Bodil Hernroth

Environmental Factors Influencing Human Viral Pathogens and Their Potential Indicator Organisms in the Blue Mussel, Mytilus edulis: the First Scandinavian Report

ABSTRACT This study was carried out in order to investigate human enteric virus contaminants in mussels from three sites on the west coast of Sweden, representing a gradient of anthropogenic influence. Mussels were sampled monthly during the period from February 2000 to July 2001 and analyzed for adeno-, entero-, Norwalk-like, and hepatitis A viruses as well as the potential viral indicator organisms somatic coliphages, F-specific RNA bacteriophages, bacteriophages infecting Bacteroides fragilis, and Escherichia coli. The influence of environmental factors such as water temperature, salinity, and land runoff on the occurrence of these microbes was also included in this study. Enteric viruses were found in 50 to 60% of the mussel samples, and there were no pronounced differences between the samples from the three sites. E. coli counts exceeded the limit for category A for shellfish sanitary safety in 40% of the samples from the sites situated in fjords. However, at the site in the outer archipelago, this limit was exceeded only once, in March 2001, when extremely high levels of atypical indole-negative strains of E. coli were registered at all three sites. The environmental factors influenced the occurrence of viruses and phages differently, and therefore, it was hard to find a coexistence between them. This study shows that, for risk assessment, separate modeling should be done for every specific area, with special emphasis on environmental factors such as temperature and land runoff. The present standard for human fecal contamination, E. coli, seems to be an acceptable indicator of only local sanitary contamination; it is not a reliable indicator of viral contaminants in mussels. To protect consumers and get verification of “clean” mussels, it seems necessary to analyze for viruses as well. The use of a molecular index of the human contamination of Swedish shellfish underscores the need for reference laboratories with high-technology facilities.

Improving marine water quality by mussel farming : a profitable solution for Swedish society

Abstract Eutrophication of coastal waters is a serious environmental problem with high costs for society globally. In eastern Skagerrak, reductions in eutrophication are planned through reduction of nitrogen inputs, but it is unclear how this can be achieved. One possible method is the cultivation of filter-feeding organisms, such as blue mussels, which remove nitrogen while generating seafood, fodder and agricultural fertilizer, thus recycling nutrients from sea to land. The expected effect of mussel farming on nitrogen cycling was modeled for the Gullmar Fjord on the Swedish west coast and it is shown that the net transport of nitrogen (sum of dissolved and particulate) at the fjord mouth was reduced by 20%. Existing commercial mussel farms already perform this service for free, but the benefits to society could be far greater. We suggest that rather than paying mussel farmers for their work that nutrient trading systems are introduced to improve coastal waters. In this context an alternative to nitrogen reduction in the sewage treatment plant in Lysekil community through mussel farming is presented. Accumulation of bio-toxins has been identified as the largest impediment to further expansion of commercial mussel farming in Sweden, but the problem seems to be manageable through new techniques and management strategies. On the basis of existing and potential regulations and payments, possible win-win solutions are suggested.

Distribution of Human Virus Contamination in Shellfish from Different Growing Areas in Greece, Spain, Sweden, and the United Kingdom

ABSTRACT Viral pollution in shellfish has been analyzed simultaneously across a wide range of geographical regions, with emphasis on the concomitant variations in physicochemical characteristics and social features. The methods for sample treatment and for the detection of human enteric viruses were optimized by the participating laboratories. The second part of this study involves the selection of a protocol for virus detection, which was validated by analyzing the distribution and concentration of human viral pathogens under diverse conditions during an 18-month period in four European countries. Shellfish-growing areas from diverse countries in the north and south of Europe were defined and studied, and the microbiological quality of the shellfish was analyzed. Human adenovirus, Norwalk-like virus, and enterovirus were identified as contaminants of shellfish in all the participating countries. Hepatitis A virus was also isolated in all areas except Sweden. The seasonal distribution of viral contamination was also described. Norwalk-like virus appeared to be the only group of viruses that demonstrated seasonal variation, with lower concentrations occurring during warm months. The depuration treatments currently applied were shown to be adequate for reducing Escherichia coli levels but ineffective for the elimination of viral particles. The human adenoviruses detected by PCR correlate with the presence of other human viruses and could be useful as a molecular index of viral contamination in shellfish.

