RESUMEN
OBJECTIVES: To develop a simple DNA sequencing test for simultaneous identification and antimicrobial resistance (AMR) detection of multiple sexually transmitted infections (STIs). METHODS: Real-time PCR (qPCR) was initially performed to identify Neisseria gonorrhoeae (NG), Chlamydia trachomatis (CT), Mycoplasma genitalium (MG) and Trichomonas vaginalis (TV) infections among a total of 200 vulvo-vaginal swab samples from female sex workers in Ecuador. qPCR positive samples plus qPCR negative controls for these STIs were subjected to single gene targeted PCR MinION-nanopore sequencing using the smartphone operated MinIT. RESULTS: Among 200 vulvo-vaginal swab samples 43 were qPCR positive for at least one of the STIs. Single gene targeted nanopore sequencing generally yielded higher pathogen specific read counts in qPCR positive samples than qPCR negative controls. Of the 26 CT, NG or MG infections identified by qPCR, 25 were clearly distinguishable from qPCR negative controls by read count. Discrimination of TV qPCR positives from qPCR negative controls was poorer as many had low pathogen loads (qPCR cycle threshold >35) which produced few specific reads. Real-time AMR profiling revealed that 3/3 NG samples identified had gyrA mutations associated with fluoroquinolone resistance, 2/10 of TV had mutations related to metronidazole resistance, while none of the MG samples possessed 23S rRNA gene mutations contributing to macrolide resistance. CONCLUSIONS: Single gene targeted nanopore sequencing for diagnosing and simultaneously identifying key antimicrobial resistance markers for four common genital STIs shows promise. Further work to optimise accuracy, reduce costs and improve speed may allow sustainable approaches for managing STIs and emerging AMR in resource poor and laboratory limited settings.
Asunto(s)
Farmacorresistencia Bacteriana/genética , Mycoplasma genitalium/genética , Neisseria gonorrhoeae/genética , Enfermedades de Transmisión Sexual/diagnóstico , Trichomonas vaginalis/genética , Antibacterianos/farmacología , Antibacterianos/uso terapéutico , Girasa de ADN/genética , Ecuador , Femenino , Fluoroquinolonas/farmacología , Fluoroquinolonas/uso terapéutico , Humanos , Macrólidos/farmacología , Mycoplasma genitalium/efectos de los fármacos , Mycoplasma genitalium/aislamiento & purificación , Secuenciación de Nanoporos , Neisseria gonorrhoeae/efectos de los fármacos , Neisseria gonorrhoeae/aislamiento & purificación , ARN Ribosómico 23S/química , ARN Ribosómico 23S/genética , ARN Ribosómico 23S/metabolismo , Reacción en Cadena en Tiempo Real de la Polimerasa , Trabajadores Sexuales , Enfermedades de Transmisión Sexual/tratamiento farmacológico , Enfermedades de Transmisión Sexual/microbiología , Trichomonas vaginalis/efectos de los fármacos , Trichomonas vaginalis/aislamiento & purificación , Vagina/microbiologíaRESUMEN
Infection with Streptococcus pneumoniae is the leading cause of death in children and burden of disease is greatest where helminth infections are also common. We investigated the impact of intestinal helminth co-infection on pneumococcal carriage; a risk factor for invasive disease. We used a mouse co-infection model and clinical data to assess the impact of co-infection on carriage density. Co-infection in mice was associated with increased pneumococcal carriage density and dissemination into lungs. Helminth-infected children also exhibited increased carriage density as compared to uninfected children. Anthelmintic treatment may be a cost-effective method of reducing pneumococcal disease burden in lower-income countries.