Effects of starter culture and sweetener on biochemical compounds and microbial diversity of kombucha tea

Kombucha tea has been claimed to have several health benefits. Many factors influence the properties of kombucha tea produced. This study focused on the effects of starter cultures (kombucha liquid broth (KLB) and cellulosic pellicle (KCP)) and sweetener (white sugar (S), honey (H) and jaggery (J)) used in the production of kombucha tea. The results showed that all kombucha teas prepared using KLB had a lower pH and a higher concentration of acetic acid during fermentation. The ethanol content for samples prepared using KLB increased (0.7 ± 0.26 mg/L to 1.73 ± 0.58 mg/L) during the fermentation period, compared to KCP which was the maximum after 72 h fermentation, and continued to decrease (2.97 ± 1.24 mg/L to 0.90 ± 0.44 mg/L). Although not too much differences in pH and ethanol content were observed when different sweetener sources were used, they did have significant differences in antioxidant properties and antimicrobial activity. Samples prepared using jaggery had the lowest antioxidant activity while kombucha tea prepared using KLB and white sugar (KLB-S) had the highest antioxidant and antibacterial activity and was mostly colonized by Acetobacteracea and Aspergillus fumigatus. Fermentation significantly increases the number of active compounds present in KLB-S from 11 to 25 compounds. New compounds such as docosanedioic acid, muramic acid and thiolactomycin were formed. Thiolactomycin, a natural antibiotic is suggested to contribute to the high antimicrobial activity of KLB-S. In conclusion, KLB and white sugar are better suited in preparing kombucha tea as more benefits and consistent results were observed

Collagen-derived cryptides : machine-learning prediction and molecular dynamic interaction against Klebsiella pneumoniae biofilm synthesis precursor

Collagen-derived cryptic peptides (cryptides) are biologically active peptides derived from the proteolytic digestion of collagen protein. These cryptides possess a multitude of activities, including antihypertensive, antiproliferative, and antibacterial. The latter, however, has not been extensively studied. The cryptides are mainly obtained from the protein hydrolysate, followed by characterizations to elucidate the function, limiting the number of cryptides investigated within a short period. The recent threat of antimicrobial resistance microorganisms (AMR) to global health requires the rapid development of new therapeutic drugs. The current study aims to predict antimicrobial peptides (AMP) from collagen-derived cryptides, followed by elucidating their potential to inhibit biofilm-related precursors in Klebsiella pneumoniae using in silico approach. Therefore, cryptides derived from collagen amino acid sequences of various types and species were subjected to online machine-learning platforms (i.e., CAMPr3, DBAASP, dPABBs, Hemopred, and ToxinPred). The peptide-protein interaction was elucidated using molecular docking, molecular dynamics, and MM-PBSA analysis against MrkH, a K. pneumoniae’s transcriptional regulator of type 3 fimbriae that promote biofilm formation. As a result, six potential antibiofilm inhibitory cryptides were screened and docked against MrkH. All six peptides bind stronger than the MrkH ligand (c-di-GMP; C2E)

