Evaluation of the clastogenicity and anticlastogenicity of the carotenoid bixin in human lymphocyte cultures

Carotenoids are regarded as effective antioxidants, antimutagenic and anticarcinogenic agents. Annatto, a red-yellow extract obtained from seeds of Bixa orellana L. is a mixture of several carotenoids and one of them bixin (BXN), is known as its major coloring compound. Studies on BXN clastogenicity and anticlastogenicity in cultured human lymphocytes have not been reported so far. Therefore, the present study was undertaken to investigate the ability of BXN to induce chromosomal aberrations in human lymphocytes in vitro and to examine the possible anticlastogenic effect of this carotenoid in chromosomal damage induced by the clastogen cisplatin (cDDP). Human blood samples were obtained from six healthy, non-smoking volunteers; two females and four males aged 18-35 years. The concentrations of BXN (1.0; 2.5; 5.0 or 10 microg/mL) tested in combination with cDDP were established on the basis of mitotic index (MI) measurements. The data showed that BXN was not cytotoxic or clastogenic, when compared to untreated control. A marked decrease in the MI values compared to the untreated control and an increased percentage of aberrant metaphases was seen in all cultures treated with cDDP. The carotenoid efficiency in reducing the inhibitory effect of cDDP on lymphocyte MI is concentration-dependent. Cultures simultaneously treated with BXN and cDDP showed a statistically significant reduction in total chromosomal aberrations and aberrant metaphases. In our experiments, BXN may have acted as an antioxidant by intercepting free radicals generated by cDDP. The data obtained in the present study suggest that dietary carotenoids may act as protective agents against clastogenic effects of antitumor agents. However, extensive studies are necessary to elucidate the mechanism of action of BXN before its therapeutic use.     

The cadherin superfamily in Anopheles gambiae: a comparative study with Drosophila melanogaster

The cadherin superfamily is a diverse and multifunctional group of proteins with extensive representation across genomes of phylogenetically distant species that is involved in cell-cell communication and adhesion. The mosquito Anopheles gambiae is an emerging model organism for the study of innate immunity and host-pathogen interactions, where the malaria parasite induces a profound rearrangement of the actin cytoskeleton at critical stages of infection. We have used bioinformatics tools to retrieve present sequence knowledge about the complete repertoire of cadherins in A. gambiae and compared it to that of the fruit fly, Drosophila melanogaster. In A. gambiae, we have identified 43 genes coding for cadherin extracellular domains that were re-annotated to 38 genes and represent an expansion of this gene family in comparison to other invertebrate organisms. The majority of Drosophila cadherins show a 1 : 1 Anopheles orthologue, but we have observed a remarkable expansion in some groups in A. gambiae, such as N-cadherins, that were recently shown to have a role in the olfactory system of the fruit fly. In vivo dsRNA silencing of overrepresented genes in A. gambiae and other genes showing expression at critical tissues for parasite infection will likely advance our understanding of the problems of host preference and hostpathogen interactions in this mosquito species.     

Energetic metabolism of Chromobacterium violaceum

Chromobacterium violaceum is a free-living microorganism, normally exposed to diverse environmental conditions; it has a versatile energy-generating metabolism. This bacterium is capable of exploiting a wide range of energy resources by using appropriate oxidases and reductases. This allows C. violaceum to live in both aerobic and anaerobic conditions. In aerobic conditions, C. violaceum is able to grow in a minimal medium with simple sugars, such as glucose, fructose, galactose, and ribose; both Embden-Meyerhoff, tricarboxylic acid and glyoxylate cycles are used. The respiratory chain supplies energy, as well as substrates for other metabolic pathways. Under anaerobic conditions, C. violaceum metabolizes glucose, producing acetic and formic acid, but not lactic acid or ethanol. C. violaceum is also able to use amino acids and lipids as an energy supply.     

Xylanases of marine fungi of potential use for biobleaching of paper pulp

Microbial xylanases that are thermostable, active at alkaline pH and cellulase-free are generally preferred for biobleaching of paper pulp. We screened obligate and facultative marine fungi for xylanase activity with these desirable traits. Several fungal isolates obtained from marine habitats showed alkaline xylanase activity. The crude enzyme from NIOCC isolate 3 (Aspergillus niger), with high xylanase activity, cellulase-free and unique properties containing 580 U l^–1 xylanase, could bring about bleaching of sugarcane bagasse pulp by a 60 min treatment at 55°C, resulting in a decrease of ten kappa numbers and a 30% reduction in consumption of chlorine during bleaching. The culture filtrate showed peaks of xylanase activity at pH 3.5 and pH 8.5. When assayed at pH 3.5, optimum activity was detected at 50°C, with a second peak of activity at 90°C. When assayed at pH 8.5, optimum activity was seen at 80°C. The crude enzyme was thermostable at 55°C for at least 4 h and retained about 60% activity. Gel filtration of the 50–80% ammonium sulphate-precipitated fraction of the crude culture filtrate separated into two peaks of xylanase with specific activities of 393 and 2,457 U (mg protein)^–1. The two peaks showing xylanase activity had molecular masses of 13 and 18 kDa. Zymogram analysis of xylanase of crude culture filtrate as well as the 50–80% ammonium sulphate-precipitated fraction showed two distinct xylanase activity bands on native PAGE. The crude culture filtrate also showed moderate activities of -xylosidase and -l-arabinofuranosidase, which could act synergistically with xylanase in attacking xylan. This is the first report showing the potential application of crude culture filtrate of a marine fungal isolate possessing thermostable, cellulase-free alkaline xylanase activity in biobleaching of paper pulp.     

