Roger M. Cole

Mycoplasma genitalium, a New Species from the Human Urogenital Tract

Two mycoplasmas recovered from human urogenital tracts were similar in their biochemical and serological properties. These organisms possessed a unique terminal structure that appeared to be associated with attachment to tissue cells and erythrocytes. The organisms fermented glucose but did not hydrolyze urea or arginine. Growth occurred at 30 to 37°. Cholesterol was required for growth. Unlike most other mycoplasmas, both strains were susceptible to thallium acetate. These two organisms were serologically distinct from other Mycoplasma species and from a group of unclassified serotypes of mycoplasmas. On the basis of these findings and other morphological, biological, and serological properties of the microorganisms, we propose that mycoplasmas with these characteristics belong to a new species, Mycoplasma genitalium. Strain G-37 (= ATCC 33530) is the type strain.

ULTRASTRUCTURE OF THE AGENT OF CITRUS “STUBBORN” DISEASE

This ultrastructural presentation supplements the characterization, in the preceding paper,’ of a new mycoplasma-like organism cultured from “stubborn” disease of orange trees. Although mycoplasma-like bodies have been frequently seen by electron microscopy of thin sections of diseased plant tissues and of insect vectors, as has been amply documented and reviewed elsewhere in this anna1,’-Q only the stubborn-associated agent has been serially cultured in artificial media10-13 and examined morphologically and ultrastructurally from cultures. A more detailed description is published elsewhere: l4 it and the present report show that the microorganism is a helical filament like the corn stunt agent.l5J6 It is also motile, possesses an outer layer of surface projections, and is further unique among wall-less microorganisms by its propagation of a tailed bacteriophage.

Human infection from an unidentified erythrocyte-associated bacterium.

A 49-year-old splenectomized man had an infection from an unidentified, gram-positive, rod-shaped bacterium that adhered to the majority of his peripheral-blood erythrocytes. On transmission electron microscopy, the bacterium was seen to be extra-erythrocytic and was 0.2 micrometer wide by 1.0 to 1.7 micrometer long. It possessed a thick, granular cell wall, a trilamellar membrane external to the cell wall and prominent mesosomes. Attempts to cultivate the organism in vitro or to duplicate the patients disease in splenectomized animals were unsuccessful. The patients response suggested that the bacterium was susceptible to cell-wall-active antibiotics and to chloramphenicol but not to tetracycline. This bacterium may be the cause of other chronic, fever-producing, multisystem diseases of unknown origin.

Transduction in group A streptococcus.

Abstract Transduction, a phage mediated genetic transfer, has been reported to occur in several species of bacteria ( Hartman and Goodgal, 1959 ; Thorne, 1962 ; Allen et , al. , 1963 ). This report presents evidence for the transduction to streptomycin resistance of Group A streptococcus by several phages. To our knowledge transduction has not been reported previously in Group A streptococcus.

Biotinylation of human C3

Purified human C3 was biotinylated using the biotinyl-N-hydroxysuccinimide imidoester (BNHS). Depending on the input of BNHS, from three to six molecules of biotin were incorporated per C3 molecule. The biotinyl-C3 retained over 90% of its specific hemolytic activity and when bound to sheep erythrocytes maintained its ability to adhere to human C3b receptors. These functions could be blocked by avidin. The biotinyl-C3 was fragmented normally to C3c and C3d in human serum and adsorption with avidin-Sepharose indicated that biotin moities were present in both fragments. Fluorescein-conjugated avidin reacted well with cell-bound biotinyl-C3b and was useful for quantitating C3 fixation by flow cytometry. Ferritin-conjugated avidin was used as a marker to characterize the distribution of biotinyl-C3b on erythrocytes by electron microscopy. These results suggest that biotinyl-C3 and avidin derivatives may be very useful tools for studies of many of the biological functions of C3.

