Bacterioplankton are recognized as important agents of biogeochemical change in marine ecosystems, yet relatively little is known about the species that make up these communities. Uncertainties about the genetic structure and diversity of natural bacterioplankton populations stem from the traditional difficulties associated with microbial cultivation techniques. Discrepancies between direct counts and plate counts are typically several orders of magnitude, raising doubts as to whether cultivated marine bacteria are actually representative of dominant planktonic species. We have phylogenetically analysed clone libraries of eubacterial 16S ribosomal RNA genes amplified from natural populations of Sargasso Sea picoplankton by the polymerase chain reaction. The analysis indicates the presence of a novel microbial group, the SAR11 cluster, which appears to be a significant component of this oligotrophic bacterioplankton community. A second cluster of lineages related to the oxygenic phototrophs--cyanobacteria, prochlorophytes and chloroplasts--was also observed. However, none of the genes matched the small subunit rRNA sequences of cultivated marine cyanobacteria from similar habitats. The diversity of 16S rRNA genes observed within the clusters suggests that these bacterioplankton may be consortia of independent lineages sharing surprisingly distant common ancestors.

Our purpose was to develop a rapid, inexpensive method of diagnosing the source of fecal pollution in water. In previous research, we identified Bacteroides-Prevotella ribosomal DNA (rDNA) PCR markers based on analysis. These markers length heterogeneity PCR and terminal restriction fragment length polymorphism distinguish cow from human feces. Here, we recovered 16S rDNA clones from natural waters that were close phylogenetic relatives of the markers. From the sequence data, we designed specific PCR primers that discriminate human and ruminant sources of fecal contamination.The inability to identify the source of fecal contamination is partly to blame for the persistent problem of fecal pollution in coastal and inland waters. Although methods exist to quantify fecal pollution, none quickly and accurately identifies the animal source. Antibiotic resistance patterns of fecal streptococci (8,16,17) and Escherichia coli ribosomal DNA (rDNA) tracking (14; D. Akre and J. Wilcox, Northwest Algal Symp. Pacific Estuarine Res. Soc. Joint Meet., 1998) have recently emerged as potentially useful, but labor-intensive, solutions to the problem. Their reliability, however, may be considerably less than 100% (16,17).Unlike these methods, which require culturing indicator organisms, detection of host-specific molecular markers does not require culturing and holds promise as a precise, rapid method for identifying sources of fecal contamination. The BacteroidesPrevotella group is one of several noncoliform bacterial groups that has been proposed as an alternative fecal pollution indicator (1, 5, 10), partly because of its abundance in feces. The use of molecular methods makes it more feasible to use anaerobic bacteria that are potentially difficult to grow, such as members of the Bacteroides-Prevotella group, as indicators.We recently identified host-specific Bacteroides-Prevotella 16S rDNA markers for humans and cows by screening fecal DNAs by length heterogeneity PCR (LH-PCR) (15) or terminal restriction fragment length polymorphism (T-RFLP) (11) analysis (2). Cloning and sequencing experiments revealed that each marker comprised multiple sequences forming host-specific gene clusters. Here, we have identified additional clones, recovered from water samples, that cluster with the fecal clones. Using the sequences from fecal and water clones, we developed cluster-specific primers that can discriminate between human and ruminant feces.Clones recovered from water samples. To identify fecal Bacteroides-Prevotella rDNA markers in water, we collected six 1-liter water samples from areas in Tillamook Bay, Oreg., that are frequently contaminated with fecal pollution. We processed the samples as previously described (2). DNAs from each water sample were amplified with Bacteroides-Prevotellaspecific primers (Bac32F and Bac708R) as described previously (2). Equal portions of PCR products from all water samples were pooled and cloned into pGEM T-Easy vectors according to the manufacturer's directions (Promega, Madison, Wis.).To locate mark...

