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

Hegde, Shivanand; Hegde, Shrilakshmi; Zimmermann, Martina E.; Flöck, Martina; Spergser, Joachim; Rosengarten, Renate; Chopra-Dewasthaly, Rohini

doi:10.1128/iai.00403-15

Cited by 10 publications

(20 citation statements)

References 67 publications

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“…Taken together, these data suggested that the Toll and IMD pathways were induced by invasion of Cedecea into mosquito cells. This is consistent with previous observations which demonstrated interplay between native microbiota and mosquito innate immune pathways [61, 66–68]. We observed dramatic modulation of effector molecules with Defensin , Cecropin and Gambicin, all significantly enhanced by Cedecea .…”

Section: Resultssupporting

confidence: 93%

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Gut-associated bacteria invade the midgut epithelium of Aedes aegypti and stimulate innate immunity and suppress Zika virus infection in cells

Hegde

Voronin

Casas-Sánchez

et al. 2019

Preprint

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23Microbiota within mosquitoes influence nutrition, immunity, fecundity, and the capacity 24 to transmit pathogens. Despite their importance, we have a limited understanding of 25 host-microbiota interactions, especially at the cellular level. It is evident bacterial 26 symbionts that are localized within the midgut also infect other organs within the 27 mosquito; however, the route these symbionts take to colonize other tissues is 28 unknown. Here, utilizing the gentamicin protection assay, we showed that the bacterial 29 symbionts Cedecea and Serratia have the capacity to invade and reside intracellularly 30 within mosquito cells. Symbiotic bacteria were found within a vacuole and bacterial 31 replication was observed in mosquito cell by transmission electron microscopy, 32indicating bacteria were adapted to the intracellular milieu. Using gene silencing, we 33 determined that bacteria exploited host factors, including actin and integrin receptors, to 34 actively invade mosquito cells. As microbiota can affect pathogens within mosquitoes, 35we examined the influence of intracellular symbionts on Zika virus (ZIKV) infection. 36Mosquito cells harbouring intracellular bacteria had significantly less ZIKV compared to 37 uninfected cells or cells exposed to non-invasive bacteria. Intracellular bacteria were 38 observed to substantially upregulate the Toll and IMD innate immune pathways, 39 providing a possible mechanism mediating these anti-viral effects. Examining mono-40 axenically infected mosquitoes using transmission electron and fluorescent microscopy 41 revealed that bacteria occupied an intracellular niche in vivo. Our results provided 42 evidence that bacteria that associate with the midgut of mosquitoes have intracellular 43 lifestyles which likely have implications for mosquito biology and pathogen infection. 44 This study expands our understanding of host-microbiota interactions in mosquitoes, 45 3 which is important as symbiont microbes are being exploited for vector control 46 strategies. 47 48 4 Introduction 49

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Section: Resultssupporting

confidence: 93%

“…Midgut-associated bacteria can affect pathogens transmitted by mosquitoes by direct or indirect interactions [59–61]. Therefore, we examined how intracellular C. neteri affected viral infection.…”

Section: Resultsmentioning

confidence: 99%

Gut-associated bacteria invade the midgut epithelium of Aedes aegypti and stimulate innate immunity and suppress Zika virus infection in cells

Hegde

Voronin

Casas-Sánchez

et al. 2019

Preprint

Self Cite

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“…agalactiae enabling survival in the mammary gland are not completely understood. We had recently identified potential pathogenicity factors by negative selection of transposon mutants that were unable to survive in the experimentally infected udders [11, 12]. Although the exact role of most of the identified factors remains to be elucidated, pyruvate dehydrogenase was shown to be involved in cell invasiveness [13], a strategy that M .…”

