Bacillus

Int J Dermatol, 1991 Nov, 30(11), 780 - 4
Characterization of circulating lymphocytes by monoclonal antibodies in childhood and adult leprosy; Sehgal VN et al.; Peripheral blood lymphocyte assays using monoclonal antibodies were done in 66 patients with leprosy, consisting of 25 children and 41 adults . The results were statistically analyzed for correlations, if any, among the different age groups and matched controls . The results, however, failed to show any significant correlation, nor was it possible to draw any conclusion as to why the disease spectrum in children tends to be incomplete (ie, there is a low incidence of the highly bacilliferous form of disease expression).

Pediatr Infect Dis J, 1991 Nov, 10(11), 832 - 6
Miliary tuberculosis in children: a review of 94 cases; Hussey G et al.; This is a retrospective review of the clinical, radiologic and laboratory features of 94 cases of childhood miliary tuberculosis seen during a 5-year period, 1985 to 1989 . A history of Bacillus Calmette-Guerin vaccination was documented in 88% of children . The median age at presentation was 10.5 months, 52% of cases occurring in those younger than 1 year . The presenting symptoms were nonspecific: cough (72%); fever (61%); loss of appetite and weight (40%); and diarrhea and vomiting (33%) . The main presenting signs were hepatomegaly (82%), splenomegaly (54%), lymphadenopathy (46%) and pyrexia (39%) . Most of the patients were malnourished and anergic . Meningitis occurred in 19% of patients and this was the only significant risk factor identified for mortality, the overall case fatality rate being 14% . The diagnosis in the vast majority was made on the clinical presentation supported by a classic miliary pattern on chest roentgenogram (91% of cases) . Mycobacterium tuberculosis was cultured in 33% of cases . In addition a review of hospital admissions from 1981 to 1989 revealed that annually miliary tuberculosis in children and adults accounted for 8.3 and 1.3%, respectively, of all tuberculosis admissions . This study confirms that miliary tuberculosis is a relatively common complication of tuberculosis in young children.

J Neurol Neurosurg Psychiatry, 1991 Nov, 54(11), 989 - 92
Whipple's disease confined to the CNS presenting with multiple intracerebral mass lesions; Wroe SJ et al.; A patient with isolated cerebral Whipple's disease presented with signs of raised intracranial pressure and multiple ring enhancing intracerebral mass lesions evident on CT and MRI imaging . Characteristic intracellular bacilliform inclusions were identified in a brain biopsy . Clinical improvement followed treatment with parenteral antibiotics for two weeks and long term sulphamethoxazole-trimethoprim . As CNS relapse of Whipple's disease may occur after several years, long term treatment should include antibiotics that are able to cross the blood-brain barrier.

Gan To Kagaku Ryoho, 1991 Nov, 18(14), 2369 - 74
{Superficial bladder cancer: prophylaxis of recurrence and progression}; Miyanaga N et al.; Superficial bladder cancer has a good prognosis compared with invasive bladder cancer . However, recurrence of the tumor is frequent and tumor stage and/or grade progress at the time of recurrence in many cases . Intravesical chemotherapy has been employed as a prophylactic method after trans-urethral resection (TUR) . Although intravesical chemotherapy has been proved to be effective in delaying the first recurrence of tumor after TUR, it cannot improve the ultimate prognosis of superficial bladder cancer . Many primary, solitary, non-invasive (Ta) and grade 1 tumors do not recur or progress in stage and grade . In these cases, prophylactic intravesical chemotherapy is not essential . Bacille Calmette-Guerin (BCG) should be considered superior overall, to any chemotherapeutic agents . Comparative studies will give information about the best clinical schedule for the treatment of superficial bladder cancer.

Indian J Exp Biol, 1991 Nov, 29(11), 1031 - 4
Fed-batch fermentation studies with Bacillus thuringiensis H-14 synthesising endotoxin; Kuppusamy M et al.; Cell yield and toxicity of B . thuringiensis H-14 was improved markedly by adopting a simple fed-batch fermentation technique based on controlling glucose concentration . Maintenance of steady glucose concentration (0.3-0.5%) in the culture medium was achieved by the continuous addition of concentrated glucose solution . Addition of glucose at 3 g/hr/l of culture starting from 3rd hr till 16th hr of fermentation was found to yield cell densities of 80 g/l (wet weight) which represented a nearly 3-fold increase over the batch mode . A fivefold increase in toxicity was obtained by fed-batch fermentation . Cultivation of B . thuringiensis H-14 to high cell densities had no negative effect on sporulation and toxin synthesis . The rate of pH drop and dissolved oxygen level were within manageable limits.

Mol Gen Mikrobiol Virusol, 1991 Nov, (11), 20 - 2
{Search for producers of restriction endonucleases with a given specificity}; Repin VE et al.; Six site-specific restriction endonucleases were isolated from Bacillus thuringiensis strains chosen of 58 strains ought purposefully for production of restriction enzymes . All six strains produce isoschisomers of Sau3A.

Mol Microbiol, 1991 Nov, 5(11), 2799 - 806
The C-terminal domain of the toxic fragment of a Bacillus thuringiensis crystal protein determines receptor binding; Honee G et al.; The insecticidal crystal proteins of Bacillus thuringiensis show a high degree of specificity . In vitro binding studies with several crystal proteins demonstrated a correlation between toxicity and binding to receptors of larval midgut epithelial cells . In order to study the domain-function relationships of the toxic fragment, hybrid crystal proteins based on CryIA(b) and CryIC were constructed . Two out of 11 hybrid proteins constructed exhibited insecticidal activity . Both dispalyed an insecticidal spectrum similar to that of the parental crystal protein from which the C-terminal part of the toxic fragment originated . In addition, in vitro binding studies directly demonstrated the involvement of the C-terminal part of the toxic fragment in receptor binding . These results demonstrate that the C-terminal part of the toxic fragment determines specific receptor binding, which in turn determines, to a large extent, the insect specificity.

Biotechnol Prog, 1991 Nov-Dec, 7(6), 516 - 25
Characterization of kappa-carrageenan gels used for immobilization of Bacillus firmus; Moon SH et al.; In this study, aimed at a biochemical and physical characterization of kappa-carrageenan gels used for entrapment of Bacillus firmus NRS 783 (a superior producer of an alkaline protease), effects of carrageenan concentration, gelation temperature, initial cell loading, and strength of the curing agent (KCl) on the properties of cell-free and cell-laden gels were examined . The physical properties of the differently prepared gels that were examined included density, free volume fraction, mechanical strength, and change in gel volume during gel curing . The biochemical characteristics studied included viability of gel-entrapped cells, cell leakage from cell-laden gels, and cell penetration into cell-free gels . For the range of carrageenan contents investigated {between 2% and 5% (w/v)}, the mechanical strength of the gels with/without KCl curing was observed to increase with an increase in carrageenan content of gels . The mechanical strength of each gel increased substantially upon extensive curing . Free volume fractions in excess of 0.8 were observed for all gels . Of cells that were viable prior to immobilization, 90-92% remained viable after formation and extensive curing of gels for cell-gel mixtures prepared at 45 degrees C . Attempts at prolonged storage of cell-laden gel beads at 0 degrees C as stock cultures of immobilized B . firmus were unsuccessful due to a significant decline in cell viability during such storage . On the basis of the cell leakage studies, the average pore sizes of 2%, 3%, 4%, and 5% gels were deduced to increase in the following order of carrageenan content (w/v): 4%, 3%, 2%, and 5% . Commensurate with the decrease in the average pore size (or the increased tightness of the gels) with increasing carrageenan content, both the extent of cell leakage and the extent of net cell penetration decreased with increasing carrageenan content for the first three gels . Owing to non-uniform distribution of free space and much larger pores, the extent of net cell penetration in 5% carrageenan gels was considerably low, while the extent of cell leakage in 5% carrageenan gels was an order of magnitude greater than the extents of cell leakage in the other three gels.

Nature, 1991 Oct 31, 353(6347), 815 - 21
Crystal structure of insecticidal delta-endotoxin from Bacillus thuringiensis at 2.5 A resolution; Li JD et al.; The structure of the delta-endotoxin from Bacillus thuringiensis subsp . tenebrionis that is specifically toxic to Coleoptera insects (beetle toxin) has been determined at 2.5 A resolution . It comprises three domains which are, from the N- to C-termini, a seven-helix bundle, a three-sheet domain, and a beta sandwich . The core of the molecule encompassing all the domain interfaces is built from conserved sequence segments of the active delta-endotoxins . Therefore the structure represents the general fold of this family of insecticidal proteins . The bundle of long, hydrophobic and amphipathic helices is equipped for pore formation in the insect membrane, and regions of the three-sheet domain are probably responsible for receptor binding.

Proc Natl Acad Sci U S A, 1991 Oct 15, 88(20), 8930 - 3
Binding of Bacillus thuringiensis proteins to a laboratory-selected line of Heliothis virescens; MacIntosh SC et al.; A laboratory-selected colony of Heliothis virescens displaying a 20- to 70-fold level of resistance to Bacillus thuringiensis proteins was evaluated to identify mechanism(s) of resistance . Brush-border membrane vesicles were isolated from larval midgut epithelium from the susceptible and resistant strains of H . virescens . Two B . thuringiensis proteins, CryIA(b) and CryIA(c), were iodinated and shown to specifically bind to brush-border membrane vesicles of both insect strains . Multiple changes in the receptor-binding parameters were seen in the resistant strain as compared with the susceptible strain . A 2- to 4-fold reduction in binding affinity was accompanied by a 4- to 6-fold increase in binding-site concentration for both proteins . Although these two B . thuringiensis proteins competed for the same high-affinity binding site, competition experiments revealed different receptor specificity toward these proteins in the resistant H . virescens line . The H . virescens strains were not sensitive to a coleopteran-active protein, CryIIIA, nor did these proteins compete with the CryIA proteins for binding . Complexity of the mechanism of resistance is consistent with the complex mode of action of B . thuringiensis proteins.

J Immunol, 1991 Oct 15, 147(8), 2802 - 8
Cellular models of macrophage tumoricidal effector mechanisms in vitro . Characterization of cytolytic responses to tumor necrosis factor and nitric oxide pathways in vitro; Klostergaard J et al.; The recently described L-arginine-dependent nitric oxide (NO) pathway has been proposed to interact synergistically with the TNF pathway in murine macrophage-mediated tumor cytotoxicity in vitro . We have employed an experimental construct in which these two pathways were independently expressed by two different effector cell populations . The TNF-dependent pathway was committed by murine 3T3 cells transfected with the cDNA encoding human pro-TNF . The NO pathway was executed by the murine EMT-6 mammary adenocarcinoma cell line treated with murine rIFN-gamma and LPS . Controls for the TNF pathway committed by the transfectant included lysis of the TNF-sensitive murine L929 cell in coculture, secretion of TNF, and absence of nitrite synthesis . For the NO pathway controls included lysis of the murine P815 mastocytoma cocultured with activated EMT-6 cells that had been pretreated with murine rIFN-gamma and LPS, production of nitrite by this activated effector cell, and an absence of TNF secretion . The target cell panel included L929, EMT-6, P815, and murine B16 melanoma and TU-5 sarcoma cell lines . All targets on this panel were susceptible to lysis by LPS-triggered murine bacillus Calmette-Guerin-activated macrophages . The 3T3 transfectant caused significant lysis of cocultured L929 and TU-5 targets . The EMT-6 effector cell only caused significant lysis of the P815 target . When both effector cells were cocultured with these target cells, lysis of the P815 target was observed to be additive or superadditive; however, for all the other targets, cytotoxicity was comparable with or subadditive compared with that seen with the 3T3 transfectant effector cell alone . Thus, these two pathways do not appear to account for the broad, potent tumoricidal activity observed for activated macrophages in vitro.

FEMS Microbiol Lett, 1991 Oct 15, 67(3), 273 - 6
Cytotoxicity of a cloned Bacillus thuringiensis subsp . israelensis CryIVB toxin to an Aedes aegypti cell line; Angsuthanasombat C et al.; A cloned CryIVB toxin was purified from a cured strain of Bacillus thuringiensis (BT) containing the cryIVB gene on the recombinant plasmid Cam135 . Solubilized protoxin was treated with Aedes gut extract or trypsin for varying times and tested for toxicity in vitro on three dipteran and one lepidopteran cell line . Treatment with the Aedes extract but not trypsin, produced an active toxin which lysed only Aedes aegypti cells out of those tested . This activation was time-dependent reaching a maximum after 6 h . Both the Aedes extract-treated and trypsin-treated toxin killed A . aegypti larvae, but this toxicity declined rapidly with increasing time of exposure to the proteolytic preparations.

J Immunol, 1991 Oct 15, 147(8), 2706 - 12
Murine T cell-stimulatory peptides from the 19-kDa antigen of Mycobacterium tuberculosis . Epitope-restricted homology with the 28-kDa protein of Mycobacterium leprae; Harris DP et al.; Fifteen overlapping synthetic peptides, spanning the entire amino acid sequence of the Mycobacterium tuberculosis 19-kDa protein, were used to identify epitopes recognized by murine T cells . Five of the 15 peptides tested were able to elicit in vitro lymph node T cell proliferative responses in C57BL/10 mice primed by footpad inoculation with homologous peptide . Analysis in congenic strains of mice revealed H-2 restriction in the response to four peptides . However, one peptide, 19.7 (residues 61 to 80), induced T cell responses in all four haplotypes tested . This peptide was also unique in being able to stimulate lymph node cells from C57BL/10 mice immunized with recombinant 19-kDa protein, killed M . tuberculosis, or live bacillus Calmette Guerin infection . T cell lines specific for peptide 19.7 were of the CD4 phenotype . Significantly, sequence analysis revealed that residues 61 to 80 of the 19-kDa protein exhibited considerable homology with a single 20-amino acid sequence (residues 120 to 140), but not with any other region of the 28-kDa protein expressed in Mycobacterium leprae . This finding is the first evidence of epitope-restricted homology between otherwise structurally unrelated microbial Ag.

Eur J Biochem, 1991 Oct 15, 201(2), 467 - 73
The electrochemical proton potential of Bacillus alcalophilus; Hoffmann A et al.; Bacillus alcalophilus strain ATCC 27647 showed usual growth characteristics, when inoculated at pH 10.4 . The cells entered the logarithmic phase at pH 10.3, and as growth continued, the pH dropped further to a value of 8.8 in the stationary phase . B . alcalophilus strain DSM 485 showed comparable growth only in the initial phase after the addition to fresh medium . The small initial growth period was succeeded by a long lag phase, where the pH continuously dropped . The cells resumed growth after the pH was about 10.0 and continued to grow accompanied by a further decrease of external pH . The bioenergetic parameters measured in the initial growth phase of the two strains at high pH (10.1-10.3) were nearly the same, i.e . delta pH = +97 to +110 mV, delta psi = -206 to -213 mV and delta microH+ = -109 to -103 mV . The inverted proton gradient of about 1.7-1.9 at high pH decreased, as the external pH dropped during growth . This led to an increase of the proton motive force (delta microH+), although the membrane potential (delta psi) also declined . The ATP/ADP ratio of strain DSM 485 was high (4.5-5.5) at fast growth during the initial and second growth period . The ratio declined to about 1.5 at the end of the lag phase . At the initial growth phase and at the end of the lag phase, the delta microH+ was, however, the same (approximately -106 mV) and considerably lower than in the middle of the second growth period (approximately -140 mV) . Fast growth, therefore, correlates with a high ATP/ADP ratio but not necessarily with a high delta microH+ . Addition of gramicidin or carbonylcyanide m-chlorophenylhydrazone stopped growth of B . alcalophilus strain DSM 485 at pH 10.3 or 9.5 and gramicidin immediately decreased the internal ATP/ADP ratio from 4.5 to 1.2 at pH 10.3.

J Biol Chem, 1991 Oct 5, 266(28), 18567 - 72
Characterization of semi-uncoupled hybrid Escherichia coli-Bacillus megaterium F1F0 proton-translocating ATPases; Scarpetta MA et al.; Cloned atp genes for the proton-translocating ATPase of the obligate aerobe Bacillus megaterium have been demonstrated to be capable of complementing Escherichia coli ATPase (unc) mutants (Hawthorne, C . A., and Brusilow, W . S . A . (1986) J . Biol . Chem . 261, 5245-5248) . To determine the minimum subunit requirements for cross-species complementation, we constructed all combinations of B . megaterium atpA, G, D, and C genes (coding for the alpha, gamma, beta, and epsilon subunits, respectively) and tested their abilities to complement two uncA (alpha subunit) and two uncD (beta subunit) mutants of E . coli . The results indicated that complementation of either uncD mutant required atpD (beta) only . Complementation of one of the uncA (alpha) mutants required atpA, G, and D (alpha, gamma, and beta) and possibly atpE (epsilon) as well . The other uncA mutant was not complemented by any combination of B . megaterium ATPase genes . Complementation of a beta mutant by atpD (beta) or atpD and C (beta epsilon) produced cells which could grow aerobically on a nonfermentable carbon source (succinate) but not anaerobically on rich medium containing glucose . These E . coli therefore had become obligate aerobes . The ability to grow anaerobically could be restored to the mutant complemented by atpD alone by growth at pH 7.5 or pH 8 in the presence of 0.1 M potassium.

Biochem J, 1991 Oct 1, 279 ( Pt 1), 213 - 21
Mechanism of acyl transfer by the class A serine beta-lactamase of Streptomyces albus G; Lamotte-Brasseur J et al.; Optimization by energy minimization of stable complexes occurring along the pathway of hydrolysis of benzylpenicillin and cephalosporin C by the Streptomyces albus G beta-lactamase has highlighted a proton shuttle that may explain the catalytic mechanism of the beta-lactamases of class A . Five residues, S70, S130, N132, T235 and A237, are involved in ligand binding . The gamma-OH group of T235 and, in the case of benzylpenicillin, the gamma-OH group of S130 interact with the carboxylate group, on one side of the ligand molecule . The side-chain NH2 group of N132 and the carbonyl backbone of A237 interact with the exocyclic CONH amide bond, on the other side of the ligand . The backbone NH groups of S70 and A237 polarize the carbonyl group of the scissile beta-lactam amide bond . Four residues, S70, K73, S130 and E166, and two water molecules, W1 and W2, perform hydrolysis of the bound beta-lactam compound . E166, via W1, abstracts the proton from the gamma-OH group of S70 . While losing its proton, the O-gamma atom of S70 attacks the carbonyl carbon atom of the beta-lactam ring and, concomitantly, the proton is delivered back to the adjacent nitrogen atom via W2, K73 and S130, thus achieving formation of the acyl-enzyme . Subsequently, E166 abstracts a proton from W1 . While losing its proton, W1 attacks the carbonyl carbon atom of the S70 ester-linked acyl-enzyme and, concomitantly, re-entry of a water molecule W'1 replacing W1 allows E166 to deliver the proton back to the same carbonyl carbon atom, thus achieving hydrolysis of the beta-lactam compound and enzyme recovery . The model well explains the differences found in the kcat . values for hydrolysis of benzylpenicillin and cephalosporin C by the Streptomyces albus G beta-lactamase . It also explains the effects caused by site-directed mutagenesis of the Bacillus cereus beta-lactamase I {Gibson, Christensen & Waley (1990) Biochem J . 272, 613-619}.

Biochem J, 1991 Oct 1, 279 ( Pt 1), 111 - 4
Interaction of beta-lactamases I and II from Bacillus cereus with semisynthetic cephamycins . Kinetic studies; Martin Villacorta J et al.; The influence of C-6 alpha- or C-7 alpha-methoxylation of the beta-lactam ring in the catalytic action of class A and B beta-lactamases has been investigated . For this purpose the kinetic behaviour of beta-lactamases I (class A) and II (class B) from Bacillus cereus was analysed by using several cephamycins, moxalactam, temocillin and related antibiotics . These compounds behaved as poor substrates for beta-lactamase II, with high Km values and very low catalytic efficiencies . In the case of beta-lactamase I, the substitution of a methoxy group for a H atom at C-7 alpha or C-6 alpha decreased the affinity of the substrates for the enzyme . Furthermore, the acylation of cephamycins was completely blocked, whereas that of penicillins was slowed down by a factor of 10(4)-10(5), acylation being the rate-determining step of the process.

J Bacteriol, 1991 Oct, 173(19), 6147 - 52
Analysis of the active center of Bacillus stearothermophilus neopullulanase; Kuriki T et al.; The active center of the neopullulanase from Bacillus stearothermophilus was analyzed by means of site-directed mutagenesis . The amino acid residues located in the active center of the neopullulanase were tentatively identified according to a molecular model of Taka-amylase A and homology analysis of the amino acid sequences of neopullulanse, Taka-amylase A, and other amylolytic enzymes . When amino acid residues Glu and Asp, corresponding to the putative catalytic sites, were replaced by the oppositely charged (His) or noncharged (Gln or Asn) amino acid residue, neopullulanase activities toward alpha-(1----4)- and alpha-(1----6)-glucosidic linkages disappeared . When the amino acids corresponding to the putative substrate-binding sites were replaced, the specificities of the mutated neopullulanases toward alpha-(1----4)- and alpha-(1----6)-glucosidic linkages were obviously different from that of the wild-type enzyme . This finding proves that one active center of neopullulanase participated in the dual activity toward alpha-(1----4)- and alpha-(1----6)-glucosidic linkages . Pullulan is a linear glucan of maltotriosyl units linked through alpha-(1----6)-glucosidic linkages . The production ratio of panose from pullulan was significantly increased by using the mutated neopullulanase which exhibited higher specificity toward the alpha-(1----4)-glucosidic linkage . In contrast, the production ratio of panose was obviously decreased by using the mutated neopullulanse which exhibited higher specificity toward the alpha-(1----6)-glucosidic linkage.

Eur J Biochem, 1991 Oct 1, 201(1), 203 - 9
Sequence-specific 1H-NMR assignments and secondary structure of the lipoyl domain of the Bacillus stearothermophilus pyruvate dehydrogenase multienzyme complex; Dardel F et al.; The lipoyl domain (residues 1-85) of the lipoate-acetyltransferase polypeptide chain of the pyruvate dehydrogenase multienzyme complex of Bacillus stearothermophilus has been subjected to detailed structural analysis by means of two-dimensional (2D) 1H-NMR spectroscopy at 400 MHz . Sequence-specific proton resonance assignments were made, but at this field strength not all of the side-chain protons could be assigned, especially from complex spin systems like those of leucine, proline and lysine residues . Measurement of short-range interproton distances identified two extensive regions of beta-sheet, each containing four anti-parallel peptide strands . The lipoyl-lysine residue (Lys42) is located in a tight turn at a corner of one sheet, the N-terminal and C-terminal residues of the domain are close together in two adjacent beta-strands in the other . The lipoylated and unlipoylated forms of the domain have almost identical spectra, indicating that there is little, if any, conformational change in the protein as a result of the post-translational modification.

J Urol, 1991 Oct, 146(4), 1164 - 7
Treatment of murine transitional cell carcinoma with intralesional interleukin 2 and murine interferon gamma; Sosnowski JT et al.; The antitumor effect of intralesionally administered recombinant interleukin-2, alone or in combination with recombinant interferon gamma was studied in murine transitional cell carcinoma, MBT2 . In the initial prophylactic model treatment was started at day one at the site of tumor inoculation . Maximal and significant reduction in tumor volume occurred in groups receiving 4,000 units of recombinant interleukin 2 and 10(7) colony forming units Bacillus Calmette Guerin (p less than 0.00001 vs saline control) . In the same experiment, a reduction in tumor incidence and increase in survival occurred in groups receiving 4,000 units of recombinant interleukin 2, 1,000 units of recombinant interleukin 2 plus 2,000 units of recombinant interferon gamma, as well as 10(7) colony forming units Bacillus Calmette Guerin relative to saline control (p less than 0.005) . The dose-response effect of recombinant interleukin 2 alone was also tested in a model of an established transitional cell carcinoma . Intralesional injection treatments were initiated after tumors were palpable . Reduction in tumor volume was observed in the group receiving 8,000 units of recombinant interleukin 2 (p = 0.01 vs saline control), but no significant advantage in survival was noted.

J Urol, 1991 Oct, 146(4), 1118 - 9
A case of granulomatous hepatitis after intravesical bacillus Calmette-Guerin administration; Graziano DA et al.; A 61-year-old man received intravesical bacillus Calmette-Guerin (BCG) as treatment for superficial bladder carcinoma . High spiking relapsing fevers began 39 days after the initial treatment . A liver biopsy revealed noncaseating granuloma . Deoxyribonucleic acid hybridization of the bone marrow was positive for Mycobacterium tuberculosis complex . Pressure exerted to instill the BCG may have favored dissemination.

Indian J Exp Biol, 1991 Oct, 29(10), 953 - 7
An enzyme linked immunosorbent assay for monitoring Bacillus sphaericus toxin; Anandkumar K et al.; Larvicidal proteins of B . sphaericus H5a5b (strain VCRC B42), purified by ion-exchange chromatography were used to raise antibodies in rabbits . The antibodies were specific in reacting to alkali-solubilized fractions from whole cells of toxic strains only . Ouchterlony immunodiffusion showed homology in toxin structure between strains of different serotype . A sandwich ELISA using avidin-biotin amplification was standardized . The lowest detectable limit was 6.25 ng/ml . Near linear quantitative binding of the antigen was found in the range 25-200 ng/ml . The growth, toxin level and LC50 values during various stages of fermentation of B . sphaericus strains 1593 and B42 were compared . There was significant correlation between LC50 values and toxin levels as measured by ELISA.

Antonie Van Leeuwenhoek, 1991 Oct-Nov, 60(3-4), 355 - 71
Formation of fermentation products and extracellular protease during anaerobic growth of Bacillus licheniformis in chemostat and batch-culture; Bulthuis BA et al.; For a relaxed (rel-), protease producing (A-type) and a stringent (rel+), not-protease producing (B-type) variant of Bacillus licheniformis we determined fermentation patterns and products, growth parameters and alkaline protease-production (if any) in anaerobic, glucose-grown chemostats and batch-cultures . Glucose is dissimilated via glycolysis and oxidative pentose phosphate pathway simultaneously; the relative share of these two routes depends on growth phase (in batch) and specific growth rate (in chemostat) . Predominant products are lactate, glycerol and acetaldehyde for A-type batches and acetaldehyde, ethanol, acetate and lactate for B-type batches . Both types show a considerable acetaldehyde production . In chemostat cultures, the fermentation products resemble those in batch-culture . From the anaerobic batches and chemostats, we conclude that the A-type (with low ATP-yield) will have a YATPmax of probably 12.9 g/mol and the B-type (with high ATP-yield) a YATPmax of about 10.1 g/mol . For batch-cultures, both types have about the same, high Yglucose (12 g/mol) . So, the slow-growing A-type has a relatively high efficiency of anaerobic growth (i.e . an efficient use of ATP) and the fast-growing B-type a relatively low efficiency of anaerobic growth . In aerobic batch-cultures, we found 48, respectively 41% glucose-carbon conversion into mainly glycerol and pyruvate, respectively acetate as overflow metabolites in the A- and B-type . In both aerobic and anaerobic batch-cultures of the A-type, protease is produced predominantly in the logarithmic and early stationary phase, while a low but steady production is maintained in the stationary phase . Protease production occurs via de novo synthesis; up to 10% of the total protease in a culture is present in a cell-associated form . Although anaerobic protease production (expressed as protease per amount of biomass) is much higher than for aerobic conditions, specific rates of production are in the same range as for aerobic conditions while, most important, the substrate costs of anaerobic production are very much higher than for aerobic conditions.

Arch Esp Urol, 1991 Oct, 44(8), 1025 - 8
Immunoprophylaxis of superficial bladder cancer: a prospective and randomized comparison of oral versus intravesical Bacillus Calmette-Guerin; Netto Junior NR et al.; A total of 84 patients with superficial transitional cell carcinoma underwent transurethral resection of bladder tumor . All patients had stage pTa or pT1 transitional cell carcinoma or carcinoma in situ without other concurrent malignancies . The patients were assigned to three treatment groups: I . Control group-transurethral resection (TUR-BT) discontinued within the study . II . Oral BCG group-TUR-BT plus BCG (Moreau) . III . Intravesical BCG group-TUR-BT plus BCG . Of 9 patients in the control group, 8 (89%) experienced tumor recurrence during a mean follow-up of 20 months . Of the 33 patients in the oral BCG group, 13 patients (39.3%) had recurrence during a mean follow-up of 39 months . Of the 42 patients in the intravesical group, 8 patients (19%) had recurrence in a 30-month mean follow-up . The incidence of complications was higher in the intravesical (33.4%) than in the oral BCG group (24.2%) . These results showed that intravesical BCG is a more effective immunotherapy; however, oral BCG can be utilized in patients who do not accept intravesical BCG administration.

Xenobiotica, 1991 Oct, 21(10), 1311 - 23
Hydroxylation and glucoside conjugation in the microbial metabolism of the diterpene sclareol; Kouzi SA et al.; 1 . The microbial metabolism of sclareol (1), a labdane diterpene ditertiary alcohol, was studied . Preliminary screening identified a number of microorganisms capable of metabolizing sclareol . 2 . Preparative scale fermentation of growing cultures of Bacillus cereus UI-1477 resulted in the production of seven metabolites which have been characterized as 3 beta-hydroxysclareol (2), 2 alpha-hydroxysclareol (3), 18-hydroxysclareol (4), 2 alpha,18-dihydroxysclareol (6), 8 alpha,13 beta-dihydroxy-labd-14-en-3 beta-O-beta-D-glucoside (5), 8 alpha,13 beta-dihydroxy-labd-14-en-18-O-beta-D-glucoside (7), and 8 alpha,13 beta-dihydroxy-labd-14-en-2 alpha-O-beta-D-glucoside (10) by chemical, enzymic, and spectral data, especially 2D-n.m.r . techniques and thermospray liquid chromatography-mass spectrometry analysis.

Acta Crystallogr B, 1991 Oct 1, 47 ( Pt 5), 707 - 30
Complex between the subtilisin from a mesophilic bacterium and the leech inhibitor eglin-C; Dauter Z et al.; The alkaline proteinase from the mesophilic bacterium Bacillus mesentericus has been crystallized in a 1:1 complex with the inhibitor eglin-C from the medical leech . The crystals have cell dimensions of a = 43.0, b = 71.9, c = 48.3 A and beta = 110.0 degrees and are in the space group P2(1) . Three-dimensional data to 2.0 A have been recorded on film from a single crystal . The orientation and position of the complex in the unit cell have been established using the refined coordinates of subtilisin Carlsberg and of eglin-C as independent models . The structure of the complex has been refined by restrained least-squares minimization . The crystallographic R factor (= sigma{magnitude of Fo - magnitude of Fc{/sigma magnitude of Fo) is 15.1% including two Ca2+ ions and 312 water molecules . The structure is discussed in terms of its physicochemical properties in solution and its relation to other Bacillus subtilisins.

Anal Biochem, 1991 Oct, 198(1), 10 - 4
A fluorescent substrate for the continuous assay of phosphatidylinositol-specific phospholipase C: synthesis and application of 2-naphthyl myo-inositol-1-phosphate; Shashidhar MS et al.; A fluorescent water-soluble substrate for phosphatidylinositol-specific phospholipase C was synthesized . The diacylglycerol moiety of the natural substrate, phosphatidylinositol, was replaced by the fluorescent moiety, 2-naphthol, resulting in the synthetic substrate, racemic 2-naphthyl myo-inositol-1-phosphate . The synthetic substrate provided a continuous fluorometric assay for the phosphatidylinositol-specific phospholipase C from Bacillus cereus . Initial rates of the cleavage of the 2-naphthyl substrate by the phospholipase measured by fluorometry were linear with time and the amount of enzyme added . The specific enzyme activity at pH 8.5 and 25 degrees C was about 0.04 mumol/min mg protein at an initial substrate concentration of 0.8 mM . 31P NMR experiments suggest that, as with phosphatidylinositol itself, cleavage of the fluorescent substrate proceeds in two steps via a myo-inositol-1,2-cyclic phosphate intermediate, and that only the D-isomer is a substrate for the B . cereus phospholipase . The synthetic substrate was stable during long-term storage as a solid in the dark at -20 degrees C . It was also stable for several weeks when stored in the dark frozen in aqueous solution near neutral pH.

FEMS Microbiol Lett, 1991 Oct 1, 67(2), 187 - 91
Taxonomic considerations of Bartonella bacilliformis based on phylogenetic and phenotypic characteristics; Birtles RJ et al.; The 16S-rRNA gene of Bartonella bacilliformis was amplified using the polymerase-chain reaction (PCR) . The amplification product was sequenced using a linear-PCR procedure and compared with other published 16S-rRNA sequences . The results of this analysis placed B . bacilliformis in the alpha subgroup of the proteobacteria, and more specifically demonstrated its close phylogenetic relationship to Rochalimaea quintana . This relationship is supported by similarities in the size and mean base composition of the genomes of the two species, and by shared phenotypic characteristics.

Can J Microbiol, 1991 Oct, 37(10), 775 - 9
Reversibility of oxygen switch-off effect on Bacillus polymyxa nitrogenase; Holzmann HP et al.; The objective of this study was to analyse in vivo the effect of oxygen on the nitrogenase of Bacillus polymyxa . The culture technique employed in this study prevented spore formation by B . polymyxa during the entire period of exposure to acetylene . Under these conditions the acetylene-reduction assay allowed quantification of nitrogenase activity over long incubation periods (44 h) . Nitrogenase activity was highest in cells harvested in the late logarithmic phase . At PO2 of 0.19 and 0.37 kPa, acetylene reduction was inhibited by 80 and 100%, respectively . This switch-off effect could be reversed through oxygen exhaustion, either by flushing the culture with N2 or by cellular respiration, suggesting a respiratory protection mechanism for the nitrogenase complex in B . polymyxa . Oxygen consumption measured by a closed-chamber respirometer showed a linear increase up to a PO2 of 0.2 kPa . Above 0.3 kPa a saturation in oxygen consumption was observed . Exposure to high oxygen pressures resulted in an irreversible loss of nitrogenase activity . The oxygen inhibition pattern was shown to be similar to that in other microaerophilic and anaerobic nitrogen-fixing microorganisms.

J Gen Microbiol, 1991 Oct, 137 ( Pt 10), 2375 - 9
Molar growth yields and bioenergetic parameters of extremely alkaliphilic Bacillus species in batch cultures, and growth in a chemostat at pH 10.5; Guffanti AA et al.; Alkaliphilic Bacillus species that grow at pH 10.5 must cope with a low protonmotive force (-50 mV) due to a reversed transmembrane pH gradient at least 2 pH units more acid inside . Here we demonstrate that strictly alkaliphilic B . firmus RAB and two strains of B . alcalophilus (ATCC 27467 and DSM 485) grow exponentially in batch cultures with a doubling time of less than 1 h in 100 mM buffered medium, while the actual medium pH remains above 10.2 . The ATCC strain continued to grow rapidly for at least 7 h, but the growth rate of the DSM strain declined dramatically after 3 h . However, both the B . alcalophilus strains, B . firmus RAB and facultatively alkaliphilic B . firmus OF4 were readily maintained for at least 24 h between pH 10.4 and 10.6 in a chemostat where nutrients were constantly replenished . A critical nutrient may be limiting in batch cultures of the DSM strain of B . alcalophilus . The facultative alkaliphile grew equally well in batch cultures at an initial pH of 7.5 or 10.5 . Its molar growth yield (23 mg dry wt mmol-1) on malate (Ymal) was the same at the two pH values and was comparable to Ymal for B . subtilis grown at neutral pH . B . firmus RAB and B . alcalophilus ATCC 27467 grown at pH 10.5 also showed Ymal values at least as high as the neutralphile, indicating efficient use of the energy source even at low protonmotive force . Moreover, the phosphorylation potential of B . firmus OF4 grown at pH 7.5 (45.2 kJ mol-1) or pH 10.5 (46 kJ mol-1) was in a conventional range for bacteria.

