Brief Summary of Minutes of Annual Meeting

Dr. Tang asked for permission to record the meeting and the present agreed.

Dr. Tang welcomed the group at 11:05 am.

Dr. Polejaeva requested to reverse the alphabetical order of the presentations (from Z to A for the present meeting) as it was discussed on the previous meeting. It was proposed to alternate the order of the presentations (A to Z and Z to A) in future meetings. All agreed.

Dr. Tang went over the minutes from the 2020 meeting. The minutes were approved.

Dr. Isom asked for a round of introductions and each of the participants at the time introduced her/himself.

Station reports started at 11:38 am. Dr. Polejaeva (Utah) had problems with the slide presentation, then Dr. Checura (South Carolina) started.

Dr. Jean M. Feugang (Mississippi), joined at 11:47 am and introduce himself.

Dr. Polajeava (Utah) presented her station report followed by Dr. Nascimento Moraes (Oklahoma), Dr. Schmidt (Montana), Dr. Geisert (Missouri), and Dr. Feugang (Mississippi).

There was a lunch break from 1:10 pm to 1:30 pm.

In the afternoon, the reports were presented as follows: Dr. Latham (Michigan), Dr. Keefer (Maryland), Dr. Bondioli (Louisiana), Dr. Youngs (Iowa), Dr. Wheeler (Illinois), Dr. Daigneault (Florida), Dr. Tian (Connecticut), Dr. Tang (Connecticut). Due to schedule conflicts, the alphabetical order of the presentation was altered for the rest of the afternoon. Dr. Looney (Arkansas) presented, followed by Dr. Berger (California).

At 3:42 pm there was an open floor for general discussion. Dr. Youngs commented on the USDA regulating GMO animals. The highlights for the subsequent discussion are:

• There is an open comment USDA website on the topic, and it is suggested that the members submit their scientific opinions as individuals.
• It is not appropriate to submit a statement to the USDA as W4171 group due to possible conflicts on the interpretation that the members may be representing their stations.

At 4:05 pm Dr. Denicol (California) presented her report.

At 4:19 pm Dr. Tang nominated Dr. Daigneault to serve as Secretary for the W4171 Technical Committee in 2022. Dr. Tian and Dr. White seconded the motion which was unanimously approved by the present members.  Dr. Daigneault accepted the position as Secretary, and Dr. Checura will serve as Chair for the W4171 Technical Committee in 2022 as agreed in the 2020 committee meeting.

The 2022 meeting for the W4171 group was discussed. It was tentatively agreed to be held concurrent with the 48th annual conference of the IETS. The exact meeting format, location and time are pending, and it will be determined based on the format and dates of the annual and satellite IETS meetings.

At 4:32 pm Dr. Wheeler opened the general discussion again, but on the topic of collaborations. A brief discussion ensued with several members offering collaborative efforts to the group.

At 4:50 pm Dr. White made closing remarks and Dr. Youngs moved the motion to adjourn the meeting. The motion was accepted and the meeting adjourned.

Respectively Submitted,

Celina M. Checura - Secretary

 W4171 Technical Committee

ACCOMPLISHMENTS:

OBJECTIVE 1
Understand the biology of gamete development, fertilization and embryogenesis including the underlying cellular and molecular mechanisms.

1.   A series of studies over several breeding seasons at multiple sites have been conducted to optimize the ability to fixed-time AI using a 6 day CO-Synch protocol using pre-synchronization. In all replications, the CIDR pre-synch has improved FTAI conception rates, in cows and heifers.  Typically, the cows with the earliest post-partum intervals have the most improved conception

2.   a.   Determined early changes in gene expression during enhanced Sertoli cell proliferation stimulated by reduced endogenous estradiol.

      b.   Identified potential receptors and ligands functioning during porcine fertilization.

      c.   Successful derivation of bovine ESC from early blastocysts.

3.   Achieved the editing of the bovine ESX1 gene in somatic cells.

4.   Provided a simple and efficient protocol for extraction of DNA from bovine with high sensitivity conducive for downstream genomic and epigenomic analyses.

5.   a. Oocyte Maturation: ethanol addition augmented the follicular and oocyte degeneration rates but increased the estradiol production and the meiotic resumption. Furthermore, the follicular metabolomic profile was similar between ethanol and rbFSH treatments but different from the control treatment in sheep.

      b.   Fertilization: dry mass is a valuable viability marker. A direct correlation between the presence of acrosome and the cell dry mass was observed, which can be used to evaluate the acrosome reaction. The three methods (Swim-Up, Gradient centrifugation and electro-channels) used to select viable spermatozoa for in vitro fertilization (IVF) result in very different sperm sub-populations; the dry mass of the sperm head is heavier in the spermatozoa selected with the electro-channel than the sperm selected by the other methods. These results show the potential of SLIM microscopy in reproductive biology. The tradeoff between cell viability and specificity can be circumvented by combining high-sensitivity phase imaging with deep learning. We trained a deep-convolutional neural network to perform semantic segmentation on quantitative phase maps. This approach, a form of phase imaging with computational specificity (PICS), allowed us to efficiently analyze thousands of sperm cells and identify correlations between dry-mass content and artificial-reproduction outcomes. Specifically, we found that the dry-mass content ratios between the head, midpiece, and tail of the cells can predict the percentages of success for zygote cleavage and embryo blastocyst formation.

c.     Embryo Culture: The power of the GLIM microscope’s ability to perform 3D reconstruction and section of the image is similar to a CT-scan for evaluation of bovine embryos. We have also discovered an interesting “turbulence” (particle motion characterized by chaotic changes in pressure or flow velocity) within the embryo.  This phenomenon is interesting because it was more prominent in compromised embryos and almost non-existent in developing embryos.

