1 NSW Agriculture, Elizabeth Macarthur Agricultural Institute, Private Mail Bag 8, Camden, NSW, Australia 2570
2 Institute for Veterinary Bacteriology, University of Berne, CH-3012 Berne, Switzerland
Correspondence
Steven P. Djordjevic
steve.djordjevic{at}agric.nsw.gov.au
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ABSTRACT |
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Abbreviations: LC, large colony type; SC, small colony type
The GenBank/EMBL accession number for the 7902 bp region of the PG50 chromosome encoding GtsA, GtsB, GtsC, LppB, ORF5 and ORF6 is AJ419906.
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INTRODUCTION |
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Mycoplasma sp. bovine group 7, Mycoplasma mycoides subsp. mycoides small colony type (SC), M. mycoides subsp. mycoides large colony type (LC), M. mycoides subsp. capri, Mycoplasma capricolum subsp. capricolum and M. capricolum subsp. capripneumoniae represent the six recognized members of the M. mycoides cluster, and species identification within this cluster has been problematic. Electrophoretic, immunoblotting and DNA hybridization studies confirm the close phylogenetic relationship between members of the cluster (Bonnet et al., 1993; Christiansen & Ernø, 1990
; Costas et al., 1987
; Olsson et al., 1990
; Rodwell, 1982
; Rodwell & Rodwell, 1978
) and serological cross-reactions between Mycoplasma sp. bovine group 7 and M. capricolum subsp. capripneumoniae (Guerin et al., 1993
; Kibe et al., 1985
; Thiaucourt et al., 1994
), and M. capricolum subsp. capricolum (Bölske et al., 1988
) have been reported. Genotyping studies using the insertion elements IS1296 and IS1634, and serological studies using antisera raised against a major surface lipoprotein (LppB) revealed the presence of two distinct clonal lineages within M. mycoides subsp. mycoides SC; a highly virulent African/Australian cluster and the moderately pathogenic cluster of strains from the re-emerging European outbreaks of contagious bovine pleuropneumonia (Cheng et al., 1995
; Frey et al., 1995
; Vilei et al., 1999
, 2000
). Detailed genetic studies comparing isolates within each of these clusters has provided evidence that the less pathogenic European isolates arose by deletion of an 8·84 kb chromosomal region, present only among isolates belonging to the African/Australian cluster. This region carries a copy of IS1634, encodes proteins for glycerol transport (GtsA, GtsB and GtsC) and a major surface lipoprotein (LppB), and also contains two ORFs encoding a putative surface lipoprotein (ORF6) and a proline-rich membrane protein (ORF5) (Vilei et al., 2000
). African/Australian strains of M. mycoides subsp. mycoides SC are capable of importing and phosphorylating glycerol (Vilei & Frey, 2001
). Glycerol 3-phosphate is presumably oxidized to dihydroxyacetone phosphate and glyceraldehyde 3-phosphate with the concomitant production of H2O2, a potent haemolysing metabolite. Deletion of gtsC and part of gtsB (two of three genes in the putative operon involved in glycerol transport) in the moderately virulent European isolate L2 is correlated with a reduced ability to produce H2O2 and haemolyse sheep erythrocytes compared to the highly virulent Afadé strain, representative of the African/Australian cluster (Houshaymi et al., 1997
; Rice et al., 2000
; Vilei & Frey, 2001
). Collectively, genes located within this region are important for virulence and have also been targets in studies to differentiate between members of the M. mycoides cluster.
It has been reported that isolates of Mycoplasma sp. bovine group 7 representative of the clonal cluster recovered from multiple animals and different tissue sites during an outbreak lysed sheep red blood cells when cultured on blood agar medium (Hum et al., 2000). The aim of this study was to identify the putative genes for glycerol transport (gtsABC), lppB, and the flanking segment containing ORF5 and ORF6 on the chromosome of PG50, and compare them with homologues in M. mycoides subsp. mycoides SC. The capacity of PG50 to transport glycerol and produce H2O2 when cultured in the presence of glycerol was also investigated.
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METHODS |
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Cloning and DNA sequence analyses.
