportunistic life history strategy of this ciliate.