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Isolation and characterisation of a mycoplasma from a kittiwake (Rissa tridactyla)

MOLLICUTES are very small prokaryotes, characterised by the absence of a cell wall. They are widely distributed as pathogens or commensal organisms of animals. Pathogenic and non-pathogenic mycoplasma flora have been described in domestic birds (chicken, turkey, duck and goose), gamebirds (pheasant, partridge, quail and pigeon) and ratites, and some mycoplasma species are important causes of economic losses in poultry production. Furthermore, numerous mycoplasma isolates have been recovered from a variety of wild birds including black and griffon vultures, buzzards, falcons, sparrows, starlings, finches, blue jays and northern mockingbirds (Peterman and others 1989, Bradbury 1998). Some of them have been elevated to the rank of species, but a number are still uncharacterised (Ley and others 1998). In July 1994, a kittiwake (Rissa tridactyla) chick cadaver was discovered close to the Sizun Cape in Finistere (France). No apparent lesion could be seen. Tracheal and cloacal swabs were inoculated in Vero and MRC5 cells, and in one-day-old suckling mice for virus isolation; no viral strain was isolated. The same specimens were also inoculated in SP-4 medium (Tully and others 1977) and incubated at 37°C. Organisms that showed growth characteristics consistent with mycoplasmas (fried egg-like appearance colonies) were isolated from the tracheal swab only; they were filter-cloned three times in SP-4 media. Biochemical tests were performed according to methods by Nougayrede and others (1985). The cloned strain catabolised glucose, reduced tetrazolium, did not hydrolyse arginine, nor produce phosphatase activity. Growth inhibition tests, first performed against sera specific for pathogenic avian Mycoplasma species including Mycoplasma gallisepticum, Mycoplasma synoviae, Mycoplasma meleagridis and Mycoplasma iowae, gave negative results. In order to further characterise the strain, the sequence of its 16S rRNA gene was determined. The Genbank/EMBL accession number for the rDNA sequence encoding the 16S rRNA from the mycoplasma isolate was AF150495, and the sequence was then compared with previously published sequences from other Mycoplasma species. A phylogenetic analysis indicated that the sequence from the kittiwake isolate belonged to the M synoviae cluster within the hominis phylogenetic group (Pettersson and others 1994, 1996). In addition to Msynoviae, this cluster includes two other Mycoplasma species: Mycoplasma bovirhinis and Mycoplasmafelis. When compared with all the available sequences from this subgroup, Mycoplasmapullorum was the closest species, but the number of nucleotide differences (51 differences over 1440 nucleotides) excluded the gull isolate. This agreed with the negative growth inhibition result forM pullorum, although a more sensitive test such as immunofluorescence or immunoperoxidase could be carried out for further confirmation. Moreover, the Mpullorum reference strain does not reduce tetrazolium whereas the kittiwake isolate did. To evaluate the pathogenicity of the strain, 60 seven-dayold specific pathogen-free chicken embryonated eggs were inoculated via the yolk sac with different dilutions of the iso-