Next Article in Journal
The Intriguing Biogeographic Pattern of the Italian Wall Lizard Podarcis siculus (Squamata: Lacertidae) in the Tuscan Archipelago Reveals the Existence of a New Ancient Insular Clade
Previous Article in Journal
A Comparison of Semilandmarking Approaches in the Visualisation of Shape Differences
 
 
Search for Articles:
Title / Keyword
Author / Affiliation / Email
Journal
Article Type
 
 
Advanced Search
 
Section
Special Issue
Volume
Issue
Number
Page
 
Journals
Animals
Volume 13
Issue 3
10.3390/ani13030384
animals-logo
Submit to this Journal Review for this Journal Propose a Special Issue
► Article Menu

Article Menu

share Share announcement Help format_quote Cite question_answer Discuss in SciProfiles thumb_up
...
Endorse textsms
...
Comment
first_page
settings
Order Article Reprints
Font Type:
Arial Georgia Verdana
Font Size:
Aa Aa Aa
Line Spacing:
Column Width:
Background:
Communication

Effect of Inoculating Two Bacteriocin-Producing Lactiplantibacillus plantarum Strains at Ensiling on In Vitro Rumen Fermentation and Methane Emissions of Alfalfa Silage with Two Dry Matter Contents

by
Ziqian Li
1,2,
Fuhou Li
1,2,
Zohreh Akhavan Kharazian
1 and
Xusheng Guo
1,2,*
1
State Key Laboratory of Grassland Agro-Ecosystems, School of Life Sciences, Lanzhou University, Lanzhou 730013, China
2
Probiotics and Biological Feed Research Center, Lanzhou University, Lanzhou 730013, China
*
Author to whom correspondence should be addressed.
Animals 2023, 13(3), 384; https://doi.org/10.3390/ani13030384
Submission received: 8 December 2022 / Revised: 13 January 2023 / Accepted: 14 January 2023 / Published: 23 January 2023
(This article belongs to the Topic Rumen Fermentation, Microbial Population and Digestibility in Ruminants)
Browse Figure
Versions Notes

Abstract

:

Simple Summary

Developing alternative antibiotics is crucial for sustainable animal production worldwide. Bacteriocin has been considered a potential alternative to antibiotics due to its lack of drug resistance, lack of residues, and environmental friendliness. In this study, we investigated the effects of inoculating two bacteriocin-producing strains, Lactiplantibacillus plantarum ATCC14917 and LP1-4, at ensiling on the in vitro ruminal fermentation characteristics and methane emissions of alfalfa silage with two different dry matter contents (355 g/kg fresh weight, moderate dry-matter content; 428 g/kg fresh weight, high dry-matter content). The results showed that inoculating with ATCC14917 and LP1-4 at ensiling reduced in vitro rumen methane production and enhanced the dry matter digestibility of ensiled alfalfa. Therefore, bacteriocin-producing L. plantarum ATCC14917 and LP1-4 inoculants can potentially mitigate ruminal methane emissions but without an adverse effect on the rumen fermentation of the inoculated alfalfa silage.

Abstract

The objective of this study was to investigate the effects of inoculating two bacteriocin-producing strains, Lactiplantibacillus plantarum ATCC14917 and LP1-4, at ensiling on the in vitro ruminal fermentation characteristics and methane production of alfalfa silage with two dry matter (DM)contents. Before ensiling, fresh alfalfa was wilted to a moderate DM content (355 g/kg) and a high DM content (428 g/kg). The wilted alfalfa was treated with (1) distilled water (control), (2) commercial strain L. plantarum MTD/1 (MTD/1), (3) bacteriocin-producing L. plantarum ATCC14917 (ATCC14917), and (4) a bacteriocin-like substance producing L. plantarum LP1-4 (LP1-4) at 1 × 105 colony forming units (CFU)/g fresh weight. After 90 d of ensiling, the silages were used for in vitro rumen fermentation. Inoculation with the two bacteriocin-producing strains at ensiling remarkably reduced (p < 0.05) in vitro ruminal CH4 production and enhanced DM digestibility compared with the control group regardless of DM content. For silages with high DM content, inoculation with the bacteriocin-producing strains even increased (p < 0.05) in vitro ruminal total volatile fatty acid production. Therefore, the bacteriocin-producing inoculants have a great potential to mitigate ruminal methane emission but without an adverse effect on rumen fermentation of the inoculated alfalfa silage.