A 5·9 kb fragment amplified from template DNA of Mycoplasma sp. bovine group 7 type strain PG50 with primers 7500bp1L and 3480bp-R was ligated into the pGEM-T vector (Promega). Ligation products were used to transform competent Escherichia coli XL-1 Blue cells and transformants were selected on LB agar containing ampicillin (100 µg ml-1), X-Gal and IPTG (30 mg l-1). Recombinant plasmids recovered from white colonies were screened for the presence of a 5·9 kb insert by digestion with NcoI and NotI. All manipulations were performed according to standard procedures (Sambrook et al., 1989). One construct, pSDW12, was isolated from a bacterial pellet using a Qiagen maxiprep column and used as template for sequencing. Sequencing was performed with a DNA sequenator AB310 and the Taq DyeDeoxy Terminator Cycle Kit (both from Applied Biosystems) using primers complementary to the SP6 and T7 promoters of the vector and primers derived from sequenced segments. Sequence coverage was extended to 7902 bp by primer walking on genomic DNA of strain PG50. The DNA and deduced amino acid sequences were analysed with the ScanProsite software (http://ca.expasy.org/tools/scanprosite/), SignalP (http://www.cbs.dtu.dk/services/SignalP-2·0/) and the TMpred software (http://www.isrec.isb-sib.ch/ftp-server/tmpred/www/TMPRED_form.html). Sequence comparisons with sequences in the GenBank and EMBL databases were made using the BLAST programs BLASTN, BLASTX and BLASTP (Altschul et al., 1990
).
Southern hybridization.
Southern blots were performed essentially as described previously (Vilei & Frey, 2001). Genomic DNA was digested with HindIII, separated electrophoretically on a 0·7 % agarose gel and transferred to positively charged nylon membrane (Roche Diagnostics). The specific DNA probes were prepared by PCR amplification of genomic DNA, adding 0·51 µl digoxigenin-11-dUTP (Roche Diagnostics) to the reaction mix.
Immunoblotting.
Bovine serum collected from cattle experimentally infected with M. mycoides subsp. mycoides SC was prepared as described previously (Abdo et al., 1998). Cell lysates of type strains representative each of the six species in the M. mycoides cluster and Mycoplasma putrefaciens type strain KS1 were separated by SDS-PAGE and blotted onto PVDF membrane. The conditions and reagents used to detect the immunoreactive proteins on the membrane have been previously described (Abdo et al., 1998
).
Production of hydrogen peroxide.
Measurements of time-dependent H2O2 production in Mycoplasma sp. bovine group 7 (PG50) and M. mycoides subsp. mycoides SC (Afadé) were carried out using the Merckoquant peroxidase test (Merck KgaA) as described previously (Vilei & Frey, 2001). Briefly, 30 ml mycoplasma cultures of PG50 and Afadé, grown to a density of approximately 108 cells ml-1, were centrifuged at 12 000 g for 10 min and the cell pellets were washed and resuspended in 10 ml isotonic HEPES buffer containing 7 mM MgCl2. For each strain, two cell suspensions each of 1·0 ml were adjusted to OD550 1·0. After starvation for 1 h at 37 °C, the isotonic buffer of one sample was adjusted to 100 µM glycerol while the buffer of the second sample was deprived of glycerol. At time intervals ranging from 5 s to 20 min the test strips were dipped for 1 s and subsequently read as suggested by the manufacturer.
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RESULTS |
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gtsA encoded a peptide of 406 aa with a predicted molecular mass of 47·4 kDa and was almost identical to the sequence for GtsA of M. mycoides subsp. mycoides SC strain Afadé (Vilei & Frey, 2001); only 15 of 406 (96·3 % identity) aa were different. It was preceded by a canonical ribosome binding sequence 16 bp upstream (nt 127133) of the ATG start codon and contained three mycoplasma-specific TGATrp codons. Although TMpred software failed to identify a significant (score more than 500) transmembrane domain, ScanProsite software identified an ATP/GTP binding site motif A (P-loop) between aa 48 and 55, and an ABC transporter family signature with an ATP binding protein motif between aa 213 and 227. GtsA sequences from PG50 and strain Afadé showed that the P-loop sequence GPSGSGKT and the ABC transporter signature LSGGQKQRVAFAKGI were identical in both species. A BLASTP analysis using GtsA from Mycoplasma sp. bovine group 7 identified, in addition to the high identity with GtsA from M. mycoides subsp. mycoides SC, 33·3 % identity and 52·6 % similar amino acids with an ABC transporter ATP-binding protein from Mycoplasma pulmonis (MyPu_4970).
