The Use of Quantitative Real Time Polymerase Chain Reaction to Quantify Some Rumen Bacterial Strains in an<i>In Vitro</i>Rumen System

Onime, Lucy A.; Zanfi, C.; Agostinis, Chiara; Bulla, Roberta; Spanghero, M.

doi:10.4081/ijas.2013.e58

Cited by 4 publications

(1 citation statement)

References 22 publications

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“…Rumen microbes can also be broadly classified based on functional activities, such as cellulolytic, amylolytic, proteolytic, lipolytic, and methanogenic ( Hungate, 1966 ). Despite the wealth of knowledge available on rumen microbial classification, little temporal dynamics data of the microbiome and by-product generation in in vitro fermentation exist, thereby impeding a more realistic depiction of the in vivo model ( Onime et al, 2013 ; Kang et al, 2017 ; Wei et al, 2022 ).…”

Section: Introductionmentioning

confidence: 99%

Temporal dynamics of volatile fatty acids profile, methane production, and prokaryotic community in an in vitro rumen fermentation system fed with maize silage

Dhakal,

Neves,

Sapkota

et al. 2024

Front. Microbiol.

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Anaerobic in vitro fermentation is widely used to simulate rumen kinetics and study the microbiome and metabolite profiling in a controlled lab environment. However, a better understanding of the interplay between the temporal dynamics of fermentation kinetics, metabolic profiles, and microbial composition in in vitro rumen fermentation batch systems is required. To fill that knowledge gap, we conducted three in vitro rumen fermentations with maize silage as the substrate, monitoring total gas production (TGP), dry matter degradability (dDM), and methane (CH4) concentration at 6, 12, 24, 36, and 48 h in each fermentation. At each time point, we collected rumen fluid samples for microbiome analysis and volatile fatty acid (VFA) analysis. Amplicon sequencing of 16S rRNA genes (V4 region) was used to profile the prokaryotic community structure in the rumen during the fermentation process. As the fermentation time increased, dDM, TGP, VFA concentrations, CH4 concentration, and yield (mL CH4 per g DM at standard temperature and pressure (STP)) significantly increased. For the dependent variables, CH4 concentration and yield, as well as the independent variables TGP and dDM, polynomial equations were fitted. These equations explained over 85% of the data variability (R2 > 0.85) and suggest that TGP and dDM can be used as predictors to estimate CH4 production in rumen fermentation systems. Microbiome analysis revealed a dominance of Bacteroidota, Cyanobacteria, Desulfobacterota, Euryarchaeota, Fibrobacterota, Firmicutes, Patescibacteria, Proteobacteria, Spirochaetota, and Verrucomicrobiota. Significant temporal variations in Bacteroidota, Campylobacterota, Firmicutes, Proteobacteria, and Spirochaetota were detected. Estimates of alpha diversity based on species richness and the Shannon index showed no variation between fermentation time points. This study demonstrated that the in vitro fermentation characteristics of a given feed type (e.g., maize silage) can be predicted from a few parameters (CH4 concentration and yield, tVFA, acetic acid, and propionic acid) without running the actual in vitro trial if the rumen fluid is collected from similar donor cows. Although the dynamics of the rumen prokaryotes changed remarkably over time and in accordance with the fermentation kinetics, more time points between 0 and 24 h are required to provide more details about the microbial temporal dynamics at the onset of the fermentation.

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Section: Introductionmentioning

confidence: 99%

Temporal dynamics of volatile fatty acids profile, methane production, and prokaryotic community in an in vitro rumen fermentation system fed with maize silage

Dhakal,

Neves,

Sapkota

et al. 2024

Front. Microbiol.

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Design and application of a synthetic DNA standard for real-time PCR analysis of microbial communities in a biogas digester

May

Koch-Singenstreu

Ebling

et al. 2015

Appl Microbiol Biotechnol

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A synthetic DNA fragment containing primer binding sites for the quantification of ten different microbial groups was constructed and evaluated as a reliable enumeration standard for quantitative real-time PCR (qPCR) analyses. This approach has been exemplary verified for the quantification of several methanogenic orders and families in a series of samples drawn from a mesophilic biogas plant. Furthermore, the total amounts of bacteria as well as the number of sulfate-reducing and propionic acid bacteria as potential methanogenic interaction partners were successfully determined. The obtained results indicated a highly dynamic microbial community structure which was distinctly affected by the organic loading rate, the substrate selection, and the amount of free volatile fatty acids in the fermenter. Methanosarcinales was the most predominant methanogenic order during the 3 months of observation despite fluctuating process conditions. During all trials, the modified quantification standard indicated a maximum of reproducibility and efficiency, enabling this method to open up a wide range of novel application options.

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Administration ofAloe arborescenshomogenate to cattle: interaction with rumen fermentation and gut absorption of aloin

Bani

Grossi

Lucini

et al. 2016

Italian Journal of Animal Science

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Aloe has long been used as a traditional medicine for its numerous beneficial properties, which are mainly ascribed to b-polysaccharides and phenolic compounds including anthraquinones, anthrones and chromones. However, few studies on large animals are currently available. The effect of whole leaf Aloe arborescens homogenate on the in vitro rumen fermentative processes was tested using alfalfa hay and barley meal as substrates. The Aloe homogeneate was added at three different concentrations (0.4, 2.0 and 10.0 g L À1 of fermentation liquid). The same homogenate was dosed (200 g) orally and through the rumen cannula to three rumen cannulated heifers and orally to six lactating dairy cows to measure the rumen degradation of aloin and the transfer of aloin from the gut into the blood, respectively. The Aloe homogenate did not affect in vitro rumen fermentations and feed digestibility. The administration of Aloe homogenate did not negatively affect animal feed intake and health neither on the cannulated heifers nor on the dairy cows. Aloin underwent a rapid degradation in the rumen milieu, and became undetectable 2 h after oral dosage. However, when Aloe homogenate was administered to dairy cows, aloin appeared in blood as early as 2 h after administration, reached a maximum after 4 h (6.2 6 5.8 lg L À1 ) and progressively decreased thereafter. These results suggest that Aloe compounds can be absorbed into the blood and encourage the study of Aloe as a potential nutraceutical in ruminants. Further studies should determine the most effective in vivo dosage. ARTICLE HISTORY

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The Use of Quantitative Real Time Polymerase Chain Reaction to Quantify Some Rumen Bacterial Strains in anIn VitroRumen System

Cited by 4 publications

References 22 publications

Temporal dynamics of volatile fatty acids profile, methane production, and prokaryotic community in an in vitro rumen fermentation system fed with maize silage

Temporal dynamics of volatile fatty acids profile, methane production, and prokaryotic community in an in vitro rumen fermentation system fed with maize silage

Design and application of a synthetic DNA standard for real-time PCR analysis of microbial communities in a biogas digester

Administration ofAloe arborescenshomogenate to cattle: interaction with rumen fermentation and gut absorption of aloin

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