The role of tetraploids in the sexual-asexual cycle in dandelions (Taraxacum).

Apomictic plants often produce pollen that can function in crosses with related sexuals. Moreover, facultative apomicts can produce some sexual offspring. In dandelions, Taraxacum, a sexual-asexual cycle between diploid sexuals and triploid apomicts, has been described, based on experimental crosses and population genetic studies. Little is known about the actual hybridization processes in nature. We therefore studied the sexual-asexual cycle in a mixed dandelion population in the Netherlands. In this population, the frequencies of sexual diploids and triploids were 0.31 and 0.68, respectively. In addition, less than 1% tetraploids were detected. Diploids were strict sexuals, triploids were obligate apomicts, but tetraploids were most often only partly apomictic, lacking certain elements of apomixis. Tetraploid seed fertility in the field was significantly lower than that of apomictic triploids. Field-pollinated sexual diploids produced on average less than 2% polyploid offspring, implying that the effect of hybridization in the 2x-3x cycle in Taraxacum will be low. Until now, 2x-3x crosses were assumed to be the main pathway of new formation of triploid apomicts in the sexual-asexual cycle in Taraxacum. However, tetraploid pollen donors produced 28 times more triploid offspring in experimental crosses with diploid sexuals than triploid pollen donors. Rare tetraploids may therefore act as an important bridge in the formation of new triploid apomicts.

A genetic linkage map of the diplosporous chromosomal region in Taraxacum officinale (common dandelion; Asteraceae).

In this study, we mapped the diplosporous chromosomal region in Taraxacum officinale, by using amplified fragment length polymorphism technology (AFLP) in 73 plants from a segregating population. Taraxacum serves as a model system to investigate the genetics, ecology, and evolution of apomixis. The genus includes sexual diploid as well as apomictic polyploid, mostly triploid, plants. Apomictic Taraxacum is diplosporous, parthenogenetic, and has autonomous endosperm formation. Previous studies have indicated that these three apomixis elements are controlled by more than one locus in Taraxacum and that diplospory inherits as a dominant, monogenic trait ( Ddd; DIP). A bulked segregant analysis provided 34 AFLP markers that were linked to DIP and were, together with two microsatellite markers, used for mapping the trait. The map length was 18.6 cM and markers were found on both sides of DIP, corresponding to 5.9 and 12.7 cM, respectively. None of the markers completely co-segregated with DIP. Eight markers were selected for PCR-based marker development, of which two were successfully converted. In contrast to all other mapping studies of apomeiosis to date, our results showed no evidence for suppression of recombination around the DIP locus in Taraxacum. No obvious evidence for sequence divergence between the DIP and non- DIP homologous loci was found, and no hemizygosity at the DIP locus was detected. These results may indicate that apomixis is relatively recent in Taraxacum.

Polytetrafluoroethylene femorotibial bypass grafting: 5-year patency and limb salvage.

For a femorotibial bypass graft the material of choice is autologous vein. The question remains whether prosthetic material is a reasonable alternative for limb salvage, if autologous vein is not available. From 1991 to 1998, 83 consecutive femorotibial bypass procedures were performed in 70 patients. Thin-walled, ringed 6-mm polytetrafluoroethylene (PTFE) was used, as autologous vein was not available. The indication for the femorotibial bypass was critical ischemia due to atherosclerotic occlusive disease in all cases. Three patients died in the hospital or within 30 days of the operation, resulting in a perioperative mortality rate of 3.6%. After 5 years, 33 patients had died (40%) and 3 patients were lost to follow-up (3.6%). Primary patency was 64.2% after 6 weeks and 18% +/- 5% after 5 years. Secondary patency was 74.1% after 6 weeks and 22.3% +/- 6% after 5 years. When we compared primary and secondary patency for distal anastomosis, there was no difference between the three crural arteries. The limb salvage rate was 61.9% +/- 7% after 5 years. From these results we conclude that, with a limb salvage of 61.9%, PTFE is an acceptable alternative for a femorotibial bypass graft in patients with critical ischemia, if autologous vein is not available.

