Quantification of bacterial groups within human fecal flora by oligonucleotide probe hybridization

To investigate the population structure of the predominant phylogenetic groups within the human adult fecal microbiota, a new oligonucleotide probe designated S-G-Clept-1240-a-A-18 was designed, validated, and used with a set of five 16S rRNA-targeted oligonucleotide probes. Application of the six probes to fecal samples from 27 human adults showed additivity of 70% of the total 16S rRNA detected by the bacterial domain probe. The Bacteroides group-specific probe accounted for 37% +/- 16% of the total rRNA, while the enteric group probe accounted for less than 1%. Clostridium leptum subgroup and Clostridium coccoides group-specific probes accounted for 16% +/- 7% and 14% +/- 6%, respectively, while Bifidobacterium and Lactobacillus groups made up less than 2%.     

Genetic divergence within the Drosophila mayaguana subcluster,a closely related triad of Caribbean species in the repleta species group

The mayaguana triad of the Drosophila repleta species group includes D. mayaguana, D. straubae, and D. parisiena, the latter two of which are very similar when examined morphologically. Many morphological characters used to define these taxa are quantitative and overlap substantially among some forms--it is only through suites of such characters that species can be identified. We apply Population Aggregation Analysis and tree building methods to five rapidly evolving gene regions--the mitochondrial AT rich region and the nuclear acetylcholinesterase, hunchback, mastermind, and vestigial loci to test the morphological species delineations within the morphocryptic mayaguana triad. We find that D. mayaguana is diagnosable using DNA sequences, but the other two species form a non-diagnosable paraphyletic assemblage. A single ecological factor, oviposition substrate, is an important diagnostic character distinguishing D. straubae from D. parisiena, highlighting the importance of examining a diverse array of data (morphological, molecular, ecological, and behavioral) when defining species limits.     

Rapid detection of human fecal Eubacterium species and related genera by nested PCR method

PCR procedures based on 16S rDNA gene sequence specific for seven Eubacterium spp. and Eggerthella lenta that predominate in the human intestinal tract were developed, and used for direct detection of these species in seven human feces samples. Three species of Eggerthella lenta, Eubacterium rectale, and Eubacterium eligens were detected from seven fecal samples. Eubacterium biforme was detected from six samples. It was reported that E. rectale, E. eligens, and E. biforme were difficult to detect by traditional culture method, but the nested PCR method is available for the detection of these species. This result shows that the nested PCR method utilizing a universal primer pair, followed by amplification with species-specific primers, would allow rapid detection of Eubacterium species in human feces.     

The endemic Cladophorales (Ulvophyceae) of ancient Lake Baikal represent a monophyletic group of very closely related but morphologically diverse species

Lake Baikal, the oldest lake in the world, is home to spectacular biodiversity and extraordinary levels of endemism. While many of the animal species flocks from Lake Baikal are famous examples of evolutionary radiations, the lake also includes a wide diversity of endemic algae that are not well investigated with regards to molecular‐biological taxonomy and phylogeny. The endemic taxa of the green algal order Cladophorales show a range of divergent morphologies that led to their classification in four genera in two families. We sequenced partial large‐ and small‐subunit rDNA as well as the internal transcribed spacer region of 14 of the 16 described endemic taxa to clarify their phylogenetic relationships. One endemic morphospecies, Cladophora kusnetzowii, was shown to be conspecific with the widespread Aegagropila linnaei. All other endemic morphospecies formed a monophyletic group nested within the genus Rhizoclonium (Cladophoraceae), a very surprising result, in stark contrast to their morphological affinities. The Baikal clade represents a species flock of closely related taxa with very low genetic differentiation. Some of the morphospecies were congruent with lineages recovered in the phylogenies, but due to the low phylogenetic signal in the rDNA sequences the relationships within the Baikal clade were not all well resolved. The Baikal clade appears to represent a recent radiation, based on the low molecular divergence within the group, and it is hypothesized that the large morphological variation results from diversification in sympatry from a common ancestor in Lake Baikal.     

Identification of group I introns within the SSU rDNA gene in species of Ceratocystiopsis and related taxa

During a recent phylogenetic study, group I introns were noted that interrupt the nuclear small subunit ribosomal RNA (SSU rDNA) gene in species of Ceratocystiopsis. Group I introns were found to be inserted at the following rDNA positions: S943, S989, and S1199. The introns have been characterized and phylogenetic analysis of the host gene and the corresponding intron data suggest that for S943 vertical transfer and frequent loss appear to be the most parsimonious explanation for the distribution of nuclear SSU rDNA introns among species of Ceratocystiopsis. The SSU rDNA data do suggest that a recent proposal of segregating the genus Ophiostoma sensu lato into Ophiostoma sensu stricto, Grosmannia, and Ceratocystiopsis has some merit but may need further amendments, as the SSU rDNA suggests that Ophiostoma s. str. may now represent a paraphyletic grouping.     

