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PubMed2004

Laboratory and simulated field evaluation of a new recombinant of Bacillus thuringiensis ssp. israelensis and Bacillus sphaericus against Culex mosquito larvae (Diptera: Culicidae).

In the laboratory, three microbial mosquito larvicidal products consisting of Bacillus thuringiensis ssp. israelensis de Barjac (Bti), Bacillus sphaericus (Neide) (Bsph) (strain 2362), and the University of California Riverside (UCR) recombinant (producing toxins of both Bacillus sphaericus and Bacillus thuringiensis ssp. israelensis) were bioassayed against larvae of Culex quinequefasciatus Say (susceptible and resistant to Bsph 2362), and Aedes aegypti (L.). Bti proved highly effective against Cx. Quinequefasciatus susceptible and resistant strains, with LC50 values of 0.009 and 0.011 ppm and LC90 values of 0.057 and 0.026 ppm for Bsph-susceptible and -resistant strains, respectively. Bti was also highly active against Ae. eagypti with LC50 and LC90 values of 0.014 and 0.055 ppm, respectively. The UCR recombinant was equally active against both Bsph-susceptible and -resistant strains of Cx. Quinquefasciatus; LC50 values were 0.005 and 0.009 and LC90 values were 0.030 and 0.043 ppm, respectively. Bti and the UCR recombinant essentially showed similar activity against Bsph-susceptible and -resistant strains. UCR recombinant showed high toxicity against Ae. eagypti with LC50 and LC90 values of 0.023 and 0.064 ppm, respectively. Bsph was highly active against susceptible strain of Cx. quinequefasciatus with LC50 and LC9o values of 0.006 and 0.024 ppm, respectively. Bsph exhibited little toxicity against Ae. eagypti larvae and also no toxicity to Bsph resistance. In the field, we evaluated four experimental corn grit formulations of Bti (VBC 60021), Bsph (VBC 60022), UCR recombinants VBC 60023 (7.89%), and VBC 60024 (1.87%) in simulated field (microcosms) against Bsph-susceptible Culex mosquitoes. Bti and low-concentrate UCR recombinant showed similar initial activity as well as persistence. Both materials provided high-to-moderate level of control for 2-7 d posttreatment at low treatment rates. At low dosages, residual activity of Bti and UCR recombinant lasted for <7 d. Bsph and high-concentrate UCR recombinant (VBC 60023), however, were more effective against natural populations of Cullex and achieved longer control (7-21 d) than the other two materials.

Animals↗

PubMed2006

Evaluation of chlorine, chlorine dioxide, and a peroxyacetic acid-based sanitizer for effectiveness in killing Bacillus cereus and Bacillus thuringiensis spores in suspensions, on the surface of stainless steel, and on apples.

Chlorine (10 to 200 microg/ml), chlorine dioxide (10 to 200 microg/ml), and a peroxyacetic acid-based sanitizer (40 and 80 microg/ ml) were evaluated for effectiveness in killing spores of Bacillus cereus and Bacillus thuringiensis in suspensions and on the surface of stainless steel and apples. Water and 5% horse serum were used as carriers for spore inoculum applied to the surface of stainless steel coupons, and 5% horse serum was used as a carrier for inoculum applied to apples. Inocula were dried on stainless steel for 5 h and on apples for 22 to 24 h before treating with sanitizers. At the concentrations of sanitizers tested, sensitivities of planktonic B. cereus and B. thuringiensis spores were similar. A portion of the spores surviving treatment with chlorine and, more markedly, chlorine dioxide had decreased tolerance to heat. Planktonic spores of both species were more sensitive to sanitizers than were spores on the surface of stainless steel or apples. At the same concentrations, chlorine was more effective than chlorine dioxide in killing spores in suspension and on stainless steel. The lethality of chlorine dioxide was markedly reduced when inoculum on stainless steel coupons was suspended in 5% horse serum as a carrier rather than water. Chlorine and chlorine dioxide at concentrations of 10 to 100 microg/ml were equally effective in killing spores on apples. Significant reductions of > or = 3.8 to 4.5 log CFU per apple were achieved by treatment with 100 microg/ml of either of the two sanitizers. The peroxyacetic acid sanitizer (40 and 80 microg/ml) was ineffective in killing Bacillus spores in the test systems investigated. Results provide information on the effectiveness of sanitizers commonly used in the food processing industry in killing Bacillus spores in suspension, on a food-contact surface, and on a ready-to-eat food.

