Freshwater mussel conservation: A global horizon scan of emerging threats and opportunities.

We identified 14 emerging and poorly understood threats and opportunities for addressing the global conservation of freshwater mussels over the next decade. A panel of 17 researchers and stakeholders from six continents submitted a total of 56 topics that were ranked and prioritized using a consensus-building Delphi technique. Our 14 priority topics fell into five broad themes (autecology, population dynamics, global stressors, global diversity, and ecosystem services) and included understanding diets throughout mussel life history; identifying the drivers of population declines; defining metrics for quantifying mussel health; assessing the role of predators, parasites, and disease; informed guidance on the risks and opportunities for captive breeding and translocations; the loss of mussel-fish co-evolutionary relationships; assessing the effects of increasing surface water changes; understanding the effects of sand and aggregate mining; understanding the effects of drug pollution and other emerging contaminants such as nanomaterials; appreciating the threats and opportunities arising from river restoration; conserving understudied hotspots by building local capacity through the principles of decolonization; identifying appropriate taxonomic units for conservation; improved quantification of the ecosystem services provided by mussels; and understanding how many mussels are enough to provide these services. Solutions for addressing the topics ranged from ecological studies to technological advances and socio-political engagement. Prioritization of our topics can help to drive a proactive approach to the conservation of this declining group which provides a multitude of important ecosystem services.

A roadmap for the conservation of freshwater mussels in Europe.

Europe has a long history of human pressure on freshwater ecosystems. As pressure continues to grow and new threats emerge, there is an urgent need for conservation of freshwater biodiversity and its ecosystem services. However, whilst some taxonomic groups, mainly vertebrates, have received a disproportionate amount of attention and funds, other groups remain largely off the public and scientific radar. Freshwater mussels (Bivalvia, Unionida) are an alarming example of this conservation bias and here we point out six conceptual areas that need immediate and long-term attention: knowledge, threats, socioeconomics, conservation, governance and education. The proposed roadmap aims to advance research, policy and education by identifying the most pressing priorities for the short- and long-term conservation of freshwater mussels across Europe.

Playing Peekaboo with a Master Manipulator: Metagenetic Detection and Phylogenetic Analysis of Wolbachia Supergroups in Freshwater Invertebrates.

The infamous "master manipulators"-intracellular bacteria of the genus Wolbachia-infect a broad range of phylogenetically diverse invertebrate hosts in terrestrial ecosystems. Wolbachia has an important impact on the ecology and evolution of their host with documented effects including induced parthenogenesis, male killing, feminization, and cytoplasmic incompatibility. Nonetheless, data on Wolbachia infections in non-terrestrial invertebrates are scarce. Sampling bias and methodological limitations are some of the reasons limiting the detection of these bacteria in aquatic organisms. In this study, we present a new metagenetic method for detecting the co-occurrence of different Wolbachia strains in freshwater invertebrates host species, i.e., freshwater Arthropoda (Crustacea), Mollusca (Bivalvia), and water bears (Tardigrada) by applying NGS primers designed by us and a Python script that allows the identification of Wolbachia target sequences from the microbiome communities. We also compare the results obtained using the commonly applied NGS primers and the Sanger sequencing approach. Finally, we describe three supergroups of Wolbachia: (i) a new supergroup V identified in Crustacea and Bivalvia hosts; (ii) supergroup A identified in Crustacea, Bivalvia, and Eutardigrada hosts, and (iii) supergroup E infection in the Crustacea host microbiome community.

Blood parasites shape extreme major histocompatibility complex diversity in a migratory passerine.

