A novel bacterium-like particles platform displaying antigens by new anchoring proteins induces efficacious immune responses.

Bacterium-like particles (BLP) are the peptidoglycan skeleton particles of lactic acid bacteria, which have high safety, mucosal delivery efficiency, and adjuvant effect. It has been widely used in recent years in the development of vaccines. Existing anchoring proteins for BLP surfaces are few in number, so screening and characterization of new anchoring proteins are necessary. In this research, we created the OACD (C-terminal domain of Escherichia coli outer membrane protein A) to serve as an anchoring protein on the surface of BLP produced by the immunomodulatory bacteria Levilactobacillus brevis 23017. We used red fluorescent protein (RFP) to demonstrate the novel surface display system's effectiveness, stability, and ability to be adapted to a wide range of lactic acid bacteria. Furthermore, this study employed this surface display method to develop a novel vaccine (called COB17) by using the multi-epitope antigen of Clostridium perfringens as the model antigen. The vaccine can induce more than 50% protection rate against C. perfringens type A challenge in mice immunized with a single dose and has been tested through three routes. The vaccine yields protection rates of 75% for subcutaneous, 50% for intranasal, and 75% for oral immunization. Additionally, it elicits a strong mucosal immune response, markedly increasing levels of specific IgG, high-affinity IgG, specific IgA, and SIgA antibodies. Additionally, we used protein anchors (PA) and OACD simultaneous to show several antigens on the BLP surface. The discovery of novel BLP anchoring proteins may expand the possibilities for creating mucosal immunity subunit vaccines. Additionally, it may work in concert with PA to provide concepts for the creation of multivalent or multiple vaccines that may be used in clinical practice to treat complex illnesses.

Effects of Catecholamine Stress Hormones Norepinephrine and Epinephrine on Growth, Antimicrobial Susceptibility, Biofilm Formation, and Gene Expressions of Enterotoxigenic Escherichia coli.

Enterotoxigenic Escherichia coli (ETEC) is a significant contributor to diarrhea. To determine whether ETEC-catecholamine hormone interactions contribute to the development of diarrhea, we tested the effects of catecholamine hormones acting on ETEC in vitro. The results showed that in the presence of norepinephrine (NE) and epinephrine (Epi), the growth of 9 out of 10 ETEC isolates was promoted, the MICs of more than 60% of the isolates to 6 antibiotics significantly increased, and the biofilm formation ability of 10 ETEC isolates was also promoted. In addition, NE and Epi also significantly upregulated the expression of the virulence genes feaG, estA, estB, and elt. Transcriptome analysis revealed that the expression of 290 genes was affected by NE. These data demonstrated that catecholamine hormones may augment the diarrhea caused by ETEC.

Protective Effect of Levilactobacillus brevis Against Yersinia enterocolitica Infection in Mouse Model via Regulating MAPK and NF-κB Pathway.

Although the use of the probiotic bacterium Lactobacillus for the treatment and prevention of diseases caused by various pathogenic bacteria has received increasing attention in recent years, its mechanism remains incompletely understood. Levilactobacillus brevis 23017 is a select probiotic strain that can regulate the immunity of host animals and resist pathogen infections. In this study, we analyzed the effect of L. brevis 23017 on Yersinia enterocolitica intestinal infection in a BALB/c mouse model and discussed its underlying mechanism. We found that in the mouse model, L. brevis 23017 prevented the damage of villi in the small intestine and decelerated weight loss after Y. enterocolitica infection. Moreover, we focused on the mechanism of the protective effect of L. brevis 23017 from the perspective of the damage and repair of the intestinal mucosal barrier. We observed that L. brevis 23017 maintained a normal mucosal barrier by altering the expression of tight junction proteins. Notably, our results indicated that L. brevis 23017 effectively promoted the secretion of the intestine-specific secretory immunoglobulin A (SIgA) by B cells via regulating cytokines and oxidative damage levels. This mechanism may be the reason for its protective role in Y. enterocolitica infection. In addition, our results demonstrated that the mechanism of L. brevis 23017 was related to antibacterial colonization and inflammation regulation and closely related to antioxidative stress and SIgA promotion. The protective effect of L. brevis 23017 on mice was related to the signaling pathway protein p38 MAPK and the phosphorylation levels of NF-κB. Our study provided novel insight into the mechanism of Lactobacillus against pathogenic bacterial infections. Such insight is of great importance for the prevention, diagnosis, and treatment of related diseases.