Bacillus cereus↗

PubMed1975

Estimates of the porosity of Bacillus licheniformis and Bacillus subtilis cell walls.

The maximum porosity of Bacillus subtilis and Bacillus licheniformis cell walls was estimated by two independent and relatively simple methods. Peptidoglycan was isolated from Bacillus subtilis cell wall preparations and used as an insoluble support for exclusion chromatography of dextrans of known average molecular size. In an alternative approach the leakage of radioactively labelled proteins from Bacillus licheniformis cells incubated in butanol-saline mixtures was measured and their size estimated by exclusion chromatography on Sephadex G-100. Under these conditions the permeability barrier of the cytoplasmic membrane was destroyed with preservation of the structural integrity of the outer cell wall. The apparent exclusion threshold of the cell wall of either organism as determined by these means corresponded to molecules with a diffusional radius of not more than 2.5 nm.

Alanine↗

PubMed2005

Non-larvicidal effects of Bacillus thuringiensis israelensis and Bacillus sphaericus on oviposition and adult mortality of Culex quinquefasciatus Say (Diptera: Culicidae).

Two microbial mosquito larvicides, Bacillus thuringiensis ssp. israelensis (Bti) and Bacillus sphaericus (Bsph), have been shown to be highly effective in controlling mosquito larvae and have been used in larvicidal programs for many years. In exploring other modes of action of these agents, we studied the ovipositional response of Bsph susceptible and resistant Culex quinquefasciatus to aqueous suspensions of Bti and Bsph water dispersible granules (WDG). We quantified the level of mortality of adult mosquitoes caused by exposure to Bti and Bsph suspensions during oviposition. Significantly lower numbers of egg rafts were laid and collected from the treatments than the control regimen. There was an inverse relationship between Bacillus product concentrations and oviposition. As the concentration of Bti or Bsph increased from 0.0 to 2.0 mg/L, treated waters received progressively fewer egg rafts. In addition to the negative effects of Bacillus on oviposition, both male and female adult mosquitoes suffered high mortality on landing and imbibing on Bti and Bsph suspensions, the extent of mortality directly proportional to concentration. These two microbial agents used solely as mosquito larvicides thus have the additional benefits of reducing mosquito oviposition and killing adult mosquitoes, especially gravid females that come in contact with the treated water either for oviposition or drinking. Reducing the number of gravid females may also result in reduced transmission rates of pathogens. The combined effects of reduced oviposition and adult mortality could result in higher control potential of these microbial agents.

Animals↗

PubMed1986

[DNA-methylases from different strains of Bacillus subtilis and Bacillus natto].

DNA-methyltransferase activity has been detected in some of Bacillus subtilis and Bacillus natto strains. Two strains of Bacillus subtilis exhibited DNA-cytosine methyltransferase activity, and the strains of Bacillus natto exhibited DNA-adenine methyltransferase activity. A possible effect of DNA-methyltransferase specificity on transformation efficiency is discussed.

Bacillus↗

PubMed1963

LETHALITY FOR MICE OF VEGETATIVE AND SPORE FORMS OF BACILLUS CEREUS AND BACILLUS CEREUS-LIKE INSECT PATHOGENS INJECTED INTRAPERITONEALLY AND SUBCUTANEOUSLY.

Lamanna, Carl (Army Research Office, Washington, D.C.) and Linda Jones. Lethality for mice of vegetative and spore forms of Bacillus cereus and Bacillus cereus-like insect pathogens injected intraperitoneally and subcutaneously. J. Bacteriol. 85:532-535. 1963.-Lethality for mice of Bacillus thuringiensis var. thuringiensis (two strains), var. sotto (one strain), var. alesti (one strain), and B. entomocidus var. entomocidus (one strain) maintained on artificial laboratory media is dependent upon the route of injection, the number of the bacilli, and their biological state, i.e., vegetative or spore form. Deaths did not occur upon subcutaneous injection of tenfold greater numbers of organisms than the ld(50) dose for the intraperitoneal route. From 10 to 100 times the number of spores were required to kill than vegetative bacilli given intraperitoneally. These observations duplicated those made with strains of B. cereus. For both types of spore forms, from 5 x 10(5) to 5 x 10(7) vegetative organisms were required for one intraperitoneal ld(50). With two strains of highly virulent B. anthracis, ld(50) values were the same for the vegetative and spore states, and for subcutaneous and intraperitoneal injections. Thus, the insect pathogenic sporeforming bacteria tested are more like B. cereus than like B. anthracis in their capacity to infect mice. The similarity to B. cereus extends to the rapidity of death after fatal injection of the insect pathogenic strains. The medical and taxonomic implications of the observations are discussed.