Chirurg, 1991 Oct, 62(10), 732 - 8
{Helicobacter pylori colonization in surgical patients}; Ludtke FE et al.; The prevalence of Helicobacter pylori (HP) was examined in 387 patients undergoing endoscopy of the upper gastrointestinal tract . Of central interest was the question to which extent surgical intervention influences the colonisation of the gastric mucosa with HP . The bacillic status was appraised using double microbiological examinations, histological determination and the CLO-test . In 229 patients a 13C-urea-breath test was also carried out (sensitivity 98%) . HP could be detected in 90% of all patients presenting with duodenal ulcers as well as in 70% of patients with gastric ulcers, whereas in those patients in whom a lesion of the upper gastrointestinal tract could be excluded through endoscopy . HP was found in only 27% . The prevalence of HP did not increase with age . In patients having undergone distal gastric resection due to gastric ulcers, HP was only rarely found (19%) in the mucosa in the vicinity of the anastomosis following removal of the apparently pathogenetically important antrum mucosa . There was no association between anastomosis ulcers and bacillic colonisation . Following selective proximal vagotomy in patients with duodenal ulcers, HP was found in 80% of all cases . In these patients there was also no association between recurrent ulceration and a HP-positive status . Our results describe the postoperative HP-status after different surgical procedures of ulcer therapy: whereas a distal gastric resection removes the antrum mucosa, which provides the necessary environmental milieu, the HP-colonisation rate after selective proximal vagotomy is similar to that in non-operated ulcer disease.

Appl Environ Microbiol, 1991 Oct, 57(10), 2816 - 20
Identification of putative insect brush border membrane-binding molecules specific to Bacillus thuringiensis delta-endotoxin by protein blot analysis; Garczynski SF et al.; Binding sites for insecticidal toxins of Bacillus thuringiensis are located in the brush border membranes of insect midguts . Two approaches were used to investigate the interactions of B . thuringiensis subsp . kurstaki HD-73 CryIA(c) toxin with brush border membrane vesicles from sensitive and naturally resistant insects: 125I-toxin-vesicle binding assays and protein blots probed with 125I-CryIA(c) toxin . In bioassays, Manduca sexta and Heliothis virescens larvae were highly sensitive, Helicoverpa zea larvae were moderately sensitive, and Spodoptera frugiperda larvae were resistant to CryIA(c) toxin . Studies of binding of 125I-CryIA(c) toxin to brush border membrane vesicles from the larval midguts revealed that all insects tested had high-affinity, saturable binding sites . Significantly, S . frugiperda larvae bind but are not killed by CryIA(c) toxin . Labeled CryIA(c) toxin incubated with protein blots identifies a major binding molecule of 120 kDa for M . sexta and 148 kDa for S . frugiperda . H . virescens and H . zea are more complex, containing 155-, 120-, 103-, 90-, and 63-kDa proteins as putative toxin-binding molecules . H . virescens also contains a minor toxin-binding protein of 81 kDa . These experiments provide information that can be applied toward a more detailed characterization of B . thuringiensis toxin-binding proteins.

Appl Environ Microbiol, 1991 Oct, 57(10), 2783 - 9
Tributyltin-resistant bacteria from estuarine and freshwater sediments; Wuertz S et al.; Resistance to tributyltin (TBT) was examined in populations from TBT-polluted sediments and nonpolluted sediments from an estuary and from fresh water as well as in pure cultures isolated from those sediments . The 50% effective concentrations (EC50s) for populations were higher at a TBT-polluted freshwater site than at a site without TBT, suggesting that TBT selected for a TBT-resistant population . In contrast, EC50s were significantly lower for populations from a TBT-contaminated estuarine site than for those from a site without TBT, suggesting that other factors in addition to TBT determine whether populations become resistant . EC50s for populations from TBT-contaminated freshwater sediments were nearly 30 times higher than those for populations from TBT-contaminated estuarine sediments . We defined a TBT-resistant bacterium as one which grows on trypticase soy agar containing 8.4 microM TBT, a concentration which prevented the growth of 90% of the culturable bacteria from these sediments . The toxicity of TBT in laboratory media was influenced markedly by the composition of the medium and whether it was liquid or solid . Ten TBT-resistant isolates from estuarine sediments and 19 from freshwater sediments were identified to the genus level . Two isolates, each a Bacillus sp., may be the first gram-positive bacteria isolated from fresh water in the presence of a high concentration of TBT . There was a high incidence of resistance to heavy metals: metal resistance indices were 0.76 for estuarine isolates and 0.68 for freshwater isolates.(ABSTRACT TRUNCATED AT 250 WORDS)

Int J Syst Bacteriol, 1991 Oct, 41(4), 510 - 5
Bacillus brevis Migula 1900 taxonomy: reassociation and base composition of DNA; Nakamura LK; Of 87 strains previously identified as Bacillus brevis Migula 1900, 58 had G + C contents of 47.0 to 51.9 mol%, a range that included the G + C content (48.7 mol%) of the type strain . The G + C contents for three other groups consisting of 5, 7, and 17 strains were 37.0 to 41.9, 42.0 to 46.9, and 52.0 mol% or higher, respectively . DNA reassociation studies showed that 25 of the 58 strains with G + C contents of 47.0 to 51.9 mol% were closely related genetically to the type strain and to each other . For the most part, this genetically related group was phenotypically homogeneous; variations in the fermentation of mannitol and mannose were observed . My results strongly suggest that many of the strains were misclassified as B . brevis . Consequently, much of the phenotypic heterogeneity of the species B . brevis Migula 1900 is not due to variations exhibited by genetically related organisms, but is the result of variability introduced by the presence of genetically unrelated strains.

Rev Saude Publica, 1991 Oct, 25(5), 367 - 70
{Susceptibility of populations of Simulium (Chirostilbia) pertinax Kollar, 1832 (Culicomorpha, Simuliidae) to eemephos and to Bacillus thuringiensis var . israelensis-based formulation}; de Andrade CF et al.; The use of wooden troughs on stream beds, artificially colonized by blackfly larvae, is proposed for larvicide evaluations . Mortality was recorded 3 or 4 hours after treatment . Larval susceptibility was also evaluated utilizing the LT50 criterion . In there field assays Simulium (C.) pertinax populations from the litoral of S . Paulo and Rio de Janeiro States were shown to be resistant to temephos, even when subjected to high concentrations . Vectobac 12 AS, a Bacillus thuringiensis var . israelensis product, was shown to be more potent against late instar larvae and efficient in concentrations higher than 7,200 ITU/l (10 min) . The LT50 to 3,744 ITU/l (10 min) was calculated as 70.9 min.

Rev Latinoam Microbiol, 1991 Oct-Dec, 33(4), 279 - 92
{Bacillus thuringiensis: biological characteristics and production perspectives}; Rodriguez Monroy M et al.; In this paper the potential activity of Bacillus thuringiensis as insecticide is reviewed . It is presented recent knowledges of its metabolism, production system and the rinetic constants already reported finally the possibilities for production in batches or continuous culture is discussed in order to improve the productivity and economical aspects.

Indian J Biochem Biophys, 1991 Oct-Dec, 28(5-6), 472 - 5
Isolation of Gln- mutants through transposon, Tn917, mutagenesis in Bacillus brevis; Mishra HS et al.; Transposon, Tn917, carried on pTV1 plasmid has been used successfully to mutagenise Bacillus brevis . The transposon showed preference for insertion at an "aro" site . A second insertional event after elimination of the preferred site with ethidium bromide/acridine orange treatment has permitted isolation of Gln- mutants in B . brevis.

J Bacteriol, 1991 Oct, 173(20), 6635 - 8
Cloning and analysis of delta-endotoxin genes from Bacillus thuringiensis subsp . alesti; Lee CS et al.; Bacillus thuringiensis subsp . alesti produced only CryIA(b)-type protoxins, and three cryIA(b) genes were cloned . One was cryptic because of an alteration near the 5' end, and the other two were very similar to each other . The protoxin encoded by one of the latter genes differed from other CryIA(b) protoxins in its greater stability and relative toxicity for two members of the order Lepidoptera.

Appl Microbiol Biotechnol, 1991 Oct, 36(1), 5 - 13
Biosurfactants from Bacillus licheniformis: structural analysis and characterization; Jenny K et al.; The surface-active compounds of the strain Bacillus licheniformis were isolated and their structure was elucidated . The high surface-active capacity of the crude extract was basically due to traces of long-chain saturated fatty acids, especially of palmitic and stearic acids, to a mixture of small amounts of hydrocarbons with chain lengths of 20 and 22 carbons, and mainly to appreciable amounts of four slightly different lipopeptides . The lipopeptides were found to be a mixture of four closely related compounds . The lipophilic part consisting of i-, n-C14 or i-, ai-C15 beta-OH fatty acids was linked to the hydrophilic peptide moiety, which contained seven amino acids (Glu, Asp, Val, three Leu and Ile) by a lactone linkage . Fifteen milligrams per litre of the purified lipopeptide product decreased the surface tension of water from 72 mN m-1 to 27 mN m-1, characterizing the product as a powerful surface-active agent that compares favourably to other (bio)surfactants . Antibiotic activity was demonstrated against bacteria and yeasts.

J Biol Chem, 1991 Sep 25, 266(27), 17954 - 8
Functional domains of Bacillus thuringiensis insecticidal crystal proteins . Refinement of Heliothis virescens and Trichoplusia ni specificity domains on CryIA(c); Ge AZ et al.; Insecticidal crystal proteins (delta-endotoxins), CryIA(a) and CryIA(c), from Bacillus thuringiensis are 82% homologous . Despite this homology, CryIA(c) was determined to have 10-fold more insecticidal activity toward Heliothis virescens and Trichoplusia ni than CryIA(a) . Reciprocal recombinations between these two genes were performed by the homolog-scanning technique . The resultant mutants had different segments of their primary sequences exchanged . Bioassays with toxin proteins from these mutants revealed that amino acids 335-450 on CryIA(c) are associated with the activity against T . ni, whereas amino acids 335-615 on the same toxin are required to exchange full H . virescens specificity . One chimeric protein toxin, involving residues 450-612 from CryIA(c), demonstrated 30 times more activity against H . virescens than the native parental toxin, indicating that this region plays an important role in H . virescens specificity . The structural integrity of mutant toxin proteins was assessed by treatment with bovine trypsin . All actively toxic proteins formed a 65-kDA trypsin-resistant active toxic core, similar to the parental CryIA(c) toxin, indicating that toxin protein structure was not altered significantly . Contrarily, certain inactive mutant proteins were susceptible to complete protease hydrolysis, indicating that their lack of toxicity may have been due to structural alterations.

Biochemistry, 1991 Sep 24, 30(38), 9195 - 200
Binding of a chaperonin to the folding intermediates of lactate dehydrogenase; Badcoe IG et al.; When Bacillus stearothermophilus LDH dimer is incubated with increasing concentrations of the denaturant guanidinium chloride, three distinct unfolded states of the molecule are observed at equilibrium {Smith, C . J., et al . (1991) Biochemistry 30, 1028-1036} . The kinetics of LDH refolding are consistent with an unbranched progression through these states . The Escherichia coli chaperonin, GroEL, binds with high affinity to the completely denatured form and more weakly to the earliest folding intermediate, thus retarding the refolding process . A later structurally defined folding intermediate, corresponding to a molten globule form, is not bound by GroEL; neither is the inactive monomer . The complex between GroEL and denatured LDH is destabilized by the binding of magnesium/ATP (Mg/ATP) or by the nonhydrolyzable analogue adenylyl imidodiphosphate (AMP-PNP) . From our initial kinetic data, we propose that GroEL exists in two interconvertible forms, one of which is stabilized by the binding of Mg/ATP but associates weakly with the unfolded protein . The other is destabilized by Mg/ATP and associates strongly with unfolded LDH . The relevance of these findings to the role of GroEL in vivo is discussed.

FEBS Lett, 1991 Sep 23, 290(1-2), 221 - 3
Overproduction, purification and crystallization of Bacillus cereus oligo-1,6-glucosidase; Watanabe K et al.; The gene coding for oligo-1,6-glucosidase from Bacillus cereus ATCC7064 has been overexpressed in Escherichia coli MV1184 cells under the control of the lac promoter in the genetically engineered plasmid pBCE4-2 . Oligo-1,6-glucosidase was purified in large quantities and was crystallized at 25 degrees C by using a hanging drop vapor diffusion method with 53% saturated ammonium sulfate . The crystals have the shape of hexagonal bipyramids and belong to the space group P6(2) or P6(4) with lattice constants of a = b = 106.1 A, c = 120.0 A and gamma = 120 degrees.

FEBS Lett, 1991 Sep 23, 290(1-2), 13 - 6
Hydrolysis of branched cyclodextrins by a cyclodextrin-hydrolyzing enzyme from Bacillus sphaericus E-244; Oguma T et al.; The action of a cyclodextrin-hydrolyzing enzyme from Bacillus sphaericus E-244 on branched alpha- and beta-cyclodextrins was investigated . Glucosyl-alpha-cyclodextrin (6-O-alpha-D-glucosylcyclomaltohexaose) and maltosyl-alpha-cyclodextrin (6-O-alpha-D-maltosylcyclomaltohexaose) were hydrolyzed to 6(3)-O-alpha-D-glucosylmaltohexaose and 6(3)-O-alpha-D-maltosylmaltohexaose, respectively . Glucosyl-beta-cyclodextrin (6-O-alpha-D-glucosylcyclomaltoheptose) and maltosyl-beta-cyclodextrin (6-O-alpha-D-maltosyclomaltohepatose) were also mainly transformed to 6(4)-O-alpha-D-glucosylmaltoheptaose and 6(4)-O-alpha-D-maltosylmaltoheptaose, respectively . These results suggest that the cyclodextrin-hydrolyzing enzyme cleaves branched alpha- and beta-cyclodextrins at an alpha-1,4 linkage which is located furthest from the branching point on the cyclodextrin ring.

Biochim Biophys Acta, 1991 Sep 20, 1079(3), 247 - 52
Duck liver malic enzyme: sequence of a tryptic peptide containing the cysteine residue labeled by the substrate analog bromopyruvate; Satterlee J et al.; Malic enzyme of duck liver is alkylated by bromopyruvate with half-of-the-sites stoichiometry, and with accompanying loss of oxidative decarboxylase and enhancement of pyruvate reductase activities as was previously shown for the pigeon enzyme (Hsu, R.Y . (1982) Mol . Cell . Biochem . 43, 3-26) . In the present work, the alkylated enzyme is shown to bind NADPH, but not L-malate in the presence of MnCl2, indicating impairment of the enzyme site for the substrate and/or divalent metal . The enzyme was differentially labeled by 3-bromo-1-{14C}-pyruvate and digested with TPCK-treated trypsin . Two peptides bearing the susceptible residue were purified by high-performance liquid chromatography and sequenced . Peptide II has the sequence of FMPIVYTPTVGLAXQQYGLAFR, corresponding to residues 86-107 (temporary numbering) of the duck enzyme; cysteine-99(x) is not detected, indicating that it is the target of modification by bromopyruvate . Peptide I is a truncated form of peptide II lacking five amino acid residues at the C-terminal . Cysteine-99 is conserved in malic enzymes from duck, rat, mouse, maize, human, Flaveria trinervia and Bacillus stearothermophilus.

Biochem Biophys Res Commun, 1991 Sep 16, 179(2), 933 - 8
The toxic moiety of the Bacillus thuringiensis protoxin undergoes a conformational change upon activation; Choma CT et al.; Proteolytic processing of the 133-kDa crystal protein (protoxin) from Bacillus thuringiensis subsp . kurstaki yields a 67-kDa insecticidal toxin . Differential scanning calorimetry was used to investigate whether the toxic moiety in the protoxin molecule has the same conformation as activated toxin . Compared to protoxin, toxin gives rise to a more complex endotherm which extends over a 10 degrees C broader temperature range and contains a component occurring at a substantially higher temperature than any unfolding transition in the protoxin endotherm . It is concluded that the toxic moiety undergoes a conformational change upon activation in which the thermal stability of at least one of its domains is significantly increased.

Biochem J, 1991 Sep 15, 278 ( Pt 3), 809 - 16
Evidence that gene G7a in the human major histocompatibility complex encodes valyl-tRNA synthetase; Hsieh SL et al.; At least 36 genes have now been located in a 680 kb segment of DNA between the class I and class II multigene families within the class III region of the human major histocompatibility complex on chromosome 6p21.3 . The complete nucleotide sequence of the 4.3 kb mRNA of one of these genes, G7a (or BAT6), has been determined from cDNA and genomic clones . The single-copy G7a gene encodes a 1265-amino-acid protein of molecular mass 140,457 Da . Comparison of the derived amino acid sequence of the G7a protein with the National Biomedical Research Foundation protein databases revealed 42% identity in a 250-amino-acid overlap with Bacillus stearothermophilus valyl-tRNA synthetase, 38.0% identity in a 993-amino-acid overlap with Escherichia coli valyl-tRNA synthetase (val RS), and 48.3% identity in a 1043-amino-acid overlap with Saccharomyces cerevisiae valyl-tRNA synthetase . The protein sequence of G7a contains two short consensus sequences, His-Ile-Gly-His and Lys-Met-Ser-Lys-Ser, which is the typical signature structure of class I tRNA synthetases and indicative of the presence of the Rossman fold . In addition, the molecular mass of the G7a protein is the same as that of other mammalian valyl-tRNA synthetases . These features and the high sequence identity with yeast valyl-tRNA synthetase strongly support the fact that the G7a gene, located within the major histocompatibility complex, encodes the human valyl-tRNA synthetase.

J Urol, 1991 Sep, 146(3), 766 - 9; discussion 769-70
Bacillus Calmette-Guerin and interleukin-2 for treatment of superficial bladder cancer; Cockett AT et al.; A total of 22 patients with bladder cancer received bacillus Calmette-Guerin (BCG) and interleukin-2 . Significant bladder tumor remissions were noted in 15 of 17 patients (88%) . Of 5 patients with carcinoma in situ 1 was noncompliant and he died of carcinoma in situ . The other 4 patients are in remission . BCG alone was instilled in 22 additional patients with superficial bladder cancer . The remission rates were encouraging . Of the 22 patients 13 (59%) had remission of the bladder tumor . A half dose of BCG (60 mg.) is adequate when given weekly for 6 weeks . Maintenance therapy is important as noted in both of our clinical arms . BCG and interleukin-2 therapy results in a higher remission rate.

FEBS Lett, 1991 Sep 9, 289(2), 148 - 50
Modification of alpha-amylase from Bacillus licheniformis by the polyaldehyde derived from beta-cyclodextrine and alpha-amylase thermostability; Morand P et al.; The cleavage of beta-cyclodextrine by sodium periodate at the seven 2-3 diols of the glucose unit gives rise to the polyaldehyde 1, used to modify alpha-amylase . The reductive modification of alpha-amylase from Bacillus licheniformis reduced the number of reactive lysine groups from 8 to 3.5 per mol of enzyme with an activity loss of 25% and increased the half-life at 80 degrees C from 4.7 to 7.0 minutes.

J Biol Chem, 1991 Sep 5, 266(25), 16356 - 62
Molecular dissection of translation initiation factor IF2 . Evidence for two structural and functional domains; Gualerzi CO et al.; By means of limited proteolysis of Bacillus stearothermophilus initiation factor IF2 and genetic manipulation of its structural gene, infB, we have been able to produce (or hyperproduce) and purify two polypeptide fragments corresponding to two structurally and functionally separate domains of the protein . The first is the G-domain (approximately 41 kDa), which makes up the central part of the molecule and contains the conserved structural elements found in all GTP/GDP-binding sites of G-proteins . This domain is resistant to proteolysis in the presence of GTP or GDP, retains the capacity to interact with the 50 S subunit, binds weakly to the 30 S subunit, and displays ribosome-dependent GTPase activity with an approximately 2-fold higher Km for GTP and the same Vmax as compared with intact IF2 . The second is the C-domain (approximately 24 kDa), which corresponds to the COOH-terminal part of IF2 and constitutes an extraordinarily compact domain containing the fMet-tRNA binding site of IF2 . In spite of its negligible affinity for the ribosomes, the C-domain weakly stimulates the ribosomal binding of fMet-tRNA, presumably by affecting the conformation of the initiator tRNA molecule.

J Appl Bacteriol, 1991 Sep, 71(3), 285 - 8
Effect of simultaneous application of heat and pressure on the survival of bacterial spores; Mallidis CG et al.; The effect of simultaneous application of moderate hydrostatic pressure (10-300 atm) and heat on the survival of the Bacillus stearothermophilus spores in a flow-through system was investigated . A high heterogeneity of the sensitization of spores to heat by pressure was found . A higher degree of reduction of heat resistance was observed at the low than at the high temperatures tested . The simultaneous application of moderate pressure and heat can not be applied for the preservation of liquid foods due to the extreme heterogeneity of spore sensitization to heat by pressure.

J Appl Bacteriol, 1991 Sep, 71(3), 202 - 6
Formulation of a flowable liquid concentrate of Bacillus thuringiensis serotype H-14 spores and crystals as mosquito larvicide; Ejiofor AO et al.; Bacillus thuringiensis serotype H-14 spores and crystals, produced in 51 fermenters, were centrifuged and resuspended in emulsified palm olein to give 3.2 x 10(11) colony forming units (cfu)/ml . The suspension was mixed with a cassava-molasses-palm olein-charcoal (CMPC-2) mixture which served as the carrier, adhesive, dispersant and protectant . The final concentration of the formulation was 3.2 x 10(9) cfu/ml . The lethal concentrations capable of killing 50% of the test population (LC50) of CMPC-2 during 0, 1 and 2 years of storage at 32 +/- 4 degrees C were 0.056, 0.058 and 0.058 mg/ml respectively as against 0.054, 0.051 and 0.054 mg/ml for the Institut Pasteur Standard-1978 (IPS-78) during the corresponding period . The chi 2 tests showed that the results were homogeneous at P = 0.05 . The relative potencies of the preparations were 964.3, 879.3 and 931 International toxic units (ITU) Aedes aegypti as compared with the 1000 ITU assigned to IPS-78 . At 95% confidence limits there was no significant difference between the potencies of CMPC-2 and IPS-78 . Field tests showed that CMPC-2 provided between 87.5 and 100% control of natural populations of Aedes spp . and Cutex spp . Sedimentation tests showed that CMPC-2 settled markedly during storage . This, therefore, required that the product be thoroughly shaken before use.

Antimicrob Agents Chemother, 1991 Sep, 35(9), 1933 - 6
In vitro activity of the new difluorinated quinolone sparfloxacin (AT-4140) against Mycobacterium tuberculosis compared with activities of ofloxacin and ciprofloxacin; Rastogi N et al.; MICs of the new fluoroquinolone drugs ofloxacin, ciprofloxacin, and sparfloxacin (AT-4140) for 10 strains of Mycobacterium tuberculosis were determined by using both a BACTEC radiometric method and testing on solid 7H11 agar medium . Radiometric MICs by 7H12 broth testing ranged from 0.5 to 1.0, 0.25 to 0.5, and 0.1 to 0.2 microgram/ml for ofloxacin, ciprofloxacin, and sparfloxacin respectively, whereas MICs in solid medium ranged from 0.5 to 1.0, 0.5 to 1.0, and 0.2 to 0.5 microgram/ml, respectively . The bactericidal action of the quinolones compared with their reported peak concentrations in human serum showed that sparfloxacin is the most bactericidal, followed by ciprofloxacin and ofloxacin . Our results suggest the potential of the new difluorinated quinolone sparfloxacin for use against the tubercle bacillus and indicate that further determination of its antimycobacterial spectrum and intracellular efficacy may be fruitful.

Quintessence Int, 1991 Sep, 22(9), 721 - 6
Evaluation of a dental unit with a built-in decontamination system; Douglas CW et al.; The efficacy of a dental unit equipped with a system that disinfects and sterilizes the water tubing by flushing with glutaraldehyde was evaluated by inserting Bacillus megaterium spores and Pseudomonas and Moraxella species into the water tubing . Up to 10(8) Pseudomonas and Moraxella organisms were killed during the disinfection cycle, but Bacillus megaterium spores were not . Up to 10(5) spores were eradicated by the sterilization cycle, although the system did not consistently kill 10(8) spores . The water tubing of the new unit was not naturally colonized by water bacteria during an 8-month period prior to the study . Evidence suggested that this was due to antimicrobial activity associated with the plastic tubing; therefore, microbial contamination of new dental units, irrespective of their design, would not be expected, until the inhibitory factor in the plastic tubing has leached out.

Kekkaku, 1991 Sep, 66(9), 577 - 87
{Preventable tuberculosis cases in Japan--a new approach to the assessment of tuberculosis control problems}; Ahiko T; I defined cases of tuberculosis which could not be prevented from infection or development of disease among infected, or could not be detected in the early stages as "preventable cases" in order to evaluate tuberculosis control efforts in the community, Japan . Among 241 bacteriologically confirmed cases with pulmonary tuberculosis newly registered from 1988 through 1989 in Yamagata Prefecture, 80 (33%) were defined as preventable cases by observing their course and the process of diagnosis . That is to say, one-third of bacillary cases could have been prevented in Yamagata where the incidence of tuberculosis was lowest in Japan, if existing prevention and control methods had been effectively used . Causes of prevention failure were investigated in detail . The most common cause was tardy detection of cases, especially due to delayed confirmation of diagnosis (so called "doctor's delay") . The delayed confirmation of diagnosis resulted from neglecting chest X-ray and sputum examination and from ignoring high risk groups . In the younger age group, it was mainly attributed to insufficient family contact examinations . Tuberculin skin tests are necessary not only for those aged 15 years and younger but also for those aged 16-29, when they are found to be household contacts of smear-positive cases . An evaluation of tuberculosis control program defining preventable cases would be a beneficial approach to the surveillance of tuberculosis.

Semin Dermatol, 1991 Sep, 10(3), 194 - 8
Bacillary angiomatosis: a systemic opportunistic infection with prominent cutaneous manifestations; LeBoit PE; Bacillary angiomatosis is an opportunistic infection with systemic manifestations . Although most cases have occurred in human immunodeficiency virus-positive patients, other immunosuppressed patients, and even seemingly immunocompetent individuals, can become infected . In addition to the well-characterized cutaneous manifestations, visceral involvement can occur and may be the only locus of infection . Lymphadenopathy, bone or soft-tissue masses, fever, and hepatosplenomegaly can be presenting signs . The causative bacterium is still unidentified, but resemblances to the rickettsiae, Rochilamea quintana, the recently identified cat-scratch disease bacillus, and Bartonella bacilliformis have been noted by various investigators . Systemic disease is treatable and can be cured with antibiotic therapy.

Infect Immun, 1991 Sep, 59(9), 2864 - 9
Effects of activated macrophages on Mycobacterium leprae; Ramasesh N et al.; Five alternative methods were used to explore in vitro the effects of normal and activated murine macrophages on the metabolic well-being of intracellular Mycobacterium leprae: fluorescein diacetate-ethidium bromide staining, ATP content, synthesis of phenolic glycolipid 1, and two techniques to quantitate oxidation of palmitic acid . In relatively short-term experiments (7 to 10 days), each of these procedures provided strong evidence that activated macrophages exerted a deleterious effect on the leprosy bacillus . These findings appear to confirm the contention that activated macrophages underlie host resistance to clinical leprosy and limitation of M . leprae growth in paucibacillary leprosy.

Eur J Biochem, 1991 Sep 1, 200(2), 545 - 51
Competitive inhibition of liver glucokinase by its regulatory protein; Vandercammen A et al.; The regulatory protein of rat liver glucokinase (hexokinase IV or D) behaved as a fully competitive inhibitor of this enzyme when glucose was the variable substrate, i.e . it increased the half-saturating concentration of glucose as a linear function of its concentration without affecting V (velocity at infinite concentration of substrate) . The inhibition by the regulatory protein and that by palmitoyl-CoA were synergistic with that by N-acetyl-glucosamine, indicating that the two former inhibitors bind to a site distinct from the catalytic site . In contrast, the effects of the regulatory protein and palmitoyl-CoA were competitive with each other, indicating that these two inhibitors bind to the same site . The regulatory protein exerted a non-competitive inhibition with respect to Mg-ATP at concentrations of this nucleotide less than 0.5 mM . At higher concentrations, the latter antagonized the inhibition by the regulatory protein partly by decreasing the apparent affinity for fructose 6-phosphate . The following anions inhibited glucokinase non-competitively with respect to glucose: Pi, sulfate, I-, Br-, No3-, Cl-, F- and acetate . Pi and sulfate, at concentrations in the millimolar range, decreased the inhibition by the regulatory protein by competing with fructose 6-phosphate . Monovalent anions also antagonized the inhibition by the regulatory protein with the following order of potency: I- greater than Br- greater than NO3- greater than Cl- greater than F- greater than acetate and their effect was non-competitive with respect to fructose 6-phosphate . Glucokinase from Buffo marinus and pig liver were, like the rat liver enzyme, inhibited by the regulatory protein, as well as by palmitoyl-CoA at micromolar concentrations . In contrast, neither compound inhibited hexokinases from rat brain, beef heart or yeast, or the low-Km specific glucokinase from Bacillus stearothermophilus.

Urology, 1991 Sep, 38(3), 271 - 9
Multicenter study of superficial bladder cancer treated with intravesical bacillus Calmette-Guerin or adriamycin . Results of long-term follow-up; Khanna OP et al.; We evaluated 158 cases of patients with superficial bladder cancers (Stages Ta, T1, and Tis) . These cases were treated with either intravesical bacillus Calmette-Guerin (BCG) (Tice strain) or Adriamycin (ADR), in a multicenter, nonrandomized study . One hundred thirty-one of these patients were followed up; the results continue to show a higher percentage of initial complete remissions with BCG (68%) than with ADR (57%) . With additional therapy, both BCG and ADR achieved complete remission in 83 percent of the patients . When 7 failures with patients taking ADR were switched to BCG and the disease cleared, the rate of complete remission for BCG rose to 85 percent . The recurrence rate per 100 patient-months was only slightly different for BCG (0.9) and ADR (0.8) . The percentage of progressions continued to be higher for BCG (8%) than for ADR (5%) . Cystectomies were performed in 2.5 percent of the BCG patients . Using the Cox regression model with covariates, we found drug treatment, tumor grade, and sex to be statistically significant in determining failures throughout the protocol . Although both BCG and ADR were effective over the course of the study, BCG is the drug of choice for residual tumor (Stages T1 and Tis).

J Urol, 1991 Sep, 146(3), 700 - 2; discussion 702-3
The management of transitional cell carcinoma in solitary renal units; Schoenberg MP et al.; Ten patients with urothelial malignancies involving a solitary functioning renal unit were treated at our center for an average of 24 months or until death . These patients were all managed by parenchyma-sparing methods, including percutaneous as well as ureteroscopic tumor resection . Of our patients 9 have received adjunctive chemotherapy in the form of bacillus Calmette-Guerin instillations . At the time of this report 5 of our patients were alive without evidence of disease, 4 were alive with evidence of either residual or recurrent neoplasia and 1 was dead of disease 5 years after original presentation . Patients with higher grade tumors or carcinoma in situ did less well than patients with low grade disease . We present an analysis of our experience with this complex patient population and discuss the implications of these data within the context of a growing literature on the topic of upper tract urothelial malignancy.

Bioorg Khim, 1991 Sep, 17(9), 1188 - 92
{BsoAI--a new site specific endonuclease from Bacillus coagulans}; Sokolov NN et al.; A new site-specific endonuclease was detected in toluene lysates of Bacillus coagulans AUCM B-732 and designated as BcoAI . The enzyme was purified by fractionation of the cell-free extract in the two-phase PEG/dextran system followed by chromatography on DEAE-sepharose and phosphocellulose and shown to be free of nonspecific nucleases and phosphatases . BcoAI has three cleavage sites on lambda DNA, but does not cleave SV40, pBR322 and pUC19 DNA . BcoAI recognizes the sequence 5' CAC decreases GTG 3' on double-stranded DNA and cleaves it as indicated by the arrow to yield blunt-ended DNA fragments . Thus, BcoAI is a true isoschizomer of PmaCI from Pseudomonas maltophila C.

Indian J Malariol, 1991 Sep, 28(3), 167 - 70
Acute toxicity of certain organochlorine, organophosphorus, synthetic pyrethroid and microbial insecticides to the mosquito fish Gambusia affinis (Baird and Girard); Mittal PK et al.; Acute toxicity of certain organochlorine, organophosphorus, synthetic pyrethroid and microbial insecticides to the mosquito fish Gambusia affinis were determined to collect baseline data for selecting the resistant strains of the fish . The synthetic pyrethroid, Lambdacyhalothrin was most toxic to the fish (LC50 = 0.0022 ppm), followed by deltamethrin, cypermethrin and fenvalerate . Organochlorine insecticides, DDT and gamma-HCH, were less toxic than the pyrethroids, and these were followed by organophosphorus insecticides, malathion, fenthion, monocrotophos and temephos . The last two insecticides were least toxic among the different chemical insecticides (LC50 greater than 80 ppm ai) . The microbial insecticide ABG-6262 (Vectolex 2.5 AS), a Bacillus sphaericus preparation, was totally harmless to the fish at 2500 microliters/l up to one week.

Indian J Malariol, 1991 Sep, 28(3), 147 - 50
Isolation and laboratory evaluation of an indigenous strain of Bacillus sphaericus (9001); Gupta DK et al.; An indigenous strain of Bacillus sphaericus H5a (9001), possessing high insecticidal properties, was isolated from diseased larvae of Culex species . This strain in comparison with the known strains of B . sphaericus, i.e., 1593 and 2362, was found to be promising against the fourth instar larvae of Anopheles culicifacies, An . stephensi, An . subpictus, Aedes aegypti and Culex quinquefasciatus . B . sphaericus 9001 is highly stable and virulent through 25 successive transfers and thus can be effectively used as a biocontrol agent against immature stages of mosquitoes.

Southeast Asian J Trop Med Public Health, 1991 Sep, 22(3), 429 - 35
Toxicity of Bacillus sphaericus strain 2362 on Mansonia spp . larvae; Petcharat J; The efficiency of Bacillus sphaericus strain 2362 (Vectolex) as larvicide against Mansonia spp . was studied . Bioassay studies showed that the toxicity of B . sphaericus on both age groups (I-II instar and III-IV instar) of Mansonia spp . larvae occurred within 24 hours . Probit analysis revealed that LC100 (one hundred per cent lethal concentration) for both age groups of M . boneae were higher than those of M . dives . Small scale field trials were done at Kreng Village, Cha-uat District, Nakhon Si Thammarat Province, one of the most serious filarial infected areas . It was indicated that 100% kill of Mansonia spp . larvae in the field occurred within 9 days after the larvicide application . When a dose of 5 times of LC100 value was used, 100% control was achieved up to about one month.

Southeast Asian J Trop Med Public Health, 1991 Sep, 22(3), 426 - 8
Laboratory evaluation of Bacillus thuringiensis H-14 against Aedes aegypti larvae in the northeast region of Thailand; Pipitgool V et al.; Laboratory bioassays using a preparation of Bacillus thuringiensis H-14 (Bt.H-14), namely Skeetal were conducted to determine their effectiveness against late 3rd/early 4th instar larvae of Aedes aegypti . The larvae were collected from municipal areas in 7 provinces, namely Burirum, Roi-Et, Khon Kaen, Ubol Ratchatani, Nakorn Phanom, Surin and Nakorn Ratchasima, in the Northeast of Thailand . It was found that for Skeetal, LC50 ranged from 128 to 151 nl/l (average 143) and LC90 ranged from 254 to 289 nl/l (average 275) . The mortality rate of Ae . aegypti larvae in the 7 provinces did not differ significantly (p greater than 0.05) at a concentration of 300 nl/l . The result of the bioassays show that the preparation of Bt.H-14 is very effective against Ae . aegypti larvae in Northeast of Thailand and the mosquito larvae in the various areas were nearly equal in susceptibility to Bt.H-14.

Yale J Biol Med, 1991 Sep-Oct, 64(5), 481 - 98
Consumption, silicosis, and the social construction of industrial disease; Rosner D et al.; In the wake of the bacterial revolution after Robert Koch identified the tuberculosis bacillus, medical and public health professionals classified the various forms of consumption and phthisis as a single disease--tuberculosis . In large measure, historians have adopted that perspective . While there is undoubtedly a great deal of truth in this conceptualization, we argue that it obscures almost as much as it illuminates . By collapsing the nineteenth-century terms phthisis and consumption into tuberculosis, we maintain that historians have not understood the effect of non-bacterial consumption on working-class populations who suffered from the symptoms of coughing, wasting away, and losing weight . In this essay, we explore how, in the nineteenth century, what we now recognize as silicosis was referred to as miners' "con," stonecutters' phthisis, and other industry-specific forms of phthisis and consumption . We examine how the later and narrower view of the bacterial origins of tuberculosis limited the medical professions' ability to diagnose and understand diseases caused by industrial dust . This paper explores the contention that developed at the turn of the century over occupational lung disease and tuberculosis and the circumstances that led to the unmasking of silicosis as a disease category.

