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Background <p>Recent advances in swine gut culturomics have substantially expanded our understanding of the cultivable repertoire and revealed the compositional complexity of the porcine gut microbiota. Nevertheless, how multiple environmental and cultivation-related factors interact to shape cultured communities under controlled nutrient conditions remains poorly understood. This study evaluated the relative and interactive contributions of intestinal segment, oxygen availability, cultivation mode, and carbon-substrate identity within a standardized nutrient-rich basal medium framework. Using inocula from the ileum and colon of Wuzhishan piglets, a roughage-tolerant Chinese indigenous breed, microbial communities were cultured on modified yeast extract casitone fatty acid (YCFA) media in which the original carbon substrate was replaced with ten alternative carbohydrates representing simple and complex substrates.</p> Results <p>Culture-dependent (CD) sequencing showed that intestinal segment was the dominant factor shaping cultured communities (<i>P</i> &lt; 0 0.001), followed by oxygen availability (<i>P</i> &lt; 0 0.001) and donor identity (<i>P</i> &lt; 0 0.001), whereas, when evaluated within a nutrient-rich basal medium, carbon-substrate identity did not function as an independent or strongly additive driver of community differentiation (<i>P</i> &gt; 0 0.05). A total of 1,308 isolates belonging to 67 genera were obtained through an empirical streaking strategy, including several rare taxa such as <i>Culturomica</i> that were below sequencing detection thresholds, revealing a disparity between CD sequencing profiles and actual strain isolation. Integration of community composition with short-chain fatty acid (SCFA) profiles further indicated that metabolic accumulation represents an additional ecological dimension influencing cultured community differentiation.</p> Conclusions <p>Our findings demonstrate that under multi-factor cultivation frameworks, ecological filters do not operate in a freely additive manner. Instead, intestinal origin, oxygen exposure, and metabolic context collectively constrain culturability, while carbon-substrate variation alone does not independently drive community clustering within nutrient-rich conditions. Together, this study provides a refined conceptual framework for interpreting culture-dependent outcomes and for designing multi-factor cultivation strategies in future swine gut culturomics research.</p>

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Exploring the predominant factors associated with shaping the cultivable swine gut microbiota of Wuzhishan piglets under in vitro cultivation conditions

  • 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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  • Yinfeng Chen,
  • Xin Zhang,
  • Xiaozhe Wang,
  • Zhenlong Wu

摘要

Background

Recent advances in swine gut culturomics have substantially expanded our understanding of the cultivable repertoire and revealed the compositional complexity of the porcine gut microbiota. Nevertheless, how multiple environmental and cultivation-related factors interact to shape cultured communities under controlled nutrient conditions remains poorly understood. This study evaluated the relative and interactive contributions of intestinal segment, oxygen availability, cultivation mode, and carbon-substrate identity within a standardized nutrient-rich basal medium framework. Using inocula from the ileum and colon of Wuzhishan piglets, a roughage-tolerant Chinese indigenous breed, microbial communities were cultured on modified yeast extract casitone fatty acid (YCFA) media in which the original carbon substrate was replaced with ten alternative carbohydrates representing simple and complex substrates.

Results

Culture-dependent (CD) sequencing showed that intestinal segment was the dominant factor shaping cultured communities (P < 0 0.001), followed by oxygen availability (P < 0 0.001) and donor identity (P < 0 0.001), whereas, when evaluated within a nutrient-rich basal medium, carbon-substrate identity did not function as an independent or strongly additive driver of community differentiation (P > 0 0.05). A total of 1,308 isolates belonging to 67 genera were obtained through an empirical streaking strategy, including several rare taxa such as Culturomica that were below sequencing detection thresholds, revealing a disparity between CD sequencing profiles and actual strain isolation. Integration of community composition with short-chain fatty acid (SCFA) profiles further indicated that metabolic accumulation represents an additional ecological dimension influencing cultured community differentiation.

Conclusions

Our findings demonstrate that under multi-factor cultivation frameworks, ecological filters do not operate in a freely additive manner. Instead, intestinal origin, oxygen exposure, and metabolic context collectively constrain culturability, while carbon-substrate variation alone does not independently drive community clustering within nutrient-rich conditions. Together, this study provides a refined conceptual framework for interpreting culture-dependent outcomes and for designing multi-factor cultivation strategies in future swine gut culturomics research.