Pathogens are one of the main forces driving the evolution and maintenance of the highly polymorphic genes of the vertebrate major histocompatibility complex (MHC). Although MHC proteins are crucial in pathogen recognition, it is still poorly understood how pathogen-mediated selection promotes and maintains MHC diversity, and especially so in host species with highly duplicated MHC genes. Sedge warblers (Acrocephalus schoenobaenus) have highly duplicated MHC genes, and using data from high-throughput MHC genotyping, we were able to investigate to what extent avian malaria parasites explain temporal MHC class I supertype fluctuations in a long-term study population. We investigated infection status and infection intensities of two different strains of Haemoproteus, that is avian malaria parasites that are known to have significant fitness consequences in sedge warblers. We found that prevalence of avian malaria in carriers of specific MHC class I supertypes was a significant predictor of their frequency changes between years. This finding suggests that avian malaria infections partly drive the temporal fluctuations of the MHC class I supertypes. Furthermore, we found that individuals with a large number of different supertypes had higher resistance to avian malaria, but there was no evidence for an optimal MHC class I diversity. Thus, the two studied malaria parasite strains appear to select for a high MHC class I supertype diversity. Such selection may explain the maintenance of the extremely high number of MHC class I gene copies in sedge warblers and possibly also in other passerines where avian malaria is a common disease.

Extreme MHC class I diversity in the sedge warbler (Acrocephalus schoenobaenus); selection patterns and allelic divergence suggest that different genes have different functions.

BACKGROUND: Recent work suggests that gene duplications may play an important role in the evolution of immunity genes. Passerine birds, and in particular Sylvioidea warblers, have highly duplicated major histocompatibility complex (MHC) genes, which are key in immunity, compared to other vertebrates. However, reasons for this high MHC gene copy number are yet unclear. High-throughput sequencing (HTS) allows MHC genotyping even in individuals with extremely duplicated genes. This HTS data can reveal evidence of selection, which may help to unravel the putative functions of different gene copies, i.e. neofunctionalization. We performed exhaustive genotyping of MHC class I in a Sylvioidea warbler, the sedge warbler, Acrocephalus schoenobaenus, using the Illumina MiSeq technique on individuals from a wild study population. RESULTS: The MHC diversity in 863 genotyped individuals by far exceeds that of any other bird species described to date. A single individual could carry up to 65 different alleles, a large proportion of which are expressed (transcribed). The MHC alleles were of three different lengths differing in evidence of selection, diversity and divergence within our study population. Alleles without any deletions and alleles containing a 6 bp deletion showed characteristics of classical MHC genes, with evidence of multiple sites subject to positive selection and high sequence divergence. In contrast, alleles containing a 3 bp deletion had no sites subject to positive selection and had low divergence. CONCLUSIONS: Our results suggest that sedge warbler MHC alleles that either have no deletion, or contain a 6 bp deletion, encode classical antigen presenting MHC molecules. In contrast, MHC alleles containing a 3 bp deletion may encode molecules with a different function. This study demonstrates that highly duplicated MHC genes can be characterised with HTS and that selection patterns can be useful for revealing neofunctionalization. Importantly, our results highlight the need to consider the putative function of different MHC genes in future studies of MHC in relation to disease resistance and fitness.

The size and shape of parasitic larvae of naiads (Unionidae) are not dependent on female size.

The naiads, large freshwater mussels (Unionida), have very long life spans, are large-bodied, and produce thousands to millions of larvae (glochidia) which typically must attach to host fish tissues to metamorphose into a juvenile mussel. Glochidia develop within a female's marsupial gill demibranch, thus their number is restricted by female size. However, larger mussels acquire more energy, which could be invested in either larger-sized glochidia, in a more glochidia, or a combination of both. The high level of host specialization seen in many naiads may constrain glochidial size and shape around a narrow optimum, while naiads that use a wide range of host fishes may be predicted to possess greater plasticity in glochidial morphology. In this paper, we investigated the relationship between maternal body size and progeny body size and shape, aided by modern digital microscopy. We analyzed the between- and within- species variation of glochidia size and shape relative to female size in four widespread species of European naiads: Anodonta anatina, Anodonta cygnea, Unio crassus and Unio tumidus. Whereas the total reproductive output is collinear with female body size, substantial differences between species in glochidia size were found within genus Anodonta, but not genus Unio where glochidial size is remarkably consistent. The glochidial shape, however, differed within both Unio and Anodonta. We interpret this constant within-species glochidial size in Unio as reflecting a constraint imposed by the likelihood of successful transmission onto and off from a narrow range of hosts, whereas their shape seems to be less constrained. The Anodonta species, inhabiting a wide spectrum of habitats and using more than twice the number of fish hosts than Unio spp., have larger glochidia with greater variation in size and shape. Our results suggest that measures of glochidial variability may also serve as an indicator of host specificity in other naiads.

