Alginate-Bentonite Encapsulation of Extremophillic Bacterial Consortia Enhances Chenopodium quinoa Tolerance to Metal Stress

Abstract

This study explores the encapsulation in alginate/bentonite beads of two metal(loid)-resistant bacterial consortia (consortium A: Pseudomonas sp. and Bacillus sp.; consortium B: Pseudomonas sp. and Bacillus sp.) from the Atacama Desert (northern Chile) and Antarctica, and their influence on physiological traits of Chenopodium quinoa growing in metal(loid)-contaminated soils. The metal(loid) sorption capacity of the consortia was determined. Bacteria were encapsulated using ionic gelation and were inoculated in soil of C. quinoa. The morphological variables, photosynthetic pigments, and lipid peroxidation in plants were evaluated. Consortium A showed a significantly higher biosorption capacity than consortium B, especially for As and Cu. The highest viability of consortia was achieved with matrices A1 (3% alginate and 2% bentonite) and A3 (3% alginate, 2% bentonite and 2.5% LB medium) at a drying temperature of 25 °C and storage at 4 °C. After 12 months, the highest viability was detected using matrix A1 with a concentration of 10⁶ CFU g^-1. Further, a greenhouse experiment using these consortia in C. quinoa plants showed that, 90 days after inoculation, the morphological traits of both consortia improved. Chemical analysis of metal(loid) contents in the leaves indicated that consortium B reduced the absorption of Cu to 32.1 mg kg^-1 and that of Mn to 171.9 mg kg^-1. Encapsulation resulted in a significant increase in bacterial survival. This highlights the benefits of using encapsulated microbial consortia from extreme environments, stimulating the growth of C. quinoa, especially in soils with metal(loid) levels that can be a serious constraint for plant growth.

Keywords: bacterial storage; extremophilic bacteria; metal(loid)s biosorption; metal(loid)s toxicity alleviation.

Figure 1

(A): Biosorption and (B): removal efficiency of the metal(loid)s by consortium A and consortium B. CI: metal(loid) mix I. CII: metal(loid) mix II. AI: metal(loid) mix I with consortium A. AII: metals(loid) mix II with consortium A. BI: metal(loid) mix I with consortium B. BII: metals(loid) mix II with consortium B. Different letters above bars indicate significant differences among treatments (p < 0.05, n = 6).

Figure 2

Survival of bacterial consortia A and B in different matrices. Matrix (A1 alginate 3% bentonite 2%), matrix A2 (alginate 3% bentonite 2% glycerol 3%), matrix A3 (alginate 3%, bentonite 2%, LB medium 2.5%), and matrix A4 (alginate 3% molasses 3%). The beads were dried at 25, 28, 30, and 35 °C for 24 h. N_t is the number of viable cells after drying, N_i is the number of viable cells at time zero. The initial cell count was approximately 10⁹ CFU g⁻¹. The different letters indicate a statistically significant difference between the groups p < 0.05.

Figure 3

Viability of bacterial consortia A and B after 12 months of storage at 4 and 24 °C using different encapsulation matrices. Matrix A1 (3% alginate and 2% bentonite) and matrix A3 (3% alginate, 2% bentonite, and 2.5% LB medium). The initial cell count was approximately 10⁸ CFU g⁻¹. Error bars indicate the standard deviation of 6 independent replicates. Lowercase letters indicate a significant difference between temperatures for each treatment and uppercase letters indicate a significant difference between matrices; p < 0.05.

Figure 4

Spearman’s rank correlation analysis between consortium, matrix, metal(loid) mix, and morphological traits of C. quinoa. The cells are colored according to the correlation coefficient. Blue represents a significant positive correlation and red represents a significant negative correlation.

Figure 5

Total chlorophyll content in C. quinoa plants WC: without consortium, CA: with consortium A, and CB: with consortium B; matrix A1 (3% alginate and 2% bentonite) and matrix A3 (3% alginate, 2% bentonite, and 2.5% LB medium) in different concentrations of metal(loid)s. (a) Control: soil without the metal(loid) mix; (b) metal(loid) mix I, and (c) metal(loid) mix II. Error bars indicate the standard deviation of 6 independent replicates. Lower-case letters indicate a significant difference between matrices for each treatment and upper-case letters indicate a significant difference between treatments p < 0.05.

Figure 6

Malondialdehyde (MDA) content in leaves and roots of C. quinoa plants. WC: without consortium, CA: with consortium A and CB: with consortium B; Matrix A1 (3% alginate and 2% bentonite) and matrix A3 (3% alginate, 2% bentonite, and 2.5% LB medium) in different concentrations of metal(loid)s. (a,d) Control: soil without the metal(loid) mix; (b,e) metal(loid) mix I and (c,f) metal(loid) mix II. Error bars indicate the standard deviation of 6 independent replicates. Lower-case letters indicate a significant difference between matrices for each treatment and upper-case letters indicate a significant difference between treatments p < 0.05.