NAD+ pool depletion as a signal for the Rex regulon involved in Streptococcus agalactiae virulence

In many Gram-positive bacteria, the redox-sensing transcriptional repressor Rex controls central carbon and energy metabolism by sensing the intra cellular balance between the reduced and oxidized forms of nicotinamide adenine dinucleotide; the NADH/NAD+ ratio. Here, we report high-resolution crystal structures and characterization of a Rex ortholog (Gbs1167) in the opportunistic pathogen, Streptococcus agalactiae, also known as group B streptococcus (GBS). We present structures of Rex bound to NAD+ and to a DNA operator which are the first structures of a Rex-family member from a pathogenic bacterium. The structures reveal the molecular basis of DNA binding and the conformation alterations between the free NAD+ complex and DNA-bound form of Rex. Transcriptomic analysis revealed that GBS Rex controls not only central metabolism, but also expression of the monocistronic rex gene as well as virulence gene expression. Rex enhances GBS virulence after disseminated infection in mice. Mechanistically, NAD+ stabilizes Rex as a repressor in the absence of NADH. However, GBS Rex is unique compared to Rex regulators previously characterized because of its sensing mechanism: we show that it primarily responds to NAD+ levels (or growth rate) rather than to the NADH/NAD+ ratio. These results indicate that Rex plays a key role in GBS pathogenicity by modulating virulence factor gene expression and carbon metabolism to harvest nutrients from the host.

YeiE Regulates Motility and Gut Colonization in Salmonella enterica Serotype Typhimurium

The ability to finely tune virulence factor gene expression is required for bacterial pathogens to successfully colonize a host. Flagellum-mediated motility is critical for many gut pathogens to establish productive infections.

Development of IMBs-qPCR Detection Method for Yersinia enterocolitica Based on the foxA Gene

Yersinia enterocolitica is an important zoonotic pathogen, which seriously endangers food safety risk. In this study, the recombinant outer membrane protein OmpF and its antibody were prepared and coupled with immunomagnetic beads (IMBs) to capture Y. enterocolitica in food samples, combining the quantitative PCR detection with primers of virulence factor gene fox A for Yersinia enterocolitica contamination. The results showed that the capture efficiency of approximately 80% using anti-OmpF antibody-immunomagnetic beads and linearly dependent capture under 10 1 -10 5 CFU/mL Y. enterocolitica . compare with less than 10% capture of other bacteria. The detection limit of 64 CFU/mL was obtained by based on fox A gene PCR detection combined with capture of the anti-OmpF antibody-immunomagnetic beads to detect Yersinia enterocolitica in artificially contaminated milk and pork samples. Comparing with the culture method, the developed IMBs-qPCR method has higher consistency, less time consuming, which providing an effective alternative method for rapid detection of Y. enterocolitica in food.

Molecular Detection of Virulence Factor Glycoprotein (Gp63) of Leishmania spp. in Phlebotomus Sand Flies

Introduction: Gp63 is the major surface glycoprotein of Leishmania which is prevalent in the promastigote stage of Phlebotomus sergenti. Glycoprotein 63 (gp63) or leishmanolysin is a zinc-dependent metalloprotease found on the surface of Leishmania. It was initially discovered in 1980and described biochemically and genetically as a surface antigen expressed in promastigotes of Leishmania species, having a range of substrates including casein, albumin, fibrinogen, haemoglobin, and gelatin. On the surface of amastigote, GP63 is present at a very low level. Glycoprotein 63 is a zinc-dependent metalloprotease that is active in the pH range of 7–10, which is neutral to alkaline. Materials and Methods: During the study, which was carried out in Al-Muthanna province from July 2017 to August 2018, about 2550 sand fly samples including 719 males and 1633 females were collected. Three species of sand flies were recorded according to morphological features which included Phlebotomus papatasi, Phlebotomus sergenti, and Sergentomyia sintoni. The caught specimens showed that P. papatasi was the most prevalent species. Two primers were used for the diagnosis of Leishmania spp. using a nested PCR technique, which was designed in a previous study for 1250 samples of sand flies from 25 locations in the study area. Results: Results showed that 13 samples were positive, containing DNA for the Leishmania parasite, and 12 samples were negative. Positive samples include 11 samples of L. major species and only 2 samples of L. tropica species. Additionally, the results showed that the L. major species was the dominant species in the study area. Positive samples (13) of sand flies had Leishmania parasite based on molecular diagnosis and virulence factor gene (Gp63) of Leishmania parasite was detected in sand flies using PCR method The results showed that virulence factor gene (Gp63) was detected on the surface of promastigote. This refers to the presence of Leishmania parasite in sand fly. The study is the first one which detected the presence of the virulence factors gene (GP63) of Leishmania in the body of sand flies. Conclusion: This study revealed the possibility of diagnosing leishmaniasis by virulence factor gene (Gp63) in leishmania promastigotes.

Upregulation of virulence genes promotes Vibrio cholerae biofilm hyperinfectivity

Vibrio cholerae remains a major global health threat, disproportionately impacting parts of the world without adequate infrastructure and sanitation resources. In aquatic environments, V. cholerae exists both as planktonic cells and as biofilms, which are held together by an extracellular matrix. V. cholerae biofilms have been shown to be hyperinfective, but the mechanism of hyperinfectivity is unclear. Here we show that biofilm-grown cells, irrespective of the surfaces on which they are formed, are able to markedly outcompete planktonic-grown cells in the infant mouse. Using an imaging technique designed to render intestinal tissue optically transparent and preserve the spatial integrity of infected intestines, we reveal and compare three-dimensional V. cholerae colonization patterns of planktonic-grown and biofilm-grown cells. Quantitative image analyses show that V. cholerae colonizes mainly the medial portion of the small intestine and that both the abundance and localization patterns of biofilm-grown cells differ from that of planktonic-grown cells. In vitro biofilm-grown cells activate expression of the virulence cascade, including the toxin coregulated pilus (TCP), and are able to acquire the cholera toxin-carrying CTXФ phage. Overall, virulence factor gene expression is also higher in vivo when infected with biofilm-grown cells, and modulation of their regulation is sufficient to cause the biofilm hyperinfectivity phenotype. Together, these results indicate that the altered biogeography of biofilm-grown cells and their enhanced production of virulence factors in the intestine underpin the biofilm hyperinfectivity phenotype.