Int J Pept Protein Res, 1991 Apr, 37(4), 325 - 30 Modification of a zinc proteinase from Bacillus mesentericus strain 76 by diethylpyrocarbonate; Stoeva S; Diethylpyrocarbonate (DEPC) inactivated the neutral zinc proteinase from Bacillus mesentericus strain 76/Bacillus subtilis (MCP 76) by ethoxycarbonylation completely . Exposure of the enzyme to DEPC together with the competitive inhibitor Z-L-phenylalanine prevented the loss of activity toward both peptide and protein substrates . Treatment with hydroxylamine restored the catalytic properties of the modified MCP 76 to that of the native enzyme . After chymotryptic digestion of ethoxycarbonylated MCP 76 in the presence and absence of Z-L-phenylalanine a single histidyl residue essential for the enzyme activity was isolated and identified as histidine 231. Ann Plast Surg, 1991 Apr, 26(4), 403 - 6 Spontaneous regression of metastatic melanoma; Hurwitz PJ; Spontaneous regression of malignant melanoma is a well-known but rare event . It seems to be caused by immunological enhancement similar to the regression observed after intralesional injection of Bacille Calmette-Guerin into cutaneous melanoma metastases . A patient is presented in whom an incisional biopsy of a metastatic melanoma in an inguinal lymph node was followed by complete disappearance of the metastasis . Reviewing the literature, several similar patients were found in whom spontaneous regression of a metastatic melanoma occurred after an incomplete excision or biopsy of the tumor . It appears that in rare instances, an unspecific mechanical stimulus may trigger an increase in immunocompetence, although laboratory evidence for this has only rarely been produced . A plea is made for serial immunological studies in consecutive patients with melanoma, to further elucidate these puzzling phenomena. J Vet Med Sci, 1991 Apr, 53(2), 281 - 6 Purification and characterization of the vascular permeability factor produced by Bacillus cereus; Shinagawa K et al.; Purification of an extracellular protein exhibiting the vascular permeability activity produced by Bacillus cereus was performed by ammonium sulfate precipitation followed by chromatography on DE-32 cellulose, Sephadex G-100, and Sephadex G-75 . The purified protein was found to be electrophoretically and antigenically almost homogeneous although it contained a trace of contaminant . The molecular weight of the protein was calculated to be 45,000 by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate . The purified protein showed vascular permeability activity and mouse lethal toxicity, and caused fluid accumulation in ligated mouse intestinal loops, whereas it did not show any hemolytic and lecithinase activities . From these findings, the purified protein is suggested to be an enterotoxin (or a diarrheagenic toxin) responsible for diarrhea caused by B . cereus in a diarrheal-type food poisoning. J Vet Med Sci, 1991 Apr, 53(2), 167 - 71 Fluid accumulation in mouse ligated intestine inoculated with the vascular permeability factor produced by Bacillus cereus; Shinagawa K et al.; Partially purified vascular permeability (VP) factor (VPF) of Bacillus cereus induced fluid accumulation in the ligated intestinal loops of mouse (MIL) and rabbit (RIL), suggesting that the VP activity may correlate with fluid accumulation in ligated intestinal loops of these animals . Fluid accumulation was observed at 6-8 hr in 55-67% of mouse intestinal loops inoculated with 40-50 immunodiffusion units (IDU) of partially purified VPF, whereas it was found at 2 hr in all loops with 400-600 IDU of partially purified VPF . In rabbit intestinal loops with 120-190 IDU of partially purified VPF, fluid accumulation was observed at 6 hr . From these findings, VPF produced by B . cereus can be easily detected in both MIL and RIL . The intestinal tissue of mouse intestinal loops was histopathologically damaged at different concentrations of the VPF to induce fluid accumulation . With 50 IDU of partially purified VPF, severe edema was found in the laminia proprial layer and submucosa . With 600 IDU of partially purified VPF, on the other hand, severe necrosis in the surface epithelium of villus and laminia proprial layer, and hyperemia in the submucosa were observed, suggesting that partially purified VPF may be cytotoxic and/or intestinecrotic. Protein Expr Purif, 1991 Apr-Jun, 2(2-3), 199 - 204 Purification and partial characterization of acyl carrier protein from Euglena gracilis variety bacillaris; Williams SG et al.; Acyl carrier protein (ACP) was purified from Euglena gracilis variety bacillaris in yields of about 1 mg/100 g (wet wt) of cells . Antibodies against the purified protein were raised in hens and isolated from eggs . Antibodies raised against Euglena ACP inhibited the Euglena chloroplast nonaggregated fatty acid synthetase using either Euglena or Escherichia coli ACP as a substrate . Comparisons with other ACPs included the following items: biologic activity, acidic pI, size, behavior in size exclusion media, and amino acid sequence of the N-terminal portion of the molecule. Protein Expr Purif, 1991 Apr-Jun, 2(2-3), 151 - 7 Hyperexpression in Escherichia coli, purification, and characterization of the metallo-beta-lactamase of Bacillus cereus 5/B/6; Shaw RW et al.; We used site-directed mutagenesis to introduce both a NdeI restriction endonuclease site and an initiator codon at the junction of the leader and structural gene sequences of the metallo-beta-lactamase of Bacillus cereus 5/B/6 . This construct allowed us to clone just the beta-lactamase structural gene sequence into an Escherichia coli expression vector . E . coli cells were transformed with the recombinant plasmid, the B . cereus beta-lactamase was expressed, and these E . coli cells were disrupted by sonic oscillation . When the resultant suspensions were clarified by ultracentrifugation, the B . cereus beta-lactamase represented 15% of the total protein in the supernatant . Subsequent gel filtration and ion-exchange chromatography allowed the first reported purification to homogeneity of the B . cereus beta-lactamase from E . coli with an 87% recovery and an overall yield of 17 mg of enzyme per liter of cell culture . The electrophoretic mobilities of the enzyme expressed in and purified from E . coli and the enzyme purified directly from B . cereus were identical in both native and sodium dodecyl sulfate gel electrophoreses . As with the B . cereus enzyme, Km and Vmax (using cephalosporin C as substrate) for the enzyme purified from E . coli were 0.39 mM and 1333 units/mg protein, respectively . Likewise, the Co(II)-reconstituted enzyme purified from E . coli, which retained 29% of the activity of the Zn(II) enzyme, had electronic absorption spectra with maxima at 347, 551, 617, and 646 nm with extinction coefficients of 900, 250, 173, and 150 M-1 cm-1, respectively. Indian J Lepr, 1991 Apr-Jun, 63(2), 203 - 8 Assessing the viability of Mycobacterium leprae by the fluorescein diacetate/ethidium bromide staining technique; Palomino JC et al.; In the present study we have evaluated the Fluorescein Diacetate/Ethidium Bromide (FDA/EB) staining technique to assess the viability of Mycobacterium leprae obtained from biopsies of leprosy patients under different periods of treatment . Bacillary suspensions were obtained from skin punch biopsies and stained with the FDA/EB solution . The average percentage of green cells seen which were deemed to be viable were: 67.2% of green cells in patients without previous treatment; 45.6% in patients with 1 to 6 months of treatment; 25.9% for patients with 7 to 12 months of treatment and 10.5% in patients with 13 to 24 months of treatment . All the patients studied were on multidrug therapy . The differences obtained in the percentages of green cells in the different groups of patients were statistically significant as determined by the Wilcoxon's test . The decrease in the percentage of green cells observed with increasing periods of treatment suggests that the FDA/EB technique correlates with the actual viability of M . leprae . The application of this technique in the routine procedures performed with Hansen's disease patients could be very useful for monitoring the effectiveness of treatment in leprosy patients. J Bacteriol, 1991 Apr, 173(7), 2354 - 65 Characterization of the Cephalosporium acremonium pcbAB gene encoding alpha-aminoadipyl-cysteinyl-valine synthetase, a large multidomain peptide synthetase: linkage to the pcbC gene as a cluster of early cephalosporin biosynthetic genes and evidence of multiple functional domains; Gutierrez S et al.; A 24-kb region of Cephalosporium acremonium C10 DNA was cloned by hybridization with the pcbAB and pcbC genes of Penicillium chrysogenum . A 3.2-kb BamHI fragment of this region complemented the mutation in the structural pcbC gene of the C . acremonium N2 mutant, resulting in cephalosporin production . A functional alpha-aminoadipyl-cysteinyl-valine (ACV) synthetase was encoded by a 15.6-kb EcoRI-BamHI DNA fragment, as shown by complementation of an ACV synthetase-deficient mutant of P . chrysogenum . Two transcripts of 1.15 and 11.4 kb were found by Northern (RNA blot) hybridization with probes internal to the pcbC and pcbAB genes, respectively . An open reading frame of 11,136 bp was located upstream of the pcbC gene that matched the 11.4-kb transcript initiation and termination regions . It encoded a protein of 3,712 amino acids with a deduced Mr of 414,791 . The nucleotide sequence of the gene showed 62.9% similarity to the pcbAB gene encoding the ACV synthetase of P . chrysogenum; 54.9% of the amino acids were identical in both ACV synthetases . Three highly repetitive regions occur in the deduced amino acid sequence of C . acremonium ACV synthetase . Each is similar to the three repetitive domains in the deduced sequence of P . chrysogenum ACV synthetase and also to the amino acid sequence of gramicidin synthetase I and tyrocidine synthetase I of Bacillus brevis . These regions probably correspond to amino acid activating domains in the ACV synthetase protein . In addition, a thioesterase domain was present in the ACV synthetases of both fungi . A similarity has been found between the domains existing in multienzyme nonribosomal peptide synthetases and polyketide and fatty acid synthetases . The pcbAB gene is linked to the pcbC gene, forming a cluster of early cephalosporin-biosynthetic genes. Wei Sheng Wu Xue Bao, 1991 Apr, 31(2), 94 - 9 {The study of Bacillus sphaericus Ts-1 protoplast-plasmid transformation of electroporation}; Wang Y et al.; This report gave the best conditions of Bacillus sphaericus Ts-1 protoplast-plasmid pHV33 electroporation . The highest transformation frequency and transformation efficiency induced by three pulse of 21 KV/cm and 10 microseconds duration applied at an interval of one sec., was 2.44 x 10(2) transformants/micrograms DNA and 3.16 x 10(-6) respectively . The saturated concentration of DNA absorbed by the protoplast was 5 micrograms DNA/10(9) cells/ml . By means of this method, pJB417, a recombinant mosquito larvicide clone, was introduced into B . subtilis 168M and B . sphaericus Ts-1 . The transformants of B . subtilis 168M with biocide activity were obtained, but the toxicity of B . sphaericus Ts-1 was not increased. Antibiot Khimioter, 1991 Apr, 36(4), 5 - 8 {Isolation and identification of an antibiotic from culture 8-86}; Melikhov VA et al.; An antibiotic complex was isolated from culture 8-86 referred to Bacillus . The complex consisted of components 8-86A and 8-86B active against gram-negative organisms . By its physico-chemical properties such as IR and UV spectra, amino acid composition, specific rotation and fatty acid composition component 8-86B was shown to be close to polymyxin F. J Mol Biol, 1991 Mar 20, 218(2), 465 - 75 Aromatic-aromatic interactions and protein stability . Investigation by double-mutant cycles; Serrano L et al.; The side-chains of phenylalanine and tyrosine residues in proteins are frequently found to be involved in pairwise interactions . These occur both within repeating elements of secondary structure and in tertiary and quaternary interactions . It has been suggested that they are important in protein folding and stability, and non-bonded potential energy calculations indicate that a typical aromatic-aromatic interaction has an energy of between -1 and -2 kcal/mol and contributes between -0.6 and -1.3 kcal/mol to protein stability . There is such an aromatic pair on the solvent-exposed face of the first alpha-helix of barnase, the small ribonuclease from Bacillus amyloliquefaciens . The edge of the aromatic ring of Tyr17 interacts with the face of that of Tyr13 . The two residues have been mutated both singly and pairwise to alanine, and their free energies of unfolding determined by denaturation with urea . Application of the double-mutant cycle analysis gives an interaction energy of -1.3 kcal/mol for the aromatic pair in the folded protein relative to solvation by water in the unfolded protein . This value is similar to that calculated from the change in surface-accessible area between the rings on the formation of the pair . Analysis of a further double-mutant cycle in which the Tyr residues are mutated to Phe indicates that the aromatic-aromatic interactions of Tyr/Tyr and Phe/Phe make identical contributions to protein stability . However, Tyr is preferred to Phe by 0.3(+/- 0.04) kcal/mol at the solvent-exposed face of the alpha-helix. J Immunol, 1991 Mar 15, 146(6), 1987 - 95 Antigenicity and immunogenicity of a multiple peptidic construction of the Schistosoma mansoni Sm-28 GST antigen in rat, mouse, and monkey . 