Untargeted Metabolomics on Skin Mucus Extract of Channa argus against Staphylococcus aureus: Antimicrobial Activity and Mechanism.
ABSTRACT: Microbial contamination is one of the most common food safety issues that lead to food spoilage and foodborne illness, which readily affects the health of the masses as well as gives rise to huge economic losses. In this study, Channa argus was used as a source of antimicrobial agent that was then analyzed by untargeted metabolomics for its antibacterial mechanism against Staphylococcus aureus. The results indicated that the skin mucus extract of C. argus had great inhibitory action on the growth of S. aureus, and the morphology of S. aureus cells treated with the skin mucus extract exhibited severe morphological damage under scanning electron microscopy. In addition, metabolomics analysis revealed that skin mucus extract stress inhibited the primary metabolic pathways of S. aureus by inducing the tricarboxylic acid cycle and amino acid biosynthesis, which further affected the normal physiological functions of biofilms. In conclusion, the antimicrobial effect of the skin mucus extract is achieved by disrupting cell membrane functions to induce an intracellular metabolic imbalance. Hence, these results conduce to amass novel insights into the antimicrobial mechanism of the skin mucus extract of C. argus against S. aureus.
Project description:Antimicrobial peptides (AMPs) constitute a broad range of bioactive compounds in diverse organisms, including fish. They are effector molecules for the innate immune response, against pathogens, tissue damage and infections. Still, AMPs from African Catfish, <i>Clarias gariepinus</i>, skin mucus are largely unexplored despite their possible therapeutic role in combating antimicrobial resistance. In this study, African Catfish Antimicrobial peptides (ACAPs) were identified from the skin mucus of African Catfish, <i>C. gariepinus</i>. Native peptides were extracted from fish mucus scrapings in 10% acetic acid (v/v) and ultra-filtered using 5 kDa molecular weight cut-off membrane. The extract was purified using C<sub>18</sub> Solid-Phase Extraction. The antibacterial activity was determined using the Agar Well Diffusion method and broth-dilution method utilizing <i>Staphylococcus aureus</i> (ATCC 25923) and <i>Escherichia coli</i> (ATCC 25922). Thereafter, Sephadex G-25 gel filtration was further utilized in bio-guided isolation of the most active fractions prior to peptide identification using Orbitrap Fusion Lumos Tribrid Mass Spectrometry. The skin mucus extracted from African Catfish from all the three major lakes of Uganda exhibited antimicrobial activity on <i>E. coli</i> and <i>S. aureus</i>. Lake Albert's <i>C. gariepinus</i> demonstrated the best activity with the lowest MIC of 2.84 and 0.71 μg/ml on <i>S. aureus</i> and <i>E. coli</i>, respectively. Sephadex G-25 peak I mass spectrometry analysis (Data are available <i>via</i> ProteomeXchange with identifier PXD029193) alongside <i>in silico</i> analysis revealed seven short peptides (11-16 amino acid residues) of high antimicrobial scores (0.561-0.905 units). In addition, these peptides had a low molecular weight (1005.57-1622.05 Da) and had percentage hydrophobicity above 54%. Up to four of these AMPs demonstrated α-helix structure conformation, rendering them amphipathic. The findings of this study indicate that novel AMPs can be sourced from the skin mucus of <i>C. gariepinus</i>. Such AMPs are potential alternatives to the traditional antibiotics and can be of great application to food and pharmaceutical industries; however, further studies are still needed to establish their drug-likeness and safety profiles.
Project description:Antimicrobial peptides (AMPs) constitute a broad range of bioactive compounds in diverse organisms, including fish. They are effector molecules for the innate immune response, against pathogens, tissue damage and infections. Still, AMPs from African Catfish, Clarias gariepinus skin mucus are largely unexplored despite their possible therapeutic role in combating antimicrobial resistance. In this study, African Catfish Antimicrobial peptides (ACAPs) were identified from the skin mucus of African Catfish, C. gariepinus. Native peptides were extracted from fish mucus scrapings in 10% acetic acid (v/v) and ultra-filtered using 5kDa molecular cut-off membrane. The extract was purified using C18 Solid Phase Extraction. The antibacterial activity was determined using the Agar Well Diffusion method and broth-dilution method utilizing Staphylococcus aureus (ATCC 25923) and Escherichia coli (ATCC 25922). Thereafter, Sephadex G-25 gel filtration was further utilized in bio-guided isolation of the most active fractions prior to peptide identification using Orbitrap Fusion Lumos Tribrid Mass Spectrometry. The skin mucus extracted from African Catfish from all the three major lakes of Uganda exhibited antimicrobial activity on E. coli and S. aureus. Lake Albert’s C. gariepinus demonstrated the best activity with the lowest MIC of 2.84 µg/mL and 0.71 µg/mL on S. aureus and E. coli respectively. Sephadex G-25 peak I mass spectrometry analysis alongside in silico analysis revealed seven short peptides (11-15 amino acid residues) of high antimicrobial scores (0.561-0.905 units). In addition, these peptides had a low molecular weight (1005.57-1622.05 Da), and had percentage hydrophobicity above 54%. Up to four of these antimicrobial peptides demonstrated α-helix structure conformation, rendering them amphipathic. The ﬁndings of this study indicate that novel antimicrobial peptides can be sourced from the skin mucus of C. gariepinus. Such antimicrobial peptides are potential alternatives to the traditional antibiotics and can be of great application to food and pharmaceutical industries; however, further studies are still needed to establish their drug-likeness and safety profiles.
