Diversity and enzymatic potential of thermophilic bacteria associated with terrestrial hot springs in Algeria

Abstract

This study aims to determine the diversity of culturable thermophilic bacteria isolated from eight terrestrial hot springs in Northeastern of Algeria using the conventional methods, SDS-PAGE fingerprinting of whole-cell proteins and 16S rRNA gene sequencing. In addition, their hydrolytic enzyme activities were also investigated. A total of 293 strains were isolated from the hot springs' water and sediment using different culture media. Overall, five distinct bacterial groups were characterized by whole-cell protein pattern analysis. Based on the 16S rRNA gene sequencing of 100 selected strains, the isolates were assigned to the following three major phyla: Firmicutes (93%), Deinococcus-Thermus (5%), and Actinobacteria (2%), which included 27 distinct species belonging to 12 different phylotypes, Aeribacillus, Aneurinibacillus, Anoxybacillus, Bacillus, Brevibacillus, Geobacillus, Laceyella, Meiothermus, Saccharomonospora, Thermoactinomyces, Thermobifida, and Thermus. The screening for nine extracellular enzymes showed that 65.87% of the isolates presented at least five types of enzyme activities, and 6.48% of strains combined all tested enzymes (amylase, cellulase, pectinase, esculinase, protease, gelatinase, lipase, lecithinase, and nuclease). It was found that Bacillus, Anoxybacillus, Aeribacillus, and Aneurinibacillus were the genera showing the highest activities. Likewise, the study showed an abundant and diverse thermophilic community with novel taxa presenting a promising source of thermozymes with important biotechnological applications. This study showed that a combined identification method using SDS-PAGE profiles of whole-cell proteins and subsequent 16S rRNA gene sequence analysis could successfully differentiate thermophilic bacteria from Algerian hot springs.

Keywords: 16S rRNA gene; Algeria; Biodiversity; Culture-dependent method; SDS-PAGE fingerprinting; Thermal spring; Thermophile; Thermozyme.

Fig. 1

Geographical location of the studied hot springs

Fig. 2

Dendrogram of similarity based on SDS-PAGE of whole-cell protein pattern analysis from thermophilic bacterial isolates. The right side of the figure presents the strain number and the clustering classification

Fig. 2

Dendrogram of similarity based on SDS-PAGE of whole-cell protein pattern analysis from thermophilic bacterial isolates. The right side of the figure presents the strain number and the clustering classification

Fig. 2

Dendrogram of similarity based on SDS-PAGE of whole-cell protein pattern analysis from thermophilic bacterial isolates. The right side of the figure presents the strain number and the clustering classification

Fig. 3

Neighbor-joining phylogenetic tree showing the relationship based on 16S rRNA gene sequence comparison of (a) Deinococcus-Thermus and (b) Actinobacteria. Bootstrap values (expressed as percentages of 1000 replications) greater than 50% are shown at nodes. The scale bar represents 2% estimated sequence divergence. The sequence of Bacillus thermoamylovorans was used as an outgroup

Fig. 4

Neighbor-joining phylogenetic tree showing the relationship between Firmicutes isolates (Paenibacillaceae and Thermoactinomycetaceae) and closely related species based on 16S rRNA gene sequence comparison. Bootstrap values expressed as percentages of 1000 replications; only bootstrap values > 50% are shown at nodes. The scale bar represents 2% estimated sequence divergence

Fig. 5

Neighbor-joining phylogenetic tree showing the relationship between Bacillaceae isolates and closely related species based on 16S rRNA gene sequence comparison. Bootstrap values expressed as percentages of 1000 replications; only bootstrap values > 50% are shown at nodes. The scale bar represents 2% estimated sequence divergence

Fig. 6

Venn diagram distribution illustrates the number of exclusive and common thermophilic bacterial isolates among the sampled sites. Each ellipse represents a sampling point

Fig. 7

Distribution of extracellular hydrolytic enzymes among isolated thermophilic bacteria. N, the number of isolates presents activities for each enzyme