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PubMed2002

DNA sequence conservation between the Bacillus anthracis pXO2 plasmid and genomic sequence from closely related bacteria.

BACKGROUND: Complete sequencing and annotation of the 96.2 kb Bacillus anthracis plasmid, pXO2, predicted 85 open reading frames (ORFs). Bacillus cereus and Bacillus thuringiensis isolates that ranged in genomic similarity to B. anthracis, as determined by amplified fragment length polymorphism (AFLP) analysis, were examined by PCR for the presence of sequences similar to 47 pXO2 ORFs. RESULTS: The two most distantly related isolates examined, B. thuringiensis 33679 and B. thuringiensis AWO6, produced the greatest number of ORF sequences similar to pXO2; 10 detected in 33679 and 16 in AWO6. No more than two of the pXO2 ORFs were detected in any one of the remaining isolates. Dot-blot DNA hybridizations between pXO2 ORF fragments and total genomic DNA from AWO6 were consistent with the PCR assay results for this isolate and also revealed nine additional ORFs shared between these two bacteria. Sequences similar to the B. anthracis cap genes or their regulator, acpA, were not detected among any of the examined isolates. CONCLUSIONS: The presence of pXO2 sequences in the other Bacillus isolates did not correlate with genomic relatedness established by AFLP analysis. The presence of pXO2 ORF sequences in other Bacillus species suggests the possibility that certain pXO2 plasmid gene functions may also be present in other closely related bacteria.

Journal Article↗
PubMed1995

Detection of cilia-associated respiratory bacillus in experimentally and naturally infected mice and rats by the polymerase chain reaction.

Cilia-associated respiratory (CAR) bacillus was detected by means of the reverse transcription (RT)-polymerase chain reaction (PCR), and the results were compared with those of indirect immunofluorescence test (IFAT) for the detection of the organism. In the experimental infections, 15 mice were in contact with mice previously inoculated with CAR bacillus. Three mice each were tested at days 3, 5, 7, 12 and 20 postexposure. On day 3 postexposure, CAR bacillus was detected in oral swab samples from all 3 mice by RT-PCR, but was not detected in any sampling sites from the mice by IFAT. Total numbers of positive samples from nasal, oral and tracheal swabs obtained through the test were 6/15, 14/15 and 8/15, respectively, by RT-PCR, and 2/15, 6/15 and 3/15, respectively by IFAT. For the detection of CAR bacillus in samples from 52 rats, 34 serum antibody negative rats by enzyme-linked immunosorbent assay were also negative by RT-PCR and IFAT except for one sample from the oral cavity, and all serum antibody positive rats were positive for the organism by RT-PCR but it could not be detected in five of them by IFAT. By means of RT-PCR, no differences in the positive rates depending on sampling sites were observed except in one rat. The RT-PCR was found to be a specific, highly sensitive and reliable procedure for detecting CAR bacillus in mice and rats. The oral cavity was the most suitable site for the diagnosis of the early stage of this infection by RT-PCR.

Animals↗
PubMed2002

Prognostic value of p53 overexpression in bladder tumors treated with Bacillus Calmette-Guerin.

Bacillus Calmette-Guerin intravesical therapy is the standard treatment of superficial bladder tumors at high risk of recurrence and progression to muscle-invasion disease. To date, there is no well established predictive factor of response to Bacillus Calmette-Guerin intravesical therapy. The prognostic value of p53 overexpression in bladder tumors is controversial. Most investigators have found no correlation between p53 status assessed before Bacillus Calmette-Guerin intravesical therapy and patient outcome. On the other hand, it is acknowledged that the persistence of p53 overexpression after Bacillus Calmette-Guerin intravesical therapy is predictive of progression in patients treated for carcinoma in situ. Since conflicting data have been reported, further evaluation of the impact of p53 overexpression in patients treated with Bacillus Calmette-Guerin intravesical therapy for bladder carcinoma is required.

Adjuvants, Immunologic↗
PubMed2003

Combined bacillus Calmette-Guerin and interferon use in superficial bladder cancer.

