Supplemental Materials: Effects of methane inhibitors on ruminal fermentation and microbial composition in vitro using inoculum from phenotypically high- and low-enteric methane emitting cows

The objective of this study was to investigate the interraction of ruminal inoculum type (obtained from high- or low-CH4-emitting phenotype cows) and potent anti-methanogenic compounds (AMC), including bromoform (CHBr3), and 3-nitropropionic acid (3NPA), chloroform (CHCl3) and a red macroalga (Rhodophyta macroalga, RM) on in vitro gas production, ruminal fermentation, and microbial composition parameters. Sixty-eight mid-lactation Holstein cows were initially screened for their enteric CH4 emissions. Out of these 68 cows, 2 were designated as high- (HM) and 2 as low- (LM) CH4 emitters. These cows were used as inoculum donors for the in vitro experiment. In vitro batch-culture incubations were carried out for 24 h. In the in vitro experiment, the low-CH4 inoculum (LMI) resulted in similar CH4 concentration in total gas as the high-CH4 inoculum (HMI), regardless of inclusion of AMC. Total gas production was 21% lower for HMI when compared with LMI, regardless of the treatment. The inclusion of all AMC resulted in a decrease in acetate and an increase in propionate and butyrate molar proportions. Of the studied inhibitors, 3NPA tended to result in a greater CH4 reduction, reduced acetate, and increased propionate concentrations when combined with LMI, than with HMI. A macroalgae inhibitor in combination with HMI, but not LMI, resulted in lower CH4 and higher propionate concentration. Both LMI inoculum and inclusion of each AMC resulted in increased butyrate concentrations. Inclusion of all AMC resulted in increased propionate concentrations in both inoculum types. Each AMC differentially affected methanogens and individual bacteria, altering H2 fluxes. Regardless of the inoculum, CHBr3 and 3NPA altered both methanogenic and bacterial communities to a greater extent than the rest of the AMC. The combination of LMI with 3NPA and HMI with RM resulted in a greater CH4 reduction than HMI with 3NPA and LMI with RM, indicating that a different set of AMC may be more effective in reducing enteric CH4 in HMI versus LMI.

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Work Title Supplemental Materials: Effects of methane inhibitors on ruminal fermentation and microbial composition in vitro using inoculum from phenotypically high- and low-enteric methane emitting cows
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Open Access
Creators
  1. Nadiia Stepanchenko
  2. Derek Wasson
  3. Nagaraju Indugu
  4. Sergio Cueva Welchez
  5. Leoni Martins
  6. Kapil Narayan
  7. Dipti Pitta
  8. Alexander Hristov
License In Copyright (Rights Reserved)
Work Type Other
Publication Date 2025
DOI doi:10.26207/73wy-fn85
Deposited January 07, 2025

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Version 1
published

  • Created
  • Updated
  • Updated Description, Publication Date Show Changes
    Description
    • The objective of this study was to investigate the interraction of rumen inoculum type (obtained from high- or low-CH4-emitting phenotype cows) and potent anti-methanogenic compounds (AMC), including bromoform (CHBr3), and 3-nitropropionic acid (3NPA), chloroform (CHCl3) and a red macroalga (SeaweedX, SX) on in vitro gas production, ruminal fermentation, and microbial composition parameters. Sixty-eight mid-lactation Holstein cows were initially screened for their enteric CH4 emissions. Out of these 68 cows, 2 were designated as high- (HM) and 2 as low- (LM) CH4 emitters. These cows were used as inoculum donors for the in vitro experiment. In vitro batch-culture incubations were carried out for 24 h. In the in vitro experiment, the low-CH4 inoculum (LMI) resulted in similar CH4 concentration in total gas as the high-CH4 inoculum (HMI), regardless of inclusion of AMC. Total gas production was 21% lower for HMI when compared with LMI, regardless of the treatment. The inclusion of all AMC resulted in a decrease in acetate and an increase in propionate and butyrate molar proportions. Of the studied inhibitors, 3NPA tended to result in a greater CH4 reduction, reduced acetate, and increased propionate concentrations when combined with LMI, than with HMI. A macroalgae inhibitor in combination with HMI, but not LMI, resulted in lower CH4 and higher propionate concentration. Both LMI inoculum and inclusion of each AMC resulted in increased butyrate concentrations. Inclusion of all AMC resulted in increased propionate concentrations in both inoculum types. Each AMC differentially affected methanogens and individual bacteria, altering H2 fluxes. Regardless of the inoculum, CHBr3 and 3NPA altered both methanogenic and bacterial communities to a greater extent than the rest of the AMC. The combination of LMI with 3NPA and HMI with SX resulted in a greater CH4 reduction than HMI with 3NPA and LMI with SX, indicating that a different set of AMC may be more effective in reducing enteric CH4 in HMI versus LMI.
    Publication Date
    • 2025
  • Added Creator Nadiia Stepanchenko
  • Added Creator Derek Wasson
  • Added Creator Nagaraju Indugu
  • Added Creator Sergio Cueva Welchez
  • Added Creator Leoni Martins
  • Added Creator Kapil Narayan
  • Added Creator Dipti Pitta
  • Added Creator Alexander Hristov
  • Added Supplementary Figures.docx
  • Added Supplementary Tables 4 and 5.xlsx
  • Updated License Show Changes
    License
    • https://rightsstatements.org/page/InC/1.0/
  • Published
  • Updated
  • Updated Creator Leoni Martins

