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The morphology and ultrastructure of two strains of Borrelia anserina were investigated by electron microscopy of negatively stained and ultrathin sectioned cells. One was a cultivable strain originally isolated in the USA and the other was originally isolated in Nigeria and maintained in chickens. The cells were regularly helical, 9-21 microns long and 0.22-0.26 microns wide with a helix wavelength of about 1.7 microns. The cells were surrounded by a surface layer and appeared to divide by binary fission. The structure of the cells from each of the two strains was identical except that those of the USA strain possessed seven flagella inserted at each end and those from the Nigerian strain had eight.

An increasing number of phenotypically heterogeneous borrelia isolates have been obtained by culture. Genetic analysis of some of them suggest that a reorganisation in delineation of the different species within the genus Borrelia may be necessary. Specifically increasing numbers of variants phenotypically different from the American type strain of B. burgdorferi have been described. In this workshop experts have discussed phenotypic and genotypic characters for (a) differentiation of B. burgdorferi strains from other borrelia and from treponemes, (b) characterization of B. burgdorferi isolates at the subspecies level. Ultrastructure was discussed by K. Hovind-Hougen, antigen structure by B. Wilske and genotypic characters by G. Baranton, A. G. Barbour, R. C. Johnson and V. Preac-Mursic. J. F. Anderson discussed differences between strains isolated from a wide range of vectors and hosts in North America. The outcome of the workshop was that B. burgdorferi may comprise different genomic species which however share common epitopes recognized by certain monoclonal antibodies. At the subspecies level a broad heterogeneity was demonstrated using methods as restriction endonuclease analysis, hybridization with whole B. burgdorferi-DNA or specific probes as well as plasmid analysis. A serotyping system based on monoclonal antibody reactivity against the outer surface protein OspA was proposed. A comparison of the different typing methods is impaired by use of different strains for analysis. In the future a broad variety of phenotypically different and defined strains need to be analysed for genotypic clusters and later on reexamined for phenotypic characters. To accomplish this close cooperation of different research groups is necessary.

Previous studies on cells of strains B31 isolated in the U.S.A. from Ixodes dammini and strain G25 isolated in Sweden from Ixodes ricinus, showed that their ultrastructure was similar, but not identical. For this reason the studies were extended to spirochetes isolated directly from patients with Lyme disease and related disorders. Included in the present study were three strains isolated from skin, blood and spinal fluid, respectively, from patients with Lyme disease, two strains from patients with erythema chronicum migrans and one strain from a patient with acrodermatitis chronica atrophicans. Three additional strains isolated in Sweden from Ixodes ricinus were also studied. All spirochetes were examined after negative straining with 1% ammonium molybdate. The cells of each individual strain were identical except for one strain isolated from a tick. This isolate was found to consist of two morphologically different spirochetes. Comparison of morphological features of cells from various isolates revealed certain differences. The cells of the different strains could be divided into at least four groups for which cell size and shape as well as number of flagella varied. By morphological criteria, all cells were found to belong to the genus Borrelia.

In 1984, two human cases of tick-borne Lyme borreliosis with considerable neurologic involvement were reported in Berlin (West). The diagnosis of Lyme borreliosis was serologically confirmed. The ticks which had transmitted the Borrelia were from Berlin (West). In the autumn of 1985, 156 ticks were collected in forests of Berlin (West) for the cultural detection of spirochetes by using BSK II medium. Three strains of spirochetes were isolated (from a pooled sample of two nymphs, and samples of one nymph and one female tick, respectively). These isolates were identified as Borrelia burgdorferi by means of SDS-PAGE, Western blot (using monoclonal antibody H 5332), microscopic agglutination test and electron microscopy. Investigations with the electron microscope showed that cells of two isolates (strains 2/B45 and 3/B56) had 8 flagella inserted at each end. The cells of the third isolate (strain 1/B29) had 9 flagella inserted at each end. This type had not been observed before.

The ultrastructure of cells of Borrelia persica was compared with that of cells of a new isolate of Barrelia (Borrelia "x"). This organism (B. "x") was isolated in a region of Iran where B. persica is already known to exist. The morphology of the cells of the two strains differed in mean cell length and cell diameter. Cells of B. persica were covered by a regularly structured surface layer, while those of B. "x" possessed an amorphous surface coat. Furthermore, the strains differed in their experimental pathogenicity for animals. It is concluded that the new isolate represents a new species of the genus Borrelia, and Borrelia baltazardi sp. nov. is proposed.