To evaluate the genetic differences and possible evolutionary trends of clinical multidrug-resistant (MDR) strains, Pseudomonas aeruginosa isolates were characterized by pulsed-field gel electrophoresis (PFGE) and evolutionary distances were estimated. A total of 85.7% of the P. aeruginosa isolates were MDR strains. Strains with the PFGE pattern A predominated; all were susceptible to amikacin and cefepime but resistant to levofloxacin and meropenem (except strain PA45 which was sensitive to meropenem). PFGE pattern H or P strains exhibited resistance to six to eight different antibiotics. PFGE pattern I or J strains were susceptible to all antibiotics tested. Two imperfect six base-pair tandem repeats, CTGGCG and CTGGCC, were found in the mutL gene. In conclusion, MDR characteristics and PFGE profiles were clearly correlated with the mutL phylogenetic tree. This indicates that mutations in mutL might contribute to genetic stability in adaptation by changing the MDR traits. Phylogenetic analysis of mutL revealed the MDR relatedness of P. aeruginosa strains.
Aim To investigate the association between chronic hepatitis C virus (HCV) infection and Mooren's ulcer. Methods Eight patients from different parts of China who were diagnosed with Mooren's ulcer at the Zhongshan Ophthalmic Center, Guangzhou (China) were screened for chronic HCV infection. Mooren's ulcer was diagnosed by the typical ulcer morphology, detailed case history, physical examination, and comprehensive laboratory tests. All patients had serological screening for HCV infection. Results Six male and two female patients were enrolled in the study. Their ages ranged from 31 to 65 years (mean 43.6713.7). None of them was reported to have any clinical evidence of chronic HCV infection before enrolment and all were negative for HCV serology. Conclusion There was no association between chronic HCV infection and Mooren's ulcer in this limited case series study.
RNA structures can interact with the ribosome to alter translational reading frame maintenance and promote recoding that result in alternative protein products. Here, we show that the internal ribosome entry site (IRES) from the dicistrovirus Cricket paralysis virus drives translation of the 0-frame viral polyprotein and an overlapping +1 open reading frame, called ORFx, via a novel mechanism whereby a subset of ribosomes recruited to the IRES bypasses downstream to resume translation at the +1-frame 13th non-AUG codon. A mutant of CrPV containing a stop codon in the +1 frame ORFx sequence, yet synonymous in the 0-frame, is attenuated compared to wild-type virus in a Drosophila infection model, indicating the importance of +1 ORFx expression in promoting viral pathogenesis. This work demonstrates a novel programmed IRES-mediated recoding strategy to increase viral coding capacity and impact virus infection, highlighting the diversity of RNA-driven translation initiation mechanisms in eukaryotes.Significance StatementViruses use alternate mechanisms to increase the coding capacity of their viral genomes. Here, we provide biochemical evidence that ribosomes recruited to the dicistrovirus cricket paralysis virus IRES undergo a bypass event to direct translation of a downstream +1 frame overlapping open reading frame, called ORFx. Mutations that block ORFx expression inhibit +1 frame translation and infection in fruit flies. These findings highlight the diversity of RNA-driven translation initiation mechanisms in eukaryotes.
A new method for labeling oligonucleotides was developed to obtain high specific activity of radioactive probes. In an oligonucleotide molecule, two sequences were designed. One sequence, the 5', contains 19 nucleotides and serves as a template for probe synthesis. The second sequence, 3', contains a consensus sequence which forms a Pst I site after forming a complementary strand with the primer. In the presence of E. coli DNA polymerase I (Klenow fragment), alpha-32P dNTP and other dNTPs, a radioactive labeled oligonucleotide was synthesized by the primer extension method. After Pst I digestion, the probe was different from its template in length by 4 bp and could be separated from each other on urea-polyacrylamide gel electrophoresis (PAGE). A radioactive oligonucleotide probe with extremely high specific activity up to 10(10) dpm/micrograms could be obtained by the use of this method. The oligonucleotide probes have been used for the detection of the Hb E mutation in this report.
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