Assessing the colonic microbiome, hydrogenogenic and hydrogenotrophic genes, transit and breath methane in constipation.

BACKGROUND: Differences in the gut microbiota and breath methane production have been observed in chronic constipation, but the relationship between colonic microbiota, transit, and breath tests remains unclear. METHODS: In 25 healthy and 25 constipated females we evaluated the sigmoid colonic mucosal and fecal microbiota using 16S rRNA gene sequencing, abundance of hydrogenogenic FeFe (FeFe-hydA) and hydrogenotrophic (methyl coenzyme M reductase A [mrcA] and dissimilatory sulfite reductase A [dsrA]) genes with real-time qPCR assays, breath hydrogen and methane levels after oral lactulose, and colonic transit with scintigraphy. KEY RESULTS: Breath hydrogen and methane were not correlated with constipation, slow colon transit, or with abundance of corresponding genes. After adjusting for colonic transit, the abundance of FeFehydA, dsrA, and mcrA were greater (P<.005) in colonic mucosa, but not stool, of constipated patients. The abundance of the selected functional gene targets also correlated with that of selected taxa. The colonic mucosal abundance of FeFe-hydA, but not mcrA, correlated positively (P<.05) with breath methane production, slow colonic transit, and overall microbiome composition. In the colonic mucosa and feces, the abundance of hydrogenogenic and hydrogenotrophic genes were positively correlated (P<.05). Breath methane production was not associated with constipation or colonic transit. CONCLUSIONS & INFERENCES: Corroborating our earlier findings with 16S rRNA genes, colonic mucosal but not fecal hydrogenogenic and hydrogenotrophic genes were more abundant in constipated vs. healthy subjects independent of colonic transit. Breath gases do not directly reflect the abundance of target genes contributing to their production.

Genetic structure of Rhododendron ferrugineum at a wide range of spatial scales.

Rhododendron ferrugineum L. (Ericaceae) is a subalpine shrub found throughout the Pyrenees and Alps at elevations of 1600-2200 m. We examined relationships between genetic and geographic distance, using 115 dominant amplified fragment length polymorphism (AFLP) markers to assess genetic structure over a wide range of spatial scales. We sampled 17 sites with distances of 4 km to more than 1000 km between them. At these scales we detected no association between geographic distance and genetic distance between populations. This suggests that genetic drift and gene flow are not in equilibrium for these populations. This pattern could have resulted from recent and rapid postglacial colonization, from more recent human disturbance, or as a function of frequent and random "natural" long-distance colonization. At two of our sites we used transects (two horizontal and two vertical with respect to slope at each site) to sample at distances ranging from 10 m to more than 5000 m. At this scale we observed a positive relationship between genetic and spatial distance along two vertical transects, one at each site. We hypothesize that isolation-by-distance at this smaller scale is a function of restricted gene flow via seed dispersal.

Phylogenetic relationships within Diadasia, a group of specialist bees.

We estimated phylogenetic relationships among species of the bee genus Diadasia, a group of new world, specialist bees. We sequenced approximately 2 kb of the mitochondrial genes cytochrome oxidase subunit I and II and tRNA leucine and approximately 1 kb of the nuclear gene elongation factor 1-alpha for 24 North American Diadasia species, 4 South American species, and five outgroup genera. Parsimony analyses of the two data sets were highly congruent. A combined analysis produced a well-resolved phylogenetic hypothesis that supported the monophyly of Diadasia, but not that of traditional subgenera: Diadasia s. str. was paraphyletic in all analyses. With one exception, the North and South American species formed separate clades, supporting previous hypotheses that two lineages of Diadasia have dispersed from South to North America: a more recent dispersal of D. ochracea and an older dispersal of the ancestor to all other North American species. Different species of Diadasia specialize on pollen from at least five different plant families; the phylogeny presented here, along with known host affinities, indicates that host-switching has been rare.

Genetic relationships and population structure of the endangered Steamboat buckwheat, Eriogonum ovalifolium var. williamsiae (Polygonaceae).

