Discriminant analysis of ribotype profiles of Escherichia coli for differentiating human and nonhuman sources of fecal pollution.

Estuarine waters receive fecal pollution from a variety of sources, including humans and wildlife. Escherichia coli is a ubiquitous bacterium in the intestines of warm-blooded animals and is used as an indicator of fecal pollution. However, its presence does not specifically differentiate sources of pollution. A total of 238 E. coli isolates from human sources (HS) and nonhuman sources (NHS) were collected from the Apalachicola National Estuarine Research Reserve, from associated sewage treatment plants, and directly from animals and tested for ribotype (RT) profile. HS and NHS isolates showed 41 and 61 RT profiles, respectively. At a similarity index of ca. 50%, HS and NHS isolates demonstrated four clusters, with the majority of HS and NHS isolates located in clusters C and D; isolates obtained directly from human and animal feces also could be grouped within these clusters. Discriminant analysis (DA) of RT profiles showed that 97% of the NHS isolates and 100% of the animal fecal isolates were correctly classified. The average rate of correct classification for HS and NHS isolates was 82%. We conclude that DA of RT profiles may be a useful method for identifying HS and NHS fecal pollution and may potentially facilitate management practices.     

Occurrence of Diaphanosoma excisum (Sars) on a sandy beach at Balramgari (Orissa), India

The occurrence of Diaphanosoma excisum (Sars), a freshwater cladoceran, in benthic samples of an intertidal sandy beach is reported. Population density was seasonal. A relatively high density was recorded from June to September (south-west monsoon season) with a maximum (46 ind 10 cm^–2) in September at a depth of 10–15 cm of sediment. A sudden decline occurred during north-east monsoon (October to January), and in the fair season (February to May), the cladocerans disappeared. Mean density varied (P<0.001) with sediment depth and season and showed a contagious dispersion. Abundance was negatively correlated with salinity (r = –0.76) but positively with POC (r =0.79) and mean grain size of the sediment (r = 0.93). The density of D. excisum was highest in fine sand.     

Geographical variation in ribotype profiles of Escherichia coli isolates from humans,swine, poultry,beef, and dairy cattle in Florida

Waters impacted by fecal pollution can exact high risks to human health and can result in financial losses due to closures of water systems used for recreation and for harvesting seafood. Identifying the sources of fecal pollution in water is paramount in assessing the potential human health risks involved as well as in assessing necessary remedial action. Recently, various researchers have used the ribotyping method to identify sources of bacterial indicators (Escherichia coli and enterococci) in environmental waters. While these studies have identified genotypic differences between human- and animal-derived indicators that are capable of differentiating organisms isolated from humans and various animal hosts, most have focused on organisms collected from a confined geographic area and have not addressed the question of whether these ribotype profiles are watershed specific or if they can be applied universally to organisms from other geographic locations. In this study, E. coli isolates were obtained from humans, beef cattle, dairy cattle, swine, and poultry from locations in northern, central, and southern Florida and were subjected to ribotyping analysis. The intent was to determine (i) if ribotype profiles are capable of discriminating the source of E. coli at the host species level and (ii) if the resulting fingerprints are uniform over an extended geographic area or if they can be applied only to a specific watershed. Our research indicated that, using a single restriction enzyme (HindIII), the ribotyping procedure is not capable of differentiating E. coli isolates from the different animal species sampled in this study. Results indicate, however, that this procedure can still be used effectively to differentiate E. coli as being either human or animal derived when applied to organisms isolated from a large geographic region.     

Extraction, structural characterisation and evaluation of hydroxycinnamate esters of orchard grass (Dactylis glomerata) as substrates for polyphenol oxidase

Polyphenol oxidase (PPO) activity has been reported in orchard grass (Dactylis glomerata); however, to date, no endogenous substrates have been identified. In the present study, we report the isolation and structural elucidation of PPO substrates in this species. The free phenol fraction was extracted, separated by reverse-phase chromatography and six potential substrates, including two hydroxycinnamate esters, were identified by UV spectrometry, electrospray ionisation-tandem mass spectrometry (LC-ESI-MSⁿ) and 1D and 2D NMR analyses (¹H NMR, ¹³C NMR, DEPT, COSY, HMQC and HMBC). Furthermore, three caffeoylquinic acids (3-CQA, 4-CQA and 5-CQA) were identified by comparison of their spectral data (ESI-MS) with those of known compounds and literature data. Five of these compounds were demonstrated to be substrates for orchard grass PPO.     