1. Introduction

It is known that the greenhouse effect of methane (CH4) is nearly 25 times that of carbon dioxide (CO2) [1]. Ruminant CH4 emissions account for 16% of global greenhouse gases and 33% of global anthropogenic CH4 emissions [2]. As reported in a previous study, 75 million tons (Tg) of CH4 was discharged from the gut of cattle around the world in 2004, and about 9 Tg of CH4 was discharged from small ruminants such as sheep and goats [3]. In addition, about 87% of CH4 in ruminants originates from the rumen and 13% from the hindgut [4,5]. Therefore, how to reduce CH4 emissions from ruminants without affecting feed digestion is always the focus worldwide.
Antibiotics have been shown to reduce ruminal CH4 emissions, but antibiotic residues in animal products and the emergence of antibiotic-resistance genes in the environment have led to the worldwide prohibition of antibiotics in animal husbandry [6,7,8]. Bacteriocins are antimicrobial peptides produced by bacteria and can accelerate the death of homologous bacteria by disrupting the lipid II metabolism of bacterial cell membranes or inhibiting the replication of nucleic acids [9,10]. They have been considered a potential alternative to antibiotics due to their lack of drug resistance, lack of residues, and environmental friendliness. Many lactic acid bacteria (LAB) produce bacteriocins. Some researchers have reported that adding bacteriocin alone in rumen culture or inoculating LAB without bacteriocin-producing ability in silage has obvious effects on reducing ruminant CH4 emissions [6,11]. Thus, the authors hypothesized that inoculating bacteriocin-producing LAB in silage probably exhibits the same role as the addition of bacteriocin alone in reducing in vitro ruminal methane emissions and improving the silage quality. However, few studies have investigated the effect of bacteriocin-producing LAB on silage fermentation and subsequent rumen digestion and CH4 emission.
Thus, this study investigated the impact of two bacteriocin-producing Lactiplantibacillus plantarum inoculants on in vitro ruminal digestibility, fermentation characteristics, and methane production of alfalfa silage with two dry matter (DM) contents. This provided a theoretical basis for the application of bacteriocin-producing L. plantarum instead of antibiotics in reducing the CH4 emission of ruminants.

2. Materials and Methods

2.1. Strain Preparation

The inoculants used in the present study included (1) L. plantarum MTD/1 (NCIMB 40027), a non-bacteriocin-producing silage inoculant, purchased from Ecosyl Products Ltd., Stokesley, UK; (2) L. plantarum ATCC14917, a class IIa bacteriocin-producing strain that was purchased from American Type Culture Collection [12]; (3) L. plantarum LP1-4, a bacteriocin-like-substance-producing strain, isolated and screened from turbot intestine originally sampled in Jinzhou, China [13]. The three strains were activated twice with a 1% (v/v) inoculation rate in MRS broth and cultured at 37 °C for 18 h before silage making.

2.2. Alfalfa Silage Preparation

Alfalfa (Medicago sativa.) was cultivated and manually harvested from four experimental fields in Dingxi (N35°58′, E104°62′), Gansu province, China, on July 2022. The harvested alfalfa from each field was divided into two portions and naturally wilted to DM contents of 355 (moderate DM content) and 428 (high DM content) g/kg fresh weight (FW), respectively. The water-soluble carbohydrates, crude protein, neutral detergent fiber (aNDF; using heat-stable α-amylase), and acid detergent fiber (ADF) contents of moderate and high DM were 31.5 vs. 36.7, 177 vs. 172, 366 vs. 391, and 283 vs. 294 g/kg DM, respectively. The wilted forages from four harvested fields at each DM content served as four experimental replicates. After that, the forages were chopped into small pieces, roughly 1−2 cm in length. The forage from each of the four fields was then treated with the following treatments: (1) distilled water (control), (2) L. plantarum MTD/1 (MTD/1), (3) L. plantarum ATCC14917 (ATCC 14917), and (4) L. plantarum LP1-4 (LP1-4) at 1 × 105 colony forming units (CFU)/g FW. Before the experiment, each LAB culture was centrifuged at 8000× g for 5 min after 24 h of cultivation, and the precipitation was suspended in sterile water to achieve an application rate of 2 × 107 CFU of viable cells/mL and subsequently evenly sprayed onto the chopped alfalfa at 5 mL/500 g FW. An equal volume of sterile water was used to treat the control. Each treated pile (approximately 500 g) was single-sealed in a plastic vacuum bag (density 0.91 to 0.93 g/cm3; vacuum degree 0.1 Mpa) and fermented for 90 d.

2.3. Lactic Acid Fermentation Characteristics and Fiber Analysis

On the designated day, a 20 g sample (including fresh samples and silages) was taken from each silo randomly, soaked with 180 mL sterile water for 24 h at 4 °C, and filtered with four layers of sterile gauze. The pH of the silage filtrate was instantly determined with a pH meter (PHSJ-3F, CANY, Shanghai, China). Lactic acid (LA) was measured according to the method of Yang et al. [14]. Approximately 20 g of fresh and silage samples were oven-dried at 65 °C for 72 h to determine DM, aNDF, and ADF contents using the methods described by Van Soest et al. [15].

