Salmonella Pullorum effector SteE regulates Th1/Th2 cytokine expression by triggering the STAT3/SOCS3 pathway that suppresses NF-κB activation.

Salmonella enterica serovar Pullorum (S. Pullorum) can regulate host immunity via special effectors that promote persistent infection and its intracellular survival. SteE as an anti-inflammatory effector is involved in the systemic infection of Salmonella in host macrophages. Macrophage activation can indirectly reflect the immune regulatory function of T helper type 1 (Th1)/T helper type 2 (Th2) cytokines. However, information concerning the regulation of Th1/Th2 cytokine expression by steE in S. Pullorum infection is limited. This study evaluates the effects of steE on the Th1/Th2 balance, STAT3/SOCS3 pathway, and NF-κB P65 activation in S. Pullorum-infected HD-11 cells and in chicken models. We demonstrated that steE diminished the expression of Th1-related cytokines (IFN-γ and IL-12) and promoted the expression of Th2-related cytokines (IL-4 and IL-10) in HD-11 cells and chicken models of S. Pullorum infection. SOCS3 silencing suppressed the function of steE in HD-11 cells and led to the imbalance of Th1/Th2-related cytokines. SteE promoted SOCS3 expression by activating STAT3 in HD-11 cells. Moreover, steE inhibited NF-κB P65 expression and blocked its translocation to the nucleus by promoting SOCS3 expression. Our results illustrated that steE regulated the expression of Th1/Th2 cytokines via modulation of the STAT3/SOCS3 and NF-κB axis, which might be associated with Th1/Th2 cell differentiation and could, therefore, be a novel therapeutic strategy against salmonellosis.

SteE Enhances the Virulence of Salmonella Pullorum in Chickens by Regulating the Inflammation Response.

Salmonella enterica serovar Pullorum (S. Pullorum) is a host-specific pathogen, which causes acute gastroenteritis with high mortality in poultry. However, the association between steE, encoded by type III secretion system 2, and Salmonella virulence is not well-understood. To elucidate the functions of steE in S. Pullorum, ΔsteE strain was constructed using the λ-Red recombination technology. Compared to that in the wild-type, the deletion of steE in S. Pullorum reduced bacterial invasion, proliferation, and late apoptosis in the infected HD-11 cells. In addition, we analyzed the mRNA expression levels of effector genes and cytokines by qRT-PCR. SteE was associated with the regulation of various effector genes and inflammatory cytokines in HD-11 cells during S. Pullorum infection. The wild-type effector steE promoted the expression of anti-inflammatory cytokines (IL-4 and IL-10) and reduced that of pro-inflammatory cytokines (IL-1ß, IL-6, and IL-12) compared to that in the ΔsteE-infected HD-11 cells and chicken spleens. Results from the chicken infection model showed that the deletion of steE resulted in significantly decreased colonization and long-term survival of the bacteria and alleviated pathological lesions compared to those in the wild-type. Further, steE increased the virulence of S. Pullorum in chickens by regulating the expression of inflammatory cytokines. Our findings provide insights into the persistent infection and autoimmunity associated with steE in S. Pullorum.

Multiplex PCR assay based on the citE2 gene and intergenic sequence for the rapid detection of Salmonella Pullorum in chickens.

Salmonella is one of the most common Gram-negative pathogens and seriously threatens chicken farms and food safety. This study aimed to establish a multiplex polymerase chain reaction (PCR) approach for the identification of different Salmonella enterica subsp. enterica. The citE2 gene and interval sequence of SPS4_00301-SPS4_00311 existed in all S. enterica subsp. enterica serovars by genomic comparison. By contrast, a 76 bp deletion in citE2 was found only in Salmonella Pullorum. Two pairs of special primers designed from citE2 and interval sequence were used to establish the multiplex PCR system. The optimized multiplex PCR system could distinguish Salmonella Pullorum and non-Salmonella Pullorum. The sensitivity of the optimized multiplex PCR system could be as low as 6.25 pg/µL and 104 colony-forming units (CFU)/mL for genomic DNA and Salmonella Pullorum cells, respectively. The developed multiplex PCR assay distinguished Salmonella Pullorum from 33 different Salmonella enterica subsp. enterica serotypes and 13 non-target species. The detection of egg samples artificially contaminated with Salmonella Pullorum, Salmonella Enteritidis, and naturally contaminated 69 anal swab samples showed that results were consistent with the culture method. These features indicated that the developed multiplex PCR system had high sensitivity and specificity and could be used for the accurate detection of Salmonella Pullorum in clinical samples.