Isolation of cells from a Marine Source Responsible for Hemocyanin Biosynthesis

Molluscan rhogocyte cells are known to synthesise the largest respiratory proteins in nature known as hemocyanin. However, attempt to investigate the cells in vitro to understand the hemocyanin biosynthesis is difficult due to the lack of current knowledge on establishing a stable cell culture. Therefore, the aims of this study were to isolate rhogocyte cells from Haliotis laevigata and assessed their characteristics, distribution, capability to grow and synthesise hemocyanin in vitro. Using flow cytometry analysis and simultaneous staining of immunofluorescence and in situ hybridisation strategy, two distinct populations of rhogocyte cells synthesizing type 1 hemocyanin was determined in the mantle tissue. Subsequently, a primary culture of heterogeneous cells was established with different parameters involving basal media, primary growth supplements, secondary growth supplements, growth temperature and seeding density. Cells cultured in MEM result in the highest cell yield compared to other basal media. It is suggested that the presence of vitamin B6 in aldehyde form (pyridoxal) instead of alcohol (pyridoxine) is responsible for promoting the cellular activity. Furthermore, addition of lipimax at 17°C resulted in higher cell-fold increase compared to FBS, knockout serum and hemolymph. Addition of secondary supplements such as amino acids cocktail, lipid concentrate, insulin-transferrin-selenium and vitamin concentrate, however, had no significant impact on the cell growth. On the other hand, evaluation of hemocyanin content using ELISA revealed significant increase of hemocyanin in the media when cells were cultured with lipimax, FBS and knockout serum. However, the hemocyanin content was at the highest concentration only after an hour of culture before decreasing significantly and stabilised around 0.14-0.16 µg/ml in the media. These results suggest that hemocyanin biosynthesis may have an inverse correlation with the cell’s growth

Non-enzymatic antioxidant from apple snail (Pomacea maculata) extract

Pomacea sp. is a freshwater gastropod that is capable of withstanding oxidative stress during extreme environmental changes. The snail enzymatic oxidative responses have already been elucidated through biochemical, transcriptomics, and proteomics analysis. However, their non-enzymatic oxidative responses have yet to be elucidated. Therefore, this study aims to characterize the antioxidant activity and identify the non-enzymatic antioxidant compounds from Pomacea maculata. To address the aims, a polar and non-polar extraction of snail-whole body extract was conducted using methanol and chloroform, respectively. The antioxidant activity of both extracts was elucidated by Folin Ciocalteau (FC), DPPH, and reducing power assay. LC-MS/MS was then used to profile both extracts. The results demonstrated that the crude methanol extract (CME) contains a higher antioxidant capacity (FC=43.22 ± 3.02 mg GAE/ g extract, DPPH IC50=0.073 mg/mL, and reducing the power of methanol and chloroform extract are 0.361 ± 0.07 and 0.051 ± 0.003 respectively). Profiling of the snail metabolites by LC-MS/MS from both extracts resulted in the identification of uric acid and phenolic compounds . The former was detected at the highest intensity in CME followed by crude chloroform extract (CCE). The phenolic compounds, however, were hypothetically identified as plant metabolites. Therefore, the study suggested that antioxidant activity exhibited by P. maculata extracts were due to non-enzymatic compounds such as uric acid and phenolic compounds originated from the animal’s metabolic activity and plants, respectively

Characterization of putative pathogenic Shewanella algae isolated from ballast water

Background and Aim: Shewanella algae is ubiquitous in marine-associated environments and has been increasingly recognized as a significant human pathogen that can cause serious infections mainly associated with exposure to seawater and ingestion of raw seafood. This study aimed to isolate and characterize S. algae from ballast water of ships berthed at Port Klang, Malaysia. Materials and Methods: Ballast water was sampled from nine ships docked at Port Klang, Malaysia. The isolates were identified and characterized based on biochemical and enzymatic properties, 16S rRNA and gyrB sequencing, biofilm formation capability, and antibiotic susceptibility. Results: A total of four S. algae isolates were isolated from four ballast water samples tentatively name Sa-BW1, Sa-BW2, Sa-BW7, and Sa-BW8. All isolates showed positive reaction for cytochrome oxidase, catalase, high tolerance to NaCl (6% and 8%), ability to grow at 42°C, and on Salmonella-Shigella agar. The strains also exhibited β-hemolytic activity on sheep blood and human blood agar, positive reaction for lipase, protease, DNase and gelatinase, strong biofilm adherence capabilities and multiple antibiotic resistances against ampicillin, carbenicillin, cephalothin, colistin, novobiocin, oxacillin, penicillin, rifampicin, and tobramycin which suggested their potential pathogenicity. Conclusion: This study demonstrated the occurrence of putative pathogen S. algae in ballast water of ships docked at Malaysian port