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The role of autophagy in the intracellular survival of Campylobacter concisus.

Burgos-Portugal JA, Mitchell HM, Castaño-Rodríguez N, Kaakoush NO - FEBS Open Bio (2014)

Bottom Line: Autophagy inhibition resulted in two- to four-fold increases in intracellular levels of C. concisus within Caco-2 cells, while autophagy induction resulted in a significant reduction in intracellular levels or bacterial clearance.C. concisus strains with low intracellular survival levels showed a dramatic increase in these levels upon autophagy inhibition.Our data collectively indicates that while autophagy is important for the clearance of C. concisus, some strains may manipulate this process to benefit their intracellular survival.

View Article: PubMed Central - PubMed

Affiliation: School of Biotechnology and Biomolecular Sciences, The University of New South Wales, Sydney, NSW 2052, Australia.

ABSTRACT
Campylobacter concisus is an emerging pathogen that has been associated with gastrointestinal diseases. Given the importance of autophagy for the elimination of intracellular bacteria and the subversion of this process by pathogenic bacteria, we investigated the role of autophagy in C. concisus intracellular survival. Gentamicin protection assays were employed to assess intracellular levels of C. concisus within Caco-2 cells, following autophagy induction and inhibition. To assess the interaction between C. concisus and autophagosomes, confocal microscopy, scanning electron microscopy, and transmission electron microscopy were employed. Expression levels of 84 genes involved in the autophagy process were measured using qPCR. Autophagy inhibition resulted in two- to four-fold increases in intracellular levels of C. concisus within Caco-2 cells, while autophagy induction resulted in a significant reduction in intracellular levels or bacterial clearance. C. concisus strains with low intracellular survival levels showed a dramatic increase in these levels upon autophagy inhibition. Confocal microscopy showed co-localization of the bacterium with autophagosomes, while transmission electron microscopy identified intracellular bacteria persisting within autophagic vesicles. Further, qPCR showed that following infection, 13 genes involved in the autophagy process were significantly regulated, and a further five showed borderline results, with an overall indication towards a dampening effect exerted by the bacterium on this process. Our data collectively indicates that while autophagy is important for the clearance of C. concisus, some strains may manipulate this process to benefit their intracellular survival.

No MeSH data available.


Related in: MedlinePlus

Intracellular percentage of C. concisus UNSWCS, ATCC 51562 and BAA-1457 following autophagy inhibition. Errors are presented as Standard Error of the Mean (SEM) based on a minimum of four biological replicates. 3-MA: 10 mM 3-methyladenine.
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f0010: Intracellular percentage of C. concisus UNSWCS, ATCC 51562 and BAA-1457 following autophagy inhibition. Errors are presented as Standard Error of the Mean (SEM) based on a minimum of four biological replicates. 3-MA: 10 mM 3-methyladenine.

Mentions: To investigate this further, three additional C. concisus strains previously shown to have naturally low intracellular percentages of bacteria (ATCC 51562: 0.00048 ± 0.00016%; UNSWCS: 0.00059 ± 0.00015%) or with no invasion (BAA-1457) were employed. Interestingly, upon addition of 10 mM 3-MA, the intracellular percentages of these strains significantly increased (P < 0.0001) to levels observed for UNSWCD (ATCC 51562: 0.51 ± 0.06%; UNSWCS: 0.72 ± 0.14%; BAA-1457: 0.66 ± 0.09%) (Fig. 2). We then examined whether host adaptation may have a similar effect on the intracellular levels of C. concisus by using bacteria that internalized into host cells for the invasion assay (re-invasion). This was performed for C. concisus UNSWCS (0.00059 ± 0.00015%), a strain with low levels of invasion. This showed that upon re-invasion of host-adapted isolates, the intracellular percentage increased approximately 8.6-fold to 0.0051 ± 0.0009%. A further attempt at re-invasion of internalized isolates did not alter the intracellular percentage (0.0045 ± 0.0011%), and a third re-invasion resulted in isolates with gentamicin resistance. These findings would suggest that the autophagy process is one of the primary regulators of intracellular levels of C. concisus strains with low invasion percentages.


