Evaluation of cellulolytic Bacillus isolates as animal probiotics and the effect of B. glycinifermentans SK 4275 on in-vitro fermentation of elephant grass

Table of Contents

Chapter 1. Introduction

1.1. Elephant grass as a feed resource for ruminants

1.2. Concerns of antibiotics as feed additives

1.3. The role of Bacillus strains as animal probiotics

1.4. Bacillus glycinifermentans and B. paralichineformis

1.5. Purpose of this study

Chapter 2. Material and Methods

2.1. Collection and preparation of elephant grass

2.2. Isolation of Bacillus strains from elephant grass

2.3. Screening for enzyme activity

2.4. Screening for probiotic properties of B. glycinifermentans SK4275 and B. Paralicheniformis SK4278 in vitro

2.4.1. Acid tolerance

2.4.2. Bile tolerance

2.4.3. Sodium chloride tolerance

2.4.4. Antimicrobial activity

2.4.5. Antibiotic sensitivity

2.4.6. Hemolytic activity

2.5. In vitro ruminal fermentation

2.5.1. Substrate, Bacillus and treatments

2.5.2. In vitro study

2.5.3. Sampling and analysis

2.6. Statistical analysis

Chapter 3 Results and discussion

3.1. Identification of isolated Bacillus strains

3.2. Enzyme activity

3.3. In vitro assessment of characteristics for survival in gastrointestinal tract

3.3.1. Acid tolerance

3.3.2. Bile tolerance

3.3.3. Antimicrobial activity

3.3.4. Sodium chloride tolerance

3.4. Biosafety assessment

3.4.1. Antibiotic sensitivity

3.4.2. Hemolytic activity

3.5. In vitro fermentation characteristics

3.5.1. pH, total gas and ammonia-N concentration

3.5.2. Volatile fatty acids

3.5.3. Total viable count in incubation fluid

3.5.4. In vitro NDF digestibility and ADF digestibility

3.6. Discussion of in vitro fermentation findings

Chapter 4. Conclusion

Research Objectives and Focus

This thesis aims to evaluate the probiotic potential and cellulolytic enzyme activity of two Bacillus strains, specifically B. glycinifermentans SK4275 and B. paralichineformis SK4278, isolated from elephant grass. The study further investigates the effects of B. glycinifermentans SK4275 on the in vitro fermentation and digestibility of elephant grass to determine its suitability as a probiotic feed additive for ruminants.

• Isolation and identification of cellulolytic Bacillus strains from elephant grass.
• In vitro assessment of probiotic properties including acid tolerance, bile tolerance, and safety (antibiotic sensitivity and hemolytic activity).
• Screening for hydrolytic enzyme activities (cellulase, amylase, xylanase, etc.).
• Evaluation of the impact of Bacillus supplementation on in vitro rumen fermentation parameters (pH, gas production, volatile fatty acids, and ammonia-N).
• Analysis of the influence of Bacillus addition on NDF and ADF digestibility of elephant grass.

Excerpt from the Book

Summary of Chapters

Chapter 1. Introduction: Discusses the importance of elephant grass as a ruminant feed and the limitations caused by its structural carbohydrates, alongside the potential role of Bacillus strains as probiotic alternatives to antibiotics.

Chapter 2. Material and Methods: Describes the procedures for isolating Bacillus strains from elephant grass, screening for their enzymatic and probiotic characteristics, and the methodology for conducting in vitro rumen fermentation trials.

Chapter 3 Results and discussion: Details the findings on bacterial identification, enzymatic potential, probiotic survivability, biosafety assessments, and the impact of Bacillus supplementation on fermentation characteristics and substrate digestibility.

Chapter 4. Conclusion: Summarizes that B. glycinifermentans SK4275 possesses favorable probiotic traits and cellulolytic activity, suggesting its potential as a beneficial additive for improving the utilization of elephant grass in ruminant nutrition.

Keywords

Bacillus glycinifermentans, elephant grass, in vitro fermentation, probiotics, cellulolytic enzyme activity, rumen digestibility, bacterial identification, acid tolerance, bile tolerance, biosafety, antibiotic sensitivity, ammonia-nitrogen, volatile fatty acids, NDF digestibility, ADF digestibility.

Frequently Asked Questions

What is the core subject of this thesis?

The thesis examines the potential of cellulolytic Bacillus strains, isolated from elephant grass, to act as probiotics that improve the fermentation and digestibility of ruminant feed.

What are the central thematic fields?

The research covers microbiology (isolation and identification of bacteria), probiotics, ruminant nutrition, and in vitro fermentation analysis.

What is the primary research goal?

The goal is to determine if selected Bacillus strains, specifically B. glycinifermentans SK4275, can improve the nutritional availability and fermentation efficiency of elephant grass through probiotic and cellulolytic actions.

What scientific methods were employed?

The study utilized 16S rRNA gene sequencing for identification, various microbiological assays for probiotic properties (acid, bile, and salt tolerance, antimicrobial and hemolytic activity), and standard in vitro rumen fermentation techniques (Tilley and Terry, 1963).

What topics are covered in the main section?

The main sections cover the isolation of bacteria, screening for enzyme activities (cellulase, amylase, xylanase, etc.), comprehensive safety assessments, and the measurement of fermentation characteristics like pH, gas production, volatile fatty acids, and fiber digestibility.

Which keywords characterize this work?

Key terms include Bacillus glycinifermentans, elephant grass, probiotics, in vitro fermentation, enzyme activity, and rumen digestibility.

How does B. glycinifermentans SK4275 impact rumen fiber digestibility?

The study demonstrates that the addition of B. glycinifermentans SK4275 significantly increased in vitro NDF and ADF digestibility of elephant grass at 24 and 48 hours of incubation.

Why is the biosafety of these Bacillus strains important?

Assessing biosafety is critical because certain Bacillus species are pathogenic. The research verified that these strains are susceptible to most antibiotics and do not exhibit hemolytic activity, ensuring their suitability as safe probiotics.