The genome sequence of a marine yeast, Metschnikowia zobellii (Uden & Cast.-Branco, 1961)

We present a genome assembly from a Metschnikowia zobellii culture (a marine yeast; Ascomycota; Saccharomycetes; Saccharomycetales; Metschnikowiaceae). The genome sequence is 13.6 megabases in span. Most of the assembly is scaffolded into 5 chromosomal pseudomolecules. The mitochondrial genome has also been assembled and is 51.12 kilobases in length.


Background
Metschnikowia zobellii (van Uden & Castelo-Branco, 1961) van Uden, 1962 is a marine ascomycete yeast in the family Metschnikowiaceae which infects small crustaceans, including copepods (Seki & Fulton, 1969).M. zobellii spores are elongated and barbed, with one spore occurring per ascus (Mendonca-Hagler et al., 1993).These needle-like spores penetrate the gut walls of grazing invertebrates where the yeast multiples, eventually killing the host (van Uden & Castelo-Branco, 1961).Metschnikowia are components of coastal marine planktonic communities that reoccur year on year (Chrismas et al., 2023) and may play an important role in plankton population dynamics.
The genome of M. zobellii was sequenced as part of the Darwin Tree of Life Project, a collaborative effort to sequence all named eukaryotic species in the Atlantic Archipelago of Britain and Ireland.This genome is one of the first marine fungal genomes available to the scientific community and will provide new insights and opportunities for marine fungal research.

Genome sequence report
The genome was sequenced from a colony of Metschnikowia zobellii collected from Church Reef, Wembury, Devon, UK (50.32,.A total of 90-fold coverage in Pacific Biosciences single-molecule HiFi long reads was generated.Primary assembly contigs were scaffolded with chromosome conformation Hi-C data. The final assembly has a total length of 13.6 Mb in 5 sequence scaffolds with a scaffold N50 of 2.8 Mb (Table 1).The whole assembly sequence was assigned to 5 chromosomal-level scaffolds.Chromosome-scale scaffolds confirmed by the Hi-C data are named in order of size (Figure 1-Figure 4; Table 2).While not fully phased, the assembly deposited is of one haplotype.Contigs corresponding to the second haplotype have also been deposited.The mitochondrial genome was also assembled and can be found as a contig within the multifasta file of the genome submission.
Metadata for specimens, spectral estimates, sequencing runs, contaminants and pre-curation assembly statistics can be found at https://links.tol.sanger.ac.uk/species/27328.

Sample acquisition and nucleic acid extraction
A colony of Metschnikowia zobellii (specimen ID MBA-191008-001A, individual gsMetZobe1) was collected from Church Reef, Wembury, Devon, UK (latitude 50.32, longitude -4.08) on 2019-10-08.The specimen was collected by Michael Cunliffe (Marine Biological Association) and grown on agar.The identifier was Ro Allen (Marine Biological Association).The sample was harvested and preserved in liquid nitrogen before processing.
DNA was extracted at the Tree of Life laboratory, Wellcome Sanger Institute (WSI).The gsMetZobe1 sample was weighed and some of the sample was set aside for Hi-C sequencing.The cells were cryogenically disrupted to a fine powder using a Covaris cryoPREP Automated Dry Pulveriser, receiving multiple impacts.High molecular weight (HMW) DNA was extracted using the Qiagen Plant MagAttract v3 DNA extraction kit.HMW DNA was sheared into an average fragment size of 12-20 kb in a Megaruptor 3 system with speed setting 30.Sheared DNA was purified by solid-phase reversible immobilisation using AMPure PB beads with a 1.8X ratio of beads to sample to remove the shorter fragments and concentrate the DNA sample.The concentration of the sheared and purified DNA was assessed using a Nanodrop spectrophotometer and Qubit Fluorometer and Qubit dsDNA High Sensitivity Assay kit.Fragment size distribution was evaluated by running the sample on the FemtoPulse system.

Sequencing
Pacific Biosciences HiFi circular consensus DNA sequencing libraries were constructed according to the manufacturers' instructions.DNA sequencing was performed by the Scientific Operations core at the WSI on a Pacific Biosciences SEQUEL II (HiFi) instrument.Hi-C data were also generated from gsMetZobe1 using the Arima2 kit and sequenced on the Illumina NovaSeq 6000 instrument.