The role of autophagy in the intracellular survival of Campylobacter concisus.

Burgos-Portugal JA, Mitchell HM, Castaño-Rodríguez N, Kaakoush NO - FEBS Open Bio (2014)

Intracellular percentage of C. concisus UNSWCS, ATCC 51562 and BAA-1457 following autophagy inhibition. Errors are presented as Standard Error of the Mean (SEM) based on a minimum of four biological replicates. 3-MA: 10 mM 3-methyladenine.
© Copyright Policy - CC BY-NC-ND
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4048850&req=5

f0010: Intracellular percentage of C. concisus UNSWCS, ATCC 51562 and BAA-1457 following autophagy inhibition. Errors are presented as Standard Error of the Mean (SEM) based on a minimum of four biological replicates. 3-MA: 10 mM 3-methyladenine.
Mentions: To investigate this further, three additional C. concisus strains previously shown to have naturally low intracellular percentages of bacteria (ATCC 51562: 0.00048 ± 0.00016%; UNSWCS: 0.00059 ± 0.00015%) or with no invasion (BAA-1457) were employed. Interestingly, upon addition of 10 mM 3-MA, the intracellular percentages of these strains significantly increased (P < 0.0001) to levels observed for UNSWCD (ATCC 51562: 0.51 ± 0.06%; UNSWCS: 0.72 ± 0.14%; BAA-1457: 0.66 ± 0.09%) (Fig. 2). We then examined whether host adaptation may have a similar effect on the intracellular levels of C. concisus by using bacteria that internalized into host cells for the invasion assay (re-invasion). This was performed for C. concisus UNSWCS (0.00059 ± 0.00015%), a strain with low levels of invasion. This showed that upon re-invasion of host-adapted isolates, the intracellular percentage increased approximately 8.6-fold to 0.0051 ± 0.0009%. A further attempt at re-invasion of internalized isolates did not alter the intracellular percentage (0.0045 ± 0.0011%), and a third re-invasion resulted in isolates with gentamicin resistance. These findings would suggest that the autophagy process is one of the primary regulators of intracellular levels of C. concisus strains with low invasion percentages.

Bottom Line: Autophagy inhibition resulted in two- to four-fold increases in intracellular levels of C. concisus within Caco-2 cells, while autophagy induction resulted in a significant reduction in intracellular levels or bacterial clearance.C. concisus strains with low intracellular survival levels showed a dramatic increase in these levels upon autophagy inhibition.Our data collectively indicates that while autophagy is important for the clearance of C. concisus, some strains may manipulate this process to benefit their intracellular survival.

View Article: PubMed Central - PubMed

Affiliation: School of Biotechnology and Biomolecular Sciences, The University of New South Wales, Sydney, NSW 2052, Australia.

ABSTRACT
Campylobacter concisus is an emerging pathogen that has been associated with gastrointestinal diseases. Given the importance of autophagy for the elimination of intracellular bacteria and the subversion of this process by pathogenic bacteria, we investigated the role of autophagy in C. concisus intracellular survival. Gentamicin protection assays were employed to assess intracellular levels of C. concisus within Caco-2 cells, following autophagy induction and inhibition. To assess the interaction between C. concisus and autophagosomes, confocal microscopy, scanning electron microscopy, and transmission electron microscopy were employed. Expression levels of 84 genes involved in the autophagy process were measured using qPCR. Autophagy inhibition resulted in two- to four-fold increases in intracellular levels of C. concisus within Caco-2 cells, while autophagy induction resulted in a significant reduction in intracellular levels or bacterial clearance. C. concisus strains with low intracellular survival levels showed a dramatic increase in these levels upon autophagy inhibition. Confocal microscopy showed co-localization of the bacterium with autophagosomes, while transmission electron microscopy identified intracellular bacteria persisting within autophagic vesicles. Further, qPCR showed that following infection, 13 genes involved in the autophagy process were significantly regulated, and a further five showed borderline results, with an overall indication towards a dampening effect exerted by the bacterium on this process. Our data collectively indicates that while autophagy is important for the clearance of C. concisus, some strains may manipulate this process to benefit their intracellular survival.

No MeSH data available.


Related in: MedlinePlus