Microbial diversity during processing of Thai traditional fermented shrimp paste, determined by next generation sequencing
Introduction
Shrimp paste, known as ‘Ka-pi’ in Thailand is one of the traditional fermented food ingredients used in Chinese and Southeast Asian cuisines, especially in Thailand for more than 100 years. It gives a unique and rich flavor to many traditional Thai dishes. Shrimp paste is produced from small planktonic shrimps (Acetes spp.) fermented with salt for 1–2 months. The fermentation process involves proteolysis from proteolytic enzymes from fish, and bacteria, which consequently releases amino acids and peptides, that contributes sensory characteristics of the fermented product (Pongsetkul, Benjakul, Vongkamjan, Sumpavapol, & Osako, 2017). It is well established that many amino acids and small peptides act as taste enhancers giving umami, sweet, salty taste, and/or kokumi in foods (Toldrá, Hui, Astiasaran, & Sebranek, 2014; Zhao, Schieber, & Gänzle, 2016). Therefore, investigating the microbial diversity in shrimp paste during processing is important for understanding the microorganisms involving in the product fermentation to obtain a better flavor quality of shrimp paste.
From the previous studies, Tetragenococcus halophillus, Tetragenococcus muriaticus, Tetragenococcus sp., Virgibacillus sp., Lentibacillus sp., and Salinicoccus sp. were found in shrimp paste (Kobayashi et al., 2003; Pakdeeto et al., 2007). However, these studies reported the microorganisms isolated by using selective culture media such as MRS agar, JCM media, which did not represent the whole microorganisms in the food of interest. To explore microbial species diversity, next generation sequencing (NGS) technique has been used as a powerful tool to study microbial community in fermented foods such as Korean chili paste (Nam, Park, & Lim, 2012), sourdough (Ercolini et al., 2013), kimchi (Jeong, Jung, Lee, Jin, & Jeon, 2013). Deep sequencing and the capacity to sequence multiple samples make metagenomic sequencing technology is an important technique in analysis of food microbiota (Ercolini et al., 2013; Jeong et al., 2013; Nam et al., 2012; Ohshima et al., 2019). Although, many studies on the applications of NGS approach in many fermented foods were widely investigated. To date, the information on Thai fermented shrimp paste has not been reported. The attempt to study on microorganisms contributed to flavor quality of shrimp paste was investigated by Qi, Dai, Wang, Sun, and Zheng (2018). The study observed the difference in microbiota profile during two-month shrimp paste fermentation. However, the information of microbiota profile in raw materials and in-processed products, and the relation of microbiota profile and its contribution on flavor have not been reported. Our study aimed to investigate dynamics of microbial diversity from raw materials, samples during preparation (before fermentation) and the probability of taste-enhancing di- or tripeptide production from those bacteria in shrimp paste. Furthermore, this study showed the difference of microbial community structure in shrimp pastes collected from various production regions in Thailand.
Section snippets
Sampling of shrimp paste samples
The shrimp paste samples included raw materials, in-process and finished product samples. The raw materials and in-process samples were collected from a production site located in Samutsakorn Province of Thailand. The raw materials were fresh planktonic shrimp and salt. The in-process samples were collected from each step of production process (Fig. 1). Fresh planktonic shrimp was mixed with salt in ratio 9:1 (%w/w) and rested at room temperature for two to 3 h (P1) and 12-h overnight (P2).
Physico-chemical analysis
At the beginning of the process (Table 1), Aw value of the mixture of shrimp and salt (P1) was 0.85 and gradually decrease during the production to 0.72 in P4, the sample before fermentation. Aw value did not change during 1-month (0.72 in P5) and 2-month fermentation (0.71 in P6) (https://link.springer.com/content/pdf/10.1007%2Fs13197-017-2804-4.pdf Pongsetkul et al., 2017). No trend of pH change during preparation and fermentation was observed. pH values of 1-month and 2-month fermented
Conclusion
The results of the present study are encouraging for the use of NGS to investigate the microbiota of shrimp paste since in raw materials, during preparation and fermentation and comparison of the commercial shrimp paste with different various regions in Thailand. The NGS data of shrimp paste showed Alkalibacterium spp., Macrococcus caseolyticus and Jeotgalicoccus psychrophilus which were not reported in cultured dependent method earlier and detected as the first time in fermented shrimp paste.
CRediT authorship contribution statement
Apiniharn Phewpan: Investigation, Writing - original draft, Data curation, Formal analysis, Software. Preecha Phuwaprisirisan: Resources, Methodology, Data curation. Hajime Takahashi: Methodology. Chihiro Ohshima: Methodology. Kannapon Lopetcharat: Software, Data curation. Punnida Techaruvichit: Writing - original draft. Suwimon Keeratipibul: Supervision, Conceptualization, Writing - review & editing.
Acknowledgements
Authors are grateful to Dr. Rabuesak Khumthong from Research and Development Center of Betagro Group for protease genes and alpha diversity analyses and Dr. Chirapiphat Phraephaisarn from Research and Development Center of Betagro Group for data sorting. This work was financially supported by Chulalongkorn University's Ratchadapisek Sompot Fund [CF5803230] and The Thailand Research Fund (Research and Researcher for Industry (RRi))[No. PHD60I0018].
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