Identification, Bioinformatics Analyses, and Expression of Immunoreactive Antigens of Mycoplasma haemofelis

Messick, Joanne B.; Santos, Andrea Pires dos

doi:10.1128/cvi.05060-11

Cited by 4 publications

(8 citation statements)

References 34 publications

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“…The majority of these genes express surface proteins in interaction with the host environment. As few of these PGFs of M. suis and M. haemofelis have been found to encode immunogenic proteins [46] (Guimaraes et al, unpublished data), rapid amino acid changes are likely to be crucial for function and for evading the host immune defenses. Therefore, while host-dependency contributes to genome shrinkage through relaxation of positive selection, other selective pressures, e.g.…”

Section: Resultsmentioning

confidence: 99%

Comparative Genomics and Phylogenomics of Hemotrophic Mycoplasmas

et al. 2014

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Hemotrophic mycoplasmas (hemoplasmas) are a group of animal pathogens of the Mollicutes class. Recently, the genomes of 8 hemoplasmas have been completely sequenced. The aim of this study was to gain a better understanding of their genomic features and relationship to other Mycoplasma species. The genome structure and dynamics of hemoplasmas were analyzed by evaluating gene synteny, adaptive evolution of paralogous gene families (PGF) and horizontal gene transfer (HGT). The Mollicutes class was then phylogenetically analyzed by constructing a distance matrix of the 16S rRNA genes and a phylogenetic tree with 32 conserved, concatenated proteins. Our results suggest that the hemoplasmas have dynamic genomes. The genome size variation (from 547 to 1,545 genes) indicates substantial gene gain/loss throughout evolution. Poorly conserved gene syntenies among hemoplasmas, positional shuffling of paralogous genes between strains, HGT, and codons under positive selection in PGFs were also observed. When compared to other Mollicutes species, the hemoplasmas experienced further metabolic reduction, and the 16S rRNA gene distance matrix of the available mollicutes suggests that these organisms presently constitute the most divergent clade within its class. Our phylogenetic tree of concatenated proteins showed some differences when compared to the 16S rRNA gene tree, but non-mycoplasma organisms, such as Ureaplasma spp. and Mesoplasma spp., continue to branch within Mycoplasma clades. In conclusion, while the hemoplasmas experienced further metabolic shrinkage through gene loss, PGFs with positively selected codons are likely beneficial to these species. Phylogeny of the mollicutes based on 16S rRNA genes or concatenated proteins do not obey the current taxonomy. The metabolism and genetic diversity of the mollicutes, the presence of HGT, and lack of standard for genus circumscription are likely to hinder attempts to classify these organisms based on phylogenetic analyses.

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Section: Resultsmentioning

confidence: 99%

Comparative Genomics and Phylogenomics of Hemotrophic Mycoplasmas

et al. 2014

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“…A total of 17 putative lipoproteins were identified in the genome of M. haemofelis (Figure 1 ). In addition, some putative surface lipoproteins were recently identified by immune screening of an expression library of M. haemofelis [ 24 ].…”

Section: Resultsmentioning

confidence: 99%

“…Gross genome validation was achieved by comparing the virtual fingerprint patterns of M. haemofelis genomic DNA to that of fragments derived from pulse-field gel electrophoresis (PFGE) using the restriction enzymes, Nru I, Sal I, and Not I, and with physical map data from an independently derived bacterial artificial chromosome (BAC) library [ 22 ]. In addition, the sequencing of 21 inserts (0.2 to 4.5 kb) from two Lambda ZAPII libraries of M. haemofelis [ 24 ] was accomplished and compared to the M. haemofelis genomic sequence.…”

Section: Methodsmentioning

confidence: 99%

Genome of Mycoplasma haemofelis, unraveling its strategies for survival and persistence