1 . Partial protection of Fischer rat after active immunization; Wolowczuk I et al.; Among the schistosome proteins characterized as vaccine candidates, an Ag of 28 kDa (Sm-28-GST) has received considerable attention . It was shown to be antigenic in humans and protective in mice, rats, hamsters, and baboons . Synthetic peptides derived from its sequence have been used to characterize the immune response to the molecule and one of these, comprising aminoacids 115-131 has been shown to incorporate both T and B cell recognition sites in a variety of experimental models . An octameric ("octopus") construction of the 115-131 peptide has been synthesized and its antigenicity and immunogenicity have been examined . The octopus construct is immunogenic in rats, mice and baboons in the presence of CFA (for rodents) and Bacille-Calmette-Guerin vaccine (for primates) as adjuvants . This clearly indicates that the construction allowed the conservation of the immune sites of the cognate protein . Moreover, anti-octopus sera from immunized Fischer rats were able to mediate platelet-, macrophage-, and eosinophil-dependent cytotoxicity toward schistosomula . Rats immunized with the 115-131 octopus were partially protected against a challenge infection with Schistosoma mansoni cercariae and this was paralleled by an increased level of IgG and more importantly, of IgE Sm-28-GST-specific antibodies. Eur J Biochem, 1991 Mar 14, 196(2), 493 - 7 The ATPase of Bacillus alcalophilus . Reconstitution of energy-transducing functions; Hoffmann A et al.; The purified ATPase of Bacillus alcalophilus (F1F0) was reconstituted into proteoliposomes by gradual removal of the detergent Triton X-100 with Amberlite XAD-2 . The reconstitution was apparently highly asymmetric with nearly 100% of the F1 portion of the ATPase becoming oriented to the outside . Similar to results obtained with the soluble enzyme, the membrane-bound ATPase required Mg2+ and methanol for maximum activity . With Ca2+ or Mg2+ without methanol, 25% and 1%, respectively, of the maximum activity were observed . The ATPase was unable to pump Na+ ions but catalyzed the translocation of protons into the reconstituted proteoliposomes . Optimum proton translocation required the presence of Mg2+, not Ca2+, as divalent metal ion . The proton pump was inhibited by dicyclohexylcarbodiimide, venturicidin and NaN3 . On incubation of the reconstituted ATPase with {14C}dicyclohexylcarbodiimide, subunit c of the enzyme complex became specifically labeled . The proteoliposomes catalyzed the Mg2(+)-dependent incorporation of {32P}phosphate into ATP by ATP/{32P}phosphate exchange . This exchange was little affected by monensin, but was completely abolished by the uncoupler carbonyl cyanide m-chlorophenylhydrazone . Protons and not Na+ are thus the coupling ions of the ATPase of B . alcalophilus. Am J Clin Pathol, 1991 Mar, 95(3), 418 - 23 Issues in cerebrospinal fluid management . Acid-fast bacillus smear and culture; Albright RE Jr et al.; Meningeal tuberculosis is an uncommon disease in the United States with an annual incidence of fewer than 200 cases . This study evaluates three approaches to improving the use of the cerebrospinal (CSF) acid-fast bacillus (AFB) smear and culture procedure: (1) education alone; (2) optional screening by which physicians can select to have the AFB analysis stopped if the initial CSF findings are unremarkable; and (3) mandatory screening before the performance of all CSF AFB analyses . With education alone, the CSF AFB culture rate decreased from 20.6% of all CSF acquisitions to 15.7% (P less than 0.001); however, the effect may have been related to a decrease in all types of AFB testing . Optional screening had no impact on the AFB testing rate . Mandatory screening significantly decreased the CSF AFB rate to 6.7% (P less than 0.001), unrelated to changes in other types of AFB testing . Laboratories that employ mandatory screening should report the screening results immediately and have a mechanism whereby physicians can bypass the screen, providing CSF AFB analysis on unremarkable fluid from high-risk patients. Cell, 1991 Mar 8, 64(5), 1017 - 23 Barnase toxin: a new chimeric toxin composed of pseudomonas exotoxin A and barnase; Prior TI et al.; We have constructed a chimeric toxin composed of Pseudomonas exotoxin A (PE) and the extracellular ribonuclease of Bacillus amyloliquefaciens, barnase . The chimeric protein, termed PE-Bar, reacted with both anti-PE and anti-barnase antisera and had both ADP ribosylation and ribonuclease activities . The chimeric toxin was cytotoxic to the murine fibroblast cell line L929 and to a murine hybridoma resistant to PE . A mutant form of PE-Bar lacking ADP-ribosylating activity was still cytotoxic to L929 cells . Because treatment of cells prelabeld with {3H}uridine resulted in a decrease in their RNA content, we conclude that this cytotoxic effect was due to the ribonuclease activity of barnase molecules that had been translocated to the cytosol . It is now possible to construct chimeric toxins with two or more enzymatic activities that can be delivered to the cytosol of the target cells. Immunol Lett, 1991 Mar, 27(3), 191 - 7 Cellular cytotoxicity of human natural killer cells and lymphokine-activated killer cells against bladder carcinoma cell lines; Wang MH et al.; The cytotoxicity of natural killer (NK) and lymphokine activated killer (LAK) cells against two human bladder tumor cell lines (BT-A and BT-B) was investigated using a fluorometric assay by labeling tumor cell DNA with Hoechst dye No . 33342 . Our results demonstrate that BT-A and BT-B cells have low sensitivity to the cytotoxic activity of mononuclear cells (MNC) and NK cells . Cytotoxicity of MNC or NK cells against both tumor cell lines is enhanced during co-culture of the effector cells with the target cells, which suggests that BT-A and BT-B cells provide the signals which could activate MNC to exert cytotoxicity . In contrast to NK cells, IL-2-generated LAK cells showed profound cytotoxicity to BT-A and BT-B within 24 h . In addition to cellular cytotoxicity to bladder tumor cells, we also tested the effect of recombinant interleukin 1 beta (rIL-1 beta), recombinant tumor necrosis factor (rTNF), and the supernatants of co-culture of MNC or LAK cells with bladder tumor cells . The results show no cytotoxic or growth-promoting activity of rIL-1, rTNF, or the crude culture supernatants on bladder tumor cells . We found that LAK cells, but not macrophages or NK cells, may play a major role in cellular cytotoxicity against the two bladder tumor cell lines tested . From this finding we conclude that activation of LAK cells may be one important mechanism induced by adjuvant bacillus Calmette-Guerin (BCG) therapy leading to effective prevention of urothelial bladder carcinoma reappearance. J Med Entomol, 1991 Mar, 28(2), 219 - 22 Spiroplasma (Mollicutes: Spiroplasmataceae) pathogenic for Aedes aegypti and Anopheles stephensi (Diptera: Culicidae); Humphery-Smith I et al.; Intrathoracic inoculation with the mosquito spiroplasma, Spiroplasma taiwanense Abalain-Colloc et al., was found to reduce significantly the survival of adult male and female Aedes aegypti (L.) and Anopheles stephensi Liston . This spiroplasma also reduced significantly the flight capacity of adult female Ae . aegypti 5-8 d after inoculation and adult female An . stephensi 4 d after inoculation . Adult female An . stephensi were incapable of flight 5 d after inoculation . As such, S . taiwanense joins Bacillus thuringiensis and B . sphaericus as bacteria known to be pathogenic for mosquito vectors. Pharm Res, 1991 Mar, 8(3), 341 - 4 Production of 5-15 microns diameter alginate-polylysine microcapsules by an air-atomization technique; Kwok KK et al.; A novel method of preparing small-sized microcapsules using a Turbotak air-atomizer is reported . Alginate-polylysine microcapsules containing Bacillus Calmette Guerin vaccine have been prepared by an adaptation of the method of Lim (1) which allows the manufacture of small-sized microcapsules . A Turbotak is used to spray sodium alginate solution into calcium chloride solution to form temporary calcium alginate microgel capsules . These temporary microgel droplets are subsequently cross-linked with polylysine to form permanent membranes . Microcapules in the size range of 5-15 microns have been produced which can be compared to an average diameter of greater than or equal to 300 microns obtained by the method reported by Lim . The microcapsule size is dependent on the conditions of operation of the Turbotak and the concentration of the sodium alginate solution . Particles within the size range 5-15 microns can be reproducibly manufactured using the conditions of operation reported here . Other size ranges below the minimum of 300 microns reported by Lim are also feasible using this technique. J Am Mosq Control Assoc, 1991 Mar, 7(1), 56 - 62 Microbial larvicides for the control of nuisance aquatic midges (Diptera: Chironomidae) inhabiting mesocosms and man-made lakes in California; Rodcharoen J et al.; Bacillus thuringiensis var . israelensis (B.t.i.) and B . sphaericus were evaluated for chironomid larvicidal activity in freshwater mesocosms . Of two B.t.i . formulations, the technical powder ABG-6164 provided excellent control of chironomines (94%) at the rate of 11.2 kg/ha whereas the liquid concentrate, Vectobac 6 AS, achieved only moderate control (57%) at the rate of 22.4 kg/ha . In contrast, similar rates of B . sphaericus products, ABG-6184 technical powder and BSP-2 flowable concentrate, produced no significant reduction . Lake studies were initiated to determine which B.t.i . formulations were practical and effective in large-scale situations . Two sinking granular corn grit formulations worked with varying degrees of success but were deemed too bulky for extensive applications . B.t.i . technical powder preparations with high potency ratings of 5,000 and 12,430 ITU/mg gave excellent control of Chironomus spp . (100% and 87%) when used at the rates of 6.7 and 2.8 kg/ha, respectively . None of the treatments reduced larval populations of the tanypodines, Procladius and Tanypus. J Gen Microbiol, 1991 Mar, 137 ( Pt 3), 667 - 77 Identification of four unique clones encoding 10 kDa proteins from Bacillus that cause phenotypic complementation of a phoA mutant strain of Escherichia coli; Lee JW et al.; A number of clones have been isolated from two Bacillus species which complement the PhoA- phenotype of Escherichia coli mutants under conditions that induce the expression of alkaline phosphatase (APase) . These clones were initially thought to carry XPases because the transformed host could hydrolyse a common APase substrate, XP (5-bromo-4-chloro-3-indolyl-phosphate) . The sequences of the open reading frames responsible for the phenotypic complementation showed no sequence similarity to ATPases of E . coli, human (bone-liver-kidney, intestinal or placental) or Bacillus . Therefore, these clones were designated as XPA (for X Phosphatase Activity) clones . Four of the clones encoded small (10 kDa), basic, hydrophobic proteins . Two of these, xpaB from B . subtilis 168 and xpaL2 from B . licheniformis MC14, shared 62% identity at both the DNA and the predicted amino acid sequence level . The fact that homologues from two Bacillus strains were cloned indicated that the screen was specific, but not for APase genes . It is clear that phenotypic complementation with cloned DNA from another genus does not ensure the identification of an APase gene . Possible mechanisms for the abnormal phenotypic complementation are discussed. J Am Vet Med Assoc, 1991 Mar 1, 198(5), 862 - 3 Prolonged milk residue in two cows after subcutaneous injections of penicillin at an extra-label dose; Krainock RJ; After cesarian section was done on 2 Holstein cows, procaine penicillin G was injected SC at an extra-label-dose . The milk contained inhibitory residue for 21 days (case 1) and 10 days (case 2) after the final penicillin injection . The Bacillus stearothermophilus disk assay was used to confirm presence of an inhibitor . Analysis to specifically identify the inhibitor substance was not performed . Other sources of residues were examined and excluded in these cases . Procaine penicillin G was assumed to be the residue substance . Delayed drug clearance was attributed to depot formation following high-dose SC penicillin administration. Gene, 1991 Mar 1, 99(1), 109 - 14 Promoter sequence analysis in Bacillus and Escherichia: construction of strong promoters in E . coli; Yamada M et al.; Many derivatives of the nprM promoter of Bacillus stearothermophilus and the strong early promoter, A3, of coliphage T3 were designed and chemically synthesized . These promoters consisted of some or all of the AT box, consensus sequence, tac promoter sequence, spacer, and lac operator . The promoter activities were assessed by their ability to express the cat gene in Escherichia coli . One of the derivatives of the A3 promoter, which contained the lac operator, was much stronger (about 3.5 times) than the tac promoter . The promoter activities in E . coli were considerably modified by the substitutions in the -43 (AT box) and -35 regions. Eur J Immunol, 1991 Mar, 21(3), 793 - 7 Mycobacterial proliferation in macrophages is prevented by incubation with lymphocytes activated in vitro with a mycobacterial antigen complex; Beschin A et al.; Antigen A60 from Mycobacterium bovis bacillus Calmette Guerin was shown to trigger both humoral and cellular immune reactions . We explored the ability of A60 to block intracellular proliferation of phagocytosed mycobacteria with a model system involving peritoneal murine macrophages infected with Mycobacterium avium . Mixed lymphocytes from lymph nodes of mice inoculated with A60 hindered intracellular proliferation of this mycobacterium, owing to A60-specific cells, proliferation of which was induced in vitro in an antigen concentration-dependent manner . The lymphokines released by A60-stimulated T lymphocytes in vitro were identified as interleukin 2 (IL2) and interferon-gamma (IFN-gamma): their production showed a clear A60 dose dependence . When supernatants of such induced lymphocyte cultures were incubated with anti-IFN-gamma antibodies, macrophage activation was prevented, whereas anti-IL 2 immunoglobulin had little effect . Treatment of infected macrophages with recombinant IFN-gamma reduced intracellular proliferation of mycobacteria, while exogenous IL 2 and tumor necrosis factor were ineffective . Therefore, A60 elicits in vitro proliferation of T lymphocytes responding specifically to this antigen with production of IFN-gamma, which in turn activates macrophages and prevents multiplication of phagocytosed mycobacteria. Biochem J, 1991 Mar 1, 274 ( Pt 2), 355 - 60 Purification and properties of adenosine 5'-phosphosulphate sulphotransferase from Euglena; Li JY et al.; Adenosine 5'-phosphosulphate sulphotransferase (APSST) was extracted from Euglena gracilis Klebs var . bacillaris mutant W10BSmL by freezing and thawing and was purified about 10,000-fold (to homogeneity) with 10.5% recovery by (NH4)2SO4 precipitation, Sephadex G-100 chromatography, Reactive Blue-agarose, Reactive Dye-agarose, DEAE-cellulose, preparative isoelectric focusing and non-inactivating SDS/PAGE . The active APSST, with a molecular mass of 102 kDa and multiple forms from pI 5.0 to 5.5, is a tetramer held together by covalent (probably disulphide) bonds . An apparent Km of the purified enzyme for adenosine 5'-phosphosulphate (APS) of 0.1 microM is obtained when dithiothreitol is used as the thiol . The enzyme is stimulated by Mg2+, Ca2+ or Ba2+, and uses almost any thiol; dithiothreitol and dithioerythritol give the highest activity . In the absence of APS, the enzyme is inactivated (and is rendered monomeric) by thiols but is protected from thiol inactivation by AMP, adenosine 5'-phosphoramidate (APA) or adenosine 5'-monosulphate (AMS), which also inhibit APSST activity somewhat . The enzyme resists inactivation by SDS in the absence of thiols; SDS stimulates APSST activity at low concentration, but high concentrations are inhibitory. Am Rev Respir Dis, 1991 Mar, 143(3), 496 - 500 Cellular immunity in current active pulmonary tuberculosis; Andrade-Arzabe R et al.; A group of 10 patients with recently diagnosed pulmonary TB were studied and compared to 10 bacillus Calmette-Guerin (BCG) immunized healthy individuals . Cellular immune mechanisms were explored in vitro utilizing fresh and precultured peripheral blood mononuclear cells exposed to PHA, PPD, and recall antigens (SK/SD and CA) . Proliferative assays were also carried out in the presence of either each patient's serum (autologous serum) or cocultured with CD3(+)-depleted adherent cells . Serum measurements of soluble interleukin-2 (IL-2) receptor and synthesis of IL-2 generated by mononuclear cells stimulated with PPD and SK/SD were also performed . Patient sera were able to inhibit autologous as well as allogeneic cell responses, and a significant adherent cell suppressive effect was observed . As a whole the group of patients showed decreased blast transformation to PPD, preserved proliferative responses to other recall antigens, and a low PPD-induced generation of IL-2 . Furthermore, as possible evidence of preactivated T cells, these patients demonstrated high soluble IL-2 receptor serum levels . Early compromise of specific cell-mediated immunity, including IL-2 abnormalities, may be of significance in newly diagnosed pulmonary TB. J Urol, 1991 Mar, 145(3), 498 - 501 Oral or intravesical bacillus Calmette-Guerin immunoprophylaxis in bladder carcinoma; D'Ancona CA et al.; A total of 71 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 3 treatment groups: control group--transurethral resection discontinued within the study, oral bacillus Calmette-Guerin (BCG) group--transurethral resection of bladder tumor plus BCG (Moreau) and intravesical BCG group--transurethral resection of bladder tumor plus BCG . Of 9 patients in the control group 8 (89%) experienced tumor recurrence during a mean followup of 20 months . Of the 28 patients in the oral BCG group 11 (39.3%) had recurrence during a mean followup of 36 months . Of the 34 patients in the intravesical group 6 (18%) had recurrence in a 24-month mean followup . The incidence of complications was higher in the intravesical (41.2%) than in the oral BCG group (28.5%) . These results show that intravesical BCG is a more effective immunotherapy; however, oral BCG can be used in patients who do not accept intravesical BCG administration. Southeast Asian J Trop Med Public Health, 1991 Mar, 22(1), 108 - 12 Fermentation of a Malaysian Bacillus thuringiensis serotype H-14 isolate, a mosquito microbial control agent utilizing local wastes; Lee HL et al.; A screening program searching for indigenous microbial control agents of mosquitos in Malaysia is initiated since 1987 and to date at least 20 isolates of mosquitocidal Bacillus thuringiensis serotypes have been obtained . Preliminary field evaluation of several isolates indicated that they are highly effective in the control of medically important mosquito species . For operational purposes, there is an urgent need to produce this agent utilizing cheap and locally available wastes through fermentation biotechnology . Fermentation studies in shake-flasks containing standard nutrient broth and soya bean waste, respectively, indicate that it takes about 37 hours for a Malaysian isolate of B . thuringiensis serotype H-14 to mature . In the grated coconut waste, fishmeal and rice bran, the bacteria took 28 hours, 26 hours and 126 hours respectively to mature . The endotoxin was harvested from the standard nutrient broth at 55 hours and at 50 hours from soybean, grated coconut waste and fishmeal . The endotoxin could only be harvested 150 hours after inoculation from rice bran medium . However, no bacterial growth was detected in palm oil effluent . In terms of endotoxin and biomass production, fishmeal appears to be a suitable medium . Variations in the pH of the fermenting media were also noted. FEMS Microbiol Lett, 1991 Mar 1, 62(2-3), 253 - 7 One-step purification procedure for UDP-N-acetylmuramyl-peptide murein precursors from Bacillus cereus; Kohlrausch U et al.; A method is described for the rapid isolation of the activated murein precursors UDP-N-acetyl-muramyl-pentapeptide (UDP-MurNAc-pentapeptide) and UDP-MurNAc-tripeptide from Bacillus cereus . After accumulation of the precursors by inhibition of murein synthesis either in the presence of vancomycin (for the pentapeptide precursor) or D-cycloserine (for the tripeptide precursor) the cells were extracted with boiling water . Prior to high pressure liquid chromatography the material was freed from acid precipitable material . UDP-MurNAc-penta- and tripeptide were separated from other components by reversed-phase HPLC on Hypersil ODS using isocratic elution conditions with sodium phosphate buffer . The precursors were obtained with at least 98% purity and a yield of about 50 mumol from a 10-l culture of B . cereus. Thorax, 1991 Mar, 46(3), 220 - 1 Successful treatment of Bacillus cereus infection with ciprofloxacin; Gascoigne AD et al.; Bacillus cereus is rarely a pulmonary pathogen but may cause pneumonia in immunocompromised patients . A patient with bronchiectasis and no recognisable immunodeficiency had this organism isolated during two infective exacerbations, once from respiratory secretions and once by blood culture . Ciprofloxacin treatment was effective on both occasions. J Bacteriol, 1991 Mar, 173(5), 1748 - 56 Effect of a 20-kilodalton protein from Bacillus thuringiensis subsp . israelensis on production of the CytA protein by Escherichia coli; Visick JE et al.; CytA, a 27-kDa cytolytic crystal protein of Bacillus thuringiensis subsp . israelensis, is produced only at very low levels by recombinant Escherichia coli cells unless a 20-kDa B . thuringiensis subsp . israelensis protein is also present (K . M . McLean and H . R . Whiteley, J . Bacteriol . 169:1017-1023, 1987; L . F . Adams, J . E . Visick, and H . R . Whiteley, J . Bacteriol . 171:521-530, 1989) . However, the data reported here demonstrate that the 20-kDa protein is not required for high-level CytA production in E . coli strains carrying mutations in rpoH, groEL, or dnaK, all of which affect the proteolytic ability of the cells . The 20-kDa protein also increases the amount of CryIVD (another B . thuringiensis subsp . israelensis crystal protein) and LacZX90 (a mutant of beta-galactosidase) made by E . coli . The latter phenomenon is attributable to an increase in the half-life of LacZX90, suggesting that the 20-kDa protein may stabilize this protein . The effect of the 20-kDa protein was also examined in vitro and in a T7 RNA polymerase expression system, and the possible significance of these results for the timing of proteolysis and of 20-kDa protein activity is discussed . Finally, the ability of a single antibody to coimmunoprecipitate CytA and the 20-kDa protein from E . coli extracts provides evidence for a protein-protein interaction that may be related to the mechanism of action of the 20-kDa protein. Cesk Gynekol, 1991 Mar, 56(2), 93 - 6 {Problems of epidemiology and diagnosis in the vaginal area . II . Measurement of vaginal pH values and its relation to vaginal diseases}; Unzeitig V et al.; The authors divided a group of 600 women into 7 sub-groups based on the results of microscopic examination of native preparations, and using indicator papers of MERCK Co . (art . no . 9542) they assessed values of the vaginal pH . The highest values were recorded in a group of women with suppurative bacterial inflammations (mean pH 5.68) and the lowest ones in women with the finding of fungi (mean pH 5.0), cocobacillary flora without leucocytes (mean pII 4.39) and the normal physiological finding of Dorderlein's bacillus (mean pH 4.15) . Values higher than 4.8 rule out the possibility of a normal finding in the vagina, lower values, however, do not rule it out entirely . Assessment of pH values in the vagina should be considered part of routine examinations of the vaginal environment. Pediatr Dermatol, 1991 Mar, 8(1), 21 - 4 Childhood leprosy in northern India; Kaur I et al.; During a period of eight years, 132 new leprosy cases were detected in children ages 3 to 19 years . Borderline tuberculoid leprosy was present in 59%, tuberculoid in 7.6%, and indeterminate type in 3.8% patients . Single skin lesions were seen in a significant number (43.9%) of patients . Bacillus-positive disease was detected more often (17.4%) than in adults . A high frequency (66.6%) of nerve involvement was also detected . Deformities were uncommon . Males were more often affected than females, especially in the ages 10 to 14 and 15 to 19 years . A history of contact was available in only 19.7% patients, and the contact was intrafamilial in 84.6%. Bull Int Union Tuberc Lung Dis, 1991 Mar, 66(1), 33 - 6 Government intervention programs in HIV/tuberculous infection . Outline of guidelines for national tuberculosis control programs in view of the HIV epidemic; Kochi A; Tuberculosis is one of the most widespread infections known in the world . WHO estimates that in 1990, 1.7 billion people, or one third of the world population, are or have been infected with the tubercle bacillus . Fortunately, few of those infected develop active forms of the disease but it is estimated that in 1990, there will be 8 million new cases and 2.9 million deaths from tuberculosis in the world . This already alarming situation of the tuberculosis problem is getting worse, mainly due to the AIDS epidemic . A basic understanding of tuberculosis/HIV epidemiology is necessary and priority actions are to be strongly recommended for application in government intervention programs . They are specified in the present article. Indian J Med Res, 1991 Mar, 93, 111 - 4 Isolation of mosquito-pathogenic Bacillus sphaericus & B . thuringiensis from the root surface of hydrophytes; Manonmani AM et al.; Attempts were made to isolate B . sphaericus and B . thuringiensis active against mosquito larvae from the root surface of hydrophytes . Out of 139 samples processed, 86 B . sphaericus and 23 B . thuringiensis isolates were obtained . Sixty two of the B . sphaericus isolates belonged to the serotype H5a5b, 2 to H6 and 22 isolates did not agglutinate with any of the 6 antisera tested . Twenty of the B . thuringiensis isolates belonged to the H14 serotype, 1 each to the H10 and H17 serotype(s) and 1 to an unknown serotype . Fifty nine of the B . sphaericus and 20 of the B . thuringiensis isolates fall under highly toxic category with the LC50 dose of 1-50 ng/ml for Culex quinquefasciatus third instar larvae. J Invertebr Pathol, 1991 Mar, 57(2), 149 - 58 Cloning and expression of Bacillus thuringiensis israelensis delta-endotoxin DNA in B . sphaericus; Bar E et al.; Bacillus thuringiensis israelensis delta-endotoxin genes were cloned into Bacillus sphaericus 2362, producing stable transformants reacting with antibody to the 28- and 65-kDa B . thuringiensis israelensis crystal proteins and approximately 10 times more toxic to Aedes mosquito larvae than the original host strain . The LC50 after 48 hr of exposure of Aedes larvae to the most active transformed clone was 0.19 microgram/ml, compared with an LC50 of 1.9 microgram/ml for B . sphaericus 2362 and less than 0.1 microgram/ml for B . thuringiensis israelensis . The cloning vector, plasmid pPL603E, was also effective in transforming B . subtilis 1E20 with B . thuringiensis israelensis DNA, producing highly toxic clones with less stable gene expression than the clones of B . sphaericus. J Exp Med, 1991 Mar 1, 173(3), 751 - 4 Listeria monocytogenes mutants lacking phosphatidylinositol-specific phospholipase C are avirulent; Camilli A et al.; A number of bacterial species secrete phosphatidylinositol-specific phospholipase C (PI-PLC) . In this report, we show