Phylogeny and antimicrobial resistance in Neisseria gonorrhoeae isolates from Rio de Janeiro, Brazil

Abstract

Resistance in Neisseria gonorrhoeae is a global public health challenge. However, little is known about N. gonorrhoeae isolates from Brazil. In this study, we characterized 116 N. gonorrhoeae isolates obtained in Rio de Janeiro between 2006 and 2015 according to antimicrobial susceptibility profiles, resistance mechanisms, and clonal diversity. We determined antimicrobial minimal inhibitory concentrations by agar dilution, and whole genome sequencing was conducted to investigate alleles related to resistance, determine multilocus sequence typing profiles, and group isolates based on core genome single nucleotide polymorphisms. Resistance to penicillin, tetracycline, ciprofloxacin, and azithromycin was observed since 2006. Resistance to penicillin was mediated by β-lactamase plasmids and chromosomal mutations in ponA and porB genes, and tetracycline resistance was mediated by TetM plasmids, and porB and rspJ mutations. Ciprofloxacin resistant isolates presented cumulative point mutations in the quinolone resistance-determining region (QRDR) of gyrA and parC. Alterations in rrl genes encoding 23S rRNA, mtrR, and the mtrR promoter region were responsible for resistance to azithromycin. Phylogenetic analysis identified seven main clades, which included isolates with similar resistance profiles that mainly belonged to a limited number of sequence types that occurred during different years. Our results demonstrated high penicillin, tetracycline, and ciprofloxacin resistance rates associated with the persistence of successful resistant gonococcal lineages, and the polyclonal emergence of azithromycin resistance. Moreover, we reinforce the importance of surveillance to monitor the evolution of this scenario and to allow the early detection of possible changes to azithromycin and ceftriaxone as effective treatment options in the city.

Evidence for Clonally Associated Increasing Rates of Azithromycin ResistantNeisseria gonorrhoeaein Rio de Janeiro, Brazil

“…However, we performed a study based on isolates obtained from 2006 to 2015 in this city indicating 17% azithromycin resistance, based on CLSI ECV [14]. In that study, isolates were recovered mainly from private clinical laboratories at their convenience, but without any screening [14]. In the present study, we describe the increase of azithromycin resistance rate and azithromycin MIC among N. gonorrhoeae isolates received by our research laboratory from 2014 to 2017.…”

“…Rio de Janeiro was not included in the Brazilian-GASP. However, we performed a study based on isolates obtained from 2006 to 2015 in this city indicating 17% azithromycin resistance, based on CLSI ECV [14]. In that study, isolates were recovered mainly from private clinical laboratories at their convenience, but without any screening [14].…”

Emergence and evolution of antimicrobial resistance genes and mutations inNeisseria gonorrhoeae

“…Five other ST-1901 isolates dating from 2001 to 2004 in Kanagawa, Japan, were sequenced using the Illumina MiSeq platform. These newly sequenced genomes were combined with those of 1,066 publicly available genomes from 14 studies covering 37 countries [8, 11, 14, 15, 19, 21–28]. The publicly available genomes were selected based on a tree constructed in our previous study [29] such that they formed a group at the core-genome level.…”

Experimental Human Infection with Neisseria gonorrhoeae

“…Genomic DNA for WGS has successfully been prepared with the ArchivePure DNA purification kit Protocol 18 (5-PRIME, USA) as instructed by the manufacturer with the following modifications: 2.2 μL of RNase A solution; 2.5 μL of proteinase K mixed thoroughly by inverting the tube at least 25 times and incubated for 40 min at 37 °C; centrifugation time increased to 6 min for all steps, as previously described [11].…”