Co-occurrence of pathogen assemblages in a keystone species the common cockle Cerastoderma edule on the Irish coast

Despite coinfections being recognized as the rule in animal populations, most studies focus on single pathogen systems. Pathogen interaction networks and the drivers of such associations are lacking in disease ecology studies. Common cockle Cerastoderma edule populations are exposed to a great diversity of pathogens, thus making them a good model system to investigate. This study examined the diversity and prevalence of pathogens from different taxonomic levels in wild and fished C. edule on the Irish coast. Potential interactions were tested focussing on abiotic (seawater temperature and salinity) and biotic (cockle size and age, and epiflora on shells) factors. No Microsporidia nor OsHV-1μVar were detected. Single infections with Haplosporidia (37.7%) or Vibrio (25.3%) were more common than two-pathogen coinfected individuals (9.5%), which may more easily succumb to infection. Fished C. edule populations with high cockle densities were more exposed to infections. Higher temperature and presence of epiflora on cockle shells promoted coinfection in warmer months. Low seawater salinity, host condition and proximity to other infected host species influenced coinfection distribution. A positive association between two Minchinia spp. was observed, most likely due to their different pathogenic effect. Findings highlight the major influence that ecological factors have on pathogen interactions and host–pathogen interplay

Biotic and abiotic factors influencing haplosporidian species distribution in the cockle Cerastoderma edule in Ireland

The Phylum Haplosporidia consists of four genera (Minchinia, Haplosporidium, Urosporidium and Bonamia) that are endoparasitic protists of a wide range of marine invertebrates including commercial bivalve species. Characterization of haplosporidian species remains a challenge due to their patchy spatial and temporal distributions, host-restricted occurrence, and poorly known life cycles. However, they are commonly associated with significant mortality events in bivalves. Due to the recent sporadic mortality events that have occurred in cockles in Europe, the objectives of this study were to determine the diversity, distribution and seasonality of haplosporidian species in Cerastoderma edule populations at several Irish sites. The role of abiotic (temperature, salinity and dissolved oxygen in water) and biotic (cockle size and age) factors as drivers or inhibitors of haplosporidian infection were also assessed. Cockles (n = 998) from the intertidal were sampled from April/July 2018 to April 2019 at three sites with no commercial fishing activity on the south coast (Celtic Sea) and one site on the northeast coast (Irish Sea) with an active commercial fishery. Screening of the cockles by molecular techniques (PCR, Sanger sequencing) and by histopathology was carried out. Two species were identified and confirmed in Irish C. edule for the first time, Minchinia mercenariae -like (14.8%) and Minchinia tapetis (29.6%). Similar to other haplosporidian parasites, the Minchinia spp. detected in our study were present year-round at all sites, except for M. tapetis in Youghal Bay (Celtic Sea). Coinfection of both Minchinia species was only observed in Cork Harbour (Celtic Sea) and Dundalk Bay (Irish Sea), where Minchinia spp. showed a higher presence compared to Youghal Bay and Dungarvan Harbour (Celtic Sea). Moreover, haplosporidians detected with generic primers, were present at all of the sample sites throughout the year but had a higher occurrence during the winter months and were positively correlated with dissolved oxygen. Likewise, smaller and older C.edule seemed to be more vulnerable to the haplosporidian infection. Furthermore, haplosporidian distribution displayed spatial variability between and within sample sites, with the highest presence being observed in cockles at one of the commercially fished Dundalk beds, while the lowest presence was observed in cockles at the second Dundalk bed that was more influenced by freshwater runoff when the tide was out. Findings from this study provide additional information on the distribution and seasonal presence of novel haplosporidian species and their potential abiotic and biotic drivers/inhibitors of infection