that the facultative intracellular bacterial pathogen, Listeria monocytogenes, contains a gene, plcA, predicting a polypeptide with 31% amino acid identity to a Bacillus thuringiensis PI-PLC . Accordingly, L . monocytogenes secretes PI-PLC activity, while a mutant with a transposon insertion in plcA lacks detectable PI-PLC activity . In addition, expression of plcA in B . subtilis resulted in secretion of PI-PLC activity . The L . monocytogenes PI-PLC-defective mutant was three logs less virulent for mice and failed to grow in host tissues . The mutant was also defective for in vitro growth in mouse peritoneal macrophages . These results strongly suggest that PI-PLC is an essential determinant of L . monocytogenes pathogenesis . Whether the PI-PLC acts on a bacterial or host substrate remains to be determined. J Neurochem, 1991 Mar, 56(3), 782 - 8 Purification and characterization of neuron-specific surface antigen defined by monoclonal antibody BM88; Patsavoudi E et al.; Monoclonal antibody BM88 recognizes a neurospecific surface antigen in the CNS and the PNS . In the present study, the antigen recognized by BM88 was immunopurified from pig brain and shown to be a 22-kDa polypeptide by reducing sodium dodecyl sulfate-polyacrylamide gel electrophoresis . Under nonreducing conditions a protein of 40 kDa was obtained, a result indicating that the antigen is composed of two polypeptide chains of equal molecular weight linked by disulfide bridges . Gel filtration of the purified antigen in the presence of Emulphogene suggested that it may be either a monomeric or a dimeric protein . However, in the presence of Triton X-100 a monomeric structure was implied . N-Glycanase digestion indicated that the protein is probably not glycosylated . The purified antigen was characterized as an integral membrane protein by hydrophobic chromatography and phase-separation experiments with Triton X-114 . The antigen, or at least the antibody binding region of the molecule, is very susceptible to protease attack, as judged by protease digestion experiments on brain membranes . By using very low concentrations of papain combined with short incubation times, the antigen was converted to a 16.3-kDa membrane-associated polypeptide as assessed by immunoblotting . This polypeptide contained the BM88 binding epitope . Soluble BM88 immunoreactive polypeptides were not obtained . Bacillus cereus phospholipase C was also unable to solubilize the antigen from the membrane . Our results suggest that the molecule, possessing at least one small extramembranous domain, is attached to the membrane via a polypeptide chain. J Am Mosq Control Assoc, 1991 Mar, 7(1), 24 - 9 Small-scale field trials of Bacillus sphaericus (strain 2362) formulations against Mansonia mosquitoes in Malaysia; Yap HH et al.; Five formulations of Bacillus sphaericus (strain 2362) including aqueous suspension BSP 1, BSP 2, technical powder ABG 6184, corncob granules ABG 6185 (potencies 2 x 10(10), 2 x 10(7), 9.5 x 10(10), 5 x 10(10), 5 x 10(10) spore/g, respectively) and wettable powder ABG 6232 (1,000 BS ITU/mg) were tested against laboratory-cultured late third/early fourth instar larvae of Mansonia uniformis in floating screened cages in small plots at swampy ditches on Penang Island, Malaysia . Mean dosage/response values at 90% mortality levels were 6.93, 95.32, 1.45, 11.92 and 2.86 liters or kg per ha, respectively, for the formulations tested . There were practically no residual effects for the formulations tested with larvae introduced at 48, 96, and 168 h post-treatment . In trials of BSP 1, ABG 6184 and ABG 6185 (1 liter or 1 kg per ha) against immature Mansonia spp . in impounded paddy field ditches, improved efficacy and residual effects were obtained with mean reductions of 93.1, 91.9 and 80.4% at days 3, 7 and 14 posttreatment, respectively. FEMS Microbiol Lett, 1991 Mar 1, 62(2-3), 293 - 6 Uptake of inorganic pyrophosphate by Bacillus megaterium; Villiotte PJ et al.; Cells of Bacillus megaterium take up inorganic pyrophosphate, employing a saturable carrier which is sensitive to sulfhydryl reagents, orthophosphate, and arsenate . Uptake is stimulated by proton ionophores, including CCCP and nigericin, indicating that proton cotransport can lead to an opposing gradient . Inhibitor sensitivity, as well a relatively high Km for inorganic pyrophosphate render it likely that uptake is mediated by an orthophosphate transport system. J Biol Chem, 1991 Feb 25, 266(6), 3955 - 60 Purification and characterization of an activator protein for methanol dehydrogenase from thermotolerant Bacillus spp; Arfman N et al.; All thermotolerant methanol-utilizing Bacillus spp . investigated by us possess a NAD-dependent methanol dehydrogenase (MDH) activity which is stimulated by a protein present in the soluble fraction of Bacillus sp . C1 cells . This activator protein was purified to homogeneity from Bacillus sp . C1 cells grown at a low dilution rate in a methanol-limited chemostat culture . The native activator protein (Mr = 50,000) is a dimer of Mr = 27,000 subunits . The N-terminal amino acid sequence revealed no significant similarity with any published sequences . Stimulation of MDH activity by the activator protein required the presence of Mg2+ ions . Plots of specific MDH activity versus activator protein concentration revealed Michaelis-Menten type kinetics . In the presence of activator protein, MDH displayed biphasic kinetics (v versus substrate concentration) toward C1-C4 primary alcohols and NAD . The data suggest that in the presence of activator protein plus Mg2+ ions, MDH possesses a high affinity active site for alcohols and NAD, in addition to an activator- and Mg2(+)-independent low affinity active site . The activation mechanism remains to be elucidated. J Mol Biol, 1991 Feb 20, 217(4), 731 - 6 Accurate measurements of coupling constants from two-dimensional nuclear magnetic resonance spectra of proteins and determination of phi-angles; Ludvigsen S et al.; A new and simple method to measure 3JHNH alpha coupling constants of proteins by adding and subtracting traces from corresponding two-dimensional nuclear Overhauser enhanced spectroscopy and two-dimensional correlated spectroscopy cross peaks after scaling is proposed . The optimal scaling for the addition and the subtraction of the two traces is obtained by minimizing an error function . The method was proven to give accurate and precise measurements of coupling constants when tested with a series of simulated spectra . The accuracy of the method was better than 0.1 Hz for all test cases including the limiting case of J = 2.0 Hz and line-width = 11.0 Hz . The accuracy of the method was better than 0.1 Hz for all test cases including The 3JHNH alpha coupling constants were measured in two-dimensional nuclear magnetic resonance spectra of the two proteins barley serine proteinase inhibitor (CI-2) and the bacterial ribonuclease (barnase) of Bacillus amyloliquefaciens . The experimentally measured coupling constants were used to calculate the constants in a Karplus equation to be: 3JHNH alpha = 6.7 cos2(phi-60) -1.3 cos(phi-60) +1.5 . These constants are in good accordance with those obtained for basic pancreatic trypsin inhibitor (BPTI) . In addition, special emphasis is given to the measurements of positive phi-angles, and to the contribution of molecular dynamics on the apparent coupling constants. J Mol Biol, 1991 Feb 20, 217(4), 737 - 50 Structure of cyclodextrin glycosyltransferase refined at 2.0 A resolution; Klein C et al.; The previously reported structural model of cyclodextrin glycosyltransferase (EC 2.4.1.19) from Bacillus circulans has been improved . For this purpose the known sequence was built into an electron density map established by multiple isomorphous replacement and subsequent solvent-flattening at 2.5 A resolution . The resulting model was refined at 2.0 A resolution using a simulated annealing refinement method . Based on 70,171 independent reflections in the range 7.0 to 2.0 A resolution, a final R-factor of 17.6% was obtained with a model obeying standard geometry within 0.013 A in bond lengths and 2.7 degrees in bond angles . The final model consists of all 684 amino acid residues, two calcium ions and 588 solvent molecules. Biochim Biophys Acta, 1991 Feb 15, 1076(3), 364 - 8 Aminoacylase I is not a glycolipid-anchored ectoenzyme in pig kidney; Greenhough KJ et al.; Subcellular fractionation of pig kidney cortex revealed that aminoacylase I (EC 3.5.1.14, N-acyl-L-amino-acid aminohydrolase) is predominantly a soluble enzyme with only 0.5% of the total activity being recovered in the membrane fraction . The aminoacylase I activity associated with the membrane preparations displayed neither rapid release following incubation with phosphatidylinositol-specific phospholipase C from Bacillus thuringiensis nor the distinctive differential pattern of detergent solubilization which was seen with glycosyl-phosphatidylinositol-anchored proteins (renal dipeptidase, alkaline phosphatase) . When fractionated by phase separation in Triton X-114, integral membrane proteins of kidney microvillar membranes partitioned predominantly (greater than 90%) into the detergent-rich phase . In contrast, only 3.7% of aminoacylase I activity associated with microvillar membranes partitioned into the detergent-rich phase . Aminoacylase I activity of pig kidney would therefore appear to be a hydrophilic protein in nature and is not, as suggested previously, a G-PI-anchored integral membrane protein. Biochem Biophys Res Commun, 1991 Feb 14, 174(3), 1077 - 83 Induction of group II-like phospholipase A2 by lipopolysaccharide in the liver of BCG-primed rat; Inada M et al.; The specific activity of phospholipase A2 (PLA2) in the liver homogenate was elevated 1.7-fold in bacillus Calmette-Guerin (BCG)-treated rats, 1.6-fold in lipopolysaccharide (LPS)-treated rats, and 2.4-fold in BCG-infected rats treated with LPS, compared with that of control rats . These increased activities were almost completely inhibited by the antibody directed against rat splenic group II PLA2 (PLA2M) but not by anti-pancreatic PLA2 antibody . The results of immunoblot analysis confirmed that the PLA2 immunochemically related to the group II enzyme was induced by treatment with BCG and/or LPS . The anti-PLA2M antibody-inhibitable PLA2 activity per a single cell was elevated not only in nonparenchymal cell fraction but also in hepatocyte fraction, as in the case of whole liver . On the contrary, the PLA2 concentration and its specific activity did not change by the same treatment both in spleen homogenate and in isolated spleen cell fractions although a 3-fold increase in spleen mass occurred by BCG treatment . These results suggested that a tissue-specific mechanism of the PLA2 induction by these inflammatory mediators may operate in liver. Eur J Biochem, 1991 Feb 14, 195(3), 631 - 5 Two structural domains as a general fold of the toxic fragment of the Bacillus thuringiensis delta-endotoxins; Convents D et al.; The unfolding by guanidine hydrochloride of the toxic fragment of a Bacillus thuringiensis toxin belonging to the CryIC class reveals a two-step denaturation under both acid and alkaline conditions . This demonstrates the existence of two structural domains as building blocks for this toxin . Protease digests performed on a CryIA(b) and CryIC B . thuringiensis toxin, under native and partially denatured conditions, confirm this conclusion . Whereas the native CryIC toxin is completely protease resistant, the CryIA(b) toxin, earlier described as consisting of two structural domains {Convents, D., Houssier, C., Lasters, I . & Lauwereys, M . (1990) J . Biol . Chem . 265, 1369-1375}, is cleaved by three proteases, resulting in at least two common fragments . This suggests that this toxin is built up of two globular units linked by a protease-susceptible linker . The detection of a stable intermediate along the denaturation curve allows us to study and compare the consecutive unfolding of the structural domains for both toxins . By addition of a protease, under conditions where such an unfolding intermediate exists, a single denaturation phase can be assigned to a specific part of the protein . These experiments lead to the conclusion that the domain whose stability is highly dependent on pH corresponds to the N-terminal half of both toxins. FEBS Lett, 1991 Feb 11, 279(1), 123 - 31 Modulation of protease specificity by a change in the enzyme microenvironment . Selectivity modification on a model substrate, purified soluble proteins and gluten; Hertmanni P et al.; Subtilisin BPN' activity on a synthetic substrate is found to decrease with the concentration of soluble additives such as sugars and polyols, the catalytic efficiency of the enzyme being related to the water activity in the reaction medium . Limited hydrolysis of B chain of insulin is followed and the cleavage priority determined . When carried out in glycerol-containing medium, both enzyme catalytic behaviour and specificity are perturbed; a different cleavage order and a selectivity restriction are observed . The experiments were generalised to purified proteins and to an insoluble protein complex . The hydrolysis kinetics of purified gliadins by pepsin and of gluten by a Bacillus neutral protease are modulated in presence of water activity depressors . Glycerol is able to increase both pepsin efficiency and gluten protein solubility . The hydrolysis order is affected by water-structuring molecules in the enzyme microenvironment and new peptides appear whatever the size and initial solubility of the substrate. Chem Pharm Bull (Tokyo), 1991 Feb, 39(2), 417 - 20 Post-translational processing of tumor necrosis factor production; Kitahara-Tanabe N et al.; To elucidate the mechanism of tumor necrosis factor (TNF) production, we analyzed proteins produced in macrophages sharing the epitope of TNF