Determination of yeast diversity in fermented Sumbawa mare’s milk using internal transcribed spacers (ITS) fragment analysis

Aims@#Molecular identification of yeast has been conducted on various fermentation products. However, the identification of yeast in fermented Sumbawa mare’s milk based on the genotyping method has not been carried out. This study was aimed to determine the diversity profile of yeasts in fermented Sumbawa mare’s milk using phenetic characters and PCR-RFLP analysis technique based on the ITS region.@*Methodology and results@#Yeast isolates were phenotypically characterized and visualized in a dendrogram using CLAD97 software. Then, the yeast DNA was extracted using heat treatment and amplified using ITS1 and ITS4 primers. The amplicons were analyzed by RFLP using HindIII and HaeIII enzymes. The phylogenetic tree was constructed using MEGA 7.0. Based on the result of grouping by phenetic analysis and PCR-RFLP, the 12 isolates were divided into four groups with different members. The results of the phenetic analysis were divided into group I (all isolates of Dompu), group II (isolate B3, B4, S3), group III (isolate B5) and group IV (isolate S1). The types of yeast that were identified molecularly and represented each group of PCR-RFLP results included in group I were Kluyveromyces marxianus D1A and K. marxianus D1B, group II: K. marxianus D7, group III: Kazachstania humilis D4, while milk from Bima and Sumbawa has one yeast species as a member of group IV, namely Pichia kudriavzevii B3. Kluyveromyces marxianus was the yeast frequently found in Sumbawa fermented mare’s milk.@*Conclusion, significance and impact of study@#Various yeast species as a consortium of the milk samples can contribute to the increasing quality of fermented Sumbawa mare’s milk.

Selection of potential lactic acid bacteria from fish intestine of mackerel (Rastrelliger kanagurta) from Lembata Regency, East Nusa Tenggara, Indonesia

Aims@#The aim of this study was to screen lactic acid bacteria (LAB) isolates with probiotic properties for tilapia (Oreochromis niloticus) from the intestines of mackerel and to analyze its ability to produce hydrolase enzymes (amylase, protease, cellulase and lipase).@*Methodology and results@#Ten samples of mackerel were collected from the Fish Auction Place, Lewoleba, Nubatukan District, Lembata Regency, East Nusa Tenggara Province, Indonesia. The process for screening were antimicrobial activity, pH tolerance test of 1, 3 and 5, bile salt concentrations of 2.5%, 5% and 7.5%, and water salinity (0.5%, 3.5% and 6.5%). The autoaggregation and semi-qualitative test of hydrolase enzymes were also carried out. In total, 21 LAB isolates were derived from mackerel intestines. Isolates that have the stronger ability to inhibit the pathogen Aeromonas hydrophila ATCC 7966 are KBP 3.3, KBP 3.3.1 and KBP 6.3, while KBP 1.3, KBP 1.1.1 and KBP 3.3.1 were able to inhibit Streptococcus agalactiae ATCC 13813. The LAB isolates that survived at the tested pH were KBP 3.3.1, KBP 3.3, KBP 1.1.1 and KBP 1.3. Results also suggest that the four isolates were tolerant to bile salt concentrations and water salinity with good survival rates (>94.7%) and had proteolytic, lipolytic and cellulolytic activities. The isolates KBP 3.3, KBP 3.3.1 and KBP 1.1.1 were identified as Weissella confusa 6250, while the isolate KBP 1.3 was identified as W. confusa C5-7.@*Conclusion, significance and impact of study@#The characteristics of four selected isolates indicate their potential as a probiotic. These potential probiotic isolates can be applied directly to fish by utilizing microencapsulation technology or adding to the feed.

Selection of potential lactic acid bacteria from fermented Sumbawa mare’s milk as starter cultures

Aims@#The aim of this study was to screen lactic acid bacteria (LAB) isolates from fermented Sumbawa mare‘s milk that meet the requirements as starter cultures, and to evaluate the effect of the selected starter culture in improving the organoleptic quality of mare‘s milk fermentation. @*Methodology and results@#The LAB isolates (13 isolates) derived from naturally fermented Sumbawa mare‘s milk were firstly screened for acidification activity. Afterwards, the selected isolates were evaluated for the starter culture criteria such as technological properties (proteolytic test, lipolytic test, and exopolysaccharide production), food safety test (hemolytic test and antibiotic sensitivity test), antimicrobial activity test. The selected culture (SC) together with yogurt starter cultures (YC) and combination between the selected isolate and a mixture of both (MC) were used to ferment fresh mare’s milk. Six LAB isolates (DB7, BC10, DC4, BC9, DC10, and BC7) were obtained from the acidification screening. Isolate BC10 was the most potential isolate as starter culture due to its ability in terms of acidification and proteolytic activity, lack of lipolytic activity, no indication of pathogenic potency, as well as able to inhibit the growth of Escherichia coli ATCC 25922. However, this isolate was resistant to antibiotics kanamycin, trimethoprim, and cinoxacin. The isolate BC10 presented 99.99% sequence similarity with respect to Lactobacillus plantarum.@*Conclusion, significance and impact of study@#The selected starter culture (isolate BC10) was able to improve the organoleptic quality of fermented mare‘s milk especially aroma compared to the other starter cultures. Therefore, L. plantarum BC10 is a potential isolate to be used as starter culture for mare’s milk fermentation.