Keio SFC Academic Society: Grants for RESEARCH PROJECTS
Grant for Research presentation at academic conference (A)Conference Report on Intelligent Systems for Molecular Biology (ISMB)
Kaoru Ogino
Keio University, Faculty of Environment and Information Studies, Third Year Undergraduate
Conference Overview:
Conference Name: Intelligent Systems for Molecular Biology (ISMB)
Date: July 12, 2024 – July 17, 2024
Venue: Palais des Congrés de Montréal, Montréal, QC, Canada
Format of Organization: Hybrid (Both in-person and online)
Research Title: Unveiling the Genomic Insight into the Exceptional Visual System of a Jumping
spider, Carrhotus xanthogramma
Presentation Format: In-person Poster Presentation
Conference Report:
I have participated in the conference, Intelligent Systems for Molecular Biology (ISMB) that washeld in Montreal, Canada and had the precious opportunity to present my research through aposter presentation. This conference is held once every year by the International Society for
Computational Biolgy (ISCB) and recognized as one of the largest bioinformatics andcomputational biology conference in the world that gathers various researchers to discuss andshowcase developments and innovations made in various fields. It was held both in-person andonline attence bringing 1,900 participants from different countries and background, making it apriceless opportunity that enabled many learning experiences.
This was my first conference and an international one, as well as being the very few
undergraduate participants, which made this experience invaluable. Being able to present myresearch on the genome assembly of the jumping spider, Carrhotus xanthogramma and thespecies identification of an unknown japanese fruitfly, Torinido-shoujoubae, through COI DNA
barcoding was rewarding. There were many opportunities to communicate with variousresearchers, professors and PhD students about my work and was able to receive insightful
feedbacks which has given me directions for the future of my research. Especially since theconference accomodates experienced scientists from various fields within the larger entity ofbiology, I was able to broaden my understanding on the research that I undertook and provided
me with potential ideas and techniques to apply to my studies.
The conference was not only about presenting my research but also learning and networking. I
attended numerous talks and discussions that covered a wide range of topics in bioinformatics
such as the use of language models in predictions of proteins or the development of algorithms
for genomic analysis. These sessions were highly informative and allowed me to have anexposure to new ideas and the latest advancements and challenges in a wide ranging field.
The attendance in ISMB 2024 was a pivotal experience in my academic journey with theexposure to the global scientific community, feedbacks on my research and the learning andnetworking opportunities have signficantly contributed to my growth as a researcher. I am deeply
grateful for this granted opportunity and look forward on applying what I have gained throughthis conference to further advance my research for a fruitful outcome.

Abstract

Poster:Methods
FUKUOKA,JAPANMorphologicalSampling

Identification

Sequencing
DataResults:
Genome Size
Estimation- SEQUEL II- HiFi Long Reads- 177,709,857,788 bp

k-mer Histogram
The graph wasproduced using
paired-end Illumina
reads with k-mer of 31.
The estimated genome
size is 3.6Gb. Bimodal
distribution can beobserved where thefirst peak represents
the heterozygous sites
while the second peak
represents
homozygous sites
which are evident in
diploid organisms.

Pre-processing
Trimming

Assembly
PECAT

Error
Correction

RepeatTill complete BUSCO
reach maximum
value

Jumping spiders (Araneae: Salticidae) exhibit exceptional visual acuity, unique courtship displays andhunting strategies, which diverge from the typical web-building tactics of other spiders. This visual system
is supported by four pairs of eyes, specializing in prey identification and motion detection.
This research aims to elucidate genetic underpinning of the visual system, physical attributes and silk
production of jumping spiders, through both behavioral and molecular genomic approach, while
conducting a comparative study with other spiders. Establishment of reliable breeding protocols isnecessary for maintaining replicability and for generating controlled populations for genetic andbehavorial analysis. As part of this effort, we identified a previously unknown Drosophila sp. feed,
Torinido-shoujoubae in Japanese, as Drosophila hydei through COI DNA barcoding. This enables themaintenance of spiders' natural diet and behavior in captivity, thereby providing more authentic context
for studying hunting strategies. Genome sequencing of Carrhotus xanthogramma, a jumping spider, wasconducted through the use of two techniques: short paired-end reads by Illumina Novaseq 6000 and longreads by PacBio Sequel II. However, the preliminary results of hybrid genome assembly using
HASLR and MaSuRCA presented significantly low quality in BUSCO coverage, indicating
challenges in obtaining a complete and accurate genome. The future direction of
our research will focus on resolving assembly issues and analyzing thegenetic basis of the jumping spider's visual system and silk
production. By comparing these genes with those ofother spiders, we aim to shed light on theevolutionary adaptations that haveenabled the remarkablecapabilities of jumpingspiders.

