Aims: The antibiofilm activity of endophytic fungus Nigrospora sphaerica CL-OP30 isolated from Swietenia macrophylla King was investigated. Methodology and results: The ability of the fungal endophytic crude extract to impede Streptococcus mutans biofilm formation was preliminarily screened with Congo red agar test. It was proven that S. mutans biofilm formation was hindered on the agar supplemented with the fungal endophytic crude extract. The antibiofilm activity of the fungal endophytic crude extract was evaluated using a microtiter plate method on both initially formed and preformed biofilm. Antibiofilm activity was recorded in a concentration-dependent pattern whereby higher concentrations reduced biofilm formation better than the lower concentrations of extract for both initially formed and preformed biofilm. The architecture of biofilm tested with fungal endophytic crude extract was also observed. Visualization under a light microscope and SEM revealed that the adherence of S. mutans biofilm treated with fungal endophytic crude extract was significantly reduced in both initially formed and preformed biofilm. In addition, observation under SEM showed that the matrices surrounding the bacterial cells were disintegrated and bacterial cells in biofilm completely lost their original shape. The overall data demonstrated that the ethyl acetate N. spaherica CL-OP30 crude extract showed good antibiofilm activity. Conclusion, significance and impact of study The antibiofilm study suggested the potential of N. sphaerica CL-OP30 crude extract against S. mutans biofilm by disrupting the biofilm formation, the disintegration of matrices surrounding the biofilm and responsible for the formation of irregular cell shape. This extract may have a promising potential to be developed as an antibiofilm agent.

Aims: Marine bacteria have been reported to produce potential natural pigment with pharmaceutical properties and their growth can be manipulated in the laboratory to increase pigment production and their antimicrobial activity. Hence, this study aimed to enhance the prodigiosin production in Serratia marcescens IBRL USM84 by improving physical conditions. Methodology and results: The quantification of the pigment produced by S. marcescens IBRL USM84, bacterial cell growth, and its antibacterial activity in the broth medium were determined using a spectrophotometry method. Meanwhile, the antibacterial effect of red pigment on MRSA cells was observed under a scanning electron microscope (SEM). This marine isolate produced the highest yield of prodigiosin (6.95 μg/mL) when cultivated in marine broth with the addition of 0.2% of agar, 25 °C incubation temperature, initial medium pH of 7, 150 rpm of agitation speed for 48 h of cultivation time under light illumination. There was an increment of 151.81% in prodigiosin production after enhancement compared to before the enhancement of cultural conditions. SEM observations revealed that severe damage to the cell’s morphologies was exposed to red pigment as indicated by the formation of small dents, which led to completely collapse and eventually, cell death. Conclusion, significance and impact of study A positive correlation between pigment production and antibacterial activity was observed in the present study. The results supported the fact that marine bacteria are a reservoir of various pigments with antimicrobial properties. Also, the pigment production by S. marcescens and its antibacterial activity were significantly influenced by physical parameters.
Prodigiosin ; Serratia marcescens ; Marine Biology

Aims: To evaluate the antibacterial efficacy of ethyl acetate extract of Aspergillus flavus IBRL-C8 against Gram-positive and Gram-negative bacteria. Methodology and results: In this experiment, an endophytic fungus which identified as A. flavus IBRL-C8 was extracted using ethyl acetate and methanol, from Senna siamea, prior to in vitro antibacterial test on eight Gram-bacteria. The results were significantly more enunciated to the ethyl acetate extract since the Gram-bacteria signified 9.0 to 20.0 mm of inhibition zones on Muller Hinton Agar (MHA) during disc diffusion assay. Minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC) of the extract were ranged from 125-1000 µg/mL and 125-2000 µg/mL, respectively. Time-kill assay depicted the ethyl acetate extract of A. flavus IBRL-C8 exceptionally retarded methicillin-resistant Staphylococcus aureus (MRSA) and also manifested extended antibacterial activity. The maximum reduction in cell numbers occurred at 2MIC concentration (250 µg/mL) during the interval time of 16 h. The malformations noticed from microscopic observations where the transformation of structural annihilation from regular spherical morphology to non-spherical shape with an irregular surface and also disruption around the cell membrane when the MRSA treated with ethyl acetate extract of A. flavus IBRL-C8. Conclusion, significance and impact of study This study proposed the ethyl acetate extract of A. flavus IBRL-C8 as a potential antibacterial agent against MRSA infection, which can be useful in pharmaceutical application.
Aspergillus flavus ; Anti-Bacterial Agents

