Identification and whole genome sequencing analysis of Bacillus subtilis K35-1, a highly efficient cellulose in forage degrading bacterium

Abstract This study reports the isolation and characterization of Bacillus subtilis K35-1, a novel cellulolytic strain with exceptional forage degradation capabilities. From eight B. subtilis isolates obtained from yak rumen fluid through Congo red screening (hydrolysis capacity = 2.61 ± 0.23), K35-...

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Main Authors: Dan Wu, Jing Feng, Dongxu Wen, Hongzhuang Wang, Sijia Lu, Kun Li, Rizwana Sultan, Bin Li
Format: Article
Language:English
Published: BMC 2025-07-01
Series:BMC Microbiology
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Online Access:https://doi.org/10.1186/s12866-025-04136-8
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author Dan Wu
Jing Feng
Dongxu Wen
Hongzhuang Wang
Sijia Lu
Kun Li
Rizwana Sultan
Bin Li
author_facet Dan Wu
Jing Feng
Dongxu Wen
Hongzhuang Wang
Sijia Lu
Kun Li
Rizwana Sultan
Bin Li
author_sort Dan Wu
collection DOAJ
description Abstract This study reports the isolation and characterization of Bacillus subtilis K35-1, a novel cellulolytic strain with exceptional forage degradation capabilities. From eight B. subtilis isolates obtained from yak rumen fluid through Congo red screening (hydrolysis capacity = 2.61 ± 0.23), K35-1 demonstrated superior enzymatic performance, achieving peak cellulase (77.26 U/mL) and hemicellulase (222.85 nmol/min/mL) activities at 36 h of fermentation. The whole genome sequencing revealed a 4.06 Mb circular chromosome (GC content 43.83%) encoding 3,980 protein-coding sequences. Comprehensive CAZy annotation identified 703 carbohydrate-active enzymes, including: 87 cellulases spanning 7 GH families (GH5, GH6, GH9, GH12, GH44, GH45, GH48) and 34 hemicellulases from 4 GH families (GH10, GH11, GH26, GH30). Comparative genomic analysis showed K35-1 possesses 40% more glycoside hydrolases than reference strains (Srivastava et al., Mol Genet Genomics 298:361–74, 2023), explaining its enhanced degradation efficiency (53.2% cellulose reduction vs. 7.3% in conventional treatments). Functional annotation revealed: 275 carbohydrate metabolism genes (KEGG), 228 cell wall/membrane biogenesis genes (COG) and 53.91% reduced-virulence mutations (PHI database). The strain’s robust enzymatic profile, coupled with minimal antibiotic resistance (11 genes, including ermB), positions K35-1 as both an efficient forage degrader and safe probiotic candidate. These findings provide a genomic foundation for developing novel feed additives to improve livestock nutrition.
