Signature-tagged mutagenesis screening revealed a novel smooth-to-rough transition determinant of Salmonella enterica serovar Enteritidis.

BACKGROUND: Salmonella enterica serovar Enteritidis (S. Enteritidis) has emerged as one of the most important food-borne pathogens for humans. Lipopolysaccharide (LPS), as a component of the outer membrane, is responsible for the virulence and smooth-to-rough transition in S. Enteritidis. In this study, we screened S. Enteritidis signature-tagged transposon mutant library using monoclonal antibody against somatic O9 antigen (O9 MAb) and O9 factor rabbit antiserum to identify novel genes that are involved in smooth-to-rough transition. RESULTS: A total of 480 mutants were screened and one mutant with transposon insertion in rfbG gene had smooth-to-rough transition phenotype. In order to verify the role of rfbG gene, an rfbG insertion or deletion mutant was constructed using λ-Red recombination system. Phenotypic and biological analysis revealed that rfbG insertion or deletion mutants were similar to the wild-type strain in growth rate and biochemical properties, but the swimming motility was reduced. SE Slide Agglutination test and ELISA test showed that rfbG mutants do not stimulate animals to produce agglutinating antibody. In addition, the half-lethal dose (LD50) of the rfbG deletion mutant strain was 106.6 -fold higher than that of the parent strain in a mouse model when injected intraperitoneally. CONCLUSIONS: These data indicate that the rfbG gene is involved in smooth-to-rough transition, swimming motility and virulence of S. Enteritidis. Furthermore, somatic O-antigen antibody-based approach to screen signature-tagged transposon mutants is feasible to clarify LPS biosynthesis and to find suitable markers in DIVA-vaccine research.

An Improved Metal-Packaged Strain Sensor Based on A Regenerated Fiber Bragg Grating in Hydrogen-Loaded Boron-Germanium Co-Doped Photosensitive Fiber for High-Temperature Applications.

Local strain measurements are considered as an effective method for structural health monitoring of high-temperature components, which require accurate, reliable and durable sensors. To develop strain sensors that can be used in higher temperature environments, an improved metal-packaged strain sensor based on a regenerated fiber Bragg grating (RFBG) fabricated in hydrogen (H2)-loaded boron-germanium (B-Ge) co-doped photosensitive fiber is developed using the process of combining magnetron sputtering and electroplating, addressing the limitation of mechanical strength degradation of silica optical fibers after annealing at a high temperature for regeneration. The regeneration characteristics of the RFBGs and the strain characteristics of the sensor are evaluated. Numerical simulation of the sensor is conducted using a three-dimensional finite element model. Anomalous decay behavior of two regeneration regimes is observed for the FBGs written in H2-loaded B-Ge co-doped fiber. The strain sensor exhibits good linearity, stability and repeatability when exposed to constant high temperatures of up to 540 °C. A satisfactory agreement is obtained between the experimental and numerical results in strain sensitivity. The results demonstrate that the improved metal-packaged strain sensors based on RFBGs in H2-loaded B-Ge co-doped fiber provide great potential for high-temperature applications by addressing the issues of mechanical integrity and packaging.

Evaluation of the Protective Immune Response Induced by an rfbG-Deficient Salmonella enterica Serovar Enteritidis Strain as a Live Attenuated DIVA (Differentiation of Infected and Vaccinated Animals) Vaccine in Chickens.

Salmonella enterica serovar Enteritidis (S. Enteritidis), one of the zoonotic pathogens, not only results in significant financial losses for the global poultry industry but also has the potential to spread to humans through poultry and poultry products. Vaccination is an effective method to prevent Salmonella infections. In this study, we constructed a live attenuated DIVA (differentiation of infected and vaccinated animals) vaccine candidate, Z11ΔrfbG, and evaluated its protective effectiveness and DIVA potential in chickens. Compared to that of the virulent wild-type strain, the 50% lethal dose (LD50) of the rfbG mutant strain increased 56-fold, confirming its attenuation. High serum levels of S. Enteritidis-specific IgG titers indicated that a significant humoral immune response was induced in the vaccinated group. After challenge, the nonvaccinated group showed serious clinical symptoms (diarrhea, depression, decreased appetite, ruffled feathers, and weight loss), pathological changes (white nodules in the liver and fatty lesions in liver cells), and death. In contrast, there were no clinical symptoms, pathological changes, or death in the 5 × 106- and 5 × 107-CFU-vaccinated groups. Z11ΔrfbG vaccination significantly reduced S. Enteritidis colonization in the spleen, liver, and cecum. In addition, the Z11ΔrfbG-vaccinated group exhibited a negative response to the serological test, whereas the virulent wild-type Z11 infection group was strongly positive for the serological test, showing a DIVA capability of Z11ΔrfbG vaccination. Overall, our findings demonstrate the viability of the rfbG mutant as a live attenuated chicken vaccine that can discriminate between animals that have been immunized and those that have been infected. IMPORTANCE S. Enteritidis is a highly adapted pathogen that causes significant economic losses in the poultry industry around the world. Vaccination is an effective method of controlling S. Enteritidis infections. Here, we demonstrated that S. Enteritidis Z11ΔrfbG has the potential to be a safe, immunogenic, and DIVA vaccine candidate for the control of Salmonella infections in chickens. Z11ΔrfbG not only provided effective protection in chickens but also distinguished between infected and vaccinated chickens by serological tests.