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We investigate the recent history and stock identity of beluga whales (Delphinapterus leucas) in Kotzebue Sound in the Chukchi Sea, a region long frequented by large numbers of belugas in summer until their near disappearance in the 1980s. Wide variation in numbers since then suggests a complex recent history that hinders recovery efforts. Analysis of teeth sampled during the historical (pre-decline) era using ancient DNA (aDNA) methods found that the original Kotzebue Sound whales were differentiated for mitochondrial DNA (mtDNA) from other summering concentrations across the Pacific Arctic revealing a demographically distinct subpopulation where long-established migratory culture likely facilitated population divergence. Analysis of microsatellite (nDNA) and mtDNA markers in belugas from the contemporary (post-decline) era revealed that whales from other stocks likely visited Kotzebue Sound, including during two low ice years when relatively large numbers of whales were present. Some mtDNA lineages were found only in Kotzebue Sound, with one recorded in both the historical and contemporary eras. Exclusion tests found a number of whales in Kotzebue Sound during the contemporary era that had nDNA genotypes unlikely to arise in other contemporary stocks in the Pacific Arctic. Our findings indicate that the Kotzebue Sound belugas comprised a unique stock of which a few remnants likely still co-occur with belugas from other larger stocks. We recommend that the US government work through the co-management process to greatly reduce or eliminate the taking of belugas, especially adult females, likely to belong to the Kotzebue Sound stock, until they recover.

Gustavo Kattan fue un científico visionario que soñó con repetir expediciones ornitológicas en las localidades históricas del Museo Americano de Historia Natural en Colombia, para estudiar los cambios en las aves del país como respuesta a cambios ambientales y en el paisaje, y así contribuir a su conservación. Construyendo sobre el trabajo de Gustavo, el programa Re-expedición Colombia es ahora realidad, con una alianza de científicos, comunidades locales, instituciones colombianas e internacionales, que recopila información histórica y reciente sobre la avifauna del país, para entender los cambios ecológicos, evolutivos y de estado de conservación que han ocurrido durante un siglo de transformación. El carácter colaborativo del programa permite fortalecer y diversificar la información depositada en las colecciones biológicas y la capacidad de investigación en ornitología. Además, basado en los conocimientos de las comunidades locales, el programa apoya iniciativas de monitoreo participativo importantes como alternativas económicas sostenibles. La producción de mapas interactivos de aves, el desarrollo de una ruta de aviturismo para ser operada por proveedores comunitarios, junto a algunos esfuerzos regionales, aportarán a la sostenibilidad y contribuirán a garantizar la conservación y bienestar de las aves, los ecosistemas que éstas habitan, y las comunidades que las protegen.

Genome editing by homology directed repair (HDR) is leveraged to precisely modify the genome of therapeutically relevant hematopoietic stem and progenitor cells (HSPCs). Here, we present a new approach to increasing the frequency of HDR in human HSPCs by the delivery of an inhibitor of 53BP1 (named "i53") as a recombinant peptide. We show that the use of i53 peptide effectively increases the frequency of HDR-mediated genome editing at a variety of therapeutically relevant loci in HSPCs as well as other primary human cell types. We show that incorporating the use of i53 recombinant protein allows high frequencies of HDR while lowering the amounts of AAV6 needed by 8-fold. HDR edited HSPCs were capable of long-term and bi-lineage hematopoietic reconstitution in NSG mice, suggesting that i53 recombinant protein might be safely integrated into the standard CRISPR/AAV6-mediated genome editing protocol to gain greater numbers of edited cells for transplantation of clinically meaningful cell populations., (© 2024. The Author(s).)

Background: Cas12a (formerly known as Cpf1), the class II type V CRISPR nuclease, has been widely used for genome editing in mammalian cells and plants due to its distinct characteristics from Cas9. Despite being one of the most robust Cas12a nucleases, LbCas12a in general is less efficient than SpCas9 for genome editing in human cells, animals, and plants., Results: To improve the editing efficiency of LbCas12a, we conduct saturation mutagenesis in E. coli and identify 1977 positive point mutations of LbCas12a. We selectively assess the editing efficiency of 56 LbCas12a variants in human cells, identifying an optimal LbCas12a variant (RVQ: G146R/R182V/E795Q) with the most robust editing activity. We further test LbCas12a-RV, LbCas12a-RRV, and LbCas12a-RVQ in plants and find LbCas12a-RV has robust editing activity in rice and tomato protoplasts. Interestingly, LbCas12a-RRV, resulting from the stacking of RV and D156R, displays improved editing efficiency in stably transformed rice and poplar plants, leading to up to 100% editing efficiency in T 0 plants of both plant species. Moreover, this high-efficiency editing occurs even at the non-canonical TTV PAM sites., Conclusions: Our results demonstrate that LbCas12a-RVQ is a powerful tool for genome editing in human cells while LbCas12a-RRV confers robust genome editing in plants. Our study reveals the tremendous potential of these LbCas12a variants for advancing precision genome editing applications across a wide range of organisms., (© 2023. The Author(s).)