Project description:The Northern snakehead (Channa argus), a member of the Channidae family of the Perciformes, is an economically important freshwater fish native to East Asia. In North America, it has become notorious as an intentionally released invasive species. Its ability to breathe air with gills and migrate short distances over land makes it a good model for bimodal breath research. Therefore, recent research has focused on the identification of relevant candidate genes. Here, we performed whole genome sequencing of C. argus to construct its draft genome, aiming to offer useful information for further functional studies and identification of target genes related to its unusual facultative air breathing. Findings: We assembled the C. argus genome with a total of 140.3 Gb of raw reads, which were sequenced using the Illumina HiSeq2000 platform. The final draft genome assembly was approximately 615.3 Mb, with a contig N50 of 81.4 kb and scaffold N50 of 4.5 Mb. The identified repeat sequences account for 18.9% of the whole genome. The 19?877 protein-coding genes were predicted from the genome assembly, with an average of 10.5 exons per gene. Conclusion: We generated a high-quality draft genome of C. argus, which will provide a valuable genetic resource for further biomedical investigations of this economically important teleost fish.
Project description:Background:The introduction of northern snakehead (Channa argus; Anabantiformes: Channidae) and their subsequent expansion is one of many problematic biological invasions in the United States. This harmful aquatic invasive species has become established in various parts of the eastern United States, including the Potomac River basin, and has recently become established in the Mississippi River basin in Arkansas. Effective management of C. argus and prevention of its further spread depends upon knowledge of current population structure in the United States. Methods:Novel methods for invasive species using whole genomic scans provide unprecedented levels of data, which are able to investigate fine scale differences between and within populations of organisms. In this study, we utilize 2b-RAD genomic sequencing to recover 1,007 single-nucleotide polymorphism (SNP) loci from genomic DNA extracted from 165 C. argus individuals: 147 individuals sampled along the East Coast of the United States and 18 individuals sampled throughout Arkansas. Results:Analysis of those SNP loci help to resolve existing population structure and recover five genetically distinct populations of C. argus in the United States. Additionally, information from the SNP loci enable us to begin to calculate the long-term effective population size ranges of this harmful aquatic invasive species. We estimate long-term Ne to be 1,840,000-18,400,000 for the Upper Hudson River basin, 4,537,500-45,375,000 for the Lower Hudson River basin, 3,422,500-34,225,000 for the Potomac River basin, 2,715,000-7,150,000 for Philadelphia, and 2,580,000-25,800,000 for Arkansas populations. Discussion and Conclusions:This work provides evidence for the presence of more genetic populations than previously estimated and estimates population size, showing the invasive potential of C. argus in the United States. The valuable information gained from this study will allow effective management of the existing populations to avoid expansion and possibly enable future eradication efforts.
Project description:<i>Staphylococcus aureus</i> is an important etiological agent that causes skin infections, and has the propensity to form biofilms, leading to significant mortality and morbidity in patients with wounds. Mucus secretion from the Giant African snail <i>Achatina fulica</i> is a potential source of biologically active substances that might be an important source for new drugs to treat resistant and biofilm-forming bacteria such as <i>S. aureus</i>. This study evaluated the effect of semi-purified fractions from the mucus secretion of <i>A. fulica</i> on the growth, biofilm formation and virulence factors of <i>S. aureus</i>. Two fractions: FMA30 (Mw >30 kDa) and FME30 (Mw 30-10 kDa) exhibited antimicrobial activity against <i>S. aureus</i> with a MIC<sub>50</sub> of 25 and 125 µg/mL, respectively. An inhibition of biofilm formation higher than 80% was observed at 9 µg/mL with FMA30 and 120 µg/mL with FME30. Furthermore, inhibition of hemolytic and protease activity was determined using a concentration of MIC<sub>20</sub>, and FME30 showed a strong inhibitory effect in the formation of clots. We report for the first time the effect of semi-purified fractions of mucus secretion of <i>A. fulica</i> on biofilm formation and activity of virulence factors such as α-hemolysin, coagulase and proteases produced by <i>S. aureus</i> strains.