gtsB encoded a 342 aa peptide with a predicted molecular mass of 39·9 kDa and varied by 13 aa (96·2 % identity) from the GtsB in M. mycoides subsp. mycoides SC strain Afadé. gtsC encoded a 267 aa peptide with a predicted molecular mass of 31·5 kDa and displayed 89·2 % identity (29 amino acid variations and two deletions) with GtsC in M. mycoides subsp. mycoides SC strain Afadé. Although database searches failed to identify ABC transporter peptide signatures, TMpred software identified six transmembrane domains in both GtsB and GtsC. BLASTP searches also demonstrated that GtsB had 33·2 % identical and 56·6 % similar amino acids to an ABC transporter permease protein in M. pulmonis (MyPu_4980). GtsC has 32·5 % identical and 54·8 % similar amino acids to another hypothetical ABC transporter permease protein in M. pulmonis (MyPu_4990).
Sequence comparison between lppB, ORF5 and ORF6 genes downstream of the glycerol uptake locus from PG50 and the Afadé strain of M. mycoides subsp. mycoides SC
An ORF (nt 32985202) encoding a peptide of 634 aa (predicted molecular mass of 71·0 kDa) showing 66·4 % aa identity (80·3 % similarity) with LppB of M. mycoides subsp. mycoides SC was identified immediately downstream of the glycerol uptake operon (Fig. 1). LppB sequences from M. mycoides subsp. mycoides SC and M. mycoides subsp. mycoides LC had 90·1 % identity (Vilei et al., 2000
). The lppB homologue was preceded by a consensus sequence for a -10 box (nt 32523258) and was followed by a sequence (nt 52245250) that could form a hairpin structure with a
G of -13·0 kcal mol-1, representing a potential transcriptional termination signal. A canonical ribosomal binding sequence preceded the lppB gene by 8 nt (nt 32833290) and the gene contained eight potential TGATrp codons. SignalP identified a prokaryotic signal peptide cleavage site spanning aa 2324 of LppB.
One hundred and eighty one nucleotides downstream of lppB was a sequence spanning 2309 nt (nt 53837691) containing two ORFs, termed ORF6 and ORF5 (Fig. 1). ORF6 (nt 55216789) was preceded by a consensus sequence for a -10 box (nt 53835388). ORF5 spanned nt 67627652 and overlapped ORF6 by 28 bp. No -10 box was identified for ORF5. A sequence that could form a hairpin structure with a
G of -16·4 kcal mol-1 spanned nt 76607691. ORF6 encoded a putative membrane protein of 422 aa (predicted molecular mass of 49·2 kDa) and was preceded by a canonical ribosomal binding sequence spanning nt 55065510. The first 217 aa of ORF6 showed 88·5 % sequence identity (90·3 % sequence similarity) with the carboxy-terminal 217 aa portion of ORF6 of M. mycoides subsp. mycoides SC, but 299 aa were missing from the N terminus compared with ORF6 of M. mycoides subsp. mycoides SC. ORF5 encoded a proline-rich peptide of 296 aa (predicted molecular mass of 32·9 kDa) which showed significant similarity to ORF5 in M. mycoides subsp. mycoides SC (86·3 % sequence identity and 88·6 % similarity). The Mycoplasma sp. bovine group 7 homologue was missing the N-terminal 199 aa. Proline constituted 14·5 % of the Mycoplasma sp. bovine group 7 ORF5 sequence.