Results:
Assembly Statistics
Input
Lengths(Coverage)

GenomeScope: Vurture et al., 2017, Bioinformatics- Novaseq 6000
- Paired-end shortreads- 227,196,730,800 bp
each

Quality
Assessment

Genomesize

Introduction: Opsin
After
AssemblywithPECATGENOMEASSEMBLY
Complete

Description

Statistics

Long reads

177,709,857,788 bp (~50x)

Paired-end (Forward)

227,196,730,800 bp (~65x)

Paired-end (Reverse)

227,196,730,800 bp (~65x)

Expected

3.2~3.6Gb

After Assembly

4.3Gb

Complete BUSCOs

51.7%

Length4,391,551,459

Number of Scaffolds

51247

N50120KB

GC Content (%)

30.81%

Genome Assembly Statistics
This table compares the statistics
between the different stages ofgenome assembly, including theoriginal sequencing reads, raw
assembly. Polishing the raw
assembly may improve theComplete BUSCO scores while
also improving the otherstatistics of the assembly.

PECAT: Nie et al., 2024, Nature Communications
Pilon: Walker et al., 2014, PloS ONE
BUSCO: Manni et al., 2021, Molecular Biology and Evolution.

Future Work

Ko

Gene Prediction and Annotation• Predict coding regions and annotate genes• BLAST analysis to assign functional information topredicted genes
Functional Genomics• Identification of Opsin Genes: insights tophotoreceptive capabilities and visual adaptations• Spidroin Gene Characterization: analyze spidroin
genes against their roles such as uses in dragline andburrow construction

Morehouse et al., 2017, The Biological Bulletin

Carrhotus xanthogramma:
Genome Size Estimation
Genome Assembly
Visual SystemOpsin

2. Faculty of Environment andInformation Studies, Keio University3. Graduate School of Media and Governance,
Keio UniversityResults:
TaxonomicDiagnosis

Comparative Genomics

Species
identification oflivefood flightless fly(Torinido−shoujoubae)by using DNA barcoding• Genomic Comparison with Other Spiders: identify
conserved and divergent regions• Phylogenetic Analysis: elucidate evolutionary
relationships between C. xanthogramma and otherspider species

Rößler et al., 2021, Frontiers in Zoology

h2 , ,31,21,
no noigO Ko
Institute for Advancedru ki Biosciences,oKeio UniversityaKa obu
1. Institute for AdvancedNBiosciences, Keio University

Further Assembly• Polishing and Refinement: further rounds of error
correction using Pilon• Alternative Assembly Tools: compare assembly tools
to assess the efficacy

1,aawgkaNa(Right, Top) Diagnosis of Torinido-shoujoubae.
A, habitus (lateral view); B, male fore tarsus(ventral view); C, left wing (ventral view). Arrowheads indicate diffuse pale areas on posterolateralabdominal bands (A), prominent fine setae on malefore tarsus (B), and tip of subcostal break (C),respectively. Scale: solid line = 1 mm; dotted line =0.2 mm.
(Right, Bottom) Male and female terminalia of
Torinido-shoujoubae. A, B, periphallic organs(caudal and caudolateral view, respectively); CE, phallic organs (ventral, ventrolateral, andlateral view, respectively); F, G, oviscapt (ventraland lateral view, respectively); H, spermatheca.
Scale: solid line = 0.1 mm.

z

A

B

COI 1490

Forward/
ReverseForward

COI 2198

Reverse

COI 1718

Forward

COI 2776

ReverseCoverage (bp)1082

Results: Phylogenetic Tree

100

NCBI (National Center forBiotechnology
Information)

Tree Constructionusing Figtree

Tree Visualization
using IQ-TREE

Figtree: http://tree.bio.ed.ac.uk/software/figtree/
IQ-TREE: Minh et al., 2020, Molecular Biology andEvolution

