Germination of concentrated suspensions of spores from Aspergillus niger

Summary Germination of condiospores fromA.niger in very concentrated suspension was required to inoculate solid fermentation media, but a germination self-inhibitor was detected in spores. It was found that the inhibitory effect depended on the composition of the medium and was reduced when glucose was used as a carbon source. The effect of the self-inhibitor was eliminated by washing the spores and using glucose and a protein nitrogen source in the germination medium. By this method it was possible to increase about 100 times (10⁶ to 10⁸) spore concentration, keeping more than 90% germination.     

Functional alpha 2-adrenoceptors in rat adipocytes: inhibition of cyclic AMP accumulation

Alpha 2-adrenoceptor activation inhibits cyclic AMP accumulation in fat cells from many species. However, the presence of alpha 2-adrenoceptors in rat adipocytes has been difficult to demonstrate. We observed that alpha 2-adrenergic activation inhibits forskolin-stimulated cyclic AMP accumulation both in rat and hamster adipocytes; UK 14304, p-amino clonidine and clonidine were the agents with higher efficacy. The effect of UK 14304 was blocked by yohimbine but not by prazosin demonstrating the involvement of alpha 2-adrenoceptors. Pertussis toxin blocked the alpha 2-adrenergic effect. Our results demonstrate the presence in rat fat cells of alpha 2-adrenoceptors coupled to adenylate cyclase via "Gi".     

Effects of postnatal age and diet on the fatty acid composition of plasma lipid fractions in preterm infants

Diet and postnatal age effect the fatty acid composition of plasma and tissue lipids. This work was designed as a transversal study to evaluate the changes in the fatty acid composition of plasma phospholipids, cholesteryl esters, triglycerides and free fatty acids in preterm infants (28-35 weeks gestational age), fed human milk (HM) and milk formula (MF) from birth to 1 month of life. Sixteen blood samples were obtained from cord, and 19 at 6-8 h after birth, 14 at 1 week and 9 at 4 weeks from HM-fed infants and 18 at 1 week and 14 at 4 weeks from MF-fed ones. Groups had similar mean birth weight, gestational age and sex ratio. The MF provided 69 kcal/dl and contained 16% of linoleic acid and 1.3% of alpha-linolenic acid on the total fat. Plasma lipid fractions were extracted and separated by thin-layer chromatography and fatty acid methyl esters were quantitated by gas liquid chromatography. In plasma phospholipids, linoleic acid (18:2 omega 6) continuously increased from birth to 1 month of age, but no changes were seen as related to type of diet; polyunsaturated fatty acids greater than 18 carbon atoms of both the omega 6 and omega 3 series (PUFA omega 6 greater than 18 C and omega 3 greater than 18 C) dropped from birth to 1 week and continued to decrease in MF-fed infants until 1 month; eicosatrienoic (20:3 omega 6), arachidonic (20:4 omega 6) and docosahexaenoic (22:6 omega 3) were the fatty acids implicated. In cholesteryl esters palmitoleic (16:1 omega 7) and oleic (18:1 omega 9) acids decreased from birth to 1 month and linoleic acid increased and arachidonic acid dropped, especially in MF fed infants. In triglycerides, palmitic, palmitoleic and stearic acid (18:0) decreased during the first month of life; oleic acid remained constant and linoleic acid increased in all infants, but arachidonic acid decreased only in those fed formula. Free fatty acids showed a similar behavior in fatty acids and in plasma triglycerides. Preterm neonates seem to have special requirements of long-chain PUFA and adapted MF should contain these fatty acids in similar amounts to those of HM to allow the maintenance of an adequate tissue structure and physiology.     

Evidence that replication initiates at only some of the potential origins in each oligomeric form of bovine papillomavirus type 1 DNA.

In a subclone of ID13 mouse fibroblasts latently infected with bovine papillomavirus type 1 (BPV-1) DNA, the viral genome occurred as a mixture of extrachromosomal circular monomers and oligomers. Multiple copies were also associated with the host cell genome, predominantly at a single site in a head-to-tail tandem array. We examined the replicative intermediates of extrachromosomal forms of BPV-1 DNA by using two-dimensional gel electrophoresis. The results obtained indicate that initiation of DNA replication occurred near the center of the EcoRI-BamHI 5.6-kilobase fragment. In some molecules, however, this fragment was replicated from one end to the other by means of a single fork initiated elsewhere. Termination also occurred within this fragment. The EcoRI-BamHI 2.3-kilobase fragment replicated as a DNA molecule containing a termination site for DNA replication and also by means of a single fork traversing the fragment from one end to the other. Thus, replication forks proceeded through these fragments in different manners, apparently depending on whether they were part of a monomer, a dimer, a trimer, or higher oligomers. These observations lead to the conclusion that initiation of DNA replication in BPV-1 DNA takes place at or close to plasmid maintenance sequence 1. From this point, replication proceeds bidirectionally and termination occurs approximately 180 degrees opposite the origin. The results obtained are consistent with one or more replication origins being quiescent in BPV-1 DNA oligomers.     