Evaluation of Potential Indicators of Viral Contamination in Shellfish and Their Applicability to Diverse Geographical Areas

ABSTRACT The distribution of the concentration of potential indicators of fecal viral pollution in shellfish was analyzed under diverse conditions over 18 months in diverse geographical areas. These microorganisms have been evaluated in relation to contamination by human viral pathogens detected in parallel in the analyzed shellfish samples. Thus, significant shellfish-growing areas from diverse countries in the north and south of Europe (Greece, Spain, Sweden, and the United Kingdom) were defined and studied by analyzing different physicochemical parameters in the water and the levels of Escherichia coli, F-specific RNA bacteriophages, and phages infecting Bacteroides fragilis strain RYC2056 in the shellfish produced, before and after depuration treatments. A total of 475 shellfish samples were studied, and the results were statistically analyzed. According to statistical analysis, the presence of human viruses seems to be related to the presence of all potential indicators in the heavily contaminated areas, where E. coli would probably be suitable as a fecal indicator. The F-RNA phages, which are present in higher numbers in Northern Europe, seem to be significantly related to the presence of viral contamination in shellfish, with a very weak predictive value for hepatitis A virus, human adenovirus, and enterovirus and a stronger one for Norwalk-like virus. However, it is important to note that shellfish produced in A or clean B areas can sporadically contain human viruses even in the absence of E. coli or F-RNA phages. The data presented here will be useful in defining microbiological parameters for improving the sanitary control of shellfish consumed raw or barely cooked.

North Atlantic Oscillation primary productivity and toxic phytoplankton in the Gullmar Fjord, Sweden (1985–1996)

The evaluation of the time–series data set of primary production, chlorophyll a (1985–1996) and toxic phytoplankton species abundance measurements (1986–1996) in the Gullmar Fjord as part of the Swedish Monitoring Program, revealed the importance of considering climatic and environmental forces among the factors that may be responsible for the observed fluctuations. The results suggested that the occurrence of toxic phytoplankton blooms in the Skagerrak may be related to changes in the phase of the North Atlantic Oscillation (NAO). The Dinophysis species were related to the NAO, temperature and salinity and the fluctuations in the phytoplankton biomass expressed as chlorophyll a were also shown to be related to the NAO. The results pointed to the need to understand further the role played by climatic and environmental forces in relation to the formation and duration of harmful algal blooms and the need for continuous quality monitoring programmes and novel statistical methods for detecting trends and cycles in ecological time–series related to environmental forces.

Immune suppression of the echinoderm Asterias rubens (L.) following long-term ocean acidification

We compared effects of exposure to predict near-future (2100) ocean acidification (OA; pH 7.7) and normal seawater (Control; pH 8.1) on immune and stress responses in the adult sea star Asterias rubens. Analyses were made after one week and after six months of continuous exposure. Following one week exposure to acidified water, the pH of coelomic fluid was significantly reduced. Levels of the chaperon Hsp70 were elevated while key cellular players in immunity, coelomocytes, were reduced by approximately 50%. Following long-term exposure (six months) levels of Hsp70 returned to control values, whereas immunity was further impaired, evidenced by the reduced phagocytic capacity of coelomocytes and inhibited activation of p38 MAP-kinase. Such impacts of reduced seawater pH may have serious consequences for resistance to pathogens in a future acidified ocean.

Tracing of Norovirus Outbreak Strains in Mussels Collected near Sewage Effluents

ABSTRACT Noroviruses from mussels collected near sewage effluents were compared with local patient outbreak strains. Sequence analyses of RNA polymerase-capsid-poly(A)-3′ (3.1-kilobase) regions confirmed the 99.9% similarity between genotype I.1 strains from mussels and patient strains from recreational-bathing outbreaks, indicating the potential usefulness of sentinel norovirus mussel studies in tracing human norovirus contamination of coastal waters.

