Three amoeba species were isolated from 3 out of 193 farmed tilapias, Oreochromis niloticus (L.), screened for the presence of free-living amoebae in parenchymatous organs. Hartmannella vermiformis Page, 1967 and Rosculus ithacus Hawes, 1963 were isolated from the kidney tissue. The third strain isolated from the liver shared morphological features of Mayorella and Platyamoeha spp. and therefore its taxonomic position has not been determined as yet. Pathogenicity of cloned strain of H. vermiformis was proved in two fish hosts.
Ninety four aquarium fishes were screened for the presence of amoebae in their internal organs. Five specimens of Ca-rassius auratus (L.) and one specimen of Xiphophorus hetleri Heckel were positive. Of the three strains which were isolated from C. auratus, successfully cloned and cultivated, one was identified as Vannella platypodia (Gläser, 1912) Page, 1976 and two strains as Rosculus ithacus Hawes, 1963. Both species are reported for the first time from organs of fish. None of them could be identified with the amoeba-like agent of goldfish granulomas described here.
Canningia spinidentis gen. et sp. n. infects the fir bark beetle Pityokteines spinidens Rtt. in Austria. The pathogen attacks mainly the fat body, Malpighian tubules, the muscles and the connective tissue of larvae and adults, and the gonads of adults. The development is haplokaryotic, with single spores. Spores are short tubular, uninucleate, with globular anchoring disc inserted subapically, laterally, in a depression of the endospore wall. Polar filament is isofilar, with 5/6 coils. Polaroplast is composed of two lamellar parts of different density. A new genus Canningia gen. n. is proposed based on differences in ultrastmc-tures of spores from Unikaryon Canning, Barker, Hammond et Nicholas, 1974.
Using degenerative primers designed on the basis of known sequences of lectin genes from different sources a fragment of genomic DNA of Borrelia burgdorferi ( strain B31) that contained a lectin-like sequence was isolated, cloned and sequenced. The presence of an open reading frame of 268 amino acids (position 1501-2304 bp) and the computer analysis of the predicted amino acid sequence showed 37% of identity and 75% of homology over region of 25 amino acids with the legume lectin proteins, including erythroagglutinating phytohcmagglutinin (РНЛ-Е) and leucoagglutinating phylohemagglutinin (PHA-L). The further analysis of the predicted amino acid sequence showed the presence of another two domains (positions 198-211 and 215-226 aa) consisting of the characteristic conserved sequence motifs for legume lectin proteins. Hemagglutinating activity was detected in lysate of В burgdorferi (strain B31) spirochete and the affinity to fetuin was determined in a hemagglutination inhibition test. Hemagglutinating activity was also found in a crude lysate of the recombinant clones carrying the fragment of B. burgdorferi genomic DNA. The inhibition of agglutinating activity by fetuin, D-galactosamine and D-mannosamine was determined using the standard procedure of hemagglutination inhibition test with native rabbit red blood cells (RBC).
When in vitro growth of Vittaforma comeae was tested using MDCK, MRC-5, XEN, L-929 and FHM cell lines, propagation occurred only in MDCK, MRC-5 and XEN cells. The intervals required for the various stages of the life cycle to develop were the same in all the cell lines tested. The MDCK cell line was selected to support the growth of V. comeae in vitro and provide the system for in vitro testing of drugs. The weekly output of V. comeae spores from the MDCK cell monolayer was monitored over a 61-week period during which there were fluctuations but no definite increase or decrease in output. Albendazole at 2.1 or 4.2 pg/ml in MEM was tested against V. comeae in MDCK cell monolayers and showed antimicrosporidial activity. The percentage of infected cells was reduced in the presence of the drug and there were ultrastmctural abnormalities in all stages of the life cycle. The drug prevents parasite division.
Investigations on the epizoic fauna of Gadus morhua (L.), Piatichthys flesus (L.) and Oncorhynchus mykiss (Walbaum) from the Kiel Fjord and Kiel Bight were carried out from September 1996 to March 1997. Smears from 120 G. morhua and 92 P. flesus caught using fish traps and trammel nets, and of 35 O. mykiss obtained from a local fish farm in the Kiel Fjord revealed the presence of three species of trichodinid ciliatcs, Trichodina claviformis sp. n., Trichodina jadranica Haider, 1964 and Trichodina raahei Loin, 1962. The new species can be distinguished from other trichodinids by its large size in combination with the characteristically shaped adhesive disc containing denticles with club-like formed thorns. The thorns are directed anteriorly and not towards the centre of the adhesive disc. As the Kiel Bight and Kiel Fjord are new locality records for T. jadranica and T. raabei, morphological data are provided for both species. Trichodina claviformis is the first record of a pcrilrichous mobiline ciliate from Atlantic cod of the Baltic Sea. An identification Icey for 16 Trichodina species occurring on Baltic Sea fishes is provided based on the morphology of the adhesive disc and other well-established features The occurrence of trichodinid ciliates on G. morhua and P. flesus in the Baltic Sea is discussed, especially considering the biology of the host and a possible host specificity of the species.
This paper summarizes work done in this laboratory over the last two years on the cloning of microsporidian rRNA by homology PCR and its subsequent use in diagnostic tests and phylogenetic studies. Using highly conserved primers in the 16S or small subunit rRNA (SSU-rRNA) these genes were cloned from human intestinal biopsies with transmission electron microscopy proven Enterocytozoon bieneusi and Septata intestinalis. The SSU-rRNA genes were then used to design and test several primer pairs for the diagnosis of microsporidian infection. Utilizing the polymerase chain reaction and primers V1 and EB45Ü Ent. bieneusi infected duodenal aspirates or intestinal biopsies could be detected. Using V I and SI500 infection with S. intestinalis could be detected. In addition to diagnostic tests, phylogenetic relationships were examined using sequence data from the fragment amplified by PCR by primer 530f in the SSU-rRNA and primer 580r in the large subunit rRNA. This data supported the placement of S. intestinalis in the family Encephalitozoonidae. In addition, it confirmed that Encephalitozoon cuniculi, E. hellem and S. intestinalis are distinct organisms. These techniques have broad applications to the study of other microsporidia and the development of a molecular phylogeny.