Zhonghua Nei Ke Za Zhi, 1991 Sep, 30(9), 572 - 4, 596
{Clinical effect and laboratory observation of ofloxacin in the treatment of typhoid fever, bacillary dysentery and gonorrhea}; Zou QY et al.; Ofloxacin was used in the treatment of 20 cases of typhoid fever, 32 cases of bacillary dysentery and 50 cases of gonococcal infection . Altogether 102 cases were treated, 53 being male and 49 female . The daily dosage was 400 mg to 600 mg, divided into two times . The result showed that the clinical effective rate for typhoid fever, bacillary dysentery and gonococcal infections was 100%, 97% and 94% respectively, while the bacterial eradication rate was 100%, 100% and 94% respectively . the bacterial eradication rate was 100%, 97% and 94% respectively . The side effects were mild in degree . The authors are of the opinion that since ofloxacin can be administered orally with only two times a day, its absorption is nearly complete and the cure rate is high, it should be considered as the drug of choice in the treatment of typhoid fever, bacillary dysentery and gonorrhea, especially in the drug resistant cases . It is suggested that this drug be used more widely.

Chem Pharm Bull (Tokyo), 1991 Sep, 39(9), 2468 - 70
Isolation of a new antitumor substance from Bacillus stearothermophilus; Kohama Y et al.; A new antitumor substance, BS-1, was isolated from the autolysate and culture filtrate of Bacillus stearothermophilus UK563 by ethylacetate extraction and HPLC . BS-1 inhibited the proliferation of mouse macrophage-like cells, P388-D1 (IC50: 4 micrograms/ml) and mouse mastocytoma, P-815 (IC50: 0.6 microgram/ml), but not that of Balb/c 3T3.

J Am Mosq Control Assoc, 1991 Sep, 7(3), 420 - 3
Delayed mortality and morphogenetic anomalies induced by the microbial control agent Bacillus thuringiensis ser . (H-14) in Culex quinquefasciatus; Mulla MS et al.; Sublethal concentrations of Bacillus thuringiensis ser . H-14 were applied to early 4th-instar larvae of Culex quinquefasciatus to assess mortality and morphogenetic aberrations in larvae, pupae and adults . At the 24 h LC10, LC25, LC50 and LC80, additional mortality occurred in surviving larvae beyond a 24 h exposure period . The cumulative mortalities increased daily and the overall mortality of larvae up to 7 days posttreatment were 12, 73, 82 and 96% at the indicated concentrations, respectively . Delayed mortality also occurred in the pupal and adult stages . Morphogenetic aberrations were noted in dead larvae and pupae but were rare in the adults . These aberrations are categorized and described . There was little or no delayed effect on the survivorship or fecundity of adults, but in all the treatments the number of emerging males was higher than females . The sex ratio in check adults was 1:1.

J Am Mosq Control Assoc, 1991 Sep, 7(3), 412 - 9
Delayed mortality and morphogenetic anomalies induced in Culex quinquefasciatus by the microbial control agent Bacillus sphaericus; Mulla MS et al.; Two preparations of Bacillus sphaericus 2362 were studied for their biological activity, delayed mortality and the induction of morphogenetic aberrations in larvae, pupae and adults of Culex quinquefasciatus . Longevity and fecundity of adult mosquitoes were also assessed . A dosage response line for B . sphaericus was established against 4th-instar larvae and sublethal concentrations (48 h LC50 and lower) were used against these larvae . Sublethal concentrations of B . sphaericus induced delayed mortality in larvae, pupae and adults . The magnitude of mortality increased in succeeding cohorts and developmental stages resulting from the surviving larvae . Only 10 and 25% overall emergence of viable adults occurred in the sublethal treatments (LC25) of 2 B . sphaericus preparations . The range of successful adult emergence was over 94% in the controls . A wide range of external morphogenetic aberrations in dead larvae, pupae and adults were noted . These aberrations and gross morphological features were quite similar to those reported for certain insect growth regulators . Sublethal concentrations had no marked effect on longevity of adults, egg deposition and hatch.

Indian J Med Res, 1991 Sep, 93, 318 - 23
Production & formulation of Bacillus thuringiensis var . israelensis & B . sphaericus 1593; Desai SY et al.; Three fermentation media each for bulk growth of B . thuringiensis var . israelensis and B . sphaericus 1593 were formulated using defatted groundnut cake (Arachis hypogea) as the first nitrogen source and gram flour (Cicer arientinum), soy bean (Glycine max) and defatted milk powder as the second nitrogen source . Medium containing gram flour showed highest toxicity (14.45 micrograms/l) in case of B . thuringiensis var . israelensis whereas medium containing milk powder was found to be highly toxic with B . sphaericus 1593 (51.39 micrograms/l) . Sustained release floating pellet formulations of B . thuringiensis var . israelensis and B . sphaericus 1593 exhibited toxicity of 77 per cent and above for 42 days at a dose of 500 micrograms/l for 4th instar larvae of Culex pipiens quinquefasciatus Say.

J Clin Microbiol, 1991 Sep, 29(9), 1777 - 9
Pneumonia and empyema infection associated with a Bacillus species that resembles B . alvei; Coudron PE et al.; An organism resembling Bacillus alvei was isolated from the lung and pleural fluid of an immunocompetent patient . The isolate differed from the type strain of B . alvei in its ability to reduce nitrate and its inability to produce dihydroxyacetone and acetylmethylcarbinol . The isolate was resistant to ciprofloxacin and showed intermediate susceptibility to vancomycin.

Appl Environ Microbiol, 1991 Sep, 57(9), 2767 - 70
Enhancement of soybean nodulation by Bacillus cereus UW85 in the field and in a growth chamber; Halverson LJ et al.; Seed treatments with Bacillus cereus UW85 increased nodulation of soybeans in three field seasons and in three different sterilized soils in the growth chamber . In the field, 28 and 35 days after planting, UW85-treated plants had 31 to 133% more nodules than untreated plants . From 49 days after planting until seed harvest, there were no significant differences between nodulation of UW85-treated plants and untreated control plants . In the growth chamber, in sterilized soil-vermiculite mixtures, at 28 days after planting, UW85 seed treatments enhanced nodulation by 34 to 61%, indicating that the increase in nodulation was not dependent on the soil flora.

Appl Environ Microbiol, 1991 Sep, 57(9), 2656 - 65
Utility of phenomenological models for describing temperature dependence of bacterial growth; Heitzer A et al.; We compared three unstructured mathematical models, the master reaction, the square root, and the damage/repair models, for describing the relationship between temperature and the specific growth rates of bacteria . The models were evaluated on the basis of several criteria: applicability, ease of use, simple interpretation of model parameters, problem-free determination of model parameters, statistical evaluation of goodness of fit (chi 2 test), and biological relevance . Best-fit parameters for the master reaction model could be obtained by using two consecutive nonlinear least-square fits . The damage/repair model proved to be unsuited for the data sets considered and was judged markedly overparameterized . The square root model allowed nonproblematical parameter estimation by a nonlinear least-square procedure and, together with the master reaction model, was able to describe the temperature dependence of the specific growth rates of Klebsiella pneumoniae NCIB 418, Escherichia coli NC3, Bacillus sp . strain NCIB 12522, and the thermotolerant coccobacillus strain NA17 . The square root and master reaction models were judged to be equally valid and superior to the damage/repair model, even though the square root model is devoid of a conceptual basis.

Scand J Immunol, 1991 Sep, 34(3), 365 - 72
MPB 64 possesses 'tuberculosis-complex'-specific B- and T-cell epitopes; Andersen AB et al.; We have developed monoclonal antibodies (MoAb) reactive with a protein from Mycobacterium tuberculosis of apparent molecular mass 24 kDa . This protein was shown to be identical with MPB 64 (Harboe et al.,) MoAb bound to four different epitopes of which two were restricted to the 'tuberculosis complex' and two were also found in mycobacteria not belonging to the 'tuberculosis complex' . The cross-reactive MoAb demonstrate that MPB 64 is present in more mycobacterial species than previously assumed . MPB 64 was shown to induce strong delayed type hypersensitivity (Dth) reactions in outbred guinea pigs immunized with M . tuberculosis and M . bovis bacille Calmette-Guerin (BCG) . No reaction was observed in animals immunized with mycobacteria not belonging to the 'tuberculosis complex' . The Dth-inducing capacity of MPB64 was compared with that of another 24 kDa protein purified from M . tuberculosis and of the previously described 38 kDa protein . The Dth responses to these three antigens were further analysed in four inbred guinea pig strains . A genetic restriction of the ability of the animals to respond to MPB 64 as well as to the 38 kDa protein was observed.

Microbiol Rev, 1991 Sep, 55(3), 425 - 36
Bacillus sphaericus as a mosquito pathogen: properties of the organism and its toxins; Baumann P et al.; In the course of sporulation, Bacillus sphaericus produces an inclusion body which is toxic to a variety of mosquito larvae . In this review we discuss the general biology of this species and concentrate on the genetics and physiology of toxin production and its processing in the midgut of the larval host . The larvicide of B . sphaericus is unique in that it consists of two proteins of 51 and 42 kDa, both of which are required for toxicity to mosquito larvae . There is a low level of sequence similarity between these two proteins, which differ in their sequences from all the other known insecticidal proteins of Bacillus thuringiensis . Within the midgut the 51- and 42-kDa proteins are processed to proteins of 43 and 39 kDa, respectively . The conversion of the 42-kDa protein to a 39-kDa protein results in a major increase in toxicity; the significance of the processing of the 51-kDa protein is not known . In contrast to the results with mosquito larvae, the 39-kDa protein is alone toxic for mosquito-derived tissue culture-grown cells, and this toxicity is not affected by the 51-kDa protein or its derivative, the 43-kDa protein . Comparisons of larvae from species which differ in their susceptibility to the B . sphaericus toxin indicate that the probable difference resides in the nature of the target sites of the epithelial midgut cells and not in uptake or processing of the toxin . A similar conclusion is derived from experiments involving tissue culture-grown cells from mosquito species which differ in their susceptibility to the B . sphaericus toxin.

Biochimie, 1991 Sep, 73(9), 1179 - 86
Glycosyl-phosphatidylinositol anchored acetylcholinesterase as substrate for phosphatidylinositol-specific phospholipase C from Bacillus cereus; Stieger S et al.; We investigated the enzymatic properties of phosphatidylinositol-specific phospholipase C (PI-PLC) from Bacillus cereus towards glycosyl-phosphatidylinositol anchored acetylcholinesterase (AChE) from bovine erythrocytes and Torpedo electric organ as substrate . The conversion of membrane from AChE to soluble AChE by PI-PLC depended on the presence of a detergent and of phosphatidylcholine . In presence of mixed micelles containing Triton X-100 (0.05%) and phosphatidylcholine (0.5 mg/ml) the rate of AChE conversion was about 3 times higher than in presence of Triton X-100 alone . Furthermore, inhibition of PI-PLC occurring at Triton X-100 concentrations higher than 0.01% could be prevented by addition of phosphatidylcholine . Ca2+, Mg2+ and sodium chloride had no effect on PI-PLC activity in presence of phosphatidylcholine and Triton X-100, whereas in presence of Triton X-100 alone sodium chloride largely increased the rate of AChE conversion . Determination of kinetic parameters with three different substrates gave Km-values of 7 microM, 17 microM and 2 mM and Vmax-values of 0.095 microM.min-1, 0.325 microM.min-1 and 56 microM.min-1 for Torpedo AChE, bovine erythrocyte AChE and phosphatidylinositol, respectively . The low Km-values for both forms of AChE indicated that PI-PLC not only recognized the phosphatidylinositol moiety of the anchor but also other components thereof.

Biochem J, 1991 Sep 1, 278 ( Pt 2), 461 - 3
A study of human erythrocyte acetylcholinesterase inhibition by chlorpromazine; Spinedi A et al.; Membrane-bound acetylcholinesterase (AChE) from the human erythrocyte is inhibited by chlorpromazine (CPZ) in a concentration range within this amphiphilic drug has been demonstrated to interact with erythrocyte membranes, causing a large spectrum of physical and structural effects; membrane solubilization with 0.5% Triton X-100 results in a complete loss of CPZ inhibitory potency . Although these observations might suggest a role of membrane lipid environment in mediating human erythrocyte AChE inhibition, we observed that CPZ retains its full inhibitory effect on the fraction of enzyme (5-6% of total) that is solubilized from erythrocytes upon treatment with phosphatidylinositol-specific phospholipase C (PI-PLC) from Bacillus thuringiensis; furthermore, Triton X-100 is able to reverse the CPZ effect also in the case of PI-PLC-solubilized enzyme . These results demonstrate unequivocally that CPZ inhibits human erythrocyte AChE through direct molecular interaction . The inhibition kinetics displayed by CPZ on human erythrocyte AChE are dependent on drug concentration: evidence is provided that this phenomenon may be related to formation of CPZ micellar aggregates.

Lett Appl Microbiol, 1991 Sep, 13(3), 171 - 4
The linear PCR reaction: a simple and robust method for sequencing amplified rRNA genes; Embley TM; The linear polymerase chain reaction was used to sequence amplified RNA genes from strains of Bacillus, Thermus and Legionella . The technique described is simple and reproducible and it works well with double standard product which has been PEG precipitated directly from PCR reactions.

Agric Biol Chem, 1991 Sep, 55(9), 2251 - 8
Purification and properties of a novel surface-active agent- and alkaline-resistant protease from Bacillus sp . Y; Shimogaki H et al.; In the course of a search for an alkaline stable protease for industrial use, an alkaline protease (protease BYA) was isolated from an alkalophilic Bacillus sp . Y, and its properties were characterized . Its optimum pH was pH 10.0-12.5, when casein was used as a substrate . In addition to the stability of protease BYA at pH 6.5-13.0, it was also very stable towards various surface-active agents, such as sodium dodecyl sulfate and sodium linear alkylbenzene sulfonate . Protease BYA was most active at 70 degrees C . The isoelectric point (pI) of protease BYA was about 10.1 . Protease BYA was characterized as a serine protease because of its sensitivity to phenylmethanesulfonyl fluoride and diisopropyl fluorophosphate . The protease seems to be related to proteases of the subtilisin family, such as subtilisin BPN', subtilisin Carlsberg, and No . 221 protease.

Arch Biochem Biophys, 1991 Aug 15, 289(1), 167 - 79
Physical studies on membrane lipids of Bacillus stearothermophilus temperature and calcium effects; Jurado AS et al.; Bacillus stearothermophilus was grown at the optimal temperature range (center, 65 degrees C), below it (48 and 55 degrees C), and above it (68 degrees C), in a complex medium with or without 2.5 mM Ca2+ . The Ca(2+)-supplement improves growth at sub- and supraoptimal temperatures and extends it to higher temperatures (Jurado et al . (1987) J . Gen . Microbiol . 133, 507-513) . The phospholipid composition of cultures obtained in the different growth conditions was studied . Phosphatidylethanolamine was always the major phospholipid (40 to 50% of the total phospholipid) . Diphosphatidylglycerol, phosphatidylglycerol, a phosphoglycolipid (pgl) and two minor phospholipids (not identified) were also found in the polar lipid extract . The pgl shows a threefold concentration increase as the growth temperature raises from 48 to 68 degrees C . The thermotropic behavior of membrane lipids was studied by differential scanning calorimetry (DSC) and by means of two fluorescent probes of fluidity, 1,6-diphenyl-1,3,5-hexatriene (DPH) and 1,3-di(2-pyrenyl)propane (2Py(3)2Py) . The results reveal similar features and clearly show a shift of the temperature range of the phase transition to higher values and an increased structural order of the bilayer, as the growth temperature rises from 55 to 68 degrees C, but an opposite effect was observed from 48 to 55 degrees C . Although the Ca(2+)-supplement to the growth medium has no detectable effect, the addition of Ca2+ to the buffer of liposomes (Ca(2+)-liposomes) has a significant ordering effect at all growth temperatures . These liposomes show a shift of the transition range to higher temperatures and the fluorescent parameters (DPH polarization and intramolecular excimerization of the 2Py(3)2Py) detected an order increase of the probes environment, along and above the main phase transition . Spectra of 31P-NMR and polarized light microscopy clearly show that the lipid extracts exhibit, in all the conditions, typical lamellar phase geometry . We concluded that B . stearothermophilus controls the membrane lipid composition to compensate for the destabilizing effect of high temperatures on the membrane organization or to provide an appropriate packing of phospholipid molecules in a stable bilayer . At high temperatures, Ca(2+)-stimulatory effect on growth is presumably due to a direct Ca2+ interaction with the membrane phospholipids, inducing an increased structural order on the bilayer . The increase of the phase transition temperature in the total lipid extracts as compared with the respective polar lipid fractions probably indicates a stabilizing effect of neutral lipids on membrane bilayers.

Carbohydr Res, 1991 Aug 12, 215(1), 127 - 36
Characterization of five isomers of branched cyclomaltoheptaose (beta CD) having degree of polymerization (d.p.) = 9: Reinvestigation of three positional isomers of diglucosyl-beta CD; Koizumi K et al.; It has been confirmed by methylation analyses and chemical syntheses that three isomers of branched cyclomaltoheptaose (beta CD) isolated from the mother liquors of a large-scale preparation of beta CD with Bacillus ohbensis cyclomaltodextrin glucanotransferase are 6(1),6(4)-di-O-(alpha-D-glucopyranosyl)-cyclomaltoheptaose (1), 6(1),6(3)-di-O-(alpha-D-glucopyranosyl)-cyclomaltoheptaose (2), and 6-O-(alpha-isomaltosyl)-cyclomaltoheptaose (4) instead of 6(1),6(2)-di-O-(alpha-D-glucopyranosyl)-cyclomaltoheptaose (3), which was erroneously characterized in an earlier paper . Compound 3 has been newly isolated from a glucosyl-beta CD mixture prepared by hydrolysis with glucoamylase of a maltosyl-beta CD mixture, synthesized from maltose and beta CD through the reverse action of pullulanase . Chromatographic behavior and spectral data (13C-n.m.r . and f.a.b.-m.s.) of these isomers of branched beta CD (1-4), as well as those of another isomer prepared by the reverse action of hydrolytic enzymes, 6-O-(alpha-maltosyl)-cyclomaltoheptaose (5), were compared.

J Biol Chem, 1991 Aug 5, 266(22), 14310 - 6
Stopped-flow and rapid-scan studies of the redox behavior of cytochrome aco from facultative alkalophilic Bacillus; Orii Y et al.; Cytochrome aco purified from an alkalophilic bacterium grown at pH 10 contains hemes a, b, and c as prosthetic groups, and their redox behavior was examined by using stopped-flow and rapid-scan techniques . Under anaerobic conditions the reduction of both heme a and c moieties with dithionite proceeded exponentially but with different rates, usually the former being reduced about 4 times faster than the latter . The reduction of protoheme was much slower, and a time-difference spectrum for this species was of a high spin type with absorption peaks at 433, 557, and 609 nm . Only the protoheme combined with CO, fulfilling the criteria for cytochrome o . Potentiometric titrations determined a midpoint potential of c heme to be 95 mV at pH 7.0 and 25 degrees C and suggested the presence of two forms of a heme with midpoint potentials of 250 and 323 mV . Cytochrome aco utilizes ascorbate plus N,N,N',N'-tetramethyl-p-phenylenediamine (TMPD) to reduce oxygen relatively rapidly without added cytochrome c (Qureshi, M . H., Yumoto, I., Fujiwara, T., Fukumori, Y., Yamanaka, T . (1990) J . Biochem . 107, 480-485) . During the steady state, however, heme a stayed almost fully reduced in contrast to a partial reduction of heme c . Even after exhaustion of the dissolved oxygen the extent of reduction of heme c was 60-70% that attained by the dithionite reduction . When ascorbate plus TMPD-reduced cytochrome aco was exposed to oxygen the reduced heme c was oxidized rapidly whereas the oxidation of reduced a heme was negligibly slow . The full reduction of heme a during the steady state and its extremely slow oxidation rendered participation of heme a in the oxidase reaction less likely . A novel peak appearing transiently around 567 nm during the reaction was tentatively ascribed to an intermediate form of protoheme, or o heme, which was thus supposed to react directly with molecular oxygen . These results suggest strongly that the main electron transfer pathway would be c----o----oxygen . A possible role of a in regulating the electron flow through the main pathway and its functional relationship to a heme in the aa3-type cytochrome oxidase were discussed.

Semin Vet Med Surg (Small Anim), 1991 Aug, 6(3), 199 - 202
Cat scratch disease: no longer a diagnostic dilemma; Margileth AM; Cat scratch disease is a relatively common cause of localized lymphadenopathy with about 80% of cases occurring in children . This self-limited infection is caused by a small pleomorphic gram-negative bacillus that has been identified in ocular granulomas, skin and lymph node specimens . Unusual manifestations of the disease, such as the oculoglandular disease of Parinaud, encephalopathy, or severe systemic disease, occur in about 12% of patients . Management consists of symptomatic treatment, occasionally aspiration of a node or selected antibiotic therapy in moderate to severely ill patients . Persistence of adenopathy for several months in a generally healthy patient with gradual, spontaneous resolution of the enlarged node is the natural course.

J Appl Bacteriol, 1991 Aug, 71(2), 130 - 3
Bacteria in food packaging paper and board; Vaisanen OM et al.; The bacteria of food packaging paper and board were studied . Most of the aerobic strains were spore-formers; members of the genus Bacillus with B . cereus group (B . cereus, B . mycoides, B . thuringiensis), B . polymyxa group (B . polymyxa, B . circulans, B . macerans, B . pabuli), B . brevis and B . licheniformis predominated . The main source of spore-forming bacteria in paper and board was the broke (rejected paper or board, which is repulped and recycled into the process) . Gram-negative bacteria were rare in paper and board in spite of their abundance in the stock . A strain of B . pumilus forming clumping, hairy spores may be of significance in aseptic packaging.

Infect Immun, 1991 Aug, 59(8), 2567 - 72
Effect of mycobacteria on sensitivity to the cytotoxic effects of tumor necrosis factor; Filley EA et al.; Unlike Mycobacterium leprae, Mycobacterium tuberculosis is not found inside cells other than macrophages and polymorphonuclear cells in vivo, yet previous work has revealed that in vitro it readily enters all cell lines tested . Moreover, these cells are not killed by the intracellular mycobacteria . We report here that when fibroblasts take up live (but not killed) M . tuberculosis H37Rv, they develop greatly increased sensitivity to the toxic effects of tumor necrosis factor (TNF) whether the cell line is inherently sensitive to TNF or not . Ultrasonically disrupted M . tuberculosis also has this property . The increased sensitivity is seen in the absence of metabolic inhibitors, although addition of emetine, an inhibitor of protein synthesis, causes the effect to manifest itself earlier and at a lower concentration of TNF . In contrast, infection with Mycobacterium bovis bacillus Calmette-Guerin induces little or no increased sensitivity to TNF, whereas Mycobacterium avium and M . tuberculosis H37Ra have intermediate sensitivities . We discuss the possibility that virulent tuberculosis strains produce a factor which distorts the normal protective function of TNF, rendering it toxic to host tissues and leading to the classical immunopathology of tuberculous lesions.

J Egypt Soc Parasitol, 1991 Aug, 21(2), 575 - 83
Effect of nutritive elements on the extracellular protein of different Bacillus strains, toxic to mosquito larvae; Rady MH et al.; The effect of 5 nutritional elements (glucose, peptone, yeast extract, beef extract and lactic acid) on the growth of the 4 tested Bacillus strains (Bacillus thuringiensis Bactimos 78, B . thuringiensis IPS 82, B . sphaericus Solvay 85 and B . sphaericus Rb 80) as well as on the concentration of their extracellular proteins were studied . The nutritive elements were individually tested either dissolved in distilled water or in a buffer base . Results indicated that the bacterial growth was higher when the nutritive elements were dissolved in a buffer base than in dist . water . The concentration of the extracellular proteins were always higher in the absence of the inorganic minerals, although the toxicity of these extracellular proteins against 3rd instar Culex pipiens larvae increased in the presence of these inorganic elements.

J Bacteriol, 1991 Aug, 173(15), 4889 - 92
Temperature-induced protein synthesis in Bacillus stearothermophilus NUB36; Wu L et al.; Cultures of Bacillus stearothermophilus subjected to a temperature shift-up or shift-down of 15 degrees C within the normal temperature range of growth (45 to 65 degrees C) enter a transient adaptation period before exponential growth at the new temperature . The de novo synthesis of some proteins coincides with the adaptation period.

J Bacteriol, 1991 Aug, 173(15), 4611 - 7
The Aeromonas hydrophila cphA gene: molecular heterogeneity among class B metallo-beta-lactamases; Massidda O et al.; An Aeromonas hydrophila gene, named cphA, coding for a carbapenem-hydrolyzing metallo-beta-lactamase, was cloned in Escherichia coli by screening an Aeromonas genomic library for clones able to grow on imipenem-containing medium . From sequencing data, the cloned cphA gene appeared able to code for a polypeptide of 254 amino acids whose sequence includes a potential N-terminal leader sequence for targeting the protein to the periplasmic space . These data were in agreement with the molecular mass of the original Aeromonas enzyme and of the recombinant enzyme produced in E . coli, evaluated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis of crude beta-lactamase preparations followed by renaturation treatment for proteins separated in the gel and localization of protein bands showing carbapenem-hydrolyzing beta-lactamase activity by a modified iodometric technique . The deduced amino acid sequence of the CphA enzyme showed regions of partial homology with both the beta-lactamase II of Bacillus cereus and the CfiA beta-lactamase of Bacteroides fragilis . Sequence homologies were more pronounced in the regions encompassing the amino acid residues known in the enzyme of B . cereus to function as ligand-binding residues for the metal cofactor . The CphA enzyme, however, appeared to share a lower degree of similarity with the two other enzymes, which, in turn, seemed more closely related to each other . These results, therefore, suggest the existence of at least two molecular subclasses within molecular class B metallo-beta-lactamases.

Zhongguo Yi Xue Ke Xue Yuan Xue Bao, 1991 Aug, 13(4), 261 - 6
{Biological assay, reliability test, error analysis and pooled calculation for assessing leucomycin yield}; Tan S; A biological assay for determining leucomycin yield was established by using a curillinear protocol (Sarcina lutea) and (2.2) dosage or (3.3) dosage protocol (Bacillus subtillis 63501) . At the same time, according to the principles of biostatistics and the methods of variance analysis, reliability tests, error analysis and pooled calculations were carried out, so as to hold the confidence limit below the internationally accepted level of 5%.

Chem Pharm Bull (Tokyo), 1991 Aug, 39(8), 2063 - 7
Action of phosphatidylinositol-specific phospholipase C from Bacillus thuringiensis is significantly influenced by coexisting lipids in substrate-detergent micelles; Kume T et al.; The effects of phosphatidylcholine (PC), phosphatidylethanolamine (PE), sphingomyelin (SM), and cholesterol on the activity of phosphatidylinositol-specific phospholipase C (PI-PLC) from Bacillus thuringiensis were studied in detail in phosphatidylinositol (PI)/detergent mixed micelles . By addition of PC, the enzymatic hydrolysis of PI was significantly stimulated in PI/Triton X-100 as well as PI/sodium deoxycholate (SDC) mixed micelles . SM stimulated enzyme activity toward PI/Triton X-100 micelles at a lower molar ratio of SM to PI, but was rather inhibitory at a ratio higher than 2.0 . The enzyme activity became significantly lower with an increase of PE or cholesterol in PI/Triton X-100 micelles . Actually, both PE and cholesterol were intensively inhibitory when added at a higher molar ratio to PI in Triton X-100-containing micelles . In the system of PI/SDC mixed micelles, not only PC but also SM, PE and cholesterol enhanced the enzymatic hydrolysis of PI . The difference between PI/Triton X-100 and PI/SDC micelles regarding the effects of these lipids on PI-PLC action, must be dependent on the physical state of micelles formed by these detergents and lipids.

J Comp Pathol, 1991 Aug, 105(2), 167 - 73
Tyzzer's disease (Bacillus piliformis) in Australian marsupials; Canfield PJ et al.; Tyzzer's disease (Bacillus piliformis infection) was diagnosed in nine marsupials (six possums, a koala, a wombat and a dasyurid) . All but two of the possums were captive . Five of the seven marsupials, for which ages were recorded, were juvenile . Affected marsupials were either found dead or were showing non-specific illness for up to two days before death . Affected livers and hearts showed gross haemorrhage and scattered areas of white discolouration . Microscopically, areas of coagulative necrosis were often associated with neutrophilic infiltrates . Intracellular, slender, faintly basophilic rods were occasionally detected in HE-stained sections . Rods were shown by silver staining techniques to be associated with the lesions in all affected animals . These rods showed a beaded appearance when stained with methenamine silver and were consistently in packets within cells, on the peripheries of lesions and scattered sparsely and irregularly within the central necrotic regions.

Appl Environ Microbiol, 1991 Aug, 57(8), 2277 - 82
Toxicity of Bacillus thuringiensis to laboratory populations of the olive fruit fly (Dacus oleae); Karamanlidou G et al.; A survey of Bacillus thuringiensis recovered from the environments of olive groves in Greece was carried out . Of 80 soil samples, 24 were found to contain B . thuringiensis with parasporal crystal inclusions; these were tested for toxicity against the olive fruit fly (Dacus oleae) . Mortality levels of larvae caused by the different isolates varied from 7 to 87% . Higher levels of mortality were observed if a mixture of relatively pure crystals and spores was used compared with the mortality resulting from either fraction alone . We were able to show that the toxicity of the most active isolate is likely to be specific for D . oleae.

J Biochem (Tokyo), 1991 Aug, 110(2), 279 - 83
Thermostable alanine racemase of Bacillus stearothermophilus: subunit dissociation and unfolding; Toyama H et al.; The guanidine hydrochloride-induced subunit dissociation and unfolding of thermostable alanine racemase from Bacillus stearothermophilus have been studied by circular dichroism, fluorescence and absorption spectroscopies, and gel filtration . The overall process was found to be reversible: more than 75% of the original activity was recovered upon reduction of the denaturant concentration . In the range of 0.6 to 1.5 M guanidine hydrochloride, the dimeric enzyme was dissociated into a monomeric form, which was catalytically inactive . The monomeric enzyme appeared to bind the cofactor pyridoxal phosphate by a non-covalent linkage, although the native dimeric enzyme binds the cofactor through an aldimine Schiff base linkage . The monomer was mostly unfolded, with the transition occurring in the range of 1.8 to 2.2 M guanidine hydrochloride.

J Immunol, 1991 Aug 1, 147(3), 1023 - 9
Identification of T cell stimulatory peptides from the 38-kDa protein of Mycobacterium tuberculosis; Vordemeier HM et al.; T cell specificity to individual antigenic epitopes could determine the distinction between protective and pathogenic host reactions in tuberculous infections . Therefore, T cell stimulatory epitopes of the Mycobacterium tuberculosis 38-kDa lipoprotein, of known structure and specificity and of prominent immunogenicity, have been examined . To identify potential T cell epitopes, eight peptides, seven of which were predicted to form amphiphatic helices, were used for immunization of various inbred mice and for elicitation of in vitro T cell proliferative responses . Three different response patterns were observed . 1) Lymph node cells from mice immunized with peptide, recombinant 38-kDa Ag, killed M . tuberculosis strain H37Ra, or live Mycobacterium bovis bacillus Calmette Guerin infection responded to peptide 38.G (residues 350 to 369) . Responses were observed in mice of H-2b, H-2d, and H-2k haplotypes . 2) Peptide 38.C (residues 201 to 220) induced proliferation of lymph node cells from 38-kDa protein-, but not from peptide-immunized mice . 3) Peptide 38.F (residues 285 to 304) only elicited a response of the homologous peptide-primed cells . Analysis of CD4+ T cell lines confirmed the distinct specificities and stimulatory features of peptides 38.F and 38.G . The described attributes of peptide 38.C and 38.G could be of potential interest for diagnostic evaluation in tuberculous infections.

J Bacteriol, 1991 Aug, 173(15), 4725 - 35
Transcriptional attenuation control of ermK, a macrolide-lincosamide-streptogramin B resistance determinant from Bacillus licheniformis; Kwak JH et al.; ermK instructs bacteria to synthesize an erythromycin-inducible 23S rRNA methylase that confers resistance to the macrolide, lincosamide, and streptogramin B antibiotics . Expression of ermK is regulated by transcriptional attenuation, in contrast to other inducible erm genes, previously described, which are regulated translationally . The ermK mRNA leader sequence has a total length of 357 nucleotides and encodes a 14-amino-acid leader peptide together with its ribosome binding site . Additionally, the mRNA leader sequence can fold in either of two mutually exclusive conformations, one of which is postulated to form in the absence of induction and to contain two rho factor-independent terminators . Truncated transcription products ca . 210 and 333 nucleotides long were synthesized in the absence of induction, both in vivo and in vitro, as predicted by the transcriptional attenuation model; run-off transcription in vitro with rITP favored the synthesis of the full-length run-off transcript over that of the 210- and 333-nucleotide truncated products . Northern (RNA) blot analysis of transcripts synthesized in vivo in the absence of erythromycin indicated that transcription terminated at either of the two inverted complementary repeat sequences in the leader that were postulated to serve as rho factor-independent terminators; moreover, no full-length transcripts were detectable in the uninduced samples . In contrast, full-length (ca . 1,200-nucleotide) transcripts were only detected in RNA samples synthesized in vivo in the presence of erythromycin . Full-length transcripts formed in the absence of induction from transcriptional readthrough past the two proposed transcription terminators would fold in a way that would sequester the ribosome binding site together with the first two codons of the ErmK methylase, reducing its efficiency in translation . This feature could therefore provide additional control of expression in the absence of induction; however, such regulation, if operative, would act only secondarily, both in time and place, relative to transcriptional control . Analysis by reverse transcriptase mapping of in vivo transcripts from two primers that bracket the transcription terminator responsible for the 210-nucleotide truncated fragment supports the transcriptional attenuation model proposed and suggests further that the synthesis of the ermK message is initiated constitutively upstream of the proposed terminator but completed inductively downstream of this site.

Bull Tokyo Dent Coll, 1991 Aug, 32(3), 99 - 110
Inorganic components and the fine structures of marginal and deep subgingival calculus attached to human teeth; Kodaka T et al.; Inorganic components and the fine structures of marginal ledge-type and deep subgingival spiny deposits in human old dental calculus were investigated by scanning electron microscopy and energy dispersive electron-probe microanalysis . The ledge-type deposits consisted of the extra- and intracellular calcifying deposits, large plate-shaped crystals, and bacillus-shaped deposits composed of hexahedrally based crystals . The spiny deposits were mainly formed by aggregations of the bacillus-shaped deposits . In the outer and middle layers of the spiny deposits, the Ca, P, and Mg concentrations were all significantly higher than those of the ledge-type deposits . A consideration of the crystal shapes and Ca, P, and Mg molar ratios reveals the following differences . Calculus components of the ledge-type deposits contained crystal types quite similar to sandy grain-shaped hydroxyapatite (HAP), plate-shaped octacalcium phosphate (OCP), and hexahedral Mg-containing whitlockite (WHT) . On the other hand, in the spiny deposits, the Mg-containing WHT type comprised a large proportion of the calculus; the HAP type was found in the outermost and inner layers; and no OCP type was detected.