Spiroplasmavirus group 1: Isolation, growth, and properties

A group S1 spiroplasmavirus, SV1/KC3·BC3, from spiroplasma strain KC3, was propagated on spiroplasma strain BC3 and purified by density gradients. Purified virus had a density of 1.21 g/cm3 in metrizamide and 1.39 g/cm3 in CsCl and contained DNA of undetermined size and conformation. The virus was stable to exposure to non-ionic detergents, pHs between 6 and 9, heating at 60°C, drying, and (partially) ether; but was sensitive to pH extremes, chloroform, and 100°C. One-step growth experiments showed, by plaque assay, virus release beginning at 60 min and continuing for at least 6 h with no decrease in host cell growth. Infection was thus nonlytic, but the occurrence of 2-mm turbid plaques on agar indicates either belated host cell death or decreased division rate. The virus differed in some of these respects from the morphologically similar group 1 acholeplasmaviruses.

A fibrinogen precipitating factor (FPF) of group A streptococci.

Summary 1) An active factor was found in streptococci which precipitates various mammalian plasmas, and human and bovine fibrinogens. Fibrinogen precipitating factor (FPF) was found in crude acid extracts of several strains of group A. and one strain of group G streptococci; extracts of staphylococci, pneumococci, and viridans streptococci have proven non-reactive. 2) Streptococcal M protein and FPF appear inseparable, although presence of hyaluronic acid in a suspension of streptococcal cells prior to acid extraction may inhibit the fibrinogen precipitating reactivity of the extract, without significantly affecting the typing reaction. FPF and staphylocoagulase share certain properties, but they are not analogous.

Electron microscopy of solubilized Acholeplasma laidlawii membrane proteins reaggregated with Mycoplasma pneumoniae glycolipids

Abstract 1. 1. The purified glycolipid haptens of Mycoplasma pneumoniae were reaggregated with Acholeplasma laidlawii membrane proteins. The process consisted of the solubilization of lipid-depleted A. laidlawii membranes and M. pneumoniae glycolipids in 20 mM sodium dodecyl sulfate, and dialysis of the solution separately or in mixtures against 20 mM Mg2+. 2. 2. The reaggregated material collected by centrifugation of the dialyzed solution of lipid-depleted A. laidlawii membrane proteins consisted of amorphous clumps, while the reaggregated M. pneumoniae glycolipids consisted of “myelin-like” globules and sheets composed of lamellae with a mean center-to-center distance of 37 A. The reaggregated material of a mixture of lipid-depleted A. laidlawii membrane proteins and M. pneumoniae glycolipids contained, in addition to the amorphous clumps representing reaggregated proteins and the “myelin-like” structures representing reaggregated glycolipids, also long membranous sheets having a triple-layered structure with a mean center-to-center distance of the dense lines of 54 A. The appearance and dimensions are closely similar to those of the original or reagregated A. laidlawii membranes. 3. 3. It is suggested that these membrane-like structures are formed by the association of A. laidlawii membrane protein and M. pneumoniae glycolipids, and that these “hybrid” structures are responsible for the increased antigenicity of the reaggregated glycolipids.

The separation and isolation of plasma membranes and mesosomal vesicles from Staphylococcus aureus.

Abstract We have separated and Isolated the plasma membranes and mesosomal vesicles of Staphylococcus aureus ATCC 6538P. Cells were grown aerobically in Difco synthetic AOAC broth, washed and resuspended in hypertonic buffer (3.45 M NaC1) containing 0.02 M MgSO4. Cell wall was removed by treatment with lytic enzyme from S. aureus, strain LS. The protoplasts were collected by centrifugation at 10,000 × g for 1 hour, resuspended in hypotonic buffer containing 0.02 M MgSO4 and lysed. The resultant plasma membranes were washed and centrifuged on a 60tr>75Z sucrose density gradient at 55,000 × g for 15 hours. Gradient patterns showed two bands of membranes. Crude mesosomes were obtained from the 10,000 × g supernatant fractions by centrifugation at 100,000 × g for 2 hours. The reddish-brown gelatinous pellet, which consisted of mesosomal vesicles and a few ribosomes, was washed and centrifuged on a 60 to 85% sucrose density gradient at 100,000 × g for 15 hours. Gradient patterns produced two bands of mesosomal...

Simultaneous occurrence of two immunological types of group A "Coxsackie" virus in a case of herpangina.

Summary Two viruses occurring simultaneously were isolated from a case of herpangina. They were identified as the H1 and H2 immunologic types of Group A Coxsackie virus.