Molecular analysis of Continuous Plankton Recorder samples, an examination of echinoderm larvae in the North Sea

Analysis of the biological time series of plankton samples collected by the Continuous Plankton Recorder (CPR) in the North Atlantic and North Sea has shown a regime shift in the plankton in this region. Both the distributions of planktonic organisms and their timing of occurrence in the seasonal cycle have changed and these changes appear to ref lect global warming. In the North Sea the planktonic larvae of echinoderms have shown a recent dramatic increase in both relative and absolute abundance and their seasonal peak of occurrence has advanced by 47 days. The identity of the echinoderm larvae involved in this change has, however, remained equivocal. The small size of many organisms like echinoderm larvae combined with incomplete taxonomic keys hinders their visual identification and their fragility often means that useful morphological features are damaged during sampling by the CPR. Here, using new molecular methods applied to CPR samples, we show that planktonic larvae of the benthic Echinocardium cordatum dominate the North Sea plankton. We argue that since this species benefits from mild winters and warmer waters their numerical increase in the plankton is consistent with recent climatic changes that appear to be affecting the wider ecology of this region.

A biological consequence of reducing Arctic ice cover: arrival of the Pacific diatom Neodenticula seminae in the North Atlantic for the first time in 800,000 years

The Continuous Plankton Recorder survey has monitored plankton in the Northwest Atlantic at monthly intervals since 1962, with an interegnum between 1978 and 1990. In May 1999, large numbers of the Pacific diatom Neodenticula seminae were found in Continuous Plankton Recorder (CPR) samples in the Labrador Sea as the first record in the North Atlantic for more than 800 000 years. The event coincided with modifications in Arctic hydrography and circulation, increased flows of Pacific water into the Northwest Atlantic and in the previous year the exceptional occurrence of extensive ice-free water to the North of Canada. These observations indicate that N. seminae was carried in a pulse of Pacific water in 1998/early 1999 via the Canadian Arctic Archipelago and/or Fram Strait. The species occurred previously in the North Atlantic during the Pleistocene from similar to 1.2 to similar to 0.8 Ma as recorded in deep sea sediment cores. The reappearance of N. seminae in the North Atlantic is an indicator of the scale and speed of changes that are taking place in the Arctic and North Atlantic oceans as a consequence of regional climate warming. Because of the unusual nature of the event it appears that a threshold has been passed, marking a change in the circulation between the North Pacific and North Atlantic Oceans via the Arctic. Trans-Arctic migrations from the Pacific into the Atlantic are likely to occur increasingly over the next 100 years as Arctic ice continues to melt affecting Atlantic biodiversity and the biological pump with consequent feedbacks to the carbon cycle.

Isolation of microplastics in biota-rich seawater samples and marine organisms

Microplastic litter is a pervasive pollutant present in aquatic systems across the globe. A range of marine organisms have the capacity to ingest microplastics, resulting in adverse health effects. Developing methods to accurately quantify microplastics in productive marine waters, and those internalized by marine organisms, is of growing importance. Here we investigate the efficacy of using acid, alkaline and enzymatic digestion techniques in mineralizing biological material from marine surface trawls to reveal any microplastics present. Our optimized enzymatic protocol can digest >97% (by weight) of the material present in plankton-rich seawater samples without destroying any microplastic debris present. In applying the method to replicate marine samples from the western English Channel, we identified 0.27 microplastics m−3. The protocol was further used to extract microplastics ingested by marine zooplankton under laboratory conditions. Our findings illustrate that enzymatic digestion can aid the detection of microplastic debris within seawater samples and marine biota.

Could benzalkonium chloride be a suitable alternative to mercuric chloride for preservation of seawater samples?