Effect of pectin on pectinases in autolysis of Botrytis cinerea

Pectic activity in autolyzed cultures of Botrytis cinerea in a medium with and without pectin was similar, but in the medium with pectin maximal activities occurred in younger cultures. The pectic activities found were polygalacturonase, polymethylgalacturonase, endo activity (pectin as substrate) and pectin lyase. The molecular weights of polygalacturonase, polymethylgalacturonase and endo activity (pectin as substrate) were 36000, 33000 and 30200 daltons respectively, and the molecular weight of pectin lyase was 18200 daltons. By gel electrophoresis four different pectic activities were detected, three in the top of the gel and one in the bottom. Two enzymes were characterized, the polygalacturonase activity (first band in the top) inhibited by Ca⁺⁺ and the pectin lyase activity (in the bottom) which was not inhibited by Ca⁺⁺. These enzymes are not induced by the presence of pectin in the medium during degradation of Botrytis cinerea.     

Thiosulfate production from cystine by the keratinolytic prokaryote Streptomyces fradiae

In cultures of Streptomyces fradiae on wool as the only source of nutrition inorganic thiosulfate (in amounts up to 0.5 mg of Na₂S₂O₃·5 H₂O/ml) was formed as the final product of metabolization of sulfur from cystine of keratin proteins. The presence of thiosulfate was proved by qualitative tests and thin-layer chromatography and estimated quantitatively by spectrophotometry, titrimetry, and capillary isotachophoresis. Metabolization of organic sulfur to thiosulfate excreted into the medium is a process not yet described in microorganisms.     

Purification and characterization of two trypsin-like enzymes from the digestive tract of anchovy Engraulis encrasicholus

1. Two trypsin-like enzymes, designated Trypsin A and B, were purified from the pyloric caeca and intestine of anchovy by (NH4)2SO4 fractionation, affinity chromatography (Benzamidine-Sepharose-6B) and ion exchange chromatography (DEAE-Sepharose). 2. Both trypsins catalyzed the hydrolysis of N-benzoyl-DL-arginine p-nitroanilide (BAPNA), p-tosyl-L-arginine methyl ester (TAME), casein and myofibrillar protein and they were inhibited by several well established trypsin-inhibitors. 3. The enzymes had mol. wts of 27,000 (Trypsin A) and 28,000 (Trypsin B). Their isoelectric points were about 4.9 (Trypsin A) and 4.6 (Trypsin B) and they had similar amino acid composition. 4. The enzymes had a pH optimum of 8-9 for the hydrolysis of BAPNA and of 9.5 for the digestion of casein and myofibrillar protein. Their activity and stability were affected by calcium ions. 5. Trypsins A and B resemble other fish trypsins in their mol. wt, pI, kinetic properties and the instability at low pH and they are similar to bovine trypsin in their dependence of Ca2+ for activity and stability.     

Isolation and biochemical characterization of Streptomyces clavuligerus mutants in the biosynthesis of clavulanic acid and cephamycin C

Summary Seven mutants of Streptomyces clavuligerus blocked in the biosynthesis of clavulanic acid, cephamycin C, or both antibiotics, have been isolated and characterized. Mutants nca1 and nca2 were unable to synthesize clavulanic acid but produced cephamycin C. Mutants nce1 and nce2 were completely blocked in cephamycin C production but formed clavulanic acid. A third group (mutants ncc1, ncc4 and ncc5) failed to produce both antibiotics. Arginase activity (forming ornithine) was very low in mutants ncc1 and ncc5. All the mutants blocked in clavulanic acid biosynthesis showed a normal ornithine--aminotransferase activity. Mutant ncc1, blocked in cephamycin biosynthesis, lacked completely lysine--aminotransferase (forming -aminoadipic acid) and isopenicillin N synthase. Two other mutants (nce2 and nce5) lacked isopenicillin N synthase. There was a good correlation between the isopenicillin N synthase and the lysine--aminotransferase activities of the nca mutants and the ability of those strains to produce cephamycin C. The condensing enzyme involved in the formation of the clavulanic acid nucleus appears to be different from the isopenicillin N synthase.Dedicated to Professor H.-J. Rehm on the occasion of his 60th birthday     

Effects of social isolation and crowding upon adrenocortical reactivity and behavior in the rat

The effects of social isolation and crowding on adrenocortical function and upon behavioral responsiveness to electric shock have been studied in male and female rats. All female experimental groups showed higher corticosterone levels and heavier adrenals than their male counterparts. The major effect of housing condition concerned the corticosterone response to stress, while basal hormone concentration was not modified. Socially housed rats showed a more intense adrenocortical response and also a greater behavioral reactivity to electric shock than the isolates.