Rapid optimization of spore production from Bacillus amyloliquefaciens in submerged cultures based on dipicolinic acid fluorimetry assay

Ren, Hang; Su, Ya-ting; Guo, Xiaohua

doi:10.1186/s13568-018-0555-x

Cited by 13 publications

(8 citation statements)

References 23 publications

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“…Therefore, the LOD may depend on desirable concentrations of TbCl 3 , and the concentrations of reagents are often adjusted before detection (Hindle and Hall 1999 ). However, in fermentation for spore production, the spore concentration often relies on the culture time and fermentation conditions, and many analytes with different spore concentrations produced in spore optimization designs must be detected simultaneously (Ren et al 2018 ). Therefore, the DPA-Tb fluorimetry assay cannot be used as a general analytical method for rapid endospore quantification.…”

Section: Discussionmentioning

confidence: 99%

“…However, the method in this study is simpler, and the fluorescent complex is readily prepared by just mixing the DPA samples, Eu, and CyDTA in the correct proportion. The method is especially suitable for the circumstances of the spore detection in fermentation optimization and quality control and also for the simultaneous detection of endospores in a large number of samples (Ren et al 2018 ).…”

Section: Discussionmentioning

confidence: 99%

“…The interference can be eliminated if the concentration is diluted to less than 10 μM. In practical samples from spore fermentation, the spore suspensions are often treated by washing and above 100-fold dilutions (Ren et al 2018 ). The possible metal ions are then greatly diluted, and their interference is likely to be negligible.…”

Section: Discussionmentioning

confidence: 99%

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Rapid and simple detection of endospore counts in probiotic Bacillus cultures using dipicolinic acid (DPA) as a marker

et al. 2018

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Spore counting in probiotic Bacillus cultures using dipicolinic acid (DPA) as a marker was studied for developing a rapid and simple detection method. The newly developed method is based on the fluorescence enhancement by a new chelating agent, which forms a complex with EuCl3 and DPA. The results showed that 1,2-cyclohexanediamine-N,N,N′N′-tetraacetic acid (CyDTA) greatly enhanced the fluorescence intensity in all selected chelating agents. The optimal composition of the fluorescence complex DPA-Eu-CyDTA had a detection limit of 0.3 nM of DPA. Metal ions in high concentrations, including Cu2+, Fe2+, Fe3+, Al3+, and Zn2+, might lower the detection sensitivity, which could be eliminated by diluting the sample with the metal ions below 10 μM. The maximum release of DPA was achieved by heating treatments at 121 °C for at least 10 min for two types of Bacillus endospores. The spore concentrations and corresponding released DPA fluorescence intensities were linearly associated (coefficient R2 = 0.9993 and 0.9995 for Bacillus subtilis MA139 and Bacillus licheniformis BL20386, respectively). The detection limit for both strains reached approximately 6800 spores/mL. The verification results showed that the DPA fluorimetry assay developed in the present study was fully consistent with the plate-counting assay. The study shows that the fluorescence complex DPA-Eu-CyDTA can be reliably used for the detection of endospores in Bacillus fermentation for the production of probiotics.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

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Rapid and simple detection of endospore counts in probiotic Bacillus cultures using dipicolinic acid (DPA) as a marker

et al. 2018

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“…Sporulation of Bacillus spp. depends on medium composition, pH, aeration, temperature, and other factors (Ren et al 2018; Posada-Uribe et al 2015).…”

Section: Determination Of the Growth Promoting Effect Of B Amyloliquefaciens 3-5 On Cucumbermentioning

confidence: 99%

Optimizing the Fermentation Conditions and Determining the Disease Control and Growth Promotion Function of Bacillus Amyloliquefaciens 3-5