Instrumental equipment unsuitable or unavailable for fieldwork as well as lack of ship space can necessitate the preservation of seawater samples prior to analysis in a shore-based laboratory. Mercuric chloride (HgCl2/ is routinely used for such preservation, but its handling and subsequent disposal incur environmental risks and significant expense. There is therefore a strong motivation to find less hazardous alternatives. Benzalkonium chloride (BAC) has been used previously as microbial inhibitor for freshwater samples. Here, we assess the use of BAC for marine samples prior to the measurement of oxygen-to-argon (O2 = Ar) ratios, as used for the determination of biological net community production. BAC at a concentration of 50 mg dm-3 inhibited microbial activity for at least 3 days in samples tested with chlorophyll a (Chl a) concentrations up to 1 mgm-3. BAC concentrations of 100 and 200 mg dm

Microbial analysis of soil and groundwater from a gasworks site and comparison to a sequenced biological reactive barrier remediation process (SEREBAR)

Aims: To investigate the distribution of a polymicrobial community of biodegradative bacteria in (i) soil and groundwater at a former manufactured gas plant (FMGP) site and (ii) in a novel SEquential REactive BARrier (SEREBAR) bioremediation process designed to bioremediate the contaminated groundwater. Methods and Results: Culture-dependent and culture-independent analyses using denaturing gradient gel electrophoresis (DGGE) and polymerase chain reaction (PCR) for the detection of 16S ribosomal RNA gene and naphthalene dioxygenase (NDO) genes of free-living (planktonic groundwater) and attached (soil biofilm) samples from across the site and from the SEREBAR process was applied. Naphthalene arising from groundwater was effectively degraded early in the process and the microbiological analysis indicated a dominant role for Pseudomonas and Comamonas in its degradation. The microbial communities appeared highly complex and diverse across both the sites and in the SEREBAR process. An increased population of naphthalene degraders was associated with naphthalene removal. Conclusion: The distribution of micro-organisms in general and naphthalene degraders across the site was highly heterogeneous. Comparisons made between areas contaminated with polycyclic aromatic hydrocarbons (PAH) and those not contaminated, revealed differences in the microbial community profile. The likelihood of noncultured bacteria being dominant in mediating naphthalene removal was evident. Significance and Impact of the Study: This work further emphasizes the importance of both traditional and molecular-based tools in determining the microbial ecology of contaminated sites and highlights the role of noncultured bacteria in the process.

Tracking biological invasions in space and time: elucidating the invasive history of the green alga Codium fragile using old DNA

With the advent of 'ancient DNA' studies on preserved material of extant and extinct species, museums and herbaria now represent an important although still underutilized resource in molecular ecology. The ability to obtain sequence data from archived specimens can reveal the recent history of cryptic species and introductions. We have analysed extant and herbarium samples of the highly invasive green alga Codium fragile, many over 100 years old, to identify cryptic accessions of the invasive strain known as C. fragile ssp. tomentosoides, which can be identified by a unique haplotype. Molecular characterization of specimens previously identified as native in various regions shows that the invasive tomentosoides strain has been colonizing new habitats across the world for longer than records indicate, in some cases nearly 100 years before it was noticed. It can now be found in the ranges of all the other native haplotypes detected, several of which correspond to recognized subspecies. Within regions in the southern hemisphere there was a greater diversity of haplotypes than in the northern hemisphere, probably as a result of dispersal by the Antarctic Circumpolar Current. The findings of this study highlight the importance of herbaria in preserving contemporaneous records of invasions as they occur, especially when invasive taxa are cryptic.

PIM-1 kinase expression in adipocytic neoplasms: diagnostic and biological implications

The differential diagnosis of soft tissue tumours poses a considerable challenge for pathologists, especially adipocytic tumours, as these may show considerable overlap in clinical presentation and morphological features with many other mesenchymal neoplasms. Hence, a specific and reliable marker that identifies adipocytic differentiation is much sought. We investigated the immunohistochemical expression of PIM-1 kinase in 35 samples of soft tissue tumours using tissue microarray technology and 49 full sections of adipocytic (n = 26) and non-adipocytic tumours (n = 23). Benign and malignant adipocytic tumours showed strong expression of PIM-1 while the non-adipocytic tumours were either negative or showed only weak staining for the protein. In myxoid liposarcomas, PIM-1 showed a distinct, unique vacuolar staining pattern, clearly outlining fine cytoplasmic lipid vacuoles. By contrast, non-adipocytic myxoid tumours (myxoma, chordoma and myxoid chondrosarcoma) did not show this vacuolar pattern of PIM-1 staining, although vacuolated cells were present on H&E. This differential expression was confirmed at a gene expression level in selected cases. Our results indicate that the expression of PIM-1 in adipose tissue may be a useful marker of adipocytic differentiation, in particular if the staining is both of high intensity and present in a unique, vacuolar pattern.

The Influence of Aspect on the Biological Colonization of Stone in Northern Ireland.