Genome assembly, curation and evaluation
Assembly was carried out with Hifiasm (Cheng et al., 2021) and haplotypic duplication was identified and removed with purge_dups (Guan et al., 2020).The assembly was then scaffolded with Hi-C data (Rao et al., 2014) using SALSA2 (Ghurye et al., 2019).The assembly was checked for contamination and corrected as described previously (Howe et al., 2021).Manual curation was performed using HiGlass (Kerpedjiev et al., 2018) andPretext (Harry, 2022).The mitochondrial genome was assembled using MitoHiFi (Uliano- Silva et al., 2023), which runs MitoFinder (Allio et al., 2020) or MITOS (Bernt et al., 2013) and uses these annotations to select the final mitochondrial contig and to ensure the general quality of the sequence.
A Hi-C map for the final assembly was produced using bwa-mem2 (Vasimuddin et al., 2019) in the Cooler file format (Abdennur & Mirny, 2020).To assess the assembly metrics,    Further, the Wellcome Sanger Institute employs a process whereby due diligence is carried out proportionate to the nature of the materials themselves, and the circumstances under  which they have been/are to be collected and provided for use.
The purpose of this is to address and mitigate any potential legal and/or ethical implications of receipt and use of the materials as part of the research project, and to ensure that in doing so we align with best practice wherever possible.The overarching areas of consideration are: • Ethical review of provenance and sourcing of the material

Suriana Sabri
Universiti Putra Malaysia, Selangor, Malaysia The paper presents a detailed genome assembly of the marine yeast Metschnikowia zobellii.This work is a significant contribution to the field, providing one of the first marine fungal genomes, which is crucial for advancing our understanding of the biology and ecology of marine fungi, particularly in the context of host-pathogen interactions.The assembly of M. zobellii is of high quality, spanning 13.6 Mb and organized into five chromosomal pseudomolecules.The use of advanced sequencing technologies, including Pacific Biosciences HiFi and Hi-C data, has resulted in a robust and comprehensive genome assembly.The inclusion of the mitochondrial genome adds further value to the study.This genome provides a foundational resource for marine fungal studies, with significant implications for understanding marine ecosystems and the dynamics of plankton populations, where M. zobellii plays a role.
The paper is well-executed, methodologically sound, and provides important insights into marine fungal genomics.Therefore, it should be accepted for indexing.

Is the rationale for creating the dataset(s) clearly described? Yes
Are the protocols appropriate and is the work technically sound?Yes

Are the datasets clearly presented in a useable and accessible format? Yes
Competing Interests: No competing interests were disclosed.
Reviewer Expertise: Microbial genomics I confirm that I have read this submission and believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard.

Yi Wu
Tianjin University, Tianjin, China The authors have provided the complete genome sequence of Metschnikowia zobellii in this study.This will provide data resources for multiple biology fields.I noticed that Metschnikowia zobellii has less than 1% genomic difference from Saccharomyces cerevisiae, which makes me curious about how this portion of the genome results in the substantial differences in the survival strategies of the two yeasts.
Overall, the paper presents comprehensive data with clear and intuitive analysis.Here are my comments: The introduction provides a good overview of the research background and objectives.However, it could further discuss the specific role of Metschnikowia zobellii in marine ecosystems and its biotechnological potential. 1.

Methods:
The methods are described in detail.However, if the authors could provide more comprehensive annotation of the assembled genome, it would greatly benefit related research fields.

2.
Is the rationale for creating the dataset(s) clearly described?Yes

Are sufficient details of methods and materials provided to allow replication by others? Yes
Are the datasets clearly presented in a useable and accessible format?Yes Competing Interests: No competing interests were disclosed.
Reviewer Expertise: Yeast, synthetic biology I confirm that I have read this submission and believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard.Care should be exercised here, as the said reference does not specify what species of Metschnikowia exhibits annual recurrence.The genus Metschnikowia is large and diverse in the ecology of its species, such that the reference may not pertain to M. zobellii (although it might, but we don't know).

○
Additional information on the strain would be useful, namely the collection where the strain was deposited and the accession number.I presume the strain is publicly available.Identification is attributed to an individual, but the method used and the supporting data (D1/D2 sequence) should be specified.I could not find a relevant GenBank deposit.A D1/D2 sequence would be essential and is not always extractable from whole genomes.