Purpose: This study determined fluid intake and physical activity behaviors among college students during the COVID-19 pandemic., Methods: College students (n = 1014; females, 75.6%) completed an online survey during the Spring 2020 academic semester following the initial global response to the COVID-19 pandemic. Academic standing, habitation situation, and University/College responses to COVID-19 were collected. Participants completed the Godin Leisure-Time Exercise Questionnaire and a 15-item Beverage Questionnaire (BEVQ-15) to determine physical activity level and fluid intake behaviors, respectively., Results: Females (1920 ± 960 mL) consumed significantly less fluid than males (2400 ± 1270 mL, p < 0.001). Living off-campus (p < 0.01) and living with a spouse/partner (p < 0.01) was associated with increased consumption of alcoholic beverages. 88.7% of participants reported being at least moderately active; however, Black/African American and Asian participants were more likely to be less active than their Caucasian/White counterparts (p < 0.05). Participants reporting no change in habitation in response to COVID-19 had a higher fluid intake (p = 0.002); however, the plain water consumption remained consistent (p = 0.116). While there was no effect of habitation or suspension of classes on physical activity levels (p > 0.05), greater self-reported physical activity was associated with greater fluid intake (std. β = 0.091, p = 0.003)., Conclusions: Fluid intake among college students during the initial response to the COVID-19 pandemic approximated current daily fluid intake recommendations. Associations between COVID-19-related disruptions (i.e., suspension of classes and changes in habitation) and increased alcohol intake are concerning and may suggest the need for the development of targeted strategies and programming to attenuate the execution of negative health-related behaviors in college students., (© 2022. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany.)

Genetically engineered animals can be produced quickly using genome editing technology. A new electroporation technique, technique for animal knockout system by electroporation (TAKE), aids in the production of genome-edited animals by introducing nucleases into intact embryos using electroporation instead of microinjection. It is difficult to confirm nuclease delivery into embryos after electroporation using the conventional TAKE method. We previously reported the successful visualization of fluorescently-labeled tracrRNA in embryos after electroporation Cas9 paired with the crRNA:tracrRNA-ATTO550 duplex. However, the amount of fluorescence signal from labeled tracrRNA in embryos did not correlate with the genome editing rate of the offspring. This study examined the visualization of Cas9 protein in embryos after electroporation and its correlation with the genome editing rate of the offspring using a fluorescent Cas9 fusion protein. The fluorescent Cas9 protein was observed in all embryos that survived following electroporation. We found that the efficiency of Cas9 protein delivery into embryos via electroporation depended on the pulse length. Furthermore, we demonstrated that the amount of fluorescent Cas9 protein detected in the embryos correlated with the genome editing efficiency of the embryos. These data indicate that the TAKE method using fluorescently-labeled nucleases can be used to optimize the delivery conditions and verify nuclease delivery into individual embryos prior to embryo transfer for the efficient production of genome-edited animals.

Though AsCas12a fills a crucial gap in the current genome editing toolbox, it exhibits relatively poor editing efficiency, restricting its overall utility. Here we isolate an engineered variant, "AsCas12a Ultra", that increased editing efficiency to nearly 100% at all sites examined in HSPCs, iPSCs, T cells, and NK cells. We show that AsCas12a Ultra maintains high on-target specificity thereby mitigating the risk for off-target editing and making it ideal for complex therapeutic genome editing applications. We achieved simultaneous targeting of three clinically relevant genes in T cells at >90% efficiency and demonstrated transgene knock-in efficiencies of up to 60%. We demonstrate site-specific knock-in of a CAR in NK cells, which afforded enhanced anti-tumor NK cell recognition, potentially enabling the next generation of allogeneic cell-based therapies in oncology. AsCas12a Ultra is an advanced CRISPR nuclease with significant advantages in basic research and in the production of gene edited cell medicines.

Drosophila Suppressor of Hairy-wing [Su(Hw)] is a multifunctional zinc finger DNA binding protein. Transcriptional regulation by Su(Hw) is essential in the ovary and testis, where Su(Hw) functions primarily as a repressor. Recently, the HP1a and Insulator Partner Protein 1 (HIPP1) was found to extensively co-localize with Su(Hw) and other insulator binding proteins in euchromatic regions of the genome, and with Heterochromatin Protein 1a (HP1a) in heterochromatic regions. As HIPP1 is the homolog of the human co-repressor Chromodomain Y-Like (CDYL), we tested its requirement in establishing transcriptional repression in flies. To this end, we generated multiple Hipp1 null alleles and a tagged derivative of the endogenous gene ( Hipp1 GFP ), using CRISPR mutagenesis. We show that HIPP1 is a widely expressed nuclear protein that is dispensable for viability, as well as female and male fertility. We find that HIPP1 and HP1a display minimum co-localization in interphase cells, and HP1a-dependent transcriptional repression of several reporter genes is HIPP1-independent, indicating that HIPP1 is not essential for HP1a-dependent heterochromatin formation. Despite Su(Hw) having a major role in promoting HIPP1 occupancy in euchromatin, we show that HIPP1 is dispensable for the transcriptional and insulator functions of Su(Hw), indicating that HIPP1 is not a critical Su(Hw) cofactor. Further studies are needed to clarify the role of HIPP1 in Drosophila development., (Copyright © 2019 Glenn, Geyer.)