Due to its high rate of local recurrence after transurethral surgery, superficial bladder cancer is often treated with adjuvant topical intravesical chemotherapy or biological agents, such as bacillus Calmette-Guerin or interferon-alpha. Recent laboratory testing has revealed that combination interferon-alpha with bacillus Calmette-Guerin results in remarkable synergy, affecting not only bladder tumors directly but also enhancing the immune response to bacillus Calmette-Guerin by orders of magnitude. Clinical studies are now demonstrating lower toxicity with combination low-dose bacillus Calmette-Guerin/interferon regimens, while providing a much needed salvage option for previous bacillus Calmette-Guerin failures.

Animals↗
PubMed2001

Cilia-Associated Respiratory (CAR) bacillus infection in veal calves and adult cattle.

Cilia-Associated Respiratory (CAR) bacillus is a filamentous bacterium that colonizes the ciliated epithelium of the respiratory tract of many animal species and that has been associated with chronic inflammatory lesions in naturally and experimentally infected rats, mice and rabbits. In the present study, the prevalence of CAR bacillus infection and histological lesions of the trachea in veal calf and adult cattle were investigated. Forty five healthy veal calves and 45 adult cattle, raised in 18 different herds were selected at slaughter. From each animal, a tracheal sample was processed for histology, stain-ed with the Warthin-Starry method to evaluate the presence of CAR bacillus, and with haematoxylin and eosin to evaluate the presence of inflammatory lesions. CAR bacillus was identified in 17 veal calves (37.7%) and in 7 adult cattle (15.5%). Inflammatory lesions were found in 42 veal calves (93.3%) and in 41 adult cattle (91.1%). Statistical analysis pointed out a significant correlation between the presence and number of CAR bacilli and the presence and number of lymphoid follicles (P = 0.0071) and the presence and severity of neutrophilic infiltrates (P = 0.0428). These results indicate that CAR bacillus infection is common in cattle and is correlated with tracheal inflammatory lesions.

Animals↗
PubMed1994

Effectiveness of sealed dental prophylaxis angles inoculated with Bacillus stearothermophilus in preventing leakage.

It was the purpose of this in vitro investigation to evaluate the effectiveness of several brands of sealed, reusable prophylaxis angles to keep internal materials within the internal portions of the head of the prophylaxis angle, and not allowing contaminates to leak out. Three brands of sealed, reusable dental prophylaxis angles were autoclaved and then taken apart under a biocontainment flow hood. Testing conditions were designed to prevent a "worst case scenario" by inoculating dental prophylaxis angles with 10(6) of the heat resistant spores of Bacillus stearothermophilus and 20% bovine serum albumin to simulate the presence of human serum. The concentration of Bacillus stearothermophilus spores was verified before testing procedures were initiated. The internal portions of the sterile prophylaxis angles were inoculated with a 1:1 mixture of the Bacillus stearothermophilus spores and bovine serum albumin, at a concentration of 1.15 x 10(6) spores/inoculation. The prophylaxis angles were reassembled under sterile conditions, and a sterile rubber cup was inserted into each of the prophylaxis angles. The prophylaxis angles were attached to a sterile dental handpiece and then submerged in a 50 ml tube containing sterile trypticase soy broth and run at 3000 rpm for 30 seconds. The tube of medium was incubated for 7 days. No growth of Bacillus stearothermophilus spores could be cultured from one of the brands of prophylaxis angles at any time during the incubation period. The other two brands of prophylaxis angles did produce some leakage of the Bacillus stearothermophilus spores.

Cross Infection↗
PubMed1995

Prevention and treatment of cilia-associated respiratory bacillus in mice by use of antibiotics.