Response of freshwater mussel recruitment to hydrological changes in a eutrophic floodplain lake.

Although eutrophication of freshwaters is a natural process, the human impact often leads to inland waters becoming overloaded with nutrients, impoverishing many valuable and vanishing habitats, such as floodplain lakes. These changes need to be reversed if the occurrence of endangered aquatic species is to be restored. In this paper we analyse the impact of a change in the water regime of a naturally eutrophic floodplain lake, which harbours a large diversity of Unionidae (large freshwater mussels), a globally threatened taxonomic group that provides important ecosystem functions and services. We found that a slight increase in the discharge from this waterbody, following the construction of an additional outflow pipe, positively influenced recruitment in three of the five mussel species inhabiting the lake. We also found that, after the construction of this additional outflow, the niches of juveniles of Anodonta cygnea and Unio spp. changed, revealing differences in their hydrological requirements. Our results suggest that, as in lotic habitats, complex hydraulic parameters are highly significant to unionid mussels in lentic conditions.

The Patterns and Puzzles of Genetic Diversity of Endangered Freshwater Mussel Unio crassus Philipsson, 1788 Populations from Vistula and Neman Drainages (Eastern Central Europe).

Mussels of the family Unionidae are important components of freshwater ecosystems. Alarmingly, the International Union for Conservation of Nature and Natural Resources Red List of Threatened Species identifies almost 200 unionid species as extinct, endangered, or threatened. Their decline is the result of human impact on freshwater habitats, and the decrease of host fish populations. The Thick Shelled River Mussel Unio crassus Philipsson, 1788 is one of the examples that has been reported to show a dramatic decline of populations. Hierarchical organization of riverine systems is supposed to reflect the genetic structure of populations inhabiting them. The main goal of this study was an assessment of the U. crassus genetic diversity in river ecosystems using hierarchical analysis. Different molecular markers, the nuclear ribosomal internal transcribed spacer ITS region, and mitochondrial DNA genes (cox1 and ndh1), were used to examine the distribution of U. crassus among-population genetic variation at multiple spatial scales (within rivers, among rivers within drainages, and between drainages of the Neman and Vistula rivers). We found high genetic structure between both drainages suggesting that in the case of the analyzed U. crassus populations we were dealing with at least two different genetic units. Only about 4% of the mtDNA variation was due to differences among populations within drainages. However, comparison of population differentiation within drainages for mtDNA also showed some genetic structure among populations within the Vistula drainage. Only one haplotype was shared among all Polish populations whereas the remainder were unique for each population despite the hydrological connection. Interestingly, some haplotypes were present in both drainages. In the case of U. crassus populations under study, the Mantel test revealed a relatively strong relationship between genetic and geographical distances. However, in detail, the pattern of genetic diversity seems to be much more complicated. Therefore, we suggest that the observed pattern of U. crassus genetic diversity distribution is shaped by both historical and current factors i.e. different routes of post glacial colonization and history of drainage systems, historical gene flow, and more recent habitat fragmentation due to anthropogenic factors.

On the reintroduction of the endangered thick-shelled river mussel Unio crassus: The importance of the river's longitudinal profile.

Freshwater mussels of the order Unionida provide important ecosystem functions and services. Unfortunately, some previously widespread species are now seriously endangered. To restore the historical range of the population of Unio crassus in the Biala River, southern Poland, the species was reintroduced into a series of 'stepping stones' joining two remnant populations. During the first phase of the study, the relationships between the abundance of U. crassus, physical habitat, and water quality were studied to assess reintroduction potential. In general, chemical water quality improved upstream from the existing population, favouring the decision for reintroduction, whereas morphological variables worsened. Mussel abundance was correlated negatively with the elevation and slope of channel, organic matter contents, and pH (exceeding 8.0), but positively with silt presence, water conductivity, and concentrations of HCO3-, Ca2+, and NO3-. During the second phase, adult individuals were introduced into one type of functional habitat-marginal channel sectors with still water and fine sediment. Despite the initial very high rate of reproduction in some parts of the upper reach of the river, the juveniles were ultimately recruited only in the lower part of the restored range, resulting in a very rapid change in recruitment at a channel slope of 1.8‰. Recruitment was positively related to silt content, conductivity, and Ca2+ and HCO3- ions, negatively to channel elevation and slope, and water pH. The host fish species showed no correlation with abiotic habitat features within the studied reach. These results imply that most of the habitat traits related to U. crassus occurrence depended on the river's longitudinal profile, not on the chemical water quality, and that final success of introduction should be evaluated after several years.

