Enumeration of methanogens with a focus on fluorescence in situ hybridization

Methanogens, the members of domain Archaea are potent contributors in global warming. Being confined to the strict anaerobic environment, their direct cultivation as pure culture is quite difficult. Therefore, a range of culture-independent methods have been developed to investigate their numbers, substrate uptake patterns, and identification in complex microbial communities. Unlike other approaches, fluorescence in situ hybridization (FISH) is not only used for faster quantification and accurate identification but also to reveal the physiological properties and spatiotemporal dynamics of methanogens in their natural environment. Aside from the methodological aspects and application of FISH, this review also focuses on culture-dependent and -independent techniques employed in enumerating methanogens along with associated problems. In addition, the combination of FISH with micro-autoradiography that could also be an important tool in investigating the activities of methanogens is also discussed.     

210Po concentration in Perna perna mussels: looking for radiation effects

Twenty ropes with 400 Perna perna mussels seeds (3 cm shell size) were set-up on floating structures at Cabo Frio Island, Arraial do Cabo, approximately 100 km northeast of Rio de Janeiro city. A rope was taken out on a monthly basis, and the shell sizes of 100 seeds were measured. The haemolymph of 10 male and 10 female individuals was taken, and the same individuals were separated for 210Po/210Pb determination. After one year of monthly sampling, no clear correlation was observed between the 210Po concentration, mussel's age and weight. A mean 210Po concentration of 155 Bq kg(-1) wet weight basis, was obtained, which is comparable with data reported in the literature. The radiation dose did not have any observable effect on the micronuclei frequency and DNA breaks in the mussels. This was probably due to the low dose rate, 0.02 mGy d(-1), in comparison with the suggested potential dose limit of 10 mGy d(-1).     

Development of the nasal olfactory organs in the larvae, settlement-stages and some adults of 14 species of Caribbean reef fishes (Labridae, Scaridae, Pomacentridae)

Larval fishes likely use a variety of settlement cues to locate and navigate toward the habitats they will inhabit as juveniles. Information about the morphology and state of development of the sensory organs of larval stages of fishes provides insight into their capabilities at the time of settlement. The peripheral olfactory organ of wild-caught late-stage larvae and early juveniles and some adults of 14 species of the Caribbean reef fishes wrasses (Labridae), parrot fishes (Scaridae) and damselfish (Pomacentridae) were examined using scanning electron microscopy and compared in terms of settlement specificity. Ages in days after hatching and days post-settlement were determined from the otoliths. Morphology of the nares and the olfactory epithelium are described for these species by stage. The separation of the anterior and posterior nares occurred before settlement in the labrids but in some specimens of scarids this separation was not complete by the time of settlement. Densities of ciliated and microvillous receptor cells and non-sensory ciliated epithelial cells were calculated. Densities of ciliated receptor cells ranged from 0.389 μm⁻² in a specimen of Thallasoma bifasciatum to 0.0057 μm⁻² in Bodianus rufus and of microvillous receptor cells from 0.038 μm⁻² in a Clepticus parrae juvenile to 0.266 μm⁻² in a juvenile Doratonotus megalepis. Densities of non-sensory cilia, also associated with high olfactory ability, were also high. The olfactory organ in wrasses is well developed prior to settlement and is comparable to that of adult fishes. The possible role of olfaction in larval schooling, reef cue detection and orientation toward habitat at settlement is discussed.

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Deleterious effects of a nonPST bioactive compound(s) from <Emphasis Type="Italic">Alexandrium tamarense</Emphasis> on bivalve hemocytes

