باکتری های تجزیه کننده تانن ها و ترکیبات فنلی در شکمبه

Author links open overlay panelSrobana Sarkar a 1, Pankaj Kumar Kumawat a 1, Randhir Singh Bhatt a, Artabandhu Sahoo a b

Show moreAdd to Mendeley

Share

Cite

https://doi.org/10.1016/j.smallrumres.2024.107272Get rights and content

Highlights

• •

  Tanninolytic bacteria were isolated from rumen contents of Malpura sheep.

• •

  Out of 24 isolates, 16 belonged to order Lactobacillales with tannase activity ranging from 1.2-12.5 mol gallic acid/min.

• •

  Total cellulolytic enzyme activity (FPase) of the isolates ranged from 0.14 to 9.80 U/mL.

• •

  Isolated tanninolytic strains effectively degraded Prosopsis cineraria leaves showing potential to be used as direct-fed microbials in ruminants for utilization of tannin-rich fibrous feeds.

Abstract

To counteract the negative effects of phytotannins, ruminants host a diverse community of bacteria that break down tannins. These bacteria not only possess tanninolytic activity but also have other functional relevance that hasn't been properly examined. Therefore, the present study was conducted to isolate and characterize tannin degrading bacteria (TDB) from rumen of sheep fed tannin-rich diet with possible fibrolytic activity. Based on the development of a clear zone on tannin-enriched agar plates, a total of 24 isolates were screened as tannin hydrolyzing bacteria. Majority of the isolates were gram positive cocci. The isolates could tolerate phenolic monomers such as ferulic acid, gallic acid, vanillic acid up to 30 mM and pyrogallol up to 10 mM, while failed to grow in presence of syringic acid, p-coumaric acid and p-hydroxybenzoic acid. Highest and lowest tannase activity was observed in TDB23 and TDB2 and TDB5 isolates, respectively. In terms of fibrolytic activity, maximum endoglucanase, exoglucanase, and FPase activity was observed for TDB9. In vitro digestibility of Prosopis cineraria leaves was considerably enhanced by inoculation of isolate TDB23 followed by TDB9. Therefore, the screened isolates demonstrated promising tannin and fibre degrading potential as measured by their tannase and fibrolytic enzyme activities, which can be further explored as direct-fed microbial in ruminants for effective utilization of tannin-rich fibrous feeds.

Introduction

Poor quality and insufficient feed supply are the main causes of low production in domesticated ruminants in arid and semiarid tropical regions (Kumawat et al., 2021). For grazing ruminants in semi-arid environments, native browsing foliages and shrubs make up a sizable share of supplemental feed resources. In addition to excessive lignification, the quantity of tannins present in these foliages and forages often limits nutrient supply to the animals by making them biologically unavailable or interfering with the proliferation and activity of rumen microbes (Patra et. al., 2012; Choudhary et al., 2018). Tannins are polyphenolic plant anti-nutritional substances that exert deleterious effects on livestock by interfering with the absorption of food protein, carbohydrates, minerals, and digestive enzymes causing reduced nutrient utilization (Bhat et al., 2013). Several studies have reported that dietary inclusion of tannin rich feeds hampers animal productivity (Singh et al., 2011, Tseu et al., 2020). Reducing these substances at the animal level is more important because the elimination of plant secondary metabolites can negatively impact plants' defense mechanisms. Therefore, various strategies have been adapted in the recent years to reduce the tannin content of feeds for better nutrient utilization in ruminants (Makkar et al., 2007).

A variety of unidentified microorganisms are present in the rumen microbiome, which can aid in the bioconversion and detoxification of various toxins present in feeds (Singh et al., 2011). Several microorganisms have been reported from the gastrointestinal tract of ruminants with the ability to degrade tannins like Streptococcus bovis (Osawa and Sly, 1991), Lonepinella koalarum (Osawa et al., 1995), Clostridium sp (McSweeney et al., 1999) and Selenomonas ruminantium (Odenyo et al., 2001). Goel et al. (2007) reported significant tannase activity in various strains of Group D streptococci (Enterococcus faecalis) isolated from goat faecal samples. Kumar et al. (2014) also reported tannin tolerant bacterial isolates from rumen of goats fed oak leaves that were 99 to 100% related to Streptococcus gallolyticus. In addition to bacteria, tanninolytic activity was also shown by anaerobic fungal isolates (Piromyces sp.) when innoculated to a media containing rumen buffalo rumen fluid and tannin rich tree leaves (Paul et al., 2006). Nevertheless, tanninolytic bacteria are more effective and fast degraders of natural tannins than fungi (Goel et al., 2011). Strategies to enhance tannin-tolerant bacteria in the rumen can improve the utilization of tannin-rich foliage and forages in ruminants. Till date, studies primarily isolated tannin-degrading bacteria from feces of ruminants, with limited characterization from the rumen and mostly focused on tanninolysis capacity. Therefore, the present study was undertaken to isolate and characterize tanninolytic bacteria from rumen contents of sheep fed tannin rich diets as well as to assess the fibrolytic potential of the isolates in order to screen direct fed microbial additives that can synergistically improve the nutrient utilization of poor quality tannin rich tropical feed resources in ruminants.