Human fecal flora: variation in bacterial composition within individuals and a possible effect of emotional stress.

Data are presented on the distribution of 101 bacterial species and subspecies among 1,442 isolates from 25 fecal specimens from three men on: (i) their normal diet and normal living conditions, (ii) normal living conditions but eating the controlled metabolic diet designed for use in the Skylab simulation and missions, and (iii) the Skylab diet in simulated Skylab (isolation) conditions. These bacteria represent the most numerous kinds in the fecal flora. Analyses of the kinds of bacteria from each astronaut during the 5-month period showed more variation in the composition of the flora among the individual astronauts than among the eight or nine samples from each person. This observation indicates that the variations in fecal flora reported previously, but based on the study of only one specimen from each person, more certainly reflect real differences (and not daily variation) in the types of bacteria maintained by individual people. The proportions of the predominant fecal species in the astronauts were similar to those reported earlier from a Japanese-Hawaiian population and were generally insensitive to changes from the normal North American diet to the Skylab diet; only two of the most common species were affected by changes in diet. However, one of the predominant species (Bacteroides fragilis subsp. thetaiotaomicron) appeared to be affected during confinement of the men in the Skylab test chamber. Evidence is presented suggesting that an anger stress situation may have been responsible for the increase of this species simultaneously in all of the subjects studied. Phenotypic characteristics of some of the less common isolates are given. The statistical analyses used in interpretation of the results are discussed.     

Peptostreptococcus hydrogenalis sp. nov. from human fecal and vaginal flora

We describe Peptostreptococcus hydrogenalis sp. nov., which is found in human feces and vaginal discharge. This new species was established on the basis of the results of DNA-DNA hybridization among anaerobic cocci. The results of different biochemical reactions also are given. The type strain of this species is strain GIFU 7662 (= JCM 7635).     

Compartmentalized dynamics within a common multi-area mesoscale manifold represent a repertoire of human hand movements

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Evolution of mating within the Candida parapsilosis species group

Candida orthopsilosis and Candida metapsilosis are closely related to Candida parapsilosis, a major cause of infection in premature neonates. Mating has not been observed in these species. We show that ～190 isolates of C. parapsilosis contain only an MTLa idiomorph at the mating-type-like locus. Here, we describe the isolation and characterization of the MTL loci from C. orthopsilosis and C. metapsilosis. Among 16 C. orthopsilosis isolates, 9 were homozygous for MTLa, 5 were homozygous for MTLα, and 2 were MTLa/α heterozygotes. The C. orthopsilosis isolates belonged to two divergent groups, as characterized by restriction patterns at MTL, which probably represent subspecies. We sequenced both idiomorphs from each group and showed that they are 95% identical and that the regulatory genes are intact. In contrast, 18 isolates of C. metapsilosis contain only MTLα idiomorphs. Our results suggest that the role of MTL in determining cell type is being eroded in the C. parapsilosis species complex. The population structure of C. orthopsilosis indicates that mating may occur. However, expression of genes in the mating signal transduction pathway does not respond to exposure to alpha factor. C. parapsilosis is also nonresponsive, even when the GTPase-activating protein gene SST2 is deleted. In addition, splicing of introns in MTLa1 and MTLa2 is defective in C. orthopsilosis. Mating is not detected. The alpha factor peptide, which is the same sequence in C. parapsilosis, C. orthopsilosis, and C. metapsilosis, can induce a mating response in Candida albicans. It is therefore likely either that mating of C. orthopsilosis takes place under certain unidentified conditions or that the mating pathway has been adapted for other functions, such as cross-species communication.     

Comparative study of bacterial groups within the human cecal and fecal microbiota

The composition of the human cecal microbiota is poorly known because of sampling difficulties. Samples of cecal fluid from eight subjects were collected via an intestinal tube. Feces were also collected. Total anaerobes, facultative anaerobes, bifidobacteria, and Bacteroides were enumerated by culture methods, and the predominant phylogenetic groups were quantified by molecular hybridization using a set of six rRNA-targeted probes. The numbers of strict anaerobes, bifidobacteria, Bacteroides, and members of the Clostridium coccoides group and Clostridium leptum subgroup were lower in the cecum. Facultative anaerobes represented 25% of total bacteria in the cecum versus 1% in the feces.     

