DNA samples from 737 tick pools, representing 6,850 Haemaphysalis longicornis and 51 Dermacentor nuttalli collected from Hebei Province, China, were analyzed by polymerase chain reaction (PCR) for the presence of spotted fever group Rickettsia. Fifty (6.9%) of 724 H. longicornis in the tick pool were positive, but no positive samples were found in 13 D. nuttalli. Sequence analysis of the partial outer membrane protein A (ompA) genes from the 10 positive samples showed 97.4-99.8% identity, but were different from the homologous sequence of Rickettsia previously deposited in GenBank. Phylogenetic analysis of ompA genes indicated that the Rickettsia detected in this study belonged to a novel haplotype, and formed a clade distinct from Rickettsia heilongjiangii, Rickettsia sibirica, and Rickettsia hulinii in China. The new strain, named Candidatus Rickettsia hebeiii, appears to represent a distinct lineage and could constitute a new species with a minimum prevalence of about 0.7% in H. longicornis from Hebei Province, China.
Genetically modified pigs are valuable models of human disease and donors of xenotransplanted organs. Conventional gene targeting in pig somatic cells is extremely inefficient. Zinc-finger nuclease (ZFN) technology has been shown to be a powerful tool for efficiently inducing mutations in the genome. However, ZFN-mediated targeting in pigs has rarely been achieved. Here, we used ZFNs to knock out the porcine α-1,3-galactosyl-transferase (GGTA1) gene, which generates Gal epitopes that trigger hyperacute immune rejection in pig-to-human transplantation. Primary pig fibroblasts were transfected with ZFNs targeting the coding region of GGTA1. Eighteen mono-allelic and four biallelic knockout cell clones were obtained after drug selection with efficiencies of 23.4% and 5.2%, respectively. The biallelic cells were used to produce cloned pigs via somatic cell nuclear transfer (SCNT). Three GGTA1 null piglets were born, and one knockout primary fibroblast cell line was established from a cloned fetus. Gal epitopes on GGTA1 null pig cells were completely eliminated from the cell membrane. Functionally, GGTA1 knockout cells were protected from complement-mediated immune attacks when incubated with human serum. This study demonstrated that ZFN is an efficient tool in creating gene-modified pigs. GGTA1 null pigs and GGTA1 null fetal fibroblasts would benefit research and pig-to-human transplantation.pig, xenotransplantation, ZFNs, GGTA1, biallelic knockout, SCNT Citation:
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