Animals↗

PubMed1963

SIMILARITY IN BASE COMPOSITIONS OF DEOXYRIBONUCLEATES FROM SEVERAL STRAINS OF BACILLUS CEREUS AND BACILLUS ANTHRACIS.

McDonald, William C. (U.S. Army Biological Laboratories, Frederick, Md.), Ira C. Felkner, Abraham Turetsky, and Thomas S. Matney. Similarity in base compositions of deoxyribonucleates from several strains of Bacillus cereus and Bacillus anthracis. J. Bacteriol. 85:1071-1073. 1963.-Studies were made on the base compositions of the deoxyribonucleates of a number of Bacillus species. Species homogeneity was quite evident from these studies since, with only one exception, several strains of the same species showed identical base compositions. It was found that the melting temperatures and corresponding percentages of guanine plus cytosine were quite similar in several strains of B. cereus and B. anthracis. These aspects are discussed as related to their possible genetic homology.

Bacillus↗

PubMed1963

TRANSDUCTION OF BACILLUS LICHENIFORMIS AND BACILLUS SUBTILIS BY EACH OF TWO PHAGES.

Taylor, Martha J. (U.S. Army Biological Laboratories, Fort Detrick, Frederick, Md.) and Curtis B. Thorne. Transduction of Bacillus licheniformis and Bacillus subtilis by each of two phages. J. Bacteriol. 86:452-461. 1963.-A second transducing bacteriophage, designated SP-15, was isolated from the same soil-sample culture filtrate that supplied the Bacillus subtilis transducing phage, SP-10, reported earlier from this laboratory. SP-10 and SP-15 differ serologically and in several other respects, but share the ability to propagate on B. subtilis W-23-S(r) (streptomycin-resistant) and B. licheniformis ATCC 9945a, and to mediate general transduction in either species when propagated homologously. Attempts to transduce between the species have failed. SP-10 forms plaques readily on both W-23-S(r) and 9945a; SP-15 forms minute plaques on W-23-S(r) and has shown no evidence of any lytic activity on 9945a. Maximal recoveries of prototrophic colonies from mixtures of SP-10 with auxotrophs of either W-23-S(r) or 9945a were obtained only when excess phage was neutralized by post-transduction treatment with specific phage antiserum. Such treatment was not necessary for maximal recovery of transductants effected by SP-15. Unlike SP-10, SP-15 propagated on W-23-S(r) did not transduce B. subtilis 168 (indole(-)). SP-15 transduced B. licheniformis more efficiently than did SP-10. Neither phage was able to transduce B. licheniformis as efficiently as it transduced B. subtilis. The differing influences of multiplicity of infection were compared for the two phages in both species.

Bacillus↗

PubMed1998

High-level expression of an endoxylanase gene from Bacillus sp. in Bacillus subtilis DB104 for the production of xylobiose from xylan.

To produce xylobiose from xylan, high-level expression of an endoxylanase gene from Bacillus sp. was carried out in Bacillus sabtilis DB104. A 1.62-kb SmaI DNA fragment, coding for an endoxylanase of Bacillus sp., was ligated into the Escherichia coli/B. subtilis shuttle vector pJH27 delta 88, producing pJHKJ4, which was subsequently transformed into B. subtilis DB104. A maximum endoxylanase activity of 105 U/ml was obtained from the supernatant of B. subtilis DB104 harboring pJHKJ4. The endoxylanase was purified to homogeneity by ion-exchange chromatography and the production profile of xylooligosaccharides from xylan by the endoxylanase was examined by HPLC with a carbohydrate analysis column. Xylobiose was the major product from xylan at 40 degrees C and its proportion in the xylan hydrolyzates increased with the reaction time; at 12 h, over 60% of the reaction products was xylobiose. These results suggest that xylobiose, which has a stimulatory effect on the selective growth of the intestinal bacterium Bifidobacterium, can be mass-produced effectively by the endoxylanase of Bacillus sp. cloned in B. subtilis.

