ISOLATION AND SCREENING OF PROTEOLYTIC BACTERIA FROM SIDOARJO SHRIMP PASTE AS PROTEASE SOURCE TO EXTRACT THE COLLAGEN FROM MILKFISH SCALES (CHANOS CHANOS)

This research aimed to isolate protease-producing bacteria from Sidoarjo shrimp paste for extracting collagen from milkfish scales. This study began with isolation, followed by screening and purification of protease-producing bacterial isolates. Further confirmation of the isolates’ proteolytic indices and the crude protease production, the enzymes’ efficacy in extracting collagen from milkfish scales were tested, followed by pathogenicity and identification using 16S rRNA molecular technique. The study has successfully isolated 15 proteolytic bacterial isolates using skimmed milk agar, but only isolates of TR-10, TR-4.1.1, and TR-15.1 exhibited prospective proteolytic activity based on their corresponding proteolytic indices of 2.96 ± 0.06, 3.10 ± 0.10, and 3.71 ± 0.48. Although the proteolytic activity of isolates TR-10 (0.22 ± 0.05 U/mL) and TR-15.1 (1.07 ± 0.14 U/mL) was high in a salt medium using peptone as the nitrogen source, only the former showed satisfactory activity to extract soluble collagen from milkfish scales. Based on the 16SrRNA, the TR-10 isolate was identified as Bacillus megaterium. The non-pathogenicity of the TR-10 bacterium signified its promising role as a protease source for the halal collagen extraction from milkfish scales.

Isolation of Aeromonas sobria JC18 from milkfish (Chanos chanos) intestine with proteolytic and cellulolytic activities for fish probiotic

Attention on the probiotic application for the improvement of fish resident intestinal microbiota has risen in the last decades. It is demonstrated that probiotics may function not only by direct inhibition of pathogenic bacteria or manipulation of enzymatic digestion of feed but also by modulation of immunity in fish. In the present study, we aimed to isolate the autochthonous microbiota of marine fish intestines as probiotic candidates based on cellulolytic and proteolytic activity. Bacteria were isolated and purified on a cellulose agar with 24h of incubation. A total of 18 bacterial strains were purified and stored in -80°C. Phenotypic screening based on the antibiotic resistance, antagonistic activity against pathogenic bacteria, resistance to an acidic environment, and ability of colonization in fish intestine found a selected strain, namely JC18. Infection test, molecular and phenotypic characterizations revealed that the JC18 isolate was a non-pathogenic Aeromonas sobria. It is hence revealed that the milkfish intestine is a potent source of proteolytic bacteria for fish probiotic screening.

Substrate preferences, phylogenetic and biochemical properties of proteolytic bacteria present in the digestive tract of Nile tilapia

Vertebrate intestine appears an excellent source of proteolytic bacteria for industrial and probiotic use. We therefore aimed obtaining the gut-associated proteolytic species of Nile tilapia. We’ve isolated twenty six bacterial strains from its intestinal tract, seven of which showed exoprotease activity with the formation of clear halos on skim milk. Their depolymerization ability was further assessed on three distinct proteins including casein, gelatin and albumin. All the isolates could successfully hydrolyze the three substrates indicating relatively broad specificity of their secreted proteases. Molecular taxonomy and phylogeny of the proteolytic isolates were determined based on their 16S rRNA gene barcoding which suggested that the seven strains belong to three phyla viz. Firmicutes, Proteobacteria and Actinobacteria, distributed across the genera Priestia, Citrobacter, Pseudomonas, Stenotrophomonas, Burkholderia, Providencia and Micrococcus. The isolates were further characterized by a comprehensive study of their morphological, cultural, cellular and biochemical properties which were consistent with the phylogenetic annotations. To reveal their proteolytic capacity alongside substrate preferences, enzyme-production was determined by the diffusion assay. The Pseudomonas, Stenotrophomonas and Micrococcus isolates appeared most promising with maximum protease production on casein, gelatin and albumin media respectively. Our findings present valuable insights into the phylogenetic and biochemical properties of gut-associated proteolytic strains of Nile tilapia.