The effects of continuous oral administration of antibiotics in mice were investigated. Sulfamerazine, ampicillin, and chlortetracycline were tested at a rate of 500 mg/L of drinking water. Mice were infected by intranasal inoculation with 10(6) bacilli of the SMR strain of cilia-associated respiratory (CAR) bacillus. The mice were treated with the antibiotics starting 1 week before, 1 week after, or 4 weeks after the inoculation, for 5, 3, or 4 weeks respectively, then were examined. The infected mice lost body weight, and this loss was prevented or regained by all of the antibiotic treatments. Serologically no antibodies were detectable in the mice administered sulfamerazine starting 1 week before the inoculation. Mice administered sulfamerazine starting 1 week after the inoculation and ampicillin or chlortetracycline starting 1 week before or after the inoculation yielded a low titer of antibodies compared with nontreated infected mice. Mice administered antibiotics starting 4 weeks after the inoculation yielded the same titer of antibodies as nontreated infected mice. No pathologic respiratory tract lesions were observed in mice administered sulfamerazine starting 1 week before the inoculation. Mice administered sulfamerazine starting 1 week after the inoculation or ampicillin starting 1 week before or after the inoculation had slight peribronchitis without CAR bacillus colonization. Mice administered chlortetracycline, starting either 1 week before or after inoculation, developed peribronchitis, with colonization of the bacillus on the airway mucosa. In mice medicated starting 4 weeks after the inoculation, respiratory tract lesions developed, but their severity was reduced. The airway mucosa in mice treated with chlortetracycline was associated with the CAR bacillus but not in mice treated with sulfamerazine and ampicillin. These findings suggest that prevention and eradication of CAR bacillus infection is possible by treatment with sulfamerazine.

Ampicillin↗
PubMed1977

Restriction and modification in Bacillus species: genetic transformation of bacteria with DNA from different species, part I.

Host specific restriction was detected in 13 Bacillus strains, when 63 strains of Bacillus subtilis and 15 other Bacillus strains were tested with phage phi 105C. These 13 strains were classified into 8 groups (M,H,C,N,E,F,G,P) by the type of restriction. M-type strains (B. subtilis Marburg 168, its derivatives, and two other strains) showed relatively weak restriction, restricting phi 105C from other groups of Bacillus by ratios of 10(-1) to 10(-3). Strains of groups H,C,N,E,F,G, and P restricted phi 105C from other groups by ratios of 10(-2) to 10(-8). It was confirmed with some of the strains that type-specific modification was endowed only by the last host. Furthermore, we isolated one restriction deficient mutant of B. subtilis Marburg 168-YS11, which had also lost its modification phenotype.

Bacillus↗
PubMed1980

Molecular cloning of heterologous chromosomal DNA by recombination between a plasmid vector and a homologous resident plasmid in Bacillus subtilis.

The difficulty experienced in the shotgun cloning of chromosomal DNA on plasmid vectors in Bacillus subtilis is analyzed and an explanation for this difficulty is offered based on an inherent property of competent cells which imposes a requirement of plasmid multimers in transformation of plasmid-free recipients (Canosi et al., 1978). A stratagem which uses cloning by recombination between the vector and a resident homologous plasmid is tested and shown to be successful. Several recombinant plasmids are obtained containing Bacillus licheniformis DNA fragments which complement aromatic amino acid mutants of Bacillus subtilis. The yield of recombinant clones ranges from 6.7 to 210 per microgram of chromosomal DNA, depending on the selection and the restriction endonuclease. The various trp clones obtained after cutting chromosomal DNA with BglII and BclI do not complement trpE and exhibit both orientations with respect to the vector. The location of several restriction endonuclease cleavage sites in the cloned trp fragments is presented, and their relationship to the genetic map of Bacillus licheniformis is described.

Bacillus↗
PubMed1984

Enzyme-linked lectinosorbent assay (ELLA) for detecting Bacillus anthracis.

An enzyme-linked lectinosorbent assay (ELLA) has been developed for the rapid identification of Bacillus anthracis. Using two different lectin-conjugates, the ELLA test can differentiate Bacillus anthracis from closely related species, such as Bacillus cereus and Bacillus cereus var. mycoides, in approximately two hours. In addition to having high specificity, the test can also detect small numbers of the bacterium.

Bacillus↗
PubMed2002

Sporulation and delta-endotoxin synthesis by Bacillus thuringiensis.

