RESEARCH ARTICLE
Selection and characterization of probiotic lactic acid bacteria and its impact on growth, nutrient digestibility, health and antioxidant status in weaned piglets Runjun Dowarah1☯¤, Ashok Kumar Verma1☯*, Neeta Agarwal1☯, Putan Singh1☯, Bhoj Raj Singh2☯
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1 Centre of Advanced Faculty Training in Animal Nutrition, ICAR-Indian Veterinary Research Institute, Izatnagar, UP, India, 2 Division of Epidemiology, ICAR-Indian Veterinary Research Institute, Izatnagar, UP, India ☯ These authors contributed equally to this work. ¤ Current address: Department of Animal Nutrition, College of Veterinary Science, Assam Agricultural University, Guwahati, Assam, India *
[email protected]
Abstract OPEN ACCESS Citation: Dowarah R, Verma AK, Agarwal N, Singh P, Singh BR (2018) Selection and characterization of probiotic lactic acid bacteria and its impact on growth, nutrient digestibility, health and antioxidant status in weaned piglets. PLoS ONE 13(3): e0192978. https://doi.org/10.1371/journal. pone.0192978 Editor: Pratyoosh Shukla, Maharshi Dayanand University, INDIA Received: September 24, 2017 Accepted: February 1, 2018 Published: March 8, 2018 Copyright: © 2018 Dowarah et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Data Availability Statement: All relevant data are within the paper and its Supporting Information files. Funding: Infrastructure facility and support for this research work was provided by Director, ICARIVRI, Izatnagar and the INSPIRE Fellowship to the first author provided by Department of Science and Technology, Govt. of India. The funder had no role in study design, data collection and analysis,
The present study was aimed to develop an effective probiotic lactic acid bacteria (LAB) from piglet feces and in vitro characterization of probiotic properties. To confirm host-species specificity of probiotics, the efficacy of isolated LAB on growth, nutrient utilization, health and antioxidant status was observed in early weaned piglets. A total of 30 LAB were isolated from feces of five healthy piglets (28d old). All isolates were Gram positive, cocco-bacilli and catalase negative. Out of thirty LAB isolates, twenty were shortlisted on the basis of their tolerance to pH (3.0, 4.0, 7.0 and 8.0) and bile salts (0.075, 0.15, 0.3 and 1.0%). Whereas, fourteen isolates were selected for further in vitro probiotic characterization due higher (P45 percent). These isolates fermented twenty-seven different carbohydrates but were negative for ONPG, citrate and malonate. Also enabled to synthesize amylase, protease, lipase and phytase. They were sensitive to penicillin, azithromycin, lincomycin, clindamycin, erythromycin, cephalothin and chloramphenicol and resistant to ciprofloxacin, ofloxacin, gatifloxacin, vancomycin and co-trimoxazole. Except three isolates, all showed antagonistic activity (>60% co-culture activity) against Escherichia coli, Salmonella Enteritidis, Salmonella serotype (ser.) Typhimurium, Staphylococcus intermedius, Staph. chromogenes, Proteus mirabillis, Areomonas veonii, Bordetella bronchioseptica and Klebsialla oxytoca. The isolate Lacp28 exhibited highest tolerance to acidic pH and bile salts (up to 0.3%), phytase activity, cell surface hydrophobicity, antagonistic activity and co-culture assay (>80% growth inhibition). Host specificity of Lacp28 was further confirmed by heavy in vitro adhesion to pig intestinal epithelium cells compared to chicken. Hence, Lacp28 was selected and identified by phylogenetic analysis of 16S rRNA as Pediococcus acidilactici strain FT28 with 100% similarity (GenBank accession nos. KU837245, KU837246 and KU837247). The Pediococcus acidilactici FT28 was selected as potential probiotic candidature for in vivo efficacy in weaned pigs. Thirty-six crossbred piglets (28d) were randomly distributed into three
PLOS ONE | https://doi.org/10.1371/journal.pone.0192978 March 8, 2018
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Phylogenetic analysis of species-specific probiotic and its impact on early weaned piglets
decision to publish, or preparation of the manuscript. Competing interests: The authors have declared that no competing interests exist.
groups (four replicates of three each) namely, basal diet without probiotics (T0) or with Lactobacillus acidophilus NCDC15 (conventional dairy-specific probiotic; T1) or Pediococcus acidilactici FT28 (swine-specific probiotic; T2). At end of the experiment, six piglets of similar body weight were selected to conduct digestion trial for estimation of nutrient digestibility. Results of the study indicated that supplementation of both probiotics improved (P