according to the priming and triggering of TNF production . Rabbit alveolar macrophages primed with Bacillus Calmette-Guerin (BCG) were isolated and cultured in vitro with 35S-methionine, and the proteins produced were analyzed using anti-rabbit TNF monoclonal antibody . Primed with BCG, alveolar macrophages synthesized two proteins with molecular sizes of 50 and 17 kilodaltons (kDa) (p50 and p17) sharing the same epitope with mature TNF within the cells . These two proteins were released into the medium where other proteins were detected without TNF-activity . Cultured with lipopolysaccharide (LPS triggering), the primed alveolar macrophages released TNF-activity into the medium where p17 together with many larger proteins was detected by immunoprecipitation . In vitro translation of messenger ribonucleic acid (mRNA) from BCG-primed macrophages showed that primary TNF has a molecular size of 28 kDa (p28) . These results suggest that active TNF of p17 is secreted when triggered via post-translational processing of the precursor molecules synthesized through priming with BCG. Curr Opin Immunol, 1991 Feb, 3(1), 91 - 6 Leprosy and cell-mediated immunity; Kaplan G et al.; The intradermal injection of the purified protein derivative of tuberculin into lepromatous leprosy patients leads to a local cell-mediated immune response and to the extensive destruction of Mycobacterium leprae . This local response also occurs after intradermal injection of recombinant human interleukin-2; when administered over an 8-day period interleukin-2 evokes a systemic cell-mediated immune response and a reduction in the bacillary burden. Curr Opin Rheumatol, 1991 Feb, 3(1), 145 - 54 Rheumatic manifestations of neoplasia; Schwarzer AC et al.; Neoplasia in an important cause of rheumatic disease . The mechanisms are either direct infiltration of the bone and joint, or indirect infiltration with manifestations distant from the site of neoplastic involvement . Many of the reports reviewed in this article center on direct associations . In particular, there is a report on polymyositis and dermatomyositis and the link with neoplasia . Cases of reflex sympathetic dystrophy are also described in association with neoplasia . There is further discussion on the link of hairy cell leukemia and vasculitis . Other case reports highlight the multiple associations of musculoskeletal disease and neoplasia . These reports include patients with subcutaneous sarcoidosis, ankylosing spondylitis, polychondritis, and systemic sclerosis . Articular manifestations of benign pleural fibromas are described . The existence of these reports, however, does not constitute proof of a causal relationship and the possibility of chance occurrences of two conditions must be considered . Finally, various therapies for cancer are associated with rheumatologic manifestations . Intravesical bacillus Calmette-Guerin has been found to cause an inflammatory polyarthritis . This form of arthritis is similar to experimentally induced adjuvant arthritis in rats that follows immunization with Freund's adjuvant containing Mycobacterium tuberculosis. An Esp Pediatr, 1991 Feb, 34(2), 129 - 31 {Contacts of children with tuberculous patients}; Castan Vidal ML et al.; One hundred and nine adults recently diagnosed of active pulmonary tuberculosis (all of them with smear positive sputum) were selected . Their household contacts under fifteen years of age were studied . 73.1 por 100 of the children were tuberculin-positive, and 33.1 por 100 of these "reactors" had developed a pulmonary tuberculosis themselves . The bacillary density in the sputum of the source case was correlated to percentage of infected and ill children among his contacts . Neonatal vaccination with BCG showed a protective effect against the illness in children under eight years of age. Clin Invest Med, 1991 Feb, 14(1), 55 - 62 Influence of biological response modifiers of bacterial origin on disease progression in the MRL-lpr model of systemic lupus erythematosus; Hart DA et al.; Murine models of systemic lupus erythematosus exhibit some, but not all of the characteristics of human disease . Disease progression in the animal models is related to autoantibodies, genetics, and inflammatory processes . In this report the effects of two bacterial biological response modifiers (BRM) on disease progression in the MRL-lpr model were investigated . The two BRM tested were C . parvum and Bacillus-Calmette-Guerin (BCG), both of which are stimulators of the reticuloendothelial system and both of which have been shown by others to influence disease progression in NZB/W mice . Treatment of 10-week-old mice with C . parvum led to transient alterations in hepatosplenomegaly and plasma proteinase regulation, which then returned to control values . Treatment with BCG led to even more transient effects on the mice . Neither BRM appeared to impact on disease-associated alterations in autoantibody titres, hepatosplenomegaly, or elevations in plasma proteinase activity . Likewise, treatment of 17-week-old MRL-lpr mice with C . parvum did not influence disease progression as evidenced by survival, autoantibody production, or hepatosplenomegaly . Therefore, in contrast to the NZB/W strain, treatment of the MRL-lpr strain with these BRM does not appear to impact on disease progression . This difference may be due to the influence of the lpr accelerator gene in this model. AIDS, 1991 Feb, 5(2), 195 - 9 Bacillus Calmette-Guérin immunization in infants born to HIV-1-seropositive mothers; Lallemant-Le Coeur S et al.; During the prospective follow-up of 64 babies at risk for perinatal HIV-1 infection because their mothers were seropositive, and of 130 control babies whose mothers were seronegative, we studied the occurrence of complications of bacillus Calmette-Guerin (BCG) immunization and its ability to induce cutaneous reactivity to tuberculin . Babies born both to HIV-1-positive and HIV-1-negative mothers received BCG immunization during their first month of life according to the Expanded Programme on Immunization (EPI) recommendations . Local and regional complications of BCG vaccine were looked for at 3, 6 and 9 months after inoculation . A tuberculin skin test was performed at 6 or 9 months of age . Most babies born to HIV-1-positive mothers were later classified as infected or uninfected according to their clinical condition and/or serological status at 18 months of age . The mean duration of the follow-up was 36 months (range 30-40 months) . No chronic or deep ulcerations at the site of injection or disseminated forms of BCG infection were observed . The frequency of BCG-related lymphadenitis in the group of HIV-1-infected children (24%) did not differ significantly from the group of uninfected children (19%; Fisher test: P = 0.73) . In contrast, the tuberculin skin test responses were positive less often in the group of HIV-1-infected children (33%) than in the uninfected group (83%; Fisher test: P = 0.007) . Because BCG vaccine appears to be safe--even when given to perinatally infected babies--continuation of the BCG immunization policies of the EPI is justified, especially in view of the growing incidence of tuberculosis as a complication of HIV infection. Appl Environ Microbiol, 1991 Feb, 57(2), 349 - 58 Insecticidal toxins from Bacillus thuringiensis subsp . kenyae: gene cloning and characterization and comparison with B . thuringiensis subsp . kurstaki CryIA(c) toxins; Von Tersch MA et al.; Genes encoding insecticidal crystal proteins were cloned from three strains of Bacillus thuringiensis subsp . kenyae and two strains of B . thuringiensis subsp . kurstaki . Characterization of the B . thuringiensis subsp . kenyae toxin genes showed that they are most closely related to cryIA(c) from B . thuringiensis subsp . kurstaki . The cloned genes were introduced into Bacillus host strains, and the spectra of insecticidal activities of each Cry protein were determined for six pest lepidopteran insects . CryIA(c) proteins from B . thuringiensis subsp . kenyae are as active as CryIA(c) proteins from B . thuringiensis subsp . kurstaki against Trichoplusia ni, Lymantria dispar, Heliothis zea, and H . virescens but are significantly less active against Plutella xylostella and, in some cases, Ostrinia nubilalis . The sequence of a cryIA(c) gene from B . thuringiensis subsp . kenyae was determined (GenBank M35524) and compared with that of cryIA(c) from B . thuringiensis subsp . kurstaki . The two genes are more than 99% identical and show seven amino acid differences among the predicted sequences of 1,177 amino acids. Gene, 1991 Feb 1, 98(1), 37 - 44 In vivo generation of hybrids between two Bacillus thuringiensis insect-toxin-encoding genes; Caramori T et al.; The parasporal crystal of Bacillus thuringiensis is composed of polypeptides highly toxic to a number of insect larvae . The structural genes (cryIA) encoding the Lepidoptera-specific toxin from different bacterial strains diverge primarily in a single hypervariable region, whereas the N-terminal and C-terminal parts of the proteins are highly conserved . In this report, we describe the generation of hybrid genes between two cryIA genes . Two truncated cryIA genes were cloned in a plasmid vector in such way as to have only the hypervariable region in common . The two truncated cryIA genes were separated by the tetracycline-resistance determinant (or part of it) . In vivo recombination between the hypervariable regions of the cryIA genes reconstituted an entire hybrid cryIA gene . Direct sequence analysis of 17 recombinant plasmids identified eleven different crossover regions which did not alter the reading frame and allowed the production of eight different hybrid proteins . The recombination events were independent from the RecA function of Escherichia coli . Some of the hybrid gene products were more specific in their insecticidal action and one had acquired a new biological activity. Gene, 1991 Feb 1, 98(1), 141 - 5 Nucleotide sequence of the LYS2 gene of Saccharomyces cerevisiae: homology to Bacillus brevis tyrocidine synthetase 1; Morris ME et al.; The Saccharomyces cerevisiae LYS2 gene, which encodes alpha-aminoadipate reductase, an essential enzyme in the yeast lysine biosynthetic pathway, has been sequenced . A large open reading frame (ORF) has been identified which can specify a 1392-amino acid protein with a deduced Mr of 155,344 . A DNA database search using the translated LYS2 ORF as a probe has revealed significant aa sequence homology to the Bacillus brevis enzyme tyrocidine synthetase 1. Mol Gen Genet, 1991 Feb, 225(2), 177 - 85 Hybrid Bacillus (1-3,1-4)-beta-glucanases: engineering thermostable enzymes by construction of hybrid genes; Olsen O et al.; Hybrid (1-3,1-4)-beta-glucanase genes were constructed by extension of overlapping segments of the (1-3,1-4)-beta-glucanase genes from Bacillus amyloliquefaciens and B . macerans generated by the polymerase chain reaction (PCR) . Four hybrid genes were expressed in Escherichia coli cells . The mature hybrid enzymes contain a 16, 36, 78, or 152 amino acid N-terminal sequence derived from B . amyloliquefaciens (1-3,1-4)-beta-glucanase followed by a C-terminal segment derived from B . macerans (1-3,1-4)-beta-glucanase . Biochemical characterization of parental and hybrid enzymes shows a significant increase in thermostability of three of the hybrid enzymes when exposed to an acidic environment thus combining two important enzyme characteristics within the same molecule . At pH 4.1, 85%-95% of the initial activity was retained after 1 h at 65 degrees C in contrast to 5% and 0% for the parental enzymes from B . amyloliquefaciens and B . macerans . After 60 min incubation at 70 degrees C, pH 6.0, the parental enzymes retained 5% or less of the initial activity whilst one of the hybrids still exhibited 90% of the initial activity . Of the parental enzymes B . macerans (1-3,1-4)-beta-glucanase had the lower specific activity while the hybrid enzymes exhibited specific activities that were 1.5- to 3-fold higher . These experimental results demonstrate that exchange of homologous gene segments from different species may be a useful technique for obtaining new and improved versions of biologically active proteins. Eur J Immunol, 1991 Feb, 21(2), 431 - 7 Regulation of tumor necrosis factor (TNF) release by murine peritoneal macrophages: role of cell stimulation and specific phagocytic plasma membrane receptors; Stein M et al.; In spite of the physiologic and pathologic importance of tumor necrosis factor (TNF), the cellular factors that govern its release by macrophages (M phi) are poorly understood, in comparison with other secretory products . We have studied the role of M phi heterogeneity and of plasma membrane receptors in regulating TNF release in vitro . Resident and various exudate murine peritoneal M phi populations were challenged with lipopolysaccharide (LPS) or different phagocytic particles, and TNF release assayed by cytotoxicity for L-929 fibroblasts . Resident peritoneal M phi (RPM phi) released a small amount of TNF in response to LPS whereas thioglycollate-elicited M phi (TPM phi) released high levels of TNF (5000 U/3 x 10(5) M phi/ml) . M phi elicited by Bio-Gel polyacrylamide beads (BgPM phi), another nonspecific inflammatory stimulus, or early in the course of intraperitoneal Bacillus Calmette-Guerin infection, before recruited cells become immunologically activated, released tenfold less TNF after the same stimulus . By contrast, TNF release in response to various phagocytic triggers was similar (approximately 300-600 U/3 x 10(5) M phi/ml) in all M phi populations including RPM phi . The response by BgPM phi to LPS was enhanced by pre-treatment in vitro with interferon-gamma or thioglycollate broth . With respect to phagocytic receptor triggering we found that complement receptor type 3 (CR3) ligation or latex uptake did not