This study aimed to compare microscopic counting, culture, and quantitative or real-time PCR (qPCR) to quantify sulfate-reducing bacteria in environmental and engineered sludge samples. Four sets of primers that amplified the dsrA and apsA gene encoding the two key enzymes of the sulfate-reduction pathway were initially tested. qPCR standard curves were constructed using genomic DNA from an SRB suspension and dilutions of an enriched sulfate-reducing sludge. According to specificity and reproducibility, the DSR1F/RH3-dsr-R primer set ensured a good quantification based on dsrA gene amplification; however, it exhibited inconsistencies at low and high levels of SRB concentrations in environmental and sulfate-reducing sludge samples. Ultimately, we conducted a qPCR method normalized to dsrA gene copies, using a synthetic double-stranded DNA fragment as a calibrator. This method fulfilled all validation criteria and proved to be specific, accurate, and precise. The enumeration of metabolically active SRB populations through culture methods differed from dsrA gene copies but showed a plausible positive correlation. Conversely, microscopic counting had limitations due to distinguishing densely clustered organisms, impacting precision. Hence, this study proves that a qPCR-based method optimized with dsrA gene copies as a calibrator is a sensitive molecular tool for the absolute enumeration of SRB populations in engineered and environmental sludge samples.

Petroleum-contaminated soils are a major environmental concern worldwide. In Ecuador, extensive oil spills in the Amazon have led to widespread hydrocarbon pollution, threatening ecosystems and posing health risks to nearby communities. Conventional remediation techniques are resource-intensive and may render soil unsuitable for future use. In contrast, mycoremediation—using fungi to degrade toxic contaminants—offers a sustainable alternative. White-rot fungi, known for their ligninolytic enzyme systems such as laccases and peroxidases, are capable of degrading a wide range of organic pollutants, including petroleum hydrocarbons. This study assessed the enzymatic activity of 16 fungal strains from the phyla Ascomycota and Basidiomycota isolated in the Ecuadorian Amazon. Plate-based screening and quantitative laccase activity assays confirmed positive enzymatic activity in all strains. The five strains with the highest enzymatic activity were Ganoderma cf. parvulum QCAM7791, Trametes menziesii QCAM7783, Trametes menziesii QCAM7788, Trametes menziesii QCAM7790, and Trametes meyenii QCAM7785, which were selected for a 60-day soil microcosm experiment under controlled laboratory conditions. These strains removed over 96% of total petroleum hydrocarbons from contaminated soil, demonstrating high biodegradation efficiency. These results highlight the promise of native fungal strains as bioremediation agents for petroleum-contaminated soils. Further studies should focus on evaluating their performance under field conditions and their potential integration into large-scale remediation strategies.

Biological treatment using sulfate-reducing bacteria (SRB) is a promising approach to remediate acid rock drainage (ARD). Our purpose was to assess the performance of a sequential system consisting of a limestone bed filter followed by a sulfate-reducing bioreactor treating synthetic ARD for 375 days and to evaluate changes in microbial composition. The treatment system was effective in increasing the pH of the ARD from 2.7 to 7.5 and removed total Cu(II) and Zn(II) concentrations by up to 99.8% and 99.9%, respectively. The presence of sulfate in ARD promoted sulfidogenesis and changed the diversity and structure of the microbial communities. Methansarcina spp. was the most abundant amplicon sequence variant (ASV); however, methane production was not detected. Biodiversity indexes decreased over time with the bioreactor operation, whereas SRB abundance remained stable. Desulfobacteraceae, Desulfocurvus, Desulfobulbaceae and Desulfovibrio became more abundant, while Desulfuromonadales, Desulfotomaculum and Desulfobacca decreased. Geobacter and Syntrophobacter were enriched with bioreactor operation time. At the beginning, ASVs with relative abundance <2% represented 65% of the microbial community and 21% at the end of the study period. Thus, the results show that the microbial community gradually lost diversity while the treatment system was highly efficient in remediating ARD.

Pesticides are widely used in agriculture, floriculture specifically, posing significant ecological and health risks. Limited research has been conducted on the presence of neonicotinoid insecticides (NNI) and atrazine, in drinking and irrigation water in agricultural regions of Latin America. This study describes targeted and non-targeted analysis of concentrations of pesticides in drinking and irrigation water sampled in 2022 and 2023 in an industrial floricultural region in Pedro Moncayo, Ecuador. In targeted analysis, we found NNI in treated drinking water in 20.5% of household tap water samples (8 out of 39), in a community well, and in 57.7% of irrigation water samples (n = 15 out of 26). Imidacloprid and thiamethoxam emerged as the most frequently detected NNI in drinking water. Atrazine was detected in two household taps and one well. In exploratory non-targeted analysis, we found 63 compounds, including insecticides, herbicides, and fungicides, including azoles, in drinking and irrigation water samples. Fungicides accounted for 59% of all compounds (37 out of 63) detected. Intense pesticide use in floriculture in proximity to residential areas and aging piped water systems may allow pesticides to leach into treated drinking water pipes and irrigation water. The presence of a wide range of pesticides, especially NNI and fungicides (azoles, specifically), in drinking and irrigation water poses health risks to community members.

El objetivo del presente estudio fue construir y operar una cámara anaeróbica no costosa para el cultivo de bacterias sulfato reductoras (BSRs) en muestras ambientales de lodos. Una vez construida la cámara, las condiciones anaeróbicas al interior de la cámara fueron evaluadas a través del monitoreo de la concentración de oxígeno y la temperatura. En la cámara anaeróbica se alcanzaron las condiciones de anaerobiosis (< 0,1% O2) y la temperatura requerida (30ºC) para el cultivo de las BSRs. A continuación, se cultivó en placa (en un medio selectivo) cuatro muestras diferentes de lodo que fueron incubadas, contadas y microscópicamente observadas. Los resultados por cultivo en placa fueron comparados con los de contaje directo en cámara de Neubauer. Habiéndose demostrado que las condiciones ambientales en la cámara son las adecuadas para el cultivo de microorganismos anaeróbicos estrictos, las BSRs también fueron observadas mediante microscopía. En base a los resultados obtenidos por cultivo en placa y por recuento en cámara de Neubauer se determinó que no hubo diferencias significativas en la muestra de suspensión bacteriana, a diferencia de las muestras de lodos. Si bien es cierto que el cultivo de microorganismos demanda tiempo, esta técnica sigue siendo útil y confiable para el conteo microbiano. Finalmente, la cámara anaeróbica construida permite el cultivo de microorganismos anaeróbicos estrictos como las BSRs, responsables de la bioprecipitación de metales.