Serratia marcescens Term Paper Example | Topics and Well Written Essays - 1250 words

Serratia marcescens - Term Paper Ex adenineleS. marcescens is susceptible to expanded-spectrum cephalosporins, such(prenominal) as ceftoxamine, ceftazidime, cefepime, and cefpirome, though different mechanicss of resistance to expanded-spectrum cephalosporins have been reported. A constitutive overproduction of Amp-C, as a result of mutation in the regulatory gene system of Amp-C expression results in resistance to expanded-spectrum cephalosporins. The another(prenominal) enzymes that confer resistance to expanded-spectrum cephalosporins are Ambler Class A extended-spectrum ?-lactamases (ESBLs) or Class B metallo-?-lactamases. innovation in chromosomal antiophthalmic factor C gene also adds to the resistance pattern to expandedspectrum cephalosporins in the Family Enterobacteriaceae. Enterobacter cloacae GC1, S marcescens GN16694 and S. marcescens HD are among the few clinical isolates producing extended-spectrum ?- Amp C ?-lactamases. Yatsuyanagi et.al., (2006), in their study ex amined the resistance mechanism of cetazidime-resistant S. marcescens strains from inpatients of a cerebral ward over a 14 month period in champion hospital in Japan. One environmental and five E.coli transformants from clinical isolates harbouring the amp-C gene, cloned from S .marcescens and E.coli AS22-6-51, which is an amp-D mutant of E.coli C600, has a cut mutation in AmpC. The cloning vector used was pBcSK+. The MICs of azetreonam, imipenem, meropenem, gentamicin, minocycline and levofloxacin were determined by broth dilution technique. The prolongation strain used for in vitro susceptibility testing was E. coli ATCC 25922. Chromosomal DNA was prepared from S .marcescens and the isolates were examined by PCR for the presence of the following genes blaTEM , blaSHV, bla CTX-M, and plasmid borne amp-C genes. Primers specific for S.marcescens isolates were used to amplify a 552-bp constituent. Chromosomal DNA infix in agarose plugs were prepared for Pulse-Field Gel Electroph oresis (PFGE). The primers amp-C Seq 5 was used to amplify 1.174 bp fragment containing the chromosomal amp C gene and Mab/F and MAb/R primers were used to amplify the 1.192 bp fragment containing the blaTEM gene. These primers were used to sequence the chromosomal amp C gene and blaTEM gene from S. marcescens isolates. The primer amp C Exp5ER was used to amplify the chromosomal gene from S. marcescens strains ES46, ES76, and SM4 by PCR. Cloning of S. marcescens chromosomal amp C gene and construction of amp C harbouring E.coli transformants were carried out. Amp-C expressing transformants were confirmed to be positive for S.marcescens chromosomal amp C gene using PCR with a discipline of Sma amp C F and Sma amp C R primers and amp C Exp5ER and ampC Exp 3 HND primers for the 1,158 bp fragment containing the amp C gene using PCR. A primer was used to introduce a hitch mutation leading to a substitution of a third motif of the amp C gene, in a PCR based site -directed mutagenesis pe rformed withal PCR in vitro. Using overnight cultures of S. marcescens, amp C ?-lactamases stimulus generalisation and enzyme assay were done using an ultrasonic disrupter and the protein content was evaluated by BCA protein assay reagent. DNA sequence data analysis were performed using an updated version of Basic Local Alignment Search Tool. The nucleotide sequences of the chromosomal amp C genes of S. marcescens were deposited in the Gene Bank data base. Results of the study showed that four strains of S.marcescens (ES11, ES31, ES42, and ES46) isolated from urine specimens showed an indistinguishable SpeI PFGE pattern, indicating that a