A repetitive element from the genome of Leptospira interrogans serovar hardjo type hardjo-bovis ('L. hardjo-bovis') was identified, cloned and sequenced. Similar sequences were shown by hybridization to be encoded by a further eight of 32 other leptospiral serovars tested. An undefined number of repetitive elements were located in the L. hardjo-bovis genome; sequence degeneracy of the elements was observed and no significant open reading frames were identified within the AT-rich (60%) 1467 bp repetitive element. The termini encoded a GC-rich 8 bp repeat motif and two variants showed rearrangements centred on these motifs. The nucleotide sequences of the chromosomal regions flanking the repetitive elements were determined but showed no similarities, with one exception which had a GAAC repeat directly adjacent to both termini. Similar hybridization patterns were shown by Southern transfers of L. hardjo-bovis total genomic digests probed with the repetitive element. Oligonucleotide primer pairs designed from sequences internal to the repetitive element and adjacent chromosomal regions were used in polymerase chain reaction experiments. With one primer pair all L. hurdjo-bovis isolates, but no other serovar, gave identical amplified products. Evidence that the repetitive element may have derived from an acquired insertion sequence that is now inactive and chromosomally fixed is discussed.

Phosphorothioate oligodeoxynucleotides belong to a class of polyanions that bind to the third variable domain (v3) of HIV-1 gp120 and inhibit infectivity of a wide variety of HIV-1 isolates. This potent v3 binding of phosphorothioate oligodeoxynucleotides, which is relatively independent of the nucleotide sequence of the oligodeoxynucleotides, decreases with chain length (below 18-mers) and is low for 8-mers. However, recent studies have observed a nucleotide sequence-dependent augmentation of phosphorothioate oligodeoxynucleotide binding to v3 for 8-mers that contain the S-dG4 motif (e.g., SdT2G4T2) and have suggested that formation of quadruple helical tetraplexes (G-tetrads) is associated with the acquisition of v3 binding ability by small phosphorothioate oligodeoxynucleotides. In the current study, a series of SdG4-containing oligodeoxynucleotides were synthesized with varying tandem length (including the 8-mer SdT2G4T2, the 12-mer SdG4T4G4, and the 28-mer SdG4(T4G4)3) and compared with phosphorothioate oligodeoxynucleotides (with similar lengths or related sequences) for (1) their inhibition of the binding of mAb 9284, which binds to the N-terminal portion of the v3 loop, (2) the values of Kc when these compounds are used as competitors of the rgp120-binding of an alkylating phosphodiester oligodeoxynucleotide probe, and (3) inhibition of HIV-1 infectivity in a cell-cell transmission model. The presence of S-dG4 motifs and the number of tandem motifs augmented v3 binding and anti-HIV-1 infectivity for small (8-mer or 12-mer oligodeoxynucleotides) but did not significantly augment the potency of 28-mers. Whereas tetraplex formation of SdT2G4T2 may contribute to its v3 binding, the 12-mer SdG4T4G4 does not migrate as the tetraplex on nonreducing gels, suggesting that S-dG4 motifs may augment anti-HIV activity by multiple mechanisms.

No abstract

Spleen lymphocytes from Lewis and Buffalo rats and peripheral blood lymphocytes from 10 human donors exhibited high levels of transformation when exposed to Mycoplasma arthritidis supernatants. In contrast, spleen lymphocytes from rabbits and guinea pigs and peripheral blood lymphocytes from sheep and calves failed to transform when exposed to M. arthritidis supernatants. The lymphocytes from all hosts were transformed in the presence of phytohemagglutinin or concanavalin A or both Serological studies failed to provide evidence that the responding hosts were presensitized against M. arthritidis antigens.

Syngeneic and allogeneic target cells were killed in the presence of CBA mouse lymphocytes and viable Mycoplasma arthritidis. Medium supplementation had no effect on the response. Nonviable M. arthritidis was also capable of stimulating lymphocytotoxicity, although to a much lesser extent. Cytotoxicity was shown to be largely dependent upon the lymphocytes, since lymphocytes preincubated with mycoplasmas and treated to remove remaining organisms were highly toxic to target cells, whereas supernatants prepared from lymphocyte/mycoplasma mixtures exhibited minimal effects. A 6-h exposure of lymphocytes to mycoplasmas at a ratio of 100:1 was sufficient for commitment to target cell killing. Functional lymphocytes were required for the reaction, since y-irradiated lymphocytes did not develop cytotoxic potential despite the fact that the mycoplasmas replicated equally well in the presence of these and untreated lymphocytes. Furthermore, lymphocytes already activated with mycoplasmas lost cytotoxic potential after disruption. The kinetics and degree of lymphocytotoxicity induced by M. arthritidis and phytohemagglutinin toward 61Cr-labeled syngeneic fibroblasts were similar. Removal of most B cells and other adherent cells by column separation did not abrogate the cytotoxic effect. Lymphocyte suspensions treated with anti-Thy 1 antiserum and complement exhibited a marked decrease in their cytotoxic potential when added to labeled target cells in the presence of M. arthritidis. We conclude that the cytotoxic reaction is dependent upon the Tlymphocyte subpopulation. Mycoplasma arthritidis is a potent arthritogenic agent causing spontaneous or experimental arthritis in rats, mice, and rabbits (11, 13, 14, 22). We recently demonstrated that M. arthritidis exerts a mitogenic effect toward normal unsensitized mouse lymphocytes (10). Many other mycoplasma species appear to exhibit similar mitogenic properties (4, 10, 17, 20). We have also shown that M. arthritidis can induce normal mouse lymphocytes to exert a cytotoxic action on allogeneic and syngeneic mouse fibroblast target cells as evidenced by release of a '1C label (1). These observations are therefore of great interest in view of the intense inflamatory response and mononuclear cell reaction seen in the joints of animals infected with this organism.