Growth and arginine metabolism of the wine lactic acid bacteria Lactobacillus buchneri and Oenococcus oeni at different pH values and arginine concentrations

During malolactic fermentation (MLF) in grape must and wine, heterofermentative lactic acid bacteria may degrade arginine, leading to the formation of ammonia and citrulline, among other substances. This is of concern because ammonia increases the pH and thus the risk of growth by spoilage bacteria, and citrulline is a precursor to the formation of carcinogenic ethyl carbamate (EC). Arginine metabolism and growth of Lactobacillus buchneri CUC-3 and Oenococcus oeni strains MCW and Lo111 in wine were investigated. In contrast to L. buchneri CUC-3, both oenococci required a higher minimum pH for arginine degradation, and arginine utilization was delayed relative to the degradation of malic acid, the main aim of MLF. This allows the control of pH increase and citrulline formation from arginine metabolism by carrying out MLF with pure oenococcal cultures and inhibiting cell metabolism after malic acid depletion. MLF by arginine-degrading lactobacilli should be discouraged because arginine degradation may lead to the enhanced formation of acids from sugar degradation. A linear relationship was found between arginine degradation and citrulline excretion rates. From this data, strain-specific arginine-to-citrulline conversion ratios were calculated that ranged between 2.2 and 3.9% (wt/wt), and these ratios can be used to estimate the contribution of citrulline to the EC precursor pool from a given amount of initial arginine. Increasing arginine concentrations led to higher rates of growth of L. buchneri CUC-3 but did not increase the growth yield of either oenococcus. These results suggest the use of non-arginine-degrading oenococci for inducing MLF.     

Gene location and bacterial sequence divergence

Previous comparison of a relatively small set of homologous genes from Escherichia coli and Salmonella typhimurium revealed that genes nearer to the origin of replication had substitution rates lower than genes closer to the replication terminus. The recently completed sequences of numerous bacterial genomes have allowed us to test whether this effect of distance from the replication origin on substitution rates, as observed for the E. coli-S. typhimurium comparison, is a general feature of bacterial genomes. Extending the analysis to all 3,000 E. coli-S. typhimurium homologs confirmed the significant association between chromosomal position and synonymous site divergence. However, the effect, though still significant, is not as dramatic as originally thought. A similar association between relative chromosomal location and synonymous substitution rate was detected in the majority of other bacterial species comparisons within alpha- and gamma- Proteobacteria, and Firmicutes but was absent in Chlamydiales. The opposite trend, i.e., a decrease in synonymous divergence with distance from the replication origin, was detected in Mycobacteria. Analysis of the patterns of nucleotide substitutions revealed that the distance effect is not affected by gene orientation and is mainly caused by an increase in rates of transversions, suggesting that this effect may not be caused by recombinational repair or biased gene conversion, as originally suggested.     

Herding and snaking by the harem stallion in domestic herds

Four herds of pony mares, each consisting of a stallion and six mares, were used to characterize the nature of herding by the stallion and the factors that induced the herding behavior. Herding behaviors were compared among four successive treatments (six mares alone, stallion added, two new mares added, and entire herd moved to a new pasture). A new treatment was initiated every 7 days and behavior was studied for 5 consecutive days (Days 1-5) for each treatment. Observations were made every 10 min during a 2-h period for each day. The extent of herding was quantitated by the mean distances between mares. The extent of snaking (herding with the head and neck extended and ears held back) was scored 0, 1, 2, or 3 (nil, minimal, intermediate, and maximal, respectively). The mean distance among the original mares on Day 1 when the mares were alone was 5.0 mare lengths and was reduced (P < 0.05) to 1.9 mare lengths when the stallion was added. The mean distance among the original mares of an established stallion/mare herd (3.8 mare lengths) was reduced (P < 0.05) on the day the herd was moved to a new pasture (1.9 mare lengths), similar to the effect of the introduction of the stallion. Scores for the extent of snaking, as well as the extent of herding, were highest (P < 0.05) on Day 1 when the stallion was added or the stallion/mare herd was moved to a new pasture. The extent of herding and snaking decreased (P < 0.05) by Day 2 and was seen only occasionally on Days 3-5. The addition of new mares to the herd did not induce herding of the original mares. However, the new mares maintained mean distances of 8-12 mare lengths from the original mares, resulting primarily from chasing by the stallion. By Day 4, the distances between the new and original mares were not different (P > 0.05) from the distances among the original mares.     

Palaeoecology of Upper Pleistocene-Holocene bryozoan and foraminiferal assemblages from Ku?dili (Kadiköy, Istanbul, Turkey)

Bryozoan and foraminiferal assemblages from a core drilled in the Kusdili (Kadiköy, Istanbul) Upper Pleistocene-Holocene sequence were studied. Assemblages are absent from the very basal and top parts but they are well represented all along the middle part of the sequence. All the species found presently live in the Mediterranean. In the area sedimentation seemingly started about 13?000 years ago in very shallow near-shore environments with the deposition of pebbly sediments rapidly evolving to mud. Near the base, the presence of erosional surface allows to recognize three successive sedimentary cycles. Bryozoan and foraminiferal assemblages found in the middle part of the sequence testify to a shallow water, estuarine or lagoonal paleo-environment, in which effects of low salinity waters, more obvious toward the top, could be related to the reestablishment of a connection between the Marmara Sea and the Black Sea through the Bosphorus, about 7000 years BP.     