Eriogonum ovalifolium var. williamsiae (Steamboat buckwheat) is a narrow endemic subshrub, known from a single locality in Washoe County, Nevada. We examined genetic structure of the only known population by analyzing patterns of allozyme variation. Our results suggest that Steamboat buckwheat has high genetic variability, with levels of variation similar to that typical of a widespread species rather than a narrow endemic. Genotype frequencies suggest that mating is random. We detected no genetic subdivision of the population. Several clones spanning up to 67 cm were found, but we do not know if such clones are common. We used allozyme data to assess the genetic similarity of var. williamsiae to five other varieties of E. ovalifolium. All six varieties are very similar allozymically with var. williamsiae being the most similar to the widespread var. ovalifolium. Although var. williamsiae and var. ovalifolium are morphologically distinct, their genetic similarity warrants further study to determine whether or not they should be treated as separate taxa. Evidence of male sterility in var. williamsiae plus other data leads us to hypothesize that this taxon might be either a hybrid or undergoing cytoplasmic introgression. Information gathered from this study, in concert with ongoing work on the breeding system of Steamboat buckwheat, should be helpful in forming management strategies for this plant.

Tests of pre- and postpollination barriers to hybridization between sympatric species of Ipomopsis (Polemoniaceae).

The Ipomopsis aggregata species complex (Polemoniaceae) includes species pairs that hybridize readily in nature as well as pairs that meet along contact zones with no apparent hybridization. Artificial hybrids can be made between I. aggregata and I. arizonica, yet morphological intermediates between these two species have not been observed in natural populations. This apparent lack of hybridization is perplexing given that plants of the two species often grow within a few metres of each other and both species have red flowers visited by the same species of hummingbirds. We used trained hummingbirds to examine pollen transfer within and between species. We also hand-pollinated flowers to examine paternal success of heterospecific and conspecific pollen, testing paternity with electrophoretic examination of seeds. Hummingbirds were not simply better at transferring pollen within than between species. Instead, I. arizonica was a better pollen donor so that considerable pollen transfer was observed from I. arizonica to I. aggregata, but very little in the opposite direction. Conversely, once pollen arrived at stigmas, I. arizonica pollen performed very poorly on I. aggregata pistils. However, pollen from I. aggregata could, in some cases, sire seeds on I. arizonica. We hypothesize that hybrids are scarce in nature, in part, because of asymmetric barriers to reproduction: little pollen transfer in one direction and poor pollen performance in the other.

Horsetails and ferns are a monophyletic group and the closest living relatives to seed plants.

Most of the 470-million-year history of plants on land belongs to bryophytes, pteridophytes and gymnosperms, which eventually yielded to the ecological dominance by angiosperms 90 Myr ago. Our knowledge of angiosperm phylogeny, particularly the branching order of the earliest lineages, has recently been increased by the concurrence of multigene sequence analyses. However, reconstructing relationships for all the main lineages of vascular plants that diverged since the Devonian period has remained a challenge. Here we report phylogenetic analyses of combined data--from morphology and from four genes--for 35 representatives from all the main lineages of land plants. We show that there are three monophyletic groups of extant vascular plants: (1) lycophytes, (2) seed plants and (3) a clade including equisetophytes (horsetails), psilotophytes (whisk ferns) and all eusporangiate and leptosporangiate ferns. Our maximum-likelihood analysis shows unambiguously that horsetails and ferns together are the closest relatives to seed plants. This refutes the prevailing view that horsetails and ferns are transitional evolutionary grades between bryophytes and seed plants, and has important implications for our understanding of the development and evolution of plants.

The phylogeny of land plants inferred from 18S rDNA sequences: pushing the limits of rDNA signal?

Previous studies of the phylogeny of land plants based on analysis of 18S ribosomal DNA (rDNA) sequences have generally found weak support for the relationships recovered and at least some obviously spurious relationships, resulting in equivocal inferences of land plant phylogeny. We hypothesized that greater sampling of both characters and taxa would improve inferences of land plant phylogeny based on 18S rDNA sequences. We therefore conducted a phylogenetic analysis of complete (or nearly complete) 18S rDNA sequences for 93 species of land plants and 7 green algal relatives. Parsimony analyses with equal weighting of characters and characters state changes and parsimony analyses weighting (1) stem bases half as much as loop bases and (2) transitions half as much as transversions did not produce substantially different topologies. Although the general structure of the shortest trees is consistent with most hypotheses of land plant phylogeny, several relationships, particularly among major groups of land plants, appear spurious. Increased character and taxon sampling did not substantially improve the performance of 18S rDNA in phylogenetic analyses of land plants, nor did analyses designed to accommodate variation in evolutionary rates among sites. The rate and pattern of 18S rDNA evolution across land plants may limit the usefulness of this gene for phylogeny reconstruction at deep levels of plant phylogeny. We conclude that the mosaic structure of 18S rDNA, consisting of highly conserved and highly variable regions, may contain historical signal at two levels. Rapidly evolving regions are informative for relatively recent divergences (e.g., within angiosperms, seed plants, and ferns), but homoplasy at these sites makes it difficult to resolve relationships among these groups. At deeper levels, changes in the highly conserved regions of small-subunit rDNAs provide signal across all of life. Because constraints imposed by the secondary structure of the rRNA may affect the phylogenetic information content of 18S rDNA, we suggest that 18S rDNA sequences be combined with other data and that methods of analysis be employed to accommodate these differences in evolutionary patterns, particularly across deep divergences in the tree of life.

