Droplet digital PCR as a tool for investigating dynamics of cryptic symbionts.

Interactions among symbiotic organisms and their hosts are major drivers of ecological and evolutionary processes. Monitoring the infection patterns among natural populations and identifying factors affecting these interactions are critical for understanding symbiont-host relationships. However, many of these interactions remain understudied since the knowledge about the symbiont species is lacking, which hinders the development of appropriate tools. In this study, we developed a digital droplet PCR (ddPCR) assay based on apicomplexan COX1 gene to detect an undescribed agamococcidian symbiont. We show that the method gives precise and reproducible results and enables detecting cryptic symbionts in low target concentration. We further exemplify the assay's use to survey seasonally sampled natural host (Pygospio elegans) populations for symbiont infection dynamics. We found that symbiont prevalence differs spatially but does not show seasonal changes. Infection load differed between populations and was low in spring and significantly increased towards fall in all populations. We also found that the symbiont prevalence is affected by host length and population density. Larger hosts were more likely to be infected, and high host densities were found to have a lower probability of infection. The observed variations could be due to characteristics of both symbiont and host biology, especially the seasonal variation in encounter rates. Our findings show that the developed ddPCR assay is a robust tool for detecting undescribed symbionts that are otherwise difficult to quantify, enabling further insight into the impact cryptic symbionts have on their hosts.

Seasonal genetic variation associated with population dynamics of a poecilogonous polychaete worm.

Poecilogonous species show variation in developmental mode, with larvae that differ both morphologically and ecologically. The spionid polychaete Pygospio elegans shows variation in developmental mode not only between populations, but also seasonally within populations. We investigated the consequences of this developmental polymorphism on the spatial and seasonal genetic structure of P. elegans at four sites in the Danish Isefjord-Roskilde-Fjord estuary at six time points, from March 2014 until February 2015. We found genetic differentiation between our sampling sites as well as seasonal differentiation at two of the sites. The seasonal genetic shift correlated with the appearance of new size cohorts in the populations. Additionally, we found that the genetic composition of reproductive individuals did not always reflect the genetic composition of the entire sample, indicating that variance in reproductive success among individuals is a likely explanation for the patterns of chaotic genetic patchiness observed during this and previous studies. The heterogeneous, unpredictable character of the estuary might maintain poecilogony in P. elegans as a bet-hedging strategy in the Isefjord-Roskilde-Fjord complex in comparison with other sites where P. elegans are expected to be fixed to a certain mode of development.

The Story of a Hitchhiker: Population Genetic Patterns in the Invasive Barnacle Balanus(Amphibalanus) improvisus Darwin 1854.

Understanding the ecological and evolutionary forces that determine the genetic structure and spread of invasive species is a key component of invasion biology. The bay barnacle, Balanus improvisus (= Amphibalanus improvisus), is one of the most successful aquatic invaders worldwide, and is characterised by broad environmental tolerance. Although the species can spread through natural larval dispersal, human-mediated transport through (primarily) shipping has almost certainly contributed to the current global distribution of this species. Despite its worldwide distribution, little is known about the phylogeography of this species. Here, we characterize the population genetic structure and model dispersal dynamics of the barnacle B. improvisus, and describe how human-mediated spreading via shipping as well as natural larval dispersal may have contributed to observed genetic variation. We used both mitochondrial DNA (cytochrome c oxidase subunit I: COI) and nuclear microsatellites to characterize the genetic structure in 14 populations of B. improvisus on a global and regional scale (Baltic Sea). Genetic diversity was high in most populations, and many haplotypes were shared among populations on a global scale, indicating that long-distance dispersal (presumably through shipping and other anthropogenic activities) has played an important role in shaping the population genetic structure of this cosmopolitan species. We could not clearly confirm prior claims that B. improvisus originates from the western margins of the Atlantic coasts; although there were indications that Argentina could be part of a native region. In addition to dispersal via shipping, we show that natural larval dispersal may play an important role for further colonisation following initial introduction.

A new haemocyanin in cuttlefish (Sepia officinalis) eggs: sequence analysis and relevance during ontogeny.

BACKGROUND: Haemocyanin is the respiratory protein of most of the Mollusca. In cephalopods and gastropods at least two distinct isoforms are differentially expressed. However, their physiological purpose is unknown. For the common cuttlefish Sepia officinalis, three isoforms are known so far, whereas for only two of them the complete mRNA sequences are available. In this study, we sequenced the complete mRNA of the third haemocyanin isoform and measured the relative expression of all three isoforms during embryogenesis to reveal a potential ontogenetic relevance. RESULTS: The cDNA of isoform 3 clearly correlates to the known Sepia officinalis haemocyanin subunits consisting of eight functional units and an internal duplicated functional unit d. Our molecular phylogenetic analyses reveal the third isoform representing a potentially ancestral haemocyanin isoform, and the analyses of the expression of haemocyanin type 3 reveal that haemocyanin type 3 only can be observed within eggs and during early development. Isoforms 1 and 2 are absent at these stages. After hatching, isoform 3 is downregulated, and isoform 1 and 2 are upregulated. CONCLUSIONS: Our study clearly shows an embryonic relevance of the third isoform, which will be further discussed in the light of the changes in the physiological function of haemocyanin during ontogeny. Taken together with the fact that it could also be the isoform closest related to the common ancestor of cuttlefish haemocyanin, the phylogeny of cuttlefish haemocyanin may be recapitulated during its ontogeny.