Water quality and microbial diversity in cisterns from semiarid areas in Brazil

Harvesting rainwater is a common practice worldwide, particularly in areas with no access to a public water supply or insufficient groundwater reserves. More than two million people living in semiarid regions of Brazil consume rainwater stored in cisterns, and little information is available regarding the water quality. Despite the initial good quality of the rainwater, its harvest and storage can introduce contaminants that must be eliminated before consumption. To evaluate the influence of handling, cistern age and precipitation on the quality of harvested rainwater, we monitored seven cisterns in the semiarid Brazilian Northeast over 4 years. Microbial and physicochemical parameters were monitored once a month, and denaturant gradient gel electrophoresis (DGGE) was performed at the end of the monitoring period. Coliform bacteria were detected in 100% of samples, while Escherichia coli were observed in 73.8%. The alkalinity and conductivity were the highest for the recently built cisterns due to the dissolution of construction materials. The DGGE of the 16S r DNA did not reveal the presence of E. coli. Instead, DGGE bands sequencing indicated that species primarily affiliated with Alphaproteobacteria were present in all cisterns, indicating the presence of microbial ecosystems capable of purifying and stabilizing the stored rainwater.

Liquid-phase denaturant gradient gel electrophoresis profiles of rumen bacteria from Brahman cross steers selected into two groups on the basis of post-weaning liveweight gain on low crude protein pasture

Two experiments were conducted to evaluate the relationship between rumen liquid-associated bacterial community structures and post-weaning liveweight gain (LWG) of Brahman crossbred steers. Bacterial diversity was assessed using denaturant gradient gel electrophoresis (DGGE). In Experiment 1, 16 steers were selected from a group of 100 steers by pairing steers with the same weaning weight, but different LWG 90 days after weaning (n = 8 highest growth, 0.21 ± 0.01 kg/day; and n = 8 lowest growth –0.07 ± 0.01 kg/day). Thereafter, steers were allocated to a 28-day pen study and fed Mitchell grass (Astrebla spp.) hay to examine DM intake and digestibility, rumen parameters and rumen microbial community in these two groups. Rumen fluid samples were taken by stomach tube 3 h after feeding on the last day of the pen phase. In Experiment 2, 12 pairs of weaned steers were selected from a group of 203 steers on the same basis as Experiment 1. The post-weaning LWG were 0.20 ± 0.03 and 0.02 ± 0.03 kg/day for the 12 highest and 12 lowest growth animals selected, respectively. Steers then grazed dry season Sabi grass (Urochloa mosambicensis) dominant pasture for 21 days, before rumen sampling 3 h after morning grazing by stomach tubing on the last day. In Experiment 1, there were no significant differences between the two groups in DM intake, digestibility, ruminal pH, total volatile fatty acid (VFA) concentration or the VFA proportion. In Experiments 1 and 2, rumen ammonia-N concentration was similar between LWG groups and there was no evidence of a relationship between liquid-phase DGGE profiles of rumen bacteria and high or low post-weaning LWG using multivariate analyses. Furthermore, the number of detected DGGE bands, the Shannon–Wiener and evenness indexes were not different between LWG groups. This DGGE analysis of the most abundant groups of rumen fluid-associated bacteria suggests that microbial populations were not related with the differences observed in post-weaning LWG within a group of weaners fed low crude protein diets.

Bifidobacterial Diversity and the Development of New Microbial Source Tracking Indicators

ABSTRACTMany studies suggest a close relationship between species ofBifidobacteriumand their hosts. Thus, species such asB. adolescentisandB. thermacidophilumsubsp.porcinumhave been proposed as potential indicators of human and porcine fecal pollution. The diversity of bifidobacteria in wastewaters (human and animal) and slurries is analyzed using nested PCR followed by denaturant gradient gel electrophoresis (DGGE). The sewage samples showed similar DGGE patterns. The predominant bands were recognized asB. adolescentis,B. longum, and two unidentified species related toB. adolescentis. A single band detected in poultry samples was identified asB. saeculare. Bifidobacterial diversity was higher within porcine and bovine samples. The main bands in porcine samples were identified asB. minimum, an unknown species, andB. thermophilum/B. thermacidophilumsubsp.porcinum. The latter species was also identified among the main bands in bovine samples together withB. pseudolongumandB. ruminantium. We then attempted to isolate the host-specific strains. DGGE bands were examined to develop specific probes to screen environmental samples by colony hybridization and further isolation of strains from positively hybridized colonies. Bifidobacterial strains that are host associated by DGGE bands to human and pig were successfully isolated from the environment:B. adolescentisfrom human sewage samples and the unidentified species related to pig from slurries and slaughterhouse wastewater. Neither the poultry-associatedB. saecularenor the ruminant-associatedB. pseudolongumcould be isolated with the current methodology, suggesting either a low prevalence in the samples or failure of the culture to grow in the media used.

Segregation of partly melted molecules: isolation of CpG islands by polyacrylamide gel electrophoresis

The technique of segregation of partly melted molecules (SPM) is a convenient and efficient method to isolate DNA fragments associated with CpG islands. The approach is conceptually simple and uses denaturant gradient gel electrophoresis to separate DNA molecules digested with restriction endonucleases. The SPM methodology has successfully been applied to the identification of genes from anonymous, unsequenced DNA fragments and CpG islands methylated in human cancer. In this article the theoretical background and practical application of the SPM method is reviewed.

Differences between Betaproteobacterial Ammonia-Oxidizing Communities in Marine Sediments and Those in Overlying Water

ABSTRACT To assess links between betaproteobacterial ammonia-oxidizing bacteria (AOB) in marine sediment and in overlying water, communities in Loch Duich, Scotland, were characterized by analysis of clone libraries and denaturant gradient gel electrophoresis of 16S rRNA gene fragments. Nitrosospira cluster 1-like sequences were isolated from both environments, but different sequence types dominated water and sediment samples. Detailed phylogenetic analysis of marine Nitrosospira cluster 1-like sequences in Loch Duich and surrounding regions suggests the existence of at least two different phylogenetic subgroups, potentially indicative of new lineages within the betaproteobacterial AOB, representing different marine ecotypes.