mediate release of significant quantities of TNF (less than 48 U/3 x 10(5) M phi/ml) by any M phi, whereas ligation of the Fc receptor for IgG1/IgG2b subclasses or of receptors for zymosan particles sufficed, in the absence of ingestion, to induce release of circa 500 U/3 x 10(5) M phi/ml TNF by all M phi tested . Our studies show that M phi vary in respect to priming for TNF release and that heterogeneity should be related to a particular triggering stimulus . Furthermore, the capacity of some M phi populations to release unusually high levels of TNF depends on immune or nonspecific stimuli subsequent to the process of inflammatory recruitment. Biochem J, 1991 Feb 1, 273 ( Pt 3), 503 - 10 Ragged N-termini and other variants of class A beta-lactamases analysed by chromatofocusing; Matagne A et al.; Four beta-lactamases excreted by Gram-positive bacteria exhibited microheterogeneity when analysed by chromatofocusing or ion-exchange chromatography . Ragged N-termini were in part responsible for the charge variants, but deamidation of an asparagine residue was also involved, at least for the Bacillus licheniformis enzyme . The activity of a contaminating proteinase could also be demonstrated in the case of Actinomadura R39 beta-lactamase . With that enzyme, proteolysis resulted in partial inactivation, but the inactivated fragments were easily separated from the active forms . With these, as with the other enzymes, the kinetic parameters of the major variants were identical with those of the mixture within the limits of experimental error, so that the catalytic properties of these enzymes can be determined with the 'heterogeneous' preparations. Semin Oncol, 1991 Feb, 18(1 Suppl 1), 27 - 38 The role of immunotherapy in colorectal cancer; Wadler S; Despite landmark advances in the molecular biology and surgical adjuvant therapy of colorectal cancer in the past decade, this illness remains a significant health hazard . Conventional therapies, including surgery, radiation therapy, and chemotherapy, have had limited utility and have often resulted in unacceptable host toxicities . Therapies dependent on potentiation of the host immune response are attractive alternatives to conventional treatment because of their greater specificity and diminished toxicity . Furthermore, the efficacy of many of these therapies has been demonstrated in preclinical models, which is important given the relative refractoriness of colorectal cancer to chemotherapy . Both active specific therapies, such as tumor vaccines, and passive therapies, such as monoclonal antibodies, have been investigated . In early clinical trials, both have demonstrated modest activity . Nonspecific immune stimulation with agents such as bacillus Calmette-Guerin or methanol extraction residue appear to have little utility, but with the development of recombinant DNA technology, specific cytokines with more precisely identified targets have been synthesized in sufficient quantities for clinical trials . Finally, combinations of biologics or combinations of biologics with conventional therapies, such as fluorouracil (5-FU)/levamisole and 5-FU/interferon, have been investigated and appear to be useful in the treatment of surgically resected and advanced colorectal carcinoma, respectively . The utility of these therapies is hampered by a limited understanding of their precise mechanisms of action and optimal conditions for administration . Nevertheless, immunotherapy of colorectal carcinoma remains an important and promising area for further clinical investigation. Proc Natl Acad Sci U S A, 1991 Feb 1, 88(3), 1054 - 8 Recombinant fusion protein identified by lepromatous sera mimics native Mycobacterium leprae in T-cell responses across the leprosy spectrum; Laal S et al.; Pooled polyvalent sera from lepromatous leprosy patients were used to screen a lambda gt11 recombinant DNA expression library of Mycobacterium leprae in order to identify the relevant antigens recognized by the human immune response . Of the 300,000 phages screened, 4 clones were identified that coded for fusion proteins of the same molecular mass . The fusion protein from clone LSR2 was tested for immunoreactivity in assays using peripheral blood cells and sera from 11 laboratory personnel and 105 patients across the leprosy spectrum . LSR2 protein appears to be predominantly a T-cell antigen . It evokes similar lymphoproliferative responses as the native bacillus both at the individual level and in the leprosy spectrum as a whole . Though only 50% of patient sera with anti-M . leprae antibodies reacted with the fusion protein, the pattern of reactivity in the antibody responses was also similar for the various clinical types . The coding regions of clones LSR1 and LSR2 are identical . They show no homology with sequences stored in data banks and encode a protein of 89 amino acids with a calculated molecular mass of approximately 10 kDa. J Bacteriol, 1991 Feb, 173(3), 1353 - 6 Efficient transformation of Bacillus thuringiensis requires nonmethylated plasmid DNA; Macaluso A et al.; The transformation efficiency of Bacillus thuringiensis depends upon the source of plasmid DNA . DNA isolated from B . thuringiensis, Bacillus megaterium, or a Dam- Dcm- Escherichia coli strain efficiently transformed several B . thuringiensis strains, B . thuringiensis strains were grouped according to which B . thuringiensis backgrounds were suitable sources of DNA for transformation of other B . thuringiensis strains, suggesting that B . thuringiensis strains differ in DNA modification and restriction . Efficient transformation allowed the demonstration of developmental regulation of cloned crystal protein genes in B . thuringiensis. Zhonghua Min Guo Wei Sheng Wu Ji Mian Yi Xue Za Zhi, 1991 Feb, 24(1), 30 - 5 Diagnostic value of measuring BCG sonicate antigen and anti-BCG antibodies in the cerebrospinal fluid and blood of patients with tuberculous meningitis; Lin HP et al.; Fourteen cerebrospinal fluid (CSF) and paired blood samples were obtained from patients with tuberculous meningitis, seven with positive culture and seven clinical/laboratory diagnosis . Another 14 paired specimens served as control, including 7 infectious meningitis and 7 non-inflammatory neurological diseases . Four groups were thus classified, including confirmed and suspected patients, and controls with infectious meningitis and neurological diseases . Measurements of Bacillus Calmette-Guerin (BCG) sonicate antigen and IgG antibody were performed using enzyme-linked immunosorbent assay . Generally, CSF BCG sonicate antigen level and anti-BCG sonicate antibody of patients with tuberculosis meningitis were higher than in control groups; greater antigen levels were found in confirmed patients than in non-inflammatory subjects (p less than 0.05), and in suspected patients than in infectious and non-inflammatory subjects (p less than 0.05 and p less than 0.01) . For serum anti-BCG sonicate levels, confirmed patients had higher antibody value than non-inflammatory subjects (p less than 0.05) . To conclude, detection of high levels of both BCG sonicate antigen and antibody in CSF and blood samples shows great value in the diagnosis of tuberculous meningitis . However, given the limited samples of the current research, more data are needed to elucidate the sensitivity and specificity of such tests. Nippon Hinyokika Gakkai Zasshi, 1991 Feb, 82(2), 290 - 6 {Intravesical instillation of bacillus Calmette-Guerin for superficial bladder carcinoma: study on significance of additional maintenance instillations of bacillus Calmette-Guerin}; Yabusaki N et al.; The efficacy of maintenance bacillus Calmette-Guerin (BCG) instillations for superficial bladder tumors was studied by prospective randomized trial . From June 1985 to October 1988, 42 newly diagnosed patients with superficial bladder carcinoma (pTa or pT1) were treated by transurethral tumor resection and subsequent five daily instillations of mitomycin C . Then they were divided into non-maintenance group (22 patients) and maintenance group (20 patients) by randomization . The patients received six weekly instillations of 80 mg of BCG . Tokyo strain (Japan BCG manufacturing Co., Tokyo, Japan), suspended in 40 ml of physiological saline, and the patients in the maintenance group received four additional instillations of BCG every three months . We could not complete the six-week course of BCG instillations in three patients due to adverse effects (two in non-maintenance group and one in maintenance group) and we lost six patients from follow-up within one year (one in non-maintenance group and five in maintenance group) . The mean follow-up period of the remaining 33 patients was 28.1 months . Of these 33 patients, six patients had been found to have recurrent tumors, and the over-all three-year non-recurrence rate was 82% . Before employing BCG, when we used only mitomycin C after TUR-Bt, the three year non-recurrence rate was 58% . This indicates prophylactic effect of BCG instillations . The stage of the initial tumor of the six recurrent cases were all pT1b . The non-recurrence rate of the patients with pT1b tumor was significantly lower than that of the patients with pTa and pT1a tumor . However, multiplicity and grade of tumors did not affect the non-recurrence rate.(ABSTRACT TRUNCATED AT 250 WORDS) J Gen Microbiol, 1991 Feb, 137 ( Pt 2), 399 - 404 Structural and stereospecific requirements for the nucleoside-triggered germination of Bacillus cereus spores; Senesi S et al.; A selection of adenosine analogues was tested for their ability to trigger germination of Bacillus cereus NCIB 8122 spores . The germination-inducing activity was governed by the structural properties of the sugar rather than the base moieties of the nucleosides . Among the sugar-modified analogues, only those containing a 2'-deoxy-D-ribose moiety promoted spore germination . Requirements for a specific molecular structure of the base were not clearly identified, although the highest activity was observed when substituents were inserted at position 6 of the purine ring . All the base-modified analogues, even those such as coformycin and 2'-deoxycoformycin with an expanded base ring, retained the germination-inducing activity of adenosine . However, of the two 2'-deoxycoformycin diastereoisomers characterized by an asymmetric carbon atom at position 8 of the homopurine ring, only the 8S-isomer induced germination, thus indicating that stereospecific configuration of the inducer, at least in the case of 2'-deoxycoformycin, appears to be essential for the initiation of spore germination . The differences in the germination-inducing activity of the various analogues tested were not affected significantly by spore activation at different temperatures, although the higher the activation temperature, the lower was the concentration of each analogue required for maximum germination. Appl Environ Microbiol, 1991 Feb, 57(2), 480 - 5 Biolistic transformation of a procaryote, Bacillus megaterium; Shark KB et al.; We present a simple and rapid method for introducing exogenous DNA into a bacterium, Bacillus megaterium, utilizing the recently developed biolistic process . A suspension of B . megaterium was spread onto the surface of nonselective medium . Plasmid pUB110 DNA, which contains a gene that confers kanamycin resistance, was precipitated onto tungsten particles . Using a biolistic propulsion system, the coated particles were accelerated at high velocities into the B . megaterium recipient cells . Selection was done by use of an agar overlay containing 50 micrograms of kanamycin per ml . Antibiotic-resistant transformants were recovered from the medium interface after 72 h of incubation, and the recipient strain was shown to contain the delivered plasmid by agarose gel electrophoresis of isolated plasmid DNA . All strains of B . megaterium tested were successfully transformed by this method, although transformation efficiency varied among strains . Physical variables of the biolistic process and biological variables associated with the target cells were optimized, yielding greater than 10(4) transformants per treated plate . This is the first report of the biolistic transformation of a procaryote. Genetika, 1991 Feb, 27(2), 238 - 46 {Inversion polymorphism in malaria mosquito Anopheles messeae . Part X . Resistance of larvae with different genotypes to toxins of crystal-forming bacteria Bacillus thuringiensis subsp . israelensis (serovar H14)}; Gordeev MI et al.; Insensibility of larvae with different chromosomal inversions to the toxins of Bacillus thuringiensis subsp . israelensis (Bti) was examined . It has been shown that larvae with inversion combinations XL0(XL1)2R0-3R0-3L0 had greater variability after treatment with Bti than larvae with inversions XL1(XL2)2R1-3R1-3L1(3L0) . The former inversion combinations were previously shown to dominate in the south of the species area and to be supported by K-selection . The latter inversion combinations form "northest" karyotypes and are supported by r-selection . It is obvious that genetic effects of treatments with Bti depend on the population's structure and the directions of natural selection . The decrease in the level of heterozygotes after treatment of larvae with Bti reflects destruction of the system of genetic homeostasis in the natural populations of A . messeae. J Biochem (Tokyo), 1991 Feb, 109(2), 211 - 6 Purification and characterization of a novel thermostable lipase from Bacillus sp; Sugihara A et al.; A thermostable lipase from Bacillus sp . has been purified to homogeneity as judged by disc-PAGE, SDS-PAGE, and isoelectric focusing . The purification included ammonium sulfate fractionation, treatment with acrinol, and sequential column chromatographies on DEAE-Sephadex A-50, Toyopearl HW-55F, and Butyl Toyopearl 650M . The purified enzyme was found to be a monomeric protein with Mr of 22,000, and pI of 5.1 . The optimal pH at 30 degrees C, and optimal temperature at pH 5.6 were 5.5-7.2, and 60 degrees C, respectively, when olive oil was used as the substrate . The substrate specificity towards simple triglycerides was broad and 1- and 3-positioned ester bonds were hydrolyzed in preference to a 2-positioned ester bond . The addition of acetone to the assay mixture in the range of 0-60% (v/v) stimulated the enzyme remarkably, whereas n-hexane had an inhibitory effect. Enferm Infecc Microbiol Clin, 1991 Feb, 9(2), 102 - 5 {Endocarditis caused cy Erysipelothrix rhusiopathiae . Study of 2 cases and review of the literature}; Azofra J et al.; Endocarditis produced by E . rhusiopathiae is a uncommon disease . Most of the infected persons (90%) work in environments with frequent exposure to E . rhusiopathiae (butchers, fisherman) . Although the clinical picture of endocarditis produced by E . rhusiopathiae is indistinguishable from other forms of subacute endocarditis, this infection has a mortality rate of 40% and a high morbidity . Microbiological diagnosis should consider the possibility of making a mistake considering that isolation of a gram-positive bacillus may represent contamination by an agent without clinical relevance . Treatment with penicillin G during 4 weeks is commonly sufficient to cure the disease. Cell Mol Neurobiol, 1991 Feb, 11(1), 219 - 30 Rapid analysis of glycolipid anchors in amphiphilic dimers of acetylcholinesterases; Toutant JP et al.; 1 . We describe two simple procedures for the rapid identification of certain structural features of glycolipid anchors in acetylcholinesterases (AChEs) . 