Bacillus subtilis↗

PubMed2000

Reidentification of facultatively alkaliphilic Bacillus firmus OF4 as Bacillus pseudofirmus OF4.

With a view toward verifying the original classification of alkaliphilic Bacillus firmus OF4, physiological and biochemical characteristics were more extensively catalogued than in original studies, and this catalog was supplemented with 16S rDNA sequence homology and more extensive DNA-DNA hybridization analyses. Phylogenetic analysis of this alkaliphile based on the comparison of multiple 16S rDNA sequences from Bacillus species indicated that this strain is most closely related to Bacillus pseudofirmus. Consistently, in the DNA-DNA hybridization analysis of the alkaliphile and Bacillus reference strains, the highest level of DNA-DNA relatedness (96%) was found between the alkaliphile and the B. pseudofirmus type strain (DSM 8715(T)). The findings support the conclusion that this alkaliphile strain is more closely related to B. pseudofirmus than to B. firmus, and we propose the future use of the designation B. pseudofirmus OF4.

Bacillus↗

PubMed1998

A DNA probe for the detection and identification of Bacillus sporothermodurans using the 16S-23S rDNA spacer region and phylogenetic analysis of some field isolates of Bacillus which form highly heat resistant spores.

The spacer regions between the 16S and 23S rRNA genes (spacer regions 1) of Bacillus sporothermodurans were PCR-amplified, cloned and sequenced. Six unique spacer sequences in four size classes were recovered from two strains, rrnA (about 190 bp), rrnB (about 303 bp), rrnC (355 bp) and rrnD (554 bp). rrnD contained two tRNA genes which were deciphered as tRNA(ala) and tRNA(ile) separated from each other by 13 nucleotides. The primary structures of the tRNA molecules clearly resembled those found in Bacillus subtilis; the tRNA(ala) genes were identical and the tRNA(ile) genes were 95% similar. The mixed rrnA and rrnB spacers when PCR-amplified from chromosomal DNA were effective as a hybridization probe for identification of B. sporothermodurans strains. However, high background signals with DNA from some other bacilli were encountered. A more discriminating probe was prepared from the cloned rrnB spacer region. Of eight aerobic, endospore-forming bacteria isolated from silage following heat enrichment, one was identified as B. sporothermodurans using the probe and its identity was confirmed from partial 16S rDNA analysis (phylotyping). This indicated that contamination in milk and dairies by B. sporothermodurans could originate from cattle feeds such as silage. Of the other seven silage strains, only two were identified conclusively by phylotyping and three represented probable new species. The latter three strains were subjected to phylogenetic analysis using almost complete 16S rDNA sequences. Branch lengths, bootstrap percentage values, and 16S rDNA similarity to other Bacillus species suggested that these isolates are likely to constitute new species within the genus Bacillus.

Animals↗

PubMed1998

Typing of Bacillus sporothermodurans and other Bacillus species isolated from milk by repetitive element sequence based PCR.

When analysing raw milk for the presence of Bacillus sporothermodurans, 11 Bacillus strains were isolated which could be differentiated from known Bacillus, Brevibacillus and Paenibacillus spp. by different primer specificity in PCR experiments, indicating that they probably belong to Bacillus species as yet undescribed. Using repetitive element sequence based PCR (REP-PCR), these 11 strains could be clearly distinguished from B. sporothermodurans as well as from each other. Eighty-five B. sporothermodurans strains were characterized by a typical REP-pattern. Using REP-PCR combined with separation on non-denaturing polyacrylamide gel electrophoresis and silver staining, individual B. sporothermodurans strains could be discriminated, which was not possible by methods previously published.

Animals↗

PubMed2004

Replacement of amino acid sequence features of a- and c-subunits of ATP synthases of Alkaliphilic Bacillus with the Bacillus consensus sequence results in defective oxidative phosphorylation and non-fermentative growth at pH 10.5.

