Biology subjects

Giulio, M.

Publications and source records attributed to Giulio, M..

4 recordsLinked to original sources

bioRxiv2025

Comparing DNA extraction methods for successful PacBio HiFi sequencing: a case study of the freshwater mussel Anodonta anatina (Bivalvia: Unionidae)

High-quality reference genomes are increasingly recognized as essential resources in biodiversity genomics and conservation. However, successful DNA extraction and long-read sequencing remain highly organism-dependent. Molluscs, a diverse phylum of invertebrates, pose particular challenges due to the presence of inhibitory compounds and the difficulty of obtaining high-molecular-weight DNA, often necessitating careful optimization of extraction protocols. Here, we present a case study on the freshwater mussel Anodonta anatina (Bivalvia: Unionidae), evaluating two preservation methods, six DNA extraction protocols, and two post-extraction clean-up steps for their effects on DNA quality and PacBio HiFi sequencing yield from foot tissue of a single individual. The PacBio Nanobind and CTAB protocols produced high-quality DNA from fresh tissue but performed poorly on flash-frozen tissue. Post-extraction clean-up generally degraded DNA and did not improve sequencing yield. Unexpectedly, the column-based Omega Mollusc Kit, although not designed for high-molecular-weight DNA, performed better than the PacBio-recommended Nanobind kit and the manual CTAB method on flash-frozen tissue. It generated high DNA quantity and purity, sufficient integrity for HiFi sequencing, and appeared to remove contaminants effectively. While the resulting DNA may be too fragmented for ultra-long read sequencing, the Omega Mollusc Kit offers a practical, cost-effective first approach for testing DNA extraction and PacBio sequencing in flash-frozen A. anatina foot tissue. When fresh tissue is available, Nanobind or CTAB were the best-performing options in our comparison. Overall, our results provide a practical starting point for protocol selection, while acknowledging that validation across other mollusc species, tissue types, and preservation methods remains important. This strategy could reduce the need for extensive protocol optimization and facilitate future mollusc genomics efforts.

genomics↗

bioRxiv2025

Beyond genetic indicators: how reproductive mode and hybridization challenge freshwater mussel conservation

Genetic diversity is a fundamental component of biodiversity that should be monitored and preserved. Unionid freshwater mussels provide essential ecosystem services but are among the most imperilled aquatic organisms worldwide. Here, we assess key genetic Essential Biodiversity Variables (EBVs)--genetic diversity, genetic differentiation, inbreeding, and effective population size--in all Anodonta species in Switzerland. After generating draft genomes for Anodonta cygnea and Anodonta anatina, we performed whole-genome resequencing of 421 individuals (243 A. anatina, 151 A. cygnea, 17 A. exulcerata and 10 Anodonta sp.) from 31 populations. While A. anatina populations followed a metapopulation structure shaped by catchment areas, genetic diversity correlated positively with waterbody size, suggesting greater vulnerability in small ponds compared with large lakes. Inbreeding levels were low, however effective population sizes were consistently below 100, indicating serious extinction risks. We also detected hybridization between A. cygnea and A. exulcerata, indicating genomic permeability between these species. Furthermore, genomic data suggested facultative selfing in A. cygnea, leading to a marked reduction in genetic diversity, increased population structure and inbreeding, and a decline in effective population size compared to the outcrossing A. anatina, emphasizing that A. cygnea faces a particularly high risk of extinction due to its reproductive strategy. Our study reveals the vulnerability of freshwater mussels and emphasizes the need for genetic indicators to reflect species-specific reproductive strategies. More broadly, it calls for conservation policies to integrate genetic monitoring and consider reproductive modes to tailor conservation efforts and better assess extinction risks.

genomics↗

bioRxiv2023

The genome of the rayed Mediterranean limpet Patella caerulea (Linnaeus, 1758)

Patella caerulea (Linnaeus, 1758) is a molluscan limpet species of the class Gastropoda. Endemic to the Mediterranean Sea, it is considered to be a keystone species in tidal and subtidal habitats due to its primary role in structuring and regulating the ecological balance of these habitats. It is currently being used as a bioindicator to assess the environmental quality of coastal marine waters and as a model species to understand adaptation to ocean acidification. Here we provide a high-quality reference genome assembly and annotation for Patella caerulea. We used a single specimen collected in the field to generate [~]30 Gb of PacBio HiFi data. The final assembly is 749.8 Mb large and contains 62 contigs, including the mitochondrial genome (14,938 bp). With an N50 of 48.8 Mb and 98% of the assembly contained in the 18 largest contigs, this assembly is near chromosome-scale. BUSCO scores were high (Mollusca: 87.8% complete; Metazoa: 97.2% complete) and similar to metrics observed for other chromosome-level Patella genomes, highlighting a possible bias in the Mollusca database for Patellids. We generated transcriptomic Illumina data from a second individual collected at the same locality, and used it together with protein evidence to annotate the genome. 23,938 protein coding gene models were found. By comparing this annotation with other published Patella annotations, we found that the distribution and median values of exon and gene lengths was comparable to other Patella species despite different annotation approaches. The present high-quality Patella caerulea reference genome is an important resource for future ecological and evolutionary studies. SignificanceReference genomes are essential resources for biodiversity conservation and management. Patella caerulea (Linnaeus, 1758) is a gastropod species occurring in the Mediterranean Sea that is currently used as a model to understand the impact of pollution and ocean acidification on marine biodiversity. Here we present a high-quality reference genome of P. caerulea, that almost reaches chromosome-level contiguity. We further provide a high-quality genome annotation supported by transcriptomic evidence. This reference genome will be of interest for researchers working on the ecology and evolution of marine biodiversity. All data is available on the public database NCBI for future use by researchers.

genomics↗

bioRxiv2023

Benchmarking sample pooling for epigenomics of natural populations

Interest in the role of DNA methylation (DNAm) has grown in ecological and evolutionary research of natural populations. While researchers are typically interested in comparing population-level variation, individual sequencing is the current standard. Natural populations have low effect sizes and thus need large sample sizes to detect differences. The cost of sequencing the necessary samples can be prohibitive in DNAm work. Pooling DNA before library preparation is a powerful tool to reduce costs but no recommendations exist for DNAm pooling in ecology-epigenetics research. We test if pooled and individual libraries provide similar global and region-specific DNA methylation signals in a natural system of response to pollution. We generated whole-epigenome data for two freshwater invasive molluscs (Corbicula flumina and Dreissena polymorpha) collected from a polluted and unpolluted locality, Lake Maggiore, Italy. Our results support that pooling effectively captures the same genome-wide and global treatment-level signals as individual libraries but we note that pooled libraries yielded orders of magnitude more input data and differentially-methylated regions (DMRs) detected compared with individual libraries. We estimated greatly lower power for regions from individual libraries compared with pooled libraries. The post-hoc process of computationally pooling data from individual libraries produced results comparable to pooled libraries in volumes but had discrepancies between DMRs. We discuss the possible causes for the discrepancies and put our results in the context of the benefits and drawbacks of sample pooling for epigenomics of natural populations.

genomics↗