Induced cell proliferation in putative haematopoietic tissues of the sea star, Asterias rubens (L.)

SUMMARY The coelomic fluid of the echinoderm Asterias rubens possesses large populations of circulating coelomocytes. This study aimed to expand the knowledge about the haematopoietic sources of these cells. Injection of the immune-stimulating molecules lipopolysaccharide (LPS) and concanavalin A (ConA) resulted in an increase in coelomocytes. To investigate if these molecules induce cell proliferation in putative haematopoietic tissues (HPTs), short-term exposure of the substitute nucleotide 5-bromo-2′-deoxyuridine (BrdU) was conducted. Immunohistochemical analysis, using fluorescein-labelled antibodies to trace BrdU, showed pronounced cell division in the coelomic epithelium and axial organ. In the pyloric caeca, not considered as an HPT, proliferation was not detected. BrdU labelling of monolayers of cells obtained by collagenase treatment of coelomic epithelium, axial organ and Tiedemann body revealed induced cell proliferation in response to both LPS and ConA while proliferation of pyloric caeca and circulating coelomocytes remained sparse. By using confocal microscopy it was observed that both the morphology and functional behaviour of cells released from explants of coelomic epithelium showed high similarity to those of circulating phagocytes. It was concluded that the increased coelomocyte numbers observed in response to LPS and ConA were reflected in an induced cell proliferation in coelomic epithelium, axial organ and Tiedemann body, which reinforces the idea that these organs are HPTs and the sources of coelomocyte renewal.

Possibility of mixed progenitor cells in sea star arm regeneration

In contrast to most vertebrates, invertebrate deuterostome echinoderms, such as the sea star Asterias rubens, undergo regeneration of lost body parts. The current hypothesis suggests that differentiated cells are the main source for regenerating arm in sea stars, but there is little information regarding the origin and identity of these cells. Here, we show that several organs distant to the regenerating arm responded by proliferation, most significantly in the coelomic epithelium and larger cells of the pyloric caeca. Analyzing markers for proliferating cells and parameters indicating cell ageing, such as levels of DNA damage, pigment, and lipofuscin contents as well as telomere length and telomerase activity, we suggest that cells contributing to the new arm likely originate from progenitors rather than differentiated cells. This is the first study showing that cells of mixed origin may be recruited from more distant sources of stem/progenitor cells in a sea star, and the first described indication of a role for pyloric caeca in arm regeneration. Data on growth rate during arm regeneration further indicate that regeneration is at the expense of whole animal growth. We propose a new working hypothesis for arm regeneration in sea stars involving four phases: wound healing by coelomocytes, migration of distant progenitor cells of mixed origin including from pyloric caeca, proliferation in these organs to compensate for cell loss, and finally, local proliferation in the regenerating arm.

Ocean acidification and host–pathogen interactions : blue mussels, Mytilus edulis, encountering Vibrio tubiashii

Ocean acidification (OA) can shift the ecological balance between interacting organisms. In this study, we have used a model system to illustrate the interaction between a calcifying host organism, the blue mussel Mytilus edulis and a common bivalve bacterial pathogen, Vibrio tubiashii, with organisms being exposed to a level of acidification projected to occur by the end of the 21st century. OA exposures of the mussels were carried out in relative long-term (4 months) and short-term (4 days) experiments. We found no effect of OA on the culturability of V. tubiashii, in broth or in seawater. OA inhibited mussel shell growth and impaired crystalline shell structures but did not appear to affect mussel immune parameters (i.e haemocyte counts and phagocytotic capacity). Despite no evident impact on host immunity or growth and virulence of the pathogen, V. tubiashii was clearly more successful in infecting mussels exposed to long-term OA compared to those maintained under ambient conditions. Moreover, OA exposed V. tubiashii increased their viability when exposed to haemocytes of OA-treated mussel. Our findings suggest that even though host organisms may have the capacity to cope with periods of OA, these conditions may alter the outcome of host-pathogen interactions, favouring the success of the latter.