A Study on the Phylogeny of the Dyer's Woad Rust Fungus and Other Species of Puccinia from Crucifers.

ABSTRACT The identity of a Puccinia species occurring on the introduced weed dyer's woad (Isatis tinctoria) was studied using sequences from the internal transcribed spacer of the nuclear ribosomal DNA. The relationship of this fungus to other Puccinia species occurring on the family Brassicaceae in Europe and North America was examined, and we tested the hypothesis that P. thlaspeos and P. monoica are correlated species. The data suggest that the Puccinia species from dyer's woad is closely related to the North American species P. consimilis and may be derived from an indigenous strain of P. consimilis that switched hosts. Thus, the Puccinia species from dyer's woad is probably native to North America and is unlikely to cause disease epidemics on indigenous plants if used as a biological control agent against dyer's woad. P. thlaspeos appears to be polyphyletic and, therefore, P. thlaspeos and P. monoica do not appear to be correlated species. Additional DNA sequence data will be needed to clarify further the phylogeny of Puccinia species on the family Brassicaceae.

Phylogenetic analysis of restriction-site variation in wild and cultivated Amaranthus species (Amaranthaceae).

Amaranthus includes approximately 60 species, of which three are cultivated as a grain source. Many wild Amaranthus species possess agriculturally desirable traits such as drought and salt tolerance, and pathogen resistance. We examined relationships among wild and cultivated Amaranthus species based upon restriction-site variation in two chloroplast DNA regions and in a nuclear DNA region. The chloroplast regions consisted of (1) an intergenic spacer in transfer RNA genes and (2) the ribulose-1,5-bisphosphate carboxylase gene with a flanking open reading frame. The nuclear region was the internal transcribed spacers ITS-1 and ITS-2 flanking the 5.8S gene in the ribosomal DNA. These regions were amplified by the polymerase chain reaction and digested with a total of 38 restriction endonucleases. We detected 11 potentially informative restriction-site mutations and seven length-polymorphisms among the 28 Amaranthus species. Parsimony analysis was used to find the shortest tree for each separate data set (chloroplast, nuclear, and length) and for two combined matrices (chloroplast/nuclear and all data sets). Overall, there was a low level of variation which generated poorly resolved trees among the 28 species. Congruence analyses revealed that the chloroplast and nuclear data sets were congruent with each other but not to the length data set. The congruence of the chloroplast and nuclear data sets suggested that cytoplasmic gene flow may not be a confounding factor in our analyses. The phylogeny also suggested that drought tolerance evolved independently several times. The molecular phylogeny provides a basis for selection of species pairs for crop development.

Phylogenetic relationships of dennstaedtioid ferns: evidence from rbcL sequences.

The ferns are an ancient group of vascular plants that have yielded a staggering array of systematic problems. Among fern classifications, the number of genera in some families has ranged over 10-fold, and some genera have been treated in up to five different families. Three main groups of leptosporangiate ferns have been recognized; the adiantoid, polypodioid, and dennstaedtioid lines. To clarify relationships among genera and families of dennstaedtioid ferns, we sequenced 1320 bp of the chloroplast gene rbcL from 45 species representing 13 families. Sequence divergence for rbcL averaged 0.9% among species within genera, 10.3% among genera within families, and 14.8% among families, suggesting that the data are appropriate for phylogenetic analysis at the generic and familial levels in ferns. Maximum parsimony analysis resulted in four shortest trees of equal length. The strict consensus tree supported many aspects of previously published hypotheses of relationship based on morphological and cytological variations. For example, the tree ferns (which form a single clade) and Hymenophyllaceae appear as sister groups to the dennstaedtioid ferns on all shortest trees. However, Polypodiaceae and adiantoid ferns, groups traditionally considered separate from the dennstaedtioid families, emerged within the dennstaedtioid clade. This analysis also suggests relationships of some problematical genera, such as Monachosorum, Calochlaena, and Lonchitis. Examination of additional DNA sequences of nuclear genes and developmental studies are needed to evaluate further the relationships suggested by phylogenetic analysis of rbcL sequence data.

