RESUMO
Neisseria gonorrhoeae resistente a quinolonas (QRNG) es un problema muy importante en salud pública por su rápido desarrollo de resistencia a quinolonas, por lo que la OMS recomienda reforzar la vigilancia de resistencia antimicrobiana para orientar el tratamiento. En Perú hay pocos reportes sobre vigilancia de QRNG, y menos aún trabajos relacionados a patrones de mutación. Este estudio busca encontrar mutaciones en la Región Determinante de Resistencia a Quinolonas (QRDR) del gen gyrA de N. gonorrhoeae a partir de 1096 muestras clínicas de orina e hisopados, colectadas entre 2012 y 2013, provenientes de 367 hombres que tuvieron sexo con hombres (HSH). Se detectaron 58 muestras positivas a N. gonorrhoeae en 45 HSH mediante el ensayo de APTIMA Combo 2, y 11 muestras positivas a QRNG de 11 HSH mediante PCR en Tiempo Real. Las bacterias resistentes a quinolonas fueron analizadas por secuenciamiento e identificamos que el patrón frecuente de mutación fue la doble mutación de Ser-91 a Phe y de Asp-95 a Gly en la QRDR del gen gyrA. En conclusión, estas dobles mutaciones en la secuencia QRDR del gen gyrA indicarían la presencia de N. gonorrhoeae con fenotipo resistente a quinolonas en muestras clínicas de HSH de Lima-Perú, resaltando que éste es el primer estudio hecho en Perú sobre esta población de alto riesgo
Neisseria gonorrhoeae resistant to quinolones (QRNG) becomes a very important problem in public health due to the development of rapid resistance to quinolones, which is why the WHO recommends reinforcing antimicrobial resistance monitoring to guide treatment. In Peru there are few reports on QRNG surveillance and still less work related to mutation patterns. This study aims to find mutations in the Determinant Region of Quinolone Resistance (QRDR) of the gyrA gene of N. gonorrhoeae from 1096 clinical samples of urine and swabs from 367 men who have sex with men (MSM) collected during the period 2012-2013. We detected 58 N. gonorrhoeae positive samples in 45 MSM by means of the APTIMA Combo 2 assay, and 11 QRNG positive samples of 11 MSM by Real Time PCR. The quinolone-resistant bacteria were analyzed by sequencing and we identified that the frequent mutation pattern was the double mutation of Ser-91 to Phe and Asp-95 to Gly in the QRDR of the gyrA gene. In conclusion, these double mutations in the QRDR sequence of the gyrA gene would indicate the presence of N. gonorrhoeae with quinolone resistant phenotype in clinical samples of MSM from Lima-Peru, noting that this is the first study done in Peru on this high population risk
RESUMO
Introdução. Quinolonas são antimicrobianos sintéticos que inibem as enzimas DNA-girase e topoisomerase IV resultando na morte bacteriana. São altamente eficazes no tratamento de infecções bacterianas, especialmente causadas por bactérias Gram negativas, e portanto amplamente utilizados na medicina humana e veterinária, na qual também são empregados como profiláticos. Porém, o uso indiscriminado e inadequado levou ao aumento de bactérias resistentes a estes compostos. Esta resistência pode ocorrer devido a mutações nas enzimas DNA-girase e topoisomerase IV, e também por genes contidos em plasmídeos. Estes últimos são os principais responsáveis pela disseminação e circulação da resistência entre o meio ambiente e o ambiente hospitalar. Objetivos. Pesquisar genes de resistência a antimicrobianos do grupo das quinolonas em bactérias Gram negativas de origem clínica e ambiental que apresentam resistência fenotípica a este grupo. Material e Métodos. 73 cepas de Enterobacteriaceae e Aeromonas sp. de origem clínica e ambiental foram selecionadas para o estudo, e avaliadas quanto à sensibilidade aos antimicrobianos do grupo das quinolonas e à pesquisa de genes de resistência a este mesmo grupo e mutações no gene que codifica a enzima DNA-girase por meio de PCR e sequenciamento. Resultados. Das 73 cepas previamente selecionadas para compor o estudo, 65 foram utilizadas, devido à exclusão de perfis clonais similares. Nestas, foram observados os genes, qnrS1 (1,5 por cento ), qnrS2 (26,2 por cento ), qnrB1 (3,1 por cento ), qnrB19 (12,3 por cento ), qnrD1 (1,5 por cento ), aac(6)-Ib-cr (10,8 por cento ), oqxA (43,1 por cento ) e oqxB (41,5 por cento ), e duas variantes determinadas qnrB-like (3,1 por cento ) e qnrB69-like (1,5 por cento ). Os genes qnrA, qnrC e qepA não foram identificados. Mutações na enzima DNA-girase foram observadas em 97,9 por cento das cepas positivas para algum dos genes pesquisados...
Introduction. Quinolones are synthetic antimicrobial agents that inhibit DNA gyrase and topoisomerase IV enzymes resulting in bacterial death. They are highly effective in the treatment of bacterial infections, especially the ones caused by Gram negative bacteria, as well as for prophylaxy. Therefore they are widely used in human and veterinary medicine. However, indiscriminate and improper use led to an increase of bacteria resistance to these compounds. This resistance can be due to mutations in DNA gyrase and topoisomerase IV enzymes and also by genes contained in plasmids, which are mainly responsible for the spread and transmission of resistance between the environment and the hospital set. Objectives. To search for genes of resistance to quinolone antimicrobial agents in Gram-negative bacteria from clinical and environmental strains that present phenotypic resistance to this group. Material and Methods. 73 strains of Enterobacteriaceae and Aeromonas spp., from clinical and environmental origin, were selected for this study, and evaluated for antimicrobial susceptibility of quinolone and search of resistance genes in this same group and also for mutations in the gene encoding the enzyme DNA gyrase by PCR and sequencing. Results. Of the 73 strains previously selected to compose this study, 65 were used, due to the exclusion of similar clonal profiles. In these, genes qnrS1 (1.5 per cent ), qnrS2 (26.2 per cent ) qnrB1 (3.1 per cent ), qnrB19 (12.3 per cent ) qnrD1 (1.5 per cent ) aac(6')-Ib-cr (10.8 per cent ) oqxA (43.1 per cent ) and oqxB (41.5 per cent ) were observed, and two variants were named as qnrB-like (3.1 per cent ) and qnrB69-like (1.5 per cent ). The qnrA, qnrC and qepA genes were not identified. Mutations in DNA gyrase enzyme were observed in 97.9 per cent of the positive strains for at least one of the genes studied. It was possible to establish the association of aac(6')-Ib-cr with class 1 integron gene in four strains...