Bernarda Calla

The Olympia oyster ( Ostrea lurida ) was once a major food source and a key component of coastal marine ecosystems and culture on the US West Coast. Overfishing and habitat contamination in the early to mid-20th century severely depleted populations of Olympia oysters, and the species is currently considered a species of concern. Although multiple restoration efforts are underway, progress understanding the species biology and recovery potential has been limited by the historical lack of genomic resources. Here, we report a chromosome-level assembly of the Olympia oyster. The assembly was built using Oxford Nanopore reads and Hi-C to obtain a highly contiguous 1.03 Gb-assembly consisting of 10 chromosomes. RNA from gills, mantle, adductor muscle, and a mix of brooding eggs and larvae were also sequenced as 150 nt long paired-end reads which were used to guide the genome annotation. Over 52 K genes were predicted in the genome, of which 34K were protein-coding. A BUSCO analysis on the assembly showed 98.9% of the mollusca_odb12 gene set were complete (with 2.1% duplicated, and 0.8% missing). The short RNA reads were also used to identify tissue-level gene expression signatures in adductor muscles, egg and brooding larvae, and mantle. Development-related genes were highly expressed in eggs and brooding larvae whereas many xenobiotic metabolism genes were present in mantle tissues, highlighting mantle’s role in the interaction of the oyster with external environments. This new genomic resource will greatly advance our understanding of this keystone mollusk and provide a critical tool for guiding restoration efforts and the development of sustainable commercial production.

Despite its broad applicability, genome-wide expression profiling may be costly when scaling up. An alternative to conventional full-length mRNA sequencing are 3’ end mRNA sequencing methods, which selectively sequence only the 3’-most end of individual transcripts, resulting in a single mapped read per transcript in the sample. This method offers some advantages including low cost, reduced amount of input sample, and independency from transcript length bias inherent to traditional full-length RNA-seq. In this study, we tested a 3’-end mRNA sequencing method (Tag-seq) in the Pacific oyster (Magallana gigas), a species of economic and ecologic relevance characterized by an unusual highly polymorphic genome, and for which available reference genomes are not yet fully functionally annotated. Our samples were collected from a farm that was undergoing a mass mortality event enabling us to evaluate changes in expression during stress conditions and mortality. Tag-seq yielded reads that mapped to a high proportion of the transcriptome, and these mapping rates were comparable to those of RNA-seq. However, Tag-seq data exhibited high dispersion due to low depth and coverage (fewer reads available). This variability resulted in a reduced number of detected differentially expressed genes, approximately half of those identified with RNA-seq. Tag-seq was robust finding known general physiological functions that differed between healthy and stressed oysters, making it a good, cost-effective choice for large studies with a high number of samples. Nevertheless, it provided a less comprehensive picture of the Pacific oyster transcriptome under stress compared to RNA-seq.

Hemolymph is vital for bivalves, contributing to their innate immune system, nutrient transport, waste elimination, and hormone regulation. Yet, traditional sampling methods are often invasive or fatal to the organisms. In this study, we evaluated the effects of repeated, non-lethal hemolymph sampling on adult Pacific oysters (Magallana gigas). Five- year-old oysters were relaxed with magnesium sulphate, and hemolymph was extracted from half of the individuals, while the other half with relaxation served as controls. Sampling took place during gonad maturation in two groups: one laboratory conditioned (approximately three months in indoor tanks with controlled environment), and one naturally conditioned (approximately seven months at the bay in Yaquina Bay, Oregon, USA). Total mortalities ranged from 10% in the laboratory groups (after four samplings) and 22% (after seven samplings) in the natural groups; most of the mortalities took place after the last sampling. Sex ratio was similar between the sampled (66%) and control groups (63%) in the laboratory setting. In the natural setting, the final sex ratio was male biased in the group that was repeatedly sampled for hemolymph (58%) compared to the non-sampled controls (28%). Our findings highlight that repeated hemolymph sampling can be performed with minimal mortality, allowing non-lethal monitoring of hemolymph physiology over time.

Background/Objectives: Sex determination and differentiation exhibit remarkable molecular diversity across taxa, driven by genetic, epigenetic, and environmental factors. Invertebrates with sequential hermaphroditism, such as the Pacific oyster (Magallana gigas), represent a poorly understood system despite their role as keystone species and contribution to a substantial aquaculture industry. Methods: To identify sex-related molecular markers during gametogenesis, we repeatedly sampled hemolymph from artificially conditioned oysters over two months, and sex phenotypes were assigned at the end of the experiment by biopsy. Results: RNA-sequencing analysis of five males and five females revealed subtle yet consistent sex-specific transcriptional signatures in hemolymph. We show that gametogenesis proceeds asynchronously among oysters, even within the same sex individuals. Complex physiological trade-offs were discovered between sexes during gonad maturation; in early stages of sexual maturation, females prioritized cell division, whereas males suppressed it. Females exhibited higher expression of solute carrier family (SLC) genes, suggesting enhanced nutrient exchange during oogenesis. Temporal dynamics highlighted differential expression of genes regulating cross-membrane ion gradients (e.g., transient receptor potential channels) and signal transduction (e.g., signal transducer and activator of transcription), previously linked to environmental sex determination (ESD) in some reptilian species. Conclusions: Together, these findings underscore that gametogenesis in Pacific oysters is complex and dynamic, and that molecular pathways of ESD may be partially conserved between invertebrate and vertebrate species.