J Biochem (Tokyo), 1991 Aug, 110(2), 267 - 73
Purification and characterization of two membrane-bound c-type cytochromes from a facultative alkalophilic Bacillus; Yumoto I et al.; The membrane fraction of the facultative alkalophilic bacterium, Bacillus YN-2000, was found to contain considerably larger amounts of two c-type cytochromes, cytochromes c-553 and c-552, when the bacterium was grown at pH 10 than when it was grown at lower pHs (pH 7-9) . In particular, cytochrome c-553 was present in a much higher amount in the cells grown at pH 10 than in those grown at pH 8 . Cytochromes c-553 and c-552, which are membrane-bound proteins, were purified to electrophoretically homogeneous states from Bacillus YN-2000 . Cytochrome c-553 showed absorption peaks at 553, 524, and 417 nm in the reduced form, and a peak at 411 nm in the oxidized form . Its molecular weight was estimated to be 10,500 from the results of SDS-polyacrylamide gel electrophoresis . However, its molecular weight was estimated to be 127,000 by gel filtration . Therefore, it seemed to occur as an oligomer in solution . The isoelectric point of cytochrome c-553 was determined to be 3.9 . Its midpoint redox potential was found to be +87 mV in the pH region from 6 to 8 . Cytochrome c-553 reacted with cytochrome c oxidase of the bacterium and the reaction was greatly accelerated in the presence of poly-L-lysine . Cytochrome c-552 showed absorption peaks at 552, 521, and 416 nm in the reduced form, and a peak at 408 nm in the oxidized form . The cytochrome molecule seemed to be composed of six different subunits, with molecular masses of 40, 32, 19, 17, 14, and 12 kDa, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Eur J Biochem, 1991 Aug 1, 199(3), 761 - 8
Two different aa3-type cytochromes can be purified from the bacterium Bacillus cereus; Garcia-Horsman JA et al.; Two aa3-type cytochromes were purified from membranes of sporulating Bacillus cereus . One of them, an aa3 complex, was found to be composed of two subunits (51 and 31 kDa), two a hemes and three copper atoms, thus being similar to the cytochrome aa3 previously purified from vegetative B . cereus {Garcia-Horsman, J . A., Barquera, B., Gonzalez-Halphen, D . & Escamilla, J . E . (1991) Mol . Microbiol . 5, 197-205} . The second isoform, a caa3 complex, was expressed in sporulating cells only, and was found to be composed of two subunits (51 and 37 kDa) . The 37-kDa subunit (subunit II) is a heme-c-containing polypeptide as shown by its peroxidase activity in SDS/PAGE gels and by its spectral features . Both subunits of the caa3 complex immunologically cross-reacted with antiserum raised against B . cereus cytochrome aa3, suggesting homology between the two enzymes . Also, the heme-c-containing subunit of the caa3 complex was reactive with anti-(bovine cytochrome c) antiserum, but not with anti-(bovine cytochrome c1) antiserum . In addition to one heme c and two hemes a, the caa3 complex contained three copper atoms . Kinetic comparison of aa3 and caa3 complexes revealed that the latter is slightly more active (k = 150 s-1) and has a lower affinity to yeast cytochrome c (Km = 76 microM) and to oxygen (Km = 2 microM) as compared with cytochrome aa3 (100 s-1, 10 microM, and 5 microM, respectively).

Am J Clin Oncol, 1991 Aug, 14(4), 291 - 7
Long-term evaluation of intrapleural bacillus Calmette-Guerin with or without adjuvant chemotherapy in completely resected stages II and III non-small-cell lung cancer; Macchiarini P et al.; Between January 1979 and December 1980, 52 patients with completely resected stages II and III non-small-cell lung cancer (NSCLC) were randomly assigned to receive either adjuvant chemotherapy (cyclophosphamide, doxorubicin, and vincristine--CAV) plus intrapleural bacillus Calmette-Guerin (BCG) (n = 26) or adjuvant CAV alone (n = 26) . Careful intraoperative staging was performed in all patients, and stratification for histology (squamous versus nonsquamous) and stage (II or III) ensured a balanced randomization for these factors . With a median follow-up time of 111 months, overall 10-year and median survival were 21% and 20 months (range 2-127 + months), respectively . Thirty-four (95%) patients relapsed in extrathoracic sites, and five (5%) developed loco-regional recurrence; their overall median disease-free interval (DFI) was 10 months (range 1-73 months) . There was a 9% and 2.5 month difference in survival (p = .76) and disease-free interval (p = .67), respectively, favoring the BCG arm . There were no significant differences in the sites and patterns of first recurrence comparing the two treatment arms . In conclusion, there is no suggestion of a significant therapeutic advantage from intrapleural BCG in conjunction with adjuvant chemotherapy for completely resected stages II and III NSCLC.

J Bacteriol, 1991 Aug, 173(16), 5010 - 6
Evidence for multiple terminal oxidases, including cytochrome d, in facultatively alkaliphilic Bacillus firmus OF4; Hicks DB et al.; The terminal oxidase content of Bacillus firmus OF4, a facultative alkaliphile that grows well over the pH range of 7.5 to 10.5, was studied by difference spectroscopy . Evidence was found for three terminal oxidases under different growth conditions . The growth pH and the stage of growth profoundly affected the expression of one of the oxidases, cytochrome d . The other two oxidases, cytochrome caa3 and cytochrome o, were expressed under all growth conditions tested, although the levels of both, especially cytochrome caa3, were higher at more alkaline pH (P.G . Quirk, A.A . Guffanti, R.J . Plass, S . Clejan, and T.A . Krulwich, Biochim . Biophys . Acta, in press) . These latter oxidases were identified in everted membrane vesicles by reduced-versus-oxidized difference spectra (absorption maximum at 600 nm for cytochrome caa3) and CO-reduced-versus-reduced difference spectra (absorption maxima at 574 and 414 nm for cytochrome o) . All three terminal oxidases were solubilized from everted membranes and partially purified . The difference spectra of the solubilized, partially purified cytochrome caa3 and cytochrome o complexes were consistent with these assignments . Cytochrome d, which has not been identified in a Bacillus species before, was tentatively assigned on the basis of its absorption maxima at 622 and 630 nm in reduced-versus-oxidized and CO-reduced-versus-reduced difference spectra, respectively, resembling the maxima exhibited by the complex found in Escherichia coli . The B . firmus OF4 cytochrome d was reducible by NADH but not by ascorbate-N,N,N',N'-tetramethyl-p-phenylenediamine in everted membrane vesicles . Cytochrome d was expressed under two conditions: in cells growing exponentially at pH 7.5 (but not at pH 10.5) and in cells stationary phase at either pH 7.5 or 10.5 . Protein immunoblots with antibodies against subunit I of the E . coli cytochrome d complex reacted only with membrane vesicles that contained spectrally identifiable cytochrome d . Additional evidence that this B . firmus OF4 cytochrome is related to the E . coli complex was obtained with a solubilized, partially purified fraction of cytochrome d that also reacted with antibodies against the subunits of the E . coli cytochrome d.

Enzyme Microb Technol, 1991 Aug, 13(8), 661 - 4
Design and use of synthetic oligonucleotide probes in the cloning of delta-endotoxin genes from Bacillus thuringiensis; Starkey MP et al.; A detailed protocol is described for the design and use of synthetic oligonucleotide probes for screening DNA libraries from Bacillus thuringiensis var . kurstaki (strain HD191) for copies of the gene (tox) encoding the insecticidal delta-endotoxin . Two homologous tox genes were identified in this organism; one of these was located on a 75-kb plasmid and the other on a second large plasmid or the bacterial chromosome . A tox gene was isolated as a 6.5-kb HindIII fragment of B . thuringiensis plasmid DNA.

Appl Microbiol Biotechnol, 1991 Aug, 35(5), 600 - 5
Cloning and sequencing of a cyclodextrin glucanotransferase gene from Bacillus ohbensis and its expression in Escherichia coli; Sin K et al.; A cyclodextrin glucanotransferase (CGTase) gene of Bacillus ohbensis was cloned in Escherichia coli and the nucleotide sequence was determined . A single open reading frame (2112 bp) with a TTG codon as an initiator was identified that encodes a typical signal peptide of 29 amino acids followed by the mature enzyme (675 amino acids), of which the partial amino acid sequences of the N-terminal region and some lysyl-endopeptidase fragments were determined by Edman degradation . The CGTase gene was expressed in E . coli under control of the lac promoter only when the upstream region containing a long inverted repeat structure (located at -108 to -67 bp from the initiation codon) was deleted . Substitution of an ATG codon for the initiation TTG triplet doubled the expression of the CGTase gene in E . coli . Enzyme preparations purified from the culture supernatant of B . ohbensis and from the periplasmic fraction of the E . coli transformant exhibited the same molecular weight (Mr) and enzymatic properties as follows: Mr, 80,000; optimum pH for activity, 5.0 (and a suboptimum at 10.0); stability between pH 6.5 and 10.0; optimum temperature for activity, 55 degrees C; and stability below 45 degrees C . The yields of the products from starch as the substrate were 25% for beta- and 5% for gamma-cyclodextrin.

J Biotechnol, 1991 Aug, 20(1), 29 - 49
Biotin formation by recombinant strains of Escherichia coli: influence of the host physiology; Sabatie J et al.; Strains of Escherichia coli were transformed with different plasmids bearing the gene clusters bioXWF and bioDAYB isolated from the Gram positive bacterium Bacillus sphaericus . These genes encode for the enzymes involved in the metabolic pathway which synthesizes biotin from the precursor pimelic acid . Transformed E . coli strains were grown in bioreactors to reach a biomass of 18 g l-1 cell dry weight in 1 litre batch culture with substrate feeding and approximately 50 g l-1 in 10 l fed batch culture . Improved yields of total vitamers and biotin formed in these processes were achieved after a comparative analysis of different culture conditions, medium compositions, host strains and expression systems . Production of 27 mg l-1 of biotin and 200 mg l-1 of vitamers was achieved in 1 litre batch culture . Using a 10-1 fed batch process, biotin and vitamer concentrations reached maximum values of 45 mg l-1 and 350 mg l-1, respectively.

FEBS Lett, 1991 Jul 29, 286(1-2), 204 - 8
Cloning, overexpression, purification and crystallisation of ribosomal protein L9 from Bacillus stearothermophilus; Vorgias CE et al.; The cloning, sequencing and overexpression of the gene coding for Bacillus stearothermophilus ribosomal protein L9 is described . The sequence corresponds directly to that presented for the protein itself by classical methods, differing at only a few amino acid positions . The purification and crystallisation of the corresponding L9 protein is presented . The crystals are isomorphous to those described for L9 obtained by conventional methods.

J Biol Chem, 1991 Jul 25, 266(21), 13634 - 9
Thermostable alanine racemase of Bacillus stearothermophilus . Construction and expression of active fragmentary enzyme; Toyama H et al.; Limited proteolysis studies on alanine racemase suggested that the enzyme subunit is composed of two domains (Galakatos, N . G., and Walsh, C . T . (1987) Biochemistry 26, 8475-8480) . We have constructed a mutant gene that tandemly encodes the two polypeptides of the Bacillus stearothermophilus enzyme subunit cleaved at the position corresponding to the predicted hinge region . The mutant gene product purified was shown to be composed of two sets of the two polypeptide fragments and was immunologically identical to the wild-type enzyme . The mutant enzyme, i.e . the fragmentary alanine racemase, was active in both directions of the racemization of alanine . The maximum velocity (Vmax) was about half that of the wild-type enzyme, and the Km value was about double . Absorption and circular dichroism spectra of the fragmentary enzyme were similar to those of the wild-type enzyme . An attempt was made to separately express in Escherichia coli a single polypeptide corresponding to each domain, but no protein reactive with the antibody against the wild-type alanine racemase was produced . Therefore, it is suggested that the two polypeptide fragments can fold into an active structure only when they are co-translated and that they correspond to structural folding units in the parental polypeptide chain.

Proc R Soc Lond B Biol Sci, 1991 Jul 22, 245(1312), 31 - 5
N-acetyl galactosamine is part of the receptor in insect gut epithelia that recognizes an insecticidal protein from Bacillus thuringiensis; Knowles BH et al.; Proteins synthesized by the bacterium Bacillus thuringiensis are potent insecticides . When ingested by susceptible larvae they rapidly lyse epithelial cells lining the midgut . In vitro the toxins lyse certain insect cell lines and show saturable, high-affinity binding to brush-border membrane vesicles (BBMVs) prepared from insect midguts . We observed that the sugar N-acetyl galactosamine (GalNAc) specifically decreased the cytolytic activity of a CryIA(c) toxin towards Choristoneura fumiferana CF1 cells, completely abolished toxin binding to Manduca sexia BBMVs, partially inhibited binding to Heliothis virescens BBMVs and had no apparent effect on binding to Pieris brassicae BBMVs . In ligand blotting experiments the toxin bound proteins of 120 kDa in M . sexta, 125 kDa in P . brassicae and numerous proteins in H . zea . Toxin binding to these proteins was specifically inhibited by GalNAc . The toxin binding proteins of M . sexta and H . zea also bound the lectin soybean agglutinin . Taken together these findings suggest that N-acetyl galactosamine might be a component of a CryIA(c) toxin receptor of CF1 cells and of at least two of the insects tested.

J Mol Biol, 1991 Jul 20, 220(2), 435 - 55
Beta-lactamase of Bacillus licheniformis 749/C . Refinement at 2 A resolution and analysis of hydration; Knox JR et al.; The crystallographic and molecular structure of the class A beta-lactamase (penicillinase) of Bacillus licheniformis 749/C has been refined with X-ray diffraction data to 2 A resolution . For the 27,330 data with F greater than or equal to 3 sigma(F), the R factor is 0.15; for all 30,090 data, R is 0.16 . The estimated co-ordinate error is 0.15 A . In the final model, the deviation of covalent bonds and angles from ideality is 0.012 A and 2.2 degrees, respectively . The model includes two molecules of 29,500 daltons each in the asymmetric unit of space group P2(1), 484 water molecules and two tetrahedral buffer anions . Overlay of the two protein molecules results in a root-mean-square difference of 0.17 A and 0.41 A for alpha-carbon atoms and for all atoms, respectively . Twenty-six water molecules fall within 0.25 A of matching water molecules associated with the second protein molecule . The reactive Ser70 is on a turn of 3(10) helix at the N terminus of a longer alpha-helix (72-83) . The penicillin-binding site near this helix contains at least seven water molecules . Upon penicillin entry, a water molecule in the oxyanion hole, hydrogen-bonded between the N terminus of helix (80-83) and beta-strand (230-238), would be displaced by the oxygen atom of the beta-lactam carbonyl group . An unexpelled molecule of water is proposed to be the catalytic water required for penicillin hydrolysis . The water is hydrogen-bonded to Glu166, a conserved residue in all beta-lactamases, and it lies 3 A from the alpha-face of a previously modeled penicillin . The position of the water-Glu166 pair is stabilized in the active site by a cis peptide bond at Pro167.

Cancer Res, 1991 Jul 15, 51(14), 3726 - 32
Modulation of fibronectin-mediated Bacillus Calmette-Guérin attachment to murine bladder mucosa by drugs influencing the coagulation pathways; Hudson MA et al.; Adjuvant intravesical Bacillus Calmette-Guerin (BCG) has proved to be an effective treatment for superficial bladder cancer . Intraluminal attachment of BCG organisms via binding to the extracellular matrix protein, fibronectin (FN), appears to be required for expression of the antitumor efficacy of BCG against a murine bladder tumor . Initial studies demonstrated that radiolabeled FN localized to the acutely injured urothelium but not to intact urothelium . These studies also demonstrated that exogenous administration of FN enhanced BCG attachment to the injured but not to the intact urothelium . Because FN has been shown to be an integral part of clot formation at sites of urothelial injury, drugs known to affect fibrin clot formation were tested for their effects on BCG attachment and antitumor efficacy in a murine bladder tumor model . A stabilizer of fibrin clot formation was shown to enhance both BCG attachment and antitumor efficacy in the same model . An increased number of BCG organisms were also retained in the lymph nodes and spleens of mice receiving fibrin clot stabilizers, suggesting indirectly that immunological mechanisms are involved in the antitumor efficacy of BCG . The data presented herein provide further support for the hypothesis that BCG attachment to the injured bladder is mediated by FN . Furthermore, modulation of BCG-FN attachment is demonstrated to be possible with drugs influencing the coagulation pathway . This attachment is shown to be required for the antitumor efficacy in a murine bladder tumor model, and thus modulation of BCG-FN attachment appears to have significant influence on the antitumor efficacy of BCG in the murine bladder tumor model.

Biochim Biophys Acta, 1991 Jul 12, 1078(3), 404 - 10
Isolation and characterization of thermostable chitinases from Bacillus licheniformis X-7u; Takayanagi T et al.; Four kinds of thermostable chitinase were isolated from the cell-free culture broth of Bacillus licheniformis X-7u by successive column chromatographies on Butyl-Toyopearl, Q-Sepharose, and Sephacryl S-200 . We named the enzymes chitinases I(89 kDa), II(76 kDa), III(66 kDa) and IV(59 kDa) . Chitinases II, III and IV possessed extremely high optimum temperatures (70-80 degrees C), showing remarkable heat stability . Chitinases II, III and IV produced (GlcNAc)2 and GlcNAc from colloidal chitin and chitinase I predominantly produced (GlcNAc)2 . The action pattern of chitinase I on PN-(GlcNAc)4 also showed a stronger propensity to cleave off the (GlcNAc)2 unit from the non-reducing end than the other three chitinases . Chitinases II, III and IV catalyzed a transglycosylation reaction that converted (GlcNAc)4 into (GlcNAc)6.

Nucleic Acids Res, 1991 Jul 11, 19(13), 3673 - 81
Methionyl-tRNA synthetase from Bacillus stearothermophilus: structural and functional identities with the Escherichia coli enzyme; Mechulam Y et al.; The metS gene encoding homodimeric methionyl-tRNA synthetase from Bacillus stearothermophilus has been cloned and a 2880 base pair sequence solved . Comparison of the deduced enzyme protomer sequence (Mr 74,355) with that of the E . coli methionyl-tRNA synthetase protomer (Mr 76,124) revealed a relatively low level (32%) of identities, although both enzymes have very similar biochemical properties (Kalogerakos, T., Dessen, P., Fayat, G . and Blanquet, S . (1980) Biochemistry 19, 3712-3723) . However, all the sequence patterns whose functional significance have been probed in the case of the E . coli enzyme are found in the thermostable enzyme sequence . In particular, a stretch of 16 amino acids corresponding to the CAU anticodon binding site in the E . coli synthetase structure is highly conserved in the metS sequence . The metS product could be expressed in E . coli and purified . It showed structure-function relationships identical to those of the enzyme extracted from B . stearothermophilus cells . In particular, the patterns of mild proteolysis were the same . Subtilisin converted the native dimer into a fully active monomeric species (62 kDa), while trypsin digestion yielded an inactive form because of an additional cleavage of the 62 kDa polypeptide into two subfragments capable however of remaining firmly associated . The subtilisin cleavage site was mapped on the enzyme polypeptide, and a gene encoding the active monomer was constructed and expressed in E . coli . Finally, trypsin attack was demonstrated to cleave a peptidic bond within the KMSKS sequence common to E . coli and B . stearothermophilus methionyl-tRNA synthetases . This sequence has been shown, in the case of the E . coli enzyme, to have an essential role for the catalysis of methionyl-adenylate formation.

Med Clin (Barc), 1991 Jul 6, 97(6), 211 - 4
{Mycobacterial infections: yield of bacillary microscopy in different clinical samples (1975-1988)}; Elcuaz Romano R et al.; BACKGROUND: The yield of microscopy examination as a quick diagnostic test in several pulmonary and nonpulmonary samples referred to the mycobacterial laboratory of a general hospital is reviewed . METHODS: During a 14-year period (1975-1988), 113,836 biological products were investigated . In 9,972 a positive culture for mycobacteria was obtained . For the microscopy examination the auramin technique was used; if positive, acid-alcohol resistance was confirmed by overstaining with the Ziehl-Neelsen technique . The culture was used as the reference method . RESULTS: Microscopic examination was positive in 34% of samples with a positive culture, being 39% for Mycobacterium tuberculosis and 10% for environmental mycobacteria . The overall specificity was 99%, the positive predictive value was 91% and the negative predictive value was 94% . In pleuropulmonary samples the sensitivity ranged from 48% in sputum and 2% in pleural biopsy, with specificity higher than 99% . In nonpulmonary samples, sensitivity, specificity and positive and negative predictive values varied with the type of sample . The false positive rate (positive microscopy with negative culture) was 0.3, and it was shown that 80% of these patients had received previous therapy . In organic fluids (pleural, peritoneal, cerebrospinal), the sensitivity was not greater than 13% . CONCLUSIONS: Sputum, bronchoaspirate and bronchoalveolar lavage were better for the diagnosis of tuberculosis than gastric aspirate . Approximately 1 in each positive microscopy examinations corresponded to environmental mycobacteria . In some nonpulmonary samples with high sensitivity the positive predictive value was low . 80% of the false positive results corresponded to previously treated patients.

J Bacteriol, 1991 Jul, 173(13), 4243 - 5
Repression of the cell wall protein gene operon in Bacillus brevis 47 by magnesium and calcium ions; Adachi T et al.; Transcription from P2, one of the major promoters of the cell wall protein gene operon of Bacillus brevis 47, was markedly enhanced at the early stationary phase of growth . MgCl2, when added at 1 to 5 mM to the medium, inhibited this enhancement of transcription as well as shedding of the cell wall protein layers from the cell surface . MgSO4 or CaCl2 showed an effect similar to that of MgCl2 . The possible coordination of the cell wall structure with regulation of the cell wall protein genes is discussed.

J Bacteriol, 1991 Jul, 173(13), 4013 - 20
Cloning and analysis of the nuclear gene for YmL33, a protein of the large subunit of the mitochondrial ribosome in Saccharomyces cerevisiae; Kang W et al.; The N-terminal amino acid sequence of a large subunit protein, termed YmL33, of the mitochondrial ribosome of the yeast Saccharomyces cerevisiae was determined . The data were obtained to synthesize two kinds of oligonucleotide primers, which were used in the polymerase chain reaction to amplify and clone the nuclear gene for this protein . By nucleotide sequencing, the cloned gene, MRP-L33, was found to encode a basic protein of 11 kDa with 98 amino acid residues . The protein encoded by this gene appears to have no leader sequence at its N terminus . The N-terminal two-thirds of the deduced amino acid sequence showed a significant degree of sequence similarity to ribosomal protein L30 of Escherichia coli and Bacillus stearothermophilus . In addition, the C-terminal one-third showed sequence similarity, though to a lesser extent, to a yeast cytoplasmic ribosomal protein termed L16 . By hybridization with the yeast chromosomes and their restriction enzyme fragments, the MRP-L33 gene was concluded to exist on chromosome XIII as a single-copy gene . Disruption of the gene by insertion of a HIS3-containing fragment showed that MRP-L33 was essential for mitochondrial function . The transcriptional level of MRP-L33 in strains with different mitochondrial genetic backgrounds was analyzed in the presence of glucose, galactose, or glycerol.

J Bacteriol, 1991 Jul, 173(13), 3966 - 76
Isolation and characterization of a novel insecticidal crystal protein gene from Bacillus thuringiensis subsp . aizawai; Chambers JA et al.; Bacillus thuringiensis subsp . aizawai EG6346, a novel grain dust isolate, was analyzed by Southern blot hybridization for its insecticidal crystal protein (ICP) gene profile . Strain EG6346 lacks previously characterized cryIA ICP genes yet does possess novel cryI-related gene sequences . A recombinant genomic plasmid library was constructed for strain EG6346 in Escherichia coli . One recombinant plasmid, pEG640, isolated from the library contained a novel ICP gene on a 5.7-kb Sau3A insert . The sequence of this gene, designated cryIF, was related to, but distinct from, the published sequences for other cryI genes . A second novel cryI-related sequence was also located on pEG640, approximately 500 bp downstream from cryIF . Introduction of cryIF into a Cry- B . thuringiensis recipient strain via electroporation enabled sufficient production of CryIF protein for quantitative bioassay analyses of insecticidal specificity . The CryIF crystal protein was selectively toxic to a subset of lepidopteran insects tested, including the larvae of Ostrinia nubilalis and Spodoptera exigua.

J Urol, 1991 Jul, 146(1), 32 - 5
A low dose bacillus Calmette-Guerin regimen in superficial bladder cancer therapy: is it effective?
Pagano F, Bassi P, Milani C, Meneghini A, Maruzzi D, Garbeglio A.
Bacillus Calmette-Guerin (BCG) intravesical therapy represents a major advance in the treatment of superficial transitional cell carcinoma of the bladder . To date, however, the optimal treatment schedule must be defined and the toxicity related to the treatment is significant . The preliminary results of a randomized ongoing study performed to evaluate the effectiveness and relative toxicity of a low dose (75 mg.) BCG regimen in the treatment of superficial bladder cancer therapy are reported . A total of 126 patients (70 for prophylaxis of recurrent stages Ta and T1 papillary tumors and 56 for treatment of carcinoma in situ or with microinfiltration of the subepithelial connective tissue) underwent a 6-week course of 75 mg . BCG (Pasteur vaccine) . An additional course was given in patients who failed to respond to the induction course . Maintenance therapy was administered in complete responders monthly for 1 year and then quarterly for 1 year . The prophylaxis group (transurethral resection plus BCG) was randomized versus transurethral resection alone (63 patients, control group) . A complete response in the prophylaxis, control and therapy groups was observed in 74, 17 and 57% of the patients, respectively, while 4, 17 and 12.5%, respectively, experienced tumor progression . The additional course of therapy increased the response rate . On the contrary, previous unsuccessful intravesical chemotherapy did not affect the response rate . In regard to toxicity, irritative disturbances (27%) and fever (17%) appeared to be significantly decreased compared with the rates reported in the literature . No major complications were experienced . In conclusion, a low dose (75 mg.) Pasteur strain BCG regimen was effective as prophylaxis against recurrent superficial papillary tumors and as treatment of carcinoma in situ or with microinfiltration of the subepithelial connective tissue . Toxicity related to the treatment appeared to be low.

J Immunol, 1991 Jul 1, 147(1), 312 - 9
Sequence and immunogenicity of the 70-kDa heat shock protein of Mycobacterium leprae; McKenzie KR et al.; The gene encoding the Mycobacterium leprae 70-kDa heat shock protein has been isolated from a cosmid library using a fragment of the clone JKL2 . Southern blot analysis of a positive clone identified a 4.4-kb fragment containing the entire coding region of the gene plus 2.4 kb upstream . Sequencing revealed the gene to encode a 621-amino acid protein, bearing 56% identity with the Escherichia coli dnaK gene product and 47% and 46% identity with the human and Caenorhabditis elegans hsp70, respectively . Comparison with the C-terminal 203 amino acids of the Mycobacterium tuberculosis 71-kDa Ag yielded 70% identity . Recombinant M . leprae p70 was produced in E . coli as a fusion protein (rp70f) with a portion of the schistosomal glutathione-S-transferase, using the expression vector, pGEX-2T . Cleavage with thrombin resulted in the release of a 70.0-kDa protein (rp70c) from the glutathione-S-transferase . Examination of the proteins by immunoblotting demonstrated that anti-M . leprae mAb, L7, and sera from lepromatous leprosy patients bound to both the cleaved and fusion proteins . We compared the T cell reactivity of the M . leprae recombinant proteins with that of mAb affinity-purified bacille Calmette-Guerin (BCG) 70-kDa Ag using proliferation assays . PBMC of BCG vaccinees responded to both M . leprae cleaved and fusion p70, though more subjects responded to the rp70c (18 of 20) than to rp70f (13 of 20) . Responses were generally higher to rp70c than to rp70f, however all responses to the M . leprae recombinant proteins were lower than to mAb affinity-purified BCG p70 . Thus, the M . leprae 70-kDa heat shock protein elicits T and B cell responses in subjects exposed to mycobacteria, despite its homology with the human hsp70.

Indian J Med Res, 1991 Jul, 93, 208 - 16
Some indices pertaining to the leprosy control programme in Tamil Nadu, based on data from a random sample of fourteen government control units; Nair NG et al.; From a random sample of 14 Government Leprosy Control Units in Tamil Nadu, information on the profile of the newly-diagnosed leprosy patients and some important aspects of the control programme in 1978-81 was collected when monotherapy with dapsone was the practice . Among the new patients, 55 per cent were males, 24 per cent were children, 6 per cent had lepromatous leprosy and 9 per cent had a deformity . About 65 per cent were detected by active case-finding methods and 25 per cent were voluntary referrals . Of the total diagnosed patients, only 68 per cent started treatment; further, of these, about 40 per cent collected drugs for at least 6 months in the first year of treatment . The average attendance at the clinic was 34 per cent of the due attendance . Coverage in the annual examination of family contacts was 57 per cent . During the 4 yr period, about 70 per cent of the villages had population surveys with a coverage of 75 per cent or more . The introduction of multi-drug therapy has provided a new impetus to the programme and therefore a similar study is called for to provide valuable information about the extent of improvement in completion rates and overall impactPIP: Researchers used 1978-1981 data from a random sample of 100 rural subcenters of 14 government leprosy control unites (LCUs) in Tamil Nadu, India to learn the status of indices of the National Leprosy Control Programme . The lepromatous rate stood at 6% (4-10% range) with males having a much higher percentage (8.6% vs . 3.9%) and children having a much lower percentage (.6% vs . 8.3%) . It was highest for those who voluntarily reported their condition (10.4%) and lowest for those detected during a routine contact examination (2.5%) . Active case finding techniques detected most new cases {65%; population survey (59%) and routine contact examination (6%)} followed by voluntary referrals (25%; 8-44% range) and miscellaneous sources . Only 68% (38-82%) of newly diagnosed cases started treatment in the same calendar year or the next . Indeed 46% of those detected by population survey and 37% of contact examination did not even register . Therefore the active case finding program does not bring full benefits to patients . It should undertake intense efforts to motivate all diagnosed patients to take treatment . Moreover, as many as 60% of patients did not collect the drugs needed for the multidrug therapy for =or + 6 months . Some of these did not even do so in the 1st year of treatment . As time passed, the levels of irregularity rose (65% in 2nd year and 71% in 3rd year) . Perhaps LCU staff should use a group approach to motivate patients to collect the needed drugs rather than an individualizes approach . In addition, 18-89% of patients at time of registration had a bacteriological examination in 12 LCUs . The median stood at 69% . The percentage of patients who have a bacteriological examination must improve since the multidrug therapy regimen is dependent on the type of leprosy the patient has (multi or pauci bacillary leprosy) .

Arch Neurobiol (Madr), 1991 Jul-Aug, 54(4), 146 - 50
{Primary neural leprosy . Report of a case}; Cremades Mira A et al.; We present the clinical manifestations and morphological characteristics in a case from a patient with familial history of leprosy that presented peripheral neuropathy without cutaneous lesions . The nerves affected showed a loss of myelinated and unmyelinated fibers as well as chronic inflammatory infiltrate . The presence of Hansen's bacillus was demonstrated in macrophages, Schwann cells, endothelial cells, and fibroblasts at light and ultrastructural levels . In this case, the demonstration of the bacillus in the biopsy of peripheral nerve confirmed the diagnosis.

J Gen Microbiol, 1991 Jul, 137 ( Pt 7), 1619 - 23
Purification and some characteristics of a calcium-binding protein from Bacillus cereus spores; Shyu YT et al.; A novel calcium-binding protein has been purified from the dormant spores of Bacillus cereus T . Purity of this protein was verified by SDS-PAGE and reversed-phase HPLC . Its calcium-binding ability was verified by a competitive calcium-binding assay using Chelex-100 resin and 45Ca autoradiography . The protein is heat-stable and is retained by hydrophobic matrices (phenyl-Sepharose) in a calcium-dependent manner . SDS-PAGE and amino acid composition indicate the molecular mass of the protein to be 24 kDa.

Eur Respir J, 1991 Jul, 4(7), 783 - 8
Association of increased mycobacterium growth inhibitory factor with antituberculous immunity; Chandrasekhar S et al.; Cell-mediated immune mechanisms (CMI) were studied in 51 patients of pulmonary tuberculosis to evaluate the role of mycobacterium growth inhibitory factor in prognosis of the infection, before and after the administration of anti-tubercular drugs . Twenty five Mantoux negative individuals who were subsequently bacille Calmette-Guerin (BCG) vaccinated and 25 Mantoux positive, non-tuberculous controls were included in the study . Their clinical assessment was compared with skin sensitivity (Mantoux); lymphocyte transformation (LT) after stimulation with phytohaemagglutinin (PHA) and purified protein derivative (PPD); macrophage migration inhibitory factor (MIF), mycobacteria growth inhibitory factor (Myco IF) and listerial growth inhibitory factor (List IF) . The tests were carried out at the beginning of the treatment and at intervals of three months, extending to one year . In the case of Mantoux positive controls, tests were carried out only once . It was found that Mantoux reaction had no correlation with LT, MIF, Myco IF and List IF . Both MIF and Myco IF, were significantly elevated in improving patients, whereas increase in List IF was not significant . An important finding was that Myco IF was at a higher level in improving patients whereas in those not responding to chemotherapy it was low.

Eur Respir J, 1991 Jul, 4(7), 778 - 82
Isoniazid resistant tuberculosis in a school outbreak: the protective effect of BCG; Shannon A et al.; An outbreak of isoniazid resistant tuberculosis occurred in a large second level school . A total of 1,160 teenage pupils were at risk . Nineteen cases of tuberculosis were diagnosed, 15 were students, 9 of whom were among 251 non-vaccinated students and 6 among 909 vaccinated students . Two cases of miliary tuberculosis, one of whom also had tuberculous (TB) meningitis, occurred in the non-vaccinated group . The number of children with Heaf grade +3 or +4 was significantly greater among children who had been given Bacille Calmette-Guerin (BCG) vaccination (8 vs 4.4%) . This suggests a boosting effect on the response in vaccinated children . The protective effect of neonatal BCG vaccination in this school outbreak suggests that it provides significant protection against tuberculosis lasting into adolescence.

Biotechniques, 1991 Jul, 11(1), 76 - 8, 80, 82-7
Bst DNA polymerase permits rapid sequence analysis from nanogram amounts of template; Mead DA et al.; A simple, rapid method is presented for the enzymatic sequence analysis of nanogram amounts of single-stranded or double-stranded DNA . This approach employs the thermostable DNA polymerase from Bacillus sterothermophilus and exploits its ability to efficiently extend all of the template-primer complex, even at low substrate concentrations . The procedure requires few pipetting steps, no preannealing step and very short reaction time . This method can significantly reduce the cost associated with DNA polymerase and the amount of template and time required to perform the enzymatic sequencing reactions . As little as a 10-ng aliquot of such sequencing reactions can be analyzed on a fluorescence-based capillary gel electrophoresis instrument recently developed in our laboratory . This highly sensitive detection, in conjunction with the ability to efficiently sequence nanogram amounts of template, strongly suggests the feasibility of direct DNA sequencing of single bacteriophage M13 plaques without prior amplification.

Antibiot Khimioter, 1991 Jul, 36(7), 19 - 22
{Various properties of RifR mutants of the plague agent}; Lebedeva SA et al.; The properties of the RifR mutants of four vaccinal and two natural strains of the plague bacillus were studied . The frequency of the mutants not growing on the complete nutrient medium after increasing the cultivation temperature to 37 degrees C averaged to 3.10(-1) . The frequency of the loss of every of the three known autonomous plasmids or combinations of the 6-mD and 65-mD plasmids was n.10(-1) . In the mutants of some strains with the preserved 47-mD plasmid sensitivity to the Ca2+ deficiency markedly lowered (but was not lost) . The mutants of the majority of the strains produced much lower quantities of bacteriocin (pesticin I) . The nutritional requirements, enzymatic activity, sensitivity to the diagnostic phage and the level of the production of fraction I, a specific antigen, in the bacillus did not change.

Angiologia, 1991 Jul-Aug, 43(4), 165 - 70
{Therapeutic procedure for tuberculous thrombophlebitis}; Batista Thomaz J; The author describe five cases of thrombophlebitis unchained for the bacillus of Koch, which first focus was not detected through the methods of search often used in this kind of work . It's a very rare peripheral vascular disease . The clinical manifestations which follows its installation has in the pain its most important symptom . The surgical resection of the vein was the most suitable conduct with the present pathological reality, once the clinical measures were unable to bring satisfactory results.

