Parkinson disease and dementia with Lewy bodies are featured with the formation of Lewy bodies composed mostly of α-synuclein (α-Syn) in the brain. Although evidence indicates that the large oligomeric or protofibril forms of α-Syn are neurotoxic agents, the detailed mechanisms of the toxic functions of the oligomers remain unclear. Here, we show that large α-Syn oligomers efficiently inhibit neuronal SNARE-mediated vesicle lipid mixing. Large α-Syn oligomers preferentially bind to the N-terminal domain of a vesicular SNARE protein, synaptobrevin-2, which blocks SNARE-mediated lipid mixing by preventing SNARE complex formation. In sharp contrast, the α-Syn monomer has a negligible effect on lipid mixing even with a 30-fold excess compared with the case of large α-Syn oligomers. Thus, the results suggest that large α-Syn oligomers function as inhibitors of dopamine release, which thus provides a clue, at the molecular level, to their neurotoxicity.

a major problem in dairy cows and beef cattle causing significant economic losses worldwide. Lesions are typically found at the volar skin proximal to the heel bulbs. Microscopic examination of biopsies or touch preparations of these lesions revealed a variety of different bacterial morphotypes including significant numbers of spirochetes which often represent the predominant morphotype. We used comparative 16s rRNA sequence analysis to determine the diversity and phylogeny of these hitherto unclassified DD spirochetes. Results indicate that those lesions looked at so far contained at least five spirochetal phylotypes, all clustering within the genus Treponema. Phylotype DDKL-4 was nearly identical (99.4% similarity) to that of a nonpathogenic human treponeme, T. phagedenis. Two phylotypes DDKL-3 and DDKL-13 were closely related to those from treponemes commonly found in human periodontitis lesions, i.e., T. denticoh and T. vincentii, exhibiting 95 and 98% similarity, respectively. The other two phylotypes, DDKL-12 and DDKL-20, had no close relatives to any cultivable treponemal species but clustered to previously described group IV oral treponemes. Preliminary analysis using in situ hybridization with fluorescently labeled oligonucleotide probes against smears from DD biopsies revealed that from all lesions analyzed so far, T. denticoh-like spirochetes were detected in the highest proportion of all spirochetal morphotypes.Digital dermatitis (DD), first described by Cheli and Mortellaro (6), is an acute or chronic ulcerative epidermitis in cattle that most commonly affects the skin immediately above the coronet between the heel bulbs (4). It is characterized clinically by an erosion of the superficial layers of the epidermis due to the loss of keratin, epithelial hyperplasia and hypertrophy, pain and swelling at the diseased sites, and a typical foul odor. Early lesions often show granulomatous strawberry-like ulcerations, whereas older lesions exhibit a grayish-brown color. DD usually leads to lameness and to a significant decrease in body weight and milk production (4). The disease is found with an incidence varying from 5 to 60% and a prevalence of 2 to 30%, rising to 80% in some herds and causing substantial economic loss in cattle dairies and the meat industry worldwide (38).Microscopic analysis of specimens taken from DD lesions revealed a variety of different bacterial morphotypes, including gram-negative rods and spirochetes. Spirochetes are often found in great numbers not only in superficial lesions but also in deeper layers of the dermis (5, 11,25). The etiology of DD is not yet established. However, the presence of high numbers of bacteria, including spirochetes, which apparently invade deeper tissues as well as successful antibiotic therapy strongly suggests a role of bacteria in the etioloLy of the disease (25). So far only a few organisms belonging to the genera Prevotella, Porphyromonas, Fusobacterium, and Bacteroides have been isolated (23). A recent paper reported the isolation of two groups of...

To determine the genetic diversity of cultivable and uncultivable spirochetes in the gingival crevice of a patient with severe periodontitis, partial 16S rRNA genes were cloned from PCR-amplified products of DNA and RNA extracted from a subgingival plaque sample. Approximately 500 bp were amplified in PCRs by using universally conserved primers with polylinker tails. Purified PCR products were cloned into Escherichia coli by using the plasmid vector pUC19. The resultant clone library was screened by colony hybridization with a radiolabeled, treponeme-specific oligonucleotide probe. The 16S rRNA inserts of 81 spirochetal clones were then sequenced by standard procedures. Sequences were compared with 16S rRNA sequences of 35 spirochetes, including the four known cultivable oral treponeme species. The analysis revealed an unexpected diversity of oral treponemes from a single patient. When 98% or greater sequence similarity was used as the definition of a species-level cluster, the clone sequences were found to represent 23 species. When 92% similarity was used as the definition, the clones fell into eight major groups, only two of which contained named species, Treponema vincentii and Treponema denticola, while Treponema pectinovorum and Treponema socranskii were not represented in any cluster. Seven of the 81 spirochetal clones were found to contain chimeric 16S rRNA sequences. In situ fluorescence hybridization with a fluorescein isothiocyanate-labeled oligonucleotide probe specific for one of the new species representing cluster 19 was used to identify cells of the target species directly in clinical samples.