A comparison of transcutaneous bilirubinometers: SpectRx BiliCheck versus Minolta AirShields.

BACKGROUND: Two devices are available for making transcutaneous estimates of serum bilirubin (SBR): the Minolta AirShields JM102 and the new SpectRx BiliCheck. OBJECTIVES: (a) To measure how well the readings produced by these devices agree with SBR measured in the laboratory; (b) to estimate for each device, the proportion of infants with clinical jaundice who would require blood sampling if the device was used as a screening tool to detect infants with SBR > or = 250 micromol/l. DESIGN: Prospective cohort study of jaundiced infants who required SBR at < or = 20 days of postnatal age. Those who had received phototherapy or exchange transfusion were excluded. SETTING: Tertiary neonatal service in South-East Scotland. INTERVENTIONS: Within 30 minutes of SBR sampling, transcutaneous bilirubinometry was performed using one Minolta and two SpectRx devices (designated A and B). RESULTS: Sixty-four neonates were enrolled, 19 of which were preterm (31-35 weeks). The 95% confidence intervals of a device reading corresponding to SBR were +/- 66.7, +/- 67.9, and +/- 66.4 micromol/l respectively. Using the devices to identify all SBR > or = 250 micromol/l would reduce SBR sampling by 23%, 16%, and 20% respectively. CONCLUSIONS: Given that SBR levels range from 50 to 400 micromol/l in jaundiced infants, the 95% confidence intervals of the devices are wide at +/- 67 micromol/l. The SpectRx can be used as a screening tool for hyperbilirubinaemia but there is no advantage in using it over the Minolta.

Skeletal muscle fibre-type shifting and metabolic profile in patients with chronic obstructive pulmonary disease.

The aim of this study was to examine the nature of fibre-type redistribution in relation to fibre metabolic profile in the vastus lateralis in chronic obstructive pulmonary disease (COPD) and COPD subtypes. Fifteen COPD patients (eight with emphysema stratified by high-resolution computed tomography) and 15 healthy control subjects were studied. A combination of myofibrillar adenosine triphosphatase staining and immunohistochemistry was used to identify pure, as well as hybrid fibre types. For oxidative capacity, fibres were stained for cytochrome c oxidase and succinate dehydrogenase activities, and glycogen phosphorylase for glycolytic capacity. The proportion of type-I fibres in COPD patients was markedly lower (16% versus 42%), especially in emphysema, and the proportion of hybrid fibres was higher (29% versus 16%) compared to controls. The proportion of fibres staining positive for oxidative enzymes was lower in COPD patients, which correlated with the proportion of type-I fibres. In COPD oxidative capacity was lower within IIA fibres. The authors conclude that fibre-type transitions are involved in the fibre-type redistribution in chronic obstructive pulmonary disease. Low oxidative capacity is closely related to the proportion of type-I fibres, but an additional reduction of oxidative enzyme activity is present within IIA fibres. Fibre-type abnormalities may be aggravated in emphysema.

Meiotic recombination in sexual diploid and apomictic triploid dandelions (Taraxacum officinale L.).

Taraxacum officinale L. (dandelion) is a vigorous weed in Europe with diploid sexual populations in the southern regions and partially overlapping populations of diploid sexuals and triploid or tetraploid apomicts in the central and northern regions. Previous studies have demonstrated unexpectedly high levels of genetic variation in the apomictic populations, suggesting the occurrence of genetic segregation in the apomicts and (or) hybridization between sexual and apomictic individuals. In this study we analysed meiosis in both sexual diploid and apomictic triploid plants to find mechanisms that could account for the high levels of genetic variation in the apomicts. Microscopic study of microsporocytes in the triploid apomicts revealed that the levels of chromosome pairing and chiasma formation at meiotic prophase I were lower than in that of the sexual diploids, but still sufficient to assume recombination between the homologues. Nomarski DIC (differential interference contrast) microscopy of optically cleared megasporocytes in the apomicts demonstrated incidental formation of tetrads, which suggests that hybridization can occur in triploid apomicts.