6.   a. In vivo and in vitro heat stress was shown not to alter global levels of DNA methylation or DNA hydroxymethylation.

      b.   The microtubule stabilizer taxol was shown to have some positive affect on normal chromosome distribution following vitrification of bovine oocytes but no positive effect on normal spindle configuration.

      c.   The alternative microtubule stabilizer Epothilone B was detrimental for both spindle configuration and chromosome distribution after vitrification of bovine oocytes.

      d.   An ultra-low-input ribosome profiling protocol was developed to systematically analyse both polysome- and nonpolysome-bound mRNA profiles of in vitro produced bovine blastocysts.

      e.   Single-cell whole genome bisulfite sequencing (scWGBS) was performed to map genome-wide DNA methylation dynamics of three classes of human trophoblast cells (TB) (cytoTB, syncytioTB, and migratory TB) during human implantation using an extended embryo culture system.

      f.    Single cell RNA sequencing was used to characterize developmental and transcriptomic differences between in vitro embryos produced in standard culture medium (IVC) with those produced using a culture medium with 75% reduced nutrients (carbohydrates and amino acids; IVRN).

7.   The application of 13C MFA to preimplantation bovine embryos presents unique challenges owing to the necessity to assess the metabolism of individual embryos (small sample size) and the complex medium. Novel labeling experiments had been designed to surmount these challenges and obtain a metabolic flux map for this complex system. This year (2020) was primarily focused on improving the mathematical model for analysis of flux networks and collecting samples according to the previously established design (varying substrates with varying labelled carbon atoms).  These experiments have been designed to probe the relative utilization of glycolysis and the pentose phosphate pathway, as well as the contributions of different carbon sources to metabolites originating at the pyruvate node.

8.   a.   RNAseq in oocytes of multiple species: RNA sequencing data were generated for mouse, bovine, and rhesus GV and MII stage oocytes.  Previously published and novel oocyte RNAseq from multiple species, including cow, are being used to compare transcriptome changes across species in a novel meta-analysis strategy seeking to identify conserved mechanisms driving oocyte maturation. Associated pathways and processes are being compared.

      b.   SMCHD1 as a maternal transcription regulator: Maternally expressed SMCHD1 plays an important role in terminating the first wave of embryonic genome activation. Additionally, transient zygotic knockdown of SMCHD1 expression negatively impacts later development, further attesting to the importance of maternally expressed and zygotically expressed SMCHD1 in correctly regulating preimplantation gene expression in a manner consistent with high developmental potency.  Ongoing studies are assessing the maturation, quality, viability, and developmental competence of oocytes that lack SMCHD1 function.

      c.   Effects of dairy cow nutrition on oocyte quality: Dairy cattle that either lose or maintain/gain body condition during lactations were compared.  Changes in serum lipid profiles were determined and oocyte transcriptomes were examined by RNA sequencing. Data are being analyzed to assess nature and mechanisms of effects.

9.  a.   Numerous up-regulated proteins in small follicles were directed to the healthy growth of both follicle and oocyte, while those in large follicles were involved in the ovulatory process. These proteins can be investigated as molecular markers of developmental competence and in vitro follicle culture. Several other proteins with potential roles during fertilization are also reported in the follicular fluid.

      b.   It was shown that balanced combinations of insulin and thyroxine concentrations are beneficial to follicle survival (low doses) and development (high doses).

      c.   The use of the follicle wall biopsy technique revealed the possibility of a real-time and repeated sampling of ovarian follicle cells and fluid of living mares, for effective evaluation of physiological processes occurring within the follicle. It was found that LH receptors in at least some of the layers of the follicle wall seems to trigger dominant follicles toward the anovulation process during the spring and fall transitional seasons.

     d.    The label-free microscopy imaging, SLIM, was found to be more informative than the AFM for sperm dimension analyses. There were interesting correlations between different compartments and dry mass of both boar and stallion spermatozoa; and positive (boar) and negative (stallion) correlations between the sperm head dry mass and the length and width of the acrosome were found in both species.

10.  Utilization of CRISPR/Cas9 gene editing has allowed knockouts of specific conceptus genes proposed to be essential in early development and establishment of pregnancy.  Research has provided information on the role of conceptus estrogen, prostaglandins, interleukin 1B2, and interferon g in maternal conceptus elongation, recognition of pregnancy, placental attachment and development during early pregnancy.  Results indicate that conceptus expression of interleukin 1B2 is essential to rapid elongation of the pig conceptus on Day 12 of pregnancy.  It would appear that the early establishment of pregnancy does not necessarily need estrogen at least to Day 24.  Establishment of pregnancy in the pig may involve both conceptus secretion of estrogen and prostaglandin E.  The role of interferon g has not been clearly established but our current data indicate conceptus production is essential for maintenance of pregnancy.

11. A simple and practical system to deliver a defined photostimulation (wavelength and intensity) has been designed.

12. FLI cytokine supplemented maturation medium improved bovine oocyte maturation, blastocyst development and initial pregnancy rate following somatic cell nuclear transfer. We are monitoring pregnancies to determine the full-term development success.

OBJECTIVE 2
Refine methods to produce animals by genetic engineering or genome editing for the improvement of livestock production efficiency and development of human biomedical models.

