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<p>Probiotics play vital roles in maintaining gut health and overall well-being of animals, and have been isolated from the guts of several animals. The caecotrophic and dynamic nature of rabbit gut make them a promising site for probiotic prospecting. This study therefore, investigated the probiotic properties and whole genome sequence profile of Lactic Acid Bacteria (LAB) isolated from the gut microflora of domestic rabbit. Gastrointestinal contents of rabbits were cultured anaerobically on de Mann Rogosa Sharpe (MRS) agar using the pour-plate method. Isolates obtained were initially identified using 16S rRNA sequencing and later using Whole genome sequencing. Probiotic properties of the LAB were determined using conventional in vitro tests such as acid and phenol tolerance, adhesion ability, susceptibility to commonly-used antibiotics and the antimicrobial activity of their cell-free supernatants against selected test microorganisms. Whole genome analysis was carried out on selected isolates to further confirm their probiotic and safety properties.</p><p>Three LAB showing probiotic properties were obtained and identified: <i>Enterococcus lactis</i> UIADO22, <i>E. lactis</i> UIADO37, and <i>Enterococcus hirae</i> UIADO30. All isolates were non-haemolytic, DNAse-negative, tolerant to pH 2, and maintaining viability in simulated duodenal juice. They were all sensitive to the tested antibiotics and their cell-free supernatants demonstrated antibacterial activity against selected test bacteria. Whole genome sequencing revealed the absence of vancomycin resistance and virulence genes (<i>IS16</i>, <i>esp</i>, <i>hyl</i>) in the LAB, confirming their safety. Probiotic potential was further supported by the detection of genes encoding bacteriocins such as <i>Enterolysin_A</i>,<i> Subtilosin_A</i>,<i> Enterocin_L50a/L50b</i>,<i> Lanthionine synthetase</i>, and <i>Enterocin_P</i>. Biosynthetic gene clusters for antimicrobial metabolites, including cyclic-lactone autoinducers, Type III polyketide synthase, terpene precursors, sactipeptides, RIPP-like peptides, and Lanthipeptide Class II, were also detected. Genes associated with stress resistance especially bile, acid and phenol (such as <i>dltA</i>, <i>dltB</i>, <i>ssb</i> and <i>dltC</i>), DNA/protein repair (such as <i>recA</i>, <i>uvrA</i>, <i>uvrB</i>,and <i>recN</i>), adhesion and anti-pathogenic activity (such <i>agrB</i>,<i> agrD</i>,<i> fsrA</i>,and <i>fbp3)</i> were detected in the genomes of the three LAB. Overall, the rabbit-derived <i>Enterococcus</i> spp. from this study exhibited both in vitro and genomic characteristics of safe, functional probiotics with strong antimicrobial potential. <i>Enterococcus hirae</i> UIADO30 is the most promising isolate because it lacks plasmids and virulence markers thereby reducing the risk of gene transfer—while carrying markers for bile and acid resistance. </p>

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Functional and Genomic insights into probiotic Lactic Acid Bacteria isolated from the Gastrointestinal microbiota of domestic rabbits (Oryctolagus cuniculus)

  • 期刊论文

  • 出版模式：

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  • 发表日期：

    2026年4月9日

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  • Oluwatosin Olubunmi Oladipo,
  • Abimbola Olumide Adekanmbi,
  • Opeyemi U. Lawal,
  • Valeria R. Parreira,
  • Bolaji Fatai Oyeyemi,
  • Olatunji Abubakar Jimoh,
  • Aderemi Akinyemi,
  • Charles Ayorinde Ologunde,
  • Ayonposi Bukola Olaoye,
  • Olugbenga David Oloruntola,
  • Mitra Soni,
  • Harmanpreet Kaur,
  • Lawrence Goodridge

摘要

Probiotics play vital roles in maintaining gut health and overall well-being of animals, and have been isolated from the guts of several animals. The caecotrophic and dynamic nature of rabbit gut make them a promising site for probiotic prospecting. This study therefore, investigated the probiotic properties and whole genome sequence profile of Lactic Acid Bacteria (LAB) isolated from the gut microflora of domestic rabbit. Gastrointestinal contents of rabbits were cultured anaerobically on de Mann Rogosa Sharpe (MRS) agar using the pour-plate method. Isolates obtained were initially identified using 16S rRNA sequencing and later using Whole genome sequencing. Probiotic properties of the LAB were determined using conventional in vitro tests such as acid and phenol tolerance, adhesion ability, susceptibility to commonly-used antibiotics and the antimicrobial activity of their cell-free supernatants against selected test microorganisms. Whole genome analysis was carried out on selected isolates to further confirm their probiotic and safety properties.

Three LAB showing probiotic properties were obtained and identified: Enterococcus lactis UIADO22, E. lactis UIADO37, and Enterococcus hirae UIADO30. All isolates were non-haemolytic, DNAse-negative, tolerant to pH 2, and maintaining viability in simulated duodenal juice. They were all sensitive to the tested antibiotics and their cell-free supernatants demonstrated antibacterial activity against selected test bacteria. Whole genome sequencing revealed the absence of vancomycin resistance and virulence genes (IS16, esp, hyl) in the LAB, confirming their safety. Probiotic potential was further supported by the detection of genes encoding bacteriocins such as Enterolysin_A, Subtilosin_A, Enterocin_L50a/L50b, Lanthionine synthetase, and Enterocin_P. Biosynthetic gene clusters for antimicrobial metabolites, including cyclic-lactone autoinducers, Type III polyketide synthase, terpene precursors, sactipeptides, RIPP-like peptides, and Lanthipeptide Class II, were also detected. Genes associated with stress resistance especially bile, acid and phenol (such as dltA, dltB, ssb and dltC), DNA/protein repair (such as recA, uvrA, uvrB,and recN), adhesion and anti-pathogenic activity (such agrB, agrD, fsrA,and fbp3) were detected in the genomes of the three LAB. Overall, the rabbit-derived Enterococcus spp. from this study exhibited both in vitro and genomic characteristics of safe, functional probiotics with strong antimicrobial potential. Enterococcus hirae UIADO30 is the most promising isolate because it lacks plasmids and virulence markers thereby reducing the risk of gene transfer—while carrying markers for bile and acid resistance.