Conservation status of freshwater mussels in Europe: state of the art and future challenges.

Freshwater mussels of the Order Unionida provide important ecosystem functions and services, yet many of their populations are in decline. We comprehensively review the status of the 16 currently recognized species in Europe, collating for the first time their life-history traits, distribution, conservation status, habitat preferences, and main threats in order to suggest future management actions. In northern, central, and eastern Europe, a relatively homogeneous species composition is found in most basins. In southern Europe, despite the lower species richness, spatially restricted species make these basins a high conservation priority. Information on freshwater mussels in Europe is unevenly distributed with considerable differences in data quality and quantity among countries and species. To make conservation more effective in the future, we suggest greater international cooperation using standardized protocols and methods to monitor and manage European freshwater mussel diversity. Such an approach will not only help conserve this vulnerable group but also, through the protection of these important organisms, will offer wider benefits to freshwater ecosystems.

Gender-Associated Mitochondrial DNA Heteroplasmy in Somatic Tissues of the Endangered Freshwater Mussel Unio crassus (Bivalvia: Unionidae): Implications for Sex Identification and Phylogeographical Studies.

Some bivalve species possess two independent mitochondrial DNA lineages: maternally (F-type) and paternally (M-type) inherited. This phenomenon is called doubly uniparental inheritance. It is generally agreed that F-type mtDNA is typically present in female somatic and gonadal tissues as well as in male somatic tissues, whereas the M-type mtDNA occurs only in male germ line and gonadal tissue. In the present study, the mtDNA heteroplasmy (for both F and M genomes) in male somatic tissues of Unio crassus (Philipsson, 1788), species threatened with extinction, has been confirmed. Taking advantage from the presence of Mcox1 marker only in male somatic tissues, we developed a new method of sex identification in this endangered species, using nondestructive tissue sampling. Probability of correct sex identification was estimated at 97.5%. The present study is the first report on gender-associated mitochondrial DNA heteroplasmy in male somatic tissues of thick-shelled river mussel and first approach to U. crassus sex identification at molecular level. Our study also confirmed the utility of paternally inherited Mcox1 gene fragment as a complementary molecular tool for resolving phylogeographical relationships among populations of thick-shelled river mussel.

Relationship between duration of repolarisation and T wave amplitude in patients with positive or negative T wave alternans.

BACKGROUND: Experimental studies documented the relationship between T wave alternans (TWA) and duration of refractoriness. To date, association between TWA and QT interval on standard ECG has not been examined. Aim. To assess the relationship between TWA and QT interval. METHODS: The study group consisted of 70 patients (57 males, mean age 56+/-16 years) with implantable cardioverterdefibrillator (ICD). TWA was measured using a high-resolution ECG obtained from surface orthogonal bipolar XYZ leads and analysed using a Fast Fourier transform. All recordings were performed during ventricular pacing at 100 betas/min. Correlation between T wave amplitude (T max) and QT interval (measured from R wave to T max) was calculated. RESULTS: TWA was found in 18 patients. In this group of patients, there was a significant positive correlation between Tmax and QT (r = 0.766), whereas in patients with negative TWA no such correlation was detected. CONCLUSIONS: (1) Positive correlation between QT and T max probably depicts the relationship between T wave amplitude and duration of repolarisation, which is associated with TWA; (2) methods used for T wave localisation, based on the identification of Q wave (with possible QT-RR correction) overestimate TWA due to periodic changes (with TWA frequency) in location of T wave in the analysed window; and (3) these results provide new insights in the genesis of TWA.

