| ObjectiveTo investigate the antimicrobial resistance of Proteus isolates, providing reasonable guidelines for proper use of antibiotics and analyze the sequence features of SXT/R3 91 integrative and conjugative element of Protes in China.Methods(1) The minimum inhibitory concentrations (MICs) of tested antibiotics were measured and interpreted using the broth microdilution method according to the instructions and standards of Clinical and Laboratory Standards Institute (CLSI).(2) A total of 123 Proteus strains were screened for the presence of SXT/R391 by targeting the int gene. Sequencing of SXT/R391 positive strains by using Next Generation Sequencing (NGS),the SXT/R391 sequences were then extracted from the whole genomes and assembly of the sequences was performed. Then analyzed the structure and function based on full SXT/R391 sequences.(3) Broth microdilution method and mating assay were performed.ResultsAll of the Proteus isolates were resistant to one or more antibiotics. The resistance percentage of Proteus mirabilis for SXT is 47.9%, Ampicillin(45.2%), Cefazolin(44.7%), and Ciprofloxacin(35.6%). For Proteus vulgaris, resistance rate of tested drugs was generally no more than 20%. Proteus isolates in this study had a lowest resistance rate for Piperacillin-tazobactam. Proteus mirabilis had a higher antibiotic resistance percentage than that of Proteus vulgaris on Cefepime, Cefotetan, Amikacin,Gentamycin, Trimethoprim-sulfamethoxazole and Ciprofloxacin. Proteus mirabilis strains from nosocomial infections had a higher resistance rate than that of food and stool,however, for Proteus vulgaris, the difference was not significant. In all the resistant strains of Proteus mirabilis, 67(35.6%) were resistant to three or more classes of antibiotics.Fifteen out of 123 strains (12.2%) were positive for int gene, 2 food isolates and 13 fecal samples were included. The 15 SXT/R3 91 sequences had highest query cover and identity with ICEs from Alteromonas macleodii, Vibrio cholerae and Proteus mirabilis. The query cover ranged from 64-99%, and identity from 97-100%. Nine and two of the sequences had highest query cover and identity with ICEs of Ptoteus and Vibrio cholerae, respectively. In terms of evolution, 15 SXT/R391 were classified into 6 different clusters, among which, ICEPmiChnl was a predominant ICE type, 5 ICEs were firstly found in our study. Besides, the results of gene annotation and structure analysis suggested that both conserved genes and inserted variable genes were present in SXT/R391 ICEs of Proteus, for example, antibiotic resistance genes and genes encoding restriction and modification system and so on. What’s more,a P-lactamase gene appeared in VRIII, which was rarely reported. The transfer frequency ranged from 3.5×10-6-2.5× 10-2.ConclusionsOur results showed that Proteus strains have been highly resistant to some commonly used antibiotics, and the phenomenon of multi-drug resistance is not rare. SXT/R391 ICEs could be transferred between Proteus and other Enterobacteriaceae, conferring resistance to the host, facilitating bacteria to survive in the environment. Therefore,we need to strengthen the continuous monitoring of antimicrobial resistance and related mobile elements among Proteus.ObjectiveA total of 187 Aeromonas strains were collected, identification of Aeromonas at species level by rpoD and gyrA gene, comparing the identification results and evaluate usefulness of gyrA gene sequence for the identification of Aeromonas strains.Getting more accurate classifications of 187 strains in this study.MethodsPCR amplification and sequencing of rpoD and gyrA gene of 187 strains of Aeromonas identified by biochemical method, constructing phylogenetic tree with 26 reference sequences that are species specific by Neighbor-Joing method.ResultsBy rpoD gene sequence analysis, 187 stains were classified into 7 species,however, by gyrA gene analysis, 9 species were identified and one indeterminate. The top three species were identical to results by rpoD gene analysis, namely, A. veronii bv.sobria, A. caviae and A. aquariorum. The added two species were A.allosaccharophila and A.jandaei. There was a consistency of 174 strains (93%) by the two genes, but 13 strains (7%) were not identical.Identification of A. veronii bv. sobria (64) and A. caviae (28) had a high concistency, between 68.8% and 85.7%. However, 95 A. hydrophila strains were reclassified as the same strain by rpoD (9.5%) and gyrA (6.3%) respectively.ConclusionsrpoD and gyrA gene were able to classify most of the Aeromonas strains accurately and consistently, however, more housekeeping genes are necessary to identify strains which were not clearly identified based on current method. |