1.      A series of studies over several breeding seasons at multiple sites have been conducted to optimize the ability to fixed-time AI using a 6 day CO-Synch protocol using pre-synchronization. In all replications, the CIDR pre-synch has improved FTAI conception rates, in cows and heifers.  Typically, the cows with the earliest post-partum intervals have the most improved conception rates along with the heifers that are prepuberal with low Reproductive Tracts Scores. 

2.       Electroporation of CRISPR/Cas9 components into pig zygotes was used to circumvent the drawbacks of microinjection. To test viability of embryos/fetuses beyond the blastocyst stage, 2-4 cell gene deleted embryos derived from in vitro matured oocytes were transferred to oviducts of market weight gilts.  Comparison of guides used for gene deletion in fetuses recovered at 24 days was evaluated to consider if more efficient and less efficient guide sequences could be evaluated with an apparent difference noted among guides appearing equivalent in development to blastocysts. Skewed gender suggested an earlier effect of AR on gender than previously detected.

3.         The transcriptome profiles of the biPSCs were studied, and the successful reprogramming status of these biPSC lines was confirmed. The history and up-to-date progress for the studies on iPSC generation and characterization for all the major farm animals were summarized and published.

4.   a.   Transgenic Animals: There is no horizontal transmission of the transgene between transgenic and non-transgenic pigs that occurs during rearing, mating, gestation, or lactation and offer a foundation for the safe integration of transgenic animals into non-transgenic commercial production systems.

      b.   Stem Cells: The addition of both platelet-rich plasma and fibrin scaffolds to autologous ASCs from liposuction improved bone healing of critical-size defects. Osteocalcin production is higher when vitamin D3, vitamin B12, and vitamin K were added to the differentiation medium compared with the standard osteogenic medium, but only 100 nM vitamin D3 treatment had a positive influence on ASC differentiation into osteoblasts. A mineralized collagen-amnion scaffold may provide a beneficial environment to aid craniomaxillofacial bone repair, especially in the course of defects presenting significant inflammatory complications.

      c.   Others: There is potential for ultra-fast product design, engineering, and testing of medical devices needed for COVID-19 emergency response. A low-cost, easy-to-produce electronic sensor and alarm system for pressure-cycled ventilators have been developed. It estimates clinically useful metrics such as pressure and respiratory rate and sounds an alarm when the ventilator malfunctions during COVID-19 intubation. A point-of-care system integrated with a smartphone for detecting live virus from nasal swab media, using a panel of equine respiratory infectious diseases as a model system for corresponding human diseases such as COVID-19, has been demonstrated. Pathogen-spiked horse nasal swab samples were correctly diagnosed using the system, with a limit of detection comparable to that of the traditional lab-based test, polymerase chain reaction, with results achieved in ∼30 minutes.

5.   The ability to obtain pregnancies and live births from transfer of fresh alpaca demi-embryos to recipients was demonstrated. 

6.   a. It was shown that pretreatment of capacitated sperm with progesterone increased fertilization after intra cytoplasmic sperm injection of bovine oocytes.

      b.   It was shown that while there was no effect on mitochondrial number, ATP production was significantly reduced as a result of vitrification and warming of bovine oocytes.  Vitrification at 24h of maturation followed by extended culture to complete maturation after warming did not improve ATP production compared to vitrification at 44h of maturation.  

      c.   An enhancing cell injection system was developed by real time confirmation of membrane penetration and embryo viability.

7.   Liver-specific genetic models have uncovered a previously unrecognized metabolic system that can help to maintain liver homeostasis under conditions of oxidative or toxic stress. The system discovered uses catabolism of the essential sulfur amino acid methionine (Met) to generate cytosolic disulfide reducing power in the liver. This is the only NADPH-independent source of cytosolic disulfide reducing power that has, as yet, been characterized.

8.   A database of Doppler measurements for normal pregnancies under different conditions (different sires; singleton vs. twins, maternal age, etc) is being developed.

9.   a. An efficient protocol for the introduction of human specific mutations into the sheep fetal fibroblast cells using CRISPR/Cas9 RNP approach has been developed.

      b.   The F508del and G542X Cystic Fibrosis sheep models were successfully produced.

      c.   Conditions for CF mutation correction in sheep fibroblast cells and ovine-bovine iSCNT preimplantation embryos in vitro were optimized.

      d.   Preliminary results on using M3814 (a DNA-dependent protein kinase inhibitor) to increase efficiency of HDR look promising.

      e.   Donor-derived spermatogenesis was sustained in NANOS2-null bucks following exogenous spermatogonial stem cell transplantation

PUBLICATIONS:

1. Refereed articles/book chapters

• Jaques, J. & Cardoso, Rodolfo & Chachere, J. & Sinha, S. & Seery, R. & Wilkes, A. & Forrest, David & Looney, Charles. (2020). Using Anti-Müllerian Hormone (AMH) as a Predictor of Ova Production for Bovine Embryo Transfer. Advances in Reproductive Sciences. 08. 36-47. 10.4236/arsci.2020.81004. v