Santos

Guimarães

Nascimento

et al. 2011

Vet Res

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Mycoplasma haemofelis is a mycoplasmal pathogen (hemoplasma) that attaches to the host's erythrocytes. Distributed worldwide, it has a significant impact on the health of cats causing acute disease and, despite treatment, establishing chronic infection. It might also have a role as a zoonotic agent, especially in immunocompromised patients. Whole genome sequencing and analyses of M. haemofelis strain Ohio2 was undertaken as a step toward understanding its survival and persistence. Metabolic pathways are reduced, relying on the host to supply many of the nutrients and metabolites needed for survival. M. haemofelis must import glucose for ATP generation and ribose derivates for RNA/DNA synthesis. Hypoxanthine, adenine, guanine, uracil and CMP are scavenged from the environment to support purine and pyrimidine synthesis. In addition, nicotinamide, amino acids and any vitamins needed for growth, must be acquired from its environment. The core proteome of M. haemofelis contains an abundance of paralogous gene families, corresponding to 70.6% of all the CDSs. This "paralog pool" is a rich source of different antigenic epitopes that can be varied to elude the host's immune system and establish chronic infection. M. haemofelis also appears to be capable of phase variation, which is particularly relevant to the cyclic bacteremia and persistence, characteristics of the infection in the cat. The data generated herein should be of great use for understanding the mechanisms of M. haemofelis infection. Further, it will provide new insights into its pathogenicity and clues needed to formulate media to support the in vitro cultivation of M. haemofelis.

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“…Finally, phagemids from positive plaques were isolated and sequenced for the identification of the insert as previously described. 17…”

Section: Mycoplasma Suis Dna Extraction and Expression Library Analysismentioning

confidence: 99%

Identification ofMycoplasma suisantigens and development of a multiplex microbead immunoassay

et al. 2014

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The aims of the current study were to identify Mycoplasma suis antigens and develop a multiplex microbead immunoassay (MIA). A M. suis-expression library was screened for immunogens using sera from infected pigs. Based on bioinformatics, putative antigens were identified within positive inserts; gene fragments were expressed and purified as polyhistidine fusion proteins, and immunoreactivity was confirmed by Western blot. Selected antigens were used to develop a MIA. Sera from noninfected and infected pigs were used to set the median fluorescent intensity (MFI) cutoffs and as positive controls, respectively. Assay specificity was tested using sera from pigs seropositive for other pathogens (2 different pigs seropositive for each pathogen). Samples from 51 field pigs and 2 pigs during the course of acute (pig 1) and chronic (pig 2) infections were tested using MIA, indirect hemagglutination assay (IHA), and quantitative polymerase chain reaction (qPCR). Sixteen reactive plaques (52 genes) were detected. A heat-shock protein (GrpE), a nicotinamide adenine dinucleotide-dependent glyceraldehyde 3-phosphate dehydrogenase (GAPN), and 4 proteins from paralogous gene families (PGFs) were identified as antigens by Western blot. While GrpE, GAPN, and 1 PGF protein were strong antigens, the others were not suitable as MIA targets. A MIA using GrpE, GAPN, and the strongly reactive PGF protein was developed. Cross-reactivity with sera from pigs infected with Mycoplasma hyopneumoniae, Porcine circovirus-2, Porcine parvovirus, Porcine reproductive and respiratory syndrome virus, and Porcine respiratory coronavirus with this MIA was not observed. Pig 2 was consistently positive by MIA and qPCR, whereas pig 1, initially negative, seroconverted before becoming qPCR positive. Only 2 samples (from pig 1) were IHA positive. Five (9.8%) field samples were qPCR positive and 40 (78.43%) were positive for all 3 MIA antigens; however, all were IHA negative. In summary, the MIA is specific and more sensitive than qPCR and IHA, providing simultaneous evaluation of antibody response to M. suis antigens.

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Identification, Bioinformatics Analyses, and Expression of Immunoreactive Antigens of Mycoplasma haemofelis

Cited by 4 publications

References 34 publications

Comparative Genomics and Phylogenomics of Hemotrophic Mycoplasmas

Comparative Genomics and Phylogenomics of Hemotrophic Mycoplasmas

Genome of Mycoplasma haemofelis, unraveling its strategies for survival and persistence

Identification ofMycoplasma suisantigens and development of a multiplex microbead immunoassay

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