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Beltsville, MD, United States

Khan A.,University of Washington | Khan A.,U.S. National Institutes of Health | Shaik J.S.,University of Washington | Behnke M.,University of Washington | And 6 more authors.
BMC Genomics | Year: 2014

Background: Toxoplasma gondii is a widespread protozoan parasite of animals that causes zoonotic disease in humans. Three clonal variants predominate in North America and Europe, while South American strains are genetically diverse, and undergo more frequent recombination. All three northern clonal variants share a monomorphic version of chromosome Ia (ChrIa), which is also found in unrelated, but successful southern lineages. Although this pattern could reflect a selective advantage, it might also arise from non-Mendelian segregation during meiosis. To understand the inheritance of ChrIa, we performed a genetic cross between the northern clonal type 2 ME49 strain and a divergent southern type 10 strain called VAND, which harbors a divergent ChrIa. Results: NextGen sequencing of haploid F1 progeny was used to generate a genetic map revealing a low level of conventional recombination, with an unexpectedly high frequency of short, double crossovers. Notably, both the monomorphic and divergent versions of ChrIa were isolated with equal frequency. As well, ChrIa showed no evidence of being a sex chromosome, of harboring an inversion, or distorting patterns of segregation. Although VAND was unable to self fertilize in the cat, it underwent successful out-crossing with ME49 and hybrid survival was strongly associated with inheritance of ChrIII from ME49 and ChrIb from VAND. Conclusions: Our findings suggest that the successful spread of the monomorphic ChrIa in the wild has not been driven by meiotic drive or related processes, but rather is due to a fitness advantage. As well, the high frequency of short double crossovers is expected to greatly increase genetic diversity among progeny from genetic crosses, thereby providing an unexpected and likely important source of diversity. © 2014 Khan et al. Source


Natilla A.,Beltsville Agricultural Research Service | Murphy C.,Beltsville Agricultural Research Service | Hammond R.W.,Beltsville Agricultural Research Service
Virus Research | Year: 2015

Viral-based nanoplatforms rely on balancing the delicate array of virus properties to optimally achieve encapsidation of foreign materials with various potential objectives. We investigated the use of Maize rayado fino virus (MRFV)-virus-like particles (VLPs) as a multifunctional nanoplatform and their potential application as protein cages. MRFV-VLPs are composed of two serologically related, carboxy co-terminal coat proteins (CP1 and CP2) which are capable of self-assembling in Nicotiana benthamiana plants into 30. nm particles with T=. 3 symmetry. The N-terminus of CP1 was targeted for genetic modification to exploit the driving forces for VLP assembly, packaging and retention of RNA in vivo and in vitro. The N-terminus of MRFV-CP1 contains a peptide sequence of 37 amino acids which has been predicted to have an alpha-helical structure, is rich in hydrophobic amino acids, facilitates CP-RNA interactions, and is not required for self-assembly. Amino acid substitutions were introduced in the 37 amino acid N-terminus by site-directed mutagenesis and the mutant VLPs produced in plants by a Potato virus X (PVX)-based vector were tested for particle stability and RNA encapsidation. All mutant CPs resulted in production of VLPs which encapsidated non-viral RNAs, including PVX genomic and subgenomic (sg) RNAs, 18S rRNA and cellular and viral mRNAs. In addition, MRFV-VLPs encapsidated GFP mRNA when was expressed in plant cells from the pGD vector. These results suggest that RNA packaging in MRFV-VLPs is predominantly driven by electrostatic interactions between the N-terminal 37 amino acid extension of CP1 and RNA, and that the overall species concentration of RNA in the cellular pool may determine the abundance and species of the RNAs packaged into the VLPs. Furthermore, RNA encapsidation is not required for VLPs stability, VLPs formed from MRFV-CP1 were stable at temperatures up to 70. °C, and can be disassembled into CP monomers, which can then reassemble in vitro into complete VLPs either in the absence or presence of RNAs. © 2014. Source

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