The known negative effects of shellfish toxin-producing dinoflagellates on feeding, burrowing and survival of some bivalve mollusks has prompted questions concerning whether they might also impair the internal defense system of affected bivalves and make them more susceptible to disease agents. The primary components of the cellular defense system are hemocytes. Many toxic dinoflagellates are too large to be ingested whole by hemocytes and would most likely be exposed to intracellular toxins only after the algae are consumed, broken down, and the water-soluble toxins, released. Therefore, we conducted a series of experiments in which hemocytes from two suspension-feeding bivalves—the Manila clam, Ruditapes philippinarum, and the softshell clam, Mya arenaria—were exposed in vitro to filtered extracts of one highly toxic paralytic shellfish toxin (PST)-producing and one nonPST-producing strain of Alexandrium tamarense (isolates PR18b, 76 ± 6 STXeq cell⁻¹ and CCMP115, with undetectable PST, respectively). We measured adherence and phagocytosis, two hemocyte attributes known to be inhibited by bacterial pathogens and other stressors. We found no measurable effect of a cell-free extract from a highly concentrated suspension of the PST-producing strain on hemocytes of either bivalve species. Instead, extract from the nonPST-producing strain had a consistent negative effect on both clams, resulting in significantly lower adherence and phagocytosis compared to strain PR18b and filtered seawater controls. The bioactive compound produced by strain CCMP115, which has yet to be characterized, may be similar to the PST-independent allelopathic compounds described for Alexandrium spp., which act on other plankters. These compounds and those produced by other harmful algae are known to cause immobilization, cellular deformation and lysis of co-occurring target organisms. Thus, nonPST producing Alexandrium spp., which do not cause paralysis and burrowing incapacitation of clams, may still produce a compound(s) that has negative effects not only on hemocytes, but on other molluscan cell types and their functions, as well.     

Transformation of hydroxylated derivatives of 2,5-dichlorobiphenyl and 2,4,6-trichlorobiphenyl by Burkholderia xenovorans LB400

The polychlorinated biphenyl (PCB)-degrading bacterium, Burkholderia xenovorans LB400, was capable of transforming three hydroxylated derivatives of 2,5-dichlorobiphenyl (2,5-DCB) (2′-hydroxy- (2′-OH-), 3′-OH-, and 4′-OH-2,5-DCB) when biphenyl was used as the carbon source (i.e., biphenyl pathway-inducing condition), although only 2′-OH-2,5-DCB was transformed when the bacterium was growing on succinate (i.e., condition non-inductive of the biphenyl pathway). On the contrary, hydroyxlated derivatives of 2,4,6-trichlorobiphenyl (2,4,6-TCB) (2′-OH-, 3′-OH-, and 4′-OH-2,4,6-TCB) were not significantly transformed by B. xenovorans LB400, regardless of the carbon source used. Gene expression analyses showed a clear correlation between the transformation of OH-2,5-DCBs and expression of genes of the biphenyl pathway. The PCB metabolite, 2,5-dichlorobenzoic acid (2,5-DCBA), was produced following the transformation of OH-2,5-DCBs. 2,5-DCBA was not further transformed by B. xenovorans LB400. The present study is significant because it provides evidence that PCB-degrading bacteria are capable of transforming hydroxylated derivatives of PCBs, which are increasingly considered as a new class of environmental contaminants.     

Economic development,institutions, and biodiversity loss at the global scale

Current accelerating biodiversity loss is by many conservationists regarded as a result of economic development. Some economists agree on this viewpoint but argue it is valid at low income levels because of the need to secure a minimum living standard. On the other hand, economic development at higher income levels can instead mitigate biodiversity loss because of improved willingness and affordability to implement measures such as protected areas. This so-called environmental Kuznets relation is tested in this study by econometric analysis of cross-sectional data on a global scale. However, the results do not support this relation. Instead, we found a relation between economic and institutional development where economic development decreases biodiversity loss at a minimum level of institutional quality, i.e. minimum capacity to implement and secure property rights for the citizens. Our results thus question policies that neglect institutional quality for reversing biodiversity loss. Further, it was found that biodiversity loss increases with invasive species, high average temperature and precipitation, and spatial autocorrelation.     