Acknowledgements
References

Altschul, S. F., Gish, W., Miller, W., Myers, E. W., & Lipman, D. J. (1990). Basic local alignment search tool. Journal of Molecular Biology, 215(3), 403–410.
https://doi.org/10.1016/S0022-2836(05)80360-2
Manni, M., Berkeley, M. R., Seppey, M., Simão, F. A., & Zdobnov, E. M. (2021). BUSCO Update: Novel and Streamlined Workflows along with Broader and Deeper Phylogenetic
Coverage for Scoring of Eukaryotic, Prokaryotic, and Viral Genomes. Molecular Biology and Evolution, 38(10), 4647–4654. https://doi.org/10.1093/molbev/msab199
Minh, B. Q., Schmidt, H. A., Chernomor, O., Schrempf, D., Woodhams, M. D., Von Haeseler, A., & Lanfear, R. (2020). IQ-TREE 2: New Models and Efficient Methods for Phylogenetic
Inference in the Genomic Era. Molecular Biology and Evolution, 37(5), 1530–1534. https://doi.org/10.1093/molbev/msaa015
4. Morehouse, N. I., Buschbeck, E. K., Zurek, D. B., Steck, M., & Porter, M. L. (2017). Molecular Evolution of Spider Vision: New Opportunities, Familiar Players. The BiologicalBulletin, 233(1), 21–38. https://doi.org/10.1086/693977
5. Nakagawa, K., Ogino, K., Katoh, T. K., and Nobuaki, K. (2024). Species identification of livefood flightless fly (Torinido-shoujoubae) through DNA barcoding. Ecology and Evolution:Nature Notes. https://doi.org/10.1002/ece3.116221
6. Nie, F., Ni, P., Huang, N., Zhang, J., Wang, Z., Xiao, C., Luo, F., & Wang, J. (2024). De novo diploid genome assembly using long noisy reads. Nature Communications, 15(1), 2964.
https://doi.org/10.1038/s41467-024-47349-7
7. Ratnasingham, S., & Hebert, P. D. N. (2007). BOLD: The Barcode of Life Data System (http://www.barcodinglife.org). Molecular Ecology Notes, 7(3), 355–364.
https://doi.org/10.1111/j.1471-8286.2007.01678.x
8. Rößler, D. C., De Agrò, M., Biundo, E., & Shamble, P. S. (2021). Hanging by a thread: Unusual nocturnal resting behaviour in a jumping spider. Frontiers in Zoology, 18(1), 23.
https://doi.org/10.1186/s12983-021-00410-3
9. Vurture, G. W., Sedlazeck, F. J., Nattestad, M., Underwood, C. J., Fang, H., Gurtowski, J., & Schatz, M. C. (2017). GenomeScope: Fast reference-free genome profiling from shortreads. Bioinformatics, 33(14), 2202–2204. https://doi.org/10.1093/bioinformatics/btx153
10. Walker, B. J., Abeel, T., Shea, T., Priest, M., Abouelliel, A., Sakthikumar, S., Cuomo, C. A., Zeng, Q., Wortman, J., Young, S. K., & Earl, A. M. (2014). Pilon: An Integrated Tool for
Comprehensive Microbial Variant Detection and Genome Assembly Improvement. PLoS ONE, 9(11), e112963. https://doi.org/10.1371/journal.pone.0112963

D. eohydei

2.
3.

99.6

99.4

Similarity Percentage (%)

BOLD (Barcode of LifeDatabase)

We would like to express our gratitude to Yuya Suzuki for the sampling, Takehiro K. Katoh for taxonomic diagnosis, Maaya Domon for
the technical support, and Keizo Takasuka for his valuable advice.

1.
D. m elanopalpa

BLAST (Basic Local
Alignment Search Tool)

BOLD: Ratnasingham et al., 2007, NatureCommunications
NCBI: Altschul et al., 1990, Journal of Molecular
Biology

Similarity Percentage of Top 50 Hits

99.8

Amplified COI geneis sequenced toobtain the sequenceby Eurofins
Genomics

725z

Results: BLAST Analysis

Sequencing

Electrophoresis

Set COI Primers

Verify the size andquality of amplified
DNA using gelelectrophoresis

Phylogenetic Analysis

kaoogi@keio.jp

Cytochrome oxidasesubunit I (COI) genes
amplified throughPCR using primers
listed below.

BLAST Alignment

nSca

Extract DNA from leg
samples using
Biomasher II withLysis buffer andProteinase K

Amplification

DNA ExtractionMethods

99.2

9998.898.6

98.4

Grants

98.2
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50

Similarity Hits
Wild Sky

Hobby club

SpringTails

charm

Bar graph of the top 50 BLAST Results on BOLD.
The gray scale bars represents all the data aligned with D. hydei. The red andorange bars are the only two outlier species found in all search of the samples
from the four companies.

OSIMO Foundation

Yamagata Prefecture

Phylogenetic Tree of Torinido-shoujoubae and known Drosophila species.
TS is an abbreviation for Torinido-shoujoubae. Values located at the node refers
to the bootstrap

JSPS KAKENHI Grant Number
JP21H02210
Tsuruoka City

JSPS KAKENHI (PAGS)

Keio SFC Academic Society
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