PCR analysis of the gtsABC and lppB loci within members of the M. mycoides cluster
Primers 7500bp1L and 3480bp-R amplified different fragment sizes from genomic DNA of different members of the M. mycoides cluster (Table 1). A 5·9 kb fragment was amplified from Mycoplasma sp. bovine group 7 isolates recovered from geographically diverse regions of the world. A 9·3 kb fragment was amplified from M. mycoides subsp. mycoides SC strains derived from the African/Australian cluster and a 0·45 kb fragment from isolates derived from the less virulent European cluster, as expected (Vilei et al., 2000
). An amplification product of 6·9 kb was observed when DNA from type strain Y-goat and field isolates of M. mycoides subsp. mycoides LC was used. Furthermore, a 7·5 kb fragment was amplified from PG3 and field isolates of M. mycoides subsp. capri (Table 1
). However, no amplification products were observed using DNA from type strains representative of the phylogenetically related species M. capricolum subsp. capripneumoniae (F38) and M. capricolum subsp. capricolum (California kid) or from the serogroup L isolate (B144P). No amplification products were obtained using DNA from phylogenetically more distantly related Mycoplasma spp., including Mycoplasma bovis, Mycoplasma agalactiae and M. putrefaciens (Table 1
).
Genetic analysis of the gtsABC and lppB loci
Variation in amplicon size demonstrated by long range PCR suggested that the chromosomal region spanning the glycerol uptake locus, lppB and the flanking segment including ORF5 and ORF6 may be prone to modification by insertion and/or deletion of gene sequences or by accumulation of sequence polymorphisms in PCR primer sites. Southern blot hybridization experiments using a suite of probes spanning regions within gtsABC, lppB, ORF5 and ORF6 derived from M. mycoides subsp. mycoides SC strain Afadé were used to determine the genetic composition of fragments amplified by long range PCR. Southern blots of HindIII-digested PG50 and field isolate 99/0361/4 of Mycoplasma sp. bovine group 7 identified fragments of 1·9 and 1·5 kb using a probe spanning a portion of gtsA. A minor RFLP (fragment sizes of 1·9 and 1·4 kb) was observed in strain PG50(54) and field isolate 99/8407/10 (Fig. 2); Afadé showed the characteristic fragments of 1·8 and 1·5 kb, whereas the European M. mycoides subsp. mycoides SC strain L2 showed bands of 3·4 kb and 1·5 kb. A probe spanning gtsB and gtsC hybridized to three fragments of 5·0, 2·5 and 1·9 kb on blots containing HindIII digests of Mycoplasma sp. bovine group 7 DNA from all four strains (Fig. 2
); fragments of 3·8, 1·8 and 1·0 kb were observed on blots containing HindIII digests of Afadé DNA, whereas L2 DNA presented a band at 3·8 kb. The presence of the 3·8 kb band in M. mycoides subsp. mycoides SC and of the 2·5 kb band in Mycoplasma sp. bovine group 7 cannot be explained by the sequence of this genetic locus. However, since the probe for gtsBC spanned most of gtsB and all of gtsC (Fig. 1
) it is conceivable that a portion(s) of this probe may share homology with domains characteristic of other ABC transporter genes known to be common in mycoplasma genomes (Chambaud et al., 2001
; Fraser et al., 1995
; Himmelreich et al., 1996
) and could explain the presence of the unpredicted hybridization signal. An lppB gene probe identified a single 5·0 kb HindIII fragment on blots of genomic DNA from all four Mycoplasma sp. bovine group 7 strains (Fig. 2
); fragments of 3·9 and 1·0 kb were evident on blots containing HindIII-digested Afadé DNA. A single 5·0 kb fragment of DNA from all four Mycoplasma sp. bovine group 7 strains hybridized with separate probes spanning regions of ORF5 and ORF6 (Fig. 2
); a single fragment of 4·4 kb hybridized with these probes in HindIII-digested Afadé DNA (Fig. 2
). As expected, probes for lppB, ORF5 and ORF6 did not react with the L2 DNA. These data showed that the genomic region encompassing genes for the glycerol uptake operon, lppB, ORF5 and ORF6 was quite stable among geographically diverse strains of Mycoplasma sp. bovine group 7.