Mitchell's (Mitchell, P. (1961) Nature 191, 144-148) chemiosmotic model of energy coupling posits a bulk electrochemical proton gradient (Deltap) as the sole driving force for proton-coupled ATP synthesis via oxidative phosphorylation (OXPHOS) and for other bioenergetic work. Two properties of proton-coupled OXPHOS by alkaliphilic Bacillus species pose a challenge to this tenet: robust ATP synthesis at pH 10.5 that does not correlate with the magnitude of the Deltap and the failure of artificially imposed potentials to substitute for respiration-generated potentials in energizing ATP synthesis at high pH (Krulwich, T. (1995) Mol. Microbiol. 15, 403-410). Here we show that these properties, in alkaliphilic Bacillus pseudofirmus OF4, depend upon alkaliphile-specific features in the proton pathway through the a- and c-subunits of ATP synthase. Site-directed changes were made in six such features to the corresponding sequence in Bacillus megaterium, which reflects the consensus sequence for non-alkaliphilic Bacillus. Five of the six single mutants assembled an active ATPase/ATP synthase, and four of these mutants exhibited a specific defect in non-fermentative growth at high pH. Most of these mutants lost the ability to generate the high phosphorylation potentials at low bulk Deltap that are characteristic of alkaliphiles. The aLys(180) and aGly(212) residues that are predicted to be in the proton uptake pathway of the a-subunit were specifically implicated in pH-dependent restriction of proton flux through the ATP synthase to and from the bulk phase. The evidence included greatly enhanced ATP synthesis in response to an artificially imposed potential at high pH. The findings demonstrate that the ATP synthase of extreme alkaliphiles has special features that are required for non-fermentative growth and OXPHOS at high pH.

Adenosine Triphosphatases↗

PubMed1999

Bacillus silvestris sp. nov., a new member of the genus Bacillus that contains lysine in its cell wall.

A Gram-positive, aerobic, rod-shaped, peritrichously flagellated, round-endospore-forming bacterium was isolated from a forest soil near Braunschweig, Lower Saxony, Germany, and designated strain HR3-23T (T = type strain). Morphologically, strain HR3-23T shows the characteristics of a member of the genus Bacillus. The spore position is terminal in a swollen sporangium. Comparative analysis of the 16S rDNA sequence shows strain HR3-23T to be most closely related to Caryophanon tenue (95.8% 16S rRNA similarity) and to Bacillus sphaericus (95.4% 16S rRNA similarity). Phylogenetically, the isolate clusters among species of Bacillus RNA group 2. The DNA G + C content of isolate HR3-23T is 39.3 mol%, the peptidoglycan type is A4 alpha (L-Lys-D-Glu), the major respiratory lipoquinone is menaquinone MK-7 and the predominant fatty acid is of the iso-C15:0 type. Based on the morphological, chemotaxonomic, physiological and phylogenetic properties, a new species, Bacillus silvestris, is proposed; strain HR3-23T is the type strain (= DSM 12223T).

Bacillus↗

PubMed2002

Bacillus pycnus sp. nov. and Bacillus neidei sp. nov., round-spored bacteria from soil.

Bacillus sphaericus sensu lato currently consists of seven or more groups of unrelated taxa, one of which is B. sphaericus sensu stricto and another of which is Bacillus fusiformis. Members of two groups (groups 6 and 7), in common with all other B. sphaericus-like organisms, are unable to grow anaerobically or to use common hexoses, pentoses and hexitols as sources of carbon, have G+C contents of 34-36 mol % and form round spores. Groups 6 and 7 can be differentiated from other B. sphaericus-like organisms by low DNA relatedness and by variations in whole-cell fatty acid composition. Unique characteristics of group 6 include the ability to oxidize beta-hydroxybutyrate, the non-requirement for biotin and thiamin and failure to grow in 5% NaCl. Distinctive traits of group 7 include the inability to oxidize pyruvate and a requirement for biotin, thiamin and cystine for growth. These data show that groups 6 and 7 represent two novel species, for which the names Bacillus pycnus sp. nov. and Bacillus neidei sp. nov., respectively, are proposed; the corresponding type strains are NRRL NRS-1691T (= JCM 11075T) and NRRL BD-87T (= JCM 11077T).

3-Hydroxybutyric Acid↗

PubMed1995

Cation-selectivity of the L-glutamate transporters of Escherichia coli, Bacillus stearothermophilus and Bacillus caldotenax: dependence on the environment in which the proteins are expressed.

