Effects of the overexpression of fructose-1,6-bisphosphate aldolase on fermentation pattern and transcription of the genes encoding lactate dehydrogenase and pyruvate formate-lyase in a ruminal bacterium, Streptococcus bovis

Asanuma, Narito; Yoshii, Toshitaka; Kikuchi, Masahiro; Hino, Tsuneo

doi:10.2323/jgam.50.71

Cited by 10 publications

(6 citation statements)

References 21 publications

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“…The higher FDP concentration in isolate S1 grown in 9 g/liter starch medium could be explained partly by its higher proportion compared to growth in 1 g/liter starch medium. Similar to a previous report (39), the FDP concentration was higher when rapidly fermentable glucose was used as the substrate. Consequently, there was more lactate produced and less production of formate with the increase of soluble starch in the media.…”

Section: Discussionsupporting

confidence: 77%

Effects of Glucose and Starch on Lactate Production by Newly Isolated Streptococcus bovis S1 from Saanen Goats

Chen

Luo

Wang

et al. 2016

Appl Environ Microbiol

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When ruminants are fed high-concentrate diets, Streptococcus bovis proliferates rapidly and produces lactate, potentially causing rumen acidosis. Understanding the regulatory mechanisms of the metabolism of this species might help in developing dietary strategies to alleviate rumen acidosis. S. bovis strain S1 was newly isolated from the ruminal fluid of Saanen dairy goats and then used to examine the effects of glucose and starch on bacterial metabolism and gene regulation of the organic acid-producing pathway in cultures at a pH of 6.5. Glucose or starch was added to the culture medium at 1 g/liter, 3 g/liter (close to a normal range in the rumen fluid), or 9 g/liter (excessive level). Lactate was the dominant acid produced during the fermentation, and levels increased with the amount of glucose or starch in a dose-dependent manner (P < 0.001). The production of formate and acetate in the fermentation media fluctuated slightly with the dose but accounted for small fractions of the total acids. The activities of lactate dehydrogenase (LDH) and ␣-amylase (␣-AMY) increased with the starch dose (P < 0.05), but the ␣-AMY activity did not change with the glucose dose. The relative expression levels of the genes ldh, pfl (encoding pyruvate formate lyase), ccpA (encoding catabolite control protein A), and ␣-amy were higher at a dose of 9 g/liter than at 1 g/liter (P < 0.05). Expression levels of pfl and ␣-amy genes were higher at 3 g/liter than at 1 g/liter (P < 0.05). The fructose 1,6-diphosphate (FDP) concentration tended to increase with the glucose and starch concentrations. In addition, the S. bovis S1 isolate fermented glucose much faster than starch. We conclude that the quantities of glucose and soluble starch had a major effect on lactate production due to the transcriptional regulation of metabolic genes. IMPORTANCEThis work used a newly isolated S. bovis strain S1 from the rumen fluid of Saanen goats and examined the effects of glucose and soluble starch on organic acid patterns, enzyme activity, and expression of genes for in vitro fermentation. It was found that lactate was the dominant product from S. bovis strain S1, and the quantities of both glucose and starch in the medium were highly correlated with lactate production and with the corresponding changes in associated enzymes and genes. Therefore, manipulating the metabolic pathway of S. bovis to alter the dietary level of readily fermentable sugar and carbohydrates may be a strategy to alleviate rumen acidosis. Streptococcus bovis is the most prominent bacterium in the rumen. Because S. bovis is lactate producing and acid tolerant, it plays an important role in rumen acidosis (1). Our previous study with dairy cows demonstrated that under normal conditions, the population of S. bovis is small, resulting in an insignificant amount of lactate production that is quickly metabolized by lactate utilizers such as Selenomonas ruminantium and Megasphaera elsdenii (2, 3). However, when a high-concentrate diet is provided, the growth rate of S. bovis i...

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Section: Discussionsupporting

confidence: 77%

Effects of Glucose and Starch on Lactate Production by Newly Isolated Streptococcus bovis S1 from Saanen Goats

Chen

Luo

Wang

et al. 2016

Appl Environ Microbiol

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“…The sources of Streptococcus bovis JB1 (ATCC 700410), 12U1 (highly transformable strain), 12U1‐ ccpA − [null mutation of the CcpA gene ( ccpA ) in 12U1], and 12U1 p (12U1‐harboring pSBE11) were described previously (Asanuma & Hino, 2002a; Asanuma et al , 2004a, b). Except where otherwise indicated, the JB1 strain was used.…”

Section: Methodsmentioning

confidence: 99%

Presence of NADP⁺-specific glyceraldehyde-3-phosphate dehydrogenase and CcpA-dependent transcription of its gene in the ruminal bacteriumStreptococcus bovis

Asanuma

Hino

2006

FEMS Microbiology Letters

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To know the role of NADP+-specific glyceraldehyde-3-phosphate dehydrogenase (GAPN) in Streptococcus bovis, the molecular properties and transcriptional control of the gene encoding GAPN (gapN) were examined. The GAPN in S. bovis was deduced to consist of 476 amino acids with a molecular mass of 51.1 kDa. The gapN gene was transcribed in a monocistronic fashion. GAPN synthesis appeared to be regulated at the transcriptional level in response to changes in growth conditions. In a mutant that lacks the ccpA gene encoding catabolite control protein A (CcpA), the gapN-mRNA level was lower than in the parent strain. A binding site of CcpA was found in the upper region of gapN. These results suggest that transcription of gapN is regulated through CcpA. Overexpression of GAPN in S. bovis did not affect the growth rate or formate-to-lactate ratio, suggesting that the flux in the glycolytic pathway is unlikely to be altered by GAPN activity. Streptococcus bovis GAPN was NADP+ dependent, but not phosphate dependent. In addition, S. bovis did not have other NADPH-producing systems such as the hexose monophosphate pathway and NADPH:NAD+ oxidoreductase. Therefore, GAPN may play an important role in NADPH production in S. bovis.

