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Mycoplasma Adleri Sp. Nov., an Isolate from a Goat

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Mycoplasma Adleri Sp. Nov., an Isolate from a Goat INTERNATIONALJOURNAL OF SYSTEMATICBACTERIOLOGY, Jan. 1995, p. 29-31 Vol. 45, No. 1 0020-7713/95/$04.00+0 Copyright 0 1995, International Union of Microbiological Societies Mycoplasma adleri sp. nov., an Isolate from a Goat RICHARD A. DEL GIUDICE,l* DAVID L. AND JOSEPH G. TULLY2 Mycoplasma Laboratory, PRIlDynCorp, Frederick Cancer Research and Development Center, National Cancer Institute, Frederick, Maryland 21 702, and Mycoplasma Section, Laboratory of Molecular Microbiology, National Institute of Allergy and Infectious Diseases, Frederick, Maryland 21 7022 Mycoplasma sp. strain G145iT (T = type strain) was isolated from a goat’s abscessed ankle. Strain G145T required cholesterol or serum for growth and possessed characteristics similar to those of other members of the genus Mycophsma. This strain was serologically distinct from previously described Mycuphsma species and from a group of currently unnamed strains thought to belong to the genus Mycophsma. Strain G145T hydrolyzed arginine, but did not hydrolyze urea or ferment glucose. The guanine-plus-cytosinecontent of the DNA was 29.6 mol%. We propose that strain 6145 (= ATCC 27948) is the type strain of a new species, for which we propose the name Mycophsma adlen’. Mycoplasma strain G145T (T = type strain) was isolated erythrocytes to colonies of strain G145T was also assessed by a technique from a goat’s open ankle joint abscess during a suppurative described previously (13). Sterol requirement. The sterol growth requirement of strain G145T was arthritis outbreak in a Maryland goat herd in 1967. In prelim- determined by a standard liquid culture technique (17, 19). inary studies, strain G145T was characterized as a new Myco- Serologic tests. We prepared hyperimmune rabbit antisera and fluorescein plasma species, and it was found to be serologically distinct conjugates to Mycoplasma species and to a collection of unclassified strains by from the previously recognized species in the genus. We using conventional techniques that have been described previously (20). All sera distributed cultures of this strain to other investigators, and it and conjugates were maintained in the Mycoplasma Section of the National Institute of Allergy and Infectious Diseases. Some equine and caprine antisera was subsequently included in various comparative studies of and fluorescent antibody conjugates were prepared as part of the Reference caprine mycoplasmas (2,6-8, lo), in which it was also found to Reagent Program of the National Institute of Allergy and Infectious Diseases be unrelated to previously described Mycoplasma species. (9). Antisera and conjugates of the following mycoplasmas were used in serologic Additional serologic comparisons of strain G145T with other tests: Mycoplasma agalactiae PG2T,M. alkalescens D12T, M. alvi IlsleyT, M. anatis newly isolated mycoplasmas have occurred over the interven- 1340T, M. anseris 1219T, M. arginini G230T, M. arthritidis PG6T, M. auris UIAT, M. bovigenitalium PG1 lT, M. bovirhinis PG43T, M. bovis DonettaT, M. bovoculi ing 27 years without any suggestion that this organism is M165/69T, M. buccale CH20247T, M. buteonis Bb/T2gT, M. califomicum ST-6T, related to any other established species. M. canadense 275CT, M. canis PG14T, M. capricolum subsp. capricolum Califor- In this paper we designate strain G145 the type strain of a nia KidT, M. capricolum subsp. capnpneumoniae F38T, M. caviae G122T, M. new Mycoplasma species, describe additional characteristics of caviphaiyngis 117CT, M. citelli RG-2CT, M. cloacale 383T, M. collis 58BT, M. columbinasale 694T, M. columbinum MMP-lT, M. columborale MMP-4T, M. strain G145, and summarize the serologic relationship between conjunctivae HRC581T, M. corogypsi BVIT, M. cottewii VET, M. cricetuli CHT, this organism and previously described species belonging to M. cynos H831T, M. dispar 462/2T, M. edwardii PG24T, M. equigenitalium T37T, the genus Mycoplasma. M. equirhinis M432/72T, M. falconis H/TIT, M. fastidiosum 4822T, M. faucium DC333T, M. felifaucium Pv,M. feliminutum BenT, M. felis COT, M. fermentans PG18T, M. jlocculare M~42~,M. gallinaceum DDT, M. gallinarum PG16T, M. MATERIALS AND METHODS gallisepticum PG31T, M. gallopavonis WRIT, M. gateae CST, M. genitalium G37T, M. glycophilum 486T, M. gvpis BlRl, M. hominis PG21T, M. hyophaiyngis Mycoplasmas, culture medium, and cultivation techniques. Strain G145iT was H3-6BFT, M. hyopneumoniae JT, M. hyorhinis BTS7T, M. hyoqnoviae S16T, M. isolated on Edward medium (12), a formulation consisting of heart infusion imitans 4229T, M. indiense 3TT, M. iners PG30T, M. iowae 695T, M. leocaptivus broth, fresh yeast extract, and 20% horse serum. For solid growth medium, 0.8% 3L2T, M. Ieophaiyngis