Aims: This present study focused on purification of fungal β-mannanase produced by Aspergillus niger USM F4 and also physicochemical characterisation of the purified enzyme. Methodology and results: The purified β-mannanase with a molecular mass of ~47.4 kDa was demonstrated on SDSPAGE gel. The enzyme signified a purification degree of 4-fold, with final specific activity of 196.42 U/mg. It reached an optimum catalytic activity at pH 4.0 and 60 °C. The thermal stability of the enzyme was up to 70 °C and maintained the 50% activity after 30 min at 80 °C. Meanwhile, the pH stability was in the range of pH 3.0-9.0 and a 30 min half-life at pH 10.0. All chemical substances manifested an inhibitory effect on purified β-mannanase, with SDS (28.16 ± 0.05% residual activity) as the strongest inhibitor, followed by cupric ion (Cu2+) (49.51 ± 0.09% residual activity). As a whole, the enzyme displayed a substrate specificity in the order of locust bean gum (LBG) > carboxymethylcellulose > soluble starch > xylan from oat spelt > α-cellulose. Its preference for LBG has generated the Km and Vmax values of 0.20 mg/mL and 9.82 U/mL, respectively. Conclusion, significance and impact of study The outcomes of our study offer potential for use at industrial scales, particularly in the oligosaccharides production that involve acid-related activity, wide-ranging temperature and pH stability.
Aspergillus niger ; beta-Mannosidase

Aims: Pigments are coloured substances that exhibit important characteristics to many industries including food, textile, cosmetics, food, pharmaceutical and also aquaculture industry. Naturally derived pigments from marine bacteria do not only exhibit the tinctorial property but are also known to possess broad range of antimicrobial activities. From the industrial point of view, the necessity to obtain suitable culture conditions for maximum yield of cell growth and pigment production is of utmost importance. Methodology and results: The effect of cultural conditions, including light, pH, temperature, agitation speed and size of inoculum on bioactivity of an epiphytic marine bacteria, Pseudoalteromonas rubra BF1A IBRL was studied using shake flask technology. The antimicrobial activity was determined using the Lorian method. As a result, prodigiosin pigment extract obtained from P. rubra BF1A IBRL showed inhibitory activity against the MRSA strain. Pseudoalteromonas rubra BF1A IBRL produced the highest level of prodigiosin and anti-MRSA activity (P<0.05) in Marine broth at initial pH of 7.6 incubated at dark condition at temperature of 26 °C, agitation speed of 120 rpm and 2% (v/v) (1 × 106 CFU/mL) of inoculums size. Conclusion, significance and impact of study A high correlation between pigmentation and antibacterial activity were observed anticipating that the pigment has its own antibacterial properties. The above findings supported the fact that epiphytic marine bacteria were fruitful source for pigmented bioactive compounds, and the physical parameters had significantly influence of the pigment production.

Aims: Pigments are coloured substances that exhibit important characteristics to many industries including food, textile, cosmetics, food, pharmaceutical and also aquaculture industry. Naturally derived pigments from marine bacteria do not only exhibit the tinctorial property but are also known to possess broad range of antimicrobial activities. From the industrial point of view, the necessity to obtain suitable culture conditions for maximum yield of cell growth and pigment production is of utmost importance. Methodology and results: The effect of cultural conditions, including light, pH, temperature, agitation speed and size of inoculum on bioactivity of an epiphytic marine bacteria, Pseudoalteromonas rubra BF1A IBRL was studied using shake flask technology. The antimicrobial activity was determined using the Lorian method. As a result, prodigiosin pigment extract obtained from P. rubra BF1A IBRL showed inhibitory activity against the MRSA strain. Pseudoalteromonas rubra BF1A IBRL produced the highest level of prodigiosin and anti-MRSA activity (P<0.05) in Marine broth at initial pH of 7.6 incubated at dark condition at temperature of 26 °C, agitation speed of 120 rpm and 2% (v/v) (1 × 106 CFU/mL) of inoculums size. Conclusion, significance and impact of study A high correlation between pigmentation and antibacterial activity were observed anticipating that the pigment has its own antibacterial properties. The above findings supported the fact that epiphytic marine bacteria were fruitful source for pigmented bioactive compounds, and the physical parameters had significantly influence of the pigment production.