Bacillus thuringiensis is distinguished from the very closely related Bacillus cereus and Bacillus anthracis by the presence of several plasmid-encoded delta-endotoxin genes. These delta-endotoxins, synthesized as protoxins, are produced in large quantities during sporulation and are packaged into intracellular inclusions. Ingestion of the inclusions by insect larvae leads to protoxin solubilization and conversion to toxins each specific for one of several orders of insects. The toxins form cation-selective channels in the membrane of cells lining the larval midgut with subsequent lethality. In most cases, delta-endotoxin synthesis and sporulation are closely coupled. The latter process in B. thuringiensis is probably virtually identical to that in Bacillus subtilis with the additional use of mother cell sporulation forms of RNA polymerase for the synthesis of the delta-endotoxins. There are other more subtle plasmid-encoded functions or plasmid interactions related to regulating protoxin synthesis. Consideration of both plasmid and chromosomal genes is thus critical for defining this organism.

Bacillus thuringiensis↗
PubMed2005

Molecular cloning and characterization of a novel gamma-CGTase from alkalophilic Bacillus sp.

We found a novel cyclodextrin glucanotransferase (CGTase) from alkalophilic Bacillus sp. G-825-6. The enzyme was expressed in the culture broth by recombinant Bacillus subtilis KN2 and was purified and characterized. The enzyme named CGTase825-6 showed 95% amino acid sequence identity with a known enzyme beta-/gamma-CGTase from Bacillus firmus/lentus 290-3. However, the product specificity of CGTase825-6 differed from that of beta-/gamma-CGTase. CGTase825-6 produced gamma-cyclodextrin (CD) as the main product, but degradation of gamma-CD was observed with prolonged reaction. The product specificity of the enzyme was positioned between gamma-CGTase produced by Bacillus clarkii 7364 and B. firmus/lentus 290-3 beta-/gamma-CGTase. It showed that the difference of product specificity was dependent on only 28 amino acid residues in 671 residues in CGTase825-6. We compared the amino acid sequence of CGTase825-6 and those of other CGTases and constructed a protein structure model of CGTase825-6. The comparison suggested that the diminished loop (Val138-Asp142) should provide subsite -8 for gamma-CD production and that Asp142 might have an important role in product specificity. CGTase825-6 should be a useful tool to produce gamma-CD and to study the differences of producing mechanisms between gamma-CD and beta-CD.

Amino Acid Sequence↗
PubMed2000

Cloning of the cyclodextrin glucanotransferase gene from alkalophilic Bacillus sp. A2-5a and analysis of the raw starch-binding domain.

The cyclodextrin glucanotransferase (CGTase) gene of alkalophilic Bacillus sp. A2-5a was cloned and expressed in Bacillus subtilis ANA-1 as a host. The DNA region included an open reading frame encoding a 704-amino-acid polypeptide with a typical raw starch-binding motif in its C-terminal region. The CGTase purified from Bacillus sp. A2-5a bound to raw starch as strongly as porcine pancreas alpha-amylase, as expected from the sequence motif. A chromosomal region (a DNA fragment of about 14.1 kbp) including the CGTase gene was also cloned and the nucleotide sequence was determined. Possible cyclodextrinase and putative cyclodextrin-binding protein genes were found in the flanking region of the CGTase gene, which implied that the novel starch-degradation pathway postulated for a gram-negative bacterium [Klebsiella oxytoca; Fiedler et al. (1996) J Mol Biol 256: 279-291] also exists in a gram-positive bacterium i.e. Bacillus.

Amino Acid Sequence↗
PubMed2004

Rapid detection and characterization of surfactin-producing Bacillus subtilis and closely related species based on PCR.

The surfactin production genetic locus ( sfp) is responsible for the ability of Bacillus subtilis to produce the lipopeptide biosurfactant, surfactin. This report demonstrates the utility of using PCR of the sfp gene as a means of identifying Bacillus species that produce surfactin. We carried out a hemolysis zone assay, quantitative HPLC and NMR in parallel to ensure that the PCR provided correct results. PCR analyses were performed for the sfp gene on 15 standard strains and 20 field-collected Bacillus spp. isolates native to Taiwan. Among the 15 standard strains, surfactin was produced by seven strains of B. subtilis and two closely related species, B. amyloliquefaciens B128 and B. circulans ATCC 4513. Of the 20 field-collected Bacillus spp. isolates; 16 strains yielded surfactin- positive results with PCR and HPLC. A good correlation was observed. Within the 16 field isolates, B. amyloliquefaciens S13 (452.5 mg/L) and B. subtilis S15 (125.6 mg/L) had high productivity of surfactin. The technique is valuable for finding out potential good yields of surfactin-producing strains. The PCR method we used could also be used to find different species or genera containing homologous genes. This is the first report of the detection of surfactin production by B. amyloliquefaciens and B. circulans based on PCR screening.