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DISCUSSION |
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Although the gtsABC locus of M. sp. bovine group 7 displays strong sequence identity with homologous genes in Mycoplasma mycoides subsp. mycoides SC strain Afadé, the regions immediately downstream of these loci differ significantly. In PG50, IS1634 and IS1296 are missing and there are truncations of large portions of the 5'-terminal halves of both ORF5 (proline-rich protein) and ORF6 (putative surface located membrane protein) compared with homologous genes in strain Afadé. However, both PG50 and strain Afadé possess the lipoprotein gene lppB. PCR and Southern blot analyses of other members of the M. mycoides cluster, including M. mycoides subsp. mycoides LC, M. mycoides subsp. capri, M. capricolum subsp. capricolum and M. capricolum subsp. capripneumoniae revealed the latter two to be most different from Mycoplasma sp. bovine group 7. Hybridization of PG50 DNA with probes spanning gtsABC, ORF5, ORF6 and lppB derived from M. mycoides subsp. mycoides SC showed that these genes were present in Mycoplasma sp. bovine group 7, while M. capricolum subsp. capricolum (California kid) DNA only yielded positive signals with probes spanning gtsB, gtsC and lppB; no signal was observed with probes spanning gtsA and ORF5, and only a very weak signal was observed with ORF6. Furthermore, immunoblotting data showed that PG50 possessed a panel of cross-reactive antigens that most closely resembled the pattern generated with lysates of M. mycoides subsp. mycoides SC strains when reacted with anti-strain Afadé serum raised during an experimental infection. The least similar patterns of cross-reactive antigens were observed with type strains California kid and KS1 of M. putrefaciens.
Salih and others examined 24 mycoplasma strains representing all six members of the M. mycoides cluster for the presence of 35 enzymes by horizontal starch gel electrophoresis. These authors concluded that Mycoplasma sp. bovine group 7 constituted a new species and was most similar to the M. mycoides subsp. mycoides SC type strain PG1 (Salih et al., 1983). Recent evidence that Mycoplasma sp. bovine group 7 is closely related to M. mycoides subsp. mycoides SC has also emerged from studies of surface accessible lipoproteins present among members of the M. mycoides cluster. The lppA gene has been characterized by DNA sequence analysis in M. mycoides subsp. mycoides SC and LC, M. mycoides subsp. capri and Mycoplasma sp. bovine group 7 (Monnerat et al., 1999
) and unlike lppB, is located at a site unrelated to the glycerol uptake locus. Comparisons of the predicted amino acid sequences of LppA from type strains of Mycoplasma sp. bovine group 7 and M. mycoides subsp. mycoides SC showed 91 % identity. However, comparisons of predicted LppA sequences of PG50 (Mycoplasma sp. bovine group 7) and California kid (M. capricolum subsp. capricolum) showed only 53 % identity (Frey et al., 1998
; Monnerat et al., 1999
). Moreover, antisera raised against recombinant LppA from M. capricolum subsp. capricolum (California kid) did not cross-react with LppA homologues in any other member of the M. mycoides cluster (Monnerat et al., 1999
). LppB sequences from M. mycoides subsp. mycoides SC and M. mycoides subsp. mycoides LC had 90·1 % identity (Vilei et al., 2000
), while LppB proteins of M. mycoides subsp. mycoides SC and Mycoplasma sp. bovine group 7 had 66·4 % sequence identity. Antisera raised against recombinant LppB from strain Afadé reacted strongly with LppB in PG50, PG1 (M. mycoides subsp. mycoides SC type strain) and Y-goat (M. mycoides subsp. mycoides LC type strain) but only weakly in Western blots of whole cell lysates of California kid, F38 (M. capricolum subsp. capripneumoniae type strain) and PG3 (M. mycoides subsp. capri type strain) (Vilei et al., 2000
).
In conclusion, our data suggest that PG50 and other strains of Mycoplasma sp. bovine group 7 possess a functional glycerol transport locus. Predicted amino acid sequences of GtsA, B and C, LppB, ORF6 and ORF5 from Mycoplasma sp. bovine group 7 showed strong sequence identity with M. mycoides subsp. mycoides SC but IS1634 and IS1296 were absent from this region in the PG50 chromosome. Southern hybridization and long range PCR studies indicated that this chromosomal region has diverged in mycoplasmas belonging to subspecies of M. capricolum.
