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Evaluation of the occurrence and prevalence of antimicrobial resistant bacteria and resistance genes in wastewater from Primary Care Units (PCU) and in urban surface water.

Grant number: 23/12641-2
Support Opportunities:Regular Research Grants
Duration: May 01, 2024 - April 30, 2026
Field of knowledge:Engineering - Sanitary Engineering - Environmental Sanitation
Principal Investigator:Maria Tereza Pepe Razzolini
Grantee:Maria Tereza Pepe Razzolini
Host Institution: Faculdade de Saúde Pública (FSP). Universidade de São Paulo (USP). São Paulo , SP, Brazil

Abstract

Antimicrobial resistance (AMR) is classified by the World Health Organization as a global public health problem (WHO 2018), driven by the use of antimicrobials in both human and animal health, as well as in food production, which leads to infections. resistance, increasing morbidity and mortality rates and treatment costs. The objectives of this proposal are to promote advances in environmental surveillance and collaborate with data of importance in Public and Environmental Health on the prevalence of ADRs/antimicrobials in urbanized communities, estimating the prevalence of resistance to antimicrobials in samples of wastewater and drinking water. stream in urbanized area; estimating the occurrence of resistance genes in samples of wastewater and stream water in an urbanized area, and providing access to data (spatial and statistical) on the occurrence of AMR and resistance genes in environmental samples. Sewage and water samples from impacted streams will be submitted to quantification of E. coli and Enterococcus. To determine the occurrence of ESBL-producing E. coli, a volume of 1000mL was concentrated on a filtering membrane (45µm, 47mm), cultivated in Petri dishes containing MacConkey agar medium supplemented with cefotaxime (4µg/mL). Likewise, the occurrence of methicillin-resistant Staphylococcus aureus will be cultivated in Petri dishes containing the mannitol salt agar medium supplemented with cefoxitin (8µg/mL). For both, selected colonies will be identified by MALDI-TOF mass spectrometry, and the antimicrobial susceptibility profile will be evaluated by disk diffusion methods, according to CLSI guidelines (2019). PCR screening for methicillin resistance gene cassettes will be performed by PCR according to Okuma et al. (2002) while ESBL production will be evaluated by disk diffusion methods, according to CLSI guidelines (2019). PCR screening for ESBL-encoding genes will be performed by previously described multiplex PCR (Dallenne et al 2010). For the determination of the genetic markers of RAM, volumes of 1000mL will be concentrated through a membrane filter technique, and, followed by DNA extraction directly from the membranes using the DNeasy PowerWater kit (Qiagen, Hilden Germany) for the search for class 1 integrases (IntI1) and genes encoding resistance to ²-lactams, Fluoroquinolones, Polymyxins, Aminoglycosides and Phosphomycins, using methods previously described in the literature. (AU)

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