Perfil microbiológico de bacterias resistentes a antibióticos en muestras provenientes de aguas residuales de PTAR La Escalerilla, Arequipa 2022
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Date
2023
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Universidad Nacional de Trujillo
Abstract
El objetivo de esta investigación fue determinar el perfil microbiológico de las bacterias resistentes a antibióticos en muestras provenientes de aguas residuales de la Planta de Tratamiento de Aguas Residuales (PTAR) “La Escalerilla”, Arequipa durante el mes de abril del 2022. Para el aislamiento se hicieron diluciones seriadas de las muestras de agua colectadas en agua peptonada (Mesquita et al., 2021), posteriormente se procedió a cultivarlas en placas con agar MacConkey y agar sangre (Mutuku, 2017). La identificación de las colonias aisladas se realizó mediante la técnica por espectrometría de masas con tiempo de vuelo (MALDI-TOF MS) con el equipo VITEK MS (Biomérieux) a través de la plataforma RUO (Research Use Only). Se aislaron 126 colonias de bacterias, la determinación del perfil microbiológico se realizó mediante sus características culturales y morfología de la colonia, realización de frotis de las colonias aisladas y coloración de Gram, la mayoría fueron bacilos Gram negativos (99.2%, 125) y coco Gram positivo (0.8%, 1). Se identificaron las 126 colonias, el 66.7% fueron Enterobacterias (Citrobacter freundii, Citrobacter sp., Enterobacter cloacae, Enterobacter ludwigii, Enterobacter sp., Escherichia coli, Hafnia alvei, Klebsiella oxytoca, Klebsiella pneumoniae, Proteous mirabilis, Proteous vulgaris/penneri, Raoultella ornithinolytica y Serratia liquefaciens), 28.6% Aeromonas spp. (Aeromonas punctata (caviae), Aeromonas sp., Aeromonas hydrophila y Aeromonas veronii), 2.4% Pseudomonas spp. (Pseudomonas oleovorans y Pseudomonas cuatrocienegasensis) y en menor porcentaje Alcaligenes faecalis, Shewanella putrefasciens y Enterococcus faecalis (0.8% cada uno). La susceptibilidad a los antibióticos (amoxicilina/acido clavulánico, ampicilina, cefoxitina, ceftriaxona, imipenem, azitromicina, ciprofloxacino, tetraciclina, estreptomicina, nitrofurantoina sulfametoxazol/trimetoprima y cloranfenicol) se evaluó mediante la técnica de Kirby-Bauer en agar Mueller-Hinton siguiendo las directrices de Clinical Laboratory Standards Institute (CLSI). El diámetro del halo de inhibición se definió como sensible (S), intermedio (I) o resistente (R) (Hartinger et al., 2021). El resultado de susceptibilidad fue analizado por cada uno de los antibióticos. El perfil de resistencia en las Enterobacterias a antibióticos fue variado, siendo mayor con ampicilina (59.5%), seguido por sulfametoxazol/trimetoprima (41.7%) y ciprofloxacino (40.5%), de esta familia la bacteria que presentó mayor número de colonias resistentes fue
Escherichia coli. Las Aeromonadaceae presentaron mayor resistencia a sulfametoxazol/trimetoprima (44.4%), seguido por ciprofloxacino (33.3%) y ceftriaxona (30.6%), de las diferentes especies de esta familia Aeromonas sp. presentó mayor número de colonias resistentes y la colonia de Enterococcus faecalis fue sensible a los cinco antibióticos aplicados. Los resultados de este trabajo de investigación evidencian la presencia de bacterias resistentes a antibióticos en el afluente de la PTAR La Escalerilla de Arequipa.