Crosses between sexual and apomictic dandelions (Taraxacum). I. The inheritance of apomixis.

Some dandelions, Taraxacum, are diplosporous gametophytic apomicts. Crosses between closely related diploid sexuals and triploid apomicts were made to study the inheritance of apomixis. Seed-set was less than one-third of that in diploid x diploid crosses, probably because of the inviability of aneuploid pollen or zygotes. Almost 90% of the viable offspring were diploid and the result of selfing, as was shown by a discriminating allozyme marker. Aneuploid outcross pollen had a mentor effect on self-pollen, causing a breakdown of the sporophytic self-incompatibility system. A similar phenomenon has been reported before in wide crosses. Of the 26 allozyme-confirmed hybrids, four were diploids, 15 were triploids and seven were tetraploids. Diploid hybrids were significantly less frequent than triploid hybrids, suggesting either low fitness of haploid pollen or more numerous formation of diploid pollen. Emasculation and bagging of flowers indicated apomictic seed-set in none of the diploid, in one-third of the triploid and in all of the tetraploid hybrids. All apomictic hybrids showed partial seed-set, but additional cross-pollination did not increase seed-set. Cytological analysis of the F2 progeny confirmed that partial apomixis was caused by semisterility and not by residual sexuality (facultative apomixis). The difference in segregation for apomixis between triploid and tetraploid hybrids may be because the triploids originated from partially reduced diploid pollen grains, whereas the tetraploids originated from unreduced triploid pollen grains.

Crosses between sexual and apomictic dandelions (Taraxacum). II. The breakdown of apomixis.

Some dandelions are diplosporous gametophytic apomicts. In order to study the inheritance and breakdown of apomixis, crosses were made between diploid sexuals and triploid apomicts. To investigate their breeding system, four nonapomictic diploid and 10 nonapomictic triploid hybrids were pollinated with diploids and the progenies were analysed. Seed fertility was significantly reduced in two diploid hybrids. Nine triploid hybrids were fertile and could be classified into three types, with respect to the composition of their progenies. Type A produced n+n hybrids. Type B produced either a mixture of n + n and 2n + n hybrids, or a mixture of pseudogamous 2n + 0 apomicts and 2n + n hybrids. Type C produced exclusively 2n + n hybrids. Inheritance of a microsatellite marker strongly suggested that 2n egg cells in type C plants were produced by a first division restitution mechanism. As in apomicts, microsporogenesis in type C plants was reductional. This suggests that type C plants are diplosporous plants that lack parthenogenesis. Such plants are very rare in other apomictic plant species. It is concluded that 'elements of apomixis', diplospory and parthenogenesis, can be uncoupled. This is inconsistent with the single-locus model for apomixis in Taraxacum as suggested by Mogie (1992). Instead, our results suggest that several loci are involved in the genetic control of apomixis in Taraxacum.

A simple method to change geometrical proportions of muscles by means of a tibial lengthening osteotomy in the rat.

A method is designed to change geometrical proportions of muscles in the rat. After a tibial osteotomy, the application of a stainless steel prosthesis resulted in an elongation of about 8% of the tibia. In 23 out of 28 operated rats a correct application of the prosthesis was maintained till the rats were sacrificed. The operation caused hardly any muscle tissue damage. No disuse of the manipulated leg was observed in the operated rats.

Enhanced cytostatic effectiveness of aniline mustard against 7,12-dimethylbenz[a]anthracene-induced rat mammary tumors during regression in response to ovariectomy.