Version 2
published

  • Created
  • Updated Description Show Changes
    Description
    • The objective of this study was to investigate the interraction of rumen inoculum type (obtained from high- or low-CH4-emitting phenotype cows) and potent anti-methanogenic compounds (AMC), including bromoform (CHBr3), and 3-nitropropionic acid (3NPA), chloroform (CHCl3) and a red macroalga (SeaweedX, SX) on in vitro gas production, ruminal fermentation, and microbial composition parameters. Sixty-eight mid-lactation Holstein cows were initially screened for their enteric CH4 emissions. Out of these 68 cows, 2 were designated as high- (HM) and 2 as low- (LM) CH4 emitters. These cows were used as inoculum donors for the in vitro experiment. In vitro batch-culture incubations were carried out for 24 h. In the in vitro experiment, the low-CH4 inoculum (LMI) resulted in similar CH4 concentration in total gas as the high-CH4 inoculum (HMI), regardless of inclusion of AMC. Total gas production was 21% lower for HMI when compared with LMI, regardless of the treatment. The inclusion of all AMC resulted in a decrease in acetate and an increase in propionate and butyrate molar proportions. Of the studied inhibitors, 3NPA tended to result in a greater CH4 reduction, reduced acetate, and increased propionate concentrations when combined with LMI, than with HMI. A macroalgae inhibitor in combination with HMI, but not LMI, resulted in lower CH4 and higher propionate concentration. Both LMI inoculum and inclusion of each AMC resulted in increased butyrate concentrations. Inclusion of all AMC resulted in increased propionate concentrations in both inoculum types. Each AMC differentially affected methanogens and individual bacteria, altering H2 fluxes. Regardless of the inoculum, CHBr3 and 3NPA altered both methanogenic and bacterial communities to a greater extent than the rest of the AMC. The combination of LMI with 3NPA and HMI with SX resulted in a greater CH4 reduction than HMI with 3NPA and LMI with SX, indicating that a different set of AMC may be more effective in reducing enteric CH4 in HMI versus LMI.
    • The objective of this study was to investigate the interraction of ruminal inoculum type (obtained from high- or low-CH4-emitting phenotype cows) and potent anti-methanogenic compounds (AMC), including bromoform (CHBr3), and 3-nitropropionic acid (3NPA), chloroform (CHCl3) and a red macroalga (Rhodophyta macroalga, RM) on in vitro gas production, ruminal fermentation, and microbial composition parameters. Sixty-eight mid-lactation Holstein cows were initially screened for their enteric CH4 emissions. Out of these 68 cows, 2 were designated as high- (HM) and 2 as low- (LM) CH4 emitters. These cows were used as inoculum donors for the in vitro experiment. In vitro batch-culture incubations were carried out for 24 h. In the in vitro experiment, the low-CH4 inoculum (LMI) resulted in similar CH4 concentration in total gas as the high-CH4 inoculum (HMI), regardless of inclusion of AMC. Total gas production was 21% lower for HMI when compared with LMI, regardless of the treatment. The inclusion of all AMC resulted in a decrease in acetate and an increase in propionate and butyrate molar proportions. Of the studied inhibitors, 3NPA tended to result in a greater CH4 reduction, reduced acetate, and increased propionate concentrations when combined with LMI, than with HMI. A macroalgae inhibitor in combination with HMI, but not LMI, resulted in lower CH4 and higher propionate concentration. Both LMI inoculum and inclusion of each AMC resulted in increased butyrate concentrations. Inclusion of all AMC resulted in increased propionate concentrations in both inoculum types. Each AMC differentially affected methanogens and individual bacteria, altering H2 fluxes. Regardless of the inoculum, CHBr3 and 3NPA altered both methanogenic and bacterial communities to a greater extent than the rest of the AMC. The combination of LMI with 3NPA and HMI with RM resulted in a greater CH4 reduction than HMI with 3NPA and LMI with RM, indicating that a different set of AMC may be more effective in reducing enteric CH4 in HMI versus LMI.
  • Published
  • Updated
  • Updated Creator Leoni Martins

Version 3
published

  • Created
  • Deleted Supplementary Figures.docx
  • Deleted Supplementary Tables 4 and 5.xlsx
  • Added Copy of H1021 Supplementary Tables 4 and 5.xlsx
  • Added Supplementary Figures.docx
  • Published
  • Updated