2.4. In Vitro Rumen Fermentation

After 90 d of fermentation, the silages with two DM contents were subject to in vitro ruminal fermentation trials. Three rumen-fistulated Hu sheep (60 ± 5 kg) were used as rumen fluid donors for in vitro culture. The sheep were rationed twice a day (08:00 and 18:00) with a total mixed ration (TMR) pellet containing 58% corn, 19% wheat bran, 18% soybean meal, 1% baking soda, and 4% vitamin and mineral supplement. The collected rumen fluid was filtered using four layers of sterile gauze, equal volume mixed and poured into sterilized bottles (1500 mL) preheated at 39 °C. CO2 was immediately infused into the sterilized bottle filled with filtrate to eliminate air, and the filtrate was transported to the laboratory within 20 min. The artificial buffer solution was prepared according to Menke and Steingass [16], with each 1 L artificial buffer solution (pH 7.0) containing 237 mL buffer solution (4.0 g/L NH4HCO3 +35.0 g/L NaHCO3), 237 mL macro element solution (5.7 g/L Na2HPO4 + 6.2 g/L KH2PO4), 0.12 mL trace element solution (13.2 g/100 mL CaCl2∙2H2O + 10.0 g/100 mL MnCl2∙2H2O + 1.00 g/100 mL CoCl2∙6H2O + 8.00 g/100 mL FeCl3 + 6H2O), 1.22 mL resazurin solution (100 mg/100 mL), 50 mL reducing agent solution (285 mg/50 mL Na2S∙7H2O + 800 mg/50 mL NaOH), and 474 mL distilled water [16]. Before blending with filtrate in a ratio of approximately 4:1 (v/v), the artificial buffer was preheated to 39 °C and continuously infused with CO2. The crushed silage samples from each silo were prepared in quadruplicate (a total of 128 subsamples, 16 replicates by a treatment). A 0.5 g sample was weighed and put into a fiber bag (F57, Ankom Technology, New York, NY, USA) and heat-sealed. Another three bags were used as a blank. All bags were pre-dried to constant weight and put into 100 mL sterile glass sealed bottles. Half of the bottles (two bottles in each silage replicate) were linked with a microbial fermentation gas production automatic recorder (Boxiang Xingwang Technology Co., Ltd., Beijing, China) and filled with 90 mL of blended in vitro fermentation solution (20 mL filtrate and 70 mL artificial buffer solution) to monitor total gas production. The remaining two bottles were linked with gas-collecting bags (500 mL) for subsequent CH4 analysis. All bottles were cultured for 48 h at 39 °C.
After incubation, the fiber bags were washed with warm water until colorless and dried at 105 °C for 3 h before calculating the in vitro DM digestibility (IVDMD). Subsequently, 0.2 mL of 25% H3PO4 containing 2-methyl butyric acid was added to 1 mL of incubated fluid for volatile fatty acid (VFA) analysis using gas chromatography (trace 1300, Thermo Fisher Scientific Inc., Singapore) with an electrical conductivity detector and capillary column (30 m × 0.32 × mm 0.50 × µm; Lan-zhou Zhongke Kaidi Chemical New Technology Co., Ltd., China) according to the method described by Chen et al. [17]. The ammonia (NH3) concentration in the incubated fluid was determined according to the method of Broderick and Kang [18]. The gas collected by the gas collecting bags was used for the determination of CH4 content using gas chromatography with a flame ionization detector and 19095P-QO3 column (30 m × 0.53 mm × 40.00 µm; Agilent Technologies Inc., Santa Clara, CA, USA) according to the method of Chen et al. [17].

2.5. Statistical Analysis

The data of silage LA fermentation, fiber content, and in vitro ruminal fermentation parameters were analyzed using the general linear model procedure of SPSS 20.0 (IBM Co., Armonk, NY, USA) according to 2 (DM) × 4 (treatment) experimental design with the fixed factors of treatment, DM content, and their interaction. DM’s effects within each treatment were analyzed using Turkey’s multiple comparisons when the interaction was significant at p < 0.05.

3. Results

3.1. Lactic Acid Fermentation Characteristics and Fiber Contents of Silages at Moderate and High DM Contents after 90 Days

There were A × D interactions (p < 0.05) for pH and ADF (Figure 1a,d). All inoculants reduced (p < 0.05) silage pH compared with the control group, and ATCC14917 had the lowest (p < 0.05) pH value at moderate DM content, while MTD/1 and ATCC14917 with the high DM content had a lower (p < 0.05) pH value than the control group and LP1-4, respectively. Compared with control, ADF contents were lower (p < 0.05) in the two bacteriocin-producing LAB treatments at moderate DM level, while at high DM silage, ADF contents in MTD/1 and ATCC14917 groups were lower (p < 0.05) than that in the other groups. On average, LA concentration was higher (p < 0.05) in moderate DM silages versus high DM silages (25.8 vs. 20.6 g/kg DM), and ATCC14917 had the highest (p < 0.05) LA concentration regardless of silage DM level (Figure 1b). All inoculants decreased (p < 0.05) aNDF contents for both DM contents compared with the control groups (Figure 1c).