J Biochem (Tokyo), 1991 Jul, 110(1), 88 - 95
Kinetics of the hydrolysis of monodispersed and micellar phosphatidylcholines catalyzed by a phospholipase C from Bacillus cereus; Ikeda K et al.; The phosphatidylcholine-hydrolyzing phospholipase C, so-called "phospholipase C" (PLC), was isolated from the culture of Bacillus cereus strain IAM 1208 . The amino-acid composition and partial N-terminal sequence of the purified enzyme were in good agreement with those expected from the nucleotide sequence for a PLC of strain ATCC 10987 {Johansen et al . (1988) Gene 65, 293-304} . The chain-length dependence of kinetic parameters for the PLC-catalyzed hydrolysis of monodispersed short-chain phosphatidylcholines (diCNPC, N = 3-6) was studied by a pH-stat assay method at 25 degrees C, pH 8.0, and ionic strength 0.2 in the presence of saturating amounts of Zn2+ (0.1 mM) . The result was compared with those for snake venom phospholipases A2 {Teshima et al . (1989) J . Biochem . 106, 518-527} . It was found that the interaction of the PLC with the head group of the substrate molecule is very important for the binding . The pH dependences of kinetic parameters for the hydrolysis of monodispersed diC5PC and mixed micelles of diC16PC with Triton X-100 were also studied under the same conditions . An ionizable group, whose pK value is perturbed from 7.77 to 8.30 by substrate binding, was found to be essential to the catalysis . This group was tentatively assigned to His 14 on the basis of the results on X-ray crystallographic and chemical modification studies {Hough et al . (1989) Nature 338, 357-360 and Little (1977) Biochem . J . 167, 399-404}.(ABSTRACT TRUNCATED AT 250 WORDS)

J Biochem (Tokyo), 1991 Jul, 110(1), 111 - 9
The nucleotide sequence for a proline-activating domain of gramicidin S synthetase 2 gene from Bacillus brevis; Hori K et al.; A fragment encoding proline-activating domain (grs 2-pro) of gramicidin S synthetase 2 (GS 2) was found in an 8.1-kilobase pairs (kb) DNA fragment of Bacillus brevis Nagano, which contained the full length of GS 1 gene (grs 1) . The clones designated GS719 and GS708, which expressed gramicidin S synthetase 1, were elucidated to express immunoreactive proteins to GS 2 antibodies with approximate molecular weights of 115,000, 105,000 (GS719), and 110,000 (GS708) . The partial purification of the gene products of these clones was carried out using DEAE-Sepharose CL-6B column chromatography . The immunoreactive proteins to GS 2 antibodies were separated from gramicidin S synthetase 1 protein and had specific proline-dependent ATP-32PPi exchange activity . The nucleotide sequence for the proline-activating domain in the 8.1-kb insert was determined . This fragment was 2,879 base pairs long, and encoded 959 amino acids . The calculated molecular weight of 111,671 was consistent with the apparent molecular weight of 115,000 found in SDS-PAGE of the immunoreactive products to GS 2 antibodies . The open reading frame for this protein followed grs 1 gene, though two were separated by a 73-base pair noncoding sequence, and remained open to the end.(ABSTRACT TRUNCATED AT 250 WORDS)

Curr Genet, 1991 Jul, 20(1-2), 5 - 8
Construction of a Saccharomyces cerevisiae strain able to ferment cellobiose; Adam AC et al.; The bglA gene, encoding a beta-glucosidase from Bacillus polymyxa, has been expressed in Saccharomyces cerevisiae under control of the CYC-GAL promoter inducible by galactose . The expression of bglA-encoded activity in the strain used as a host was not sufficient to allow its growth with cellobiose as a carbon source . However, a recessive mutation in a gene designated cem1 has been obtained which, combined with the expression of beta-glucosidase activity, allows the growth of S . cerevisiae on cellobiose . The expression of the blgA gene in a cem1 strain confers on S . cerevisiae the capability for an efficient fermentation of cellobiose, as detected by the formation of CO2.

Semin Surg Oncol, 1991 Jul-Aug, 7(4), 221 - 9
Immunotherapy of renal cell cancer; Crusinberry R et al.; Immunotherapy of metastatic renal adenocarcinoma (RCC) is currently an alternative to cytotoxic chemotherapy . Bacillus Calmette-Guerin has been associated with a 22% response rate in small series, but no large-scale clinical trials have been completed . Transfer factor, in combination with other immunotherapeutic and chemotherapeutic compounds, has a reported 13% incidence of response . Tumor vaccines have caused clinical response in only 5% of patients while monoclonal antibodies have produced partial remission in one of nine patients . Immune RNA has been associated with a 14% overall incidence of response . Tumor necrosis factor has not as yet been studied in any large-scale clinical investigations but preliminary studies are not promising . Leukocyte-derived and recombinant interferons alone have produced responses in 10-20% of patients with tolerable toxicity . Combinations of interferons or with cytotoxic chemotherapy have produced slightly improved responses with short duration and substantial toxicity . Adoptive immunotherapy using Interleukin-2 alone, or with IL-2 plus lymphocyte-activated killer cells, or tumor infiltrating lymphocytes or interferons have produced clinical responses in 10-30% of patients treated . Combinations of specific forms of immune therapy may hold promise for better rates of clinical response in the future.

Semin Surg Oncol, 1991 Jul-Aug, 7(4), 211 - 6
Immunotherapy of breast cancer: a review of the development of cell-specific therapy; Lytle GH; A brief history of immunotherapy for breast cancer is presented, with emphasis on how theories developed as the field of immunology became more sophisticated . Non-specific therapies, such as Bacillus Calmette-Guerin, levamisole, interferon, interleukin, and others are reviewed . A form of cell-specific immunotherapy is then presented, and some current results are summarized . Problems and proposals for the future development of immunotherapy for breast cancer are then presented.

Int J Hyperthermia, 1991 Jul-Aug, 7(4), 643 - 51
Bacterial endotoxin lipopolysaccharide modulates synthesis of the 70 kDa heat stress protein family; Tomasovic SP et al.; Murine bacillus Calmette-Guerin activated macrophages release several monokines when triggered by the bacterial endotoxin lipopolysaccharide (LPS); this has recently been reported to be strongly influenced by the sequence of hyperthermic and LPS treatments . In the work reported here, it was found that LPS treatment markedly modulated the rate of synthesis of proteins in the heat stress protein (HSP) 70 family in these macrophages . The rate of synthesis of the HSP 70 family was slightly reduced if the cells were incubated with LPS 4 h prior to heating at 43 degrees C for 1 h, but was greatly reduced as the triggering time approached the initiation of heating and was nearly completely abrogated if the LPS triggering immediately preceded or followed heating . Near-normal rates of HSP 70 synthesis occurred if the triggering was delayed until 1-2 h after the heating ended . The LPS-triggered release of tumour necrosis factor (TNF) was also reduced as the time of LPS addition approached the heating time, but this depressed release preceded the effects on HSP 70 synthesis and did not recover for up to 3 h after heating . The effects of LPS on HSP 70 synthesis also occurred in a murine monocytic cell line, PU5-1.8, which releases TNF in response to LPS, and in a murine fibroblast cell line, NIH/3T3 . This indicates that these effects are not restricted to cells of monocyte or macrophage lineage . The nature of the transcriptional or translational mechanisms controlling these responses is unknown, but these data may contribute to the understanding of (1) the regulation of the HSP 70 family and (2) TNF processing in stressed cells.

J Assoc Off Anal Chem, 1991 Jul-Aug, 74(4), 704 - 6
Symposium on microbiology update: old friends and new enemies . Bacillus cereus; Jackson SG; Bacillus cereus is an environmentally ubiquitous, Gram-positive, spore-forming bacillus responsible for 2 distinct foodborne disease syndromes as well as other manifestations of pathogenicity . The rapid-onset, "emetic," foodborne-disease syndrome is associated with an emetic toxin; the delayed-onset, "diarrheal" syndrome is associated with elaboration of enterotoxin . The majority of methods for detection of these toxins have relied on in vivo testing . More recent work on purification of enterotoxin facilitated the development of a rapid, specific, fluorescent immunodot assay and a tissue culture screening assay for enterotoxin . Work on characterization and detection of emetic toxin is ongoing.

Can J Microbiol, 1991 Jul, 37(7), 513 - 20
Occurrence of antibiotic and metal resistance and plasmids in Bacillus strains isolated from marine sediment; Belliveau BH et al.; Eleven hundred Bacillus strains isolated from marine sediment from the Minas Basin, Nova Scotia, Canada, were purified on LB agar supplemented with ampicillin, chloramphenicol, erythromycin, streptomycin, tetracycline, or mercuric chloride . Seventy-seven isolates were examined for plasmid DNA, and for resistance to 11 antibiotics, HgCl2, and phenylmercuric acetate . Minimum inhibitory concentrations of Ag, Cd, Co, Cu, and Zn were also determined . Forty-three percent of antibiotic- and mercury-resistant strains contained one or more plasmids ranging from 1.9 to 210 MDa . Fifty-four percent carried plasmids greater than 20 MDa, and 97% were resistant to two or more metals . There was no correlation between plasmid content and resistance either to antibiotics or to mercurial compounds in these strains . Mercury-resistant isolates were unable to transform Hg2+ to volatile Hg0 by virtue of a mercuric reductase enzyme system (mer) . Strains resistant to Hg2+ were investigated for their ability to produce H2S and intracellular acid-labile sulfide when grown in the absence and presence of HgCl2 . Lower levels of H2S and intracellular sulfide were detected only in metal-resistant strains grown in the presence of HgCl2, suggesting that cellular sulfides complexed with Hg2+ in these strains.

J Invertebr Pathol, 1991 Jul, 58(1), 33 - 9
Two new isolates of Bacillus thuringiensis pathogenic to Spodoptera litura; Whitlock VH et al.; Both the standard Bacillus thuringiensis kurstaki (HD-1) and the formulated commercial product resulted from this strain have shown limited pathogenicity against the tobacco cutworm (Spodoptera litura) . However, two new isolates of Bacillus thuringiensis (K-2074 and K-2178) isolated from Taiwan have been identified through an active screening program to be highly pathogenic against the tobacco cutworm . In this paper, we present results of characterization and the pathogenicity of these two new isolates.

J Thorac Imaging, 1991 Jul, 6(3), 6 - 13
Legionnaires' disease; Fairbank JT et al.; The Legionella bacillus is a relatively common pulmonary pathogen that has been responsible for a number of outbreaks of respiratory illness this century . Not until 1976, however, after exciting epidemiologic and microbiologic investigation, was the organism isolated and identified . Legionnaires' disease does not have a characteristic radiographic appearance, but certain features may alert the clinician to its presence . It often rapidly progresses to a lobar pneumonia that may not respond immediately to treatment . The radiographic findings lag behind clinical improvement, and radiographic resolution is prolonged . Organ transplantation patients often present with ill-defined, rounded, pleura-based opacities that may simulate pulmonary infarction and can cavitate.

J Gen Virol, 1991 Jul, 72 ( Pt 7), 1735 - 9
Characterization of the genome of cacao swollen shoot virus; Lot H et al.; Cacao swollen shoot disease has been known to be caused by a small non-enveloped bacilliform virus for more than 25 years . Purification using a combination of celite filtration, polyethylene glycol concentration and sucrose density gradient centrifugation has yielded concentrated preparations of purified cacao swollen shoot virus (CSSV) . Results of nuclease sensitivity tests indicated that the CSSV genome consists of dsDNA which has two single-stranded regions . The approximate size of CSSV DNA calculated from restriction enzyme digests is 7.4 kbp . It is very likely that CSSV is a member of the commelina yellow mottle virus group.

G E N, 1991 Jul-Sep, 45(3), 190 - 5
{Abdominal tuberculosis}; Salazar S et al.; 20 cases of abdominal tuberculosis (TB) were evaluated; from these, 10% with intestinal TB without peritoneal involvement and 90% presenting TB of peritoneal localization . 80% of the patients showed clinical manifestation in other organs . Pleura-lung alterations were found in 83% of the cases after X-ray chest examination . The diagnosis of abdominal TB was based on finding of caseating tuberculoid granuloma (65%), anti-TB therapeutic response (30%) and positive observation of acid-fast bacillus in sputum (5%) . The conclusions from this review are that: 1) Patients with chronic illness, negative cultures and clinical evidence of infectious etiology are highly suspicious; 2) Analysis of pathologic specimens was the most accurate diagnostic method; 3) when abdominal TB is suspected a stepwise methodology must be followed to confirm diagnosis; 4) Anti-TB therapy must be started as soon as diagnosis is confirmed.

Pneumoftiziologia, 1991 Jul-Sep, 40(3), 48 - 51
{The attitude of pulmonary tuberculosis patients towards treatment, a factor in the efficacy of chemotherapy}; Bicica V et al.; The paper reports on the efficiency at the first treatment of pulmonary tuberculosis chemotherapy in a homogeneous group of 124 bacillary men, belonging to the territory of the Tb out-patient clinic no . 2, all admitted in the Galati Hospital of pulmonary diseases, between 1985 and 1986 . This group was divided into 4 subgroups, as a function of the degree of participation to chemotherapy (intermittent treatment 2/7 intensive, for a short period-6 months), i.e.: a) cooperative-conformist in 66 patients; b) cooperative-over-particular in 20 patients without therapeutic failure; c) non-cooperative by interest 21 (16.93%) with 11 failures (52.3%); d) completely noncooperative (nonsocial) 17-13.71% with 13 failures (76.48%) . In the 124 patients, 26 failures were recorded (20.97%) but in 19.35% the therapeutic failure was determined by non-compliance; of them, 20 patients left the hospital without the physician's advice . For improving compliance, the individual health education, cooperation with psychologist or psychiatrist, the final compulsory isolation of the nonsocial patients are recommended.

Biochimie, 1991 Jul-Aug, 73(7-8), 855 - 60
Characterization and primary structure of proteins L28, L33 and L34 from Bacillus stearothermophilus ribosomes; Kruft V et al.; The complete amino acid sequences of 3 proteins from the 50S subunit of Bacillus stearothermophilus ribosomes were determined by N-terminal sequence analysis and by sequencing of overlapping fragments obtained from enzymatic digestions and chemical cleavages . The proteins BstL28, BstL33 and BstL34, named according to the equivalent proteins in Escherichia coli ribosomes, consist of 60, 49, and 44 amino acid residues and have calculated molecular masses of 6811.0, 5908.6, and 5253.9 Da, respectively . They are highly basic with a content of positively charged residues ranging between 29% for L33 and 45% for L34 . The 3 proteins were positioned in the 2-dimensional map of B stearothermophilus 50S ribosomal proteins . The electrophoretic mobilities confirm sizes and net charges deduced from the sequences.

Anticancer Res, 1991 Jul-Aug, 11(4), 1625 - 8
Potentiation of antitumor activity of bleomycin towards solid tumors in mice by Bacillus thuringiensis subsp . israelensis toxin; Yokoyama Y et al.; We previously reported that a 25-kDa Bacillus thuringiensis subsp . israelensis (BTI) toxin is cytotoxic to cultured tumor cells and can also potentiate the cytotoxic effect of some antitumor agents in vitro . In this study, we examined the in vivo effect of BTI toxin on the potentiation of antitumor activity of bleomycin (BLM) against solid tumor-bearing mice . Three tumor cell lines, Ehrlich carcinoma, B16 melanoma and Meth A fibrosarcoma, were employed . It was shown that dose of BTI toxin, ineffective alone, potentiated the antitumor activity of BLM when used in combination.

J Assoc Off Anal Chem, 1991 Jul-Aug, 74(4), 649 - 51
Lipid globule staining to aid in differentiating Bacillus species; Harmon SM et al.; The use of the lipid globule stain to aid in differentiating the Bacillus cereus group (i.e., B . cereus, B . cereus var . mycoides, and B . thuringiensis) from other Bacillus species was investigated . Smears from colonies grown on suitable agar were made on precleaned slides, stained, and examined microscopically for characteristic deep blue lipid globules . The study included a total of 649 cultures of Bacillus species plus 143 incompletely characterized Bacillus isolates from food . Only B . cereus, B . cereus var . mycoides, B . thuringiensis, B . megaterium, and B . sphaericus were consistently positive for lipid globules, although at times, a few cells of B . aneurinolyticus and B . thiaminolyticus were also positive . The lipid globule stain procedure is of value in differentiating Bacillus species, especially when performed by an experienced analyst and used in conjunction with tests for cell and spore morphology.

J Clin Microbiol, 1991 Jul, 29(7), 1299 - 302
Molecular characterization and proposal of a neotype strain for Bartonella bacilliformis; Brenner DJ et al.; Bartonella bacilliformis, the etiologic agent of bartonellosis, was characterized biochemically and by DNA hybridization, guanine-plus-cytosine content, genome size, and 16S rRNA sequencing . DNAs from the two strains in our collection exhibited 97% relatedness in hydroxyapatite reactions done at 55 degrees C (optimal reassociation criterion) and 100% relatedness in reactions done at 70 degrees C (stringent reassociation criterion) . There was no evidence of divergence within the related sequences . B . bacilliformis DNA showed no relatedness to the cat scratch disease bacillus or to a strain of a second species in the same genus as the cat scratch disease bacillus in hybridization reactions done at 65 degrees C . The guanine-plus-cytosine contents of DNAs from the two B . bacilliformis strains were 39 and 40 mol% . Time course reassociation, done by determining spectrophotometrically the time required for one-half of the denatured DNA to form duplexes, indicated that B . bacilliformis has a genome size of approximately 4 x 10(8) . The 16S rRNA sequence analysis indicated that B . bacilliformis is in the alpha-2 subgroup of the purple bacteria, class Proteobacteria, and that its closest relatives are Rochalimaea quintana and Brucella abortus . Strain KC583 (= Herrer 020/F12,63 = ATCC 35685) is proposed as the type strain of B . bacilliformis.

EMBO J, 1991 Jul, 10(7), 1607 - 18
Crystal structure of Penicillium citrinum P1 nuclease at 2.8 A resolution; Volbeda A et al.; P1 nuclease from Penicillium citrinum is a zinc dependent glyco-enzyme consisting of 270 amino acid residues which cleaves single-stranded RNA and DNA into 5'-mononucleotides . The X-ray structure of a tetragonal crystal form of the enzyme with two molecules per asymmetric unit has been solved at 3.3 and refined at 2.8 A resolution to a crystallographic R-factor of 21.6% . The current model consists of 269 amino acid residues, three Zn ions and two N-acetyl glucosamines per subunit . The enzyme is folded very similarly to phospholipase C from Bacillus cereus, with 56% of the structure displaying an alpha-helical conformation . The three Zn ions are located at the bottom of a cleft and appear to be rather inaccessible for any phosphate group in double-stranded RNA or DNA substrates . A crystal soaking experiment with a dinucleotide gives clear evidence for two mononucleotide binding sites separated by approximately 20 A . One site shows binding of the phosphate group to one of the zinc ions . At both sites there is a hydrophobic binding pocket for the base, but no direct interaction between the protein and the deoxyribose . A cleavage mechanism is proposed involving nucleophilic attack by a Zn activated water molecule.

J Gen Microbiol, 1991 Jul, 137 ( Pt 7), 1635 - 9
Phosphatidyltransferase activity in Bacillus megaterium; Morii H et al.; Phosphatidyl transfer between phosphatidylethanolamine, phosphatidylglycerol or phosphatidylserine as donors and primary hydroxyl acceptors including ethanolamine, glycerol, serine and Triton X-100 has been shown to be catalysed by membrane particles derived from Bacillus megaterium strains ATCC 13632 and ATCC 14581 . The rate of cardiolipin synthesis from phosphatidylglycerol in the presence of ethanolamine was an order of magnitude greater than that of phosphatidylethanolamine formation . Cardiolipin synthesis from phosphatidylethanolamine in the presence of glycerol was also observed, and was 1.5-fold greater than the formation of phosphatidylglycerol . Similar heat lability, effects of pH and of Triton X-100 for phosphatidyl transfer and cardiolipin synthesis indicate that both reactions were catalysed by cardiolipin synthase.

Plasmid, 1991 Jul, 26(1), 67 - 73
Conjugal transfer of recombinant transposon Tn916 from Escherichia coli to Bacillus stearothermophilus; Natarajan MR et al.; A gene for thermostable amylase has been inserted at the BstXI site of Tn916 . Mating experiments demonstrated that unlike Tn916, the recombinant transposon, designated Tn916A, could transfer from Escherichia coli to Bacillus stearothermophilus BR219 in broth matings, resulting in chromosomal integration of the transposon and expression of the amylase at significant levels.

J Bacteriol, 1991 Jul, 173(14), 4526 - 9
IS231A from Bacillus thuringiensis is functional in Escherichia coli: transposition and insertion specificity; Hallet B et al.; A kanamycin resistance gene was introduced within the insertion sequence IS231A from Bacillus thuringiensis, and transposition of the element was demonstrated in Escherichia coli . DNA sequencing at the target sites showed that IS231A transposition results in direct repeats of variable lengths (10, 11, and 12 bp) . These target sequences resemble the terminal inverted repeats of the transposon Tn4430, which are the preferred natural insertion sites of IS231 in B . thuringiensis.

Agric Biol Chem, 1991 Jul, 55(7), 1695 - 9
Enzymatic properties of dipeptidyl carboxypeptidase from Bacillus pumilus; Nagamori Y et al.; Enzymatic properties of dipeptidyl carboxypeptidase (DCP) from Bacillus pumilus were investigated . The enzyme was more active on tri- and tetrapeptides than angiotensin-converting enzyme (ACE) from rabbit lung . The presence of chloride ion is essential for the hydrolysis . The Km value of angiotensin I for the enzyme was 0.119 x 10(-3) M . The enzyme was not inhibited by the mammalian ACE inhibitors lisinopril and enalaprilat . The enzyme is readily inhibited by EDTA but restored by Co2+, Mn2+, and Zn2+ . Therefore, it seems to be a zinc-metallo protease.

J Biotechnol, 1991 Jul, 19(2-3), 221 - 40
Genetic manipulation of Bacillus amyloliquefaciens; Vehmaanpera J et al.; Application of modern gene technology to strain improvement of the industrially important bacterium Bacillus amyloliquefaciens is reported . Several different plasmid constructions carrying the alpha-amylase gene (amyE) from B . amyloliquefaciens were amplified in this species either extrachromosomally or intrachromosomally . The amyE gene cloned on a pUB110-derived high copy plasmid pKTH10 directed the highest yields both in rich laboratory medium and in crude industrial medium . The alpha-amylase activity, when compared with the parental strain, was enhanced up to 20-fold in the pKTH 10 transformant . This strain showed decreased activities for other exoenzymes, such as proteases and beta-glucanase suggesting common limiting resources in the processing of these enzymes . Deletions were made in vitro in genes encoding neutral (nprE), alkaline (aprE) protease and beta-glucanase (bglA) . The engineered genes were cloned into the thermosensitive plasmid pE194, and the resulting plasmids were used to replace the corresponding wild type chromosomal genes in B . amyloliquefaciens by integration-excision at non-permissive temperature . The double mutant deficient in the major proteases (delta nprE delta aprE) showed about a 2-fold further enhancement in alpha-amylase production in the industrial medium compared with the relevant wild type backgroud, both when plasmid-free and when transformed with pKTH10; this strain also produced elevated levels of the chromosomally-encoded beta-glucanase; pKTH10 was stably maintained both in the wild type strain and in the delta nprE delta aprE mutant . We suggest that the higher yields in alpha-amylase and beta-glucanase in the delta nprE delta aprE strain are primarily due to improved access to limiting resources, and that decreased proteolytic degradation may have had a secondary role in retaining the high activity obtained.

Cancer, 1991 Jun 15, 67(12), 3024 - 8
ABO(H) antigens and beta-2 microglobulin in transitional cell carcinoma . Predictors of response to intravesical bacillus Calmette-Guerin; Sanders H et al.; The response of patients with superficial transitional cell carcinoma of the bladder (STCB) to intravesical chemotherapy is variable; some patients enjoy a long period without recurrence, whereas others have recurrence of tumor within 2 years of removal of the primary lesion . Previously, others have demonstrated that the loss of normal cell surface antigens, such as ABO(H) blood group antigens or beta-2 microglobulin (B2M) has been correlated with more aggressive behavior by tumor . In this study, using immunohistochemical techniques, the authors evaluated the initial pretreatment biopsy specimen of bladder tumors for the presence of ABO(H) antigens and B2M . Data from this sample patient population, all with biopsy-proven STCB, indicate that expression of these two markers is predictive of a therapeutic response to prophylactic intravesical bacillus Calmette-Guerin (BCG) (Tice strain) after resection, and that expression of the two markers is of greater predictive value than expression of either antigen alone.

J Biol Chem, 1991 Jun 25, 266(18), 11909 - 14
Characterization of recombinant Bacillus megaterium cytochrome P-450 BM-3 and its two functional domains; Li HY et al.; Bacillus megaterium cytochrome P-450BM-3 and its two functional domains, the heme and flavin domains, have been purified and characterized using an Escherichia coli expression system . Recombinant P-450BM-3 behaves both spectrally and enzymatically the same as the enzyme produced from the natural host, B . megaterium, and another E . coli system recently described (Bouddupalli, S . S., Estabrook, R . W., and Peterson, J . A . (1990) J . Biol . Chem . 265, 4233-4239) . Reduction of the flavins in P-450BM-3 domain with NADPH appears to be very similar to microsomal P-450 reductases where two reducing equivalents are consumed to fully reduce the FMN while the FAD is converted to the semiquinone in an one electron reduction . NADPH reduction of the heme occurs only in the presence of substrate suggesting, by analogy with the cytochrome P-450CAM system, a possible increase in iron redox potential of the heme upon substrate binding which facilitates electron transfer from the flavins to the heme . The flavin domain retains a high level of cytochrome c reductase activity and also reacts with NADPH to give a 3-electron reduced product . The heme domain retains the ability to bind substrate and generates the characteristic 450-nm absorption band upon reduction in the presence of CO . The heme domain has been crystallized and a preliminary set of x-ray diffraction data obtained.

Biochim Biophys Acta, 1991 Jun 24, 1078(2), 127 - 32
Characterization of Bacillus stearothermophilus cyclodextrin glucanotransferase in ascorbic acid 2-O-alpha-glucoside formation; Tanaka M et al.; In this study, we characterized cyclodextrin glucanotransferase (CGTase) from Bacillus stearothermophilus in L-ascorbic acid-2-O-alpha-D-glucoside (AA-2G) formation and compared its enzymological properties with those of rat intestinal and rice seed alpha-glucosidases which had the ability to form AA-2G . CGTase formed AA-2G efficiently using alpha-cyclodextrin (alpha-CD) as a substrate and ascorbic acid (AA) as an acceptor . Several AA-2-oligoglucosides were also formed in this reaction mixture, and they could be converted to AA-2G by the additional treatment of glucoamylase . The optimum temperature for AA-2G formation was 70 degrees C and its optimum pH was around 5.0 . CGTase also utilized beta- and gamma-CDs, maltooligosaccharides, dextrin, amylose, glycogen and starch as substrates, but not any disaccharides except maltose . CGTase showed the same acceptor specificity as two alpha-glucosidases, whereas its hydrolyzing activity towards AA-2G was very low compared with those of alpha-glucosidases . Cleavage profiles of AA-2-oligoglucosides by CGTase present a possible mechanism for AA-2G formation that CGTase transfers a glucose-hexamer to an acceptor at the first step and then a glucose is stepwisely removed from the non-reducing end of the product through glucoamylase-like action of this enzyme . These results indicate that CGTase is able to synthesize AA-2G more efficiently than rat and rice alpha-glucosidases and utilization of this enzyme makes the mass production of AA-2G possible.

Biochim Biophys Acta, 1991 Jun 24, 1078(2), 133 - 8
The effect of kirromycin on the metal ion coordination in complexes of elongation factor Tu from Bacillus stearothermophilus as inferred from the 17O-55Mn superhyperfine interaction; Kalbitzer HR et al.; The binding of the antibiotic kirromycin to elongation factor Tu (EF-Tu) leads to a severe line broadening of the Q-band manganese EPR lines of EF-Tu.Mn, EF-Tu.Mn.GDP, as well as EF-Tu.Mn.GDP.Pi indicating that the coordination sphere of the metal ion is changed by this interaction . The number of coordinated water molecules was determined from the 17O-55Mn superhyperfine coupling observable in H2(17)O enriched water; it is pH-dependent in the EF-Tu.Mn.GDP complex, at pH 6.8 probably four water molecules are coordinated with the protein bound manganese, at pH 8.4 one of the water molecules is replaced by a functional group from the protein . Independent of the pH, probably four and five water molecules are bound to the metal ion in the EF-Tu.Mn and in the EF-Tu.Mn.GDP.Pi complex, respectively . Kirromycin does not influence the number of water molecules bound to EF-Tu.Mn.GDP and EF-Tu.Mn.GDP.Pi, but leads to an increase of the number of water molecules coordinated to the metal ion in EF-Tu.Mn . The 17O-55Mn superhyperfine coupling constant in Mn(H2O)6 was determined from the EPR-spectra as 0.24 mT.

Pharm Weekbl Sci, 1991 Jun 21, 13(3), 130 - 6
Microbiological aspects of heat sterilization of drugs . III . Heat resistance of spore-forming bacteria, isolated from large-volume parenterals; Boom FA et al.; In order to calculate the minimum sterilization process conditions to obtain the generally accepted sterility level (less than 1.10(-6) probability of microbial survival), we determined the bioburden and its heat resistance of 500 ml large-volume parenteral bottles over a period of 5 years . For the bioburden determination 1,832 bottles were examined by the membrane filtration method . Mean bioburden was 9.36 colony-forming units/bottle . Of the colony-forming units isolated 118 were heat resistant (0.69%) . These were spore-forming Bacillus species . Of the isolated Bacillus species heat resistance was determined in 5% glucose, 0.9% sodium chloride and 8% amino acids solution . D values greater than 1 min at 105 degrees C were found for 2, 5 and 4 different Bacillus species in glucose 5%, sodium chloride 0.9% and amino acids 8%, respectively . 2 Bacillus species showed a D value over 2 min at 105 degrees C in all three media . D values at 110 degrees C in sodium chloride 0.9% for these 2 Bacillus species were 1.8 and 2.6 min and in amino acids 8% 0.9 and 1.7 min, respectively . The minimum sterilization process time at 110 degrees C, calculated with the experimentally determined bioburden and D values is less than 25 min . When introducing reduced exposure times/temperatures, each individual manufacturer should assess the bioburden . The time-consuming determination of the heat resistance of bioburden isolates is not always necessary . By dividing the isolated colony-forming units in a 'heat-resistant' group and a 'not-heat-resistant' group, changing from standard overkill sterilization procedures to processes with lower F0 values is possible.

Biochim Biophys Acta, 1991 Jun 18, 1065(2), 250 - 60
Early response of cultured lepidopteran cells to exposure to delta-endotoxin from Bacillus thuringiensis: involvement of calcium and anionic channels; Schwartz JL et al.; The role of ion channels in the initial steps following exposure of SF-9 lepidopteran insect cells in culture to the delta-endotoxin CryIC from the insecticidal bacterium Bacillus thuringiensis was investigated using single ionic channel measurements and microspectrofluorescence of the calcium-sensitive probe fura-2 . It was found that: (1) the toxin triggers an immediate rise in intracellular calcium; (2) the surge is due to calcium entering the cells via calcium channels; (3) the toxin recruits or introduces anionic channels in the cell's plasma membrane in a time-dependent manner . These channels, not seen in the absence of the toxin, are induced by toxin exposure to either side of the cell membrane . They have a conductance of 26 picosiemens (pS) and are mainly permeable to chloride . This study provides the first evidence of the primary role of calcium and chloride ions in the action of delta-endotoxin on cultured insect cells.

Biochim Biophys Acta, 1991 Jun 17, 1058(2), 131 - 40
Protonophore-resistance and cytochrome expression in mutant strains of the facultative alkaliphile Bacillus firmus OF4; Quirk PG et al.; Two protonophore-resistant mutants, designated strains CC1 and CC2, of the facultative alkaliphile Bacillus firmus OF4 811M were isolated . The ability of carbonyl cyanide m-chlorophenylhydrazone (CCCP) to collapse the protonmotive force (delta mu H+) was unimpaired in both mutants . Both resistant strains possessed elevated respiratory rates when grown at pH 7.5, in either the presence or absence of CCCP . Membrane cytochromes were also elevated: cytochrome o in particular in strain CC1, and cytochromes aa3, b, c and o in strain CC2 . Strain CC2 also maintained a higher delta mu H+ than the others when grown in the absence of CCCP . When grown in the presence of low concentrations of CCCP, strains CC1 and CC2 both maintained higher values of delta mu H+ than the wild-type parent and correspondingly higher capacities for ATP synthesis . In large-scale batch culture at pH 10.5, both mutant strains grew more slowly than the parent and contained significantly reduced levels of cytochrome o . Cells of stran CC1 also displayed a markedly altered membrane lipid composition when grown at pH 10.5 . Unlike previously characterized protonophore-resistant strains of B . subtilis and B . megaterium, neither B . firmus mutant possessed any ability above that of the parent strain to synthesize ATP at given suboptimal values of delta mu H+ . Instead, both resistant alkaliphile strains maintained a higher delta mu H+ and a correspondingly higher delta Gp than the parent strain when growing in sublethal concentrations of CCCP, apparently as a result of mutational changes affecting respiratory chain composition . Also of note in both the mutant and the wild-type strains was a marked elevation in the level of one of the multiple terminal oxidases, an aa3-type cytochrome, during growth at pH 7.5 in the presence of CCCP or during growth at pH 10.5, i.e . two conditions that reduce the bulk delta mu H+.

Proc Natl Acad Sci U S A, 1991 Jun 15, 88(12), 5119 - 23
Resistance to the Bacillus thuringiensis bioinsecticide in a field population of Plutella xylostella is due to a change in a midgut membrane receptor; Ferre J et al.; The biochemical mechanism for resistance to Bacillus thuringiensis crystal proteins was studied in a field population of diamondback moths (Plutella xylostella) with a reduced susceptibility to the bioinsecticidal spray . The toxicity and binding characteristics of three crystal proteins {CryIA(b), CryIB, and CryIC} were compared between the field population and a laboratory strain . The field population proved resistant (greater than 200-fold compared with the laboratory strain) to CryIA(b), one of the crystal proteins in the insecticidal formulation . Binding studies showed that the two strains differ in a membrane receptor that recognizes CryIA(b) . This crystal protein did not bind to the brush-border membrane of the midgut epithelial cells of the field population, either because of strongly reduced binding affinity or because of the complete absence of the receptor molecule . Both strains proved fully susceptible to the CryIB and CryIC crystal proteins, which were not present in the B . thuringiensis formulation used in the field . Characteristics of CryIB and CryIC binding to brush-border membranes of midgut epithelial cells were virtually identical in the laboratory and the field population.

Biochem J, 1991 Jun 15, 276 ( Pt 3), 801 - 7
Irreversible inactivation of beta-lactamase I from Bacillus cereus by chlorinated 6-spiroepoxypenicillins; Gledhill L et al.; On incubation of the chlorinated 6-spiroepoxypenicillin anilides (I) and (II) {formula: see text} with beta-lactamase 1 from Bacillus cereus, three distinct processes are observed . The inhibitors act as (a) substrates, the turnover of which respectively results in a single product, namely 6-substituted 2(H)-3,4-dihydro-1,4-thiazine, (b) a transiently inhibited enzyme complex, and finally (c) an irreversibly inactivated enzyme complex . Although differing only in their stereochemistry at one centre, the anilide (K) is a more potent irreversible inactivator of beta-lactamase I than is compound (II) . Analysis of irreversibly inactivated beta-lactamase I by isoelectric focusing and inspection of peptide fragmentation maps indicated that irreversible inactivation appears to be accompanied by covalent modification . These studies reveal that the chlorinated 6-spiroepoxypenicillin anilide (I) is a mechanism-based beta-lactamase inhibitor.

Gene, 1991 Jun 15, 102(1), 83 - 5
Cloning the BstVI restriction-modification system in Escherichia coli; Vasquez C et al.; A standard DNA modification methyltransferase (MTase) selection protocol was followed to clone the BstVI restriction and modification system from Bacillus stearothermophilus in Escherichia coli . Both genes were contained in a 4.4-kb EcoRI fragment from B . stearothermophilus V chromosomal DNA . The heterologous expression of these genes did not depend on their orientation in the vector, suggesting that the genes are expressed in E . coli under the control of promoters located on the cloned fragment . Subcloning experiments demonstrated that the bstVIR gene was expressed in the absence of its cognate MTase.

