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Fulltext Artificial Neural Networks (ANNs) and Response Surface Methodology (RSM) Approach for Modelling the Optimization of Chromium (VI) Reduction by Newly Isolated Acinetobacter radioresistens Strain NS-MIE from Agricultural Soil

Ram Talib NS, Halmi MIE, Abd Ghani SS, Zaidan UH, Shukor MYA

Biomed Res Int, 2019;2019:5785387.
PMID: 31240217 DOI: 10.1155/2019/5785387

Numerous technologies and approaches have been used in the past few decades to remove hexavalent chromium (Cr[VI]) in wastewater and the environment. However, these conventional technologies are not economical and efficient in removing Cr(VI) at a very low concentration (1-100 ppm). As an alternative, the utilization of bioremediation techniques which uses the potential of microorganisms could represent an effective technique for the detoxification of Cr(VI). In this study, we reported a newly isolated bacterium identified as Acinetobacter radioresistens sp. NS-MIE from Malaysian agricultural soil. The chromate reduction potential of strain NS-MIE was optimized using RSM and ANN techniques. The optimum condition predicted by RSM for the bacterium to reduce hexavalent chromium occurred at pH 6, 10 g/L ppm of nutrient broth (NB) concentration and 100 ppm of chromate concentration while the optimum condition predicted by ANN is at pH 6 and 10 g/L of NB concentration and of 60 ppm of chromate concentration with chromate reduction (%) of 75.13 % and 96.27 %, respectively. The analysis by the ANN model shows better prediction data with a higher R2 value of 0.9991 and smaller average absolute deviation (AAD) and root mean square error (RMSE) of 0.33 % and 0.302 %, respectively. Validation analysis showed the predicted values by RSM and ANN were close to the validation values, whereas the ANN showed the lowest deviation, 2.57%, compared to the RSM. This finding suggests that the ANN showed a better prediction and fitting ability compared to the RSM for the nonlinear regression analysis. Based on this study, A. radioresistens sp. NS-MIE exhibits strong potential characteristics as a candidate for the bioremediation of hexavalent chromium in the environment.
Fulltext Improvement of Ficin-Based Inhibitive Enzyme Assay for Toxic Metals Using Response Surface Methodology and Its Application for Near Real-Time Monitoring of Mercury in Marine Waters

Uba G, Manogaran M, Gunasekaran B, Halmi MIE, Shukor MYA

Int J Environ Res Public Health, 2020 11 19;17(22).
PMID: 33227985 DOI: 10.3390/ijerph17228585

Potentially toxic metals pollution in the Straits of Malacca warrants the development of rapid, simple and sensitive assays. Enzyme-based assays are excellent preliminary screening tools with near real-time potential. The heavy-metal assay based on the protease ficin was optimized for mercury detection using response surface methodology. The inhibitive assay is based on ficin action on the substrate casein and residual casein is determined using the Coomassie dye-binding assay. Toxic metals strongly inhibit this hydrolysis. A central composite design (CCD) was utilized to optimize the detection of toxic metals. The results show a marked improvement for the concentration causing 50% inhibition (IC50) for mercury, silver and copper. Compared to one-factor-at-a-time (OFAT) optimization, RSM gave an improvement of IC50 (mg/L) from 0.060 (95% CI, 0.030-0.080) to 0.017 (95% CI, 0.016-0.019), from 0.098 (95% CI, 0.077-0.127) to 0.028 (95% CI, 0.022-0.037) and from 0.040 (95% CI, 0.035-0.045) to 0.023 (95% CI, 0.020-0.027), for mercury, silver and copper, respectively. A near-real time monitoring of mercury concentration in the Straits of Malacca at one location in Port Klang was carried out over a 4 h interval for a total of 24 h and validated by instrumental analysis, with the result revealing an absence of mercury pollution in the sampling site.
Bioluminescent method for the rapid screening of toxic heayy metals in environmental samples using Photobacterium leiognathi strain AK-MIE

Kassim A, Halmi MIE, Gani SSA, Zaidan UH, Othman R, Mahmud K, et al.

Ecotoxicol Environ Saf, 2020 Jun 15;196:110527.
PMID: 32278138 DOI: 10.1016/j.ecoenv.2020.110527

Assessment of eco-toxicant using bioluminescent bacterial assay is a widely used and globally accepted method. In this work, a new luminescent bacterium was isolated from squid (Loligo duvauceli) and identified as Photobacterium leiognathi strain AK-MIE using 16S rRNA, phylogeny analysis. The predicted optimum conditions by RSM were 2.76% (w/v) NaCl, 2.28% (w/v) peptone, 0.34% (w/v) yeast extract, and pH 6.83 with 541,211.80 RLU of luminescent production whereas the predicted optimum conditions by ANN were 2.21% (w/v) NaCl, 2.27% (w/v) peptone, 0.39% (w/v) yeast extract, and pH 6.94 which produced 541,986.20 RLU. The validation analysis of both RSM and ANN show 0.60% and 0.69% deviation from the predicted results indicating that both models provided good quality predictions with ANN showing a superior data fitting capability for non-linear regression analysis. Toxicity tests show strain AK-MIE was sensitive to mercury (concentration causing 50% inhibition or IC50 of 0.00978 mgL-1), followed by cadmium (IC50 of 0.5288 mgL-1), copper IC50 of (0.8117 mgL-1), silver (IC50 of 1.109 mgL-1), and lead (IC50 of 10.71 mgL-1) which are more sensitive than previously isolated luminescent bacteria, suggesting that strain AK-MIE has the potential to be used in toxicity assessment of heavy metals in the environment. Based on the field trial results, several sediment samples from industrial areas in Bangi, Selangor managed to inhibit the bioluminescence of strain AK-MIE. Validation method carried out using ICP-MS proved the presence of several toxic heavy metal elements.
Fulltext Evaluation of conventional and response surface level optimisation of n-dodecane (n-C12) mineralisation by psychrotolerant strains isolated from pristine soil at Southern Victoria Island, Antarctica

Habib S, Ahmad SA, Johari WLW, Shukor MYA, Alias SA, Khalil KA, et al.