3.2. CH4 Production and Fermentation Characteristics of In Vitro Ruminal Fermentation of Alfalfa Silage

There were A × D interactions (p < 0.05) for total gas, CH4, CH4-to-total gas, in vitro IDVMD, total VFA, butyrate, and valerate (Table 1). Silages ensiled at moderate DM content increased (p < 0.05) CH4-to-total gas ratio, acetate-to-propionate ratio, IDVMD, CH4, total VFA, acetate, butyrate, isobutyrate, and isovalerate concentrations compared with the silages at high DM content regardless of treatments. All inoculants reduced (p < 0.05) CH4 concentrations compared with the control group at both DM contents; ATCC4917 with the moderate DM content and the two bacteriocin-producing LAB treatments with the high DM content had lower (p < 0.05) CH4 concentrations than MTD/1. The two bacteriocin-producing LAB decreased (p < 0.05) CH4-to-total gas ratio compared with the control group at moderate DM content, while a lower (p < 0.05) CH4-to-total gas ratio was observed in all inoculants at high DM content. The CH4-to-total gas ratio in ATCC4917 with moderate DM content and the two bacteriocin-producing LAB groups with high DM content was lower (p < 0.05) than that in MTD/1. IDVMD was increased (p < 0.05) in all inoculated silages with moderate DM content and the two bacteriocin-producing LAB treated-silages with high DM content. Acetate concentration and acetate to propionate ratio in the two bacteriocin-producing LAB treatments were lower (p < 0.05) than in the control group at both DM contents. LP1-4 with the moderate DM content and the two bacteriocin-producing LAB with the high DM content increased (p < 0.05) propionate concentrations compared with the control group. Butyrate concentrations in LP1-4 with the moderate DM content and the two bacteriocin-producing LAB treated-silages with the high DM content were lower (p < 0.05) than in the control group. Compared with the control group, inoculating the two bacteriocin-producing LAB in silage increased (p < 0.05) the ruminal isobutyrate and isovalerate concentrations only at high DM content. Valerate concentration in the two bacteriocin-producing LAB treated-silages was lower (p < 0.05) than that in the control group at moderate DM content, while an increased (p < 0.05) valerate concentration was observed in the two bacteriocin-producing LAB treatments at high DM content.

4. Discussion

The present study showed that alfalfa silages ensiled at high DM content increased pH values and decreased LA concentrations compared with the silages at moderate DM content regardless of treatments, consistent with previous studies [19,20,21]. Moreover, the lowest pH and the highest LA concentration in ATCC14914 regardless of the DM contents, showed that the silage fermentation effect of the ATCC14917 was the most desirable. In addition, the fiber contents in all inoculant treatments, regardless of DM contents, can be explained by the impact of pH because low pH can enhance the ability of plant cellulase to degrade structural carbohydrates during ensiling [22,23].
The in vitro ruminal fermentation trial showed that IDVMD, CH4, acetate, and butyrate concentrations with moderate DM content were higher than those with high DM content regardless of treatments, which contributed to the higher aNDF content in moderate DM content versus high DM content (393 vs. 387 g/kg DM) because high fiber content determined the increase in in vitro ruminal acetate concentration, accompanied by the increase in CH4 production [24]. Moreover, elevated acetate concentration at moderate DM content, regardless of treatments, may suggest an enhancement of cellulose bacteria’s role in acetate fermentation, thereby increasing digestibility [24]. As expected, ATCC14917 and LP1-4 reduced CH4 productions by 68.8% and 43.9% at moderate DM content, respectively, and they reduced CH4 productions by 63.6% and 78.6% at high DM content, respectively. This was consistent with the results of the application of LAB or nisin in vitro trials [6,11]. Moreover, the two bacteriocin-producing strains enhanced the forage DM digestibility at both DM contents. However, Shen et al. [6] reported that nisin decreased CH4 production without increasing digestibility, which was distinct from the result of inoculation with LAB. Oskoueian et al. [25] showed that L. plantarum, a silage inoculant, could diminish CH4 emissions and improve DM digestibility. Generally, a decrease in CH4 production is usually accompanied by a reduction in digestibility [24]. In previous studies, the inhibition of the ruminal fibrolytic bacteria such as Ruminococcus flavefaciens and Ruminococcus albus by nisin resulted in an inhibition of hydrogen forming through the acetyl-coA pathway, thereby reducing methane emission [26,27]. However, ATCC14917 in this study could produce a large amount of LA in addition to bacteriocins during ensiling, thereby probably enhancing the electron-accepting reaction in the rumen [28]. This allows the metabolic hydrogen produced by glucose during rumen fermentation to synthesize VFA (mainly propionate) instead of being utilized by methanogens to generate CH4 [29]. This can be confirmed by the higher in vitro ruminal propionate concentration, lower acetate and butyrate concentrations and acetate to propionate ratio in ATCC14917 treatment. Previous studies also confirmed that LA produced by L. plantarum during ensiling could promote the availability of hydrogen via the acrylate metabolic pathway to produce more propionate, which leads to a reduction in the acetate and butyrate metabolic pathway for CH4 production [30,31]. The increased isobutyrate and isovalerate concentrations in the two bacteriocin-producing LAB treatments at high DM content were consistent with the increased DM digestibility because the increase in branched-chain fatty acid (BCVFA) could enhance the digestion capacity of fibrobacteria, thereby improving DM digestibility [32]. Moreover, all inoculants and the two bacteriocin-producing strains decreased aNDF and ADF, respectively, which also improved the DM digestibility of in vitro fermentation [33]. In addition, the increased BCVFA concentrations in the two bacteriocin-producing LAB treatments at high DM content also imply enhanced deaminations of amino acids [24]. However, the NH3 concentration in the present study remained constant in the two treatments, and the NH3 concentration in the in vitro rumen only depends on the balance between the rate of formation and utilization of NH3 by microorganisms, suggesting that microbial protein synthesis was increased.
The bacteriocin concentration during silage fermentation was not determined in the present study. In future research, the qualitative or quantitative analysis of bacteriocin concentration in silages and rumen-incubated fluid will be checked by using high-performance liquid chromatography/mass spectrometry (HPLC-MS)