Degradation of human intestinal glycosphingolipids by extracellular glycosidases from mucin-degrading bacteria of the human fecal flora

Certain normal strains of human fecal bacteria are unique in producing extracellular glycosidases that degrade the oligosaccharide chains of gut mucin glycoproteins. We have studied the action of such glycosidases partially purified from the cell-free supernates of five of these strains on intestinal glycosphingolipids isolated from human meconium. The glycolipids were sialosyl-lactosylceramide, lactosylceramide, and fucolipids with A, B, H, Lea, or Leb blood group determinants. In addition to the strain-specific high blood group A-degrading activities (Ruminococcus torques strains VIII-239 and IX-70), B-degrading activity (Ruminococcus AB strain VI-268), and H-degrading activities (all strains) corresponding to alpha 1-3-N-acetylgalactosaminidase, alpha 1-3-galactosidase and alpha 1-2-fucosidase, respectively, all strains also degraded sialosyl-lactosylceramide and Lea and Leb antigenic glycolipids, indicating the presence of alpha 2-3-neuraminidases and alpha 1-4-fucosidases. Enzyme preparations from Bifidobacterium infantis strain VIII-240 and R. torques strain VIII-239 hydrolyzed the Lea active glycolipid directly to lactosylceramide, suggesting the presence of endo-beta 1-3-N-acetylglucosaminidase activities. Similar endo-beta-N-acetylglucosaminidase activities were identified in four of the five enzyme preparations. The enzymes produced by R. AB strain VI-268 lacked this activity as well as beta 1-3-galactosidase, and thus degradation stopped at lactotetraosylceramide. With enzyme preparations from the other strains lactosylceramide was the single major degradation product from complex glycosphingolipids with less than 30% further degradation to glucosylceramide within 48 h. We conclude that glycosidases from mucin-degrading strains of human enteric bacteria degrade oligosaccharide chains of lactoseries fucolipids and gangliosides of intestinal origin primarily to lactosylceramide. Since several genera of enteric bacteria bind preferentially to lactosylceramide in vitro, mucin-degrading strains may have an important ecological role in host-microbial associations in the human gut.     

Study of closely related species within the Physalaemus cuvieri group (Anura): contribution of microsatellite markers

Various species of the Physalaemus cuvieri group of frogs are difficult to distinguish morphologically, making molecular analysis an attractive alternative for indentifying members of this group, which is considered to be at risk because of loss of habitat. The genetic structure of natural populations of P. ephippifer and P. albonotatus species was investigated and analyzed, together with that of five previously studied populations of P. cuvieri. Nine microsatellite loci were used in the analyses. The overall G(ST) value (0.46) revealed high genetic variation among the populations, as expected for different species. Bayesian analysis implemented by the STRUCTURE software clustered the seven populations into seven groups (K = 7). All the P. albonotatus and P. ephippifer specimens were grouped into a single cluster, both species showing clear differentiation from P. cuvieri. The different grouping based on these microsatellites of some P. cuvieri individuals from Porto Nacional and from Passo Fundo suggests that they could be a new species, indicating a necessity for taxonomic reevaluation. Despite the intrinsic difficulties in analyzing closely related species, the nine microsatellite loci were found to be adequate for distinguishing these three species of the P. cuvieri group and their populations.     

Cecal and fecal bacterial flora of the Mongolian gerbil and the chinchilla

The Mongolian gerbil is being increasingly used as a laboratory animal and as a pet. Both chinchillas and gerbils are used as animal models for otitis media and other otic research. Previously, only incomplete information was available regarding the indigenous bacterial flora of the lower intestinal tracts of these coprophagic animals. Using the strict anaerobic methodology of the Virginia Polytechnic Institute Anaerobe Laboratory, we studied the predominant bacterial flora of the cecum and fecal pellets of the gerbil and the chinchilla and the bacterial flora of digesta pellets in the proximal colon. We found species of the following anaerobic genera in high dilutions of gerbil fecal pellets: Bifidobacterium, Clostridium, Propionibacterium, Lactobacillus, and Bacteroides. Only lactobacilli were found in high dilutions of digesta from the upper colon, although the cecum yielded Peptostreptococcus, Bifidobacterium, Clostridium, Lactobacillus, Propionibacterium, and Bacteroides species from high dilutions of cecal contents. The facultatively anaerobic and aerobic flora isolated consisted of species of Bacillus, Streptococcus, Staphylococcus, Acinetobacter, Alcaligenes, Escherichia, Pasteurella, and Pseudomonas plus several unidentifiable organisms. Species of Bifidobacterium, Bacteroides, Eubacterium, and anaerobic Lactobacillus were isolated from chinchillas.     

Genetic diversity within Cryptosporidium parvum and related Cryptosporidium species.