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ACKNOWLEDGEMENTS |
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REFERENCES |
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Alexander, P. G., Slee, K. J., McOrist, S., Ireland, L. & Coloe, P. J. (1985). Mastitis in cows and polyarthritis and pneumonia in calves caused by Mycoplasma species bovine group 7. Aust Vet J 62, 135136.[Medline]
Altschul, S. F., Gish, W., Miller, W., Myers, E. W. & Lipman, D. J. (1990). Basic local alignment search tool. J Mol Biol 215, 403410.[CrossRef][Medline]
Bannerman, E. S. & Nicolet, J. (1971). Isolation and identification of porcine Mycoplasma in Switzerland. Schweiz Arch Tierheilkd 113, 697710.[Medline]
Bölske, G., Msami, H., Humlesjö, N. E., Ernø, H. & Jönsson, L. (1988). Mycoplasma capricolum in an outbreak of polyarthritis and pneumonia in goats. Acta Vet Scand 29, 331338.[Medline]
Bonnet, F., Saillard, C., Bové, J. M., Leach, R. H., Rose, D. L., Cottew, G. S. & Tully, J. G. (1993). DNA relatedness between field isolates of Mycoplasma F38-group, the agent of contagious caprine pleuropneumonia, and strains of Mycoplasma capricolum. Int J Syst Bacteriol 43, 597602.[Abstract]
Chambaud, I., Heilig, R., Ferris, S. & 9 other authors (2001). The complete genome sequence of the murine respiratory pathogen Mycoplasma pulmonis. Nucleic Acids Res 29, 21452153.
Cheng, X., Nicolet, J., Poumarat, F., Regalla, J., Thiaucourt, F. & Frey, J. (1995). Insertion element IS1296 in Mycoplasma mycoides subsp. mycoides small colony identifies a European clonal line distinct from the African and Australian strains. Microbiology 141, 32213228.[Abstract]
Christiansen, C. & Ernø, H. (1990). RFLP in rRNA genes of Mycoplasma capricolum, the caprine F38-like group and the bovine serogroup 7. Zentbl Bakteriol 20, 479488.
Connole, M. D., Laws, L. & Hart, R. K. (1967). Mastitis in cattle caused by a mycoplasma sp. Aust Vet J 43, 157162.[Medline]
Costas, M., Leach, R. H. & Mitchelmore, D. L. (1987). Numerical analysis of PAGE protein patterns and the taxonomic relationships within the Mycoplasma mycoides cluster. J Gen Microbiol 133, 33193329.[Medline]
Djordjevic, S. P., Forbes, W. A., Forbes-Faulkner, J., Kuhnert, P., Hum, S., Hornitzky, M. A., Vilei, E. M. & Frey, J. (2001). Genetic diversity among Mycoplasma species bovine group 7: clonal isolates from an outbreak of polyarthritis, mastitis, and abortion in dairy cattle. Electrophoresis 22, 35513561.[CrossRef][Medline]
Fraser, C. M., Gocayne, J. D., White, O. & 25 other authors (1995). The minimal gene complement of Mycoplasma genitalium. Science 270, 397403.[Abstract]
Frey, J., Cheng, X., Kuhnert, P. & Nicolet, J. (1995). Identification and characterisation of IS1296 in Mycoplasma mycoides subsp. mycoides SC and presence in related mycoplasmas. Gene 160, 95100.[CrossRef][Medline]
Frey, J., Cheng, X., Monnerat, M.-P., Abdo, E.-M., Krawinkler, M., Bölske, G. & Nicolet, J. (1998). Genetic and serological analysis of the immunogenic 67-kDa lipoprotein of Mycoplasma sp. bovine group 7. Res Microbiol 149, 5564.[CrossRef][Medline]
Gonçalves, R., Regalla, J., Nicolet, J., Frey, J., Nicholas, R., Bashiruddin, J., De Santis, P. & Gonçalves, A. P. (1998). Antigen heterogeneity among Mycoplasma mycoides subsp. mycoides SC isolates: discrimination of major surface proteins. Vet Microbiol 63, 1328.[CrossRef][Medline]
Guerin, C., Thiaucourt, F., Mady, V., Bréard, A. & Lefèvre, P. C. (1993). Rapid diagnosis of contagious caprine pleuropneumonia in pleural fluids by immunoblotting assay. Small Rumin Res 12, 193200.