L-Glutamate transport by the H(+)-glutamate and Na(+)-glutamate symport proteins of Escherichia coli K-12 (GltPEc and GltSEc, respectively) and the Na(+)-H(+)-glutamate symport proteins of Bacillus stearothermophilus (GltTBs) and Bacillus caldotenax (GltTBc) was studied in membrane vesicles derived from cells in which the proteins were either homologously or heterologously expressed. Substrate and inhibitor specificity studies indicate that GltPEc, GltTBs and GltTBc fall into the same group of transporters, whereas GltSEc is distinctly different from the others. Also, the cation specificity of GltSEc is different; GltSEc transported L-glutamate with (at least) two Na+, whereas GltPEc, GltTBs and GltTBc catalysed an electrogenic symport of L-glutamate with > or = two H+, i.e. when the proteins were expressed in E. coli. Surprisingly studies in membrane vesicles of B. stearothermophilus and B. caldotenax indicated a Na(+)-H(+)-L-glutamate symport for both GltTBs and GltTBc. The Na+ dependency of the GltT transporters in the Bacillus strains increased with temperature. These observations suggest that the conformation of the transport proteins in the E. coli and the Bacillus membranes differs, which influences the coupling ion selectivity.

Amino Acid Transport Systems, Acidic↗

PubMed1988

[Cloning in cells of Bacillus subtilis and study of the characteristics of promoter regions of DNA of Bacillus mesentericus and Phage SPO2].

11 fragments of Bacillus mesentericus and phage SPO1 DNA containing promoters were cloned in Bacillus subtilis. Nucleotide sequences of these fragments were determined and S1 mapping of transcriptional start points was performed. It was found that some fragments contained tandem or overlapping promoters. Nucleotide sequences of Bacillus mesentericus promoters were compared with known promoters from bacilli. Promoter activities were determined in different phases of cell growth. Nucleotide sequences of several promoters coincided with known sigma 43, sigma 29, sigma 28-promoters. Nucleotide sequences of other promoters differ greatly from those used by the known forms of Bacillus subtilis RNA polymerase. The existence of at least four other forms of bacilli RNA polymerase specific for these promoters is suggested.

Bacillus subtilis↗

PubMed1976

The primary structure of Bacillus subtilis and Bacillus stearothermophilus 5 S ribonucleic acids.

The nucleotide of Bacillus stearothermophilus 5 S RNA is pC-C-U-A-G-U-G-A-C-A-A-U-A-G-C-G-(G-A-G-A-G-G-)-A-A-A-C-A-C-C-C-G-U-U-C-C-C-A-U-C-C-C-G-A-A-C-A-C-G-G-A-A-G-U-U-A-A-G-C-U-C-U-C-C-A-G-C-G-C-C-G-A-U-G-G-U-A-G-U-U-G-G-G-G-C-C-A-G-C-G-C-C-C-C-U-G-C-A-A-G-A-G-U-A-G-G-U-C-G-U-U-G-C-U-A-G-G-COH; the nucelotide sequence of Bacillus subtilis 5 S RNA is pU-U-U-G-G-U-G-G-C-G-A-U-A-G-C-G-A-A-G-A-G-G-U-C-A-C-A-C-C-C-G-U-U-C-C-C-A-U-A-C-C-G-A-A-C-A-C-G-G-A-A-G-U-U-A-A-G-C-U-C-U-U-C-A-G-C-G-C-C-G-A-U-G-G-U-A-G-U-C-G-G-G-G-G-U-U-U-C-C-C-C-C-U-G-U-G-A-G-A-G-U-A-G-G-A-C-G-C-C-G-C-C-A-A-G-COH. Comparison of the sequence of B. stearothermophilus 5 S RNA to the sequence of B. subtilis 5 S RNA and to that of Bacillus megaterium 5 S RNA (Pribula, C. D., Fox, G. E., Woese, C. R., Sogin, M., and Pace, N. (1974) FEBS Lett. 44, 322-323) indicates that the 5 S RNA isolated from the thermophile contains unique nucleotide sequences not found in 5 S RNAs isolated from the two mesophilic species of genus Bacillus.

Bacillus subtilis↗