Genomics and cellulolytic, hemicellulolytic, and amylolytic potential of Iocasia fonsfrigidae strain SP3-1 for polysaccharide degradation

Abstract

Background Cellulolytic, hemicellulolytic, and amylolytic (CHA) enzyme-producing halophiles are understudied. The recently defined taxon Iocasia fonsfrigidae consists of one well-described anaerobic bacterial strain: NS-1T. Prior to characterization of strain NS-1T, an isolate designated Halocella sp. SP3-1 was isolated and its genome was published. Based on physiological and genetic comparisons, it was suggested that Halocella sp. SP3-1 may be another isolate of I. fronsfrigidae. Despite being geographic variants of the same species, data indicate that strain SP3-1 exhibits genetic, genomic, and physiological characteristics that distinguish it from strain NS-1T. In this study, we examine the halophilic and alkaliphilic nature of strain SP3-1 and the genetic substrates underlying phenotypic differences between strains SP3-1 and NS-1T with focus on sugar metabolism and CHA enzyme expression. Methods Standard methods in anaerobic cell culture were used to grow strains SP3-1 as well as other comparator species. Morphological characterization was done via electron microscopy and Schaeffer-Fulton staining. Data for sequence comparisons (e.g., 16S rRNA) were retrieved via BLAST and EzBioCloud. Alignments and phylogenetic trees were generated via CLUTAL_X and neighbor joining functions in MEGA (version 11). Genomes were assembled/annotated via the Prokka annotation pipeline. Clusters of Orthologous Groups (COGs) were defined by eegNOG 4.5. DNA-DNA hybridization calculations were performed by the ANI Calculator web service. Results Cells of strain SP3-1 are rods. SP3-1 cells grow at NaCl concentrations of 5-30% (w/v). Optimal growth occurs at 37 °C, pH 8.0, and 20% NaCl (w/v). Although phylogenetic analysis based on 16S rRNA gene indicates that strain SP3-1 belongs to the genus Iocasia with 99.58% average nucleotide sequence identity to Iocasia fonsfrigida NS-1T, strain SP3-1 is uniquely an extreme haloalkaliphile. Moreover, strain SP3-1 ferments D-glucose to acetate, butyrate, carbon dioxide, hydrogen, ethanol, and butanol and will grow on L-arabinose, D-fructose, D-galactose, D-glucose, D-mannose, D-raffinose, D-xylose, cellobiose, lactose, maltose, sucrose, starch, xylan and phosphoric acid swollen cellulose (PASC). D-rhamnose, alginate, and lignin do not serve as suitable culture substrates for strain SP3-1. Thus, the carbon utilization profile of strain SP3-1 differs from that of I. fronsfrigidae strain NS-1T. Differences between these two strains are also noted in their lipid composition. Genomic data reveal key differences between the genetic profiles of strain SP3-1 and NS-1T that likely account for differences in morphology, sugar metabolism, and CHA-enzyme potential. Important to this study, I. fonsfrigidae SP3-1 produces and extracellularly secretes CHA enzymes at different levels and composition than type strain NS-1T. The high salt tolerance and pH range of SP3-1 makes it an ideal candidate for salt and pH tolerant enzyme discovery.

Hydrolysis of ionic liquid–treated substrate with an Iocasia fonsfrigidae strain SP3-1 endoglucanase

“…In this study, we demonstrate that a recently reported endoglucanase, designated IfCelS12A (Heng et al 2022 ), from the haloalkaliphile Iocasia fonsfrigidae strain SP3-1 (Heng et al 2019 , 2022 ) used in conjunction with a cocktail of enzymes from Clostridium thermocellum and in the presence of a mobile enzyme sequestration platform (MESP) (Mitsuzawa et al 2009 ; Ceballos et al 2014 ; Ceballos 2023 ) enhances enzymatic breakdown of multiple substrates including rice straw (RS) in IL. Our data further demonstrate that IfCelS12A has a mode of action distinct from other glycoside hydrolases within the GH12 family of enzymes.…”

“…Discovery and characterization of cellulolytic enzymes with unique attributes, such as stability at elevated temperatures and tolerance to low pH, is an active area of research in applied microbiology (e.g., Elleuche et al 2015 ; Husain 2017 ; Thapa et al 2020 ; Dadwal et al 2021 ; Gan et al 2022 ). For this study, we characterized an endoglucanase (i.e., IfCelS12A) derived from the halophile I. fonsfrigidae strain SP3-1 isolated from a hypersaline pond in Thailand (Heng et al 2022 ).…”

Screening, identification, and mechanism analysis of starch-degrading bacteria during curing process in tobacco leaf

“…The bacteria of Bacillus can secrete various amylases, which can degrade starch and produce organic acids ( Ma et al, 2023 ). Moreover, it has been found that certain anaerobic bacteria within the Anaeromyxobacter genus can degrade starch into organic acids and gases, such as acetic acid, propionic acid, butyric acid, hydrogen, and carbon dioxide ( Heng et al, 2022 ). The production of these organic acids and gases was found to further promote starch degradation.…”

A Novel D-Psicose 3-Epimerase from Halophilic, Anaerobic Iocasia fonsfrigidae and Its Application in Coconut Water

“…Iocasia is a new genus of the family Halanaerobiaceae . Thus far, only one species with two strains, Iocasia fonsfrigidae strain NS-1 and strain SP3-1, has been reported [ 10 ]. Although these two strains have no confirmed toxicity, strain SP3-1 (this study) showed the highest similarity with another close-up genus in the family Halanaerobiaceae , namely, Halocella cellulosilytica DSM7362 T , at 92.7%.…”

“…Iocasia fonsfrigidae strain SP3-1 was isolated from a salt evaporation pond as an extremely halophilic, anaerobic bacterium that grows well in a medium containing 20% ( w / v ) NaCl. The whole genome sequence of this bacterium revealed the presence of a possible DPEase gene [ 10 ] for which the amino acid sequence of the DPEase from I. fonsfrigidae strain SP3-1 (IfDPEase) has a low similarity to other ketose 3-epimerase members. Therefore, we speculated that the IfDPEase may represent a new function in the ketose 3-epimerase family.…”