5. Conclusions

ATCC14917 and LP1-4 had better effects than MTD/1 in reducing in vitro rumen methane production and improving the DM digestibility of alfalfa silage. Therefore, the bacteriocin-producing L. plantarum ATCC14917 and LP1-4 inoculants have a great potential to be used as silage additives to improve silage fermentation quality and mitigate methane emissions but without an adverse effect on rumen fermentation of the inoculated silage.

Author Contributions

Conceptualization, X.G.; methodology, Z.L.; validation, X.G. and Z.L.; formal analysis, Z.L.; investigation, Z.L.; data curation, Z.L.; writing—original draft preparation, Z.L.; writing—review and editing, X.G., F.L. and Z.A.K.; visualization, X.G.; supervision, X.G.; project administration, X.G.; funding acquisition, X.G. All authors have read and agreed to the published version of the manuscript.

Funding

This work was financially supported by the National Natural Science Foundation of China (31872417).

Institutional Review Board Statement

Not applicable.

Informed Consent Statement

Not applicable.

Data Availability Statement

The data presented in this study are available upon request from the corresponding author.

Conflicts of Interest

The authors declare no conflict of interest.

References

  1. Zhou, Y.; Mao, H.; Jiang, F.; Wang, J.; Liu, J.; Mcsweeney, C.S. Inhibition of rumen methanogenesis by tea saponins with reference to fermentation pattern and microbial communities in Hu sheep. Anim. Feed Sci. Technol. 2011, 166–167, 93–100. [Google Scholar] [CrossRef]
  2. Scheehle, E.A.; Kruger, D. Global anthropogenic methane and nitrous oxide emissions. Energy J. 2006, 27, 33–44. [Google Scholar] [CrossRef]
  3. McMichael, A.J.; Powles, J.W.; Butler, C.D.; Uauy, R. Energy and health 5-Food, livestock production, energy, climate change, and health. Lancet 2007, 370, 1253–1263. [Google Scholar] [CrossRef]
  4. Murray, R.M.; Bryant, A.M.; Leng, R.A. Rates of production of methane in the rumen and large intestine of sheep. Br. J. Nutr. 1976, 36, 1–14. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  5. Lan, W.; Yang, C. Ruminal methane production: Associated microorganisms and the potential of applying hydrogen-utilizing bacteria for mitigation. Sci. Total Environ. 2019, 654, 1270–1283. [Google Scholar] [CrossRef] [PubMed]
  6. Shen, J.; Liu, Z.; Yu, Z.; Zhu, W. Monensin and nisin affect rumen fermentation and microbiota differently in vitro. Front. Microbiol. 2017, 8, 1111. [Google Scholar] [CrossRef] [Green Version]
  7. Michael, C.A.; Dominey-Howes, D.; Labbate, M. The Antimicrobial resistance crisis: Causes, consequences, and management. Front. Public Health 2014, 2, 145. [Google Scholar] [CrossRef] [PubMed]
  8. Gillor, O.; Nigro, L.M.; Riley, M.A. Genetically engineered bacteriocins and their potential as the next generation of antimicrobials. Curr. Pharm. Design 2005, 11, 1067–1075. [Google Scholar] [CrossRef] [Green Version]
  9. Santos, J.C.; Sousa, R.C.; Otoni, C.G.; Moraes, A.R.; Souza, V.G.; Medeiros, E.A.; Espitia, P.J.; Pires, A.C.; Coimbra, J.S.; Soares, N.F. Nisin and other antimicrobial peptides: Production, mechanisms of action, and application in active food packaging. Innov. Food Sci. Emerg. 2018, 48, 179–194. [Google Scholar] [CrossRef]
  10. Cleveland, J.; Montville, T.J.; Nes, I.F.; Chikindas, M.L. Bacteriocins: Safe, natural antimicrobials for food preservation. Int. J. Food Microbiol. 2001, 71, 1–20. [Google Scholar] [CrossRef]
  11. Direkvandi, E.; Mohammadabadi, T.; Dashtizadeh, M.; Alqaisi, O.; Salem, A.Z.M. Lactobacillus plantarum as feed additive to improvement in vitro ruminal biofermentation and digestibility of some tropical tree leaves. J. Appl. Microbiol. 2021, 131, 2739–2747. [Google Scholar] [CrossRef] [PubMed]
  12. Liu, W.; Zhang, L.; Yi, H.; Shi, J.; Xue, C.; Li, H.; Jiao, Y.; Shigwedha, N.; Du, M.; Han, X. Qualitative detection of class IIa bacteriocinogenic lactic acid bacteria from traditional chinese fermented food using a ygngv-motif-based assay. J. Microbiol. Meth. 2014, 100, 121–127. [Google Scholar] [CrossRef] [PubMed]
  13. Ma, G.; Ma, H.; Lu, X.; Liu, J.; Sun, Y.; Bai, F.; Li, J. Screening for broad-spectrum antagonistic lactic acid bacteria from intestine of turbot and identification of bacteriocin produced by it. Food Sci. 2019, 6, 159–165. [Google Scholar] [CrossRef]