• Berger, T Nitta-Oda, BJ. 2020. Sertoli cell proliferation in juvenile boars and microRNA. Livestock Science, 233(March): 103954.
• Berger, T, Tang, S, Tu, L, Soto, D, Conley, A, Nitta-Oda, B. 2020. Changes in testicular gene expression following reduced estradiol synthesis: a complex pathway to increased porcine Sertoli cell proliferation. Molecular and Cellular Endocrinology.
• Candelaria JI, Rabaglino MB, Denicol AC. Ovarian preantral follicles are responsive to FSH as early as the primary stage of development. J Endocrinol. 2020; JOE-20-0126.R2. doi:10.1530/JOE-20-0126.
• The Induced Pluripotent Stem Cells from Farm Animals. Su, Y., Zhu, J., Salman, S., & Tang, Y*. (2020). Journal of animal science. https://doi.org/10.1093/jas/skaa34
• Daigneault, B. W. 2020. Dynamics of paternal contributions to early embryo development in large animals. Biol Reprod http://dx.doi.org/doi:10.1093/biolre/ioaa182
• Daigneault, B. W., S. K. Rajput, and G. W. Smith. 2020. Simple workflow for genome and methylation analyses of ejaculated bovine spermatozoa with low sperm input. Biotechniques 68: 155-158.http://dx.doi.org/doi:10.2144/btn-2019-0121
• Rubessa, M., Kandel, M.E., Schreiber, S., Meyers, S., Beck, D.H., Popescu, G., and
• Wheeler, M.B. 2020. Sperm selection methods for bovine in vitro embryo production, characterized by spatial light interference microscopy. Systems Biology in Reproductive Medicine. https://doi.org/10.1080/19396368.2019.1701139
• Rubessa, M., Wheeler, M.B. (2020) Label-free microscopy: a non-invasive new
• tool to assess gametes and embryo quality. Theriogenology Volume 150, 1 July 2020, Pages 241-246. https://doi.org/10.1016/j.theriogenology.2020.01.065
• Dewey, M. J., E. M. Johnson, S. T. Slater, D. J. Milner, M. B. Wheeler and B. A. C. Harley (2020). Mineralized collagen scaffolds fabricated with amniotic membrane matrix increase osteogenesis under inflammatory conditions. Regenerative Biomaterials. Online 4/8/20, doi.org/10.1093/rb/rbaa005
• William P. King, Jennifer Amos, Magdi Azer, Daniel Baker, Rashid Bashir, Catherine Best, Eliot Bethke, Steve Boppart, Elizabeth Bralts, Rachael Deitkus, Gary Durak, Stefan Elbel, Greg Elliott, Jake Fava, Nigel Goldenfeld, Mitch Goodman, Courtney Hayes, Nicole Herndon, Shandra Jamison, Harley Johnson, Mark Johnson, John Kolaczynski, Tonghun Lee, Sergei Maslov, Davis McGregor, Derek Milner, Jonathan Mosley, Ujjal Mikherjee, Andy Musser, Max Newberger, Lucas O’Bryan, Jerry O’Leary, Alex Pagano, Michael Philpott, Brian Pianfetti, Alex Pille, Marcello Rubessa, Sam Rylowicz, Stephanie Selting, Sridar Seshadr, Cliff Shipley, Brian Stewart, Rachel Switzky, Sameh Tawfick, Matthew B. Wheeler, Karen White, Sarah Womack, Eric Wood, Charles Wood, Abigail Wooldridge. (2020). Emergency Ventilator for COVID-19. PLoS ONE 15(12): e0244963. https://doi.org/10.1371/journal.pone.0244963
• Kandel, M., Y. He, M. Rubessa, M. Wheeler and G. Popescu (2020). High sensitivity SLIM imaging and deep learning to correlate sperm morphology and fertility outcomes (Conference Presentation), SPIE BIOS, vol. 11249, Quantitative Phase Imaging VI. https://doi.org/10.1117/12.2550470
• Sun, F., Ganguli, A., Nguyen, J., Brisbin, R., Shanmugam, K., Hirschberg, D. L., Wheeler, M. B., Bashir, R., Nash, D. M., Cunningham, B. T. (2020).  Smartphone-Based Multiplex 30-minute Nucleic Acid Test of Live Virus from Nasal Swab Extract. Lab Chip 20, 1621-1627 DOI: 10.1039/D0LC00304B.
• Bane T.A., Bertels J.C., Polkoff K.M., Rubessa M. and Wheeler M.B. (2020). The Effects of Vitamin D3, Vitamin B12, and Vitamin K on the Differentiation of Porcine Adipose-Derived Stem Cells (ASC) into Osteoblasts. Journal of Biology Engineering and Medicine Volume 2(1): 1-5 doi:10.31487/j.JBEM.2020.01.01
• Rubessa, M., Feugang, J.M., Kandel, M.E., Schreiber, S., Hessee, J., Salerno, F., Meyers, S., Chu, I., Popescu, G., Wheeler, M.B. (2020). High-throughput sperm assay using label free microscopy: morphometric comparison between different sperm structures in boar and stallion spermatozoa. Animal Reproduction Science 219, August 2020, 106509, online June 16, 2020, https://doi.org/10.1016/j.anireprosci.2020.10650
• Kandel, M.E., Rubessa, M., He, Y.R., Schreiber, S., Meyers, S., Naves, L.M., Sermersheim, M.K., Sell, G.S., Szewczyk, M.J., Wheeler, M.B., Popescu, G. (2020). Deep convolutional neural networks reveal slight differences in spermatozoon ultra-structure correlating with reproductive outcomes. Proceedings of the National Academy of Sciences (P.N.A.S.) 117 (31) 18302-18309; DOI:10.1073/pnas.2001754117
• Mosley, J.F., Hurley, W.L., Rodriguez-Zas, S., Wheeler, M.B. (2020). Evaluation of risks from environmental contact with transgenic livestock. J Vet Med Res 7(3): 1190. 9 pages. https://www.jscimedcentral.com/VeterinaryMedicine/veterinarymedicine-7-1190.pdf
• Maki, A.J., Rabel, R.A.C., Rubessa, M., Ercolin, A.C., Roballo, K.S., Cooper, J.J., Wheeler, M.B. (2020). Autologous Adipose-Derived Stem Cells, Platelet-rich Plasma, and Fibrin Enhance Healing of Mandibular Bone Defects in Swine. International Journal of Regenerative Medicine Volume 3(2): 2-9. DOI: 10.31487/j.RGM.2020.02.01
• Corey, R.M., Widloski, E.M., Null, D., Ricconi, B., Johnson, M.A., White, K.C.,

