Turtle Insights into the Evolution of the Reptilian Karyotype and the Genomic Architecture of Sex Determination

Sex chromosome evolution remains an evolutionary puzzle despite its importance in understanding sexual development and genome evolution. The seemingly random distribution of sex-determining systems in reptiles offers a unique opportunity to study sex chromosome evolution not afforded by mammals or birds. These reptilian systems derive from multiple transitions in sex determination, some independent, some convergent, that lead to the birth and death of sex chromosomes in various lineages. Here we focus on turtles, an emerging model group with growing genomic resources. We review karyotypic changes that accompanied the evolution of chromosomal systems of genotypic sex determination (GSD) in chelonians from systems under the control of environmental temperature (TSD). These transitions gave rise to 31 GSD species identified thus far (out of 101 turtles with known sex determination), 27 with a characterized sex chromosome system (13 of those karyotypically). These sex chromosomes are varied in terms of the ancestral autosome they co-opted and thus in their homology, as well as in their size (some are macro-, some are micro-chromosomes), heterogamety (some are XX/XY, some ZZ/ZW), dimorphism (some are virtually homomorphic, some heteromorphic with larger-X, larger W, or smaller-Y), age (the oldest system could be ~195 My old and the youngest \u3c 25 My old). Combined, all data indicate that turtles follow some tenets of classic theoretical models of sex chromosome evolution while countering others. Finally, although the study of dosage compensation and molecular divergence of turtle sex chromosomes has lagged behind research on other aspects of their evolution, this gap is rapidly decreasing with the acceleration of ongoing research and growing genomic resources in this group

Phytochemical screening, antimicrobial activity and cytotoxicity of Nepalese medicinal plants Swertia chirayita and Dendrobium amoenum

Research on medicinal plants are important to Nepal because most of its rural population relies on it as mode of medicine. Medicinal plants namely Swertia chirayita and Dendrobium amoenum were collected from mid hills of Nepal. The present study was undertaken to find the antimicrobial activity, phytochemical presence and their cytotoxicity in different extraction medium. The percentage yield from the plants were highest in warm methanol extraction with 12.6%, followed by ethyl acetate and lowest was for cold methanol. Plant extract showed the presence of antioxidants like alkaloid, terpenoids, flavonoids, tannin, glycosides. The Brine Shrimp Bioassay of methanol and ethyl acetate extract showed cytotoxicity. Chiraito extract showed LC50 of 199 ppm for Dhunche sample, 128.82 ppm for Daman sample and 131.82 ppm of Illam sample. The antibacterial activity of methanol extract of Chiraito and Dendrobium amoenum showed significant bioactivity by inhibiting growth of microbial species selected for the test. The zone of inhibition shown by the extracts was comparable to the standard antibiotics. Similarly, methanol extract of Chiraito also showed significant antifungal activity with the zone of inhibition comparable to amphotericin.Nepal Journal of Biotechnology. Dec. 2015 Vol. 3, No. 1: 48-5

Establishment and characterization of turtle liver organoids provides a potential model to decode their unique adaptations

Painted turtles are remarkable for their freeze tolerance and supercooling ability along with their associated resilience to hypoxia/anoxia and oxidative stress, rendering them an ideal biomedical model for hypoxia-induced injuries (including strokes), tissue cooling during surgeries, and organ cryopreservation. Yet, such research is hindered by their seasonal reproduction and slow maturation. Here we developed and characterized adult stem cell-derived turtle liver organoids (3D self-assembled in vitro structures) from painted, snapping, and spiny softshell turtles spanning ~175My of evolution, with a subset cryopreserved. This development is, to the best of our knowledge, a first for this vertebrate Order, and complements the only other non-avian reptile organoids from snake venom glands. Preliminary characterization, including morphological, transcriptomic, and proteomic analyses, revealed organoids enriched in cholangiocytes. Deriving organoids from distant turtles and life stages demonstrates that our techniques are broadly applicable to chelonians, permitting the development of functional genomic tools currently lacking in herpetological research. Such platform could potentially support studies including genome-to-phenome mapping, gene function, genome architecture, and adaptive responses to climate change, with implications for ecological, evolutionary, and biomedical research

Isolation of Arsenic Resistant Escherichia coli from Sewage Water and Its Potential in Arsenic Biotransformation

Arsenic contamination in drinking water from ground water poses a threat to the health of a large population in developing countries in Asia. This has sparked great interests in the potential of different microbes in arsenic resistance and removal from water. This study involves isolation of arsenic resistant Escherichia coli from sewage water from Kathmandu University and investigation of its attributes. Arsenic resistant E. coli was successfully isolated which could survive in high concentration of arsenic. The maximum tolerance of arsenite was 909.79 mg/L (sodium arsenite) and 3120.1 mg/L arsenate (sodium arsenate) which is well above most natural concentration of arsenic in ground water. This particular E. coli tolerated multiple heavy metal like silver nitrate, cobalt sulphate, cadmium chloride, nickel chloride, mercury chloride, copper sulphate, and zinc chloride at concentration 20 µM, 1 mM, 0.5mM, 1mM, 0.01 mM, 1 mM, and 1 mM respectively which are concentrations known to be toxic to E. coli. Biotransformation of arsenite to arsenate was also checked for by a qualitative silver nitrate technique. This E. coli was able to transform arsenate to arsenite. It showed some sensitivity to Ciprofloxacin, Gentamicin and Nalidixic Acid. As E. coli and its genome are very widely studied, these particular properties have a lot of potential in microbial remediation or microbial recovery of metals and possible recombination approaches

