Dose-response effect of crude extracts produced by actinobacteria on in vitro rumen fermentation

Abstract

Actinobacteria have been researched as a source that produces crude extracts, which contain bioactive compounds able to act as antimicrobial agents. The present investigation evaluated the dose-response effect of two crude extracts, obtained at Caatinga rhizosphere (Caat) and Rhizophora mangle (AMC), on in vitro ruminal fermentation by: cumulative gas production, digestibility of dry (IVDMD) and organic matter (IVOMD), and short-chain fatty acids concentration (SCFA). Three multiparous Holstein dairy cows with ruminal fistula were used as the inoculum donors and fed a basal diet consisting of corn silage, soybean meal, urea, ground corn and mineral supplement. Ruminal fluid samples were incubated in glass bottles with 1 g of the dried and milled diet, a buffer solution, and the crude extracts evaluated in four doses (0.3, 0.6, 0.9 and 1.20 mg/10 mL inoculum) in a randomized block design, and the donators were considered as blocks with random effects. Additionally, negative controls were used. The results were expressed as average values based on triplicate analyses. Decreased cumulative gas production was observed according to linear dose response at 24, 48 and 72 h of incubation with the addition of Caat extract. The IVOMD showed a linear decrease at 72 h of incubation with dose Caat inclusion. Furthermore, the inclusion of Caat extract linearly reduced butyric and isovaleric acid concentrations, as well as acetate:propionate ratio. Finally, the Caat inclusion increased the propionic acid concentration in comparison to AMC extract. However, the inclusion of AMC extract did not affect any of the analyzed variables at the used doses. The Caat extract could be used as a modulator of in vitro ruminal fermentation, since it reduced acetate:propionate ratio and cumulative gas production. ResumoAs actinobactérias têm sido pesquisadas como fonte produtoras de extratos brutos que contêm compostos bioativos capazes de atuar como agentes antimicrobianos. O presente trabalho investigou o efeito dose-resposta de dois extratos brutos, AMC e Caat, na fermentação ruminal in vitro por: produção cumulativa de gás, digestibilidade in vitro da matéria seca (IVDMD) e matéria orgânica (IVOMD) e concentração de ácidos graxos de cadeia curta (SCFA). Três vacas leiteiras da raça Holandesa, multíparas e portadoras de fístula ruminal foram utilizadas como doadoras de inóculo ruminal e foram alimentadas com uma dieta basal composta por silagem de milho, farelo de soja, ureia, milho moído e suplemento mineral. As amostras de inóculo ruminal foram incubadas em garrafas de vidro com 1 g da dieta seca e moída, solução tampão e os extratos brutos avaliados em quatro doses (0,3, 0,6, 0,9 e 1,20 mg/10 mL de inóculo) em delineamento em blocos casualizados, sendo as doadoras consideradas os blocos como efeito aleatório. Além disso, foram utilizados controles negativos para a correção da produção de gás. Os resultados foram expressos como valores médios com base em análises triplicadas. A diminuição da produção cumulativa de gás foi observada de aco...

In vitro evaluation of novel crude extracts produced by actinobacteria for modulation of ruminal fermentation

“…While several food In vitro evaluation of novel crude extracts produced by actinobacteria for modulation of ruminal fermentation Alves et al 7 additives-including ionophores, oil extracts, and essential oils-have been commercially available for the past decade, to our best knowledge, this is the first study on the use of crude extracts derived from actinobacteria in the modulation of in vitro ruminal fermentation. This study was the first to report the CH 4 and NH 3 -N concentration, after the in vitro dose-response effects reported previously by our group (Alves et al, 2018), although previous studies have used essential oils or chitosan as alternatives to ionophores (Benchaar et al, 2006;Khorrami et al, 2015;Tomkins et al, 2015;Vendramini et al, 2016).…”

“…However, the reduction of total gases after the inclusion of extracts may vary according the in vitro assay characteristics, such as substrate quality, anaerobic conditions, buffer quality, as well as the included dose (Alves et al, 2018;Bueno et al, 2005;Rymer et al, 1999). In the present study, we evaluated two crude extracts diluted in DMSO, and probably this dilution may have affected the magnitude of the crude extract mode of action in comparison with a pure extract.…”

“…Our research group (Crevelin et al, 2013) recently characterized the AMC crude extract, the composition of which indicated the presence of macrotetrolids, with bafilomycin as the main compound. The identification of main secondary metabolites present in actinobacteria Caat 2-63 (actinomycin D) was based on the dereplication data employed in high-resolution mass spectrometry (HRMS) and collision-induced dissociation (CID) experiments, as described by Alves et al (2018).…”

“…In vitro evaluation of novel crude extracts produced by actinobacteria for modulation of ruminal fermentation Alves et al 3 composed of corn silage as forage, soybean meal, urea, ground corn, and mineral supplement, in a forage:concentrate ratio of 60:40, formulated based on the nutritional requirements of lactating cows producing 30 kg of milk/day, 3.5% fat, and 3.1% protein, according to NRC (2001). The composition of the substrate (diet) used in the in vitro fermentation assay was determined by the chemical analysis previously reported by Alves et al (2018).…”

“…A previous companion paper (Alves et al, 2018) evaluated the dose-response of AMC and Caat crude extracts, and an optimum dose response for the crude extracts was observed at 1.2 mg/25 mL ruminal inoculum 160 mL (glass bottle) with a reduction of gas production during 72 h of in vitro fermentation.…”