Aims: Marine bacteria are a great source of natural pigments, which can be used as colouring agent in food, textile, cosmetics and aquaculture industry to overcome the drawbacks poses by the synthetic pigments. The aim of the study is to identify the potential bio pigment producer, determine the antimicrobial susceptibilities, and characterize the pigment produced. Methodology and results: In this study, the surface attached marine bacteria isolated from the surface of seaweed, Enteromorpha sp. has been identified as Pseudoalteromonas rubra BF1A IBRL through the molecular identification step. This species produced intracellular and extracellular red pigment with antibacterial activity. The susceptible bacteria include Bacillus subtilis, Bacillus cereus, Methicillin Resistant Staphylococcus aureus, Staphylococcus aureus and also Acinetobacter anitratus with inhibition zone ranges from 7.33 to 10.33 mm, whereas Minimum inhibitory concentration (MIC) values ranges from 0.055 to 8.88 mg/mL. The UV/vis analysis indicated that the maximal absorbance of ISO and DE pigment extract were at 531 and 534 nm, respectively. Based on the antimicrobial activity, the extracellular extract poses greater antibacterial activity, thus was selected as the potential pigment extract and were further evaluated. The Thin Layer Chromatography (TLC) profile of the DE extract showed one major band under visible light ((Rf = 0.87) and the bioautography analysis of the pigmented band showed positive activity against both Gram-positive and Gram-negative bacteria. The pigment in DE extract was identified as prodigiosin based on the spectroscopic properties, presumptive test and HPLC analysis. Conclusion, significance and impact of study This study highlights the dual benefits of the P. rubra BF1A IBRL pigment extract, which exhibited both tinctorial and pharmacology benefits, thus it can be act as colouring agent with own preservative value in food, textile, or cosmetics industries.

Aims: A simple in vitro model system was applied in this study assessing the dynamics of the microbial community associated with the shrimp gut system to understand the changes that influence dietary variables. Methodology and results: The diversity and abundance of microbiome were monitored within two different treatment slurries inoculated with shrimp faecal samples as to mimic the effect of diet manipulation, and 16S rRNA gene of MiSeq Illumina-based sequencing was applied. The different diets tested were a commercial standard diet and a prodigiosin added diet. There was very clear separation between the commercial standard diet and prodigiosin added diet as revealed by the total viable counts (TVC) and sequencing data. It suggested that the microbial community of the shrimp gut system exhibited a dynamic response with the treatments and allochthonous bacterial present. The prodigiosin added diet was clearly separated from the commercial standard diet serving as a potential shrimp feed additive. The sequencing data analysis showed that members of the genera Vibrio, Shigella and Photobacterium became predominant on the commercial standard diet treatment. The prodigiosin-added diet treatments indicated an abundance of members of the genera Micrococcus, Arthrobacter, and Shigella. Conclusion, significance and impact of study In vitro model system-based testing of diets could be a useful method to determine the potential effect of diet manipulation on shrimp gut system microbiome members.

Aims: Endophytes are microorganisms residing in the living tissues of the host plant and may contribute to their hostplant by producing a plethora of bioactive compounds that provide survival value to the plant. This study aimed toevaluate the antimicrobial activity of Aspergillus sp. IBRL MP15 CCL, an endophytic fungus isolated from Swieteniamacrophylla leaf.Methodology and results: The antimicrobial activity was evaluated with disc diffusion and a colorimetric brothmicrodilution test against 15 organisms comprising of 4 Gram-positive bacteria and 4 Gram-negative bacteria, 4 fungiand 3 yeast. On disc diffusion assay, the fungal extract was shown to inhibit the growth of 7 test bacteria and 3 testyeast. The antibacterial activity was more pronounced with extract from fungal culture with host plant extractsupplementation with significantly larger inhibition zones on all susceptible test microorganisms. The minimal inhibitoryconcentration of the extract ranged from 250 to 4000 μg/mL indicating different level of susceptibility of the testedpathogens against the fungal extract. The killing kinetic study shows that antimicrobial activity of the fungal extract isconcentration dependent and it can act as bactericidal at higher concentration.Conclusion, significance and impact of study: The findings of this study suggest that Aspergillus sp. IBRL MP15CCL can be a promising source of antimicrobial agent to be further studied and developed

To extract the bioactive compound from Enteromorpha intestinalis (E. intestinalis) and determine its in vitro antimicrobial activity. Methods: E. intestinalis was extracted by methanol and subjected to antimicrobial screening. The antimicrobial activity was studied by using disc diffusion and broth dilution method. The effect of the extract on the growth profile of the bacterial was also examined via time-kill assay. Microscopy observations using SEM was done to determine the major alterations in the microstructure of methicillin-resistant Staphylococcus aureus (MRSA). Results: The results showed methanolic extract of E. intestinalis exhibited a favourable antimicrobial activity against tested bacteria with produced inhibition zone ranging from 8.0-19.0 mm. However, all the tested fungi and yeast were resistant to the extract treatment. Time kill assay suggested that methanolic extract of E. intestinalis had completely inhibited MRSA growth and also exhibited prolonged antibacterial activity. The main abnormalities noted from the microscopic observations were the structural deterioration in the normal morphology and complete collapsed of the bacteria cells after 36 h of treatment. Conclusions: The significant antibacterial activity shown by crude extract suggested its potential against MRSA infection. The extract may have potential to develop as antibacterial agent in pharmaceutical use.