[The state of working conditions in light of control recommendations issued by the State Labor Inspectorate--selected problems]. / Stan warunków pracy w swietle ustalen kontrolnych PIP--wybrane problemy.

In enterprises covered by the scheme of the intensified supervision of the State Labor Inspectorate (SLI), working conditions are analyzed every year, and the measures to reduce or eliminate existing hazards are evaluated. The enterprises with high rates of accidents at work and employees exposed to hazardous chemical and physical factors present in the work environment are classified in this category. The list of such enterprises is subjected to annual verification. Enterprises with potential high or increased risk for serious industrial accidents involving noxious chemicals or dangerous for the local environment are also taken into account. The supervision of the State Labor Inspectorate, lasting already for a number of years, greatly contributed--this is confirmed by numerous inspections--to the sustained improvement of working conditions in enterprises of this category. In accord with the suggestions made by parliamentarians, the issue of intensified supervision of enterprises, representing various branches of the national economy, characterized by the highest rates of occupational hazards has been incorporated into the SLI Plan of Action for the years 2001-2003. In January 2001, district inspectorates selected 137 enterprises of various sectors with the highest hazard rates to be covered by the SLI scheme in the nearest future. The systematic control and promotion activities will be aimed at eliminating or substantial reduction of reported hazards. In the previous year, the SLI tried to find out how far the employers fulfill their obligation to check technical conditions in their enterprises, as well as to discuss the possibility of further use of asbestos-containing products. In the opinion of the SLI, the employers do not comply with these requirements, or they are not willing to bear costs of removal or repair of asbestos-containing products. They postpone endlessly all necessary repairs. It is still impossible to evaluate the situation and to explicitly state that after completing works involving asbestos it is removed from premises designed for people. This is of paramount importance for safety of people at schools, kindergartens, high schools, as well as of hospital personnel and patients. In Polish regulations, the maximum admissible concentrations of asbestos for this kind of premises have no as yet been set. The State Labor Inspector has submitted a motion to the Minister of Health on setting permissible concentrations of asbestos dust in premises meant for people. The regulation (still binding) of March 12, 1996 on admissible concentrations and intensities of factors harmful to the human health, emitted by construction materials, devices and equipment in premises intended for people does not take account of asbestos.

Territory choice during the breeding tenure of male sedge warblers.

A territorial male can shift the location of its territory from year to year in order to increase its quality. The male can base its decision on environmental cues or else on its breeding experiences (when territory shift is caused by breeding failure in previous seasons). We tested these possible mechanisms of territory choice in the sedge warbler (Acrocephalus schoenobaenus), a territorial migrating passerine that occupies wetlands. This species bases its territory choices on an environmental cue: tall wetland vegetation cover. We found that the magnitude of territory quality improvement between seasons (measured as the area of tall wetland vegetation) increased throughout the early stages of a male's breeding career as a result of territory shifts dependent on the earliness of arrival. The distance the territory was shifted between seasons depended negatively on the previous year's territory quality and, less clearly, on the previous year's mating success. On the other hand, previous mating or nesting success had no influence on territory quality improvement between seasons as measured in terms of vegetation. The results imply that tall wetland vegetation is a long-term, effective environmental cue and that a preference for territories in which this type of landcover prevails has evolved into a rigid behavioral mechanism, supplemented by short-term individual experiences of breeding failure.

Lead interaction: rare cause of oversensing during implantation procedure of implantable cardioverter-defibrillator system.

A variety of etiologies can cause erroneous detection in patients with implantable cardioverter defibrillator (ICDs). Interaction between two endocardial leads is rare and uncommon in causing electrical noise. During a reimplantation procedure of an ICD system in a 68-year-old man, additional electrical signals could be detected. The interaction between two endocardial defibrillation leads was identified as the cause of sensing problems. When it is not possible to extract the nonfunctional endocardial lead during implantation of the new electrode, it should be implanted away and not in parallel from the old one to avoid interaction between them. A variety of etiologies can cause erroneous detection in patiens with ICD. Interaction between two endocardial leads is rare and uncommon to cause electrical noise.