Substrate preferences, phylogenetic and biochemical properties of proteolytic bacteria present in the digestive tract of Nile tilapia

Vertebrate intestine appears an excellent source of proteolytic bacteria for industrial and probiotic use. We therefore aimed obtaining the gut-associated proteolytic species of Nile tilapia. We have isolated twenty six bacterial strains from its intestinal tract, seven of which showed exoprotease activity with the formation of clear halos on skim milk. Their depolymerization ability was further assessed on three distinct proteins including casein, gelatin and albumin. All the isolates could successfully hydrolyze the three substrates indicating relatively broad specificity of their secreted proteases. Molecular taxonomy and phylogeny of the proteolytic isolates were determined based on their 16S rRNA gene barcoding which suggested that the seven strains belong to three phyla viz. Firmicutes, Proteobacteria and Actinobacteria, distributed across the genera Priestia, Citrobacter, Pseudomonas, Stenotrophomonas, Burkholderia, Providencia and Micrococcus. The isolates were further characterized by a comprehensive study of their morphological, cultural, cellular and biochemical properties which were consistent with the phylogenetic annotations. To reveal their proteolytic capacity alongside substrate preferences, enzyme-production was determined by the diffusion assay. The Pseudomonas, Stenotrophomonas and Micrococcus isolates appeared most promising with maximum protease production on casein, gelatin and albumin media respectively. Our findings present valuable insights into the phylogenetic and biochemical properties of gut-associated proteolytic strains of Nile tilapia.

Yellow fever disease severity is driven by an acute cytokine storm modulated by an interplay between the human gut microbiome and the metabolome

Infection with Yellow Fever (YF) can lead to multiple outcomes ranging from death from total organ failure to clearance of viremia and survival. The mechanisms underlying these differences in clinical outcomes have yet to be defined. We had access to a cohort of YF infected subjects that showed these range of outcomes. An unbiased integrated OMICs approach was used to identify pathways and effector molecules that drive the severe disease and death as compared to resolution of infection. We used the MELD and SIC score as objective markers of disease severity. We show that a specific signature of upregulated innate pro-inflammatory cytokines significantly demarcates subjects with severe disease leading to death from subjects who clear virus. Pathogen sequencing showed heightened levels of proteolytic bacteria at the i.e Actinobacteria and these were correlated to lower levels of tryptophan and tyrosine amino acids measured by untargeted metabolomics. These two features were significantly associated to MELD scores synonymous of milder disease. Propionate a bacterial metabolite that triggers Treg differentiation that can as well limit the hyperimmune activation associated to severe outcome was also associated to improved outcome. Our results suggest a model whereby proteolytic bacteria limit the availability of the aromatic amino acid pool available for cytokine production thereby preventing the induction of the cytokine storm that is associated to severe disease and death.

Komposisi Bakteri Saluran Pencernaan Belut Rawa (Synbranchus Bengalensis)

Eels are a group of snake- shaped fish belonging to the Synbranchidae. Bacteria in groups of fish such as eels can be found on the body surface and digestive tract. Some bacteria are pathogenic, while a number of other bacteria are beneficial, but researches in this topic has not been optimal. This study examined the bacterial composition of the digestive tract of swamp eel (Synbranchus bengalensis) using descriptive exploratory methods. The results showed that the composition of bacteria found in the digestive tract of swamp eels was diverse but dominated by gram-positive bacteria; BR2, BR3, BR4, BR5. In addition, some of these bacteria belong to the group of proteolytic bacteria; BR2 and BR4, lipolytic bacteria; BR2, and amylolytic bacteria; BR1 which hydrolyzes digestive enzymes can act as probiotics in the digestive tract of swamp eels (Synbranchus bengalensis).Keywords: swamp eels, microflora digestive, Synbranchus bengalensis