  14. Yang, F.; Wang, Y.; Zhao, S.; Wang, Y. Lactobacillus plantarum inoculants delay spoilage of high moisture alfalfa silages by regulating bacterial community composition. Front. Microbiol. 2020, 11, 1989. [Google Scholar] [CrossRef]
  15. Van Soest, P.J.; Robertson, J.B.; Lewis, B.A. Methods for dietary fiber, neutral detergent fiber, and nonstarch polysaccha-rides in relation to animal nutrition. J. Dairy Sci. 1991, 74, 3583–3597. [Google Scholar] [CrossRef]
  16. Menke, K.H.; Steingass, H. Estimation of the energetic feed value obtained from chemical analysis and in vitro gas production using rumen fluid. Anim. Res. Dev. 1988, 28, 7–55. [Google Scholar]
  17. Chen, L.; Dong, Z.; Li, J.; Shao, T. Ensiling characteristics, in vitro rumen fermentation, microbial communities and aerobic stability of low-dry matter silages produced with sweet sorghum and alfalfa mixtures. J. Sci. Food Agr. 2019, 99, 2140–2151. [Google Scholar] [CrossRef]
  18. Broderick, G.A.; Kang, J.H. Automated simultaneous determination of ammonia and total amino acids in ruminal fluid and in vitro media. J. Dairy Sci. 1980, 63, 64–75. [Google Scholar] [CrossRef]
  19. Hu, W.; Schmidt, R.J.; McDonell, E.E.; Klingerman, C.M.; Kung, L.M. The effect of Lactobacillus buchneri 40788 or Lactobacillus plantarum MTD-1 on the fermentation and aerobic stability of corn silages ensiled at two dry matter contents. J. Dairy Sci. 2009, 92, 3907–3914. [Google Scholar] [CrossRef] [Green Version]
  20. Ke, W.; Ding, Z.; Li, F.; Xu, D.; Bai, J.; Muhammad, I.; Zhang, Y.; Zhao, L.; Guo, X. The effects of malic or citric acid on the fermentation quality, proteolysis and lipolysis of alfalfa silage ensiled at two dry matter contents. J. Anim. Physiol. Anim. Nutr. 2021, 106, 988–994. [Google Scholar] [CrossRef]
  21. Zhang, Y.; Huang, Z.; Vyas, D.; Adesogan, A.T.; Francoc, M.; Ke, W.; Li, F.; Bai, J.; Ding, Z.; Guo, X. Antioxidant status, chemical composition and fermentation profile of alfalfa silage ensiled at two dry matter contents with a novel Lactobacillus plantarum strain with high-antioxidant activity. Anim. Feed Sci. Technol. 2021, 272, 114751. [Google Scholar] [CrossRef]
  22. Heron, S.J.E.; Edwards, R.A.; McDonald, P. Changes in the nitrogenous components of gamma-irradiated and inoculated ensiled ryegrass. J. Sci. Food Agric. 1986, 37, 979–985. [Google Scholar] [CrossRef]
  23. Heron, S.J.E.; Owen, T.R. Review of the effects of ‘Ecosyl’ silage inoculant on in vivo digestibilities and metabolizable energy of grass silages. In Management Issues for the Grassland Farmer in the 1990’s; Mayne, C.S., Ed.; Occasional Symposium No 25; British Grassland Society: Birmingham, UK, 1991; pp. 230–233. [Google Scholar]
  24. Palarea-Albaladejo, J.; Rooke, J.A.; Nevison, I.M.; Dewhurst, R.J. Compositional mixed modeling of methane emissions and ruminal volatile fatty acids from individual cattle and multiple experiments. J. Anim. Sci. 2017, 95, 2467–2480. [Google Scholar] [CrossRef] [PubMed]
  25. Oskoueian, E.; Jahromi, M.F.; Jafari, S.; Shakeri, M.; Le, H.H.; Ebrahimi, M. Manipulation of rice straw silage fermentation with different types of lactic acid bacteria inoculant affects rumen microbial fermentation characteristics and methane production. Vet. Sci. 2021, 8, 100. [Google Scholar] [CrossRef]
  26. Wolin, M.J. Interactions between the bacterial species of the rumen. In Digestion and Metabolism in the Ruminant; McDonald, I.M., Warner, A.C.I., Eds.; University of New England Publishing Unit: Sydney, Australia, 1975; pp. 135–148. [Google Scholar]
  27. Belaich, J.P.; Bruschi, M.; Garcia, J.L. Microbiology and Biochemistry of Strict Anaerobes Involved in Interspecies Hydrogen Transfer; Belaich, J.P., Bruschi, M., Garcia, J., Eds.; Plenum Public Corporation: New York, NY, USA, 1990. [Google Scholar]
  28. Stewart, C.S.; Bryant, M.P. The Rumen Bacteria in the Rumen Microbial Ecosystem; Hobson, P.N., Ed.; Elsevier Applied Science: New York, NY, USA, 1988; pp. 21–75. [Google Scholar]
  29. Baker, S.K. Rumen methanogens, and inhibition of methanogenesis. Aust. J. Agric. Res. 1999, 50, 1293–1298. [Google Scholar] [CrossRef]
  30. Wolin, M.J.; Miller, T.L. Microbe-microbe interactions. In The Rumen Microbial Ecosystem; Hobson, P.N., Stewart, C.S., Eds.; Springer Science & Business Media: London, UK, 1997; pp. 467–488. [Google Scholar]