The rate and type of biological colonization of stone is influenced by a wide array of environmental factors in addition to substrate characteristics. A series of experiments was designed to compare the rate and type of biological colonization of stone at varying locations over a 21-month time period. Exposure
trials were set up at nine different sites across Northern Ireland that covered a wide variety of environmental conditions. To determine aspect-related differences in colonization, blocks of Peakmoor sandstone and Portland limestone were placed on the north- and south-facing sides of purpose-designed exposure racks. Colorimetry and visual analysis were carried out on collected samples at increasing time intervals. Results showed significantly different rates of darkening and greening over time between north-facing and south-facing blocks, for both sandstone and limestone. This difference is likely to be representative of the fact that in Northern Ireland’s wet climate and northern-latitude position, the north face of a building will receive less direct sunlight. Therefore north-facing blocks, once wet, will remain damp for much longer than blocks on other façades. This slow-drying phenomenon is much more hospitable for biological colonization and continued growth than the hostile environment of rapid wetting and drying cycles experienced on the south face.

Development of a solid-phase extraction coupling chemiluminescent enzyme immunoassay for determination of organophosphorus pesticides in environmental water samples

Solid-phase extraction (SPE) and direct competitive chemiluminescence enzyme immunoassay (dcCL-EIA) were combined for the detection of organophosphorus pesticides (OPs) in environmental water samples. dcCL-EIA based on horseradish peroxidase labeled with a broad-specificity monoclonal antibody against OPs was developed, and the effects of several physicochemical parameters on dcCL-EIA performance were studied. SPE was used for the pretreatment of water samples to remove interfering substances and to concentrate the OP analytes. The coupling of SPE and dcCL-EIA can detect seven OPs (parathion, coumaphos, phoxim, quinalphos, triazophos, dichlofenthion, and azinphos-ethyl) with the limit of quantitation below 0.1 ng/mL. The recoveries of OPs from spiked water samples ranged from 62.5% to 131.7% by SPE-dcCL-EIA and 69.5% to 112.3% by SPE-HPLC-MS/MS. The screening of OP residues in real-world environmental water samples by the developed SPE-dcCL-EIA and their confirmatory analysis using SPE-HPLC-MS/MS demonstrated that the assay is ideally suited as a monitoring method for OP residues prior to chromatographic analysis.

Development of a Broad-Specificity Monoclonal Antibody-Based Immunoaffinity Chromatography Cleanup for Organophosphorus Pesticide Determination in Environmental Samples

An immunoaffinity chromatographic (IAC) method for the selective extraction and concentration of 13 organophosphorus pesticides (OPs, including coumaphos, parathion, phoxim, quinalphos, dichlofenthion, triazophos, azinphos-ethyl, phosalone, isochlorthion, parathion-methyl, cyanophos, disulfoton, and phorate) prior to analysis by high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) was developed. The IAC column was prepared by covalently immobilizing a monoclonal antibody with broad specificity for OPs on CNBr-activated Sephrose 4B. The column capacity ranged from 884 to 2641 ng/mL of gel. The optimum elution solvent was 0.01 M phosphate-buffered saline containing 80% methanol. The breakthrough volume of the IAC column was found to be 400 mL. Recoveries of OPs from spiked environmental samples by IAC cleanup and HPLC-MS/MS analysis ranged from 60.2 to 107.1%, with a relative standard deviation below 11.1%. The limit of quantitation for 13 OPs ranged from 0.01 to 0.13 ng/mL (ng/g). The application of IAC cleanup coupled to HPLC-MS/MS in real environmental samples demonstrated the potential of this method for the determination of OP residues in environmental samples at trace levels.

Dual-colour HER2/chromosome 17 chromogenic in situ hybridisation assay enables accurate assessment of HER2 genomic status in gastric cancer and has potential utility in HER2 testing of biopsy samples

Determination of HER2 protein expression by immunohistochemistry (IHC) and genomic status by fluorescent in situ hybridisation (FISH) are important in identifying a subset of high HER2-expressing gastric cancers that might respond to trastuzumab. Although FISH is considered the standard for determination of HER2 genomic status, brightfield ISH is being increasingly recognised as a viable alternative. Also, the impact of HER2 protein expression/genomic heterogeneity on the accuracy of HER2 testing has not been well studied in the context of gastric biopsy samples.