Sprague-Dawley rats bearing 7,12-dimethylbenz[a]anthracene (DMBA)-induced mammary tumors were treated with either of two aromatic alkylating agents, aniline mustard or melphalan, alone or combined with ovariectomy. Both drugs were applied once a week for 8 weeks. Eight-four percent of the tumors responded to ovariectomy, 38% regressing completely and 46% regressing partially. Aniline mustard, though virtually ineffective as a single agent, appeared synergistic with ovariectomy: a 100% regression rate (72% complete, 28% partial) was observed for this combination. Treatment with melphalan was as effective as ovariectomy, but the combination of melphalan with ovariectomy was no more effective than either treatment alone. The end product of aniline mustard metabolism, p-hydroxyaniline mustard O-glucuronide, may be more extensively activated by beta-glucuronidase in hormonally regressing than in growing or stationary tumors. Intratumoral levels of beta-glucoronidase occurring in DMBA-induced tumors 4 days after ovariectomy were found to be similar to those in the aniline mustard-sensitive mouse plasma cell tumor ADJ/PC6. It remains to be more extensively studied whether an effect of endocrine treatment on tumor beta-glucuronidase levels, and possibly on intracellular distribution of enzyme, could be used therapeutically. An effectively scheduled cytostatic treatment (with a drug conjugate such as that formed metabolically from aniline mustard) in conjunction with ovariectomy might be effective in the treatment of hormone-responsive breast cancer.

Chaotropic resolution of high molecular weight (type I) NADH dehydrogenase, and reassociation of flavin-rich (type II) and flavin-poor subunits.

1. Type-I NADH dehydrogenase (Complex I) was solubilized and dissociated into subunits by NaClO4. NADH slows the dissociation. On subsequent stepwise addition of (NH4)2SO4 the dissociation is partly reversed, as is to be expected from the opposing effects of ClO-4 and SO-24, which are on the salting-in and salting-out sides, respectively, of the lyotropic series. 2. In consequence, the aggregates of subunits that are separated by (NH4)2-SO4 fractionation consist of randomly associated subunits as well as fragments of Type I enzyme. The fraction precipitating at 27% satd. (NH4)2SO4 is flavin-poor, that remaining soluble at 55% satd. (NH4)2SO4 flavin-rich and those separating between 27 and 55% satd. (NH4)2SO4 intermediate in composition. 3. The fraction remaining soluble at 55% satd. (NH4)2SO4 contains the purified low-molecular-weight iron-sulphur flavoprotein (Type-II dehydrogenase). It is a dimer consisting of one molecule of FMN, one 28-kilodalton and one 56-kilodalton subunit per protomer. Work of others indicates that it contains 4 Fe and 4 acid-labile S atoms per molecule of FMN. Sometimes the fraction remaining soluble at 55% satd. (NH4)2SO4 contained an additional small subunit (12 kilodaltons) and four additional Fe and acid labile S atoms per protomer. The sedimentation coefficients (s020,w) of the two preparations were 5.3 and 6.6 S, respectively, with calculated frictional ratios of 1.5 and 1.24, respectively. 4. The intermediate fractions are mixtures of the various subunits present in Complex I. Specifically a fraction separating at 55% satd. (NH4)2SO4 was found to be a mixture of two fragments, the pure iron-sulphur flavoprotein and a 26-S fragment that contained per protomer four subunits of 12 kilodaltons, one each of 28, 32, 56 and 77 kilodaltons, one molecule of FMN and 20 Fe and acid-labile S atoms. It was probably tetrameric or even larger. 5. The oxidoreductase activity of the intermediate fractions is dependent on the protein concentration, the activity with ferricyanide increasing and that with ferricytochrome c decreasing with increasing protein concentration. This is interpreted as an increased association of subunits present in the intermediate fractions. Similar results are obtained when flavin-rich and flavin-poor fractions are mixed. The association is cooperative. NADH favours the association of the subunits. 6. Association of the subunits is accompanied by a 10-fold increase in k2 (rate constant for intramolecular electron flow), a 10-fold decrease of the accessibility of ferricyanide to the reduced enzyme and a 10(4)-fold decrease of the accessibility of ferricytochrome c. The Ks (NADH) is also decreased. Although the changes are in the direction to be expected from a conversion of Type II enzyme to Type I, the value of k2 is still much less than in the latter enzyme.