To assess the genetic diversity in Cryptosporidium parvum, we have sequenced the small subunit (SSU) rRNA gene of seven Cryptosporidium spp., various isolates of C. parvum from eight hosts, and a Cryptosporidium isolate from a desert monitor. Phylogenetic analysis of the SSU rRNA sequences confirmed the multispecies nature of the genus Cryptosporidium, with at least four distinct species (C. parvum, C. baileyi, C. muris, and C. serpentis). Other species previously defined by biologic characteristics, including C. wrairi, C. meleagridis, and C. felis, and the desert monitor isolate, clustered together or within C. parvum. Extensive genetic diversities were present among C. parvum isolates from humans, calves, pigs, dogs, mice, ferrets, marsupials, and a monkey. In general, specific genotypes were associated with specific host species. A PCR-restriction fragment length polymorphism technique previously developed by us could differentiate most Cryptosporidium spp. and C. parvum genotypes, but sequence analysis of the PCR product was needed to differentiate C. wrairi and C. meleagridis from some of the C. parvum genotypes. These results indicate a need for revision in the taxonomy and assessment of the zoonotic potential of some animal C. parvum isolates.     

Persistence, extinction and different species pools within the flora of lake islands in western Ireland

The vascular flora on twenty-nine lake islands in Lough Corrib, western Ireland was surveyed in 1992–93. Thirteen of these islands had been surveyed by the author in 1974 (Roden, 1979). Data on species–area curves and species turnover between 1974 and 1992 are presented. Species numbers on each island did not change greatly in the 18-year interval and extinctions were most common on smaller islands. It is known that six of the islands surveyed are less than 150 years old and their flora must have immigrated over open water during that period. It is shown that this group of species has a different log species/log area regression than the remaining flora, with a much shallower slope (low Z value). The proportion of less widespread species was greatest on islands nearest to the mainland. The implication of different slopes in different species groups and the restriction of turnover to rare species is discussed with reference to the island Theory of Biogeography.     

Leaf hydraulic conductance and mesophyll conductance are not closely related within a single species

Stomata represent one resistor in a series of resistances for carbon and water exchange between the leaf and the atmosphere; the remaining resistors occurring within the leaf, commonly represented as mesophyll conductance to CO₂, g_m, and leaf hydraulic conductance, k_Leaf. Recent studies have proposed that g_m and k_Leaf may be coordinated across species because of shared pathways. We assessed the correlation between g_m and k_Leaf within cotton, under growth CO₂ partial pressure and irradiance treatments and also with short‐term variation in irradiance and humidity. g_m was estimated using two isotopic techniques that allowed partitioning of total g_m (Δ¹³C‐g_m) into cell wall plus plasma membrane conductance (Δ¹⁸O‐g_m) and chloroplast membrane conductance (g_cm). A weak correlation was found between Δ¹³C‐g_m and k_Leaf only when measured under growth conditions. However, Δ¹⁸O‐g_m was related to k_Leaf under both short‐term environmental variation and growth conditions. Partitioning g_m showed that g_cm was not affected by short‐term changes in irradiance or correlated with k_Leaf, but was strongly reduced at high growth CO₂ partial pressure. Thus, simultaneous measurements of g_m, k_Leaf and g_cm suggest independent regulation of carbon and water transport across the chloroplast membrane with limited coordinated regulation across the cell wall and plasma membrane.     

Cecal and fecal bacterial flora of the Mongolian gerbil and the chinchilla.

The Mongolian gerbil is being increasingly used as a laboratory animal and as a pet. Both chinchillas and gerbils are used as animal models for otitis media and other otic research. Previously, only incomplete information was available regarding the indigenous bacterial flora of the lower intestinal tracts of these coprophagic animals. Using the strict anaerobic methodology of the Virginia Polytechnic Institute Anaerobe Laboratory, we studied the predominant bacterial flora of the cecum and fecal pellets of the gerbil and the chinchilla and the bacterial flora of digesta pellets in the proximal colon. We found species of the following anaerobic genera in high dilutions of gerbil fecal pellets: Bifidobacterium, Clostridium, Propionibacterium, Lactobacillus, and Bacteroides. Only lactobacilli were found in high dilutions of digesta from the upper colon, although the cecum yielded Peptostreptococcus, Bifidobacterium, Clostridium, Lactobacillus, Propionibacterium, and Bacteroides species from high dilutions of cecal contents. The facultatively anaerobic and aerobic flora isolated consisted of species of Bacillus, Streptococcus, Staphylococcus, Acinetobacter, Alcaligenes, Escherichia, Pasteurella, and Pseudomonas plus several unidentifiable organisms. Species of Bifidobacterium, Bacteroides, Eubacterium, and anaerobic Lactobacillus were isolated from chinchillas.