Comparison of Mycoplasma agalactiae isolates by pulsed field gel electrophoresis, SDS-PAGE and immunoblotting

Tola, Sebastiana; Idini, G.; Manunta, Daniela; Casciano, Ida; Rocchigiani, Angela Maria; Angioi, A.; Leori, Guido

doi:10.1111/j.1574-6968.1996.tb08490.x

Cited by 44 publications

(34 citation statements)

References 19 publications

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“…All strains examined had the same protein pattern (Fig. 1); this pattern was also similar to that previously described by Tola et al (26). Ninety-six sera obtained from naturally infected sheep from different areas of Sardinia were tested by immunoblotting; the antibody response was qualitatively homogeneous in all sera (data not shown).…”

Section: Resultssupporting

confidence: 58%

Mapping Antigenic Sites of an Immunodominant Surface Lipoprotein of Mycoplasma agalactiae , AvgC, with the Use of Synthetic Peptides

Santona¹,

Carta²,

Fraghí³

et al. 2002

Infect Immun

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As a first step toward the design of an epitope vaccine to prevent contagious agalactia, the strongly immunogenic 55-kDa protein of Mycoplasma agalactiae was studied and found to correspond to the AvgC protein encoded by the avgC gene. The avg genes of M. agalactiae, which encode four variable surface lipoproteins, display a significant homology to the vsp (variable membrane surface lipoproteins) genes of the bovine pathogen Mycoplasma bovis at their promoter region as well as their N-terminus-encoding regions. Some members of the Vsp family are known to be involved in cytoadhesion to host cells. In order to localize immunogenic peptides in the AvgC antigen, the protein sequence was submitted to epitope prediction analysis, and five sets of overlapping peptides, corresponding to five selected regions, were synthesized by Spot synthesis. Reactive peptides were selected by immunobinding assay with sera from infected sheep. The three most immunogenic epitopes were shown to be surface exposed by immunoprecipitation assays, and one of these was specifically recognized by all tested sera. Our study indicates that selected epitopes of the AvgC lipoprotein may be used to develop a peptide-based vaccine which is effective against M. agalactiae infection.

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Section: Resultssupporting

confidence: 58%

Mapping Antigenic Sites of an Immunodominant Surface Lipoprotein of Mycoplasma agalactiae , AvgC, with the Use of Synthetic Peptides

Santona¹,

Carta²,

Fraghí³

et al. 2002

Infect Immun

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show abstract

“…Name of isolates, years of isolation, countries of origin, and sequence types (MLST) are specified to the right of the branches. Isolates previously analyzed were added to the study (2). Black dots indicate the isolates analyzed in this study.…”

Section: Figmentioning

confidence: 99%

“…Sequence type 5 was found to be the ancestral genotype. European isolates previously analyzed were added to the study (2).…”

Section: Fig 3 Burst Analysis Of 104 Worldwide Mycoplasma Agalactiae mentioning

confidence: 99%

Molecular Characterization of Mycoplasma agalactiae Reveals the Presence of an Endemic Clone in Spain

2013

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Mycoplasma agalactiae isolates from Spain were genetically characterized to investigate their genomic diversity and to better understand their relationship to isolates from other countries. Molecular typing revealed a high genomic homogeneity in Spanish M. agalactiae isolates, which clearly shows the circulation of one endemic clonal population. Mycoplasma agalactiae is the main etiologic agent of contagious agalactia (CA), a serious syndrome affecting small ruminants; the World Organization for Animal Health must be notified of its occurrence because of its high economic significance worldwide. The first genomic studies showed little genomic diversity within M. agalactiae species, apart from that provided by antigenic variation (1, 2). Recently, the development of new sequence-based typing systems has revealed more genetic heterogeneity than previously thought (3-5). To investigate the genomic diversity of Spanish M. agalactiae isolates and to elucidate their relationship to those from other geographic areas, we analyzed isolates from Spain using pulsed-field gel electrophoresis (PFGE), which has been demonstrated to be robust and discriminative for typing different species of mycoplasmas (6-9), including M. agalactiae (3, 10); we also used the most recently developed sequence-based typing techniques, such as multilocus variable-

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“…Variability between M. agalactiae strains and isolates has been shown to exist at the protein level (5,44,49); however, little is known about the genetic systems that generate surface variation in this species. In a previous study to develop M. agalactiae-specific monoclonal antibodies (MAbs) for diagnostic purposes, four MAbs were shown to immunostain M. agalactiae colonies, revealing typical sectorial staining associated with the presence of variable epitopes (5).…”

mentioning

confidence: 99%

Characterization of a Multigene Family Undergoing High-Frequency DNA Rearrangements and Coding for Abundant Variable Surface Proteins in Mycoplasma agalactiae

et al. 2000

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A family of abundant surface proteins (Vpmas [variable proteins of Mycoplasma agalactiae]) undergoing phase variation in M. agalactiae has been characterized using monoclonal antibodies and specific polyclonal sera. Two expressed members of 39 kDa (Vpma39) and 34 kDa (Vpma34), which varied in expression between clones of a lineage, shared a common amino-terminal sequence but were immunologically distinct. An aminoterminal oligonucleotide probe identified multiple vpma genes which were clustered within a 14-kb ClaI genomic fragment. Rearrangements were found to have occurred within the vpma locus between clones which correlated with changes in their Vpma phenotype. Two neighboring vpma genes were cloned and sequenced from one M. agalactiae clonal variant expressing Vpma39. The two genes, vpmaX and vpmaY, were orientated divergently and shared highly homologous 5 untranslated regions, 25-amino-acid (aa) lipoprotein leader sequences, and amino-terminal sequences. The vpmaY gene coded for 346 aa and 84% of the open reading frame, comprised of 1.5 units of a large repeat of 186 aa. Although the sequence for an entire second vpmaY repeat was present, it was prematurely terminated by insertion of two nucleotides. The vpmaX gene encoded 221 aa and possessed 102 aa of the 186-aa repeat of vpmaY. Many of the features in common between the vpma genes were also found to be shared by the vsp genes of M. bovis, which also undergo DNA rearrangements concomitant with phenotypic changes. Since M. bovis is the closest phylogenetic relative to M. agalactiae, the vpma and vsp gene families most probably represent homologous systems.

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Comparison of Mycoplasma agalactiae isolates by pulsed field gel electrophoresis, SDS-PAGE and immunoblotting

Cited by 44 publications

References 19 publications

Mapping Antigenic Sites of an Immunodominant Surface Lipoprotein of Mycoplasma agalactiae , AvgC, with the Use of Synthetic Peptides

Mapping Antigenic Sites of an Immunodominant Surface Lipoprotein of Mycoplasma agalactiae , AvgC, with the Use of Synthetic Peptides

Molecular Characterization of Mycoplasma agalactiae Reveals the Presence of an Endemic Clone in Spain

Characterization of a Multigene Family Undergoing High-Frequency DNA Rearrangements and Coding for Abundant Variable Surface Proteins in Mycoplasma agalactiae

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