Section: Introductionmentioning

confidence: 99%

Comprehensive RNA-Seq Profiling to Evaluate the Sheep Mammary Gland Transcriptome in Response to Experimental Mycoplasma agalactiae Infection

et al. 2017

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Mycoplasma agalactiae is a worldwide serious pathogen of small ruminants that usually spreads through the mammary route causing acute to subacute mastitis progressing to chronic persistent disease that is hard to eradicate. Knowledge of mechanisms of its pathogenesis and persistence in the mammary gland are still insufficient, especially the host-pathogen interplay that enables it to reside in a chronic subclinical state. This study reports transcriptome profiling of mammary tissue from udders of sheep experimentally infected with M. agalactiae type strain PG2 in comparison with uninfected control animals using Illumina RNA-sequencing (RNA-Seq). Several differentially expressed genes (DEGs) were observed in the infected udders and RT-qPCR analyses of selected DEGs showed their expression profiles to be in agreement with results from RNA-Seq. Gene Ontology (GO) analysis revealed majority of the DEGs to be associated with mycoplasma defense responses that are directly or indirectly involved in host innate and adaptive immune responses. Similar RNA-Seq analyses were also performed with spleen cells of the same sheep to know the specific systemic transcriptome responses. Spleen cells exhibited a comparatively lower number of DEGs suggesting a less prominent host response in this organ. To our knowledge this is the first study that describes host transcriptomics of M. agalactiae infection and the related immune-inflammatory responses. The data provides useful information to further dissect the molecular genetic mechanisms underlying mycoplasma mastitis, which is a prerequisite for designing effective intervention strategies.

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“…These studies have demonstrated the role of the vpma locus in M. agalactiae pathogenesis [ 8 ]. Also, recently we attempted to identify potential pathogenicity factors of M. agalactiae in the natural sheep host by negative selection of transposon mutants [ 9 ]. Other independent studies using transposon mutants were able to show the role of different genomic loci in interaction with host cells [ 10 ] and the results obtained were further evaluated in vivo to conclude that the NIF locus was important for the pathogenicity of M. agalactiae [ 11 ].…”

Section: Introductionmentioning

confidence: 99%

“…Systemic spread of M. agalactiae after intramammary infection is well established [ 12 ]. In a recent experimental infection trial using pools of transposon mutants for intramammary infection of lactating ewes, we identified transposon mutants absent at the local site of infection and lymph nodes (LNs) [ 9 ]. The present study is an extension of the latter work, where we have further assessed the presence or absence of specific mutants in several inner organs distant from the site of inoculation.…”

Section: Introductionmentioning

confidence: 99%

Genetic loci of Mycoplasma agalactiae involved in systemic spreading during experimental intramammary infection of sheep

Hegde

Zimmermann

Flöck

et al. 2016

Vet Res

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Mycoplasmas are amongst the most successful pathogens of both humans and animals yet the molecular basis of mycoplasma pathogenesis is poorly understood. This is partly due to the lack of classical virulence factors and little similarity to common bacterial pathogenic determinants. Using Mycoplasma agalactiae as a model we initiated research in this direction by screening a transposon mutant library in the natural sheep host using a negative selection method. Having successfully identified putative factors involved in the colonization of local infection and lymphogenic sites, the current study assessed mutants unable to spread systemically in sheep after experimental intramammary infection. Analysis of distant body sites for complete absence of mutants via SSM PCR revealed that additional set of genes, such as pdhB, oppC, oppB, gtsB, MAG1890, MAG5520 and MAG3650 are required for systemic spreading apart from those that were necessary for initial colonization. Additional in vitro studies with the mutants absent at these systemic sites confirmed the potential role of some of the respective gene products concerning their interaction with host cells. Mutants of pdhB, oppC and MAG4460 exhibited significantly slower growth in the presence of HeLa cells in MEM medium. This first attempt to identify genes exclusively required for systemic spreading provides a basis for further in-depth research to understand the exact mechanism of chronicity and persistence of M. agalactiae.Electronic supplementary materialThe online version of this article (doi:10.1186/s13567-016-0387-0) contains supplementary material, which is available to authorized users.

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

Cited by 10 publications

References 67 publications

Gut-associated bacteria invade the midgut epithelium of Aedes aegypti and stimulate innate immunity and suppress Zika virus infection in cells

Gut-associated bacteria invade the midgut epithelium of Aedes aegypti and stimulate innate immunity and suppress Zika virus infection in cells

Comprehensive RNA-Seq Profiling to Evaluate the Sheep Mammary Gland Transcriptome in Response to Experimental Mycoplasma agalactiae Infection

Genetic loci of Mycoplasma agalactiae involved in systemic spreading during experimental intramammary infection of sheep

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