Atherina punctata and Atherina lagunae (Pisces,Atherinidae), new species in the Mediterranean Sea. 1. Biometric investigations of three Atherinid species

Discriminative canonical analysis of 87 biometric parameters in the marine and lagoon atherinids of 'Atherina boyeri' complex from the Mediterranean Sea helps defining three distinct atherinid groups: marine punctuated, marine unpunctuated and lagoon atherinids. Each atherinid group constitutes a clearly independent original entity. Besides, each one is a more or less heterogeneous group with geographical disparities, characterising specimen collected from France and Tunisia. Concordance of biometric, biochemical and genetic results as well help define two new species of atherinids: Atherina punctata = punctuated marine atherinids, and Atherina lagunae = atherinids living in lagoon environments.     

RAPD analysis of herbicide-resistant Brasilian rice lines produced via mutagenesis

Over the last two decades, mutational techniques have become one of the most important tools available to progressive rice- breeding programs. In a mutation-breeding program initiated in 1999 at the Instituto Agron?mico of Campinas, SP, Brazil, a rice line, IAC103, was selected for mutational studies with gamma radiation and ethyl methyl sulfonate mutagenesis, with the aim of developing a herbicide-resistant crop. After mutagenesis, surviving plants were exposed to glufosinate to check for herbicide resistance, which was examined up to the second generation. A detailed RAPD analysis was made of the resistant plants. Eighty Operon technology primers were tested and 10 were selected for a detailed study of RAPD markers that could tag herbicide resistance genes. Resistant and susceptible lines produced variation in the RAPD patterns and certain bands were found only in certain lines. These results suggest genetic ligation that will be confirmed through a genetic segregation study.     

Expansions and contractions in 36-bp minisatellites by gene conversion in yeast

The instability of simple tandem repeats, such as human minisatellite loci, has been suggested to arise by gene conversions. In Saccharomyces cerevisiae, a double-strand break (DSB) was created by the HO endonuclease so that DNA polymerases associated with gap repair must traverse an artificial minisatellite of perfect 36-bp repeats or a yeast Y' minisatellite containing diverged 36-bp repeats. Gene conversions are frequently accompanied by changes in repeat number when the template contains perfect repeats. When the ends of the DSB have nonhomologous tails of 47 and 70 nucleotides that must be removed before repair DNA synthesis can begin, 16% of gene conversions had rearrangements, most of which were contractions, almost always in the recipient locus. When efficient removal of nonhomologous tails was prevented in rad1 and msh2 strains, repair was reduced 10-fold, but among survivors there was a 10-fold reduction in contractions. Half the remaining events were expansions. A similar decrease in the contraction rate was observed when the template was modified so that DSB ends were homologous to the template; and here, too, half of the remaining rearrangements were expansions. In this case, efficient repair does not require RAD1 and MSH2, consistent with our previous observations. In addition, without nonhomologous DSB ends, msh2 and rad1 mutations did not affect the frequency or the distribution of rearrangements. We conclude that the presence of nonhomologous ends alters the mechanism of DSB repair, likely through early recruitment of repair proteins including Msh2p and Rad1p, resulting in more frequent contractions of repeated sequences.     

Association of the CHE2 locus with body mass index and butyrylcholinesterase activity

The butyrylcholinesterase (BChE; EC 3.1.1.8) activities of two electrophoretic bands of the CHE2 C5+ phenotype--C5 and C(OF) (other forms)--were quantified by densitometry in 100 individuals. The activity data suggested that, in addition to determining C5, the CHE2*C5+ allele also increases the level of other BChE forms. Since the relative activity of C5 showed the highest correlation coefficient with weight when compared with the other BChE activity variables (total, absolute C5, and absolute C(OF)), its median activity level was used for the classification of CHE2 C5+ phenotypes (faint and intense). Mean body mass index (BMI) was compared among the CHE2 locus phenotypes-controlled by sex, age, and ethnic group. It was shown that the intense CHE2 C5+ phenotype presents a significantly lower (p < 0.001) mean BMI (23.2) than the other phenotypes (faint CHE2 C5+ = 25.2; CHE2 C5- = 25.4). It seems that the relative COF activity is positively associated with fat storage, since CHE2 C5- and faint CHE2 C5+ phenotypes showed higher mean BMI than the intense CHE2 C5+ phenotype. Our hypothesis is that the presence of C5 in a relatively high proportion leads to less fat storage.     

Application of AFLP, RAPD and ISSR markers to genetic mapping of European and Japanese larch

Genetic linkage maps have been increasingly developed for a wide variety of plants, using segregating populations such as F₂s or backcrosses between inbred lines. These pedigrees are rarely available in outbred species like forest trees which have long generation times. Thus genetic mapping studies have to use peculiar pedigrees and markers in appropriate configurations. We constructed single-tree genetic linkage maps of European larch (Larix decidua Mill.) and Japanese larch [Larix kaempferi (Lamb.) Carr.] using segregation data from 112 progeny individuals of an hybrid family. A total of 266 markers (114 AFLP, 149 RAPD and 3 ISSR loci) showing a testcross configuration, i.e.heterozygous in one parent and null in the other parent, were grouped at LOD 4.0, θ=0.3. The maternal parent map (L. decidua)consisted of 117 markers partitioned within 17 linkage groups (1152 cM) and the paternal parent map (L. kaempferi) had 125 markers assembled into 21 linkage groups (1206 cM). The map distance covered by markers was determined by adding a 34.7-cM independence distance at the end of each group and unlinked marker. It reached 2537 cM and 2997 cM respectively for European larch and Japanese larch, and represented respectively a 79.6% and 80.8% coverage of the overall genome. A few 3:1 segregating markers were used to identify homologous linkage groups between the European larch and the Japanese larch genetic maps. The PCR-based molecular markers allowed the construction of genetic maps, thus ensuring a good coverage of the larch genome for further QTL detection and mapping studies. Received: 15 March 1999 / Accepted: 29 March 1999