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Biology subjects

Takatsuka, S.

Publications and source records attributed to Takatsuka, S..

2 recordsLinked to original sources

Millisecond-scale behaviours of plankton quantified in situ and in vitro using the Event-based Vision Sensor (EVS)

The Event-based Vision Sensor (EVS) is a bio-inspired sensor that captures detailed motions of objects, developed with the applicability to become the eyes of machines and especially self-driving cars. Compared to conventional frame-based image sensors as employed in video cameras, EVS has an extremely fast motion capture equivalent to 10,000-fps even with standard optical settings and additionally has high dynamic ranges for brightness and also lower consumption of memory and energy. These features make the EVS an ideal method to tackle questions in biology, such as the fine-scale behavioural ecology. Here, we developed 22 characteristic features for analysing the motions of aquatic particles from the raw data of the EVS, and deployed the EVS system in both natural environments and laboratory aquariums to test its applicability to filming and analysing plankton behaviour. Our EVS monitoring in turbid water at the bottom of Lake Biwa, Japan identified several particles exhibiting distinct cumulative trajectory with periodicities in their motion (up to 16 Hz), suggesting that they were living organisms with rhythmic behaviour. We also carried out EVS monitoring in the deep sea aided by infrared lighting to minimise influence on behaviour, and observed particles with active motion and periodicities over 40 Hz. Furthermore, we used the EVS to observe laboratory cultures of six species of zooplankton and phytoplankton, confirming that they have species-specific motion periodicities of up to 41 Hz. We applied machine learning to automatically classify particles into five categories (four categories of zooplankton plus passive particles), which achieved an accuracy up to 86%. Our attempts to use the EVS for biological observations, especially focusing on its millisecond-scale temporal resolution and wide dynamic range provide a new avenue to investigate rapid and periodical motion and behaviour in small organisms. Given its compact size with low consumption of battery and memory, the EVS will likely be applicable in the near future for the automated monitoring of the behaviour of plankton by edge computing on autonomous floats, as well as quantifying rapid cellular-level activities under microscopy.

animal behavior and cognition↗

Cas9/CRISPR genome editing to demonstrate the contribution of Cyp51A Gly138Ser to azole resistance in Aspergillus fumigatus

Azole resistance in Aspergillus fumigatus is predominantly associated with increased expression of Cyp51A (lanosterol 14-demethylase), the target enzyme of azole antifungal agents, or with single-nucleotide polymorphisms (SNPs) in cyp51A. Although several SNPs that may be linked to low susceptibility in azole-resistant isolates have previously been reported, few studies have been conducted to conclusively demonstrate the contribution of SNPs to decreased azole susceptibility. An A. fumigatus strain was isolated from the sputum of a 74-year-old male receiving long-term voriconazole treatment for chronic progressive pulmonary aspergillosis. Etest antifungal susceptibility testing showed low susceptibility to voriconazole, itraconazole, and posaconazole. Nucleotide sequencing of cyp51A from this isolate revealed the mutations Gly138Ser (GGC[->]AGC) and Asn248Lys (AAT[->]AAA) compared with the cyp51A of azole-susceptible isolates. PCR-amplified DNA fragments containing cyp51A with or without the mutations of interest and a hygromycin marker were simultaneously introduced along with the Cas9 protein and in vitro-synthesized single-guide RNA into protoplasts of the azole-resistant/susceptible strains. Etest azole susceptibility testing of recombinant strains showed an increased susceptibility via the replacement of Ser138 by glycine. In contrast, azole susceptibility was slightly decreased when a Ser138 mutation was introduced into the azole-susceptible strain AfS35, indicating that the serine at position 138 of Cyp51A contributes to low susceptibility in the azole-resistant isolate. Genetic recombination, which has been hampered thus far in clinical isolates, can now be achieved using Cas9/CRISPR genome editing. This technique could be useful to investigate the contribution of other SNPs of cyp51A to azole resistance.

microbiology↗