A simple and efficient transfection protocol for Cryptosporidium parvum using Polyethylenimine (PEI) and Octaarginine

The transfection of Cryptosporidium represents a major challenge, and current protocols are based on electroporation of freshly excysted sporozoites using a rather large amount of plasmid DNA which typically has a very poor yield. In this study, we report a fast and simple protocol for transfection of Cryptosporidium parvum that takes advantage of the DNA condensing power of the poly cationic polymer polyethylenimine (PEI) and the gene delivery property of the short cell-penetrating peptide octaarginine. Our novel protocol requires a very low amount of plasmid DNA and does not necessitate special laboratory equipment to be performed. Transfection appears to be more efficient in oocysts just triggered for excystation than the excysted sporozoites. Altogether, the application of octaarginine with PEI allows efficient transfection. To the best of our knowledge, this is the first report on an electroporation-free protocol for transfection of sporozoites of a Cryptosporidium species.

Preserving Genome Integrity during the Early Embryonic DNA Replication Cycles

During the very early stages of embryonic development chromosome replication occurs under rather challenging conditions, including a very short cell cycle, absence of transcription, a relaxed DNA damage response and, in certain animal species, a highly contracted S-phase. This raises the puzzling question of how the genome can be faithfully replicated in such a peculiar metabolic context. Recent studies have provided new insights into this issue, and unveiled that embryos are prone to accumulate genetic and genomic alterations, most likely due to restricted cellular functions, in particular reduced DNA synthesis quality control. These findings may explain the low rate of successful development in mammals and the occurrence of diseases, such as abnormal developmental features and cancer. In this review, we will discuss recent findings in this field and put forward perspectives to further study this fascinating question.

Vegetation dynamics during the Pleistocene–Holocene transition in the central Great Plains, USA

The Holocene–Pleistocene transition in the upland loess-mantled regions of the central Great Plains is punctuated by the Brady Soil, which separates the late-Pleistocene Peoria Loess and the Holocene Bignell Loess. Previous research on the Brady Soil at the Old Wauneta Roadcut site in Southwestern Nebraska has produced paleoenvironmental information based on well-constrained luminescence and radiocarbon ages, stable carbon isotope data, and chemical and physical data. While the research indicated high effective moisture during formation of the Brady Soil and a shift to warm-season C4 vegetation from the cool-season C3-dominated vegetation of the Peoria Loess, those data do not provide any detail as to plant community composition and significant underlying climatic inferences. Assemblages of phytoliths and other biosilicates extracted from the Brady Soil provide specific information on vegetation communities and indicate shifts of plant taxa comprising these assemblages. Short-cell phytolith count data reveal a shift from dominance of Pooideae (C3) grasses, with relatively large numbers of arboreal dicot spheres and a few Cyperaceae (sedge) present in a savannah or open woodland in the Bølling-Allerød, to a mixed, open Chloridoideae (C4) and Pooideae (C3) grassland in the early-Holocene. Stipa-type Pooideae, a cool-season grass preferring drier soil conditions, marks the onset of the Younger Dryas. Large-cell phytoliths such as long cells, bulliforms, and trichomes, provided further definition of the climate history. This comprehensive biosilicate study of the Brady Soil has provided a more detailed paleoclimatic reconstruction than that generated with bulk sediment-derived δ13C data, or even with short-cell phytolith data alone.