Bacillus↗
PubMed1990

Purification and characterization of Bacillus thuringiensis var. tenebrionis insecticidal proteins produced in E. coli.

Native and single amino acid variants of the Bacillus thuringiensis var. tenebrionis insecticidal proteins were expressed in Escherichia coli, purified and examined for biological and biochemical properties. A novel, pH dependent, preferential precipitation method was implemented to purify Escherichia coli produced Bacillus thuringiensis var. tenebrionis proteins, which are active against Colorado potato beetle (Leptinotarsa decemlineata) larvae. Cysteine residues of the native Bacillus thuringiensis var. tenebrionis protein were replaced by serine residues by site-directed mutagenesis to investigate the biological and structural importance of the individual cysteine residues. Sulfhydryl determination of the native and amino acid variant Bacillus thuringiensis var. tenebrionis proteins revealed that the native protein contains no disulfide bonds. Modification of the carboxyl terminal cysteine residue (amino acid 540) caused complete inactivation of the protein. Native, truncated and single amino acid variants (other than at amino acid 540) exhibited insecticidal activities comparable to each other and to solubilized crystals from the original strain.

Animals↗
PubMed1990

Efficiency of transcriptional terminators in Bacillus subtilis.

To promote more efficient synthesis of heterologous gene products in a Bacillus subtilis host, we have developed a system for rapidly testing the effect of a putative terminator on in vivo gene expression. Terminator structures from the Bacillus amyloliquefaciens amyE gene, the Bacillus licheniformis penP gene, the B. subtilis bglS gene, and the Bacillus thuringiensis cry gene were subcloned and inserted into a vector in such a way as to disrupt expression of the cat-86 gene. Comparisons are made between gene expression levels and the stabilities of the respective stem-loop structures.

Bacillus subtilis↗
PubMed2006

N-thiolated beta-lactams: a new family of anti-Bacillus agents.

This report describes the evaluation of N-thiolated beta-lactam antibiotics as potential anti-Bacillus agents. N-Thiolated beta-lactams are a new family of antibacterials that previously have been found to selectively inhibit the growth of Staphylococcus bacteria over many other genera of microbes. From the data presented herein, these lactams similarly inhibit a variety of Bacillus species, including Bacillus anthracis. The preliminary structure-activity studies suggest that there is a need to balance the lipophilic character of the C3/C4 groups in order to obtain optimal anti-Bacillus activity. Elongation or extensive branching of the organothio substituent diminishes antibacterial effects, with the sec-butylthio derivative providing the strongest activity.

Anti-Bacterial Agents↗
PubMed2026

Spatially resolved multi-omics analysis of indigenous Bacillus-fortified high-temperature Daqu.

Layer-dependent patterns associated with indigenous Bacillus fortification on high-temperature Daqu remain unclear. Here, six indigenous functional Bacillus strains were combined to fortify Daqu at three inoculation levels (QH4, QH5, QH6), with non-fortified as the control (CK). Upper, middle, and lower shelf-layer samples were profiled by physicochemical measurements, volatilomics, organic acid analysis, untargeted metabolomics, 16S/ITS amplicon sequencing, and metagenomics. PERMANOVA showed significant effects of treatment, spatial layer, and their interaction on physicochemical, volatile, bacterial, and fungal profiles (P = 0.001). Among the three inoculation levels, QH5 showed the most balanced performance: QH5_M exhibited the highest observed mean peak temperature (63.3 °C; +4.5 °C relative to CK_M), and its group-mean temperature remained ≥ 60 °C for seven consecutive days. Multi-omics analyses indicated coordinated, non-linear, and layer-dependent differences associated with indigenous Bacillus fortification, with QH5_M showing the most pronounced combined thermal, pyrazine, substrate, microbial, and predicted functional profile. These findings indicate that moderate indigenous Bacillus fortification was associated with distinct layer-dependent thermal and flavor profiles and coordinated microbial, metabolic, and predicted functional differences.

Bacillus↗