Amos, J.R., Pagano, A.,  Oelze, M.L., Switzky, R.D., Wheeler, M.B., Bethke, E.,  Shipley, C.F.,  Singer, A.C. (2020). Low-Complexity System and Algorithm for an Emergency Ventilator Sensor and Alarm. IEEE Transactions on Biomedical Circuits and Systems, doi: 10.1109/TBCAS.2020.3020702.

• Renato Félix da Silva, Laritza Lima, Rebeca Rocha, Ivina Brito, Gerlane Silva, Hudson Correia, Giovanna Rodrigues, Anna Clara Ferreira, Diana Nunes-Pinheiro, Arlindo Moura, Larissa Silveira, Edson G. Lo Turco, Matthew B. Wheeler, Ana Paula Rodrigues, Claudio Campello. (2020). In vitro long-term culture of isolated ovine preantral follicle: influence of ethanol on steroid production, oocyte meiotic resumption and metabolomic profile. Research in Veterinary Science, accepted 30 October 2020, In Press, doi.org/10.1016/j.rvsc.2020.10.029
• Benaissa MH, Y Bentria, N Mimoune, T Kernif, A Boukhelkhal, CR Youngs, R Kaidi, B Faye, and Y Halis. Seroprevalence and risk factors for Trypanosoma evansi, the causative agent of surra, in the dromedary camel (Camelus dromedarius) population in southeastern Algeria. Onderstepoort Journal of Veterinary Research 87(1), a1891. https://doi.org/10.4102/ojvr.v87i1.1891
• Benaissa MH, N Mimoune, CR Youngs, R Kaidi, and B Faye. 2020. First report of Chlamydophila abortus infection in the dromedary camel (Camelus dromedarius) population in eastern Algeria. Comparative Immunology, Microbiology and Infectious Diseases 73:101557,  https://doi.org/10.1016/j.cimid.2020.101557
• Lutz JC, SL Johnson, KJ Duprey, PJ Taylor, HW Vivanco, DP Salazar, MM Gonzales, and CR Youngs. 2020. Birth of a live cria after transfer of a vitrified-warmed alpaca (Vicugna pacos) preimplantation embryo. Frontiers in Veterinary Science 7:581877, https://doi.org/10.3389/fvets.2020.581877
• Esencan E, Jiang Z, Wang T, Zhang M, Soylemez-Imamoglu G, Seli E. Impaired mitochondrial stress response due to CLPP deletion is associated with altered mitochondrial dynamics and increased apoptosis in cumulus cells. Reproduction Sciences. 2020 Jan 14. PMID: 31939198.
• Li Y, Sun J, Ling Y, Hao M, Chen Z, Fang F, Liu Y, Cao H, Ding J, Cao Z, Zhang X, Bondioli K, Jiang Z, Zhang Y. Transcriptional profiles of oocytes during maturation and embryos during preimplantation development in vivo in the goat. Reprod Fertil Dev. 2020 Apr;32(7):714-725. PMID: 32317096.
• Li Z, Zhao S, Nelakanti RV, Lin K, Wu TP, Alderman III MH, Guo C, Wang P, Zhang M, Wang M, Jiang Z, Wang Y, Li H, Xiao AZ. N6-methyladenine in DNA antagonizes SATB1 in early development. Nature 2020 Jul: 583, 625–630. PMID: 32669713.
• Ming H, Sun J, Pasquariello R, Gatenby L, Herrick J, Yuan Y, Pinto C, Bondioli KR, Krisher RL, Jiang Z. The landscape of accessible chromatin in bovine oocytes and early embryos. Epigenetics. 2020 Jul 14. PMID:32663104.
• Smith R, Jiang Z, Susor A, Ming Hao, Tait J, Conti M, Lin CJ. The H3.3 chaperone Hira complex orchestrates oocyte developmental competence. bioRxiv. 2020 May 26, doi: https://doi.org/10.1101/2020.05.25.114124.
• Chung J, Sriram G, Keefer CL.Nanoparticle technology improves in-vitro attachment of cattle (Bos taurus) trophectoderm cells. Biotechnol Lett. 2020 Nov;42(11):2083-2089.

doi: 10.1007/s10529-020-02926-w. Epub 2020 Jun 3.

