Reema Singh1,2, Sumudu R Perera1,2, George S Katselis3, Paulos Chumala3, Irene Martin4, Anthony Kusalik5, Kristen M Mitzel1, Jo-Anne R Dillon1,2. 1. Department of Biochemistry, Microbiology and Immunology, 2D01 Health Science Building, 107 Wiggins Road, University of Saskatchewan, Saskatoon, SK, Canada. 2. Vaccine and Infectious Disease Organization-International Vaccine Centre, University of Saskatchewan, 120 Veterinary Road, Saskatoon, SK, Canada. 3. Department of Medicine, Division of the Canadian Centre for Health and Safety in Agriculture, 1246 Health Sciences E-Wing, 104 Clinic Place, University of Saskatchewan, Saskatoon, SK, Canada. 4. National Microbiology Laboratory, Streptococcus and STI Unit, Public Health Agency of Canada, 1015 Arlington Street, Winnipeg, MB, Canada. 5. Department of Computer Science, 176 Thorvaldson Building, 110 Science Place, University of Saskatchewan, Saskatoon, SK, Canada.
Abstract
BACKGROUND: Seven structurally related β-lactamase-producing plasmids have been characterized in penicillinase-producing Neisseria gonorrhoeae (PPNG) isolates. We characterized a variant (i.e. pJRD20, Canada type) of the Africa-type (pJD5) plasmid isolated from N. gonorrhoeae strain 8903. OBJECTIVES: To compare the DNA sequence of pJRD20 with that of pJD5 and pJD4 (Asia-type) and their TEM-1 β-lactamases. METHODS: N. gonorrhoeae 8903 was identified as part of the Gonococcal Antimicrobial Surveillance Program in Canada. β-Lactamase production was assessed using nitrocefin. MICs were determined by agar dilution and Etest methods (CLSI). The DNA sequences of pJRD20, pJD5 and pJD4 were assembled and annotated. The structure of TEM-1 and its penicillin-binding properties were determined by in silico molecular modelling and docking. TEM-1 proteins were characterized by western blot, mass spectrometry and ampicillin hydrolysis assays. RESULTS: N. gonorrhoeae 8903 exhibited intermediate susceptibility to penicillin with slow β-lactamase activity (i.e. 35 min to hydrolyse nitrocefin). Except for a novel 6 bp deletion starting at the G of the ATG start codon of blaTEM-1, the DNA sequence of pJRD20 was identical to that of pJD5. The TEM-1 β-lactamase produced by pJRD20 is 24 kDa and hydrolyses ampicillin only after several hours. CONCLUSIONS: This unusual PPNG isolate might have been characterized as a non-PPNG owing to its low MIC of penicillin and its very slow hydrolysis of nitrocefin. Given the unusual nature of its TEM-1 β-lactamase, laboratories might consider extending the duration of nitrocefin hydrolysis assays.
BACKGROUND: Seven structurally related β-lactamase-producing plasmids have been characterized in penicillinase-producing Neisseria gonorrhoeae (PPNG) isolates. We characterized a variant (i.e. pJRD20, Canada type) of the Africa-type (pJD5) plasmid isolated from N. gonorrhoeae strain 8903. OBJECTIVES: To compare the DNA sequence of pJRD20 with that of pJD5 and pJD4 (Asia-type) and their TEM-1 β-lactamases. METHODS:N. gonorrhoeae 8903 was identified as part of the Gonococcal Antimicrobial Surveillance Program in Canada. β-Lactamase production was assessed using nitrocefin. MICs were determined by agar dilution and Etest methods (CLSI). The DNA sequences of pJRD20, pJD5 and pJD4 were assembled and annotated. The structure of TEM-1 and its penicillin-binding properties were determined by in silico molecular modelling and docking. TEM-1 proteins were characterized by western blot, mass spectrometry and ampicillin hydrolysis assays. RESULTS:N. gonorrhoeae 8903 exhibited intermediate susceptibility to penicillin with slow β-lactamase activity (i.e. 35 min to hydrolyse nitrocefin). Except for a novel 6 bp deletion starting at the G of the ATG start codon of blaTEM-1, the DNA sequence of pJRD20 was identical to that of pJD5. The TEM-1 β-lactamase produced by pJRD20 is 24 kDa and hydrolyses ampicillin only after several hours. CONCLUSIONS: This unusual PPNG isolate might have been characterized as a non-PPNG owing to its low MIC of penicillin and its very slow hydrolysis of nitrocefin. Given the unusual nature of its TEM-1 β-lactamase, laboratories might consider extending the duration of nitrocefin hydrolysis assays.
Authors: Ana Cehovin; Keith A Jolley; Martin C J Maiden; Odile B Harrison; Christoph M Tang Journal: J Infect Dis Date: 2020-11-09 Impact factor: 5.226
Authors: Ivan Bodoev; Maja Malakhova; Julia Bespyatykh; Dmitry Bespiatykh; Georgij Arapidi; Olga Pobeguts; Victor Zgoda; Egor Shitikov; Elena Ilina Journal: Genes (Basel) Date: 2021-08-25 Impact factor: 4.096