Turtle Insights into the Evolution of the Reptilian Karyotype and the Genomic Architecture of Sex Determination

Sex chromosome evolution remains an evolutionary puzzle despite its importance in understanding sexual development and genome evolution. The seemingly random distribution of sex-determining systems in reptiles offers a unique opportunity to study sex chromosome evolution not afforded by mammals or birds. These reptilian systems derive from multiple transitions in sex determination, some independent, some convergent, that lead to the birth and death of sex chromosomes in various lineages. Here we focus on turtles, an emerging model group with growing genomic resources. We review karyotypic changes that accompanied the evolution of chromosomal systems of genotypic sex determination (GSD) in chelonians from systems under the control of environmental temperature (TSD). These transitions gave rise to 31 GSD species identified thus far (out of 101 turtles with known sex determination), 27 with a characterized sex chromosome system (13 of those karyotypically). These sex chromosomes are varied in terms of the ancestral autosome they co-opted and thus in their homology, as well as in their size (some are macro-, some are micro-chromosomes), heterogamety (some are XX/XY, some ZZ/ZW), dimorphism (some are virtually homomorphic, some heteromorphic with larger-X, larger W, or smaller-Y), age (the oldest system could be ~195 My old and the youngest This article is published as Bista, Basanta, and Nicole Valenzuela. "Turtle Insights into the Evolution of the Reptilian Karyotype and the Genomic Architecture of Sex Determination." Genes 11, no. 4 (2020): 416. doi: 10.3390/genes11040416.</p

Karyotypic Evolution of Sauropsid Vertebrates Illuminated by Optical and Physical Mapping of the Painted Turtle and Slider Turtle Genomes

Recent sequencing and software enhancements have advanced our understanding of the evolution of genomic structure and function, especially addressing novel evolutionary biology questions. Yet fragmentary turtle genome assemblies remain a challenge to fully decipher the genetic architecture of adaptive evolution. Here, we use optical mapping to improve the contiguity of the painted turtle (Chrysemys picta) genome assembly and use de novo fluorescent in situ hybridization (FISH) of bacterial artificial chromosome (BAC) clones, BAC-FISH, to physically map the genomes of the painted and slider turtles (Trachemys scripta elegans). Optical mapping increased C. picta’s N50 by ~242% compared to the previous assembly. Physical mapping permitted anchoring ~45% of the genome assembly, spanning 5544 genes (including 20 genes related to the sex determination network of turtles and vertebrates). BAC-FISH data revealed assembly errors in C. picta and T. s. elegans assemblies, highlighting the importance of molecular cytogenetic data to complement bioinformatic approaches. We also compared C. picta’s anchored scaffolds to the genomes of other chelonians, chicken, lizards, and snake. Results revealed a mostly one-to-one correspondence between chromosomes of painted and slider turtles, and high homology among large syntenic blocks shared with other turtles and sauropsids. Yet, numerous chromosomal rearrangements were also evident across chelonians, between turtles and squamates, and between avian and non-avian reptiles.This aritcle is published as Lee, Ling S., Beatriz M. Navarro-Domínguez, Zhiqiang Wu, Eugenia E. Montiel, Daleen Badenhorst, Basanta Bista, Thea B. Gessler, and Nicole Valenzuela. 2020. "Karyotypic Evolution of Sauropsid Vertebrates Illuminated by Optical and Physical Mapping of the Painted Turtle and Slider Turtle Genomes" Genes 11, no. 8: 928. https://doi.org/10.3390/genes11080928. Posted with permission. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/)

Characterization of the First Turtle Organoids: A Model for Investigating Unique Adaptations with Biomedical Potential

Painted turtles are remarkable for their well-developed freeze tolerance and associated resilience to hypoxia/anoxia, oxidative stress, and ability to supercool. They are, therefore, an ideal model for biomedical research on hypoxia-induced injuries (including strokes), tissue cooling during extensive surgeries, and organ cryopreservation. Yet, the seasonal reproduction and slow maturation of turtles hinder basic and applied biomedical research. To overcome these limitations, we developed the first adult stem cell-derived turtle hepatic organoids, which provide 3D self-assembled structures that mimic their original tissue and allow for in vitro testing and experimentation without constantly harvesting donor tissue and screening offspring. Our pioneering work with turtles represents the first for this vertebrate Order and complements the only other organoid lines from non-avian reptiles, derived from snake venom glands. Here we report the isolation and characterization of hepatic organoids derived from painted, snapping, and spiny softshell turtles spanning ∼175 million years of evolution, with a subset being preserved in a biobank. Morphological and transcriptomics revealed organoid cells resembling cholangiocytes, which was then compared to the tissue of origin. Deriving turtle organoids from multiple species and life stages demonstrates that our techniques are broadly applicable to chelonians, permitting the development of functional genomic tools currently missing in most herpetological research. When combined with genetic editing, this platform will further support studies of genome-to-phenome mapping, gene function, genome architecture, and adaptive responses to climate change, among others. We discuss the unique abilities of turtles, including their overwintering potential, which has implications for ecological, evolutionary, and biomedical research.This is a pre-print of the article Zdyrski, Christopher, Vojtech Gabriel, Thea B. Gessler, Abigail Ralston, Itzel Sifuentes-Romero, Debosmita Kundu, Sydney Honold et al. "Characterization of the First Turtle Organoids: A Model for Investigating Unique Adaptations with Biomedical Potential." bioRxiv (2023): 2023-02. DOI: 10.1101/2023.02.20.527070. Attribution-NonCommercial-NoDerivatives 4.0 International (CC BY-NC-ND 4.0). Copyright 2023. The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. Posted with permission