Philos Trans R Soc Lond B Biol Sci, 1991 May 29, 332(1263), 171 - 6
Pathway and stability of protein folding; Fersht AR et al.; We describe an experimental approach to the problem of protein folding and stability which measures interaction energies and maps structures of intermediates and transition states during the folding pathway . The strategy is based on two steps . First, protein engineering is used to remove interactions that stabilize defined positions in barnase, the RNAse from Bacillus amyloliquefaciens . The consequent changes in stability are measured from the changes in free energy of unfolding of the protein . Second, each mutation is used as a probe of the structure around the wild-type side chain during the folding process . Kinetic measurements are made on the folding and unfolding of wild-type and mutant proteins . The kinetic and thermodynamic data are combined and analysed to show the role of individual side chains in the stabilization of the folded, transition and intermediate states of the protein . The protein engineering experiments are corroborated by nuclear magnetic resonance studies of hydrogen exchange during the folding process . Folding is a multiphasic process in which alpha-helices and beta-sheet are formed relatively early . Formation of the hydrophobic core by docking helix and sheet is (partly) rate determining . The final steps involve the forming of loops and the capping of the N-termini of helices.

Nature, 1991 Jun 6, 351(6326), 479 - 82
Humoral and cell-mediated immune responses to live recombinant BCG-HIV vaccines; Aldovini A et al.; Several viral and bacterial live recombinant vaccine vehicles are being developed to produce a new generation of vaccines against a broad spectrum of infectious diseases . The human tuberculosis vaccine Mycobacterium bovis bacillus Calmette-Guerin (BCG) has features that make it a particularly attractive live recombinant vaccine vehicle . BCG and other mycobacteria are highly effective adjuvants, and the immune response to mycobacteria has been studied extensively . With nearly two billion immunizations, BCG has a long record of safe use in man . It is one of the few vaccines that can be given at birth, it engenders long-lived immune responses with only a single dose, and there is a worldwide distribution network with experience in BCG vaccination . Recently developed molecular genetic tools and methods for mycobacteria have provided the means to introduce foreign genes into BCG . Here we report that a variety of human immunodeficiency virus type 1 polypeptides can be expressed in BCG recombinants under the control of the mycobacterial hsp70 promoter and that the foreign polypeptides produced in BCG can induce antibody and T-cell responses . These results demonstrate that BCG can be used as a live recombinant vaccine vehicle to induce immune responses to pathogen proteins produced by the bacillus.

Nature, 1991 Jun 6, 351(6326), 456 - 60
New use of BCG for recombinant vaccines; Stover CK et al.; BCG, a live attenuated tubercle bacillus, is the most widely used vaccine in the world and is also a useful vaccine vehicle for delivering protective antigens of multiple pathogens . Extrachromosomal and integrative expression vectors carrying the regulatory sequences for major BCG heat-shock proteins have been developed to allow expression of foreign antigens in BCG . These recombinant BCG strains can elicit long-lasting humoral and cellular immune responses to foreign antigens in mice.

J Biol Chem, 1991 Jun 5, 266(16), 10162 - 7
Generation of EPR-detectable nitrosyl-iron complexes in tumor target cells cocultured with activated macrophages; Drapier JC et al.; After immunostimulation, murine macrophages oxidize L-arginine into nitric oxide (NO) which acts as an effector molecule . In this study, we attempted to establish whether activated macrophage-derived NO forms paramagnetic complexes in tumor target cells which do not express by themselves the L-arginine:NO pathway . Accordingly, murine L1210 leukemia cells were cocultivated with activated peritoneal macrophages from Bacillus-Calmette-Guerin-infected mice, or activated in vitro with interferon-gamma . In control experiments, macrophages were prevented from producing nitrogen oxides by incubation with NG-monomethyl-L-arginine, a specific inhibitor of the L-arginine:NO pathway . After coculture, L1210 cells were removed from adherent macrophage monolayers and analyzed by electron paramagnetic resonance at 77 K . In the L1210 cells cultured with activated macrophages, we detected a signal typical of nitrosyl-iron-sulfur complexes, with g values of 2.041 and 2.015 . This signal was not present when L1210 cells were either cultured alone or cocultured with activated macrophages in the presence of NG-monomethyl-L-arginine . Mitochondria from activated macrophage-injured L1210 cells also exhibited the signal with g values of 2.041 and 2.015 . These results show that when tumor target cells undergo cell-to-cell contact with activated macrophages during culture, the macrophages promote target cell nitrosylation in compartments like mitochondria.

Eur J Biochem, 1991 Jun 1, 198(2), 429 - 35
The cytosolic and glycosomal glyceraldehyde-3-phosphate dehydrogenase from Trypanosoma brucei . Kinetic properties and comparison with homologous enzymes; Lambeir AM et al.; The protozoan haemoflagellate Trypanosoma brucei has two NAD-dependent glyceraldehyde-3-phosphate dehydrogenase isoenzymes, each with a different localization within the cell . One isoenzyme is found in the cytosol, as in other eukaryotes, while the other is found in the glycosome, a microbody-like organelle that fulfils an essential role in glycolysis . The kinetic properties of the purified glycosomal and cytosolic isoenzymes were compared with homologous enzymes from other organisms . Both trypanosome enzymes had pH/activity profiles similar to that of other glyceraldehyde-3-phosphate dehydrogenases, with optimal activity around pH 8.5-9 . Only the yeast enzyme showed its maximal activity at a lower pH . The glycosomal enzyme was more sensitive to changes in ionic strength below 0.1 M, while the cytosolic enzyme resembled more the enzymes from rabbit muscle, human erythrocytes and yeast . The affinity for NAD of the glycosomal enzyme was 5-10-fold lower than that of the cytosolic, as well as the other enzymes . A similar, but less pronounced, difference was found for its affinity for NADH . These differences are explained by a number of amino acid substitutions in the NAD-binding domain of the glycosomal isoenzyme . In addition, the effects of suramin, gossypol, agaricic acid and pentalenolactone on the trypanosome enzymes were studied . The trypanocidal drug suramin inhibited both enzymes, but in a different manner . Inhibition of the cytosolic enzyme was competitive with NAD, while in the case of the glycosomal isoenzyme, with NAD as substrate, the drug had an effect both on Km and Vmax . The most potent inhibitor was pentalenolactone, which at micromolar concentrations inhibited the glycosomal enzyme and the enzymes from yeast and Bacillus stearothermophilus in a reversible manner, while the rabbit muscle enzyme was irreversibly inhibited.

Urology, 1991 Jun, 37(6), 557 - 60
Granulomatous renal mass during endovesical BCG therapy for bladder carcinoma . Diagnosis by fine-needle aspiration; De Boisgisson P et al.; One case of pseudotumoral granulomatous renal mass during endovesical bacillus Calmette-Guerin (BCG) therapy for superficial bladder neoplasm is reported . Such an adverse effect is exceptional and is clearly related to a vesicorenal reflux in our patient . In this case ultrasound-guided fine-needle aspiration was able to settle the diagnosis and avoid surgery . The patient responded to triple antituberculous therapy.

J Urol, 1991 Jun, 145(6), 1316 - 24
Internalization of bacille Calmette-Guerin by bladder tumor cells; Becich MJ et al.; Mechanisms by which intravesical bacille Calmette-Guerin (BCG) treatment mediates antitumor activity are currently poorly understood . We have determined that both human bladder tumor (T-24) and mouse bladder tumor (MBT-2) cells are capable of internalizing the BCG and have characterized this process in vitro . The internalization of BCG by T-24 and MBT-2 cells was verified by histochemistry and electron microscopy . Time dependent internalization of BCG was observed with a maximum occurring at three hrs . Internalization was significantly inhibited by both incubation at 4C and cytochalasin B; conditions known to inhibit phagocytosis . Ultrastructural studies suggested that BCG were transported to membrane bound intracellular compartments and were degraded . The compartments containing the degraded mycobacteria labeled with the fluid phase marker HRP which is known to be transported to lysosomes in a variety of cell types . To determine if the internalization process occurred in vivo, we examined bladder washings of patients treated with intravesical BCG . The majority of the cells in the bladder washings were inflammatory cells which contained ingested BCG . In addition, internalized and degraded BCG were identified in urothelial cells . These data demonstrate that BCG are internalized by transitional epithelial cells both in vitro and in vivo . The internalization process is inhibitable by temperature and microfilament disruption . The potential therapeutic implications of this process and its relationship to antitumor activity of BCG therapy are currently being investigated.

Southeast Asian J Trop Med Public Health, 1991 Jun, 22(2), 160 - 4
Evaluation of Bacillus sphaericus formulations against the vector of bancroftian filariasis; Arunachalalm N et al.; Three different formulations of Bacillus sphaericus viz, Spherimos, Vectobac and Spherifix, were evaluated for their efficacy and residual activity against Culex quinquefasciatus breeding in polluted disused wells . Spherimos, a flowable concentrate formulation, exerted 96-100% control when treated at the dosage of 10 l/ha for 17 days, whereas the effective residual activity lasted up to 67 days at 15 l/ha . In the case of Vectolex, a granular formulation, the residual activity lasted up to 56 days with the dosage of 30 l/ha and up to 66-77 days with higher dosages of 45 and 60 l/ha . The residual activity of Spherifix, a floating controlled release formulation, lasted up to 67 days with a dosage of 10 kg/ha.

J Gen Microbiol, 1991 Jun, 137 ( Pt 6), 1397 - 400
An efficient cyanide-degrading Bacillus pumilus strain; Meyers PR et al.; A Gram-positive, aerobic, endospore-forming bacterium was isolated by an enrichment technique for the ability to degrade cyanide and was identified as a Bacillus pumilus strain . The bacterium rapidly degraded 100 mg l-1 of free cyanide in the absence of added inorganic and organic substances . The ability to degrade cyanide was linked to the growth phase and was not exhibited before late exponential/early stationary phase . Cyanide-degrading activity could not be induced before this time by the addition of 20 mg cyanide l-1 . Production of the cyanide-degrading activity required 0.01 mg Mn2+ l-1 and did not occur at Mn2+ concentrations below 0.002 mg l-1 . Cyanide-degrading activity was intracellular and cell-free extracts rapidly degraded cyanide.

Biol Chem Hoppe Seyler, 1991 Jun, 372(6), 437 - 42
Sialidase in the guinea pig pulmonary parenchyma . Increased activity in the cytosolic and microsomal subcellular fractions after stimulation with Bacillus Calmette Guérin; Terzidis-Trabelsi H et al.; The sialidase activity was assayed in the guinea pig pulmonary parenchyma after removal of bronchoalveolar cells by washing . After differential centrifugation of the crude tissue homogenate, sialidase activities were measured in the subcellular fractions using the fluorogenic substrate 2-(4-methylumbelliferyl)-alpha-D-N-acetylneuraminate . Sialidase activities were found in the lysosomal-enriched (17,000 x g pellet), in the microsomal (105,000 x g pellet) and in the cytosolic (105,000 x g supernatant) fractions . Microsomal and lysosomal forms of sialidase had an optimum activity at pH 3.6-3.8, whereas the optimum for the cytosolic form was pH 4.6 . The activity of all three forms was inhibited by Cu2+, whereas 1 mM Zn2+ and 0.5 mM Ca2+ activated the lysosomal and the cytosolic forms, respectively . In the crude homogenate taken from lungs of Bacillus Calmette Guerin-(BCG-) stimulated guinea pigs, the sialidase activity was increased by 43% (p = 0.025) 3 weeks after the end of the treatment . The cytosolic (+246%) and microsomal (+51%) sialidase activities were significantly increased, whereas the lysosomal sialidase activity was not changed significantly by BCG stimulation.

Biol Chem Hoppe Seyler, 1991 Jun, 372(6), 381 - 3
Microbial metabolism of quinoline and related compounds . IX . Degradation of 6-hydroxyquinoline and quinoline by Pseudomonas diminuta 31/1 Fa1 and Bacillus circulans 31/2 A1; Bott G et al.; Two strains, using 6-hydroxyquinoline as sole source of energy, carbon and nitrogen, have been isolated . These bacteria, designated 31/1 Fa1 and 31/2 A1, are also able to degrade quinoline . According to their physiological properties strain 31/1 Fa1 has been identified as Pseudomonas diminuta and strain 31/2 A1 as Bacillus circulans . 6-Hydroxy-2-oxo-1,2-dihydroquinoline was found as intermediate in the degradation of 6-hydroxyquinoline and quinoline . 2-Oxo-1,2-dihydroquinoline was the first metabolite in the degradation of quinoline.

Biochem J, 1991 Jun 1, 276 ( Pt 2), 401 - 4
Site-directed mutagenesis of dicarboxylic acids near the active site of Bacillus cereus 5/B/6 beta-lactamase II; Lim HM et al.; An amino acid residue functioning as a general base has been proposed to assist in the hydrolysis of beta-lactam antibiotics by the zinc-containing Bacillus cereus beta-lactamase II {Bicknell & Waley (1985) Biochemistry 24, 6876-6887} . Oligonucleotide-directed mutagenesis of cloned Bacillus cereus 5/B/6 beta-lactamase II was used in an 'in vivo' study to investigate the role of carboxy-group-containing amino acids near the active site of the enzyme . Substitution of asparagine for the wild-type aspartic acid residue at position 81 resulted in fully functional enzyme . An aspartic acid residue at position 90 is essential for beta-lactamase II to confer any detectable ampicillin and cephalosporin C resistance to Escherichia coli . Conversion of Asp90 into Asn90 or Glu90 lead to the synthesis of inactive enzyme, suggesting that the spatial position of the beta-carboxy group of Asp90 is critical for enzyme function.

Rev Rhum Mal Osteoartic, 1991 Jun, 58(6), 433 - 9
{Bacillary trochanteritis . Apropos of 30 cases}; Franceschi JP et al.; Tuberculous trochanteritis is rare . The authors report 30 cases, 2/3 of which were seen in sanatoria . They occurred in four cases out of five in patients with long standing tuberculosis but were sometimes the presenting feature . Development of an abscess of the trochanteric serous bursa is a virtually constant complication . Their minimal functional consequences and slow progression endow them with an apparently benign nature which masks the risk of secondary complications (4 cases of coxalgia) . Radiological appearances, minimal in early forms, are currently becoming more abundant by virtue of new medical imaging techniques . Specific antibiotic therapy is adequate in two cases out of three, completed in advanced and aggressive forms by local procedures dominated by surgical excision (13 cases).

J Am Mosq Control Assoc, 1991 Jun, 7(2), 313 - 5
A simple technique for determining relative toxicities of Bacillus thuringiensis var . Israelensis formulations against larval blackflies (Diptera: Simuliidae); Barton WE et al.; A laboratory system for assaying the potency of Bacillus thuringiensis var . israelensis formulations against larval blackflies was developed . An orbital shaker was used to create a water current in 250-ml Erlenmeyer flasks containing the test larvae . This system produced dose-mortality relationships with acceptable statistical parameters.

J Am Mosq Control Assoc, 1991 Jun, 7(2), 307 - 9
Aerially applied, liquid Bacillus thuringiensis var . Israelensis (H-14) for control of spring Aedes mosquitoes in Michigan; Knepper RG et al.; Liquid B.t.i . (Vectobac 12AS), when mixed with water at a 1:3 ratio and applied by helicopter at a rate of 1.17 liters B.t.i . (4.68 liters mix) per ha, was 99% effective in a small (mass median diameter on dye cards: 178 microns) droplet size but ineffective (65%) in a large (553 microns) droplet against spring Aedes larvae in snowmelt pools . There were about 6 times as many smaller droplets as larger ones impacting the treated pools, which probably explained the difference in effectiveness for the 2 treatment regimes . Results indicate that liquid formulations of B.t.i . could be aerially applied for spring Aedes control at a considerable cost savings and efficiency over aerially applied, granular formulations.

J Am Mosq Control Assoc, 1991 Jun, 7(2), 260 - 81
Perspectives on management of pestiferous Chironomidae (Diptera), an emerging global problem; Ali A; In recent years, adult Chironomidae emerging from some urban natural or man-made habitats have increasingly posed a variety of nuisance and economic problems, and in some situations medical problems to humans in different parts of the world . Although there are an estimated 4,000 or more species of chironomid midges worldwide, less than 100 species have been reported to be pestiferous . Among midge control methods, numerous laboratory and field studies have been conducted on the use of organochlorines, organophosphates (OPs), pyrethroids and insect growth regulators (IGRs) . Field use of OP insecticides such as chlorpyrifos, temephos and others in the USA and Japan has generally resulted in larval control for 2-5 wk or longer with application rates below 0.56 kg AI/ha (USA) and less than 1-5 ppm (Japan) . Frequent use of some OP insecticides in the USA and Japan has caused their increased tolerance in several midge species . The IGRs, diflubenzuron and methoprene, provide alternate means for midge control . These IGRs in some situations suppressed adult midge emergence by greater than 90% at rates less than 0.3 kg AI/ha . A number of parasites and pathogens have been reported from midges in different parts of the world . Bacillus thuringiensis serovar . israelensis is effective against some midge species, but at rates at least 10x or higher established for mosquito larvicidal activity . The flatworm, Dugesia dorotocephala, and some fish species offer a good potential for midge control in some situations . In large habitats covering hundreds or thousands of ha, information on the basic ecology of larval midges and adult behavior is essential for formulating midge control criteria . More research is needed on the biological and physical and cultural control of these pestiferous insects.

J Am Mosq Control Assoc, 1991 Jun, 7(2), 188 - 93
Compatibility of Bacillus thuringiensis var . Israelensis and Bacillus sphaericus with the fungal pathogen Lagenidium giganteum (Oomycetes: Lagenidiales); Orduz S et al.; Larvae of Culex quinquefasciatus were exposed to infection by Lagenidium giganteum and various concentrations of B.t.i . or B . sphaericus . The resulting larval mortalities, percentages of infected dead larvae and percentages of larval body regions containing the fungus were compared . Overall, the effectiveness of Lagenidum giganteum against the larvae was not significantly affected by the presence of B.t.i . or B . sphaericus, and the fungal and bacterial agents were compatible . In experiments using 3-day-old larvae, the extent of growth of the fungus in the infected larvae and the percentage of the larvae infected were related to the concentration of B.t.i . in the range of 0.057-0.456 ITU/ml tested but were not related to the concentration of B . sphaericus in the range of 0.6-4.8 x 10(4) spores/ml . With larvae of various ages treated with a low concentration of B.t.i . (0.114 ITU/ml), exposure to the fungus increased the mortality rate in early but not late instars . After single and multiple applications of B.t.i . and B . sphaericus in the presence of the fungus, followed by drying and reflooding, the fungus persisted and reinfected larvae while the B . sphaericus persisted but the B.t.i . did not.

J Am Mosq Control Assoc, 1991 Jun, 7(2), 176 - 9
Field trials with Vectolex (Bacillus sphaericus) and Vectobac (Bacillus thuringiensis (H-14)) against Anopheles gambiae and Culex quinquefasciatus breeding in Zaire; Karch S et al.; Under field conditions in Kinshasa, Zaire, an aqueous suspension of Bacillus thuringiensis (H-14), Vectobac (12-AS), lost most of its larvicidal activity at all concentrations after 48 h against Culex quinquefasciatus breeding in polluted gutter water and Anopheles gambiae breeding in clear water irrigation ponds . However, good control of Cx . quinquefasciatus was obtained using a granular formulation of B . sphaericus, Vectolex-G (ABG-6185), at concentrations of 10-30 kg/ha . High concentrations of Vectolex-G gave excellent control of An . gambiae breeding in irrigation ponds . The Vectobac-G was less active against An . gambiae than Vectolex-G, in spite of good dispersion of Vectobac-G particles.

An Med Interna, 1991 Jun, 8(6), 273 - 80
{Cefotaxime treatment of osteomyelitis of the foot in the diabetic}; Herranz A et al.; Osteomyelitis at the base of an ulcer the foot of diabetic patient is a serious complication usually produced because of the patient's neglect, and entailing difficulties in diagnosis and treatment . Several factors (neurological and vascular ...) favor the onset of the initial ulcer and its evolution, the ulcer subsequently converting into the main door for soft tissue infection with extension to the contiguous bone, with a bad prognosis . Cefotaxime is a 3rd generation cephalosporin, active against coccus gram positive and the majority of aerobic gram negative bacillus . This antibiotic was used in 25 diabetic patients with osteomyelitis at a dosage of 2 gr . IV a/d, during at least 30 days, adding metronidazole when anaerobic bacteria were isolated . The number of patients cured were 21 (84%), one improvement (4%), one resistant (4%) and two relapses (8%) . There were no secondary effects.

Nippon Hinyokika Gakkai Zasshi, 1991 Jun, 82(6), 900 - 6
{The significance of bacillus Calmette-Guerin in the therapy of carcinoma in situ of the bladder}; Kurozumi T et al.; We treated ten patients with carcinoma in situ of the bladder (primary type, 6 and secondary type, 4) by intravesical bacillus Calmette-Guerin therapy . All patients received 8 weekly instillation, and among them 3 patients were followed by additional instillation monthly for 7 months . Complete regression (negative biopsies and cytology study) was observed after 8 weekly instillation in all patients . To elucidate the mechanism of action of BCG on carcinoma in situ, the biopsied specimens after BCG instillation were examined light and electronmicroscopically . It was speculated that BCG might act on cancer cells in two ways: one was sloughing and denudation of the cancer cells by acute tuberculous cystitis, and the other was a role of the macrophages through immune reaction . In our study, the toxicity and complications seemed to be much severer than previously reported . Vesicoureteral reflux was observed in 6 patients . Decreased bladder capacity noted in all patients, among them radical cystectomy and colocystoplasty were performed in 2 patients under the diagnosis of bladder contracture . Although complete regression was observed in all patients after 8 weekly BCG instillation, the duration being free from cancer cells was variable . And we also discussed on the additional therapy in such patients in the literature.

Appl Environ Microbiol, 1991 Jun, 57(6), 1758 - 63
Methylated sulfur compounds in microbial mats: in situ concentrations and metabolism by a colorless sulfur bacterium; Visscher PT et al.; The concentrations of the volatile organic sulfur compounds methanethiol, dimethyl disulfide, and dimethyl sulfide (DMS) and the viable population capable of DMS utilization in laminated microbial ecosystems were evaluated . Significant levels of DMS and dimethyl disulfide (maximum concentrations of 220 and 24 nmol cm3 of sediment-1, respectively) could be detected only at the top 20 mm of the microbial mat, whereas methanethiol was found only at depth horizons from 20 to 50 mm (maximum concentration of 42 nmol cm3 of sediment-1) . DMS concentrations in the surface layer doubled after cold hydrolysis of its precursor, dimethylsulfoniopropionate . Most-probable-number counts revealed 2.2 x 10(5) cells cm3 of sediment-1, in the 0- to 5-mm depth horizon, capable of growth on DMS as the sole source of energy . An obligately chemolithoautotrophic bacillus designated strain T5 was isolated from the top layer of the marine sediment . Continuous culture studies in which DMS was the growth-limiting substrate revealed a maximum specific growth rate of 0.10 h-1 and a saturation constant of 90 mumol liter-1 for aerobic growth on this substrate.

Lepr Rev, 1991 Jun, 62(2), 158 - 70
Patterns of erythropoiesis and anaemia in leprosy; Sen R et al.; A total of 128 leprosy patients were investigated for the morphological type of anaemia, the underlying disturbances in iron metabolism and patterns of erythropoiesis and other cytomorphological changes in the bone marrow . The anaemia was a mild to moderate degree in paucibacillary (PB) leprosy, while in multibacillary (MB) leprosy it was of a severe degree . Iron deficiency was observed in only a few patients . Impaired iron utilization as observed in a anaemia of a chronic disorder was a common finding in MB leprosy (41.7%) and more so in new cases (50%) . Megaloblastic erythropoiesis was also more frequent in MB leprosy (45.2%) as compared to PB leprosy (16%), accounting for the severe degree of anaemia in the former type . In 17.2% of the total patients (MB, 21.4%; PB, 9%) both megaloblastic erythropoiesis and features of impaired iron utilization were observed in bone marrow . Disturbances in iron metabolism and erythropoiesis were also observed but to a lesser degree in patients receiving specific antileprosy treatment . Irrespective of the type of disease and duration of treatment, increasing frequency of acid-fast bacillia (AFB) positivity and granulomas was observed in the bone marrow with an increasing severity of anaemia.

Plant Mol Biol, 1991 Jun, 16(6), 1035 - 50
Analysis of unstable RNA transcripts of insecticidal crystal protein genes of Bacillus thuringiensis in transgenic plants and electroporated protoplasts; Murray EE et al.; We have examined expression of several insecticidal crystal protein (ICP) genes of Bacillus thuringiensis in transgenic tobacco plants and electroporated carrot protoplasts . We determined that low levels of lepidopteran toxin cryIA(b) ICP gene expression in plants and electroporated carrot cells is due to RNA instability . We used a series of 3' deleted by cryIA(b) constructs directed by the cauliflower mosaic virus 35S promoter to demonstrate that this instability is minimally contained in the first 579 bases of the gene in both systems . This instability may result from 5'----3' as well as 3'----5' RNA metabolism . The coleopteran toxic cryIIIA gene was also examined in electroporated carrot cells, and found to be poorly expressed . A model for improvement of ICP RNA stability in plants is presented.

J Vet Med Sci, 1991 Jun, 53(3), 469 - 74
Purification and some properties of a Bacillus cereus mouse lethal toxin; Shinagawa K et al.; A mouse lethal toxin (MLT) produced by Bacillus cereus isolated in vomiting-type food poisoning was purified by chromatography on DEAE-Sephadex A-25 followed by gel filtration on Sephadex G-75 . Purified MLT possessed a molecular weight of 33,000-34,000 . It showed mouse lethality and hemolytic (HL) activity on sheep and rabbit erythrocytes; the latter erythrocytes were more weakly hemolyzed than the former ones . However, fluid accumulation in mouse ligated intestinal loops was not induced by purified MLT at the highest concentration used . Both MLT and HL activities were stable at pH 6-9, during storage at -20 degrees C for 8 weeks, and resistant to papain, cholesterol, lecithin, and dithiothreitol treatments . Most activity was lost during storage at 4 degrees C or 25 degrees C for 2 weeks or upon treatment with trypsin, trypanblue, or ethanol . The activities were resistant to heating at 37 degrees C for 5 min, less resistant at 98 degrees C for 5 min, and sensitive at 60 degrees C for 5 min . It can be concluded from the results that MLT is different from the diarrheagenic toxin produced by B . cereus isolated in diarrheal-type food poisoning, but is similar to, if not identical, hemolysin II.

J Vet Med Sci, 1991 Jun, 53(3), 419 - 22
Development of murine monoclonal antibodies against an enterotoxin produced by Bacillus cereus; Shinagawa K et al.; Three murine monoclonal antibodies (MAbs) were prepared against an enterotoxin (ET) produced by Bacillus cereus . Although these MAbs were found to react with the ET, their specificities appeared to be different in competitive binding assays . One of the MAbs (D-8), which was highly reactive with the ET, will be useful in developing immunological methods to detect crude ET and to isolate the ET in high yield.

Gig Sanit, 1991 Jun, (6), 26 - 8
{Viability of Bacillus and Pseudomonas bacteria in nitrate-polluted marine and river water}; Alton LV; Influence of nitrates on some bacterial species development was strictly depended on temperature and levels of water pollution . The bacterial activity in the river water increased with nitrate concentrations from 0.2 to 2 g/l . The concentrations of nitrates 20 g/l inhibited the bacterial development.

Rev Saude Publica, 1991 Jun, 25(3), 184 - 7
Susceptibility of Aedes aegypti larvae to temephos and Bacillus thuringiensis var israelensis in integrated control; de Andrande CF et al.; The susceptibility of field collected Aedes aegypti larvae was evaluated in terms of median lethal time (LT50) and final mortality, when treated with temephos, Bacillus thuringiensis var israelensis as well as mixtures of these two agents . Third instar larvae were shown to be more susceptible than early and late fourth instar ones to the entomopathogen . Survival of some individuals when exposed to temephos suggest possible resistance . Temporal synergism in early fourth instar larvae was detected when they were exposed to mixtures of Bti-temephos . The possibility of this integrated treatment is commented on.

Drugs, 1991 Jun, 41(6), 832 - 56
Current concepts in the pathogenesis of leprosy . Clinical, pathological, immunological and chemotherapeutic aspects; Meyers WM et al.; In recent years there have been notable advances in the laboratory investigation and field management of leprosy . Progress, however, continues to be hindered by the lack of efficient methods for early diagnosis and implementation of control and treatment measures . Diagnosis is still made on the same principles as a century ago (clinical and histopathological findings), and only 1 in 3 known patients worldwide receives optimal chemotherapy . In 1988, nearly 1 in 10 newly diagnosed patients already had debilitating deformities . Contributing factors are operational, administrative and financial difficulties in implementing multidrug therapeutic regimens, inadequately trained personnel, and lack of priority and political commitment to leprosy control . The formulation and implementation of multidrug therapy is the most important development in leprosy in the past 10 years . Dapsone monotherapy was the mainstay for treatment and control for approximately 40 years, but secondary dapsone-resistant strains, first noted in 1964, now infect as many as 50% of all new patients . Multidrug regimens recommended by the WHO consist of various combinations of therapy using dapsone, rifampicin, clofazimine and a thionamide . Duration of therapy is limited to 6 months for paucibacillary and 2 years or more for multibacillary patients; relapse rates thus far are low . The average cost of treatment worldwide, including the cost of drugs, is estimated at $US150 per patient . The recent annual drop of nearly 8% in newly registered patients may be due to the implementation of these therapeutic regimens . Newer drugs that may be introduced into these regimens include fluoroquinolones, minocycline and clarithromycin . While knowledge of the microbiology of the leprosy bacillus and host response has advanced remarkably, there is little improvement in the understanding or amelioration of social aspects of leprosy . Better treatment and control reduces the stigma, but improvements in the attitudes of patients and society towards leprosy are as important as advances in medical science in achieving ultimate eradication of the disease.

J Antibiot (Tokyo), 1991 Jun, 44(6), 579 - 81
Leuhistin, a new inhibitor of aminopeptidase M, produced by Bacillus laterosporus BMI156-14F1 . II . Structure determination of leuhistin; Yoshida S et al.; Leuhistin, a new inhibitor of aminopeptidase M, has been isolated from the culture broth of Bacillus laterosporus BMI156-14F1 . The structure of leuhistin was determined by NMR studies . X-Ray and chemical analysis confirmed the absolute structure to be (2R,3S)-3-amino-2-hydroxy-2-(1H-imidazol-4-ylmethyl)-5-methylhe xanoic acid.

J Antibiot (Tokyo), 1991 Jun, 44(6), 573 - 8
Leuhistin, a new inhibitor of aminopeptidase M, produced by Bacillus laterosporus BMI156-14F1 . I . Taxonomy, production, isolation, physico-chemical properties and biological activities; Aoyagi T et al.; Leuhistin has been isolated from the culture broth of Bacillus laterosporus BMI156-14F1 as part of a program designed to find microorganism-produced inhibitors of aminopeptidase M (AP-M) . It was purified by use of column chromatography on Sepabeads SP206, Amberlite IRC-50, MCI gel CHP-20P and Sephadex G-10 and then isolated as colorless needles . Leuhistin inhibits AP-M strongly and it also inhibits AP-A and AP-B weakly . It is competitive with the substrate, and the inhibition constant (Ki) was 2.3 x 10(-7) M.

J Bacteriol, 1991 Jun, 173(11), 3374 - 81
Deletion by in vivo recombination shows that the 28-kilodalton cytolytic polypeptide from Bacillus thuringiensis subsp . israelensis is not essential for mosquitocidal activity; Delecluse A et al.; The cytA gene encoding the 28-kDa polypeptide of Bacillus thuringiensis subsp . israelensis crystals was disrupted in the 72-MDa resident plasmid by in vivo recombination, thus indicating that homologous recombination occurs in B . thuringiensis . The absence of the 28-kDa protein in B . thuringiensis did not affect the crystallization of the other toxic components of the parasporal body (68-, 125-, and 135-kDa polypeptides) . The absence of the 28-kDa protein abolished the hemolytic activity of B . thuringiensis subsp . israelensis crystals . However, the mosquitocidal activity of the 28-kDa protein-free crystals did not differ significantly from that of the wild-type crystals when tested on Aedes aegypti and Culex pipiens larvae . The 28-kDa protein contributed slightly to the toxicity to Anopheles stephensi larvae . This indicates that the 28-kDa protein is not essential for mosquitocidal activity, at least against the three species tested.

FEMS Microbiol Lett, 1991 Jun 1, 65(1), 31 - 5
Sequence of an operon containing a novel delta-endotoxin gene from Bacillus thuringiensis; Wu D et al.; An insect larval toxin designated CryII is produced by several subspecies of Bacillus thuringiensis and differs from the other major delta-endotoxins in these bacteria in its size, toxicity profile and presence as part of an operon with three open reading frames (ORF) . Such an operon from a novel B . thuringiensis isolate has been cloned and differs from one previously characterized in the following ways: (a) the size and number of amino acid repeats in one of the ORFs; (b) the smaller size of the CryII protoxin and the presence of a unique 110-kDa CryII-related antigen; and (c) high larvicidal activity for a particular Lepidopteran but low activity for a Dipteran . Various subclones of this operon were introduced into a plasmid-free B . thuringiensis strain and only the cryII gene was found to be necessary for protoxin accumulation.

Eur J Biochem, 1991 Jun 1, 198(2), 527 - 34
The H(+)-motive and Na(+)-motive respiratory chains in Bacillus FTU subcellular vesicles; Kostyrko VA et al.; Respiration-dependent pumping of Na+ and H+ into the inside-out subcellular vesicles of alkalotolerant and halotolerant Bacillus FTU grown at alkaline pH was studied . The vesicles were shown to be competent in Na+ and H+ transport coupled to ascorbate oxidation via N,N,N',N'-tetramethyl-p-phenylenediamine or diaminodurene . The uphill Na+ uptake is strongly stimulated by either protonophores or valinomycin, whereas H+ uptake is stimulated by valinomycin and completely inhibited by protonophores . The salt of a penetrating weak base and of the penetrating weak acid, diethylammonium acetate, potentiates the stimulating effect of protonophores on Na+ uptake and abolishes H+ uptake . Na+ transport, supported by ascorbate oxidation, is resistant to 2-heptyl-4-hydroxyquinoline N-oxide, but sensitive to Ag+ and Na+ ionophore, N,N'-dibenzyl-N,N'-diphenyl-1,2-phenylenediacetamide . Micromolar concentrations of cyanide specifically inhibit the H+ uptake but does not affect Na+ uptake . These cyanide concentrations are shown to cause 70% inhibition of respiration, complete reduction of alpha-type cytochromes and partial reduction of c/b-type cytochromes . To inhibit the remaining respiratory activity and Na/ uptake, approximately 100-fold higher cyanide concentrations are necessary . High cyanide concentrations cause some additional increase in absorbance in the region of cytochromes c and/or b . In the presence of a high cyanide concentration, Na+ uptake can be supported by NADH oxidation by fumarate . This Na+ transport is stimulated by protonophores and diethylammonium acetate, being sensitive to very low concentrations of 2-heptyl-4-hydroxyquinoline N-oxide and Ag+ . The NADH-fumarate reductase reaction is also found to be competent in H+ uptake, which is inhibited by protonophores and by much higher 2-heptyl-4-hydroxyquinoline N-oxide concentrations, and is resistant to Ag+ . It is inferred that Bacillus FTU possesses two respiratory chains: the H(+)-motive and the Na(+)-motive, which strongly differ in their inhibitor sensitivities . Each chain comprises at least two energy-coupling sites which are localized in their initial and terminal segments . It has been indicated that common redox carrier(s) are present in the two chains.

Cytotechnology, 1991 Jun, 6(2), 115 - 20
Immunoglobulin production stimulating factor-II alpha (IPSF-II alpha) is glyceraldehyde-3-phosphate dehydrogenase like protein; Sugahara T et al.; Amino acid sequence of the 36 KD protein which is the active subunit of immunoglobulin production stimulating factor-II alpha (IPSF-II alpha) derived from Burkitt's lymphoma Namalwa cells was analyzed for the 20 amino acids from N-terminus . The N-terminal amino acid sequence of this protein coincided very closely with glyceraldehyde-3-phosphate dehydrogenase (GPD; EC 1.2.1.12) derived from various origins . Especially, it was completely homologous with that of human liver GPD . Several GPD's derived from human erythrocyte, rabbit muscle and Bacillus stearothermophilus also stimulated IgM production of hybridomas, as well as IPSF-II alpha . Conversely, IPSF-II alpha had GPD enzymic activity as strong as rabbit muscle and B . stearothermophilus, and stronger than human erythrocyte GPD . These results suggested that 36 KD subunit of IPSF-II alpha was a GPD, or GPD like protein . The level of mRNA for IgM was not enhanced by IPSF-II alpha in hybridoma cells, though the IgM productivity of the cell was remarkably stimulated by the protein, indicating that IPSF-II alpha does not stimulate immunoglobulin production by enhancement of transcription.

