The Bacillus anthracis chromosome contains four conserved, excision-proficient, putative prophages

BACKGROUND: Bacillus anthracis is considered to be a recently emerged clone within the Bacillus cereus sensu lato group. The B. anthracis genome sequence contains four putative lambdoid prophages. We undertook this study in order to understand whether the four prophages are unique to B. anthracis and whether they produce active phages. RESULTS: More than 300 geographically and temporally divergent isolates of B. anthracis and its near neighbors were screened by PCR for the presence of specific DNA sequences from each prophage region. Every isolate of B. anthracis screened by PCR was found to produce all four phage-specific amplicons whereas none of the non-B. anthracis isolates, produced more than one phage-specific amplicon. Excision of prophages could be detected by a PCR based assay for attP sites on extra-chromosomal phage circles and for attB sites on phage-excised chromosomes. SYBR-green real-time PCR assays indicated that prophage excision occurs at very low frequencies (2 × 10(-5 )- 8 × 10(-8)/cell). Induction with mitomycin C increased the frequency of excision of one of the prophages by approximately 250 fold. All four prophages appear to be defective since, mitomycin C induced culture did not release any viable phage particle or lyse the cells or reveal any phage particle under electron microscopic examination. CONCLUSION: The retention of all four putative prophage regions across all tested strains of B. anthracis is further evidence of the very recent emergence of this lineage and the prophage regions may be useful for differentiating the B. anthracis chromosome from that of its neighbors. All four prophages can excise at low frequencies, but are apparently defective in phage production

Paring Down HIV Env: Design and Crystal Structure of a Stabilized Inner Domain of HIV-1 gp120 Displaying a Major ADCC Target of the A32 Region

SummaryEvidence supports a role of antibody-dependent cellular cytotoxicity (ADCC) toward transitional epitopes in the first and second constant (C1-C2) regions of gp120 (A32-like epitopes) in preventing HIV-1 infection and in vaccine-induced protection. Here, we describe the first successful attempt at isolating the inner domain (ID) of gp120 as an independent molecule that encapsulates the A32-like region within a minimal structural unit of the HIV-1 Env. Through structure-based design, we developed ID2, which consists of the ID expressed independently of the outer domain and stabilized in the CD4-bound conformation by an inter-layer disulfide bond. ID2 expresses C1-C2 epitopes in the context of CD4-triggered full-length gp120 but without any known neutralizing epitope present. Thus, ID2 represents a novel probe for the analysis and/or selective induction of antibody responses to the A32 epitope region. We also present the crystal structure of ID2 complexed with mAb A32, which defines its epitope

Cholera Toxin and Heat-Labile Enterotoxin Activate Human Monocyte-Derived Dendritic Cells and Dominantly Inhibit Cytokine Production through a Cyclic AMP-Dependent Pathway

Cholera toxin (CT) and heat-labile enterotoxin (LT) are powerful mucosal adjuvants whose cellular targets and mechanism of action are unknown. There is emerging evidence that dendritic cells (DC) are one of the principal cell types that mediate the adjuvant effects of these toxins in vivo. Here we investigate the effects of CT and LT on the maturation of human monocyte-derived DC (MDDC) in vitro. We found that an enzymatically active A domain is necessary for both CT and LT to induce the maturation of MDDC and that this activation is strictly cyclic AMP (cAMP) dependent. ADP-ribosylation-defective derivatives of these toxins failed to induce maturation of MDDC, whereas dibutyryl-cyclic-3′,5′-AMP and Forskolin mimic the maturation of MDDC induced by CT and LT. In addition, an inhibitor of cAMP-dependent kinases, Rp-8-Br-cAMPs, blocked the ability of CT, LT, and Forskolin to activate MDDC. CT, LT, dibutyryl-cyclic-3′,5′-AMP, and Forskolin also dominantly inhibit interleukin 12 and tumor necrosis factor alpha production by MDDC in the presence of saturating concentrations of lipopolysaccharide. Taken together, these results show that the effects of CT and LT on MDDC are mediated by cAMP

Sequencing Bacillus anthracis Typing Phages Gamma and Cherry Reveals a Common Ancestry

The genetic relatedness of the Bacillus anthracis typing phages Gamma and Cherry was determined by nucleotide sequencing and comparative analysis. The genomes of these two phages were identical except at three variable loci, which showed heterogeneity within individual lysates and among Cherry, Wβ, Fah, and four Gamma bacteriophage sequences