Cockle health, disease connectivity and trophic interaction dynamics

Parasites and pathogens are an essential part of the community structure, and their transmission and development are affected not only by their host’s immune system and host population density, but also by multiple biological and environmental drivers that impact on the host-parasite dynamics. There are many gaps in the understanding of these processes and how they influence one another. Given their ecological and economic importance and their exposure to fluctuating and stressful coastal environments, common cockles Cerastoderma edule are ideal model organisms to investigate these dynamics within a natural framework. Therefore, this thesis is intended to present C. edule as a model pathosystem to study marine epizootics and their increased risk of occurrence due to the current changing environment. This thesis develops the understanding of the biological and environmental factors that influence the distribution and dynamics of the host-parasite interplay. In this study, single and multiple parasite and pathogen infections plus over a range of study areas were utilised to better understand these dynamics. The initial part of the study describes a previously undocumented geographic range on the northeast and south coasts of Ireland of two haplosporidian species, Minchinia tapetis and Minchinia mercenariae-like, that have been recently characterised in C. edule. In order to understand the Minchinia:cockle disease dynamics, species-specific primers were developed and these parasites were detected by PCR and confirmed by sequencing. The histology, in turn, confirmed the presence of spore-like stages in the connective tissues of C. edule. A range of factors including host condition and environmental drivers affected both spatial and temporal patterns of these haplosporidian infections in C. edule. Based on statistical modelling, high dissolved oxygen in seawater and small cockle size were the main drivers of the haplosporidian prevalence in the study. The detailed examination of a wider microbial community within C. edule by PCR and histology revealed that single infections with Haplosporidia or Vibrio were more common than coinfected individuals, as expected by random chance. During this second study, no microsporidian species nor ostreid herpesvirus-1 microVar (OsHV-1 μVar) or variants were detected. Based on statistical modelling, coinfected individuals with Haplosporidia and Vibrio occurred because the same risk factors (increased seawater temperature, reduced salinity, and poor host condition) promoted their presence and increased the probability of infection by both pathogen groups. Coinfection with both Minchinia species was overrepresented in our samples and a positive association between M. tapetis and M. mercenariae-like was statistically confirmed. Findings emphasise the need for a more holistic approach in pathological studies, considering the diverse pathogen community within the cockle and their relationship with the environment. The transmission of the microbial community within C. edule to shorebird populations via consumption of cockles and the existing connectivity with the environment (sediment) were also assessed by molecular techniques (PCR and Sanger sequencing) during a third study. Identical strains of Vibrio splendidus were identified in C. edule and, for the first time, in shorebird faecal samples. The consumption of C. edule, thus, may be a source of this Vibrio species in shorebirds. Our findings add evidence to the significant role migratory birds may have as carriers of infectious agents, enabling their transport and dispersal. The findings also support the role of the sediment as an environmental reservoir for Vibrio, which may have an impact on the infaunal community within the sediment responsible for different ecosystem services. Ultimately, in a study that spanned the east and west coast of Ireland and the Welsh coast, patterns of occurrence of Minchinia and Vibrio spp. in C. edule were associated with environmental stressors (high concentration of nitrates and warm temperatures), which may have affected host susceptibility and increased pathogen prevalence. These findings highlight the impact of site-specific environmental stress and habitat degradation on pathogen emergence, particularly in coastal marine environments. In conclusion, this thesis develops a greater understanding of C. edule as a pathosystem and its potential for the early detection of emergent infectious agents, which will have important implications for the sustainability of wild and cultured bivalve populations and the functioning of coastal ecosystems in a changing environmental context

Connectivity dynamics in Irish mudflats between microorganisms including Vibrio spp., common cockles Cerastoderma edule, and shorebirds

Shellfish, including the key species the common cockle Cerastoderma edule, living and feeding in waters contaminated by infectious agents can accumulate them within their tissues. It is unknown if microbial pathogens and microparasites can subsequently be transmitted via concomitant predation to their consumers, including shorebirds. The objective of this study was to assess if pathogens associated with C. edule could be detected seasonally in the faeces of shorebirds that feed on C. edule and in the physical environment (sediment) in which C. edule reside, along the Irish and Celtic Seas. Two potentially pathogenic global groups, Vibrio and Haplosporidia, were detected in C. edule. Although Haplosporidia were not detected in the bird faeces nor in the sediment, identical strains of Vibrio splendidus were detected in C. edule and bird faecal samples at sites where the oystercatcher Haematopus ostralegus and other waders were observed to be feeding on cockles. Vibrio spp. prevalence was seasonal and increased in C. edule and bird faecal samples during the warmer months, possibly due to higher seawater temperatures that promote the replication of this bacteria. The sediment samples showed an overall higher prevalence of Vibrio spp. than the bird faecal and C. edule samples, and its detection remained consistently high through the sites and throughout the seasons, which further supports the role of the sediment as a Vibrio reservoir. Our findings shed light on the fact that not all pathogen groups are transmitted from prey to predator via feeding but bacteria such as V. splendidus can be. As most of the wading birds observed in this study are migratory, the results also indicate the potential for this bacterium to be dispersed over greater geographic distances, which will have consequences for areas where it may be introduced

Summer-drought constrains the phenology and growth of two coexisting Mediterranean oaks with contrasting leaf habit: implications for their persistence and reproduction

