MalF is essential for persistence of Mycoplasma gallisepticum in vivo

Tseng, Chi-Wen; Kanci, Anna; Citti, Christine; Rosengarten, Renate; Chiu, Chien-Ju; Chen, Zhenghong; Geary, Steven J.; Browning, Glenn F.; Markham, Philip F.

doi:10.1099/mic.0.067553-0

Cited by 19 publications

(26 citation statements)

References 58 publications

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“…The identified mutants could serve as promising candidates for detailed investigations to develop useful intervention strategies against M. agalactiae and possibly also against M. bovis and other ruminant pathogens. Efforts made so far in understanding the pathogenicity of mycoplasmas using Tn mutagenesis were based mostly on in vitro studies, with very few exceptions (6,22,23). This study presents an in vivo analysis of several different Tn mutants in the natural ovine host to identify genes that likely play a significant role in M. agalactiae host colonization and pathogenesis without having a comparable disadvantage during in vitro competitive growth.…”

Section: Discussionmentioning

confidence: 99%

“…Transposition studies also have helped in deciphering the role of different genetic factors involved in motility, cytadherence, and defense systems of mycoplasmas (19)(20)(21). Although transposon mutagenesis is attained in many mycoplasmas, it is only in M. gallisepticum that Tn mutant libraries have been screened in vivo for identifying pathogenicity determinants using the chicken infection model (22,23). However, there is no report of any such study in ruminant mycoplasmas, which is imperative in understanding and preventing infections caused by this economically important group of mycoplasmas.…”

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confidence: 99%

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Simultaneous Identification of Potential Pathogenicity Factors of Mycoplasma agalactiae in the Natural Ovine Host by Negative Selection

Hegde

Zimmermann

et al. 2015

Infect Immun

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Mycoplasmas possess complex pathogenicity determinants that are largely unknown at the molecular level. Mycoplasma agalactiae serves as a useful model to study the molecular basis of mycoplasma pathogenicity. The generation and in vivo screening of a transposon mutant library of M. agalactiae were employed to unravel its host colonization factors. Tn4001mod mutants were sequenced using a novel sequencing method, and functionally heterogeneous pools containing 15 to 19 selected mutants were screened simultaneously through two successive cycles of sheep intramammary infections. A PCR-based negative selection method was employed to identify mutants that failed to colonize the udders and draining lymph nodes in the animals. A total of 14 different mutants found to be absent from >95% of samples were identified and subsequently verified via a second round of stringent confirmatory screening where 100% absence was considered attenuation. Using this criterion, seven mutants with insertions in genes MAG1050, MAG2540, MAG3390, uhpT, eutD, adhT, and MAG4460 were not recovered from any of the infected animals. Among the attenuated mutants, many contain disruptions in hypothetical genes, implying their previously unknown role in M. agalactiae pathogenicity. These data indicate the putative role of functionally different genes, including hypothetical ones, in the pathogenesis of M. agalactiae. Defining the precise functions of the identified genes is anticipated to increase our understanding of M. agalactiae infections and to develop successful intervention strategies against it.

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Section: Discussionmentioning

confidence: 99%

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confidence: 99%

Simultaneous Identification of Potential Pathogenicity Factors of Mycoplasma agalactiae in the Natural Ovine Host by Negative Selection

Hegde

Zimmermann

et al. 2015

Infect Immun

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“…These mutants contained insertions in genes encoding the cytadhesin GapA and its accessory protein, CrmA. Three other virulence-associated determinants in M. gallisepticum, lpd, malF, and mslA, which has recently been shown to encode a novel oligonucleotide binding protein, have also been identified previously using transposon mutagenesis (5)(6)(7)(8). Two of these (malF and mslA) are members of ABC transporter operons.…”

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“…In a previous study, we showed that several signature-tagged (ST) mutants of M. gallisepticum that were recovered infrequently from chickens inoculated with pools of mutants were significantly attenuated when assessed for their virulence in isolation (5). These mutants contained insertions in genes encoding the cytadhesin GapA and its accessory protein, CrmA.…”