Biochim Biophys Acta, 1991 May 30, 1078(1), 68 - 76
Purification and properties of two forms of ATP sulfurylase from Euglena; Li JJ et al.; Two forms of ATP sulfurylase have been purified to homogeneity from mitochondria (ATPSm) and cells (ATPSc) of Euglena gracilis Klebs var . bacillaris Cori (aplastidic mutant W10BSmL) . Both forms are monomeric, ATPSc is 52.3 kDa and ATPSm is 55 kDa . The pI is 7.9 for ATPSc and 5.8 for ATPSm . Therefore, ATPSm binds to DEAE-cellulose at pH 7.4; ATPSc does not . After cleavage by CNBr, the two forms of ATP sulfurylase show different sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) patterns, suggesting that they differ in amino acid sequence . ATPSm is mainly associated with the mitochondrial membrane and ATPSc is mainly soluble in the cells . Both enzymes require similar conditions in the molybdolysis assay, but show different pH optima when sulfate is used as substrate . ATPSc is more sensitive to adenosine 5'-phosphosulfate (APS) inhibition than ATPSm in the SO2-4 incorporation reaction . In the reverse reaction, ATPSc requires much higher concentrations of PPi and MgCl2 to saturate the reaction than ATPSm . The data indicate that the two enzymes are quite distinct and may have different roles in cell metabolism.

Biochemistry, 1991 May 28, 30(21), 5151 - 6
Kinetic and thermodynamic properties of wild-type and engineered mutants of tyrosyl-tRNA synthetase analyzed by pyrophosphate-exchange kinetics; Wells TN et al.; The first step of the reaction catalyzed by the aminoacyl-tRNA synthetases is the formation of enzyme-bound aminoacyl adenylate . The steady-state kinetics of this step has conventionally been studied by measuring the rate of isotopic exchange between pyrophosphate and ATP . A simple kinetic analysis of the pyrophosphate-exchange reaction catalyzed by the tyrosyl-tRNA synthetase from Bacillus stearothermophilus is given in which all the observed rate and binding constants can be assigned to identifiable physical processes under a variety of limiting conditions . The free energies of binding to the enzyme of tyrosine, ATP, and the transition state for tyrosyl adenylate formation can be measured in relatively straightforward experiments . The excellent agreement between parameters measured in these experiments and those from earlier pre-steady-state kinetics confirms that the intermediates isolated in the presteady state are kinetically competent . The dissociation constant of ATP from the unligated enzyme, a constant that has previously been experimentally inaccessible, has been measured for wild-type and several mutant enzymes . The changes in enthalpy and entropy of activation on mutation have been measured by a rapid procedure for mutants that have altered contacts with tyrosine and ATP . Those mutants that have large changes of enthalpy and entropy of binding are likely to have structural changes and so warrant further examination by protein crystallography.

Nucleic Acids Res, 1991 May 25, 19(10), 2615 - 21
An analysis of the sequence of an infectious clone of rice tungro bacilliform virus, a plant pararetrovirus; Hay JM et al.; The nucleotide sequence of an infectious clone of rice tungro bacilliform virus (RTBV) DNA has been determined . The circular genome has 8002 bp and one strand contains four open reading frames (ORFs) . One ORF is potentially capable of encoding a protein of 24 kD (P24) and has no initiation (ATG) codon . The other three ORFs potentially encode proteins of 12 kD, 194 kD and 46 kD (P12, P194, P46) respectively . The functions of P24, P12 and P46 are unknown . Comparative analyses with retroviruses and Commelina yellow mottle virus suggest that the 194 kD putative product is a polyprotein that is proteolytically cleaved to yield the virion coat protein, a protease and replicase (reverse transcriptase and RNase H) characteristic of retroelements . The DNA sequence reveals other features which strongly support our belief that RTBV is a pararetrovirus . These include sequences at the mapped positions of two discontinuities in the virion DNA which are complementary to tRNA metinit and purine-rich, and may be the priming sites for minus- and plus-strand DNA synthesis respectively . As the positions of likely transcriptional signals suggest, a full-length viral transcript is observed by northern analysis . The predicted folding of the 645 bp 5'-region of this RNA resembles that of caulimoviruses . Comparisons with other reverse transcribing elements are discussed.

Biochim Biophys Acta, 1991 May 23, 1058(1), 52 - 5
Two structurally different cytochromes c from Bacillus azotoformans: on the evolution of gram-positive bacteria; Hreggvidsson GO; c-552 and split-alpha c-555 cytochromes from Bacillus azotoformans are classified on the basis of partial sequence information . The haem-containing polypeptides are postulated to be structurally equivalent to small IC and ID subclass cytochromes found in purple bacteria.

Biochem J, 1991 May 15, 276 ( Pt 1), 129 - 33
A continuous fluorescence-displacement assay for triacylglycerol lipase and phospholipase C that also allows the measurement of acylglycerols; Wilton DC; A new continuous fluorescence-displacement assay for enzymes that release long-chain fatty acids {Wilton (1990) Biochem . J . 266, 435-439} is described in detail for pig pancreatic triacylglycerol lipase . The assay involves the displacement of the highly fluorescent fatty acid probe 11-(dansylamino)undecanoic acid from rat liver fatty acid-binding protein by long-chain fatty acids released as a result of enzyme activity . The assay is surprisingly effective for triacylglycerol lipase, allowing the expression of full activity with low concentrations of substrates in the absence of detergents . The initial rate of decrease in fluorescence is linearly related to enzyme concentration, and activity can be detected in the assay down to concentrations of 10 pg of pure enzyme/ml . The assays demonstrated the quantitative conversion of limiting amounts of substrate into the monacylglycerol . This observation allowed the assay to be used to measure substrates such as triacylglycerols and particularly 1,2-diacylglycerols at concentrations down to about 0.1 microM . Because phospholipase C releases 1,2-diacylglycerols, the coupling of this enzyme to excess lipase allowed the measurement of pure phospholipase C from Bacillus cereus at concentrations down to about 2 ng/ml, and the initial rate of fall in fluorescence in the assay was linearly related to enzyme activity.

J Natl Cancer Inst, 1991 May 15, 83(10), 682 - 94
Adjuvant intravesicular pharmacotherapy for superficial bladder cancer; Lum BL et al.; In 1990, bladder cancer, excluding carcinoma in situ, was estimated to contribute 49,000 cases of cancer . In men 75 years old or older, it became the fifth leading cause of cancer deaths . Of patients with bladder cancer, 75%-80% initially present with superficial bladder tumors . Treatment of these tumors has three objectives: 1) to eradicate existing disease, 2) to provide prophylaxis against tumor recurrence, and 3) to avoid deep invasion into the muscle layers of the bladder . Transurethral resection is the primary treatment to eradicate superficial bladder tumors, but 40%-80% of these tumors recur . Because of these high recurrence rates, adjuvant intravesicular pharmacotherapy with cytotoxic and immunomodulatory drugs has gained widespread use . The past two decades of clinical investigations in superficial bladder cancer have provided valuable information on the biology and treatment of the disease . Multivariate analyses have indicated that tumor grade and stage are the most important prognostic variables commonly available to the clinician to identify the patient at greatest risk of developing muscle-invasive or metastatic bladder cancer . These studies have also identified groups at low risk for tumor recurrence and invasive bladder cancer . Randomized trials have shown that recurrence rates are decreased by adjuvant intravesicular pharmacotherapy with a number of drugs: bacillus Calmette-Guerin vaccine (BCG), doxorubicin, ethoglucid (Epodyl), mitomycin-C, teniposide, and thiotepa . However, few studies indicate that adjuvant intravesicular pharmacotherapy can prevent progression to invasive bladder cancer in the high-risk patient with superficial bladder cancer . Additional clinical trials are needed to determine whether such therapy can prevent invasive and metastatic bladder cancer and improve disease-free survival in this group . In addition, the identification of tests (e.g., monoclonal antibody tests, chromosomal analyses, and tumor marker assays) that can help to identify high-risk patients is needed to better develop therapeutic strategies for superficial bladder cancer.

J Mol Biol, 1991 May 5, 219(1), 123 - 32
Crystal structure of a barnase-d(GpC) complex at 1.9 A resolution; Baudet S et al.; The ribonuclease excreted by Bacillus amyloliquefaciens, Barnase, was co-crystallized with the deoxy-dinucleotide d(GpC) . The crystal structure was determined by molecular replacement from a model of free Barnase previously derived by Mauguen et al . Refinement was carried out using data to 1.9 A resolution . The final model, which has a crystallographic R factor of 22%, includes 869 protein atoms, 38 atoms from d(GpC), a sulfate ion and 73 water molecules . Only minor differences from free Barnase are seen in the protein moiety, the root-mean-square C alpha movement being 0.45 A . The dinucleotide has a folded conformation . It is located near the active site of the enzyme, but outside the protein molecule and making crystal packing contacts with neighboring molecules . The guanine base is stacked on the imidazole ring of active site His102, rather than binding to the so-called recognition loop as it does in other complexes of guanine nucleotides with microbial nucleases . The deoxyguanosine is syn, with the sugar ring in C-2'-endo conformation; the deoxycytidine is anti and C-4'-exo . In addition to the stacking interaction, His102 hydrogen bonds to the free 5' hydroxyl, which is located near the position where the 3' phosphate group is found in other inhibitors of microbial ribonucleases . While the mode of binding observed with d(GpC) and Barnase would be non-productive for a dinucleotide substrate, it may define a site for the nucleotide product on the 3' side of the hydrolyzed bond.

Pediatr Infect Dis J, 1991 May, 10(5), 359 - 65
Protective efficacy of neonatal Bacillus Calmette-Guérin vaccination against tuberculosis; Sirinavin S et al.; This matched case-control study was performed in two hospitals in Bangkok to evaluate the protective efficacy of neonatal Bacillus Calmette-Guerin (BCG) vaccination in Thai children and to find factors that might explain the reported variation in estimates of the protective efficacy of BCG . Cases were defined as children 3 months to 14 years of age who had tuberculosis and controls consisted of one to four children who were born in the same year and had the same district of residence as the case . A matched analysis with a variable number of controls per case was performed on 75 cases and 207 controls . Conditional logistic regression was performed to adjust for the potential confounding effects of household tuberculosis exposure and socioeconomic status . Forty-eight percent of cases had nonrespiratory tuberculosis . Laboratory-confirmed evidence for tuberculosis was found in 48% of cases . The adjusted protective efficacy of neonatal BCG vaccination was 83% (95% confidence limits, 35%, 96%) . It was 96% (95% confidence limits, 66%, 100%) when only 36 matched sets of laboratory-confirmed cases were analyzed . Subgroup analyses and literature reviews indicated that the accuracy of tuberculosis diagnosis, types of tuberculosis, duration after vaccination and household tuberculosis exposure contribute to variation in the reported protective efficacy of neonatal BCG vaccination.

J Invertebr Pathol, 1991 May, 57(3), 343 - 51
Photoprotection of Bacillus thuringiensis kurstaki from ultraviolet irradiation; Cohen E et al.; Irradiation of Bacillus thuringiensis var . kurstaki HD1 at 300-350 nm for up to 12 hr using a photochemical reactor results in a rapid loss of its toxicity to larvae of Heliothis armigera . Photoprotection of the toxic component was obtained by adsorption of cationic chromophores such as acriflavin (AF), methyl green, and rhodamine B to B . thuringiensis . AF gave the best photoprotection and a level of 0.42 mmol/g dye absorbed per gram of B . thuringiensis was highly toxic even after 12 hr of ultraviolet (uv) irradiation as compared to the control (77.5 and 5% of insect mortality, respectively) . Ultraviolet and Fourier-transform infrared spectroscopic studies indicate molecular interactions between B . thuringiensis and AF . The nature of these interactions and energy or charge transfer as possible mechanisms of photoprotection are discussed . It is speculated that tryptophan residues are essential for the toxic effect of B . thuringiensis . It is suggested that photoprotection is attained as energy is transferred from the excited tryptophan moieties to the chromophore molecules.

J Invertebr Pathol, 1991 May, 57(3), 325 - 33
Isolation and identification of Bacillus sphaericus strains pathogenic for mosquito larvae; Guerineau M et al.; Three selective media for the isolation of Bacillus sphaericus have been compared . BATS medium and a formulation employing adenosine as the principal carbon source were the most effective for the recovery of spores of strain 1593 . Anthranilic acid as the principal carbon source was less efficient . Eighty-four strains were isolated from mud samples using these media and were identified by computer . Identifications were confirmed for representative strains using DNA sequence homology . Most were B . sphaericus sensu stricto or members of an unnamed group . However, one strain (BSE 18) was identified as the DNA homology group IIB and this organism was found to be highly toxic toward larvae of Culex pipiens . Southern hybridization of BSE 18 DNA to a probe prepared from the cloned toxin gene from strain 1593 revealed that BSE 18 contained a typical gene for the 41.9-kDa toxin.

Public Health, 1991 May, 105(3), 229 - 38
Primary health care and immunisation in Iran; Nasseri K et al.; The Primary Health Care (PHC) network of Iran consists of a rural and an urban branch . While the rural branch presently covers a sizeable portion of the rural population, the urban PHC project is in its early stages of implementation . The Expanded Programme on Immunisation (EPI) in Iran, which started as an independent and vertical project in early 1983, is being gradually integrated into the PHC network as the latter expands . Results of the second PHC programme review of Iran shows that immunisation coverage of children has improved appreciably since the first PHC review, especially for BCG which stands at 56.3% . Complete immunisation at first birthday in the rural areas with the PHC services is 44.1%, whereas for urban areas other than Teheran it is 28.1% . While the high coverage in the rural areas is attributed to the 'active' approach and vigilance of the providers of immunisation (i.e . the community health workers and the vaccinators of the mobile teams), the higher coverage in the capital city of Teheran is attributed to the involvement of private paediatricians and the generally higher social, economic, and educational status as well as higher interest of mothers . It is noticed that the results of cluster sampling for determination of immunisation coverage in large metropolitan areas of the developing world must be interpreted with much care . The reason is that in these areas extreme fluctuations in the crude birth rate are common and therefore results tend to over-represent the attributes of the segment of population with lower birth rate . It is also argued that complete immunisation might not be the best indicator for assessing the progress of the immunisation efforts . These and other findings are discussed in detail . are discussed in detailPIP: Childhood immunization in Iran was assessed by a WHO EPI (World Health Organization, Expanded Program on Immunization) cluster survey method covering 2118 children aged 12-23 months in 1987 . Complete immunization was defined as a minimum of 3 DPT (diphtheria, pertussis, tetanus), 3 OPV (oral polio vaccine), 1 BCG (Bacillus Calmette-Guerin), and 1 live attenuated measles vaccine by age 1 year . Iran's Primary Health Care system consists of a rural branch operated by mobile male and female teams, and an urban branch still in the process of changing from cure-oriented care to emphasis on health education, nutrition, and maternal-child health services . Complete immunization coverage by age 1 was better in rural areas, 44.1%, than in urban areas, 28.2%, and Teheran 34.9% . There was no relationship between immunization coverage and infant mortality rate, which is dominated by diarrhea . The reason for better coverage in rural areas is that village workers actively search out, visit and immunize children, while in urban areas provide physicians dominate care, but do not insist on immunization . Furthermore, in Teheran, BCG is not routinely given, which lowered the overall immunization coverage rate there .

Sci China B, 1991 May, 34(5), 593 - 8
Location of 43 kD mosquito-larvicidal toxin gene of highly toxic Bacillus sphaericus 10; Liu Y et al.; The direct evidence of the location of the mosquito-larvicidal gene of Bacillus sphaericus 10 (isolated from Jiangsu Province of China) on megaplasmid pFW1 was given by molecular cloning . The clone (pFL109) containing the 1.4 kb HindIII DNA fragment from pFW1 expressed the mosquito-larvicidal toxin protein (43 kD) . The location of the gene coding for the 43 kD toxin protein within the XhoI B fragment on the restriction map of pFW1 was confirmed by Southern blotting using the 1.4 kb HindIII DNA fragment as a probe . The non-toxic strains of Bacillus sphaericus were revealed to be 43 kD toxin gene deletion mutants by Southern and Western analyses . The 1.4 kb HindIII fragment of pFL109 can be used as a probe for differentiating the non-toxic strains of Bacillus sphaericus from the toxic ones.

J Clin Microbiol, 1991 May, 29(5), 906 - 10
Application of a polymerase chain reaction for the detection of Mycobacterium leprae in skin tissues; de Wit MY et al.; The polymerase chain reaction (PCR) based on the selective amplification of a 530-bp fragment of the gene encoding the proline-rich antigen of Mycobacterium leprae was applied on sections of fixed or frozen biopsy samples from leprosy patients . A simple procedure for the extraction of DNA from M . leprae in clinical specimens that provided suitable template DNA for amplification was developed . When PCR was applied on frozen sections, positive amplification in samples from all untreated acid-fast bacillus (AFB)-positive patients and in samples from 56% of the untreated AFB-negative patients could be detected, while biopsy samples from patients with skin diseases other than leprosy were all PCR negative . With neutral Formalin-fixed biopsy samples, positive amplification in 92% of the samples from untreated AFB-positive patients and in 61% of the samples from untreated AFB-negative patients could be detected by PCR . Biopsy samples exposed to mercuric chloride or nonbuffered formaldehyde containing fixatives were not suitable for application of PCR . This PCR holds promise as a tool for studies on M . leprae infection.

Mol Gen Genet, 1991 May, 227(1), 60 - 4
In vivo transcriptional pattern in the infC operon of Bacillus stearothermophilus; Falconi M et al.; By Northern blot and primer extension analyses it was shown that in Bacillus stearothermophilus the genes infC, rpmI and rplT constitute a single transcriptional unit; the promoter and the transcriptional start-point used in vivo were identified and the half-life of the transcript (1.2 min) was determined . No indication of multiple initiation sites nor of differential stability of different regions of the transcript was found . The results suggest that Escherichia coli and B . stearothermophilus have a different pattern of transcription around the infC gene cluster and that differential translational efficiency within the infC-rpmI-rplT transcriptional unit accounts for the different levels of IF3, L35 and L20 expression . The rare AUU initiation codon is the only strictly conserved element of the several peculiar transcriptional and translational features found in E . coli infC . Upon changing this codon to AUG, we found a ca . 30-fold increased expression of B . stearothermophilus infC, which is similar to that previously found with E . coli infC (i.e . 40-fold), and provided evidence that regulation of infC expression through its rare AUU initiation codon might be a general phenomenon in bacteria.

J Clin Oncol, 1991 May, 9(5), 729 - 35
Improved survival in patients with poor-prognosis malignant melanoma treated with adjuvant levamisole: a phase III study by the National Cancer Institute of Canada Clinical Trials Group; Quirt IC et al.; Five hundred forty-three patients with completely resected malignant melanoma who were considered to have a significant risk of developing recurrent disease were randomized to one of four study groups . One group received levamisole 2.5 mg/kg on 2 consecutive days weekly for 3 years, a second group received bacillus Calmette-Guerin (BCG) for 3 years . A third group alternated 8-week courses of BCG and levamisole for 3 years and a fourth group underwent clinical assessment at the same frequency as the three treatment groups . The median duration of follow-up is 8.5 years . The percentage of reduction in the death rate and the recurrence rate in the treatment groups compared with the control group was calculated using the Cox proportional hazards model and adjusted for age, sex, and stage as covariants . The patients treated with levamisole were estimated to have a 29% reduction in both the death rate (P = .08) and the recurrence rate (P = .09) compared with patients receiving no further treatment . Fifty-five patients discontinued levamisole early because of gastrointestinal intolerance or arthralgia, myalgia, fever, and immune leukopenia . The patients treated with BCG alternating with levamisole experienced a 10% reduction in the death rate and a 6% reduction in the recurrence rate, and the patients treated with BCG alone experienced a 4% reduction in the death rate and a 3% increase in the recurrence rate compared with the control group . The degree of improvement experienced by the patients that were treated by levamisole is of sufficient magnitude to warrant further investigation of this dose of levamisole as adjuvant treatment in patients with melanoma.

J Biochem (Tokyo), 1991 May, 109(5), 763 - 9
Characterization and location of the L-proline activating fragment from the multifunctional gramicidin S synthetase 2; Kurotsu T et al.; Gramicidin S synthetase 2 (GS2) derived from Bacillus brevis is a multifunctional single polypeptide (Mr 280,000) with a 4'-phosphopantetheine residue covalently bound to the enzyme . When GS2 was treated with trypsin or chymotrypsin, fragments with some activity were liberated . The molecular mass of the L-proline activating fragment was 114 kDa on SDS-PAGE . This fragment, when incubated with gramicidin S synthetase 1 (GS1) in the presence of phenylalanine and proline, produced D-Phe-L-Pro dipeptide . The fragment accepted D-phenylalanine from GS1 in the absence of L-proline . The L-proline activating fragment was shown to lack pantothenic acid by microbiological assay . On the other hand, the L-leucine activating fragment, which was partially purified, contained a large amount of pantothenic acid, although it did not form the D-Phe-L-Pro dipeptide . These results indicate that the L-proline activating site is located near an acceptor site for D-phenylalanine on GS2, but that it is not adjacent to a 4'-phosphopantetheine group . The N-terminal sequence (15 amino acid residues) of the L-proline activating fragment obtained by trypsin treatment was identical with that of GS2, indicating that the L-proline activating site is located at the N-terminus of the native synthetase . The N-terminal sequence of GS2 has been matched with the amino acid sequence deduced from the nucleotide sequence 71 bp downstream of the stop codon of the GS1 gene except that the first initiator methionine was not detected.

J Biochem (Tokyo), 1991 May, 109(5), 678 - 83
Nucleotide sequence of the gene encoding NADH dehydrogenase from an alkalophile, Bacillus sp . strain YN-1; Xu XM et al.; The gene encoding NADH dehydrogenase from an alkalophile, Bacillus sp., was cloned and sequenced . The cloned DNA fragment contained an open reading frame of 1,557 nucleotides which encodes a polypeptide composed of 519 amino acid residues (Mr 55,830) . The predicted amino acid sequence was consistent with the partial amino acid sequences including the N-terminal and C-terminal sequences determined in a previous study . Sequence comparison with other flavoenzymes revealed high homology between the present dehydrogenase and Escherichia coli thioredoxin reductase.

Biull Eksp Biol Med, 1991 May, 111(5), 525 - 8
{Determination of antibodies to cross-reacting antigens of microorganisms in oncological diseases}; Autenshlius AI et al.; It was studied antibodies to saprophytic microorganism Bacillus megaterium H glycoprotein in healthy, oncological, non-oncological gastrointestinal patients and animals . High level of antibodies to microbial glycoprotein in blood sera was revealed in cancer and precancer patients . Analogical results were obtained in mice A/Sn and Balb/c with inductive or transplanted tumors . It has been suggested the use of Bacillus megaterium H glycoprotein (M . m . 65-70 kD) for immunological monitoring.

FEMS Microbiol Lett, 1991 May 1, 64(1), 1 - 5
Improved methods for purification of an enterotoxin produced by Bacillus cereus; Shinagawa K et al.; An enterotoxin (ET) produced by Bacillus cereus was purified by ammonium sulfate precipitation followed by chromatography on SP-Sephadex C-25, chromatofocusing, and gel filtration on Sephacryl S-300 . Purified ET was electrophoretically homogeneous with a molecular mass of 45,000 and with an isoelectric point of 5.5 . It showed vascular permeability activity in rabbits, was lethal to mice, and caused fluid accumulation in mouse ligated intestinal loops . It showed cytotoxicity to Vero cells, but did not show any hemolytic or phospholipase C activities . These biological activities were found to be easily inactivated by heating at 56 degrees C for 5 min, by digestion with trypsin and pepsin, and by exposure to pH below 3.0 or higher than 11.

Mol Gen Genet, 1991 May, 227(1), 137 - 43
Preprosubtilisin Carlsberg processing and secretion is blocked after deletion of amino acids 97-101 in the mature part of the enzyme; Schulein R et al.; During an investigation into the substrate specificity and processing of subtilisin Carlsberg from Bacillus licheniformis, two major independent findings were made: (i) as has been shown previously, a stretch of five amino acids (residues 97-101 of the mature enzyme) that loops out into the binding cleft is involved in substrate binding by subtilisin Carlsberg . In order to see whether this loop element also determines substrate specificity, the coding region for these five amino acids was deleted from the cloned gene for subtilisin Carlsberg by site-directed mutagenesis . Unexpectedly the resulting mutant preproenzyme (P42c, Mr = 42 kDa) was not processed to the mature form (Mr = 30 kDa) and was not released into the medium by a protease-deficient B . subtilis host strain; rather, it accumulated in the cell membrane . This result demonstrates that the integrity of this loop element, which is very distant from the processing cleavage sites in the preproenzyme, is required for secretion of subtilisin Carlsberg . (ii) In culture supernatants from B . subtilis harbouring the cloned wild-type subtilisin Carlsberg gene the transient appearance (at 0-3 h after onset of stationary phase) of a processing intermediate (P38c, Mr = 38 kDa) of this protease could be demonstrated . P38c very probably represents a genuine proform of subtilisin Carlsberg.

Biochem J, 1991 May 1, 275 ( Pt 3), 545 - 53
Envelope alteration of Escherichia coli HB101 carrying pEAP31 caused by Kil peptide and its involvement in the extracellular release of periplasmic penicillinase from an alkaliphilic Bacillus; Aono R; The plasmid pEAP31 contains the colicin E1 kil gene . Peptidoglycan and outer-membrane components (lipopoly-saccharide, proteins and phosphatidylethanolamine) decreased concurrently in the envelope fraction from Escherichia coli HB101 carrying pEAP31 during the stationary phase of growth . At almost the same time . D-alanine residues in peptidoglycan decreased . The Kil peptide is suggested to affect, directly or indirectly, the turnover of peptidoglycan in stationary phase and, as a result, to cause partial exfoliation of the outer membrane . Periplasmic proteins are liberated from E . coli HB101 (pEAP31) probably because of the exfoliation of outer membrane.

Infect Immun, 1991 May, 59(5), 1778 - 84
Characterization of the components of hemolysin BL from Bacillus cereus; Beecher DJ et al.; Previously we described the partial purification of a novel hemolysin from Bacillus cereus and showed that hemolytic activity required the combined action of at least two components, called B and L to signify their cell-binding and cell-lytic roles in this activity . On further purification, as described in the present article, a combination of anion-exchange chromatography and polyacrylamide gel electrophoresis separated three proteins, B, L1, and L2 (35, 36, and 45 kDa, respectively) . Individually, these proteins were inactive in hemolytic and vascular permeability assays, and combinations of B and L1 or B and L2 were either inactive or slightly active . Combinations of all three moieties produced the unique ring-shaped zone of hemolysis, a previously described characteristic of hemolysin BL, as well as edema and bluing in the vascular permeability assay . Since the vascular permeability assay is known to correlate with enterotoxicity, these results suggest that hemolysin BL is enterotoxigenic . Furthermore, the molecular weights and isoelectric point values of the hemolysin BL components are consistent with those described by others for the multicomponent diarrheal enterotoxin of B . cereus . Immunofluorescent staining of B-treated erythrocytes confirmed that B binds to cells as an initial step required before the L components can act to cause cell lysis.

Mol Gen Mikrobiol Virusol, 1991 May, (5), 29 - 31
{Genetic properties of the nonflagellar Bacillus thuringiensis mutant}; Bogdanova TL et al.; The nonflagellar mutant has been selected in Bacillus thuringiensis strain 33-69 var . galleriae . The absence of flagella in the mutant cells is confirmed by electron microscopy, by the specific "trench transit" test . The mutant preserved the biochemical characteristics of the parent strain, possesses the same phage sensitivity but lower sporulation ability . Fla+ phenotype is cotransduced with the His+ one in general transduction.

Emerg Med Clin North Am, 1991 May, 9(2), 327 - 34
Cat-scratch disease; Roberge RJ; Cat-scratch disease is a relatively common disorder that is the result of infection with a recently identified gram-negative bacillus . Although it is overwhelmingly a benign, self-limited illness requiring only supportive care, serious complications can occur and the disease may be life-threatening on occasion . Although randomized clinical trials of antibiotic therapy for CSD do not exist, CSD has demonstrated in vitro susceptibility to several antibiotics, and gentamicin has been reported beneficial in a few patients with systemic manifestations of the illness . Therefore, pending clinical trials, gentamicin may be worthy of consideration in CSD patients with serious manifestations of this illness . CSD is one of the most common causes of adenopathy and should be considered in all patients who present with unilateral or regional lymphadenopathy.

Anticancer Res, 1991 May-Jun, 11(3), 1253 - 8
Intravesical Bacillus Calmette-Guérin treatment for superficial bladder cancer: results after 15 years of experience; van der Meijden AP et al.; Superficial bladder cancer has a high recurrence rate and considerable progression rate . The treatment currently consists of resection and adjuvant intravesical chemotherapy or immunotherapy . Intravesical instillation of Bacillus Calmette-Guerin (B.C.G.) is considered to be one of the most successful immunotherapies in man . Durable response rates of 60-70% are achieved . Toxicity is more pronounced in comparison with intravesical chemotherapy . In this article we describe the experience with B.C.G . during the last 15 years . No consensus has been reached yet about the ideal treatment scheme, appropriate strain and optimal dosage . The mechanisms of action are complicated and still not completely understood.

Eur J Epidemiol, 1991 May, 7(3), 291 - 3
Rickettsiae and rickettsioses in Portugal; Bacellar F et al.; The only rickettsiae recorded in Portugal till now were Rickettsia conorii and Coxiella burnetii . Boutonneuse fever is one of the most important transmissible diseases in Portugal . Though the annual number of cases is not exactly known, it is estimated to be not far from 20,000 in some years . Q fever is the other rickettsiosis widely disseminated throughout the country . The serological prevalence and the incidence of those rickettsioses in Portugal are presented in this communication . In recent research in southern Portugal, about 4,000 adult ticks of nine species were screened by the haemocyte test for rickettsiae and rickettsia-like organisms (RLO) . In addition to R . conorii three microscopically different RLO were observed . Two of them, i.e . ovoid and bacillary-like, were positive in the immunofluorescence test with spotted fever (R . conorii) antiserum . The first occurred mainly in Rhipicephalus sanguineus ticks, the second one also in other tick species . The latter agent was cultivated in half-engorged R . sanguineus females and in Vero cells . The third organism was found in R . sanguineus, where it exhibited a massive infestation in haemocytes resembling that seen in experimentally infected ticks with C . burnettii, but not being this agent . The investigation of the isolates and their identification and characterization are being continued.

Arch Histol Cytol, 1991 May, 54(2), 145 - 61
The ultrastructure of human cumulus-corona cells at the time of fertilization and early embryogenesis . A scanning and transmission electron microscopic study in an in vitro fertilization program; Nottola SA et al.; We observed the ultrastructure of the cumulus-corona cells (CC cells) surrounding: 1) human preovulatory oocytes unfertilized after in vitro insemination and 2) in vitro-fertilized polypronuclear ova (PO) at the pronuclear stage (3 pronuclei) and during early cleavage, at the 3-8 cell stage (cleaving PO) . All the samples were obtained from women who underwent pharmacological hormonal stimulation during in vitro-fertilization procedures . Both cell groups were composed of irregularly rounded CC cells, showing an oval nucleus with one or more reticular nucleoli . Spermatozoa in close contact with CC cells were also seen . Linear and annular gap junctions between neighbouring cells were present, particularly in Group 1 . Lipid droplets were present in both groups, appearing slightly more numerous and electron-dense in Group 2 . In Group 1, mitochondria were numerous, polymorphic, and provided with cristae varying from lamellar to tubular . In Group 2, mitochondria also showed polymorphism, with bacilliform organelles with tubular cristae being predominant . In both groups cisternae and associated vacuoles and vesicles belonging to the Golgi complex were scattered in the cytoplasm of CC cells . Similarly, tubular and vesicular profiles of smooth endoplasmic reticulum were abundant and uniformly distributed throughout the cytoplasm of CC cells of both groups . In contrast, the abundant rough endoplasmic reticulum in Group 1 was formed by parallel stacks of flattened cisternae, whereas it was less plentiful and not arranged in stacks in Group 2 . The CC-cell surface appeared covered by numerous membrane expansions in both groups . The expansions in Group 1 were mainly composed of blebs of various sizes and a few short microvilli, whereas in Group 2 numerous microvilli covered the cell surface . These observations demonstrate that a gradual establishment and maintainance of a steroidosynthetic capability (luteinization) takes place in CC cells, particularly during and shortly after fertilization, as occurs contemporaneously in the granulosa cells of the postovulatory follicle . Our results may be considered as ultrastructural confirmation of the capability of the CC cells to produce small amounts of steroids (estrogens and mainly progesterone) . These hormones, alone or together with other substances (proteins, nutrients, growth factors?), might--around the fertilization time--act positively upon the early embryo itself, as well as on the microenvironment in which the embryo develops, both in vivo and in vitro conditions.

Yeast, 1991 May-Jun, 7(4), 337 - 46
Optimization of Bacillus alpha-amylase production by Saccharomyces cerevisiae; Ruohonen L et al.; Production of Bacillus amyloliquefaciens alpha-amylase by Saccharomyces cerevisiae using the multicopy plasmid pAAH5 and ways of improving the yields of secreted enzyme were studied . In standard non-buffered medium, alpha-amylase was rapidly inactivated but stabilization of the pH at 6 led to stable accumulation of alpha-amylase in the culture medium . Removal of 1100 bp of the upstream sequence of the ADH1 promoter present on pAAH5 resulted in delayed but increased alpha-amylase production: 29-fold in selective medium, two-fold in non-selective medium . With the original ADH1 promoter, accumulation of alpha-amylase in the medium started to level off before the cultures reached stationary phase and was very low when exponentially growing cells were transferred from glucose to ethanol . This coincided with the appearance of a mRNA larger than the alpha-amylase messenger . With the shortened promoter, the normal-size alpha-amylase mRNA was detected under all growth conditions and alpha-amylase was efficiently secreted into the medium also late in stationary phase and after transfer to ethanol . Highest total yields of alpha-amylase were obtained with the short promoter in non-selective glucose-containing medium; this may be explained by the greater final cell density obtained . However, the production of alpha-amylase per cell mass was higher in ethanol-containing selective medium . Seventy to eighty per cent of the alpha-amylase activity was secreted into the medium independent of the total amount produced.

Br Poult Sci, 1991 May, 32(2), 399 - 404
Pharmacotoxicological aspects of nitrate and nitrite in domestic fowls; Atef M et al.; 1 . The effects of nitrates and nitrites on growth, erythrocytic count, liver and kidney functions, humoral and cell mediated immune responses in cockerels were investigated . 2 . Sodium nitrate (4.2 g/kg diet) and sodium nitrite (1.7 g/kg) retarded growth, caused methaemoglobinaemia and changes in erythrocytic count, serum concentrations of glutamic-pyruvic transaminase, creatinine and urea . 3 . Cockerels given nitrate or nitrite in the food had reduced haemagglutination responses after injection of sheep erythrocytes and a reduced delayed hypersensitivity reaction to purified protein derivative tuberculin following sensitisation to Bacille Calmette Guerin . 4 . Nitrates and nitrites are environmental pollutants present in food and water and they may contribute to the aetiology of liver and kidney diseases and problems related to failure of immunity in domestic fowls.