13 páginas, 9 figuras, 5 tablas.-- El PDF es la versión post-print.This study analyses how coexisting evergreen and deciduous oaks adjust their phenology to cope with the stressful Mediterranean summer conditions. We test the hypothesis that the vegetative and reproductive growth of the winter deciduous (Quercus faginea Lam.) is more affected by summer drought than that of the evergreen [Quercus ilex L. subsp. ballota (Desf.) Samp.]. First, we assessed the complete aboveground phenology of both species during two consecutive years. Shoot and litter production and bud, acorn and secondary growth were monitored monthly. Second, we identified several parameters affected by summer conditions: apical bud size, individual leaf area (LA), leaf mass per area (LMA) and acorn yield in both species, and leaf-fall in Q. faginea; and analysed their variation over 10 years. Q. ilex performed up to 25% of shoot growth and most leaf development during summer, whereas Q. faginea completed most of both phenophases during spring. Secondary growth was arrested in summer under drought conditions. Approximately, 30–40% of bud and 40–50% of acorn growth was undertaken during summer in both species. Summer drought related to differences in LA, LMA and leaf senescence, but not to acorn yield. Both species had similar year-to-year patterns of acorn production, though yields were always lower in Q. faginea. Bud size decreased severely in both species during extremely dry years. In Q. ilex, bud size tended to alternate between years of large and small buds, and these patterns were followed by opposite trends in stem length. In Q. faginea, bud size was more stable through time. Q. ilex was more phenologically active during summer than Q. faginea, indicating a higher tolerance to drought. Furthermore, bud and fruit growth (the only two phenophases that both species performed during summer) were more severely affected by summer drought in Q. faginea than in the evergreen. The differential effects of summer drought on key phenophases for the persistence (bud growth) and colonization ability (fruit production) of both species may have consequences for their coexistence.This study was possible thanks to the collaboration within the GLOBIMED network (Ministerio de Educación y Ciencia, Spain) and it was supported by the MEC-CICyT projects AGF96-0399, CGL2007-66066-C04/BOS and CGL2008- 04847-C02-01, DGA projects P-038/96 and GA-LC-011/2008, and INIA projects RTA2005-00100-C02-00 and SUM2006-00025-00-00. JJC acknowledges the support of the ‘‘Fundación Aragón I+D’’. SP and RM were funded by MEC by a postdoc (SEUI-FECYT) and a Juan de la Cierva contract, respectively. JA was funded by DGA.Peer reviewe

Seasonal Variability of Dry Matter Content and Its Relationship with Shoot Growth and Nonstructural Carbohydrates

30 páginas, 3 figuras, 4 tablas.[EN]• This study assesses how different phases of shoot growth underlie seasonal change in leaf and stem dry matter content (LDMC and SDMC, respectively) of 12 woody Mediterranean species. The relationship between LDMC and nonstructural carbohydrate (NSC) concentrations is also explored and the seasonal vs interspecies variability of LDMC compared. • LDMC, SDMC and shoot elongation rate (SER) were measured on a monthly basis for a minimum of 12 months. Bud growth rate (BGR) and NSC concentrations were also assessed in several of the study species. • LDMC and SDMC decreased during shoot elongation in spring and increased in summer, showing a significant negative correlation with SER, but were unrelated to BGR. Half of the species analysed showed a positive relationship between LDMC and NSC. Seasonal fluctuations of LDMC within species were higher than interspecies differences, and species ranking was significantly affected by the month of sampling, except during winter months. • Seasonal changes in LDMC and SDMC are mainly related to shoot elongation phenology, and NSC sink–source relationships between old and growing organs can explain this relationship in some species. Owing to the high seasonal variability in LDMC, it is recommended that samples for comparative purposes should be collected as close to the winter as possible.The authors are grateful to Peter Millard, Owen Atkin and four anonymous referees for helpful comments on earlier versions of the manuscript, and to Patricia Fustero and Elena Lahoz for their help with plant sampling and processing. Mark Brewer from BioSS provided helpful advice on REML analysis. SP and RM were funded by MEC by a postdoc contract (SEUIFECYT) and a Juan de la Cierva contract, respectively. JA was founded by DGA. This study was supported by the research projects SUM 2006-00025-00-00 and RTA 2006-00100- CO2-00 (INIA) and the project CGL 2007-66066-CO4-02/ BOS (CICyT). JJC acknowledges the support of Fundación ‘Aragón I+D’.Peer reviewe