Genetic and evolutionary analysis of emerging HoBi-like pestivirus.

HoBi-like pestivirus, an emerging species within the Pestivirus genus, is an important pathogen associated with a variety of clinical manifestations of ruminants, especially cattle. HoBi-like pestiviruses were identified in several countries and from different hosts, and raised concerns with regard to their acute and persistent infections, which is implicated in economic losses for cattle farmers. However, the transmission path, codon usage bias, and host adaptation of the virus has not been studied. Hence, we performed the analysis the spatio-temporal transmission based on the available 5'-UTR sequences of HoBi-like pestivirus, and then conducted codon analysis of the complete coding sequence of the virus. The results show the virus appeared in 1952 (95% HPD: 1905-1985) and may have originated in India. In addition, Italy is the hub for the spread of the virus. Moreover, six potential recombination events and two complex recombination events were discovered. Analysis of codon usage patterns revealed that the effective number of codon (ENC) values with an average of 50.85, and the codon usage bias is greatly affected by natural selection, which is different from the previous BVDV-1, 2. Finally, codon adaptation index (CAI) analysis shows that pigs may be the potential origin species of the HoBi-like pestivirus. These findings will contribute to more effective control of the spread of the virus, extend the knowledge about the genetic and evolutionary features of HoBi-like viruses and provide some information for vaccine research.

Detection, genetic, and codon usage bias analyses of the VP2 gene of mink bocavirus.

Mink bocavirus 1 (MiBoV1), a novel virus detected from the feces of domestic minks in China in 2016, may be related to gastrointestinal diseases. However, its prevalence and genetic characteristics are poorly described. In this study, we examined 192 samples collected from minks in the major mink industry province from northern China. PCR results showed that 10 samples (5.2%) were positive for MiBoV1, and 60% of MiBoV1-positive samples were co-infected with Aleutian mink disease virus or mink circovirus. MiBoV1 was detected in six serum samples. Sequence analysis demonstrated that the VP2 gene of MiBoV1 was highly conserved and had low viral diversity over the VP2 region and unique nucleotide mutations. Phylogenetic analysis of the VP2 sequence demonstrated that MiBoV1 strains formed two clades and were grouped with California sea lion bocavirus, Canine bocavirus, and Feline bocavirus. Codon usage analysis revealed that most of the preferentially used codons in MiBoV1 were A- or U-ended codons, and no evident codon usage bias was found. This study provides evidence that MiBoV1 has a low prevalence in Jilin and Hebei provinces in China. Moreover, it contributes information regarding the expansion of the limited mink bocavirus sequence and determines the codon usage bias of the VP2 gene for the first time. Epidemiological surveillance is necessary to understand the importance and evolution of MiBoV1.

Detection and molecular characterization of canine circovirus circulating in northeastern China during 2014-2016.

Canine circovirus (canine CV) is an etiological agent associated with diarrhea, hemorrhagic gastroenteritis and vasculitis. Although canine CV has been identified and characterized in southern China in recent years, its epidemiology in other regions of China and its precise molecular characteristics have not been examined. In this study, we examined 141 fecal specimens collected from domestic dogs with or without diarrhea in Heilongjiang province, Northeastern China, during 2014 to 2016. A total of 18 out of 141 samples were found to be positive for canine CV by real-time quantitative PCR. In the diarrhea samples, canine CV was detected in coinfections with canine parvovirus 2. More importantly, two different canine CV strains were detected in one sample. Five canine CV genomes were successfully amplified. Sequence analysis showed that there were two unique amino acid changes in the Rep protein (N39S in the K1 strain, and T71A in the XF16 strain). Phylogenetic analysis indicated that canine CV could be divided into four genotypes, and specific nucleotide mutations could be used for confirming the four genotypes. Moreover, recombination analysis revealed that a total of eight recombination events were found in five genomic sequences. Molecular evolution analysis showed that the canine CV has been under purifying selection. This study provides evidence that at least three genotypes of canine CV are co-circulating in China. Continuous epidemiological surveillance is therefore necessary to understand their importance for the evolution of canine CV.