The therapeutic use of albumin.

In this review, we condense and summarize the results of studies on the therapeutic use of human albumin to promote the more efficient use of this costly resource. Reports of major controlled and uncontrolled therapeutic trials, reviews, and summary articles published in English between 1972 and 1991 were identified through library and MEDLINE searches. Case series, prospective studies, and blinded therapeutic trials were identified from the bibliographies of these sources. All sources were critically evaluated for information about the comparative physiologic results and patient outcomes of the therapeutic use of albumin solutions, crystalloid solutions, and volume expanders other than albumin. The therapeutic use of albumin is of marginal benefit for many conditions for which it has been administered, apparently because of the body's capacity to quickly compensate for rapid colloid osmotic shifts. Human studies show little or no demonstrable value for albumin when it is administered for nutritional supplementation, wound healing, perioperative fluid replacement, treatment of early thermal injury, or therapy during extensive retroperitoneal surgery (including aortic aneurysm resection). Therapeutic albumin has well-defined value in several special circumstances: large-volume paracentesis in cirrhotic patients, acute nephrotic syndromes with diuretic resistance, organ transplantation, and plasmapheresis. Additional studies are needed to compare the efficacy of albumin with other volume expanders. For most purposes, balanced crystalloid solutions are satisfactory substitutes for colloid volume expanders and can be obtained at a fraction of the cost of colloid volume expanders.

Estimates of gene flow among populations, geographic races, and species in the Ipomopsis aggregata complex.

Interpopulational gene flow within a species can reduce population differentiation due to genetic drift, whereas genetic exchange among taxa can impede speciation. We used allozyme data to estimate gene flow within and among geographic races and species of perennial herbs in the Ipomopsis aggregata complex (Polemoniaceae). Estimates of interpopulational gene flow within taxa from two methods (F statistics and private alleles) were correlated with one another. Gene flow among populations within each geographic race (subspecies) of I. aggregata was relatively high (Nm greater than approximately 1.0). Gene flow was also high among populations of I. arizonica and among four northern populations of I. tenuituba. However, gene flow was low (Nm less than 1.0) for I. tenuituba when a population representing subsp. macrosiphon was included. This is consistent with previous findings that subsp. macrosiphon has had an independent origin and is reproductively, as well as geographically, isolated. A recently developed model, based on hierarchical F statistics, was employed to estimate genetic exchange among taxa. Gene flow estimates were generally high among races of I. aggregata (dNmrace greater than 1.0) but were low among subspecies of I. tenuituba (dNmrace less than 1.0). Consistent with morphological evidence, estimates of interspecific gene flow were moderate between I. aggregata and I. tenuituba, which hybridize in several areas. However, contrary to morphological evidence, we estimated relatively high levels of interspecific gene flow involving I. arizonica. Our results suggest that I. arizonica has hybridized with other species without the transfer of morphological traits.(ABSTRACT TRUNCATED AT 250 WORDS)

Practical approach to the treatment of hypothyroidism.

Hypothyroidism most commonly is the result of primary thyroid failure due to autoimmune thyroiditis, but also may follow treated Graves' disease or may occur as transient hypothyroidism. Laboratory evaluation of thyroid function has been refined by the development of ultrasensitive thyroid-stimulating hormone determinations, allowing accurate diagnosis of primary hypothyroidism and assessment of treatment response. Determining whether therapy is appropriate for patients with subclinical hypothyroidism or euthyroid sick syndrome requires clinical judgment in addition to laboratory evaluation. Thyroid hormone replacement for hypothyroidism is best accomplished with levothyroxine. Factors that may affect serum thyroxine levels include thyroid binding globulin levels, drug interactions and variations in thyroxine absorption and clearance.

Electrophoretic Evidence for Genetic Diploidy in the Bracken Fern (Pteridium aquilinum).

Analysis of isozyme variability demonstrates that bracken fern (Pteridium aquilinum) has a diploid genetic system and expresses solely disomic inheritance patterns. Electrophoretic data indicate that genetically variable progeny are produced in natural populations after intergametophytic mating rather than by a process involving recombination between duplicated unlinked loci. Although some enzymes are encoded by more than one locus, this has resulted from subcellular compartmentalization of isozymes, and there is no evidence of extensive gene duplication resulting from polyploidy. The conclusions reached in this report differ from those which propose polyploidy as an adaptive mechanism for maintaining genetic variability in Pteridium and other homosporous pteridophytes.