Pycnogenol prevents potassium dichromate (K2Cr2O7)-induced oxidative damage and nephrotoxicity in rats

Environmental and occupational exposure to chromium compounds, especially hexavalent chromium [Cr(VI)], is widely recognized as a potential nephrotoxic in humans and animals. Its toxicity is associated with overproduction of free radicals, which induces oxidative damage. Recent evidence indicates that Pycnogenol^® (PYC), French maritime pine bark extract, exhibits antioxidant potential and protects against various oxidative stressors. The aim of the present study was to examine the modulating impacts of PYC on potassium dichromate (K₂Cr₂O₇)-induced oxidative damage and nephrotoxicity in rats. Male Wistar rats were divided into four groups. The first group was control, the second group was control plus pre-treated with PYC (10 mg/kg, body weight; in saline; intraperitoneally; once daily for 3 weeks) as drug control and the third group was saline pre-treated plus treated with a single injection of K₂Cr₂O₇ (15 mg/kg, body weight; in saline; intraperitoneally) as toxicant group. The fourth group was PYC pre-treated plus K₂Cr₂O₇ injected. Forty-eight hours after K₂Cr₂O₇-treatment, blood was drawn for estimation of renal injury markers in serum. Rats were then sacrificed, and their kidneys were dissected for biochemical and histopathological assays. K₂Cr₂O₇-treated rats showed significant increases in markers of renal injury in serum, including blood urea nitrogen (BUN), serum creatinine (Scr), and alkaline phosphatase (ALP), which were significantly (P < 0.05) decreased by PYC pre-treatment. Moreover, prophylactic pre-treatment of rats with PYC significantly (P < 0.05) ameliorated increased thiobarbituric reactive substances (TBARS), malonaldehyde (MDA) and protein carbonyl (PC), and decreased levels of glutathione (GSH) and catalase activity in the kidney homogenate of K₂Cr₂O₇-treated rats. These results were also supported and confirmed with histopathological findings. The study suggests that PYC is effective in preventing K₂Cr₂O₇-induced oxidative mediated nephrotoxicity, but more studies are needed to confirm the effects of PYC as a nephroprotective agent.     

Phylum Verrucomicrobia representatives share a compartmentalized cell plan with members of bacterial phylum Planctomycetes

Background  

The phylum Verrucomicrobia is a divergent phylum within domain Bacteria including members of the microbial communities of soil and fresh and marine waters; recently extremely acidophilic members from hot springs have been found to oxidize methane. At least one genus, Prosthecobacter, includes species with genes homologous to those encoding eukaryotic tubulins. A significant superphylum relationship of Verrucomicrobia with members of phylum Planctomycetes possessing a unique compartmentalized cell plan, and members of the phylum Chlamydiae including human pathogens with a complex intracellular life cycle, has been proposed. Based on the postulated superphylum relationship, we hypothesized that members of the two separate phyla Planctomycetes and Verrucomicrobia might share a similar ultrastructure plan differing from classical prokaryote organization.     

Isolation, identification and quantitation of hydroxycinnamic acid conjugates, potential platform chemicals, in the leaves and stems of Miscanthus × giganteus using LC-ESI-MSn

Miscanthus × giganteus is a source of platform chemicals and bioethanol through fermentation. Cinnamates in leaves and stems were analysed by LC–ESI-MSⁿ. Free phenols were extracted and separated chromatographically. More than 20 hydroxycinnamates were identified by UV and LC–ESI-MSⁿ. Comparative LC–MS studies on the leaf extract showed isomers of O-caffeoylquinic acid (3-CQA, 4-CQA and 5-CQA), O-feruloylquinic acid (3-FQA, 4-FQA and 5-FQA) and para-coumaroylquinic acid (3-pCoQA and 5-pCoQA). Excepting 3-pCoQA, all were also detected in stem. 5-CQA dominated in leaf; a mandelonitrile–caffeoylquinic acid dominated in stem. Three minor leaf components were distinguished by fragmentation patterns in a targetted MS² experiment as dicaffeoylquinic acid isomers. Others (M_r 516) were tentatively identified as hexosylcaffeoyl-quinates. Three positional isomers of O-caffeoylshikimic acid were minor components. p-Hydroxybenzaldehyde was also a major component in stem. This is the first report of the hydroxycinnamic acid profile of leaves and stems of M. × giganteus.