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We describe a new PCR-based method for distinguishing human and cow fecal contamination in coastal waters without culturing indicator organisms, and we show that the method can be used to track bacterial marker sequences in complex environments. We identified two human-specific genetic markers and five cow-specific genetic markers in fecal samples by amplifying 16S ribosomal DNA (rDNA) fragments from members of the genus Bifidobacterium and the Bacteroides-Prevotella group and performing length heterogeneity PCR and terminal restriction fragment length polymorphism analyses. Host-specific patterns suggested that there are species composition differences in the Bifidobacterium and Bacteroides-Prevotella populations of human and cow feces. The patterns were highly reproducible among different hosts belonging to the same species. Additionally, all host-specific genetic markers were detected in water samples collected from areas frequently contaminated with fecal pollution. Ease of detection and longer survival in water made BacteroidesPrevotella indicators better than Bifidobacterium indicators. Fecal 16S rDNA sequences corresponding to our Bacteroides-Prevotella markers comprised closely related gene clusters, none of which exactly matched previously published Bacteroides or Prevotella sequences. Our method detected host-specific markers in water at pollutant concentrations of 2.8 ؋ 10؊5 to 2.8 ؋ 10 ؊7 g (dry weight) of feces/liter and 6.8 ؋ 10 ؊7 g (dry weight) of sewage/liter. Although our aim was to identify nonpoint sources of fecal contamination, the method described here should be widely applicable for monitoring spatial and temporal fluctuations in specific bacterial groups in natural environments.

Brain development in autism follows an abnormal pattern, with accelerated growth in early life that results in brain enlargement in childhood. Brain volume in adolescents and adults with autism is, however, normal, and appears to be due to a slight decrease in brain volume for these individuals at the same time that normal children are experiencing a slight increase.

Striatal atrophy begins many years prior to diagnosable HD, and assessment of atrophy on MRI may be very useful in both predicting HD onset and in tracking progression in future therapeutic trials in preclinical subjects.

A rapid sequencing method for ribosomal RNA was applied to the resolution of evolutionary relationships among Metazoa. Representatives of 22 classes in 10 animal phyla were used to infer phylogenetic relationships, based on evolutionary distances determined from pairwise comparisons of the 18S ribosomal RNA sequences. The classical Eumetazoa are divided into two groups. Cnidarians arose from a protist ancestry different from the second group, the Bilateria. Within the Bilateria, an early split gave rise to Platyhelminthes (flatworms) and the coelomate lineage. Coelomates are thus monophyletic, and they radiated rapidly into four groups:chordates, echinoderms, arthropods, and eucoelomate protostomes.

The purpose of this study was to examine host distribution patterns among fecal bacteria in the order Bacteroidales, with the goal of using endemic sequences as markers for fecal source identification in aquatic environments. We analyzed Bacteroidales 16S rRNA gene sequences from the feces of eight hosts: human, bovine, pig, horse, dog, cat, gull, and elk. Recovered sequences did not match database sequences, indicating high levels of uncultivated diversity. The analysis revealed both endemic and cosmopolitan distributions among the eight hosts. Ruminant, pig, and horse sequences tended to form host-or host group-specific clusters in a phylogenetic tree, while human, dog, cat, and gull sequences clustered together almost exclusively. Many of the human, dog, cat, and gull sequences fell within a large branch containing cultivated species from the genus Bacteroides. Most of the cultivated Bacteroides species had very close matches with multiple hosts and thus may not be useful targets for fecal source identification. A large branch containing cultivated members of the genus Prevotella included cloned sequences that were not closely related to cultivated Prevotella species. Most ruminant sequences formed clusters separate from the branches containing Bacteroides and Prevotella species. Host-specific sequences were identified for pigs and horses and were used to design PCR primers to identify pig and horse sources of fecal pollution in water. The primers successfully amplified fecal DNAs from their target hosts and did not amplify fecal DNAs from other species. Fecal bacteria endemic to the host species may result from evolution in different types of digestive systems.