• Paes, V. M., de Figueiredo, J. R., Ryan, P. L., Willard, S. T., Feugang, J. M. (2020). Comparative analysis of porcine follicular fluid proteomes of small and large ovarian follicles. Biology, 9(5). https://api.elsevier.com/content/abstract/scopus_id/85085521478
• Paes, V. M., Lima, L. F., Ferreira, A. C.A., Lobo, C. H., Alves, B. G., Rodrigues, A. P.R., Oliveira, A. C., Figueiredo, J. R., Feugang, J. M. (2020). The subtle balance of insulin and thyroxine on survival and development of in vitro cultured caprine preantral follicles enclosed in ovarian tissue. Theriogenology, 147, 10-17.

https://api.elsevier.com/content/abstract/scopus_id/85079386717

• Ishak, G. M., Dutra, G. A., Gastal, G. D.A., Elcombe, M. E., Gastal, M. O., Park, S. B., Feugang, J. M., Gastal, E. L. (2020). Deficiency in proliferative, angiogenic, and LH receptors in the follicle wall: implications of season toward the anovulatory condition. Domestic Animal Endocrinology, 70.

https://api.elsevier.com/content/abstract/scopus_id/85072712691

• Rubessa, M., Feugang, J. M., Kandel, M. E., Schreiber, S., Hessee, J., Salerno, F., Meyers, S., Chu, I., Popescu, G., Wheeler, M. B. (2020). High-throughput sperm assay using label-free microscopy: morphometric comparison between different sperm structures of boar and stallion spermatozoa. Animal Reproduction Science, 219. https://api.elsevier.com/content/abstract/scopus_id/85086603197
• Geisert R. D., and T. E. Spencer. 2021. Chapter 1: Introduction. In: Placentation in Mammals: Tribute to EC Amoroso’s contributions to Viviparity. Advances in Anatomy, Embryology and Cell Biology, Springer. (In Press).
• Geisert R. D., A. E. Meyer, C. A. Pfeiffer, D. N. Johns, K. Lee, K. D. Wells, T. E. Spencer, and R. S. Prather. 2021. Gene Editing to Investigate Role of Conceptus Factors in Establishment of Pregnancy. Reproduction (In Press).
• Pfeiffer C. A., A. E. Meyer, K. E. Brooks, P. R. Chen, J. Milano-Foster, L. D. Spate, J. A. Benne, R. Cecil, M. S. Samuel, L. A. Ciernia, M. F. Smith, K. D. Wells, T. E. Spencer, R. S. Prather, and R. D. Geisert. 2020. Ablation of conceptus PTGS2 provides a new understanding of early pregnancy events in the pig. Biol Reprod 102: 475-488. doi.org/10.1093/biolre/ioz192.
• Miller CG, Schmidt EE. Sulfur Metabolism Under Stress. Antioxid Redox Signal. 2020 Dec 1;33(16):1158-1173. doi: 10.1089/ars.2020.8151. Epub 2020 Aug 14. PubMed PMID: 32799544; PubMed Central PMCID: PMC7699002.
• Dóka É, Ida T, Dagnell M, Abiko Y, Luong NC, Balog N, Takata T, Espinosa B, Nishimura A, Cheng Q, Funato Y, Miki H, Fukuto JM, Prigge JR, Schmidt EE, Arnér ESJ, Kumagai Y, Akaike T, Nagy P. Control of protein function through oxidation and reduction of persulfidated states. Sci Adv. 2020 Jan;6(1):eaax8358. doi: 10.1126/sciadv.aax8358. eCollection 2020 Jan. PubMed PMID: 31911946; PubMed Central PMCID: PMC6938701.
• Shearn CT, Fennimore B, Orlicky DJ, Gao YR, Saba LM, Battista KD, Aivazidis S, Assiri M, Harris PS, Michel C, Merrill GF, Schmidt EE, Colgan SP, Petersen DR. Cholestatic liver disease results increased production of reactive aldehydes and an atypical periportal hepatic antioxidant response. Free Radic Biol Med. 2019 Nov 1;143:101-114. doi: 10.1016/j.freeradbiomed.2019.07.036. Epub 2019 Aug 1. PubMed PMID: 31377417; PubMed Central PMCID: PMC6848778.
• Ciccarelli M., Giassetti M.I., Miao D., Oatley M.J., Robbins C., Lopez-Biladeau B., Waqas M.S., Tibary A., Whitelaw B., Lillico S., Park C.H., Park K.E., Telugu B., Fan Z., Liu Y., Regouski M., Polejaeva I.A. & Oatley J.M. (2020) Donor-derived spermatogenesis following stem cell transplantation in sterile NANOS2 knockout males. Proc Natl Acad Sci U S A 117, 24195-204.
• Perisse I.V., Fan Z., Singina,G.N., White, K.L., and Polejaeva I.A.* (2020) Improvements in Gene Editing Technology Boosts Its Applications in Livestock. Frontiers in Genetics 11,

2. Books, non-refereed book chapters, proceedings, instructional media, theses/dissertations

• Noga, N., C. Looney. 2020. Embryo Collection and Embryo Transfer. In: Bovine Reproduction. Hopper, R. M. John Wiley & Sons.
• Berger, T. Juvenile boars and testicular microRNA. 2020. Mendeley Data.
• Berger, T Nitta-Oda, B. 2020. Alteration in porcine testicular gene expression in response to reduced testicular estradiol synthesis. NCBI Gene Expression Omnibus.
• Placentation in Mammals: Tribute to EC Amoroso’s contributions to Viviparity. 2021. R. D. Geisert and T. Spencer. Advances in Anatomy, Embryology and Cell Biology, Springer. (In Press).

