preprints.org > biology and life sciences > biology and biotechnology > doi: 10.20944/preprints202310.0945.v1

Preprint Article Version 1 Preserved in Portico This version is not peer-reviewed

Version 1 : Received: 16 October 2023 / Approved: 16 October 2023 / Online: 16 October 2023 (13:15:21 CEST)

: With the increase in demand for non-dairy starter cultures and probiotic bacteria as carriers, the use of legumes (also called pulses) as an alternative has gained momentum. In this study, we investigated the diversity of bacterial communities in samples of pigeon pea (Cajanus cajan L. Millsp.) soaked in water for 12 h and 24 h. We soaked 500 g of pigeon pea in sterile distilled water at room temperature (± 25 °C) for 12 h and 24 h; 10 mL of the soaking water was then collected to measure the bacterial diversity using a metataxonomic analysis. The V1–V9 regions on the 16S ribosomal RNA gene were amplified using 27F and 1492R primers under specific polymerase chain reaction conditions for the bacterial identification. Genomic DNA (130 ng) was sequenced on a R9.4 flow cell by Oxford Nanopore Technologies using a GridION sequencer. Library preparations were initiated using a Native Barcoding Kit 24 V14 (SQK-NBD114.24). Primary data were acquired using MinKNOW version 22.05.7. A total of 13 bacterial families and 89 genera were identified in the pigeon pea sample soaked for 12 h; 26 families and 90 genera were identified in the pigeon pea sample soaked for 24 h. Among the bacterial families identified, the five predominant families in both samples were Enterobacteriaceae, Erwiniaceae, Yersiniaceae, Pectobacteriaceae, and Lactobacillaceae. According to the relative abundance of the identified bacterial genera, the following nine genera were predominant in both samples: Enterobacter, Klebsiella, Citrobacter, Pantoea, Kosakonia, Pseudoenterobacter, Pluralibacter, Leclercia, and Kluyvera. At a genus level, a slight increase in the abundance of Klebsiella, Kosakonia, and Pluralibacter and a slight decrease in the abundance of Citrobacter were observed after prolonged incubation from 12 h to 24 h. The values of five diversity indices revealed that the sample soaked in water for 24 h had a richer bacterial abundance and diversity than the 12 h sample. Shannon and Simpson values revealed a higher bacterial diversity in the sample collected at 24 h than the sample collected at 12 h. Species observations and abundance-based coverage estimator (ACE) values demonstrated that the sample collected at 24 h harbored a higher bacterial richness than the sample collected at 12 h. These findings indicated that the bacterial diversity in the pigeon pea samples increased with the soaking time. The bacterial communities during the soaking of the pigeon pea samples were dominated by the Enterobacteriaceae family and Enterobacter genus. The presence of bacterial genera such as Lacticaseibacillus, Lentilactobacillus, and Secundilactobacillus was notable because of their importance as starter cultures for fermented plant-based milk products, including pigeon pea beverages for lactose-intolerant individuals or individuals with malnutrition.

pigeon pea; bacterial diversity; metataxonomic analysis

Biology and Life Sciences, Biology and Biotechnology

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