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spelling doaj-art-8bb311fe56ad40c1864478794261a1bb2025-08-20T04:01:53ZengBMCBMC Microbiology1471-21802025-07-0125111010.1186/s12866-025-04136-8Identification and whole genome sequencing analysis of Bacillus subtilis K35-1, a highly efficient cellulose in forage degrading bacteriumDan Wu0Jing Feng1Dongxu Wen2Hongzhuang Wang3Sijia Lu4Kun Li5Rizwana Sultan6Bin Li7State Key Laboratory of Hulless Barley and Yak Germplasm Resources and Genetic Improvement, and Institute of Animal Husbandry and Veterinary Medicine, Tibet Academy of Agricultural and Animal Husbandry SciencesState Key Laboratory of Hulless Barley and Yak Germplasm Resources and Genetic Improvement, and Institute of Animal Husbandry and Veterinary Medicine, Tibet Academy of Agricultural and Animal Husbandry SciencesState Key Laboratory of Hulless Barley and Yak Germplasm Resources and Genetic Improvement, and Institute of Animal Husbandry and Veterinary Medicine, Tibet Academy of Agricultural and Animal Husbandry SciencesState Key Laboratory of Hulless Barley and Yak Germplasm Resources and Genetic Improvement, and Institute of Animal Husbandry and Veterinary Medicine, Tibet Academy of Agricultural and Animal Husbandry SciencesCollege of Veterinary Medicine, Nanjing Agricultural UniversityCollege of Veterinary Medicine, Nanjing Agricultural UniversityDepartment of Pathology, Faculty of Veterinary Science, Cholistan University of Veterinary and Animal Sciences (CUVAS)State Key Laboratory of Hulless Barley and Yak Germplasm Resources and Genetic Improvement, and Institute of Animal Husbandry and Veterinary Medicine, Tibet Academy of Agricultural and Animal Husbandry SciencesAbstract This study reports the isolation and characterization of Bacillus subtilis K35-1, a novel cellulolytic strain with exceptional forage degradation capabilities. From eight B. subtilis isolates obtained from yak rumen fluid through Congo red screening (hydrolysis capacity = 2.61 ± 0.23), K35-1 demonstrated superior enzymatic performance, achieving peak cellulase (77.26 U/mL) and hemicellulase (222.85 nmol/min/mL) activities at 36 h of fermentation. The whole genome sequencing revealed a 4.06 Mb circular chromosome (GC content 43.83%) encoding 3,980 protein-coding sequences. Comprehensive CAZy annotation identified 703 carbohydrate-active enzymes, including: 87 cellulases spanning 7 GH families (GH5, GH6, GH9, GH12, GH44, GH45, GH48) and 34 hemicellulases from 4 GH families (GH10, GH11, GH26, GH30). Comparative genomic analysis showed K35-1 possesses 40% more glycoside hydrolases than reference strains (Srivastava et al., Mol Genet Genomics 298:361–74, 2023), explaining its enhanced degradation efficiency (53.2% cellulose reduction vs. 7.3% in conventional treatments). Functional annotation revealed: 275 carbohydrate metabolism genes (KEGG), 228 cell wall/membrane biogenesis genes (COG) and 53.91% reduced-virulence mutations (PHI database). The strain’s robust enzymatic profile, coupled with minimal antibiotic resistance (11 genes, including ermB), positions K35-1 as both an efficient forage degrader and safe probiotic candidate. These findings provide a genomic foundation for developing novel feed additives to improve livestock nutrition.https://doi.org/10.1186/s12866-025-04136-8Bacillus subtilisCellulose degrading bacteriaWhole genome
spellingShingle Dan Wu
Jing Feng
Dongxu Wen
Hongzhuang Wang
Sijia Lu
Kun Li
Rizwana Sultan
Bin Li
Identification and whole genome sequencing analysis of Bacillus subtilis K35-1, a highly efficient cellulose in forage degrading bacterium
BMC Microbiology
Bacillus subtilis
Cellulose degrading bacteria
Whole genome
title Identification and whole genome sequencing analysis of Bacillus subtilis K35-1, a highly efficient cellulose in forage degrading bacterium
title_full Identification and whole genome sequencing analysis of Bacillus subtilis K35-1, a highly efficient cellulose in forage degrading bacterium
title_fullStr Identification and whole genome sequencing analysis of Bacillus subtilis K35-1, a highly efficient cellulose in forage degrading bacterium
title_full_unstemmed Identification and whole genome sequencing analysis of Bacillus subtilis K35-1, a highly efficient cellulose in forage degrading bacterium
title_short Identification and whole genome sequencing analysis of Bacillus subtilis K35-1, a highly efficient cellulose in forage degrading bacterium
title_sort identification and whole genome sequencing analysis of bacillus subtilis k35 1 a highly efficient cellulose in forage degrading bacterium
topic Bacillus subtilis
Cellulose degrading bacteria
Whole genome
url https://doi.org/10.1186/s12866-025-04136-8
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