3. Abstracts

• Oosthuizen, N., Fontes, P., Filho, R., Dahlen, C.R., Grieger, D.M., Hall, J.B., Lake, S., Looney, C., Mercadante, V.R., Neville, B.W. and Perry, G., 2020. 147
• Presynchronization and delayed fixed-time artificial insemination increases pregnancy rates with sex-sorted semen in replacement beef heifers. Journal of Animal Science, 98(Supplement_4), pp.115-116.
• Vazquez, J. Berger, T. 2020. Unveiling the proteome of sperm-oocyte plasma membrane interaction using a porcine model. Society for the Study of Reproduction 2020 Annual Meeting.
• Zacanti, K, Park, I, Rowe, JD, McNabb, BR, Ross, PJ, Maga, EA, Berger, TJ. 2020. Pig androgen receptor knockout fetuses via CRISPR/ Cas9 technology. Society for the Study of Reproduction 2020 Annual Meeting
• Schefer, L., Naves, L. M., Ramires Ferreira, C., Pradebon da Silva, E., Sobreira, T. J., Vieiro Marchioretto, P., Lima Verde Leal, C., Rubessa, M., and Wheeler, M. B. (2020). Lipid profiling of bovine blastocysts produced in vitro with and without a stimulator of cyclic guanosine monophosphate synthesis by multiple reaction monitoring profiling. L. Reproduction, Fertility and Development.  32(1): 158-159.
• Schefer, L., Naves, L. M., Ramires Ferreira, C., Pradebon da Silva, E., Sobreira, T. J., Vieiro Marchioretto, P., Lima Verde Leal, C., Rubessa, M., and Wheeler, M. B. (2020). Lipid profiling of bovine oocytes matured with different stimulation of cyclic GMP synthesis by multiple reaction monitoring profiling. Reproduction, Fertility and Development.  32(1): 226.
• Sermersheim, M.K., Lindsey, B. R., Naves, L. M., Rubessa, M., and Wheeler, M. B. (2020). Relationship between corpus luteum blood flow evaluated via colour Doppler ultrasound and pregnancy rate in bovine embryo transfer recipients. Reproduction, Fertility and Development. 32(1): 176.
• Pasqual, B. M., Correa, E. E., da Silva, E. P., Sermersheim, M. K., Naves, L. M., Sell, G. S., Lindsey, B.D., Rubessa, M., and Wheeler, M.B. (2020). Relationship between oestrus manifestation and pregnancy rates on recipients used in an in vitro-produced embryo transfer program. Reproduction, Fertility and Development. 32(1): 128-129.
• Lutz JC, SL Johnson, KJ Dupree, PJ Taylor, HW Vivanco-Mackie, MD Ponce-Salazar, M Miguel-Gonzales, and CR Youngs. 2020. Pregnancy from a vitrified-warmed alpaca preimplantation embryo. Reprod. Fertil. Dev. 32(2):128. https://doi.org/10.1071/RDv32n2Ab7
• Diaz, FA, Gutierrex, EJ, Foster, BA, Hardin PT and KR Bondioli. Effect of in vivo and in vitro heat stress on DNA methylation and DNA hydrooxymethylation of bovine oocytes and embryos. Reprod. Fertil. Dev. January 2021.
• Gatenby, L and KR Bondioli. Progesterone induced acrosome reaction and fertilization rates with bovine intracytoplasmic sperm injection.  Fertil. Dev.  January 2021
• Girka, E and KR Bondioli. Effect of taxol and epothilone B on meiotic spindle stabilization in vitrified bovine ooctyes.  Fertil. Dev.  January 2021.
• Goheen B, Rajput S, Logsdon DM, Yuan Y, Schoolcraft WB, Jiang Z, and Krisher RL. A reduced nutrient culture environment improves bovine blastocyst development and alters mitochondrial metabolism as elucidated by single cell RNA sequencing. Fertility Sterility. 2020, P-566.
• Gutierrez, EJ, Diaz, FA and KR Bondioli. Effect of vitrification of porcine oocytes on ATP production and mitochondrial copy number. Reprod. Fertil. Dev. January 2021.
• Logsdon DM, Ming H, Sun J, R. Schoolcraft WB, Krisher RL, Jiang Z, Yuan Y. Single cell multi-omics of peri-implantation stage human embryos reveals similarities between early trophoblast differentiation and neuronal behavior. 53th Annual conference of Society for the Study of Reproduction (SSR), 2020.
• Mor A, Gatenby L, Dzekunskas E, Zhu L, Bondioli KR, Jiang Z, and Seli E. Enhancing cell injection systems by real time confirmation of cytoplasmic penetration. Fertility Sterility. 2020, O-174.
• Zhu L, Ming H, Liu S, Iyyappan R, Liano ED, Dvoran M, Chen Q, Susor A, Zhou T, and Jiang Z. High-resolution ribosome profiling reveals translational selectivity in the mammalian blastocyst. Reprod, Fertil. Dev. 3 December 2020.
• Lugar D, Adhikari S, Keefer CL, and Sriram G. 13C Metabolic Flux Analysis of Pre-Implantation Bovine Embryos: Surmounting Challenges Due to Small Sample Size and Complex Media, American Chemical Society National Meeting, March 22-26, 2020
• Littlejohn, B. P., Mercer, K. B., Bowers, K. J., Messman, R. D., Contreras-Correa, Z. E., Reid, D., Feugang, J. M., Vann, R. C., Lemley, C. O. (2020). Influence of chronic melatonin supplementation on bovine testis physiology. Southern Section ASAS.
• D. Geisert, A. E. Meyer., C. A. Pfeiffer, D. N. Johns, T. E. Spencer, and R. S. Prather. 2020. Early Embryonic Loss is an Important Mechanism for Maximizing Litter Size in the Pig. Physiology and Endocrinology Symposium National ASAS Meeting, Madison, Wisconsin
• Johns D. N., C. G. Lucas, P. R. Chen, S. D. Perry, C. A. Pfeiffer, L. D. Spate, R. F. Cecil, K. M. Wells, T. E. Spencer, R. S. Prather, and R. D. Geisert. 2020. Porcine Conceptus Expression of Interferon Deltas and Gamma During Early Pregnancy. 53rd Annual Meeting of SSR.
• Johns D. N., C. G. Lucas, P. R. Chen, L. D. Spate, C. A. Pfeiffer, R. F. Cecil, M. A. Fudge, S. D. Perry, K. M. Whitworth, R. F. Cecil, K. M. Wells, T. E. Spencer, R. S. Prather, and R. D. Geisert. 2020. Conceptus Interferon Gamma is Essential for Pregnancy Maintenance in the Pig. The 17th Annual Gilbert S. Greenwald Symposium on Reproduction and Regenerative Medicine, Kansas City, MO.
• Gholson S. K., A. L. Schmelzle, J. A. Green, M. F. Smith and R. D. Geisert. 2020. Altering sexual behavior of rats through neonatal treatment with an aromatase inhibitor, estrogen receptor antagonist and testosterone. Midwest ASAS Meeting, March 2-4, 2020.
• Checura CM, Pratt SL, Campbell LV, Farmer K, Loughlin G, Mitchell M, Sanford A, Treske A, Malter H. 197 - The use of photostimulation to enhance oocyte cytoplasmic maturation. Reproduction, Fertility and Development. 2020;32(2):227.
• Checura CM, Atkinson K, Boulos N, Crowfoot E, Friend H, Jackson S, Lubowicki S, Silva C, Walker E. 124 - Utilization of Doppler Ultrasonography for the Study of Fetal Development in Sheep. FOCI meeting; Clemson University; 2020, 55.
• Goldberg M, Mitchell M, Pratt SL, Malter H, Checura CM. The Use of Photostimulation to Improve In Vitro Fertilization. HSC Showcase 2020; Health Sciences Center at Prisma Health; Greenville, SC, USA: 2020; 57.
• Fan Z., Regouski M., Liu Y., Keim J., Perisse I.V., Oatley J.M., Polejaeva I.A. (2020) Generation of NANOS2 knockout goats using CRISPR/Cas9 and SCNT techniques. Reproduction, Fertility and Development 32 (2) 193 - 194.
• Perisse I.V., Fan Z., Van Wettere A., Wang Z., Harris A., White K., Polejaeva I.A. (2020) Introduction of F508del human mutation into the CFTR gene of sheep fetal fibroblasts using CRISPR/Cas9 ribonucleoprotein. Reproduction, Fertility and Development 32 (2) 192 - 193.
• Singina G.N.*, Shedova E.N., Polejaeva I.A. and Taradajnic T.E. (2020) Effect of cytokines during in vitro maturation of bovine oocytes on the development potential of partheno-genetic embryos. Reproduction, Fertility and Development 32 (2) 222.
• Keim J., Zhang W., Liu Y., Rutigliano H., Zhou A., Polejaeva I.A. (2020) Analyzing metabolomic profile of bovine IVF and SCNT embryos through Raman spectroscopy. Reproduction, Fertility and Development 32 (2) 169 - 170.
• Bunch K., Perisse I.V., Fan Z., Wang Z., Harris A., White K.L., Polejaeva I.A. (2020) Construction of candidate vectors for correction of the intestinal CFTR gene expression in Cystic Fibrosis sheep fetal fibroblast cells. The 14th Utah Conference on Undergraduate Research, Utah State University, Logan, Utah, USA, February 7, 2020.

