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Isolation and Identification of Waterborne Antibiotic-Resistant Bacteria and Molecular Characterization of their Antibiotic Resistance Genes
Published on: March 3, 2023
High Frequency of the EMRSA-15 Clone (ST22-MRSA-IV) in Hospital Wastewater
Vanessa Silva1,2,3,4, Jessica Ribeiro1,2,3, Jaqueline Rocha5
1Microbiology and Antibiotic Resistance Team (MicroART), Department of Veterinary Sciences, University of Trás-os-Montes and Alto Douro (UTAD), 5000-801 Vila Real, Portugal.
Abstract:
Hospital wastewaters often carry multidrug-resistant bacteria and priority pathogens, such as methicillin-resistant Staphylococcus aureus (MRSA). Pathogens and antibiotic resistance genes present in wastewaters may reach the natural environment facilitating their spread. Thus, we aimed to isolate MRSA from wastewater of 3 hospitals located in the north of Portugal and to characterize the isolates regarding the antimicrobial resistance and genetic lineages. A total of 96 wastewater samples were collected over six months. The water was filtered, and the filtration membrane was immersed in BHI broth supplemented with 6.5% of NaCl and incubated. The inoculum was streaked in ORSAB agar plates for MRSA isolation. The isolates susceptibility testing was performed against 14 antimicrobial agents. The presence of resistance and virulence genes was accessed by PCR. Molecular typing was performed in all isolates. From the 96 samples, 28 (29.2%) were MRSA-positive. Most isolates had a multidrug-resistant profile and carried the mecA, blaZ, aac(6')-Ie-aph(2″)-Ia, aph(3')-IIIa, ermA, ermB, ermC, tetL, tetM, dfrA dfrG and catpC221 genes. Most of the isolates were ascribed to the immune evasion cluster (IEC) type B. The isolates belonged to ST22-IV, ST8-IV and ST105-II and spa-types t747, t1302, t19963, t6966, t020, t008 and tOur study shows that MRSA can be found over time in hospital wastewater. The wastewater treatment processes can reduce the MRSA load. The great majority of the isolates belonged to ST22 and spa-type t747 which suggests the fitness of these genetic lineages in hospital effluents.
Insights
Methicillin-resistant Staphylococcus aureus (MRSA) was detected in 29.2% of hospital wastewater samples in Portugal. Most MRSA isolates were multidrug-resistant and belonged to specific genetic lineages, highlighting the need for wastewater monitoring.
Area of Science:
- Environmental microbiology
- Clinical microbiology
- Antimicrobial resistance surveillance
Background:
- Hospital wastewaters are a significant source of multidrug-resistant bacteria (MDRB) and priority pathogens, including methicillin-resistant Staphylococcus aureus (MRSA).
- The presence of these resistant strains and their genes in wastewater poses a risk of dissemination into natural environments.
- Effective monitoring of MRSA in hospital effluents is crucial for understanding and mitigating the spread of antibiotic resistance.
Purpose of the Study:
- To isolate and characterize MRSA from the wastewater of three hospitals in northern Portugal.
- To determine the antimicrobial resistance profiles and genetic lineages of the isolated MRSA strains.
- To assess the prevalence of MRSA in hospital wastewater over a six-month period.
Main Methods:
- Collection and filtration of 96 wastewater samples over six months.
- Isolation of MRSA using selective broth enrichment and ORSAB agar.
- Antimicrobial susceptibility testing against 14 agents, PCR for resistance/virulence genes, and molecular typing (MLST, spa typing).
Main Results:
- MRSA was detected in 28 (29.2%) of the 96 wastewater samples.
- Most isolates exhibited multidrug resistance and carried numerous resistance genes (e.g., mecA, ermA, tetM, dfrA).
- Predominant genetic lineages included ST22-IV and ST8-IV, with spa-types t747 and t008 being most common, suggesting their adaptation to hospital effluents.
Conclusions:
- MRSA is consistently present in hospital wastewater, underscoring its role as a reservoir for resistant bacteria.
- Wastewater treatment processes can potentially reduce the load of MRSA.
- Specific MRSA lineages, particularly ST22-IV and spa-type t747, demonstrate significant fitness in hospital effluent environments.
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