Int J Food Microbiol, 1991 May, 13(1), 47 - 54
Effect of surface sterilization, fumigation and gamma irradiation on the microflora and germination of barley seeds; Ramakrishna N et al.; Sodium hypochlorite (NaOCl) and mercuric chloride (HgCl2) surface sterilization, methyl bromide and propylene oxide fumigation and gamma irradiation treatments were compared for their effectiveness in killing microorganisms on or within barley seeds . Surface sterilization with 12.5, 25 or 50% (v/v) NaOCl for 5, 15 or 30 min, decreased Fusarium spp., Epicoccum purpurascens, and Bacillus spp . but did not kill Alternaria alternata . However, surface sterilization with 0.1 or 0.2% (w/v) HgCl2 for 3 min significantly decreased A . alternata, Fusarium spp . and E . purpurascens but Bacillus spp . were only killed by 0.3% (w/v) HgCl2 used for 10 min, which also decreased seed germination . Aspergillus flavus inoculated onto barley seeds as spores, was completely killed by surface sterilization with NaOCl but not with HgCl2, while Fusarium culmorum was killed by both NaOCl and HgCl2 treatments . Fumigation with methyl bromide yielding a concentration-time product of 3000 mg h l-1 or with propylene oxide giving a concentration-time product of 2400 mg h l-1 eliminated all filamentous fungi but Bacillus spp . and yeasts survived, and both treatments adversely affected seed germination . Gamma irradiation at a dose of 4 kGy eliminated most Alternaria, Fusarium and Epicoccum spp . but a dose of 12 kGy was required to kill Bacillus spp., yeasts and Aureobasidium pullulans . Germination was improved slightly up to a dose of 8 kGy but gradually decreased with increase in dosage to 15 kGy of gamma irradiation.

J Bacteriol, 1991 May, 173(9), 2776 - 85
Cloning, sequencing, and expression of a gene encoding a 100-kilodalton mosquitocidal toxin from Bacillus sphaericus SSII-1; Thanabalu T et al.; A cosmid library was prepared from a partial BamHI digest of total DNA from Bacillus sphaericus SSII-1 . Two hundred fifty Escherichia coli clones were screened for toxicity against larvae of the mosquito Culex quinquefasciatus . One toxic clone, designated pKF2, was chosen for further study . Two toxic subclones, designated pXP33 and pXP34, obtained by ligating PstI-derived fragments of pKF2 into pUC18, contained the same 3.8-kb fragment, but in opposite orientations . Sequence analysis revealed the presence of an open reading frame corresponding to a 100-kDa protein and the 3' end of a further open reading frame having significant homology to open reading frames of transposons Tn501 and Tn21 . The sequence of the SSII-1 toxin was compared with those of known toxins and was found to show regional homology to those of ADP-ribosyltransferase toxins . The distribution of the toxin gene among other B . sphaericus strains was examined.

Mikrobiol Zh, 1991 May-Jun, 53(3), 38 - 42
{The flotation characteristics of Bacillus cells and spores}; Stabnikova EV et al.; Variations in hydrophobicity of the surface of bacillary cells and their capacity to flotation in the process of batch cultivation have been studied . It is shown that hydrophobicity of the cell surface increases in the course of batch cultivation of Bacillus thuringiensis, B . licheniformis and B . megaterium . Hydrophobicity of spores of the mentioned cultures is considerably higher than that of the vegetative cells . The increase of hydrophobicity of bacillary cells positively correlated with their capacity to flotation . That is why the use of flotation for the age fractionation of bacillary cells is possible: spores are concentrated in the foam while vegetative cells remain in the culture liquid.

Proc Natl Acad Sci U S A, 1991 May 1, 88(9), 3762 - 6
Assembly and deacetylation of N-acetylglucosaminyl-plasmanylinositol in normal and affected paroxysmal nocturnal hemoglobinuria cells; Hirose S et al.; Decay-accelerating factor (DAF) is anchored in cell membranes by a glycosyl-plasmanylinositol (GPI) moiety that is transferred to it en bloc in the rough endoplasmic reticulum . To analyze the biochemical reactions involved in preassembly of this structure, a human hematopoietic cell-free system was employed . Incubation of cell extracts with UDP-{3H}GlcNAc and butanol partitioning of reaction mixtures yielded two products similar in TLC mobility to intermediates described in Trypanosoma brucei . Both species were sensitive to Bacillus thuringiensis phosphatidylinositol-specific phospholipase C, indicative of association of {3H}GlcNAc label with a plasmanylinositol-containing acceptor . In contrast to trypanosome intermediates, which contain phosphatidylinositol (1,2-diacylglycerophosphoinositol), however, alkali treatment and phospholipase A2 digestion generated butanol-phase products characteristic of glycosylated plasmanylinositol (1-alkyl-2-acylglycerophosphoinositol) . Kinetic and pulse-chase experiments indicated that the slower-migrating species was a product of the faster and that it, but not the faster, was sensitive to both GPI-specific phospholipase D and nitrous acid deamination, consistent with conversion of GlcNAc- to GlcN-plasmanylinositol . Accordingly, acetic anhydride acetylation retransformed the slower species back to the faster . Further incubation with cell extracts converted the slower species into more polar products . Lysates of normal and of affected blood leukocytes from two paroxysmal nocturnal hemoglobinuria (PNH) patients supported assembly of the two intermediates within 1 min . Thus, the initial enzymes mediating human GPI-anchor assembly are GlcNAc-plasmanylinositol transferase and GlcNAc-plasmanylinositol deacetylase, their substrates contain plasmanylinositols, and the products of their activities are normal in affected PNH cells.

Agric Biol Chem, 1991 May, 55(5), 1259 - 63
Cloning and expression of the sarcosine oxidase gene from Bacillus sp . NS-129 in Escherichia coli; Koyama Y et al.; The gene coding for a thermostable sarcosine oxidase (EC 1.5.3.1) was isolated from Bacillus sp . NS-129 . The primary structure of sarcosine oxidase deduced from the nucleotide sequence was a protein composed of 387 amino acids with molecular weight 42,955 . When the sarcosine oxidase was overproduced to about 35% of soluble protein in E . coli under the control of a lac promoter, the sarcosine oxidase activity of the crude extract was increased 3-fold by the addition of FAD . This indicates that most of the enzyme is accumulated in an active form, a flavinless aporotein, in the cell.

Proc Natl Acad Sci U S A, 1991 Apr 15, 88(8), 3111 - 5
Site-specific integration of mycobacteriophage L5: integration-proficient vectors for Mycobacterium smegmatis, Mycobacterium tuberculosis, and bacille Calmette-Guérin; Lee MH et al.; Mycobacteriophage L5, a temperate phage of mycobacteria, integrates site-specifically into the Mycobacterium smegmatis chromosome . We have identified the int gene and attP site of L5, characterized the chromosomal attachment site (attB), and constructed plasmid vectors that efficiently transform M . smegmatis through stable site-specific integration of the plasmid into the bacterial genome . These integration-proficient plasmids also efficiently transform slow-growing mycobacteria such as the pathogen Mycobacterium tuberculosis and the vaccine strain bacille Calmette-Guerin (BCG) . The ability to easily generate stable recombinants in these slow-growing mycobacteria without the requirement for continual selection is of particular importance for the construction of recombinant BCG vaccines and for the isolation and characterization of mycobacterial pathogenic determinants in animal model systems . Integration vectors of this type should be of general use in a number of additional bacterial systems where temperate phages have been identified.

Nucleic Acids Res, 1991 Apr 25, 19(8), 1825 - 9
Overexpression, purification and crystallization of BamHI endonuclease; Jack WE et al.; The type II restriction endonuclease BamHI has been expressed in E . coli, producing 100-fold more enzyme than the wild type Bacillus amyloliquefaciens H strain . This high yield has facilitated purification to homogeneity of large amounts of the enzyme, along with its crystallization in a form which diffracts to at least 1.9 A in X-ray analysis.

J Biol Chem, 1991 Apr 25, 266(12), 7864 - 9
A barbiturate-regulated protein binding to a common sequence in the cytochrome P450 genes of rodents and bacteria; He JS et al.; Analyses of the 5' regulatory sequences of genes encoding barbiturate-inducible cytochromes P450BM-1 and P450BM-3 from Bacillus megaterium and of the 5' sequences of genes for barbiturate-inducible P450b and P450e of the rat revealed a string of 17 base pairs in each of the genes that shared a high degree of sequence identity . Labeled oligonucleotide probes of each of these four sequences were tested in gel retardation assays with protein obtained from B . megaterium grown either in the presence or absence of barbiturates or with protein from nuclear extracts from livers of rats left untreated or injected with phenobarbital . Each of the four 17-mers bound strongly to a single protein from bacteria grown in the absence of barbiturates, but this binding was dramatically reduced with protein from pentobarbital- or phenobarbital-grown cells . Conversely, the probes complexed weakly to one protein band from nuclear extracts from untreated rats but much more strongly with protein from phenobarbital-treated rats . Similar effects could be obtained by prolonged incubation with phenobarbital of either soluble protein from the bacteria grown in the absence of barbiturates or nuclear extract protein from untreated rats . Deletion analysis of the 5'-flanking region of the P450BM-1 gene of B . megaterium revealed a putative repressor binding site located within a 24-base pair DNA segment that included the 17-base pair sequence involved in barbiturate-regulated protein binding.

Eur J Biochem, 1991 Apr 23, 197(2), 337 - 43
Molecular cloning, expression and nucleotide sequence of the endo-beta-1,3-1,4-D-glucanase gene from Bacillus licheniformis . Predictive structural analyses of the encoded polypeptide; Lloberas J et al.; A Bacillus licheniformis gene coding for an endo-beta-1,3-1,4-D-glucanase have been cloned in Escherichia coli and sequenced . The open reading frame contains a sequence of 731 bp, encoding a polypeptide of 243 amino acid residues, with a molecular mass of 27404 Da (24418 Da without the putative signal peptide), which corresponds to the enzyme we had previously isolated and characterized . The signal peptide is functional in E . coli . More than 60% of the endo-beta-1,3-1,4-D-glucanase activity is extracellular or periplasmic . The polypeptide is highly similar to other reported Bacillus beta-glucanases . Several structural predictive analyses (secondary structure, hydropathic plots, similarity with other related enzymes, etc.) have been performed . From these analyses we assign a tentative three-functional-domain structure for the enzyme (signal peptide, substrate binding and catalytic domains) and a putative lysozyme-like active site.

FEBS Lett, 1991 Apr 22, 282(1), 13 - 6
Improving the thermostability of the neutral protease of Bacillus stearothermophilus by replacing a buried asparagine by leucine; Eijsink VG et al.; Amino acids buried in the hydrophobic interior of a protein with polar side chain atoms, which are not involved in hydrogen bonding or electrostatic interactions, have an adverse effect on protein stability . Replacing such residues by hydrophobic ones may render a protein more stable . Asparagine 241, which is buried in the neutral protease of Bacillus stearothermophilus, was replaced by leucine by site-directed mutagenesis . This mutation increased the stability of the protein by 0.7 +/- 0.1 degree.

J Mol Biol, 1991 Apr 20, 218(4), 703 - 4
Crystallization of the hybrid Bacillus (1-3,1-4)-beta-glucanase H(A16-M); Keitel T et al.; The extremely thermostable hybrid Bacillus (1-3,1-4)-beta-glucanase H(A16-M) has been crystallized with polyethylene glycol by vapour diffusion . The single crystals diffract to a resolution of 2.2 A . The protein crystallizes in the orthorhombic space group P2(1)2(1)2(1) with a = 70.22(7) A, b = 72.56(1) A, c = 49.97(1) A, and has one molecule per asymmetric unit.

Biochem Biophys Res Commun, 1991 Apr 15, 176(1), 356 - 63
Fluidity of bacterial membrane lipids monitored by intramolecular excimerization of 1.3-di(2-pyrenyl)propane; Jurado AS et al.; Intramolecular excimer formation of 1,3-di(2-pyrenyl)propane was used to study the fluidity of liposomes prepared from membrane polar lipids of Bacillus stearothermophilus . On the basis of spectral data, local polarity and polarizability parameters were established suggesting that the probe molecules are located well inside the membranes, but displaced towards the polar head groups of the phospholipid molecules . The excimerization rate is very sensitive to lipid phase transitions and pretransitions of synthetic pure lipid bilayers . In bacterial lipids from cultures grown at 55 and 68 degrees C, thermal profiles of excimer to monomer intensity ratios (I'/I) show a broad transition which is displaced to higher temperatures in response to the increase of the growth temperature; these results correlate well with differential scanning calorimetry data and fluorescence polarization of diphenylhexatriene . Additionally, lipid bilayers of bacteria grown at 68 degrees C exhibit a decreased membrane fluidity, as monitored by both fluorescent probes.

Proc Natl Acad Sci U S A, 1991 Apr 15, 88(8), 3324 - 8
Modification of the coding sequence enhances plant expression of insect control protein genes; Perlak FJ et al.; Increased expression of the insect control protein genes of Bacillus thuringiensis in plants has been critical to the development of genetically improved plants with agronomically acceptable levels of insect resistance . The expression of the cryIA(b) gene was compared to partially modified (3% nucleotide difference) and to fully modified (21% nucleotide difference) cryIA(b) and cryIA(c) genes in tobacco and tomato . The modified genes increased the frequency of plants that produced the proteins at quantities sufficient to control insects and dramatically increased the levels of these proteins . Among the most highly expressing transformed plants for each gene, the plants with the partially modified cryIA(b) gene had a 10-fold higher level of insect control protein and plants with the fully modified cryIA(b) had a 100-fold higher level of CryIA(b) protein compared with the wild-type gene . Similar results were obtained with the fully modified cryIA(c) gene in plants . Specific sequences of the partially modified cryIA(b) gene were analyzed for their ability to affect cryIA(b) gene expression in tobacco . The DNA sequence of a single region was identified as important to the improvement of plant expression of the cryIA(b) gene . The increased levels of cryIA(b) mRNA were not directly proportional to the increased levels of CryIA(b) protein in plants transformed with the modified cryIA(b) genes, indicating that the nucleotide sequence of these genes had an effect in improving their translational efficiency in plants.

FEMS Microbiol Lett, 1991 Apr 15, 63(2-3), 205 - 9
Identity of hemolysins produced by Bacillus thuringiensis and Bacillus cereus; Honda T et al.; A hemolysin (Bt-hemolysin) produced by Bacillus thuringiensis var . kurstaki HD-1 producing crystalline toxin(s) was purified by successive treatments of ammonium sulfate (45-65%) and column chromatography using DEAE-cellulose, Sephadex G-75 and KB-002 (a hydroxyapatite column for fast protein liquid chromatography) . A hemolysin (Bc-hemolysin) produced by B . cereus HG-6A was also purified by the same procedure . The purified Bt-hemolysin and Bc-hemolysin, both of which are thiol-activated hemolysins, were biologically, physicochemically and immunologically identical . These findings provide further evidence of the similarity of B . thuringiensis, which is being used as a biological insecticide, to B . cereus, a toxigenic organism of food poisoning.

J Immunol, 1991 Apr 15, 146(8), 2754 - 62
Early appearing gamma/delta-bearing T cells during infection with Calmétte Guérin bacillus; Inoue T et al.; To search for a potential role of TCR gamma/delta T cells in host-defense against mycobacterial infection, we analyzed the kinetics, repertoire, specificity, and cytokine production of gamma/delta T cells in the peritoneal exudate cells (PEC), lymph node (LN) cells and spleen cells during an i.p . infection with a sublethal dose (5 x 10(5) of viable Bacillus Calmette-Guerin (BCG) in mice . In the PEC on day 7 after infection, approximately 26% of the CD3+ cells were CD4-CD8-, most of which expressed TCR gamma/delta on their surface . However, the PEC on day 28 contained an increased number of alpha/beta T cells that were CD4+8- or CD4-8+ and the proportion of gamma/delta T cells in the PEC reciprocally decreased to 18% of the CD3+ cells . The kinetics of gamma/delta and alpha/beta T cells in the LN during BCG infection showed in much the same pattern as that seen in the PEC . When purified CD4-CD8- cells in the LN on day 7 after BCG infection were cultured with sonicated BCG lysate, PPD derived from Mycobacterium tuberculosis or recombinant 65 kDa heat shock protein derived from Mycobacterium bovis, the gamma/delta T cells on this stage significantly proliferated and secreted IL-2 in response to sonicated BCG lysate and PPD but not to 65 kDa heat shock protein . V gene segment usage analysis with PCR method revealed that purified protein derivative-reactive gamma/delta T cells preferentially used V gamma 1/2/V delta 6, whereas gamma/delta T cells polyclonally expanded in response to the BCG lysate . These results suggest that gamma/delta T cells specific for mycobacterial antigens preceding alpha/beta T cells in appearance during infection may serve as a first line of defense against mycobacterial infection.

Appl Environ Microbiol, 1991 Apr, 57(4), 981 - 6
The solubility of inclusion proteins from Bacillus thuringiensis is dependent upon protoxin composition and is a factor in toxicity to insects; Aronson AI et al.; Bacillus thuringiensis subsp . aizawai HD133 is one of several strains particularly effective against Plodia interpunctella selected for resistance to B . thuringiensis subsp . kurstaki HD1 (Dipel) . B . thuringiensis subsp . aizawai HD133 produces inclusions containing three protoxins, CryIA(b), CryIC, and CryID, and the CryIC protoxin has been shown to be active on resistant P . interpunctella as well as on Spodoptera larvae . The CryIA(b) protoxin is very similar to the major one in B . thuringiensis subsp . kurstaki HD1, and as expected, this protoxin was inactive on resistant P . interpunctella . A derivative of B . thuringiensis subsp . aizawai HD133 which had been cured of a 68-kb plasmid containing the cryIA(b) gene produced inclusions comprising only the CryIC and CryID protoxins . Surprisingly, these inclusions were much less toxic for resistant P . interpunctella and two other Lepidoptera than those produced by the parental strain, whereas the soluble protoxins from these strains were equally effective . In contrast, inclusions from the two strains were about as active as soluble protoxins for Spodoptera frugiperda larvae, so toxicity differences between inclusions may be due to the solubilizing conditions within particular larval guts . Consistent with this hypothesis, it was found that a higher pH was required to solubilize protoxins from inclusions from the plasmid-cured strain than from B . thuringiensis subsp . aizawai HD133, a difference which is probably attributable to the absence of the CryIA(b) protoxin in the former . The interactions of structurally related protoxins within an inclusion are probably important for solubility and are thus another factor in the effectiveness of B . thuringiensis isolates for particular insect larvae.

Appl Environ Microbiol, 1991 Apr, 57(4), 1075 - 81
Characterization of mosquitocidal activity of Bacillus thuringiensis subsp . fukuokaensis crystal proteins; Yu YM et al.; The mosquitocidal crystals of Bacillus thuringiensis subsp . fukuokaensis were isolated and bioassayed against fourth-instar larvae of two mosquito species . The 50% lethal concentration values of the crystals to Aedes aegypti and Culex quinquefasciatus were 4.1 and 2.9 micrograms/ml, respectively . In addition, the solubilized crystals had hemolytic activity; 50 micrograms/ml was the lowest detectable level . Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis revealed that the crystals consisted of polypeptides of 90, 86, 82, 72, 50, 48, 37, and 27 kDa . When the solubilized inclusion was treated with C . quinquefasciatus midgut brush border membrane vesicles or Manduca sexta gut juice, only one major protein was detected . This protein retained mosquitocidal activity but had no detectable hemolytic activity . Immunological analysis of this subspecies and the subspecies israelensis, kyushuensis and darmstadiensis by using polyclonal antisera raised against the whole-crystal protein of B . thuringiensis subsp . fukuokaensis revealed that the proteins in subsp . fukuokaensis are distinct from proteins in the other subspecies because little cross-reaction was observed . Analysis of the plasmid pattern showed that the crystal protein genes are located on a plasmid of 130 MDa . Analysis of plasmid and chromosomal DNA from subsp . fukuokaensis showed little homology to the 72-kDa toxin gene (PG-14) of B . thuringiensis subsp . morrisoni . However, some of the proteins of B . thuringiensis subsp . fukuokaensis are homologous to other B . thuringiensis toxins because N-terminal amino acid analysis revealed that the 90-kDa protein is encoded by a cryIV gene type.

Appl Environ Microbiol, 1991 Apr, 57(4), 1000 - 5
Transduction of certain genes by an autonomously replicating Bacillus thuringiensis phage; Walter TM et al.; A derivative of Bacillus thuringiensis subsp . kurstaki HD1 (HD1-9) released transducing phage (TP21) from late exponential cultures . Three of seven markers tested were transduced into Bacillus cereus, but only two of these (cysC and trpB/F) were transduced at a frequency of more than 100 times the reversion rates . A limited transduction capacity was given further support in that few chromosomal markers were carried in the HD1-9 lysate, as demonstrated by Southern hybridization . Restriction fragments from the phage DNA and from total B . thuringiensis DNA hybridized to an insertion sequence (IS231-like) probe, which may provide a region of homology for transduction . All of the B . cereus transductants contained the phage as a 44-kb plasmid, and each could transduce both the cys and trp genes to other B . cereus auxotrophs, albeit at lower frequencies than those for the B . thuringiensis transducing phage . In some cases, especially for cys, the transduced gene was integrated into the chromosome of the recipient, whereas the trp gene in many cases appeared to be lost with curing of the 44-kb plasmid . In addition, some B . cereus transductants lost prototrophy but retained a 44-kb plasmid, consistent with the presence of TP21 helper phage . These phage may mediate the subsequent transduction from B . cereus phototrophs . TP21 replicates as a plasmid and, at least under the conditions studied, selectively transfers markers to B . cereus.

Gene, 1991 Apr, 100, 85 - 93
Co-expression of a Saccharomyces diastaticus glucoamylase-encoding gene and a Bacillus amyloliquefaciens alpha-amylase-encoding gene in Saccharomyces cerevisiae; Steyn AJ et al.; A glucoamylase-encoding gene (STA2) from Saccharomyces diastaticus and an alpha-amylase-encoding gene (AMY) from Bacillus amyloliquefaciens were cloned separately into a yeast-integrating shuttle vector (YIp5), generating recombinant plasmids pSP1 and pSP2, respectively . The STA2 and AMY genes were jointly cloned into YIp5, generating plasmid pSP3 . Subsequently, the dominant selectable marker APH1, encoding resistance to Geneticin G418 (GtR), was cloned into pSP3, resulting in pSP4 . For enhanced expression of GtR, the APH1 gene was fused to the GAL10 promoter and terminated by the URA3 terminator, resulting in pSP5 . Plasmid pSP5 was converted to a circular minichromosome (pSP6) by the addition of the ARS1 and CEN4 sequences . Laboratory strains of Saccharomyces cerevisiae transformed with plasmids pSP1 through pSP6, stably produced and secreted glucoamylase and/or alpha-amylase . Brewers' and distillers' yeast transformed with pSP6 were also capable of secreting amylolytic enzymes . Yeast transformants containing pSP1, pSP2 and pSP3 assimilated soluble starch with an efficiency of 69%, 84% and 93%, respectively . The major starch hydrolysis products produced by crude amylolytic enzymes found in the culture broths of the pSP1-, pSP2- and pSP3-containing transformants, were glucose, glucose and maltose (1:1), and glucose and maltose (3:1), respectively . These results confirmed that co-expression of the STA2 and AMY genes synergistically enhanced starch degradation.

Jikken Dobutsu, 1991 Apr, 40(2), 231 - 3
A serological survey on Bacillus piliformis infection in laboratory rabbits in Japan; Goto K et al.; A total of 544 rabbit sera obtained from 6 commercial breeding facilities and 9 research institutions during 1985-1990 were tested for Bacillus piliformis antibody by an enzyme-linked immunosorbent assay (ELISA) and indirect immunofluorescent antibody test (IFAT) . The antibody was detected in 53 (14.2%) rabbits from 3 breeding facilities and 30 (17.4%) rabbits from 6 research institutions, indicating the prevalence of B . piliformis infection among laboratory rabbits in Japan . The overall agreement with ELISA for immune status was 96.9% (527/544) with IFAT . In tests of the ability of ELISA and IFAT to quantitate antibody, a correlation coefficient (r) of 0.86 (P less than 0.01) was obtained by plotting the measured average log of the ELISA titer against the corresponding log of the IFAT titer.

Eur J Pediatr, 1991 Apr, 150(6), 423 - 4
Bacille Calmette-Guérin--associated neonatal hepatitis; Simma B et al.; We describe a full-term immunocompetent neonate who developed jaundice at 3 weeks of age . Physical examination disclosed no abnormalities . Laboratory investigations showed direct reacting hyperbilirubinaemia and elevated liver enzymes . Liver biopsy revealed a non-caseating granulomatous hepatitis . The patient made an uneventful recovery within 4 weeks without therapy . Bacille Calmette-Guerin hypersensitivity reaction is suggested as the reason for this neonatal hepatitis.

J Gen Virol, 1991 Apr, 72 ( Pt 4), 757 - 61
Rice tungro disease is caused by an RNA and a DNA virus; Jones MC et al.; We present evidence that rice tungro spherical virus (RTSV) has a genome of polyadenylated single-stranded RNA of about 10 kb whereas rice tungro bacilliform virus (RTBV) contains double-stranded circular DNA . RTBV DNA has been mapped and shown to have two discontinuities, one in each strand, at specific sites; it thus resembles that of the caulimoviruses . Gel electrophoresis of RTSV preparations revealed two protein bands (Mr 35K and 26K) . RTBV yielded two major protein bands of 37K and 33K together with several minor species of higher and lower Mr which react with antiviral serum.

J Bacteriol, 1991 Apr, 173(8), 2498 - 505
Menaquinol-nitrate oxidoreductase of Bacillus halodenitrificans; Ketchum PA et al.; When grown anaerobically on nitrate-containing medium, Bacillus halodenitrificans exhibited a membrane-bound nitrate reductase (NR) that was solubilized by 2% Triton X-100 but not by 1% cholate or deoxycholate . Purification on columns of DE-52, hydroxylapatite, and Sephacryl S-300 yielded reduced methyl viologen NR (MVH-NR) with specific activities of 20 to 35 U/mg of protein that was stable when stored in 40% sucrose at -20 degrees C for 6 weeks . 3-{(3-cholamidopropyl)dimethylammonio}-2-hydroxypropone-1-sulfonat e (CHAPSO) and dodecyl-beta-D-maltoside stimulated enzyme activity three- to fourfold . Membrane extractions yielded purified NR that separated after electrophoresis into a 145-kDa alpha subunit, a 58-kDa beta subunit, and a 23-kDa gamma subunit . The electronic spectrum of dithionite-reduced, purified NR displayed peaks at 424.6, 527, and 557 nm, indicative of the presence of a cytochrome b, an interpretation consistent with the pyridine hemochrome spectrum formed . Analyses revealed a molybdenum-heme-non-heme iron ratio of 1:1:8 for the NR and the presence of molybdopterin . Electron paramagnetic resonance (EPR) signals characteristic of iron-sulfur centers were detected at low temperature . EPR also revealed a minor signal centered in the g = 2 region of the spectra . Upon reduction with dithionite, the enzyme displayed signals at g = 2.064, 2.026, 1.906, and 1.888, indicative of the presence of low-potential iron-sulfur centers, which resolve most probably as two {4Fe-4S}+1 clusters . With menadiol as the substrate for nitrate reduction, the Km for nitrate was 50-fold less than that seen when MVH was the electron donor . The cytochrome b557-containing enzyme from B . halodenitrificans is characterized as a menaquinol-nitrate:oxidoreductase.

J Urol, 1991 Apr, 145(4), 738 - 40
Prospective randomized comparison of intravesical with percutaneous bacillus Calmette-Guerin versus intravesical bacillus Calmette-Guerin in superficial bladder cancer; Lamm DL et al.; Conflicting reports of the necessity for percutaneous bacillus Calmette-Guerin (BCG) administration with intravesical BCG prompted us to evaluate its benefit in a randomized prospective comparison of intravesical versus intravesical with percutaneous BCG therapy . Intravesical Tice BCG was given in a dose of 50 mg . with or without percutaneous BCG weekly for 6 weeks, at 8, 10 and 12 weeks, at 6 months and every 6 months thereafter . Tumor recurrence was documented in 13 of 30 patients (43%) receiving only intravesical BCG and in 15 of 36 patients (42%) receiving intravesical plus percutaneous BCG . The addition of percutaneous BCG to intravesical therapy did not increase treatment efficacy in this study.

Exp Cell Res, 1991 Apr, 193(2), 320 - 30
Stage-dependent localization of LHCP II apoprotein in the Golgi of synchronized cells of Euglena gracilis by immunogold electron microscopy; Osafune T et al.; We have localized LHCP II apoprotein in the Golgi and thylakoids of Euglena gracilis Klebs var . bacillaris Cori and strain Z Pringsheim by electron microscopy using a specific antibody and protein A-gold . Using synchronized cells (light, 14 h:dark, 10 h) we show that thylakoids are always immunoreactive . There is no reaction in the Golgi at 0 h (the beginning of the light period) but immunoreaction appears in the Golgi soon thereafter, rises to a peak at 8 h and declines to zero by 16 h (2 h into the dark period) . The peak in immunoreaction in the Golgi immediately precedes the peak in cellular 14C-labeling of thylakoid LHCP II apoprotein seen by Brandt and von Kessel (Plant Physiol . (1983) 72, 616), supporting our suggestion that processing in the Golgi precedes deposition of LHCP apoprotein in the thylakoids . Substitution of preimmune serum for antiserum eliminates the immunoreaction in the Golgi, and thylakoids of synchronized cells of mutant Gr1BSL which lacks LHCP II apoprotein show no immunoreaction in the Golgi or thylakoids at any stage . Random observations indicate that the compartmentalized osmiophilic structure (COS) shows an immunoreaction with anti-LHCP II apoprotein antibody at 1 h into the light period (when the Golgi is not immunoreactive) and at 10 h into the light period (when the Golgi is fully reactive), suggesting that the COS remains immunoreactive throughout the cell cycle.

Eur Respir J Suppl, 1991 Apr, 13, 47s - 59s
Immune reactions in allergic alveolitis; Salvaggio JE; This article discusses newer cellular immunological and immunoregulatory events operative in the pathogenesis of "allergic alveolitis" (aa) . An early increase in neutrophils and chemotactic factors followed by an influx of macrophages and lymphoid cells with ultimate production of granulomas have been demonstrated in the bronchoalveolar lavage (BAL) fluids of experimental animals and of man in aa . Transfer of specifically sensitized lymph node and spleen cells intraperitoneally followed by antigen challenge via the respiratory tract route has resulted in production of pulmonary lesions closely resembling those observed in human aa in several animal species . Activated alveolar macrophages and T-cells, local lymphokine production, and elevated immunoglobulin levels in BAL fluid have also been demonstrated repeatedly in experimental animal models of these diseases . High levels of interleukin-1 have also been demonstrated in aqueous extracts prepared from pulmonary granulomatous lesions . Other studies have indicated a role for macrophage-derived lipoxygenase products in production of experimental pulmonary granulomatous inflammation . Mononuclear cell pulmonary infiltrates have been inhibited by corticosteroids, the use of antimacrophage serum, neonatal thymectomy, cobra factor venom, and cyclosporin . A T suppressor factor and other suppressor factors from adherent cell populations have also been shown to dampen or modulate experimental granuloma formation . Appropriately sensitized animals which demonstrate chronic pulmonary granulomatous inflammation can also become "desensitized" following a series of antigen challenges . The desensitization produced is immunospecific and nontransferable with immune serum . Studies of bronchoalveolar lavage fluids in man have demonstrated a very high percentage of lymphocytes in BAL fluids . The percentage of suppressor cytotoxic T-cells is usually above 40% and CD4:CD8 ratios are reversed . Evidence also suggests that suppressor lymphocytic alveolitis is a chronic event in patients with aa . Immunogenetic studies of granulomatous pneumonitis resembling aa in different strains of mice have shown that certain high-responder strains develop intense granulomas after inoculation with killed bacille Calmette-Guerin (BCG), whereas other low-responder strains do not . The intensity of the pulmonary granuloma formation in this model has been shown to be a dominant and polygenic trait and to be linked to the immunoglobulin heavy chain locus (IgH) . Strains that develop intense, chronic granulomatous inflammation have usually proved to be anergic, and the anergy also appears to be under genetic control.(ABSTRACT TRUNCATED AT 400 WORDS)

J Appl Bacteriol, 1991 Apr, 70(4), 315 - 24
Differentiation of dairy strains of the Bacillus cereus group by phage typing, minimum growth temperature, and fatty acid analysis; Vaisanen OM et al.; A total of 130 Bacillus strains were isolated from dairy products, the dairy environment and from packaging boards and board-producing machines . Ninety-eight of these were members of the B . cereus group (B . cereus, B . mycoides and B . thuringiensis) as determined by whole cell fatty acid composition . Fatty acid composition did not differentiate between the three species . Of the 98 strains, which were indistinguishable by biochemical tests, 87 could be assigned into 21 different phage types (11 strains remained untypable) when tested with 12 B . cereus, B . mycoides and B . thuringiensis phages . The distribution of phage types between strains from different sources showed that the source of contamination of the dairy products was of milk origin and not from the packaging materials . Most strains isolated from the dairy products were able to grow below 10 degrees C, whereas strains from the dairy environment and from board mills had higher minimum growth temperatures.

Biotechnol Appl Biochem, 1991 Apr, 13(2), 231 - 7
Effect of phenolic compounds and oleuropein on the germination of Bacillus cereus T spores; Tassou CC et al.; The phenolic compounds extracted from olives with ethyl acetate inhibited germination and outgrowth of Bacillus cereus T spores . Purified oleuropein, a well-characterized component of olive extract, inhibited these processes also . The addition of oleuropein and olive extracts 3 or 5 min after germination began, immediately decreased the rate of change of phase bright to phase dark spores and delayed significantly outgrowth.

Eur J Immunol, 1991 Apr, 21(4), 1047 - 52
Characterization of the induction of persistence of major histocompatibility complex class II by hybrids of macrophages from bacillus Calmette Guerin-resistant mice; Faris M et al.; Peritoneal macrophages (M phi) from mice that are resistant to infection by Mycobacterium bovis (strain BCG) (Bcgr) can be induced to express major histocompatibility complex (MHC) class II glycoproteins (I-A) continuously upon treatment with 100 units of recombinant interferon-gamma (rIFN-gamma) . In contrast, M phi from mice that are susceptible to BCG (Bcgs) express I-A transiently . Persistent expression of I-A does not require the continued synthesis of the glycoprotein . Thus, treatment with cycloheximide (CHX) reduces I-A expression by M phi that express I-A transiently but does not affect the expression of I-A that is persistently expressed . It was not possible, in these studies, to characterize the induction of persistence independent of MHC class II expression because of the 24-48 h required for MHC class II synthesis and cycling to the cell surface . During this time, persistence was also induced . To characterize persistence independent of MHC class II induction we have produced M phi-M phi somatic cell hybrids that express I-A constitutively by fusing cells from a Bcgs M phi cell line with M phi from Bcgr mice . Treatment of some of the hybrids with CHX reduced MHC class II expression . The M phi hybrids required treatment with high doses of rIFN-gamma to induce CHX-resistant I-A expression . The induction of the persistence of I-A, following the addition of rIFN-gamma, required a short burst of protein synthesis as well as the presence of rIFN-gamma for at least 3 h . The addition of actinomycin D simultaneously with rIFN-gamma did not prevent the induction of the persistence of I-A expression by one of the M phi hybrids (F6.4) . In contrast, the induction of persistence of I-A expression required a longer period of induction than was observed for hybrid F6.4, which was attributed to the requirement for new RNA and protein synthesis by the A1.8 hybridoma.

J Bacteriol, 1991 Apr, 173(8), 2548 - 55
Cell wall assembly in Bacillus megaterium: incorporation of new peptidoglycan by a monomer addition process; Gally DL et al.; The pattern of cross-linking in the peptidoglycan of Bacillus megaterium has been studied by the pulsed addition of radiolabeled diaminopimelic acid . The distribution of label in muropeptides, generated by digestion with Chalaropsis muramidase and separated by high-performance liquid chromatography, stabilized after 0.15 of a generation time . The proportion of label in the acceptor and donor positions of isolated muropeptide dimers stabilized over the same period of time . The results have led to the formulation a new model for the assembly of peptidoglycan into the cylindrical wall of B . megaterium by a monomer addition process . Single nascent glycan peptide strands form cross-linkages only with material at the inner surface of the wall . Maturation is a direct consequence of subsequent incorporation of further new glycan peptide strands, and there is no secondary cross-linking process . The initial distribution of muropeptides is constant . It follows that the final pattern of cross-linking in the wall is determined solely by, and can be forecast from, this repetitive pattern of incorporation . In a modified form, this model can also be applied to assembly of cell walls in rod-shaped gram-negative bacteria.

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