  31. Gamo, Y.; Mii, M.; Zhou, X.G.; Sar, C.; Santoso, B.; Arai, I.; Kimura, K.; Takahashi, J. Effects of lactic acid bacteria, yeasts and galactooligosaccha-ride supplementation on in vitro rumen methane production. In Proceedings of the 1st International Conference on Greenhouse Gases and Animal Agriculture (GGAA), Obihiro, Japan, 7–11 November 2001; Takahashi, J., Young, B.A., Eds.; 2001; pp. 7–11. [Google Scholar]
  32. Feng, Y. Nutrition of Ruminants; Feng, Y., Ed.; Science Press: Beijing, China, 2004. [Google Scholar]
  33. Charmley, E. Towards improved silage quality—A review. The Canadian veterinary journal. Can. J. Anim. Sci. 2001, 81, 157–168. [Google Scholar] [CrossRef]
Animals 13 00384 g001 550
Figure 1. Lactic acid fermentation characteristics and fiber contents of alfalfa silages at moderate and high DM contents after 90 days. (a) PH, (b) LA, (c) aNDF, and (d) ADF. LA, lactic acid; aNDF, neutral detergent fiber analyzed with heat-stable α-amylase; ADF, acid detergent fiber; DM, dry matter. a–d: Different lowercase letters indicate significant differences among treatments in the same DM level (p < 0.05). A–B: Different capital letters indicate significant differences between DM levels in the same treatment (p < 0.05).
Figure 1. Lactic acid fermentation characteristics and fiber contents of alfalfa silages at moderate and high DM contents after 90 days. (a) PH, (b) LA, (c) aNDF, and (d) ADF. LA, lactic acid; aNDF, neutral detergent fiber analyzed with heat-stable α-amylase; ADF, acid detergent fiber; DM, dry matter. a–d: Different lowercase letters indicate significant differences among treatments in the same DM level (p < 0.05). A–B: Different capital letters indicate significant differences between DM levels in the same treatment (p < 0.05).
Animals 13 00384 g001
Table
Table 1. In vitro ruminal fermentation characteristics of silage inoculated with bacterial strains.
Table 1. In vitro ruminal fermentation characteristics of silage inoculated with bacterial strains.
Iterms 1M H Effects 3
ControlMTD/1ATCC-14917LP1-4MeanControlMTD/1ATCC-14917LP1-4MeanSEM 2ADA × D
pH6.676.676.746.756.716.696.686.726.706.700.0140.3590.6820.801
Total gas (mL/g DM)103 Ba86.6 Bb83.5 b99.0 a93.0124 Aa106 Ab90.5 c92.5 c10318.473<0.001<0.001<0.001
CH4 (mL/g DM)11.3 a7.34 b3.53 c6.34 Abc7.149.88 a7.23 b3.60 c2.11 Bc5.710.977<0.0010.0030.006
CH4-to-total gas (% mL)11.0 a8.49 Aab4.27 c7.29 Abc7.778.00 a6.85 Bb3.92 c2.29 Bc5.271.151<0.001<0.0010.011
IDVMD (g/kg DM)65.5 Ab70.3 Aa71.7 Aa69.3 Aa69.252.1 Bb54.5 Bb63.4 Ba61.5 Ba57.92.609<0.001<0.0010.001
NH3 (mg/100 mL)22.924.324.825.124.223.223.127.526.225.02.3440.0060.2300.277
Total VFA (mM)72.0 A68.6 A67.8 A68.0 A69.154.2 Bab50.4 Bb57.0 Ba57.5 Ba54.84.4090.033<0.0010.006
Acetate (mM)40.7 Aa37.5 Aab36.9 Ab36.4 Ab37.930.8 Ba29.2 Bb28.7 Bb29.2 Bb29.51.032<0.001<0.0010.171
Propionate (mM)10.3 b10.7 b12.1 ab13.2 a11.611.0 b11.0 b14.9 a14.6 a12.91.060<0.0010.0070.233
Acetate to propionate3.98 Aa3.50 Aab3.05 Abc2.76 Ac3.322.78 Ba2.54 Bab2.13 Bbc1.80 Bc2.320.055<0.001<0.0010.703
Butyrate (mM)15.1 Aa12.8 Aab12.3 Aab11.3 Ac12.98.52 Ba5.76 Bc7.37 Bb7.38 Bb7.260.9580.001<0.0010.039
Isobutyrate (mM)1.201.18 A1.29 A1.441.280.66 b0.61 Bb1.04 Ba1.14 a0.860.0620.0410.0010.577
Valerate (mM)2.30 a2.01 Bab1.77 Bb1.84 Bb1.982.00 b2.62 Aab3.16 Aa3.23 Aa2.750.0450.027<0.001<0.001
Isovalerate (mM)2.453.30 A3.37 A3.90 A3.261.15 b1.19 Bb1.86 Ba1.90 Ba1.520.2660.012<0.0010.486
1 CH4, methane; IDVMD, in vitro dry matter digestibility; NH3, ammonia; VFA, volatile fatty acids. 2 SEM, standard error of the means. a–c Different lowercase letters indicate significant treatment differences (p < 0.05). A,B Different uppercase letters indicate significant differences in dry matters (p < 0.05). 3 A, additive; D, dry matter; A × D, the interaction between additive and dry matter; M, moderate dry matter content; H, high dry matter content.
Disclaimer/Publisher’s Note: The statements, opinions and data contained in all publications are solely those of the individual author(s) and contributor(s) and not of MDPI and/or the editor(s). MDPI and/or the editor(s) disclaim responsibility for any injury to people or property resulting from any ideas, methods, instructions or products referred to in the content.