Project description:Bacopa monnieri Linn. (Plantaginaceae), a well-known medicinal plant, is widely used in traditional medicine system. It has long been used in gastrointestinal discomfort, skin diseases, epilepsy and analgesia. This research investigated the in vitro antimicrobial activity of Bacopa monnieri leaf extract against Staphylococcus aureus and the interaction of possible compounds involved in this antimicrobial action.Non-edible plant parts were extracted with ethanol and evaporated in vacuo to obtain the crude extract. A zone of inhibition studies and the minimum inhibitory concentration (MIC) of plant extracts were evaluated against clinical isolates by the microbroth dilution method. Docking study was performed to analyze and identify the interactions of possible antimicrobial compounds of Bacopa monnieri in the active site of penicillin binding protein and DNA gyrase through GOLD 4.12 software.A zone of inhibition studies showed significant (p?<?0.05) inhibition capacity of different concentrations of Bacopa monnieri's extract against Staphylococcus aureus. The extract also displayed very remarkable minimum inhibitory concentrations (?16 ?g/ml) which was significant compared to that (?75 ?g/ml) of the reference antibiotic against the experimental strain Staphylococcus aureus. Docking studies recommended that luteolin, an existing phytochemical of Bacopa monnieri, has the highest fitness score and more specificity towards the DNA gyrase binding site rather than penicillin binding protein.Bacopa monnieri extract and its compound luteolin have a significant antimicrobial activity against Staphylococcus aureus. Molecular binding interaction of an in silico data demonstrated that luteolin has more specificity towards the DNA gyrase binding site and could be a potent antimicrobial compound.
Project description:The high antimicrobial ability and low toxicity of zinc-aminoclay (ZnAC) are claimed in our previous reports. In this study, we formulate a novel hand gel based on ZnAC and Opuntia humifusa (O. humifusa) extract, which is a high moisturizing agent. The antimicrobial activity, cytotoxicity, moisturizing effect, and clinical skin irritation of the hand gel are evaluated. The hand gel with 0.5 wt.% ZnAC and 1.0 v/v% O. humifusa extract can kill more than 99% Escherichia coli (gram-negative bacteria) and Staphylococcus aureus (gram-positive bacteria) after 24 h. Toxicity evaluation shows that, the hand gel does not affect the viability of mammalian HaCaT cells. Additionally, skin moisture is increased by applying the hand gel while its viscosity is at the standard level of commercial products. The hand gel has a skin irritation index of 0.0 and is classified as a non-irritating product. We successfully formulated hand gel from ZnAC, glucomannan, glycerol, and O. humifusa extract. Owing to the high antimicrobial activity and skin protection of hand gels, they are suitable to be used as hand sanitizers in restaurants, hospitals, and homes effectively.
Project description:Northern snakehead, <i>Channa argus</i>, is a commercially important food fish species in China. In the present study, the complete mitochondrial genome of <i>C.argus</i> from the Baima Hu Lake was characterized. It is 16,558 bp in length, consist of 22 tRNA genes, 13 PCD genes, 2 rRNA genes, and 1 D-loop region. The overall base composition of the <i>C. argus</i> mitogenome is 27.26% A, 24.21% T, 31.58% C and 16.95% G, exhibits a similar AT bias (51.47%) feature to other vertebrate mitogenomes. The phylogenetic analysis showed that <i>C. argus</i> clustered in genus <i>Channa</i>. The present resultes provide useful information to population genetics and conservation biology studies of <i>Channa</i> fishes.
Project description:A critical challenge in the study of botanical natural products is the difficulty of identifying multiple compounds that may contribute additively, synergistically, or antagonistically to biological activity. Herein, it is demonstrated how combining untargeted metabolomics with synergy-directed fractionation can be effective toward accomplishing this goal. To demonstrate this approach, an extract of the botanical goldenseal ( Hydrastis canadensis) was fractionated and tested for its ability to enhance the antimicrobial activity of the alkaloid berberine (4) against the pathogenic bacterium Staphylococcus aureus. Bioassay data were combined with untargeted mass spectrometry-based metabolomics data sets (biochemometrics) to produce selectivity ratio (SR) plots, which visually show which extract components are most strongly associated with the biological effect. Using this approach, the new flavonoid 3,3'-dihydroxy-5,7,4'-trimethoxy-6,8- C-dimethylflavone (29) was identified, as were several flavonoids known to be active. When tested in combination with 4, 29 lowered the IC50 of 4 from 132.2 ± 1.1 ?M to 91.5 ± 1.1 ?M. In isolation, 29 did not demonstrate antimicrobial activity. The current study highlights the importance of fractionation when utilizing metabolomics for identifying bioactive components from botanical extracts and demonstrates the power of SR plots to help merge and interpret complex biological and chemical data sets.