| Anthrax is a zoonotic disease caused by Bacillus anthracis. Anthrax epidemic usually happened among herbivore, and human can be infected by contacting infected animal and their products or anthrax spore in air and soil. The anthrax toxins include edema toxin (ET) and lethal toxin (LT) which are critical virulence factors produced by Bacillus anthracis. ET is composed of protective antigen (PA) and edema factor (EF), while a mixture of PA and lethal factor (LF) forms LT. PA is the pivotal protein of the anthrax toxin complex and the major immunogen. Studies in animal models indicate that the immune response to PA is central to protection against Bacillus anthracis. The anti-PA immunoglobulin G (IgG) antibody concentrations and dilutional titers are, therefore, the most commonly reported marker of human immune responses to anthrax vaccines and B. anthracis infection. Many detection methods are based on protective antigen. Many serologic detection methods are based on this theory.In this study, the accuracy of the published anthrax toxin protein gene was verified by the theory that ribosomal binding site (RBS) plays an important role in the regulation of the initiation of gene translation and is often located in front of the initiation codon. The RBS sequence of Bacillus anthracis was confirmed to be GCACCUUCC, by comparing the16S rRNA sequence of published complete genome sequence of five Bacillus anthracis. It was found that, among the203annotated coding sequences (CDSs), RBS can be found in124of them(61.08%) in the plasmid pXO1of the strain’Ames ancestor’. The distance between RBS and the initiation codon is similar to the normal distribution (mean to7). By searching the high frequency RBS though the constrained condition of plasmid pXO1, we acquired many opening reading frames (ORF). Then by the comparison of these ORFs obtained and the annotated CDSs, we found the completed or partial coincidence rate was63.89%. And there are RBS both in front of the toxin protein genes and toxins regulated genes. So, it is concluded that the published toxin genes are accurate.In this study, Sera both of the clinical samples and healthy controls were detected by anthrax lethal toxin neutralization assay (TNA) and enzyme linked immunosorbent assay (ELISA). Among the131clinical samples, only45were positive (34.35%) by TNA. None of the healthy controls was positive. There was no significant difference in the seroprevalence between groups sorted by days after the onset of symptoms. Among these sera,83clinical samples and100healthy controls were detected by ELISA.76of the83clinical samples were positive (91.57%) and7of the100health controls were positive (7%). By comparing the results of TNA and ELISA, we got the following conclusions. The ELISA results of all the TNA-positive sera were positive and the TNA results of all the ELISA-negative sera were also negative. While, the ELISA result of some TNA-negative were positive. The ELISA titers of the TNA-positive sera are higher than the TNA-negative ones. With regard to the45sera of TNA-positive and ELISA-positive, anti-PA IgG antibody titer and ED50TNA titer was correlated (R2=0.461) indicates that LT neutralization activity correlated with anti-PA IgG.This study provides a reference for the establishment of the anthrax toxin protein expression system and acquires the data of anthrax toxin-neutralizing antibody level of anthrax clinical cases first. The result of this study provides the scientific guidance for the serological diagnosis of anthrax and scientific basis for determining immune protective effect of the key areas and high-risk groups of anthrax surveillance in our country. |
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