The Italian Twin Project: from the personal identification number to a national twin registry.

The unique opportunity given by the "fiscal code", an alphanumeric identification with demographic information on any single person residing in Italy, introduced in 1976 by the Ministry of Finance, allowed a database of all potential Italian twins to be created. This database contains up to now name, surname, date and place of birth and home address of about 1,300,000 "possible twins". Even though we estimated an excess of 40% of pseudo-twins, this still is the world's largest twin population ever collected. The database of possible twins is currently used in population-based studies on multiple sclerosis, Alzheimer's disease, celiac disease, and type 1 diabetes. A system is currently being developed for linking the database with data from mortality and cancer registries. In 2001, the Italian Government, through the Ministry of Health, financed a broad national research program on twin studies, including the establishment of a national twin registry. Among all the possible twins, a sample of 500,000 individuals are going to be contacted and we expect to enrol around 120,000 real twin pairs in a formal Twin Registry. According to available financial resources, a sub sample of the enrolled population will be asked to donate DNA. A biological bank from twins will be then implemented, guaranteeing information on future etiological questions regarding genetic and modifiable factors for physical impairment and disability, cancers, cardiovascular diseases and other age related chronic illnesses.     

A low-resolution 3D model of the tetrameric alcohol dehydrogenase from Sulfolobus solfataricus

We describe the computation of a model of the thermophilic NAD-dependent homotetrameric alcohol dehydrogenase from the archaeon Sulfolobus solfataricus (SsADH). Modeling is based on the knowledge that each monomer contains two Zn ions with catalytic and structural function, respectively. In the database of known structures, proteins with similar functions are either dimers containing two zinc ions per monomer or tetramers with one zinc ion per monomer. In any case, the sequence identity of the target to the possible templates is low. A threading procedure is therefore developed which includes constraints taking into account residue conservation both at the zinc ion binding and at the monomer-monomer interaction sites in the tetrameric unit. The model is consistent with previously reported data. Furthermore, cross-linking experiments are described which support the computed tetrameric model.     

Xanthine oxidase and aldehyde oxidase: a simple procedure for the simultaneous purification from rat liver

Aldehyde oxidase (AO) and xanthine oxidase (XO) are cytosolic enzymes that have been involved in some pathological conditions and play an important role in the biotransformation of drugs and xenobiotics. The increasing interest in these enzymes demands for a simple and rapid procedure for their purification. This paper describes for the first time a method that allows simultaneous purification of both enzymes from the same batch of rat livers. It involves few steps, is reproducible and offers high enzyme yields with high specific activities. The rat liver homogenate was fractionated by heat denaturation and by ammonium sulphate precipitation to give a crude extract containing both enzymes. This extract was chromatographed on an Hydroxyapatite column that completely separated AO from XO. Further purification of XO by anion exchange chromatography on a Q-Sepharose Fast Flow column resulted in a highly purified (1200-fold) preparation, with a specific activity of 3.64 U/mg and with a 20% yield. AO was purified about 1000-fold at a yield of 15%, with a specific activity of 3.48 U/mg, by affinity chromatography on Benzamidine-Sepharose 6B. The purified enzymes gave single bands of approximately 300 kDa on a polyacrylamide gel gradient electrophoresis and displayed the characteristic absorption spectra of highly purified enzymes.     

Novel Proteorhodopsin variants from the Mediterranean and Red Seas

Proteorhodopsins, ubiquitous retinylidene photoactive proton pumps, were recently found in the widespread uncultured SAR86 bacterial group in oceanic surface waters. To survey proteorhodopsin diversity, new degenerate sets of proteorhodopsin primers were designed based on a genomic proteorhodopsin gene sequence originating from an Antarctic fosmid library. New proteorhodopsin variants were identified in Red Sea samples that were most similar to the original green-light absorbing proteorhodopsins found in Monterey Bay California. Unlike green-absorbing proteorhodopsins however, these new variants contained a glutamine residue at position 105, the same site recently shown to control spectral tuning in naturally occurring proteorhodopsins. Different proteorhodopsin variants were also found in the Mediterranean Sea. These proteorhodopsins formed new and distinctive proteorhodopsin groups. Phylogenetic analyses show that some of the new variants were very different from previously characterized proteorhodopsins, and formed the deepest branching groups found so far among marine proteorhodopsins. The existence of these varied proteorhodopsin sequences suggests that this class of proteins has undergone substantial evolution. These variants could represent functionally divergent paralogous genes, derived from the same or similar species, or orthologous proteorhodopsins that are distributed amongst divergent planktonic microbial taxa.