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The oppD Gene and Putative Peptidase Genes May Be Required for Virulence in Mycoplasma gallisepticum

et al. 2017

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Relatively few virulence genes have been identified in pathogenic mycoplasmas, so we used signature-tagged mutagenesis to identify mutants of the avian pathogen Mycoplasma gallisepticum with a reduced capacity to persist in vivo and compared the levels of virulence of selected mutants in experimentally infected chickens. Four mutants had insertions in one of the two incomplete oppABCDF operons, and a further three had insertions in distinct hypothetical genes, two containing peptidase motifs and one containing a member of a gene family. The three hypothetical gene mutants and the two with insertions in oppD 1 were used to infect chickens, and all five were shown to have a reduced capacity to induce respiratory tract lesions. One oppD 1 mutant and the MGA_1102 and MGA_1079 mutants had a greatly reduced capacity to persist in the respiratory tract and to induce systemic antibody responses against M. gallisepticum. The other oppD 1 mutant and the MGA_0588 mutant had less capacity than the wild type to persist in the respiratory tract but did elicit systemic antibody responses. Although M. gallisepticum carries two incomplete opp operons, one of which has been acquired by horizontal gene transfer, our results suggest that one of the copies of oppD may be required for full expression of virulence. We have also shown that three hypothetical genes, two of which encode putative peptidases, may be required for full expression of virulence in M. gallisepticum. None of these genes has previously been shown to influence virulence in pathogenic mycoplasmas. KEYWORDS Mycoplasma gallisepticum, oppD, virulence Mycoplasma gallisepticum is the most important mycoplasmal pathogen in poultry (1, 2) and a particularly useful model of mycoplasmal pathogenesis, in part as a typical representative of the Pneumoniae phylogenic group, which includes the important human pathogen M. pneumoniae. There have been relatively few studies attempting to identify virulence genes in mycoplasmas, in part because of the limited availability of tools for such studies. Signature-tagged mutagenesis (STM) is a useful technique for identifying genes likely to be involved in virulence, as it enables the rapid comparison of populations of mutants used to inoculate animals with those recovered from these animals during the course of infection, allowing identification of mutants with a reduced capacity to persist in experimentally infected animals (3, 4). However, the examination of only pools of mutants may result in a failure to distinguish between a mutation in a gene that is essential in vivo and a mutation in a gene that reduces the capacity of the mutant to compete with other mutants in vivo. In addition, it is not possible to fully assess the virulence of individual mutants within populations. Therefore, STM needs to be complemented with direct comparisons of isolated mutants for their capacity to infect, persist, and cause disease.

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“…M. gallisepticum is pathogenic in other species, causing infectious sinusitis in turkeys and conjunctivitis in house finches (1,2). Aside from primary attachment proteins GapA and CrmA, fibronectin binding proteins PlpA and Hlp3, sugar transport permease MalF, and dihydrolipoamide dehydrogenase Lpd, little is understood about specific bacterial factors affecting survival and persistence of this important avian pathogen in its natural host (3)(4)(5)(6).…”

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Global Changes in Mycoplasma gallisepticum Phase-Variable Lipoprotein Gene vlhA Expression during In Vivo Infection of the Natural Chicken Host

et al. 2016

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Mycoplasma gallisepticum is the primary etiologic agent of chronic respiratory disease in poultry, a disease largely affecting the respiratory tract and causing significant economic losses worldwide. Immunodominant proteins encoded by members of the variable lipoprotein and hemagglutinin (vlhA) gene family are thought to be important for mechanisms of M. gallisepticum-host interaction, pathogenesis, and immune evasion, but their exact role and the overall nature of their phase variation are unknown. To better understand these mechanisms, we assessed global transcriptomic vlhA gene expression directly from M. gallisepticum populations present on tracheal mucosae during a 7-day experimental infection in the natural chicken host. Here we report differences in both dominant and minor vlhA gene expression levels throughout the first week of infection and starting as early as day 1 postinfection, consistent with a functional role not dependent on adaptive immunity for driving phase variation. Notably, data indicated that, at given time points, specific vlhA genes were similarly dominant in multiple independent hosts, suggesting a nonstochastic temporal progression of dominant vlhA gene expression in the colonizing bacterial population. The dominant expression of a given vlhA gene was not dependent on the presence of 12-copy GAA trinucleotide repeats in the promoter region and did not revert to the predominate vlhA gene when no longer faced with host pressures. Overall, these data indicate that vlhA phase variation is dynamic throughout the earliest stages of infection and that the pattern of dominant vlhA expression may be nonrandom and regulated by previously unrecognized mechanisms.

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MalF is essential for persistence of Mycoplasma gallisepticum in vivo

Cited by 19 publications

References 58 publications

Simultaneous Identification of Potential Pathogenicity Factors of Mycoplasma agalactiae in the Natural Ovine Host by Negative Selection

Simultaneous Identification of Potential Pathogenicity Factors of Mycoplasma agalactiae in the Natural Ovine Host by Negative Selection

The oppD Gene and Putative Peptidase Genes May Be Required for Virulence in Mycoplasma gallisepticum

Global Changes in Mycoplasma gallisepticum Phase-Variable Lipoprotein Gene vlhA Expression during In Vivo Infection of the Natural Chicken Host

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