MIC/MBC resistance fingerprints to As(III) in Bacillus and Pseudomonas as bioindicators across water and solid matrices in southern Perú

Arsenic (As) contamination in soils and waters is a critical challenge to human health, agricultural productivity, and ecological integrity. In soil-water systems, As can modify microbial community structure and physicochemical properties; therefore, indicators that integrate As availability and biological stress across heterogeneous matrices are needed. This study evaluated whether phenotypic As resistance patterns in environmental bacteria can be used as bioindicators and whether native, As-tolerant Bacillus and Pseudomonas stains could support recovery-oriented assessments.

Soil/sediment and water samples were collected at six sites across three high-Andean areas in southern Perú: Desaguadero (Puno; deep well and spring), Sicuani (Cusco; two springs), and Espinar (Cusco; Salado River). Solid matrices included riverbed sediments, saturated solids at spring outlets, and excavated well soils (drill cuttings/spoil around the wellhead). In soil/sediment, pseudo-total As was determined by aqua regia digestion. In waters, dissolved As, was quantified alter filtration (0.45 µm) and acidification (HNO₃ (pH < 2).

Tolerance assays were performed on nutrient agar amended with 100, 1000, 1500, and 2000 mg L^-1 As (III) at 30°C for 24-72 h to estimate the minimum inhibitory concentration (MIC). The minimum bactericidal concentration (MBC) was then determined by subculture in As-free broth (triplicate; OD₆₀₀). A total of 59 isolates were obtained: Bacillus (n = 40, water = 12, solid matrix = 28) and Pseudomonas (n = 19, water = 3, solid matrix = 16). Biochemical profiling assigned Bacillus to the B. cereus complex (cereus/thuringiensis), B. subtilis group, and Bacillus spp.; Pseudomonas to P. aeruginosa, P. stutzeri, P. mendocina and Pseudomonas spp.

Bacillus showed higher resistance than Pseudomonas: with growth observed in 37/40, 28/40, 20/40 and 4/40 isolates at 1000, 1500 and 2000 mg L^-1, respectively, and higher tolerance enriched in solid matrix isolates (1500 mg L^-1: 17/28 vs 3/12; 2000 mg L^-1: 4/28 vs 0/12). In Pseudomonas, growth occurred in 16/19, 9/19 3/19 and 0/19 isolates at the same concentrations. The most tolerant isolates were B2539 (Bacillus sp.; MBC = 2200 mg L^-1) and P2501 (P. aeruginosa), with an MBC = 1400 mg L^-1). These results support MIC/MBC “resistance fingerprints” as quantitative bioindicators to compare sites and matrices in As-affected environments.

Keywords: arsenic; microbial bioindicators; riverbed sediment; springs, soil; Bacillus; Psedomonas; souther Perú.