Three monoclonal antibodies (MAbs) were prepared against an arthritogenic strain of Mycoplasma hominis isolated from the joint aspirates of a patient with chronic septic arthritis. Immunoblots of polyacrylamide gel-electrophoresed proteins before and after surface proteolysis showed that the predominant antigenic determinants were on surface-exposed polypeptides. These polypeptides have extensive hydrophobic characteristics, as demonstrated by Triton X-114 phase partitioning. The electrophoresed proteins from cells grown in medium containing [14C]palmitate were blotted onto nitrocellulose which was both reacted with the MAbs and exposed to X-ray film. Superimposable bands on both the immunoblots and the exposed film suggested that the proteins might be acylated. The MAbs were further tested for reactivity with 16 other strains of M. hominis isolated from patients with septic arthritis (1 strain), septicemia (10 strains), or nongonococcal urethritis (1 strain); from the cervix (1 strain), rectum (1 strain), or surgical wound (1 strain) of patients; and from a contaminated cell culture. No single protein was consistently recognized from strain to strain, although a 94-kDa protein from 16 of the 17 strains tested was bound by at least one of the MAbs. The apparent antigenic heterogeneity among strains of M. hominis, including those isolated from the same tissue source and/or from patients with the same type of clinical disease, might be misleading in that all strains express epitopes associated with a discrete number of proteins to which one, two, or all three MAbs bind. The expression of the epitopes on multiple proteins from the same or different strains may reflect a mechanism for generating antigenic diversity.

The expression of surface proteins by 14 successive Mycoplasma hominis isolates obtained from the synovial fluid of a chronically infected septic arthritis patient was examined. Marked differences in the expression of surface proteins, as determined by monoclonal antibodies raised against the first isolate, were observed. However, identical restriction patterns and virtually identical hybridization patterns with probes containing the conserved genes of the Mycoplasma capricolum rRNA operon and the Escherichia coli elongation factor Tu suggest that the protein differences might reflect antigenic variation by M. hominis during infection.

The high concentrations of sulfated glycolipids in the human endometrium (6) and in mammalian male germ cells (10) suggest an advantage for organisms that can use these glycolipids as receptors. Lingwood and coworkers recently found that M. hominis binds to 2 ,ug of sulfogalactoglycerolipid and sulfogalactosylceramide, or sulfatide, on thin-layer plates (7). However, it was not demonstrated that this attachment is a specific adhesin-receptor interaction rather than some form of nonspecific, perhaps ionic, binding.To better understand the ability of M. hominis to colonize and cause disease, we screened a variety of glycoconjugates for the ability to serve as receptors for M. hominis adherence. We also examined the specificity and avidity of M. hominis attachment to sulfatide.All saccharides, polysaccharides, and glycoproteins were purchased from Sigma Chemical Company (St. Louis, Mo.), as were cholesterol, cholesterol sulfate, phosphatidylcholine, and bovine serum albumin (BSA). The glycolipids GM1, asialo-GM1, GD1a, GDib, GT1b, GM2, GM3, galactosylceramide, glucosylceramide, lactosylceramide, globoside, and sulfatide were also obtained from Sigma. Asialo-GM2, trihexosylceramide, and Forssman antigen were purchased from BioCarb Inc., Gaithersburg, Md.M. hominis 1620 (passage level 3 to 5) was used predominantly in this study. Strain 1620 was originally isolated from the synovial tissue of a chronically infected septic arthritis patient (13). For comparison where noted, reference strain PG21, a rectal isolate which has been passed in the labora-* Corresponding author. tory for over 30 years, was used in some studies, as was patient blood isolate strain H5488 at passage level 3. All cultures were grown and metabolically labeled with [3H]palmitic acid (12 to 17 Ci/mmol; Dupont-New England Nuclear) as previously described (2). After harvesting and two washes in 0.05 M Tris-HCl (pH 7.6)-110 mM sodium chloride (TBS), the organisms were suspended in 10 ml of buffer, passed four times through a 23-gauge needle, and further diluted with buffer to approximately 108/ml. Depending on the experiment, this dilution provided approximately 106 cpm/ml.Thin-layer chromatography (TLC) overlays were performed as a modification of the method described in reference 5. Briefly, glycolipids and lipids were separated on aluminum-backed silica gel high-performance plates (Merck, Darmstadt, Germany) with chloroform-methanol-water (60: 35:8). Following chromatography, the plates were blocked with 1% BSA in TBS (TBS-BSA) for 1 h. Plates were then incubated for 3 h at 370C with approximately 108 CFU of unlabeled M. hominis per ml in TBS-BSA. After 30 min of washing in TBS, the plates were incubated overnight at 40C with monoclonal antibodies (1:1,000 dilution) specific for surface-exposed, protease-sensitive, integral membrane proteins of M. hominis (11). After another wash in TBS-BSA, the plates were incubated for 1 h at room temperature with

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13C‐NMR studies on the effect of glucose metabolism on arginine hydrolysis in Mycoplasma fermentans cells have been performed using a continuous perfusion technique. With this procedure we were able to show, in the presence of glucose, the rapid accumulation of lactic acid and, in the presence of arginine, the formation of citrulline that is apparently further metabolized. As the accumulation of lactate and the breakdown of arginine were observed in the simultaneous presence of both substrates, it is suggested that the glucose utilization has little or no effect on the deimination of arginine to citrulline.