Background: The genetic diversity of a population defines its ability to adapt to episodic and fluctuating environmental changes. For species of agricultural value, available genetic diversity also determines their breeding potential and remains fundamental to the development of practices that maintain health and productivity. In this study, we used whole-genome resequencing to investigate genetic diversity within and between naturalized and captively reared populations of Pacific oysters from the US Pacific coast. The analyses included individuals from preserved samples dating to 1998 and 2004, two contemporary naturalized populations, and one domesticated population. Results: Despite high overall heterozygosity, there was extremely low but significant genetic divergence between populations, indicative of high gene flow and/or little variability from founding events. The captive population, which was reared for over 25 years was the most genetically distinct population and exhibited reduced nucleotide diversity, attributable to inbreeding. Individuals from populations that were separated both geographically and temporally did not show detectable genetic differences, illustrating the consequences of human intervention in the form translocation of animals between farms, hatcheries and natural settings. Fifty-nine significant F _ST outlier sites were identified, the majority of which were present in high proportions of the captive population individuals, and which are possibly associated with domestication. Conclusion: Pacific oysters in the US Pacific coast harbor high genetic heterozygosity which obscures weak population structure. Differences between these Pacific oyster populations could be leveraged for breeding and might be a source of adaptation to new environments.

Aspergillus fungi are ubiquitous inhabitants of colonies of the western honey bee (Apis mellifera), where they interact with bees in associations ranging from parasitism to possible mutualism. Aspergillus Flavi fungi are frequently found in bee bread (pollen processed for longterm storage) and are thought to contribute to food preparation, processing, preservation, and digestion. Conditions in the hive are challenging for fungi due, in part, to xeric and acidic properties of bee bread and the omnipresence of propolis, an antimicrobial product manufactured by bees from plant resins. We used quantitative and qualitative assays to determine whether A. flavus isolated from bee bread demonstrates tolerance for hive environmental conditions in terms of temperature, pH, osmotic pressure, and propolis exposure. Comparisons made use of three strains of A. flavus: a fungal biocontrol product not known from beehives (AF36), a strain isolated from bee bread (AFBB) in hives from central Illinois, and a pathogenic strain from a honey bee colony displaying symptoms of stonebrood (AFPA). Strain AFBB displayed higher tolerance of acidic conditions, low matric potential (simulating xeric substrate), and propolis exposure than did other strains. A genomic comparison between this new strain and the reference NRRL‐3357 showed that AFBB, like AF36, might be blocked from carrying out aflatoxin biosynthesis. Sequence comparisons also revealed several missense variants in genes that encode proteins regulating osmotolerance and osmotic pressure in Aspergillus spp., including SakA, SskB, GfdA, and TcsB/Sln1. Collectively, results of our laboratory assays and genetic analyses are consistent with the suggestion that the strain isolated from bee bread is adapted to the bee bread environment and may have persisted due to a coevolutionary relationship between Aspergillus and A. mellifera. This finding bolsters recent concerns about the effects of fungicide use near bee colonies and broadens the ecological importance of highly adaptable fungal strains.

Guadalupe Fernandez-Milmanda, Bernarda Calla, Mary Williams; Mind the middleman: How receptor-like cytoplasmic kinases mediate plant immunity, Plant Physiology,

Social behaviour is defined as the interaction between two or more individuals of the same species and includes “all behavior that tends to bring individuals together” ( Whishaw et al., 2006). Unicellular organisms can be highly social, frequently forming structured groups and expressing collective behaviour. For example, bacteria are capable of quorum sensing, a mechanism by which cells communicate to sense population density ( Miller and Bassler, 2001). In a more unique example, the Gram-negative bacteria Myxococcus xanthus expresses cooperative feeding by hunting for food in swarms and forms fruiting bodies upon starvation ( Dworkin and Kaiser, 1985). Microorganisms can also socialize by forming cell aggregates, such as biofilms attached to surfaces ( Stoodley et al., 2002) or surface-free aggregates ( Cai, 2020) in response to changes in environmental conditions.

Legumes can form symbiotic associations with rhizobia, which are nitrogen-fixing bacteria that colonize roots by forming facultative organs termed nodules. The development of nodules depends on a concerted interchange of signals between the plant and the symbiotic bacteria. Ultimately, legumes benefit from this association by being able to use nitrogen that would otherwise be unavailable, while rhizobia successfully multiply by utilizing plant resources mainly in the form of sugars ( Oldroyd et al., 2011). The question remains, however, how plants distinguish these beneficial bacteria from bacteria that cause disease.