4. Miscellaneous publications (semi-technical/lay publications)

• Hahn, M., C. Looney. 2020. Precision Agriculture in Dairy. In: Arkansas Cattle Business.
• Noga, N., C. Looney. 2020. Defined Breeding Season. In: Arkansas Cattle Business.
• Looney, C., N. Noga. 2020. Ultrasound Scanning to Measure Body Composition in Beef Cattle. FSA 3106. University of Arkansas, Fayetteville, AR.
• Sep 14, 2020 “Surrogate sires' could create specially bred livestock” The Gardian.
• Sep 14, 2020 “Gene editing to produce ‘super dad’ livestock” BBC News.
• Sep 14, 2020 “Scientists create gene-edited animals as 'surrogate sires' to boost food production”
• Daily Mail.
• Sep 14, 2020 “Gene-edited livestock 'surrogate sires' successfully made fertile”
• Science Daily.
• Sep 14, 2020 “First gene-edited livestock created that can serve as ‘surrogate sires’” Irish examiner.
• Sep 15, 2020 “Scientists create ‘surrogate sire’ animals to pass on genetic traits of donor males.
• Yorkshire Post, UK.
• Sep 15, 2020 “GM ‘super livestock’ to meet food shortage” The Daily Telegraph.
• Sep 15, 2020 “Stem cell hopes of beefing up livestock” Daily Mail.
• Sep 15, 2020 “Gene-edited livestock could boost food production in growing world” The Scotsman.
• Sep 15, 2020 “CRISPR-Cas9 Used to Generate “Surrogate Sire” Livestock that Produce Only Donor Sperm” Genetic Engineering & Biotechnology News.
• Sep 16, 2020 “Gene-edited livestock surrogate sires promise to transform selective breeding”