LL2T, M. Iipofaciens R171T, M. lipophilum MaByT, M. (wt/vol) agarose was added to Edward medium; the inoculated media were maculosum PGIST, M. meleagridis 17529T,M. moatsii l~fK405~,M. mobile 163KT, incubated at 35°C under an anaerobic atmosphere (GasPak system; BBL M. molare H542T, M. muris RII14T, M. mustelae MX9T, M. mycoides subsp. Microbiology Systems, Cockeysville, Md.). The strain was purified by filtration mycodes B3, M. mycoides subsp. Capri PG3T, M. neurolyticum Type AT, M. cloning (18). Early-passage cultures were grown on liquid medium without opalescens MH5408T, M. orale CH19299T,M. ovipneumoniae Y98T, M. oxoniensis penicillin or other antibiotics for at least five consecutive passages; each 128T, M. penetrans GTU54T, M. phocacerebrale 1049T, M. phocarhinis 852T, M. subculture was plated onto blood agar and incubated at 35°C. After 2 to 4 days, phocidae 105T, M. pirum 70-159T, M. pneumoniae FHT, M. primatum HRC292T, the plates were examined for bacterial colonies. M. pullorum CKKT, M. pulmonis PG34T, M. putrefaciens KS-lT, M. salivarium Morphological studies. Liquid cultures were examined by dark-field micros- PG20T, M, simbae LXT, M. spermatophilum AH159T, M. spumans PG13T, M. copy. Cellular morphology was also assessed after Gram staining and observation sualvi Mayfield BT, M. subdolum TBT, M. synoviae WW 1853T, M. testudinis by light microscopy. The ultrastructure of the cells was also determined by 01008T, M. yeatsii GIHT, M. verecundum 107T,Mycobacterium sp. strain Califor- electron microscopy, as described previously (5). nia calf (joint strain), Mycoplasma sp. strain B5P (bovine joint strain), Myco- Filtration studies. The filterability of strain G145T was assessed by using 24- to plasma sp. strain M7806 (cat oropharynx strain), Mycoplasma sp. strain 3306 48-h-old liquid cultures (18). A series of cellulose acetate membrane filters (ovine genital strain), Mycoplasma sp. strain HRC291 (primate throat strain), (Millipore Corp., Bedford, Mass.) with average pore sizes rated at 800,450,300, Mycoplasma sp. strain 3446 (bovine fetus strain), Mycoplasma sp. strain B689 and 220 nm were used. Serial 10-fold dilutions of filtrates and control liquid (dog throat strain), Mycoplasma sp. strain GM257A (caprine ear strain), and cultures were inoculated onto agar medium, and the resulting preparations were Mycoplasma sp. strain Utah C (tortoise nasal strain). The M. agalactiae PG2T, M. incubated at 35°C to determine the viable titers of the filtrates, which were anatis 1340T,M. arginini G230T,M. arthritidis PG6T,M. bovigenitalium PG1lT, M. expressed as CFU per milliliter. bovirhinis PG43T, M. buccale CH20247T, M. canis PG14T, M. faucium DC333T, Tests for biological and biochemical properties. The procedures used to M. fermentans PG18T, M. gallinarum PG16T, M. gallisepticum PG31T, M. hominis determine glucose fermentation, arginine hydrolysis, urea hydrolysis, phos- PG21T, M. hyorhinis BT57T, M. iners PG30T, M. lipophilum MaByT, M. maculo- phatase activity, tetrazolium reduction, serum liquefaction, and the film and spot sum PGIST, M. meleagridis 17529T, M. neurolyticum Type AT, M. orale reaction have been described previously (1, 3). Hemadsorption of guinea pig CH19299T, M. pneumoniae FHT, M. pulmonis PG34T, M. salivarium PG20T, and M. spumans PG13T reagents were National Institutes of Health reference reagents; all other reagents were prepared by the Mycoplasma Section of the Laboratory of Molecular Microbiology at the National Institute of Allergy and * Corresponding author. Mailing address: PRI/DynCorp, NCI- Infectious Diseases. Agar colonies of strain G14ST were tested by both the disc FCRDC, P.O. Box B, Frederick, MD 21702. Phone: (301) 846-1385. growth inhibition technique (4) and the plate immunofluorescence procedure t Present address: 8602 Cinnamon Creek, San Antonio, TX 78284. (11, 14), using antisera and fluorescein-conjugated antisera, respectively, to the 29 30 DEL GIUDICE ET AL. INT.J. SYST.BACTERIOL. FIG. 1. Electron micrograph of a thin section of a strain G145T cell pellet, showing the cellular morphology and the three-layer cytoplasmic membrane (arrowheads). Magnification, X 100,000. Bar = 100 nm. species listed above. One minor modification was made; strain G145T agar was 4.6 X lo6 CFU/ml. The titers of filtrates after passage colonies were examined only in an immunofluorescence test with conjugated through filters having pore sizes of 800, 450, 300, and 220 nm antiserum to M. agalactiae PG2T, since the unconjugated antiserum to M. agalactiae exhibited low homologous potency in growth inhibition tests. were 3.89 x lo6, 1.76 X lo5,8.9 X lo4, and 4.1 X lo4 CFU/ml, remectivelv. Biochemical and biological properties. Strain G145T hydro- RESULTS AND DISCUSSION Morphology and cultural properties. Liquid cultures of strain G145T contained numerous pleomorphic,
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