Section snippets

Site of study and specimen collection

The present experiment was carried out in semi-arid regions of ICAR-Central Sheep and Wool Research Institute, Rajasthan, India. For the adaptive proliferation of tannin degrading/adapting microbes in the rumen, 25 Malpura sheep were maintained on a tanniferous diet consisting of Prosopsis cineraria (Total tannins- 98.5, condensed tannins- 62.2 and hydrolysable tannins- 36.3 g/kg dry matter) and Zizipus numularia (Total tannins- 60.4, condensed tannins- 36.9 and hydrolysable tannins- 26.3 g/kg

Morphological and biochemical attributes

A total of 80 rumen content samples were included in the study. Out of 67 isolates of tannin degrading bacteria, 24 isolates showing clear zones on agar plates containing tannic acid as substrate were presumptively selected for further analysis and named as TDB1 - TDB24. The morphological and biochemical characteristics of the isolates are presented in Table 1. Most of the isolates were Gram positive except TDB1, TDB3, TDB13, TDB17, TDB21, and TDB24 which were Gram negative. Isolates were

Morphological and biochemical attributes

Most of the isolates were cocci and diplococci morphologically, while others were rod-shaped bacilli. Similarly, Osawa and Mitsuoka (1990) and Goel et al. (2007) isolated diplococcic or chain of cocci from feces of koala and goat, respectively capable of degrading tannin protein complex. Kumar et al. (2014) also reported tannin degrading rod shaped coccobacilli from rumen of goats fed Quercus semicarpifolia leaves. Goel et al. (2015) isolated streptococci from cattle and reported that these

Conclusions

From the present findings it can be concluded that the tannin degrading bacterial isolates from sheep rumen contents showed promising fibrolytic activity. Isolate TDB23 had high content of tannase and FPase which synergistically improved the digestibility of tannin rich Prosopis cineraria leaves. Hence, this isolates can be explored as direct fed microbes or trans-inoculants into other animal species for enhancing nutrient utilization of tannin rich low quality feeds.

Uncited references

(Aguilar et al., 2007, Kumar et al., 2013)

CRediT authorship contribution statement

Pankaj Kumar Kumawat: Methodology, Investigation. Srobana Sarkar: Writing – original draft, Project administration, Funding acquisition, Formal analysis, Data curation, Conceptualization. Artabandhu Sahoo: Writing – review & editing, Validation, Supervision, Project administration. Randhir Singh Bhatt: Visualization, Validation, Formal analysis, Data curation.

Declaration of Competing Interest

On behalf of all the authors, I declare that no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Acknowledgement

Authors are thankful to Director, ICAR-Central Sheep and Wool Research Institute, Avikanagar for providing necessary research facilities required during this study. The financial assistance provided by National Centre for Veterinary Type Cultures Network Project is gratefully acknowledged.

Declaration of Competing Interest

It is to certify that there is no actual or potential conflict of interest including any financial, personal or other relationships with other people or organizations within three years of beginning the

References (43)

• G. Goel et al.

  Degradation of tannic acid and purification and characterization of tannase from Enterococcus faecalis

  Int. Biodeterior. Biodegradation

  (2011)

• G. Goel et al.

  Phenotypic characterization of tannin–protein complex degrading bacteria from faeces of goat

  Small Rumin. Res.

  (2007)

• G.G. Gross

  Biosynthesis of hydrolyzable tannins

  Comprehensive Natural Products Chemistry

  (1999)

• V. Mlambo et al.

  The effectiveness of adapted rumen fluid versus PEG to ferment tannin-containing substrates in vitro

  Anim Feed Sci. Technol.

  (2007)

• Y. Nishitani et al.

  Genotypic analyses of lactobacilli with a range of tannase activities isolated from human feces and fermented foods

  Syst. Appl. Microbiol.

  (2004)

• A.A. Odenyo et al.

  Characterization of tannin-tolerant bacterial isolates from East African ruminants

  Anaerobe

  (2001)

• R. Osawa et al.

  Lonepinella koalarum gen. nov., sp. nov., a new tannin-protein complex degrading bacterium

  Syst. Appl. Microbiol.

  (1995)

• L.J. Porter et al.

  The conversion of procyanidins and prodelphinidins to cyanidin and delphinidin

  Phytochem

  (1985)

• E. Sasaki et al.

  Isolation of tannin-degrading bacteria isolated from feces of the Japanese large wood mouse, Apode musspeciosus, feeding on tannin-rich acorns

  Syst. Appl. Microbiol.

  (2005)

• S. Sharma et al.

  A spectrophotometric method for assay of tannase using rhodanine

  Anal. Biochem.

  (2000)