2 . Treatment with alkaline hydroxylamine (that cleaves ester-linked acyl chains but not ether-linked alkyl chains) converts molecules possessing a diacylglycerol, but not those with an alkylacylglycerol, into hydrophilic derivatives . AChEs in human and bovine erythrocytes possess an alkylacylglycerol (Roberts et al., J . Biol . Chem . 263:18766-18775, 1988; Biochem . Biophys . Res . Commun . 150:271-277, 1988) and are not converted to hydrophilic dimers by alkaline hydroxylamine . Amphiphilic dimers of AChE from Drosophila, from mouse erythrocytes, and from the human erythroleukaemia cell line K562 also resist the treatment with hydroxylamine and likely possess a terminal alkylacylglycerol . This indicates that the cellular pool of free glycolipids used as precursors of protein anchors is distinct from the pool of membrane phosphatidylinositols (which contain diacylglycerols) . 3 . Pretreatment with alkaline hydroxylamine is required to render the amphiphilic AChE from human erythrocytes susceptible to digestion by Bacillus thuringiensis phosphatidylinositol-specific phospholipase C (PI-PLC) (Toutant et al., Eur . J . Biochem . 180:503-508, 1989) . We show here that this is also the case for the AChE from mouse erythrocytes, which therefore likely possesses an additional acyl chain in the anchor that prevents the action of PI-PLC . 4 . In two sublines of K562 cells (48 and 243), we observed that AChE either was directly susceptible to PI-PLC (243) or required a prior deacylation by alkaline hydroxylamine (48) . This suggests that glycolipid anchors in AChE of K562-48 cells, but not those in AChE of K562-243 cells, contain the additional acylation demonstrated in AChE from human erythrocytes . These observations illustrate the cell specificity (and the lack of species-specificity) of the structure of glycolipid anchors. Hindustan Antibiot Bull, 1991 Feb-Nov, 33(1-4), 19 - 25 Role of side chain moiety in the hydrolysis of penicillins by beta-lactamase; Ambedkar SS et al.; Various beta-lactam compounds and structurally related moieties were examined as substrates of beta-lactamase from Bacillus cereus 5/B NCTC 9946 . The enzyme was specific for penicillins and none of the cephalosporins were hydrolysed . Electronic environment of allylic carboxy group in dihydrothiazine ring restricts the acceptance of cephalosporins as substrates . The efficiency of hydrolysis of penicillins is dependent on dense resonating electronic environment of phenyl ring present in the side chain, flexibility of the side chain and the distance between the phenyl ring and carbonyl group in the side chain. Immunol Invest, 1991 Feb, 20(1), 33 - 43 The effect of alpha 2 macroglobulin-proteinase complexes on macrophage Ia expression in vivo; DeStefano A et al.; Alpha-2-Macroglobulin (alpha 2M) is a major plasma proteinase inhibitor . It can also regulate the function of cells of the immune system, including macrophage expression of Ia antigens in tissue culture systems . The present work was done to assess the effect of alpha 2M-trypsin complexes (alpha 2M-t) on macrophage Ia expression in vivo . Bacillus Calmette-Guerin-infected mice were injected intraperitoneally with 100nM alpha 2M-t, phosphate buffered saline (PBS), or bovine serum albumin (BSA) in PBS . The peritoneal cells were harvested by lavage from 3 to 6 days after injection . Differential cell counts were performed, and macrophage Ia antigen expression determined by indirect immunofluorescence . Injection of either alpha 2M-t or BSA solutions tended to increase the number of total cells and lymphocytes harvested, without changing the number of macrophages harvested . alpha 2M-t injection reduced the proportion of macrophages which were Ia positive from 60 to 37% on day 3 after injection, and to 20% Ia positive on day 6 . The reduction in Ia positive macrophages was statistically significant when compared to either PBS or BSA injected groups . In summary, in vivo exposure to alpha 2M-t can alter macrophage function . alpha 2M-proteinase complexes formed during the course of coagulation or inflammation may play a physiologic role as regulators of the immune response. Protein Expr Purif, 1991 Feb, 2(1), 51 - 8 High-level expression in Escherichia coli and rapid purification of phosphatidylinositol-specific phospholipase C from Bacillus cereus and Bacillus thuringiensis; Koke JA et al.; The construction of four vectors for high-level expression in Escherichia coli of the phosphatidylinositol-specific phospholipase C from Bacillus cereus or Bacillus thuringiensis is described . In all constructs the coding sequence for the mature phospholipase is precisely fused to the E . coli heat-stable enterotoxin II signal sequence for targeting of the protein to the periplasm . In one set of plasmids expression of the B . cereus or B . thuringiensis enzyme is under control of the E . coli alkaline phosphatase promoter, while in a second set of plasmids expression is under control of a lac-tac-tac triple tandem promoter . A simple and rapid procedure for complete purification of the phospholipase C overproduced in E . coli, involving isolation of the periplasmic proteins by osmotic shock followed by a single column chromatography step, is described . The largest quantity of purified enzyme, 40-60 mg per liter culture, is obtained with the plasmid expressing the B . cereus enzyme under control of the lac-tac-tac promoter . Lower quantities are obtained with the plasmids containing the alkaline phosphatase promoter (15-20 and 4-6 mg/liter for the B . cereus and B . thuringiensis enzymes, respectively) and with the plasmid expressing the B . thuringiensis phospholipase under control of the lac-tac-tac promoter (15-20 mg/liter) . A comparison of the functional properties of the recombinant phospholipases with the native enzymes isolated from B . cereus or B . thuringiensis culture supernatant shows that they are identical with respect to their catalytic functions, viz., cleavage of phosphatidylinositol and cleavage of the glycosyl-phosphatidylinositol membrane anchor of bovine erythrocyte acetylcholinesterase. Mol Microbiol, 1991 Feb, 5(2), 367 - 72 Identification of phosphatidylinositol-specific phospholipase C activity in Listeria monocytogenes: a novel type of virulence factor? Mengaud J, Braun-Breton C, Cossart P. A phospholipase C which cleaves phosphatidylinositol and glycosylphosphatidylinositol (GPI) anchors was identified in Listeria monocytogenes . This 36 kDa protein is encoded by the gene plcA, and is homologous to the Bacillus cereus, Bacillus thuringiensis and eukaryotic phosphatidylinositol-specific phospholipases C (PI-PLC) . Expression of the plcA gene in Escherichia coli correlates with the appearance of PI-PLC activity in the cells . In Listeria monocytogenes, the activity is secreted to the culture medium . PI-PLC activity was only found in the two pathogenic species of the genus Listeria, namely L . monocytogenes and L . ivanovii . PI-PLC activity was lost and virulence decreased when the plcA gene was disrupted in the chromosome . This suggests that the PI-PLC of L . monocytogenes might be involved in virulence. J Biotechnol, 1991 Feb, 17(2), 121 - 31 Production of 5-methyluridine by immobilized thermostable purine nucleoside phosphorylase and pyrimidine nucleoside phosphorylase from Bacillus stearothermophilus JTS 859; Hori N et al.; 5-Methyluridine was produced continuously from thymine and inosine by immobilized enzymes, which consisted of thermostable purine nucleoside phosphorylase and thermostable pyrimidine nucleoside phosphorylase obtained from Bacillus stearothermophilus JTS 859 . The process was carried out in a column reactor at 60 degrees C for 17 d without any bacterial contamination under non-aseptical conditions . Half-lives of the activity of the immobilized enzymes were 47 d and 4.5 d at 60 degrees C and 70 degrees C, respectively, although half-life of the crude enzyme was only 14 h at 70 degrees C. Biochemistry, 1991 Jan 29, 30(4), 1028 - 36 Detection and characterization of intermediates in the folding of large proteins by the use of genetically inserted tryptophan probes; Smith CJ et al.; L-Lactate dehydrogenase from Bacillus stearothermophilus was rebuilt by using site-directed mutagenesis to produce an enzymically active, tryptophan-less enzyme by replacing all the wild-type tryptophans (80, 150, and 203) by tyrosines . Nine single tryptophan-containing active enzymes were constructed from this enzyme by genetically replacing one of the tyrosines 36, 85, 147, 190, 203, 237, 248, 279, or 285 by tryptophan . The equilibrium and the time-resolved tryptophan fluorescence intensity and anisotropy were used to report unfolding events in guanidine hydrochloride (GHCl) monitored from these nine defined positions . Three structural transitions, half complete at 0.55, 1.7, and 2.8 M GHCl, were identified and defined four folding intermediates, I (native), II (expanded monomer 1), III (expanded monomer 2), and IV (random coil), stable at 0, 1, 2.2, and 4 M GHCl, respectively . Intermediate II is a globular monomer . All the probed alpha-helices and most of the beta-structure was intact . There was an increase in the rate but not the extent of the mobilities of six of the probed tryptophan side chains, indicating loss of tertiary structure . Circular dichroism (CD) showed all the secondary structure to be intact . Intermediate III is monomeric and still globular, but the tryptophan anisotropy indicated an increase mobility at positions 36, 85, 190, 203, 279, and 285 . Helix alpha-B is further disrupted but helices alpha-1F, alpha-2G, and alpha 3G were still rigid . CD showed half the secondary structure to be still intact . Intermediate IV is a random coil in which all tryptophans have complete rotational freedom and the helix CD signal is lost.(ABSTRACT TRUNCATED AT 250 WORDS) J Biol Chem, 1991 Jan 15, 266(2), 880 - 5 Cloning, sequencing, and overexpression of genes for ribosomal proteins from Bacillus stearothermophilus; Ramakrishnan V et al.; Although a low resolution model for the arrangement of the proteins of the small and large ribosomal subunits is known, a detailed mechanistic understanding of the function of the ribosome awaits a high resolution structure of its components . While crystals have been obtained of several ribosomal proteins from Bacillus stearothermophilus, determination of atomic resolution structures of these proteins is impeded by the difficulty of obtaining large amounts of native proteins for crystallographic or NMR studies . We describe here the cloning and overexpression in Escherichia coli of the genes for ribosomal proteins S5, L6, L9, and L18 from B . stearothermophilus . S5 is extremely toxic to E . coli when overexpressed, and we have taken advantage of a new tightly regulated expression system to obtain high yields (more than 100 mg of pure protein/liter of culture) of this protein . The B . stearothermophilus S5 produced in E . coli crystallizes, and the crystals are identical to those obtained from the native protein . The crystals diffract to 2-A resolution. FEMS Microbiol Lett, 1991 Jan 15, 61(2-3), 271 - 5 Cloning of a chromosomal alpha-amylase gene from Bacillus stearothermophilus; Jorgensen PL et al.; We have cloned and sequenced a gene for a heat-stable alpha-amylase from a natural isolate of Bacillus stearothermophilus . Previously, it had been shown that B . stearothermophilus amylase genes may be harboured on indigenous plasmids . We have found that our isolate harbours the amylase gene only on the chromosome and not on its indigenous plasmid. J Biol Chem, 1991 Jan 15, 266(2), 1170 - 6 Mechanism of inhibition of adenylate cyclase by phospholipase C-catalyzed hydrolysis of phosphatidylcholine . Involvement of a pertussis toxin-sensitive G protein and protein kinase C; Diaz-Laviada I et al.; The phospholipase C-mediated hydrolysis of phosphatidylcholine has been shown recently to be activated by a number of agonists . Muscarinic receptors, which trigger various signal transduction mechanisms including inhibition of adenylate cyclase through Gi, have been shown to be potent stimulants of this novel phospholipid degradative pathway . We demonstrate here, by exogenous addition of Bacillus cereus