This research aimed to determine the microbiological profile of antibiotic-resistant bacteria in wastewater samples collected from the "La Escalerilla" Wastewater Treatment Plant (WWTP) in Arequipa in April 2022. The isolation process involved serial dilutions of collected water samples in peptone water (Mesquita et al., 2021), followed by cultivation on MacConkey agar and blood agar plates (Mutuku, 2017). The identification of isolated colonies was carried out using Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry (MALDI-TOF MS) with the VITEK MS system (Biomérieux) through the Research Use Only (RUO) platform. A total of 126 bacterial colonies were isolated. The microbiological profiling was based on cultural characteristics, colony morphology, smears of isolated colonies, and Gram staining. The majority of the isolated colonies were Gram-negative bacilli (98.4%, 124), followed by Gram-negative coccobacilli (0.8%, 1), and Gram-positive cocci (0.8%, 1). Out of the 126 isolated colonies, 66.7% were identified as Enterobacteriaceae, encompassing species such as Citrobacter freundii, Citrobacter sp., Enterobacter cloacae, Enterobacter ludwigii, Enterobacter sp., Escherichia coli, Hafnia alvei, Klebsiella oxytoca, Klebsiella pneumoniae, Proteus mirabilis, Proteus vulgaris/penneri, Raoultella ornithinolytica, and Serratia liquefaciens. Additionally, 28.6% were identified as Aeromonas spp. including Aeromonas punctata (caviae), Aeromonas sp., Aeromonas hydrophila, and Aeromonas veronii. Pseudomonas spp. accounted for 2.4% of the isolated colonies, specifically Pseudomonas oleovorans and Pseudomonas cuatrocienegasensis. A smaller percentage of colonies were identified as Alcaligenes faecalis, Shewanella putrefaciens, and Enterococcus faecalis (0.8% each). Antibiotic susceptibility testing was performed using the Kirby-Bauer method on Mueller-Hinton agar, following the guidelines of the Clinical Laboratory Standards Institute (CLSI). The diameter of the inhibition zone determined the susceptibility as sensitive (S), intermediate (I), or resistant (R) based on the methodology by Hartinger et al. (2021). Resistance profiles were analyzed for each antibiotic. In Enterobacteriaceae, varying resistance patterns were observed, with higher resistance rates to ampicillin (59.5%), followed by sulfamethoxazole/trimethoprim (41.7%), and ciprofloxacin (40.5%). Among this group, Escherichia coli exhibited the highest number of resistant colonies. Among Aeromonadaceae, the greatest resistance was seen against sulfamethoxazole/trimethoprim (44.4%), followed by ciprofloxacin (33.3%) and ceftriaxone (30.6%). Within this family, Aeromonas sp. showed the highest number of resistant colonies. Notably, the Enterococcus faecalis colony demonstrated susceptibility to all five tested antibiotics. The findings of this research highlight the presence of antibiotic-resistant bacteria in the influent of the "La Escalerilla" WWTP in Arequipa.
This research aimed to determine the microbiological profile of antibiotic-resistant bacteria in wastewater samples collected from the "La Escalerilla" Wastewater Treatment Plant (WWTP) in Arequipa in April 2022. The isolation process involved serial dilutions of collected water samples in peptone water (Mesquita et al., 2021), followed by cultivation on MacConkey agar and blood agar plates (Mutuku, 2017). The identification of isolated colonies was carried out using Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry (MALDI-TOF MS) with the VITEK MS system (Biomérieux) through the Research Use Only (RUO) platform. A total of 126 bacterial colonies were isolated. The microbiological profiling was based on cultural characteristics, colony morphology, smears of isolated colonies, and Gram staining. The majority of the isolated colonies were Gram-negative bacilli (98.4%, 124), followed by Gram-negative coccobacilli (0.8%, 1), and Gram-positive cocci (0.8%, 1). Out of the 126 isolated colonies, 66.7% were identified as Enterobacteriaceae, encompassing species such as Citrobacter freundii, Citrobacter sp., Enterobacter cloacae, Enterobacter ludwigii, Enterobacter sp., Escherichia coli, Hafnia alvei, Klebsiella oxytoca, Klebsiella pneumoniae, Proteus mirabilis, Proteus vulgaris/penneri, Raoultella ornithinolytica, and Serratia liquefaciens. Additionally, 28.6% were identified as Aeromonas spp. including Aeromonas punctata (caviae), Aeromonas sp., Aeromonas hydrophila, and Aeromonas veronii. Pseudomonas spp. accounted for 2.4% of the isolated colonies, specifically Pseudomonas oleovorans and Pseudomonas cuatrocienegasensis. A smaller percentage of colonies were identified as Alcaligenes faecalis, Shewanella putrefaciens, and Enterococcus faecalis (0.8% each). Antibiotic susceptibility testing was performed using the Kirby-Bauer method on Mueller-Hinton agar, following the guidelines of the Clinical Laboratory Standards Institute (CLSI). The diameter of the inhibition zone determined the susceptibility as sensitive (S), intermediate (I), or resistant (R) based on the methodology by Hartinger et al. (2021). Resistance profiles were analyzed for each antibiotic. In Enterobacteriaceae, varying resistance patterns were observed, with higher resistance rates to ampicillin (59.5%), followed by sulfamethoxazole/trimethoprim (41.7%), and ciprofloxacin (40.5%). Among this group, Escherichia coli exhibited the highest number of resistant colonies. Among Aeromonadaceae, the greatest resistance was seen against sulfamethoxazole/trimethoprim (44.4%), followed by ciprofloxacin (33.3%) and ceftriaxone (30.6%). Within this family, Aeromonas sp. showed the highest number of resistant colonies. Notably, the Enterococcus faecalis colony demonstrated susceptibility to all five tested antibiotics. The findings of this research highlight the presence of antibiotic-resistant bacteria in the influent of the "La Escalerilla" WWTP in Arequipa.
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Keywords
Planta de tratamiento de aguas residuales, resistencia a antibióticos, Enterobacteriaceae