Share and Cite

MDPI and ACS Style

Li, Z.; Li, F.; Kharazian, Z.A.; Guo, X. Effect of Inoculating Two Bacteriocin-Producing Lactiplantibacillus plantarum Strains at Ensiling on In Vitro Rumen Fermentation and Methane Emissions of Alfalfa Silage with Two Dry Matter Contents. Animals 2023, 13, 384. https://doi.org/10.3390/ani13030384

AMA Style

Li Z, Li F, Kharazian ZA, Guo X. Effect of Inoculating Two Bacteriocin-Producing Lactiplantibacillus plantarum Strains at Ensiling on In Vitro Rumen Fermentation and Methane Emissions of Alfalfa Silage with Two Dry Matter Contents. Animals. 2023; 13(3):384. https://doi.org/10.3390/ani13030384

Chicago/Turabian Style

Li, Ziqian, Fuhou Li, Zohreh Akhavan Kharazian, and Xusheng Guo. 2023. "Effect of Inoculating Two Bacteriocin-Producing Lactiplantibacillus plantarum Strains at Ensiling on In Vitro Rumen Fermentation and Methane Emissions of Alfalfa Silage with Two Dry Matter Contents" Animals 13, no. 3: 384. https://doi.org/10.3390/ani13030384

Note that from the first issue of 2016, this journal uses article numbers instead of page numbers. See further details here.

Article Metrics

Back to TopTop