Soo-Je Park
Department of Biology, Jeju National University, Jeju-si, Jeju-do, South Korea In this study, the authors described a high-completeness (i.e., high-qulity) genome of a marine yeast, Metschnikowia zobellii as a pathogen for Daphnia magna.The genome sequences were generated by HiFi technique.The genome is reported to be 13.6 Mb and comprised of 5 scaffolds.Also, the mitochondrial genome was assembled into 51.1 Kb.Protocols used were clearly presented and all data is publicly available within ENA (project: PRJEB52022).
Minor comments: For the reader, the authors should be provided with protocols for specific purposes.

○
It is requested that the specific goals for the analysis tools used be defined, rather than a general statement such as "The genome was analyzed in the BlobToolKit environment."For example, Busco was used to estimate genome completeness, and Hifiasm was used for de novo assembly.This information should be included in Table 3. Reviewer Expertise: environmental genomics, environmental microbiology, ecogenomics.
I confirm that I have read this submission and believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard.

Figure 1 .
Figure 1.Genome assembly of Metschnikowia zobellii, gsMetZobe1.1:metrics.The BlobToolKit Snailplot shows N50 metrics and BUSCO gene completeness.The main plot is divided into 1,000 size-ordered bins around the circumference with each bin representing 0.1% of the 13,653,384 bp assembly.The distribution of scaffold lengths is shown in dark grey with the plot radius scaled to the longest scaffold present in the assembly (3,378,874 bp, shown in red).Orange and pale-orange arcs show the N50 and N90 scaffold lengths (2,836,740 and 2,093,737 bp), respectively.The pale grey spiral shows the cumulative scaffold count on a log scale with white scale lines showing successive orders of magnitude.The blue and pale-blue area around the outside of the plot shows the distribution of GC, AT and N percentages in the same bins as the inner plot.A summary of complete, fragmented, duplicated and missing BUSCO genes in the saccharomycetes_odb10 set is shown in the top right.An interactive version of this figure is available at https://blobtoolkit.genomehubs.org/view/gsMetZobe1.1/dataset/gsMetZobe1_1/snail.

Figure 4 .
Figure 4. Genome assembly of Metschnikowia zobellii, gsMetZobe1.1:Hi-C contact map of the gsMetZobe1.1 assembly, visualised using HiGlass.Chromosomes are shown in order of size from left to right and top to bottom.An interactive version of this figure may be viewed at https://genome-note-higlass.tol.sanger.ac.uk/l/?d=MjYP2bWrTqimY9doy8ZPjA.

Reviewer Report 13
August 2024 https://doi.org/10.21956/wellcomeopenres.22144.r90273© 2024 Wu Y.This is an open access peer review report distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Reviewer Report 05
August 2024 https://doi.org/10.21956/wellcomeopenres.22144.r90279© 2024 Lachance M. This is an open access peer review report distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.Marc-André Lachance University of Western Ontario, Ontario, Canada A high definition sequence of Metschnikowia zobellii is presented.The following are comments that might help improve the final version of the article: In the background paragraph, a reference to Seki and Fulton 1969 is cited in support of a statement regarding Metschnikowia zobellii.Examination of the reference reveals that the species was identified as M. krissii.Having said this, the identification tools available in 1969 (and the data in the 1969 paper) would suggest that a great deal of prudence should be exercised in reporting identifications to the species level performed in the pre-sequencing era.○ Secondly, a statement is made, referring to Chrismas et al., 2023, on the occurrence of Metschnikowia in nature.

○
Is the rationale for creating the dataset(s) clearly described?YesAre the protocols appropriate and is the work technically sound?YesAre sufficient details of methods and materials provided to allow replication by others?YesAre the datasets clearly presented in a useable and accessible format?Yes Competing Interests: No competing interests were disclosed.

Table 3 . Software tools: versions and sources. Software tool Version Open Peer Review Current Peer Review Status: Version 1
This is an open access peer review report distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

the protocols appropriate and is the work technically sound? Yes Are sufficient details of methods and materials provided to allow replication by others? Yes Are the datasets clearly presented in a useable and accessible format? Yes Competing Interests:
(Opulente et al., 2024)an already existing genome sequence for M. zobellii, although there is one: GCA_030572635.1.The sequence has been used in a large phylogeny of hundreds of ascomycetous yeasts(Opulente et al., 2024)1.It might be useful to give a short comparison of the two datasets.No competing interests were disclosed.
○ I do ○ Are

have read this submission and believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard, however I have significant reservations, as outlined above.
Reviewer Report 25 July 2024 https://doi.org/10.21956/wellcomeopenres.22144.r87867© 2024 Park S. This is an open access peer review report distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.