Identification, Biochemical Characterization, and Safety Attributes of Locally Isolated Lactobacillus fermentum from Bubalus bubalis (buffalo) Milk as a Probiotic

Sana Abid; Arshad Farid; Rameesha Abid; Mujeeb Ur Rehman; Walaa F Alsanie; Majid Alhomrani; Abdulhakeem S Alamri; Syed Mohammed Basheeruddin Asdaq; Daniel Ingo Hefft; Saddam Saqib; Muhammad Muzammal; Sabrin Abdelrahman Morshedy; Mashael W Alruways; Shakira Ghazanfar

doi:10.3390/microorganisms10050954

Identification, Biochemical Characterization, and Safety Attributes of Locally Isolated Lactobacillus fermentum from Bubalus bubalis (buffalo) Milk as a Probiotic

Microorganisms. 2022 Apr 30;10(5):954. doi: 10.3390/microorganisms10050954.

Authors

Sana Abid¹, Arshad Farid², Rameesha Abid^{3

4}, Mujeeb Ur Rehman⁵, Walaa F Alsanie^{6

7}, Majid Alhomrani^{6

7}, Abdulhakeem S Alamri^{6

7}, Syed Mohammed Basheeruddin Asdaq⁸, Daniel Ingo Hefft⁹, Saddam Saqib¹⁰, Muhammad Muzammal², Sabrin Abdelrahman Morshedy¹¹, Mashael W Alruways¹², Shakira Ghazanfar⁴

Affiliations

¹ Department of Biology, Faculty of Science & Technology, Virtual University, Lahore 54000, Pakistan.
² Gomal Centre of Biochemistry and Biotechnology, Gomal University, D.I.Khan 29050, Pakistan.
³ Department of Biotechnology, University of Sialkot, Sialkot 51310, Pakistan.
⁴ National Institute of Genomics and Advanced Biotechnology (NIGAB), National Agricultural Research Centre, Park Road, Islamabad 45500, Pakistan.
⁵ Department of Pharmacy, Quaid-i-Azam University, Islamabad 45500, Pakistan.
⁶ Department of Clinical Laboratory Sciences, The Faculty of Applied Medical Sciences, Taif University, Al Hawiyah 21944, Saudi Arabia.
⁷ Centre of Biomedical Sciences Research (CBSR), Deanship of Scientific Research, Taif University, Al Hawiyah 21944, Saudi Arabia.
⁸ Department of Pharmacy Practice, College of Pharmacy, AlMaarefa University, Dariyah 13713, Saudi Arabia.
⁹ Reaseheath College, University Centre Reaseheath, Nantwich CW5 6DF, UK.
¹⁰ Department of Biotechnology, Mohi- ud-Din Islamic University, Nerian Sharif 12080, Pakistan.
¹¹ Fish and Animal Production Department, Faculty of Agriculture (Saba Basha), Alexandria University, Alexandria 21526, Egypt.
¹² Department of Clinical Laboratory Sciences, College of Applied Medical Sciences, Shaqra University, Shaqra 15273, Saudi Arabia.

Abstract

The demand of functional foods is on the rise, and researchers are trying to develop nutritious dairy products by using well-characterized strains of bacteria. In this study, we identified locally isolated strains of Lactobacillus fermentum from Bubalus bubalis (Nilli Ravi buffalo) milk and evaluated their potential as probiotics in food products like fermented milk. Fifteen Lactobacillus strains were initially isolated, and only four strains (NMCC-2, NMCC-14, NMCC-17, and NMCC-27) were examined for morphological and biochemical characterizations due to their ability of gas production in Durham tubes. Moreover, these strains were selected for further probiotic characterizations due to their extreme morphological resemblance with lactic acid bacteria for their antimicrobial activity, enzymatic potential, autoaggregation capability, hydrophobicity, and acid and bile tolerance. All selected isolates showed significant probiotic potential. However, NMCC-14 and NMCC-17 strains showed maximum probiotic potential. The isolates (NMCC-2, NMCC-14, NMCC-17, and NMCC-27) were identified as Lactobacillus fermentum utilizing 16S rRNA gene sequencing. The in vivo safety study of NMCC-14 (dose: 1010 CFU/day/mice; 21 days, orally) showed no histological dysfunctions in a mouse model. Pathogenic bacterial enzymes reduced the beneficial bacterial load in the host gastrointestinal tract. These results suggest that the NMCC-14 strain is safe and can be potentially used as a probiotic. Moreover, fermented milk was prepared by using the NMCC-14 strain. The results revealed that NMCC-14 strain-based fermented milk had significantly (p < 0.05) higher protein content (4.4 ± 0.06), water-holding capacity (WHC), and dynamic viscosity as compared to non-fermented milk. The results suggest that L. fermentum NMCC-14 is safe and nontoxic; hence, it can be a beneficial supplement to be used for the development of dairy products to be subjected to further clinical testing.

Keywords: Lactobacillus fermentum; fermented milk; identification; probiotics; safety attributes.

Grants and funding

The authors gratefully acknowledge the National Institute for Genomics and Advanced Biotechnology (NIGAB), National Agricultural Research Centre, Islamabad, Pakistan, for providing financial support and lab facility for this research work. Abdulhakeem S. Alamri would like to acknowledge Taif university for support, No. TURSP (2020/288). Syed Mohammed Basheeruddin Asdaq wishes to express his gratitude to AlMaarefa University in Riyadh, Saudi Arabia, for providing support (TUMA-2021-1) for this study.