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“…Correspondingly, we observed a decrease in FBP in S. bovis S1 growing in glucose-limited media as compared to glucose-rich media. LDH activity can be induced by FBP in S. bovis ( Wolin, 1964 ), and FBP concentration is positively correlated with the levels of ldh -mRNA and inversely correlated with the pfl -mRNA levels ( Asanuma et al, 2004b ). In this study, the increased lactate and decreased formate production in glucose-rich media might be associated with the increased FBP concentration.…”

Section: Discussionmentioning

confidence: 99%

Transcriptome Analysis Reveals Catabolite Control Protein A Regulatory Mechanisms Underlying Glucose-Excess or -Limited Conditions in a Ruminal Bacterium, Streptococcus bovis

et al. 2021

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Ruminants may suffer from rumen acidosis when fed with high-concentrate diets due to the higher proliferation and overproduction of lactate by Streptococcus bovis. The catabolite control protein A (CcpA) regulates the transcription of lactate dehydrogenase (ldh) and pyruvate formate-lyase (pfl) in S. bovis, but its role in response to different carbon concentrations remains unclear. To characterize the regulatory mechanisms of CcpA in S. bovis S1 at different levels of carbon, herein, we analyzed the transcriptomic and physiological characteristics of S. bovis S1 and its ccpA mutant strain grown in glucose-excess and glucose-limited conditions. A reduced growth rate and a shift in fermentation pattern from homofermentation to heterofermentation were observed under glucose-limited condition as compared to glucose-excess condition, in S. bovis S1. Additionally, the inactivation of ccpA significantly affected the growth and end metabolites in both conditions. For the glycolytic intermediate, fructose 1,6-bisphosphate (FBP), the concentration significantly reduced at lower glucose conditions; its concentration decreased significantly in the ccpA mutant strain. Transcriptomic results showed that about 46% of the total genes were differentially transcribed between the wild-type strain and ccpA mutant strain grown in glucose-excess conditions; while only 12% genes were differentially transcribed in glucose-limited conditions. Different glucose concentrations led to the differential expression of 38% genes in the wild-type strain, while only half of these were differentially expressed in the ccpA-knockout strain. Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses showed that the substrate glucose concentration significantly affected the gene expression in histidine metabolism, nitrogen metabolism, and some carbohydrate metabolism pathways. The deletion of ccpA affected several genes involved in carbohydrate metabolism, such as glycolysis, pyruvate metabolism, fructose and mannose metabolism, as well as in fatty acid biosynthesis pathways in bacteria grown in glucose-excess conditions; this effect was attenuated under glucose-limited conditions. Overall, these findings provide new information on gene transcription and metabolic mechanisms associated with substrate glucose concentration and validate the important role of CcpA in the regulation of carbon metabolism in S. bovis S1 at differential glucose availability.

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Effects of the overexpression of fructose-1,6-bisphosphate aldolase on fermentation pattern and transcription of the genes encoding lactate dehydrogenase and pyruvate formate-lyase in a ruminal bacterium, Streptococcus bovis

Cited by 10 publications

References 21 publications

Effects of Glucose and Starch on Lactate Production by Newly Isolated Streptococcus bovis S1 from Saanen Goats

Effects of Glucose and Starch on Lactate Production by Newly Isolated Streptococcus bovis S1 from Saanen Goats

Presence of NADP⁺-specific glyceraldehyde-3-phosphate dehydrogenase and CcpA-dependent transcription of its gene in the ruminal bacteriumStreptococcus bovis

Transcriptome Analysis Reveals Catabolite Control Protein A Regulatory Mechanisms Underlying Glucose-Excess or -Limited Conditions in a Ruminal Bacterium, Streptococcus bovis

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Effects of the overexpression of fructose-1,6-bisphosphate aldolase on fermentation pattern and transcription of the genes encoding lactate dehydrogenase and pyruvate formate-lyase in a ruminal bacterium, Streptococcus bovis

Cited by 10 publications

References 21 publications

Effects of Glucose and Starch on Lactate Production by Newly Isolated Streptococcus bovis S1 from Saanen Goats

Effects of Glucose and Starch on Lactate Production by Newly Isolated Streptococcus bovis S1 from Saanen Goats

Presence of NADP+-specific glyceraldehyde-3-phosphate dehydrogenase and CcpA-dependent transcription of its gene in the ruminal bacteriumStreptococcus bovis

Transcriptome Analysis Reveals Catabolite Control Protein A Regulatory Mechanisms Underlying Glucose-Excess or -Limited Conditions in a Ruminal Bacterium, Streptococcus bovis

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Presence of NADP⁺-specific glyceraldehyde-3-phosphate dehydrogenase and CcpA-dependent transcription of its gene in the ruminal bacteriumStreptococcus bovis