phosphatidylcholine-hydrolyzing phospholipase C, that phosphatidylcholine breakdown mimics the ability of carbachol to inhibit adenylate cyclase . This effect is sensitive to pertussis toxin and is entirely dependent on the presence of protein kinase C . This kinase is also required for the inhibition by carbachol of adenylate cyclase . These results suggest that the activation of phosphatidylcholine breakdown by phospholipase C may play an important role linking or favoring the coupling muscarinic receptors to Gi . Results presented here also show that phospholipase C-mediated hydrolysis of phosphoinositides by exogenous addition of Bacillus thuringiensis phosphoinositide-hydrolyzing phospholipase C does not affect adenylate cyclase, despite the fact that protein kinase C is translocated to an extent similar to that produced by the hydrolysis of phosphatidylcholine . According to the results shown here, both phospholipases also differ in their ability to down-regulate protein kinase C as well as to phosphorylate p80 and to transmodulate the binding of epidermal growth factor, two well established effects of protein kinase C in Swiss 3T3 fibroblasts . This emphasizes the complexity, from a functional point of view, of protein kinase C activation "in vivo." Carbohydr Res, 1991 Jan 15, 209, 145 - 53 On the role of histidine residues in cyclodextrin glycosyltransferase: chemical modification with diethyl pyrocarbonate; Bender H; Ethoxyformylation with diethyl pyrocarbonate of approximately 1.5 His residues per molecule of enzyme reduced the cyclising activity of both the alpha-cyclodextrin glycosyltransferase from Klebsiella pneumoniae strain M 5 al and the beta-cyclodextrin glycosyltransferase from Bacillus circulans strain 8 by greater than 90% . Pre-incubation with substrate protected the enzymes from ethoxyformylation . Digestion of starch by the modified enzymes resulted in a delayed formation of cyclodextrins (cyclomalto-oligosaccharides, CDs), but a marked increase in the production of reducing saccharides . Similarly, coupling of alpha CD and maltose and successive disproportionation yielded mainly glucose and malto-oligosaccharides . The results are discussed in the context of the role of conserved His residues for binding of substrate and the transfer reactions. Dev Comp Immunol, 1991 Winter, 15(1-2), 27 - 32 Antibacterial activity of Eisenia fetida andrei coelomic fluid: III--Relationship within the polymorphic hemolysins; Roch P et al.; The antibacterial activity exhibited by 10 different hemolytic, genetic families was established by measuring the inhibition of spontaneous in vitro growth by cell-free coelomic fluid toward 2 bacteria which are pathogenic for the earthworm: Bacillus megaterium (Gram +) and Aeromonas hydrophila (Gram -) . Only two families (B and K) displayed potent inhibitory activities . This finding is consistent with the fact that the B family occurs most frequently in both natural as well as in industrial breedings . Nevertheless, evidence of a poor antibacterial defense in some frequent families suggests the existence of alternative antibacterial mechanisms. Salud Publica Mex, 1991 Jan-Feb, 33(1), 70 - 6 {Tuberculous meningitis: a 10-year analysis at the "Dr . Federico Gómez" Children's Hospital of Mexico}; Karam Bechara J et al.; Of all the forms of tuberculous infection in children, the most frequent is the pulmonary, but the tuberculous meningitis is the most severe and mortal, it occurs mainly in children under five years old, and the highest mortality is in children under two . The results of a retrospective study carried out at the Children Hospital of Mexico "Dr . Federico Gomez" about all the patients hospitalized with tuberculous meningitis diagnose during the January 1975 to December 1985 period were presented . One hundred and eighteen cases were studied: the majority (80%) corresponded to children under four years old, a percentage of 79 presented some degree of malnutrition, and 86 per cent showed clinical data of neurological affection . The confirmation of the diagnose was made in the majority of cases through laboratory and cabinet studies . Forty per cent showed no alteration in the chest X-rays, and the isolation of the bacillus was in a very low percentage (15%) . All the patients were treated with the antituberculous drugs mentioned, with a better development in the ones associated with steroids . The hospital stay was over 30 days in 15 per cent of the cases . The mortality of the series reviewed was 44.5 per cent. J Gen Microbiol, 1991 Jan, 137 ( Pt 1), 41 - 8 Purification and properties of an acid endo-1,4-beta-glucanase from Bacillus sp . KSM-330; Ozaki K et al.; A novel acid cellulase (endo-1,4-beta-glucanase, EC 3.2.1.4) was found in a culture of Bacillus sp . KSM-330 isolated from soil . One-step chromatography on a column of CM-Bio-Gel A yielded a homogeneous enzyme, as determined by silver staining of both sodium dodecyl sulphate (SDS) and nondenaturing gels . The enzyme had a molecular mass of 42 kDa, as determined by SDS-polyacrylamide gel electrophoresis . The isoelectric point was higher than pH 10 . The N-terminal amino acid sequence of the enzyme was Val-Ala-Lys-Glu-Met-Lys-Pro-Phe-Pro-Gln-Gln-Val-Asn-Tyr-Ser-Gly-Ile-Leu- Lys-Pro . This enzyme had an optimum pH for activity of 5.2, being active over an extremely narrow range of pH values, from 4.2 to 6.9; below and above these pH values no activity was detectable . The optimum temperature at pH 5.2 was around 45 degrees C . The enzyme efficiently hydrolysed carboxymethylcellulose (CMC) and lichenan, but more crystalline forms of cellulose, curdlan, laminarin, 4-nitrophenyl-beta-D-glucopyranoside and 4-nitrophenyl-beta-D-cellobioside were barely hydrolysed . The enzymic activity was inhibited by Hg2+ but was not affected by other inhibitors of thiol enzymes, such as 4-chloromercuribenzoate . N-ethylmaleimide and monoiodoacetate . N-Bromosuccinimide abolished the enzymic activity, and CMC protected the enzyme from inactivation by this tryptophan-specific oxidant . It is suggested that a tryptophan residue(s) is involved in the mechanism of action of the Bacillus cellulase and that the inhibition of enzymic activity by Hg2+ is ascribable to interactions with the tryptophan residue(s) rather than with thiol group(s). Pharmacotherapy, 1991, 11(2 ( Pt 2)), 64S - 71S Monotherapy versus combination antimicrobial therapy: a review; Barriere SL; In view of the newer antibiotics that can cover the spectrum of organisms in mixed infections, single agents are now a viable option for antimicrobial therapy . In addition, monotherapy is relatively nontoxic and possibly less costly than combination therapy . Combinations may be more effective in preventing the emergence of resistance, however, and can also provide synergistic effects . They are a necessity in mycobacterial infections, enterococcal endocarditis, and deep-seated pseudomonal infections, as well as in patients with gram-negative bacillary infection with profound granulocytopenia. Exp Pathol, 1991, 41(3), 147 - 50 Bacillus Calmette-Guérin (BCG) influences cell proliferation and glycosaminoglycans of chondrocyte cultures; Kittlick PD et al.; There are only few reports on the correlation between bacterial products and the GAG pattern of cartilage . Mycobacteria bovis (BCG) were applied to chondrocyte monolayer cultures for one week . The following parameters did change: cell proliferation increased, glycosaminoglycan synthesis and secretion decreased, hyaluronic acid in secreted and cell-associated glycosaminoglycans increased, a correlation between the degree of these changes and the degree of cell differentiation seems to exist . The contact of bacteria like BCG to chondrocytes may change the cellular metabolism . On the tissue level this may injure articular cartilage and thus support the concept of predamaged cartilage that is readily susceptible to further degradation. Acta Otorrinolaringol Esp, 1991 Jan-Feb, 42(1), 75 - 7 {Primary laryngeal tuberculosis caused by Mycobacterium bovis}; Contreras Sanchez JD et al.; It is suggested that laryngeal tuberculosis is a common complication of pulmonary tuberculosis . The most frequent germ is Mycobacterium tuberculosis . We present a case in which a infrequent bacillus, Mycobacterium bovis, was isolated without lung afectation . Considerations about morphologic and microbiologic findings are discussed. Urol Res, 1991, 19(1), 35 - 8 Induction of controlled prostatic tissue necrosis by bacille Calmette-Guérin derivatives; Morales A et al.; Intraprostatic administration of live bacille Calmette-Guerin (BCG) in humans has been found to produce tumor necrosis; unfortunately, the number of severity of complications have made its clinical use prohibitive . Previous studies have shown that soluble and microparticulate components present in the supernatants obtained after centrifugation of a reconstituted BCG preparation exhibit similar immunogenicity to the one shown by live bacteria . The supernatants, however, are not associated with disseminated infection of the progressive regional tissue destruction observed with the use of viable vaccine . Experiments were conducted to determine the effect of intraprostatic injection of BCG and its supernatants . Adult dogs, after positive conversion to protein purified derivative (PPD), were randomly assigned to three groups . Under direct vision and with digital rectal control, intraprostatic injections of various agents were given as follows: group I, normal saline; group II, live BCG; group III, 200 micrograms of BCG supernatants . Two months later the animals were sacrificed, and the prostates removed in toto and submitted for a thorough histological examination . Extensive but variable tissue necrosis was noted in groups II and III . No histological alterations were present in group I . The histological picture of the animals receiving BCG supernatants conclusively demonstrated circumscribed necrosis of the gland . Side effects and complications were present in animals receiving live BCG but conspicuously absent in the ones receiving supernatants . The observed effectiveness and safety of BCG supernatants for intraprostatic administration in an experimental system may lead to a simple, safe, and efficacious therapeutic modality for localized carcinoma of the prostate in humans. Food Addit Contam, 1991 Jan-Feb, 8(1), 65 - 9 Residues of doxycycline and oxytetracycline in eggs after medication via drinking water to laying hens; Yoshimura H et al.; Doxycycline (DOTC) and oxytetracycline (OTC) were dissolved in drinking water (0.5 g/l) and supplied to laying hens for 7 consecutive days . Eggs laid were collected daily during and after medication, and the antibiotic concentrations in the yolk and albumin were determined by the cup-plate method with Bacillus cereus var . mycoides ATCC 11778 . The concentrations of both antibiotics were increased in yolk day by day with the advance in medication, reached peaks 2 days after withdrawal and then declined gradually . Mean peak concentrations in the yolk were 6.70 micrograms/g for DOTC and 1.42 micrograms/g for OTC . Peak concentrations in the albumin occurred in the middle stage of medication, where the mean values were 12.24 micrograms/g for DOTC and 1.03 micrograms/g for OTC . DOTC was detected in albumin until 24 days after withdrawal and for 2 days more in yolk than in albumin . OTC was detected in yolk until 9 days after withdrawal . The depletion period of OTC was shorter for the albumin, where the residue disappeared in all eggs 6 days after withdrawal . In spite of similarities between DOTC and OTC in structure, DOTC was deposited in higher concentrations and lasted for a longer period in eggs . This characteristic was considered due to its greater lipophilicity, closely correlated with oral absorption and tissue penetration. J Invertebr Pathol, 1991 Jan, 57(1), 82 - 93 Peptide mapping of different Bacillus thuringiensis toxin gene products by CNBr cleavage in SDS-PAGE gels; Pang AS et al.; A cyanogen bromide fragmentation reaction of 3 hr in sodium dodecyl sulfate gel slices was used for the degradation of the toxins coded by the three cryIA genes from Bacillus thuringiensis kurstaki . Peptide patterns diagnostic for each toxin gene product were observed . Treatment longer than 3 hr led to the weakening and disappearance of protein bands . In 9 out of 16 wild B . thuringiensis strains tested, it was determined that only one of the cryIA genes was being expressed; in 6 strains, one to two genes were identified, with the presence of an additional gene possible and masked because of the overlap of peptide bands; in one strain none of the genes was expressed. J Invertebr Pathol, 1991 Jan, 57(1), 101 - 8 Chemical modification of Bacillus thuringiensis subsp . thuringiensis (HD-524) trypsin-activated endotoxin: implication of tyrosine residues in lepidopteran cell lysis; Yan XJ et al.; A purified protein fraction from a solubilized and trypsin-digested extract of Bacillus thuringiensis subsp . thuringiensis (HD-524) fermentation powder was lytic to cells from several lepidopteran lines . Maximum yield was obtained by alkaline carbonate-thiocyanate solubilization of washed powder followed by trypsin digestion and Sephacryl (S-300) chromatography . The alkaline carbonate-solubilized fraction consisted predominantly of two bands on sodium dodecyl sulfate-polyacrylamide gel electrophoresis with MW of 144 +/- 0.9 kDa and 134 +/- 1.4 kDa . After trypsin treatment and column chromatography, the cytolytic fraction consisted of a major band with a MW of 60.0 +/- 1.8 kDa and a minor band of 69 +/- 0.9 kDa . Cells from Trichoplusia ni (TN368) were most susceptible to lysis with 50% of cells lysed at 3 micrograms/ml, followed by Spodoptera frugiperda cells (SF21AE) exhibiting 50% cell lysis at 5 micrograms/ml and Lymantria dispar cells (Ld652Y) showing 40% lys