Himmelreich, R., Hilbert, H., Plagens, H., Pirkl, E., Li, B. C. & Herrmann, R. (1996). Complete sequence analysis of the genome of the bacterium Mycoplasma pneumoniae. Nucleic Acids Res 24, 44204449.
Houshaymi, B. M., Miles, R. J. & Nicholas, R. A. (1997). Oxidation of glycerol differentiates African from European isolates of Mycoplasma mycoides subspecies mycoides SC (small colony). Vet Rec 140, 182183.[Medline]
Hum, S., Kessell, A., Djordjevic, S. P., Rheinberger, R., Horntizky, M., Forbes, W. & Gonsalves, J. (2000). Mastitis, polyarthritis and abortion caused by Mycoplasma species bovine group 7 in dairy cattle. Aust Vet J 78, 744750.[Medline]
Kibe, M. K., Bidwell, D. E., Turp, P. & Smith, G. R. (1985). Demonstration of cross-reactive antigens in F38 and related mycoplasmas by enzyme-linked immunosorbent assay (ELISA) and immunoblotting. J Hyg 95, 95106.
Monnerat, M.-P., Thiaucourt, F., Nicolet, J. & Frey, J. (1999). Comparative analysis of the lppA locus in Mycoplasma capricolum subsp. capricolum and Mycoplasma capricolum subsp. capripneumoniae. Vet Microbiol 69, 157172.[CrossRef][Medline]
Olsson, B., Bölske, G., Bergström, K. & Johansson, K.-E. (1990). Analysis of caprine mycoplasmas and mycoplasma infections in goats using two-dimensional electrophoresis and immunoblotting. Electrophoresis 11, 861869.[Medline]
Rice, P., Houshaymi, B. M., Nicholas, R. A. & Miles, R. J. (2000). A rapid biochemical test to aid identification of Mycoplasma mycoides subspecies mycoides small colony (SC) strains. Lett Appl Microbiol 30, 7074.[CrossRef][Medline]
Rodwell, A. W. (1982). The protein fingerprints of mycoplasmas. Rev Infect Dis 4, 817.
Rodwell, A. W. & Rodwell, E. S. (1978). Relationships between strains of Mycoplasma mycoides subsp. mycoides and capri studied by two-dimensional gel electrophoresis of cell proteins. J Gen Microbiol 109, 259263.
Salih, M. M., Ernø, H. & Simonsen, V. (1983). Electrophoretic analysis of isoenzymes of mycoplasma species. Acta Vet Scand 24, 1433.[Medline]
Sambrook, J., Fritsch, E. F. & Maniatis, T. (1989). Molecular Cloning: a Laboratory Manual, 2nd edn. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory.
Shiel, M. J., Coloe, P. J., Worotniuk, B. & Burgess, G. W. (1982). Polyarthritis in a calf associated with a group 7 mycoplasma infection. Aust Vet J 59, 192193.
Simmons, G. C. & Johnston, L. A. Y. (1963). Arthritis in calves caused by Mycoplasma sp. Aust Vet J 39, 1114.
Thiaucourt, F., Bölske, G., Libeau, G., LeGoff, C. & Lefèvre, P. C. (1994). The use of monoclonal antibodies in the diagnosis of contagious caprine pleuropneumonia (CCCP). Vet Microbiol 41, 191203.[Medline]
Vilei, E. M. & Frey, J. (2001). Genetic and biochemical characterization of glycerol uptake in Mycoplasma mycoides subsp. mycoides SC: its impact on H2O2 production and virulence. Clin Diagn Lab Immunol 8, 8592.
Vilei, E. M., Nicolet, J. & Frey, J. (1999). IS1634, a novel insertion element creating long, variable-length direct repeats which is specific for Mycoplasma mycoides subsp. mycoides small-colony type. J Bacteriol 181, 13191323.
Vilei, E. M., Abdo, E.-M., Nicolet, J., Botelho, A., Gonçalves, R. & Frey, J. (2000). Genomic and antigenic differences between the European and African/Australian clusters of Mycoplasma mycoides subsp. mycoides SC. Microbiology 146, 477486.
Received 17 May 2002;
revised 8 October 2002;
accepted 9 October 2002.
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