Molecular Epidemiology of Plasmid-Mediated Types 1 and 3 Fimbriae Associated with Biofilm Formation in Multidrug Resistant Escherichia coli from Diseased Food Animals in Guangdong, China.

Types 1 and 3 fimbriae in Enterobacteriaceae play versatile roles in bacterial physiology including attachment, invasion, cell motility as well as with biofilm formation and urinary tract infections. Herein, we investigated the prevalence and transmission of plasmid-mediated types 1 and 3 fimbriae from 1753 non-duplicate Enterobacteriaceae from diseased food Animals. We identified 123 (7.01%) strong biofilm producers and all was identified as E. coli. WGS analysis of 43 selected strong biofilm producers revealed that they harbored multiple ARGs, including ESBLs, PMQR and mcr-1. The gene clusters mrkABCDF and fimACDH encoding types 1 and 3 fimbriae, respectively, were identified among 43 (34.96%) and 7 (5.7%) of 123 strong biofilm isolates, respectively. These two operons were able to confer strong biofilm-forming ability to an E. coli weak-biofilm forming laboratory strain. Plasmid analysis revealed that mrk and fim operons were found to co-exist with ARGs and were primarily located on IncX1 and IncFII plasmids with similar backbones, respectively. mrkABCDF operons was present in all of 9457 Klebsiella pneumoniae using archived WGS data, and shared high homology to those on plasmids of 8 replicon types and chromosomes from 6 Enterobacteriaceae species from various origins and countries. In contrast, fimACDH operons was present in most of Enterobacter cloacae (62.15%), and shared high homology to those with only a small group of plasmids and Enterobacteriaceae species. This is the first comprehensive report of the prevalence, transmission and homology of plasmid-encoded type 1 and 3 fimbriae among the Enterobacteriaceae. Our findings indicated that plasmid-encoded mrkABCDF and fimACDH were major contributors to enhanced biofilm formation among E. coli and these two operons, in particular mrk could be as a potential anti-biofilm target. IMPORTANCE Biofilms allow bacteria to tolerate disinfectants and antimicrobials, as well as mammalian host defenses, and are therefore difficult to treat clinically. Most research concerning biofilm-related infections is typically focused on chromosomal biofilm-associated factors, including types 1 and 3 fimbriae of biofilm-forming Enterobacterium. However, the transmission and homology of the mobile types 1 and 3 fimbriae among Enterobacteriaceae is largely unknown. The findings revealed that the plasmid-encoded type 3 fimbriae encoded by mrkABCDF and type 1 fimbriae encoded by fimACDH were major contributors to enhancing biofilm formation among strong biofilm E. coli from diseased food producing animals. Additionally, mrk operon with high homology at an amino acid sequence was present both on plasmids of various replicon types and on chromosomes from diverse Enterobacteriaceae species from numerous origins and countries. These findings provide important information on the transmission of the mobile types 1 and 3 fimbriae among Enterobacteriaceae, indicating a potential antibiofilm target.

Cloned Guangxi Bama minipig (Sus scrofa) and its offspring have normal reproductive performance.

Xenotransplantation is a rapidly expanding field of research, and cloned miniature pigs are considered to be good model animals for its development. Although many animal species have been cloned, the success rate is very low, especially in the pig. To optimize the protocols for somatic cell nuclear transfer in the Guangxi Bama minipig, the relationship between cell cycle synchronization and nuclear histone acetylation levels were investigated. The results showed that the cells were efficiently synchronized by either serum starvation or contact inhibition. The level of nuclear histone acetylation in G0/G1 donor cells had similar variation trends in serum starvation and contact inhibition groups. When the synchronized donor cells were introduced into the enucleated oocytes, 8.8% (serum starvation group) or 9.7% (contact inhibition group) of the reconstructed embryos developed to blastocysts. After embryo transfer, one healthy male Guangxi Bama minipig was obtained. To evaluate the fertility of the cloned pig and its offspring, a series of mating experiments were done. Ninety-eight F1 generation crossbred piglets were born, of which 93 piglets survived. Also, the F1 pigs gave birth to 22 F2 generation piglets, of which 14 piglets survived. In conclusion, a Guangxi Bama minipig was successfully cloned from cultured newborn male gonad fibroblast cells, and the cloned minipig and its offspring had normal fertility.