Abstract
The pharmaceutical industry shows an emerging interest in formulas that contain live and beneficial microorganisms, also known as probiotics or pharmabiotics, which in many cases, are host-specific. The resistance to higher temperature is an essential feature of these microorganisms when working on the design of products for vaginal formula. In order to obtain a high number of viable cells and a prolonged shelf life in the designed product, it is required to apply technological procedures using high temperatures or abrupt changes of them, which result in conditions that are different from the optimal growth temperature and can affect the metabolic capabilities of the bacteria when administered to the host in order to reestablish the ecological mucosa. The aim of this work was to evaluate the behavior of 30 different species and strains of autochthonous beneficial vaginal lactobacilli (BVL) when exposed to high temperatures, determine their survival capabilities and analyze their pre-adaptation to those temperatures, in order that they still maintain their viability after technological processes and further conservation. BVL were exhibited to temperatures higher than optimal, with the purpose of evaluating their growth kinetics and parameters. Later, they were exposed to higher temperatures, and then, returned to their optimal, to determine if they were able to grow again. The strains that showed higher resistance were selected, and their viability and beneficial properties studied further. The growth kinetics of strains exposed to higher temperatures showed different patterns, which provided evidence that the thermal adaptation is strain-dependent and is not related to any particular species and/or metabolic group in which the strains were taxonomically classified. The pre-adaptive step allowed the growth of some of the strains, preserving their viability and probiotic properties after the high temperatures were applied. The results shows that BVL can be exposed to high temperatures used in different technological processes that are applied for pharmabiotic formulations, such as spray dried or vacuum rotary evaporation, and/or during the conservation period. The results obtained indicate that some specific BVL strains resist high temperatures and grow afterwards at optimal conditions.
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Acknowledgements
This work was supported by CONICET (PIP 545 and 744) and ANPCYT (PICT 4324) and was performed under the cooperation agreement between Mincyt and BMBF (01DN14006). Some of the strains were licensed through a CONICET-ANIDRAL agreement.
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JAS. Experimental procedures. References acquisition. Draft writing. Edition of results. AM. Experimental procedures. Draft of results. BW. Validation. Statistical evaluation. MEFN: Conceptualization; Original draft; Formal analysis; Funding acquisition; Investigation; Methodology; Supervision; Writing - Review & editing.
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Jessica Alejandra Silva and Antonella Marchesi have participated equally performing experimental protocols and in the preparation of graphs and tables.
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Fig. 1A–B. Supp. material. Experimental protocol applied to determine the resistance to different temperatures. 1B. Pre-adaptation of the strains to higher temperatures and further growth at 37 °C (PDF 537 kb)
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Fig. 4. Supp. material. Growth Kinetics of vaginal lactobacilli strains incubated 9 hours at 50 °C and transferred to 37 °C. The methods are described in the text and plotted in Figure 1B (supp. mat.) (PDF 74 kb)
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Fig. 5. Supp. material. Growth Kinetics of vaginal lactobacilli strains incubated 9 hours at 55 °C and transferred at 37 °C. The methods are described in the text and plotted in Fig 1B (supp. mat.) (PDF 70 kb)
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Fig. 6. Supp. material. Self-aggregation pattern of vaginal Lactobacillus strains: (a) L. gasseri CRL1320 and (b) L. rhamnosus CRL1511 monitored spectrophotometrically (A600nm) during 4 h. Continue black plot represents the pattern before (B-HT) and grey line after exposure at high temperature (A-HT). Self-aggregation patterns of selected BVL strains B-HT (c) and A-HT (d). Different letters indicate statistically significant differences (P < 0.05) in the OD values determined at 4 h B-HT and A-HT for each strain (PDF 328 kb)
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Fig. 7. Supp. material. Hydrophobicity (toluene and xylene) of Lactobacillus. In the 3D graph the columns represent the percentage of hydrophobicity with xylene or toluene of selected BVL strains before (B-HT) and after (A-HT) the exposure to higher temperatures. The degree of hydrophobicity was represented as the score: high (71–100%), medium (36–70%), and low (0–35%). Different letters indicate statistically significant differences (P < 0.05) for each strain in a same solvent B-HT and A-HT (PDF 430 kb)
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Silva, J.A., Marchesi, A., Wiese, B. et al. Screening of autochthonous vaginal beneficial lactobacilli strains by their growth at high temperatures for technological applications. Antonie van Leeuwenhoek 113, 1393–1409 (2020). https://doi.org/10.1007/s10482-020-01431-4
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DOI: https://doi.org/10.1007/s10482-020-01431-4