Ma¹,

Yang²,

Cai³

et al. 2021

Preprint

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Bacillus spp. are helping to develop towards sustainable agriculture and have become a research hotspot in the field of plant pathology because they have great development potential such as control fungal plant diseases. Bacillus Amyloliquefaciens 3-5 was used as antagonistic bacteria in this experiment. The optimal medium for solid-state fermentation of B. amyloliquefaciens 3-5 contained bran (35%), rice hull powder (40%), corn gluten (20%), bean flour (15%), corn starch (1.5%), beef extract (2.5%) and MgSO4 (1.5%), and the optimal fermentation conditions included an inoculum of 6%, a solids content of 36 g/L, a feed-to-water ratio of 1:1, a fermentation temperature of 32 ℃, an initial pH of 7.0 and fermentation time of 44 h. When the dosage of the B. amyloliquefaciens 3-5 agent was 10%, the thick stems, root length and plant height of cucumber were significantly higher than those of the control (P＜0.05), and the growth rates were 77.45 %, 35.77 %, and 53.33 %, respectively in pot experiments. Compared with the control, and the preventive effect and therapeutic effect on cucumber Fusarium wilt were 72.09 % and 48.83 % by the application of B. amyloliquefaciens 3-5 agent，which showed that cucumber Fusarium wilt was successfully controlled by a newly isolated strain of B. Amyloliquefaciens 3-5. These results suggested that the prevention and control effect of B. amyloliquefaciens 3-5 agent on cucumber Fusarium wilt could not be underestimated.

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“…The proposed low-cost medium was optimized to have a concentration that maximized spore production using the central composite design. In order to further promote spore production, the following inorganic salts were added: 164 mg/l Ca(NO [26,27]. To evaluate the effectiveness of the proposed medium, it was compared to the existing commercial Difco sporulation medium (DSM), which is composed of 8 g/l nutrient broth (Difco), 1 g/l KCl, and 0.25 g/l MgSO were then added in the form of filter-sterilized solutions.…”

Section: Confirmation Experiments and Comparison With A Commercial Mediummentioning

confidence: 99%

Low-cost cultivation and sporulation of alkaliphilic Bacillus sp. strain AK13 for self-healing concrete

Hong¹,

Kim²,

Park³

2019

Journal of Microbiology and Biotechnology

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The alkaliphilic, calcium carbonate precipitating Bacillus sp. strain AK13 can be utilized in concrete for self-repairing. A statistical experimental design was used to develop an economical medium for its mass cultivation and sporulation. Two types of screening experiment were first conducted to identify substrates that promote the growth of the AK13 strain: the first followed a one-factor-at-a-time factorial design and the second a two-level full factorial design. Based on these screening experiments, barley malt powder and mixed grain powder were identified as the substrates that most effectively promoted the growth of the AK13 strain from a range of 21 agricultural products and by-products. A quadratic statistical model was then constructed using a central composite design and the concentration of the two substrates was optimized. The estimated growth and sporulation of Bacillus sp. strain AK13 in the proposed medium were 3.08 ± 0.38 × 10 8 and 1.25 ± 0.12 × 10 8 CFU/ml, respectively, which meant that the proposed low-cost medium was approximately 45 times more effective than the commercial medium in terms of the number of cultivatable bacteria per unit price. The spores were then powdered via a spray-drying process to produce a spore powder with a spore count of 2.0 ± 0.7 × 10 9 CFU/g. The AK13 spore powder was mixed with cement paste, yeast extract, calcium lactate, and water. The yeast extract and calcium lactate generated the highest CFU/ml for AK13 at a 0.4:0.4 ratio compared to 0.4:0.25 (the original ratio of the B4 medium) and 0.4:0.8. Twenty-eight days after the spores were mixed into the mortar, the number of vegetative cells and spores of the AK13 strain had reached 10 6 CFU/g within the mortar. Cracks in the mortar under 0.29 mm were healed in 14 days. Calcium carbonate precipitation was observed on the crack surface. The mortar containing the spore powder was thus concluded to be effective in terms of healing micro-cracks.

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Rapid optimization of spore production from Bacillus amyloliquefaciens in submerged cultures based on dipicolinic acid fluorimetry assay

Cited by 13 publications

References 23 publications

Rapid and simple detection of endospore counts in probiotic Bacillus cultures using dipicolinic acid (DPA) as a marker

Rapid and simple detection of endospore counts in probiotic Bacillus cultures using dipicolinic acid (DPA) as a marker

Optimizing the Fermentation Conditions and Determining the Disease Control and Growth Promotion Function of Bacillus Amyloliquefaciens 3-5

Low-cost cultivation and sporulation of alkaliphilic Bacillus sp. strain AK13 for self-healing concrete

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