Bioethanol-gasoline fuel blends: exhaust emissions and morphological characterization of particulate from a moped engine

This study was aimed at evaluating the effects of gasoline-ethanol blends on the exhaust emissions in a catalyst-equipped four-stroke moped engine. The ethanol was blended with unleaded gasoline in at percentages (10, 15, and 20% v/v). The regulated pollutants and the particulate matter emissions were evaluated over the European ECE R47 driving cycle on the chassis dynamometer bench. Particulate matter was characterized in terms of total mass collected on filters and total number ofparticles in the range 7 nm-10 microm measured by electrical low-pressure impactor (ELPI). In addition, particle-phase polycyclic aromatic hydrocarbons (PAHs) emissions were evaluated to assess the health impact of the emitted particulate. Finally, an accurate morphological analysis was performed on the particulate by high-resolution transmission electron microscope (TEM) equipped with a digital image-processing/data-acquisition system. In general, CO emission reductions of 60-70% were obtained with 15 and 20% v/v ethanol blends, while the ethanol use did not reduce hydrocarbon (HC) and NOx emissions. No evident effect of ethanol on the particulate mass emissions and associated PAHs emissions was observed. Twenty-one PAHs were quantified in the particulate phase with emissions ranging from 26 to 35 microg/km and benzo[a]pyrene equivalent (BaPeq) emission factors from 2.2 to 4.1 microg/km. Both particulate matter and associated PAHs with higher carcinogenic risk were mainly emitted in the submicrometer size range (<0.1 microm). On the basis of the TEM observations, no relevant effect of the ethanol use on the particulate morphology was evidenced, showing aggregates composed ofprimary particles with mean diameters in the range 17.5-32.5 nm.     

A rapid method for screening of the Stockholm Convention POPs in small amounts of human plasma using SPE and HRGC/HRMS

A rapid analytical screening method allowing simultaneous analysis of 23 persistent organic pollutants (POPs) in human plasma was developed. Sample preparation based on solid-phase extraction (SPE) with additional clean-up using small multilayer silica gel columns. SPE was performed using a custom made polystyrene-divinylbenzene sorbent for the extraction of chlorinated and brominated POPs. Special efforts to reduce sample volume and improve speed and efficiency of the analytical procedure were made. Determination of 16 polychlorinated biphenyls (PCBs), 5 organochlorine (OC) pesticides, octachlorinated dibenzo-p-dioxin (OCDD) and polybrominated diphenyl ether (BDE #47) in 0.5 mL human plasma was performed by using high resolution gas chromatography coupled to high resolution mass spectrometry (HRGC/HRMS). Recovery of POPs ranged between 46% and 110%, and reproducibility was below 25% relative standard deviation (RSD) for all target compounds, except for trans-nonachlor and OCDD, which were present only at low levels. Limits of detection (LOD) were for the PCBs between 0.8 and 117.7 pg mL⁻¹ plasma and for the OC pesticides between 5.9 and 89.1 pg mL⁻¹ plasma. The LOD for OCDD and BDE #47 were 1.4 pg mL⁻¹ plasma, and 9.2 pg mL⁻¹ plasma, respectively. The presented method was successfully applied to 1016 human plasma samples from an epidemiological study on cardiovascular disease.     

Walkerton's Contaminated Water Supply System: A Forensic Approach to Identifying the Source

In May 2000 an investigation of a contaminated water distribution system in the town of Walkerton, Ontario, was performed by the Bruce-Grey-Owen Sound Health Unit (BGOSHU), with the assistance of Health Canada and the Ontario Ministry of Health and Long-Term Care. The purpose of the investigation was to determine the scope, likely cause and contributing factors of an Escherichia coli 0157:H7 outbreak. This investigation-represented the first documented outbreak of E. coli 0157:H7 associated with a treated municipal water supply in Canada and the largest multi-bacterial waterborne outbreak in Canada. The investigation proceeded on an expedited basis to determine the extent of damages to the water supply system. Environmental forensic techniques were incorporated into hydrogeological studies, environmental transport assessment, genetic characterization, and fate and transport of contaminants. The probable sources of bacteria contamination included the use of cast-iron pipes, the thickness and concentration of bio-film sediment and mineral, the infiltration of contaminated surface runoff due to heavy rain or flooding, flow velocity related to the diameter of the pipe, and/or surface water.