AUTHOR=Bharanidharan Rajaraman , Thirugnanasambantham Krishnaraj , Ibidhi Ridha , Baik Myunggi , Kim Tae Hoon , Lee Yookyung , Kim Kyoung Hoon 

TITLE=Metabolite Profile, Ruminal Methane Reduction, and Microbiome Modulating Potential of Seeds of Pharbitis nil

JOURNAL=Frontiers in Microbiology

VOLUME=13

YEAR=2022

URL=https://www.frontiersin.org/journals/microbiology/articles/10.3389/fmicb.2022.892605

DOI=10.3389/fmicb.2022.892605

ISSN=1664-302X

ABSTRACT=<p>We identified metabolites in the seeds of <italic>Pharbitis nil</italic> (PA) and evaluated their effects on rumen methanogenesis, fiber digestibility, and the rumen microbiome <italic>in vitro</italic> and <italic>in sacco</italic>. Four rumen-cannulated Holstein steers (mean body weight 507 ± 32 kg) were used as inoculum donor for <italic>in vitro</italic> trial and live continuous culture system for <italic>in sacco</italic> trial. PA was tested <italic>in vitro</italic> at doses ranging from 4.5 to 45.2% dry matter (DM) substrate. The <italic>in sacco</italic> trial was divided into three phases: a control phase of 10 days without nylon bags containing PA in the rumen, a treatment phase of 11 days in which nylon bags containing PA (180 g) were placed in the rumen, and a recovery phase of 10 days after removing the PA-containing bags from the rumen. Rumen headspace gas and rumen fluid samples were collected directly from the rumen. PA is enriched in polyunsaturated fatty acids dominated by linoleic acid (C18:2) and flavonoids such as chlorogenate, quercetin, quercetin-3-<italic>O</italic>-glucoside, and quinic acid derivatives. PA decreased (<italic>p</italic> &lt; 0.001) methane (CH<sub>4</sub>) production linearly <italic>in vitro</italic> with a reduction of 24% at doses as low as 4.5% DM substrate. A quadratic increase (<italic>p</italic> = 0.078) in neutral detergent fiber digestibility was also noted, demonstrating that doses &lt; 9% DM were optimal for simultaneously enhancing digestibility and CH<sub>4</sub> reduction. <italic>In sacco</italic>, a 50% decrease (<italic>p</italic> = 0.087) in CH<sub>4</sub> coupled with an increase in propionate suggested increased biohydrogenation in the treatment phase. A decrease (<italic>p</italic> &lt; 0.005) in ruminal ammonia nitrogen (NH<sub>3</sub>-N) was also noted with PA in the rumen. Analysis of the rumen microbiome revealed a decrease (<italic>p</italic> &lt; 0.001) in the Bacteroidetes-to-Firmicutes ratio, suggesting PA to have antiprotozoal potential. At the genus level, a 78% decrease in <italic>Prevotella</italic> spp. and a moderate increase in fibrolytic <italic>Ruminococcus</italic> spp. were noted in the treatment phase. <italic>In silico</italic> binding of PA metabolites to <italic>cyclic GMP-dependent protein kinase</italic> of <italic>Entodinium caudatum</italic> supported the antiprotozoal effect of PA. Overall, based on its high nutrient value and antiprotozoal activity, PA could probably replace the ionophores used for CH<sub>4</sub> abatement in the livestock industry.</p>