Diversity in coffee assessed with SSR markers: structure of the genus Coffea and perspectives for breeding.

The present study shows transferability of microsatellite markers developed in the two cultivated coffee species (Coffea arabica L. and C. canephora Pierre ex Froehn.) to 15 species representing the previously identified main groups of the genus Coffea. Evaluation of the genetic diversity and available resources within Coffea and development of molecular markers transferable across species are important steps for breeding of the two cultivated species. We worked on 15 species with 60 microsatellite markers developed using different strategies (SSR-enriched libraries, BAC libraries, gene sequences). We focused our analysis on 4 species used for commercial or breeding purposes. Our results establish the high transferability of microsatellite markers within Coffea. We show the large amount of diversity available within wild species for breeding applications. Finally we discuss the consequences for future comparative mapping studies and breeding of the two cultivated species.     

Possible Case of Maternal Transmission of Feline Spongiform Encephalopathy in a Captive Cheetah

Feline spongiform encephalopathy (FSE) is considered to be related to bovine spongiform encephalopathy (BSE) and has been reported in domestic cats as well as in captive wild cats including cheetahs, first in the United Kingdom (UK) and then in other European countries. In France, several cases were described in cheetahs either imported from UK or born in France. Here we report details of two other FSE cases in captive cheetah including a 2^nd case of FSE in a cheetah born in France, most likely due to maternal transmission. Complete prion protein immunohistochemical study on both brains and peripheral organs showed the close likeness between the two cases. In addition, transmission studies to the TgOvPrP4 mouse line were also performed, for comparison with the transmission of cattle BSE. The TgOvPrP4 mouse brains infected with cattle BSE and cheetah FSE revealed similar vacuolar lesion profiles, PrP^d brain mapping with occurrence of typical florid plaques. Collectively, these data indicate that they harbor the same strain of agent as the cattle BSE agent. This new observation may have some impact on our knowledge of vertical transmission of BSE agent-linked TSEs such as in housecat FSE, or vCJD.     

Radioisotopic assay of picomolar amounts of coenzyme A

A two-step method of determining reduced coenzyme A (CoASH) concentrations in tissue or cell extracts is described. In the first step, CoASH is reacted with acetylphosphate in a reaction catalyzed by phosphotransacetylase to yield acetyl-CoA. Acetyl-CoA is then condensed with [14C]oxaloacetate by citrate synthase to give [14C]citrate. This method allows the measurement of 10-200 pmol of CoASH. By omitting the phosphotransacetylase step, measurement of the same amount of acetyl-CoA is possible.     

Mass fragmentographic determination of xanthine and hypoxanthine in biological fluids

The method presented for the simultaneous determination of xanthine and hypoxanthine, uses mass-fragmentography in the electron impact (EI) mode, after the gas chromatographic separation of butylated derivatives. Butylation, rather than methylation, is used in order to avoid interference coming from exogenous caffeine, which is frequently encountered. [7,9-¹⁵N]Xanthine is used as the internal standard, and for each sample, a blank is obtained by xanthine oxidase reaction. In the biological fluids studied the sensitivity was about 50 ng/ml.     

Inactivation of human placental aromatase by 6 alpha- and 6 beta-hydroperoxyandrostenedione

The epimeric 6 alpha- and 6 beta-hydroperoxy derivatives of androstendione caused irreversible inactivation of human placental aromatase. Microsomes from term-placentae were first preincubated in the presence of increasing concentrations of the hydroperoxides. The microsomes were then washed free of steroids and its residual aromatase activity was assayed by the tritium-exchange method to [3H]water. Aromatase activity decreased in a time-, and concentration-dependent manner; the axial, beta-hydroperoxy epimer was the slightly stronger inactivator. Less inactivation occurred when during the preincubation stage the natural aromatase substrate, androstenedione, or the anti-oxidant, dithiothreitol, was added. The sulfhydryl reagent, p-hydroxy-mercuribenzoate, decreased this protective effect. The inactivation is not dependent on the presence of NADPH. Both steroids induced a Type I difference spectrum with a Ks of 0.167 microM and 0.163 microM for the 6 alpha-, and the 6 beta-hydroperoxyandrostenedione, respectively. We suggest that these 6-hydroperoxyandrogens may function as active-site directed inhibitors and inactivators of estrogen synthetase through oxidation of cysteine residues.     

Microbial Isotopic Fractionation of Perchlorate Chlorine

Perchlorate contamination can be microbially respired to innocuous chloride and thus can be treated effectively. However, monitoring a bioremediative strategy is often difficult due to the complexities of environmental samples. Here we demonstrate that microbial respiration of perchlorate results in a significant fractionation (~−15‰) of the chlorine stable isotope composition of perchlorate. This can be used to quantify the extent of biotic degradation and to separate biotic from abiotic attenuation of this contaminant.     

The nuclear genome is involved in heteroplasmy control in a mitochondrial mutant strain of Drosophila subobscura.

Most (78%) mitochondrial genomes in the studied mutant strain of Drosophila subobscura have undergone a large-scale deletion (5 kb) in the coding region. This mutation is stable, and is transmitted intact to the offspring. This animal model of major rearrangements of mitochondrial genomes can be used to analyse the involvement of the nuclear genome in the production and maintenance of these rearrangements. Successive backcrosses between mutant strain females and wild-type males yield a biphasic change in heteroplasmy level: (a) a 5% decrease in mutated genomes per generation (from 78 to 55%), until the nuclear genome is virtually replaced by the wild-type genome (seven to eight crosses); and (b) a continuous decrease of 0.5% per generation when the nuclear context is completely wild-type. In parallel with these changes, NADH dehydrogenase activity, which is halved in the mutant strain (five subunits of this complex are affected by the mutation), gradually increases and stabilizes near the wild-type activity. A return to a nuclear context is accompanied by the opposite phenomena: progressive increase in heteroplasmy level and stabilization at the value seen in the wild-type strain and a decrease in the activity of complex I. These results indicate that the nuclear genome plays an important role in the control of heteroplasmy level and probably in the production of rearranged genomes.