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Immune reaction and survivability of salmonella typhimurium and salmonella infantis after infection of primary avian macrophages.

ABSTRACT: Salmonella serovars are differentially able to infect chickens. The underlying causes are not yet fully understood. Aim of the present study was to elucidate the importance of Salmonella Pathogenicity Island 1 and 2 (SPI-1 and -2) for the virulence of two non-host-specific, but in-vivo differently invasive, Salmonella serovars in conjunction with the immune reaction of the host. Primary avian splenic macrophages were inoculated with Salmonella enterica sub-species enterica serovar (S.) Typhimurium and S. Infantis. The number and viability of intracellular bacteria and transcription of SPI-1 and -2 genes by the pathogens, as well as transcription of immune-related proteins, surface antigen expression and nitric oxide production by the macrophages, were compared at different times post inoculation. After infection, both of the Salmonella serovars were found inside the primary macrophages. Invasion-associated SPI-1 genes were significantly higher transcribed in S. Infantis- than S. Typhimurium-infected macrophages. The macrophages counteracted the S. Infantis and S. Typhimurium infection with elevated mRNA expression of inducible nitric oxide synthase (iNOS), interleukin (IL)-12, IL-18 and lipopolysaccharide-induced tumor necrosis factor alpha factor (LITAF) as well as with an increased synthesis of nitric oxide. Despite these host cell attacks, S. Typhimurium was better able than S. Infantis to survive within the macrophages and transcribed higher rates of the SPI-2 genes spiC, ssaV, sifA, and sseA. The results showed similar immune reactions of primary macrophages after infection with both of the Salmonella strains. The more rapid and stronger transcription of SPI-2-related genes by intracellular S. Typhimurium compared to S. Infantis might be responsible for its better survival in avian primary macrophages.

SUBMITTER: Braukmann M

PROVIDER: S-EPMC4374797 | biostudies-literature | 2015

REPOSITORIES: biostudies-literature

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Immune reaction and survivability of salmonella typhimurium and salmonella infantis after infection of primary avian macrophages.

Braukmann Maria M Methner Ulrich U Berndt Angela A

PloS one 20150326 3

Salmonella serovars are differentially able to infect chickens. The underlying causes are not yet fully understood. Aim of the present study was to elucidate the importance of Salmonella Pathogenicity Island 1 and 2 (SPI-1 and -2) for the virulence of two non-host-specific, but in-vivo differently invasive, Salmonella serovars in conjunction with the immune reaction of the host. Primary avian splenic macrophages were inoculated with Salmonella enterica sub-species enterica serovar (S.) Typhimuri ...[more]

PMID: 25811871

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Project description:Macrophages are central for the immune control of intracellular microbes. Heme oxygenase 1 (HO-1, hmox) is the first and rate limiting enzyme in the breakdown of heme originating from degraded senescent erythrocytes and heme-proteins, yielding equal amounts of iron, carbon monoxide and biliverdin. HO-1 is strongly up-regulated in macrophages in response to inflammatory signals, including bacterial endotoxin. In view of the essential role of iron for the growth and proliferation of intracellular bacteria along with known effects of the metal on innate immune function, we examined whether HO-1 plays a role in the control of infection with the intracellular bacterium Salmonella Typhimurium. We studied the course of infection in stably-transfected murine macrophages (RAW264.7) bearing a tetracycline-inducible plasmid producing hmox shRNA and in primary HO-1 knockout macrophages. While uptake of bacteria into macrophages was not affected, a significantly reduced survival of intracellular Salmonella was observed upon hmox knockdown or pharmacological hmox inhibition, which was independent of Nramp1 functionality. This could be traced to limitation of iron availability for intramacrophage bacteria along with enhanced stimulation of innate immune effector pathways, including the formation of reactive oxygen and nitrogen species and increased TNF-α expression. Mechanistically, these latter effects result from intracellular iron limitation with subsequent activation of NF-κB and further inos, tnfa and p47phox transcription along with reduced formation of the anti-inflammatory and radical scavenging molecules, CO and biliverdin as a consequence of HO-1 silencing. Taken together our data provide novel evidence that the infection-driven induction of HO-1 exerts detrimental effects in the early control of Salmonella infection, whereas hmox inhibition can favourably modulate anti-bacterial immune effector pathways of macrophages and promote bacterial elimination.

Project description:The host-specific Salmonella serovar S. Pullorum (SP) modulates the chicken immune response to a Th2-biased response associated with persistent infection. This is different from the Th1-biased immune response induced by the genetically close serovar, S. Enteritidis (SE). Based on core genome differences between SP and SE, we used three complementary bioinformatics approaches to identify SP genes, which may be important for stimulation of the immune response. Defined mutants were constructed in selected genes, and the infection potential and ability of mutants to stimulate cytokine production in avian derived HD11 macrophages were determined. Deletion of large genomic regions unique to SP did not change infection potential nor immune stimulation significantly. Mutants in genes with conserved single nucleotide polymorphisms (SNPs) between the two serovars in the region 100 bp upstream of the start codon (conserved upstream SNPs [CuSNPs]) such as sseE, osmB, tolQ, a putative immune antigen, and a putative persistent infection factor, exhibited differences in induction of inflammatory cytokines compared to wild-type SP, suggesting a possible role of these CuSNPs in immune regulation. Single nucleotide SP mutants correcting for the CuSNP difference were constructed in the upstream region of sifA and pipA. The SNP corrected pipA mutant expressed pipA at a higher level than the wild-type SP strain, and the mutant differentially caused upregulation of proinflammatory cytokines. It suggests that this CuSNP is important for the suppression of proinflammatory responses. In conclusion, this study has identified putative immune stimulating factors of relevance to the difference in infection dynamics between SP and SE in avian macrophages. IMPORTANCE Salmonella Pullorum is host specific to avian species, where it causes life-threatening infection in young birds. It is unknown why it is host restricted and causes systemic disease, rather than gastroenteritis normally seen with Salmonella. In the present study, we identified genes and single nucleotide polymorphisms (SNPs; relative to the broad-host-range type Salmonella Enteritidis), which affected survival and immune induction in macrophages from hens suggesting a role in development of the host specific infection. Further studies of such genes may enable understanding of which genetic factors determine the development of host specific infection by S. Pullorum. In this study, we developed an in silico approach to predict candidate genes and SNPs for development of the host-specific infection and the specific induction of immunity associated with this infection. This study flow can be used in similar studies in other clades of bacteria.

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Dataset Information

Immune reaction and survivability of salmonella typhimurium and salmonella infantis after infection of primary avian macrophages.

Publications

Immune reaction and survivability of salmonella typhimurium and salmonella infantis after infection of primary avian macrophages.

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