Microb Cell Fact, 2018 Mar 17;17(1):44.
PMID: 29549881 DOI: 10.1186/s12934-018-0889-8

BACKGROUND: Biodegradation of hydrocarbons in Antarctic soil has been reported to be achieved through the utilisation of indigenous cold-adapted microorganisms. Although numerous bacteria isolated from hydrocarbon-contaminated sites in Antarctica were able to demonstrate promising outcomes in utilising hydrocarbon components as their energy source, reports on the utilisation of hydrocarbons by strains isolated from pristine Antarctic soil are scarce. In the present work, two psychrotolerant strains isolated from Antarctic pristine soil with the competency to utilise diesel fuel as the sole carbon source were identified and optimised through conventional and response surface method.
RESULTS: Two potent hydrocarbon-degraders (ADL15 and ADL36) were identified via partial 16S rRNA gene sequence analysis, and revealed to be closely related to the genus Pseudomonas and Rhodococcus sp., respectively. Factors affecting diesel degradation such as temperature, hydrocarbon concentration, pH and salt tolerance were studied. Although strain ADL36 was able to withstand a higher concentration of diesel than strain ADL15, both strains showed similar optimal condition for the cell's growth at pH 7.0 and 1.0% (w/v) NaCl at the conventional 'one-factor-at-a-time' level. Both strains were observed to be psychrotrophs with optimal temperatures of 20 °C. Qualitative and quantitative analysis were performed with a gas chromatograph equipped with a flame ionisation detector to measure the reduction of n-alkane components in diesel. In the pre-screening medium, strain ADL36 showed 83.75% of n-dodecane mineralisation while the reduction of n-dodecane by strain ADL15 was merely at 22.39%. The optimised condition for n-dodecane mineralisation predicted through response surface methodology enhanced the reduction of n-dodecane to 99.89 and 38.32% for strain ADL36 and strain ADL15, respectively.
CONCLUSIONS: Strain ADL36 proves to be a better candidate for bioaugmentation operations on sites contaminated with aliphatic hydrocarbons especially in the Antarctic and other cold regions. The results obtained throughout strongly supports the use of RSM for medium optimisation.
Fulltext Biodegradation of Petroleum Sludge by Methylobacterium sp. Strain ZASH

Harumain ZAS, Mohamad MAN, Nordin NFH, Shukor MYA

Trop Life Sci Res, 2023 Jun;34(2):197-222.
PMID: 38144383 DOI: 10.21315/tlsr2023.34.2.10

A bacterium was isolated from sludge-contaminated soil in a petroleum refinery and tested for its ability to degrade aliphatic hydrocarbon compounds present in petroleum sludge. The isolate was grown on minimal salt media agar supplemented with 1% (w/v) petroleum sludge. The isolate was tentatively identified as Methylobacterium s p. s t rain ZASH based on the partial 16s rDNA molecular phylogeny. The bacterium grew optimally between the temperatures of 30°C and 35°C, pH 7 and 7.5, 0.5% and 1.5% (v/v) Tween 80 as the surfactant, and between 1% and 2% (w/v) peptone as the nitrogen source. The constants derived from the Haldane equation were μmax = 0.039 hr-1, Ks = 0.385% (w/v) total petroleum hydrocarbons (TPH) or 3,850 mg/L TPH, and Ki =1.12% (w/v) TPH or 11,200 mg/L. The maximum biodegradation rate exhibited by this strain was 19 mg/L/hr at an initial TPH concentration of 10,000 mg/L. Gas chromatography analysis revealed that after 15 days the strain was able to degrade all aliphatic n-alkanes investigated with different efficiencies. Shorter n-alkanes were generally degraded more rapidly than longer n-alkanes with 90% removal for C-12 compared to only 30% removal for C-36. The addition of sawdust did not improve bacterial degradation of petroleum hydrocarbons, but it assisted in the removal of remaining undegraded hydrocarbons through adsorption.
Fulltext Biodecolourisation of Reactive Red 120 as a Sole Carbon Source by a Bacterial Consortium-Toxicity Assessment and Statistical Optimisation

Manogaran M, Yasid NA, Othman AR, Gunasekaran B, Halmi MIE, Shukor MYA

Int J Environ Res Public Health, 2021 Mar 02;18(5).
PMID: 33801387 DOI: 10.3390/ijerph18052424

The application of microorganisms in azo dye remediation has gained significant attention, leading to various published studies reporting different methods for obtaining the best dye decolouriser. This paper investigates and compares the role of methods and media used in obtaining a bacterial consortium capable of decolourising azo dye as the sole carbon source, which is extremely rare to find. It was demonstrated that a prolonged acclimation under low substrate availability successfully isolated a novel consortium capable of utilising Reactive Red 120 dye as a sole carbon source in aerobic conditions. This consortium, known as JR3, consists of Pseudomonas aeruginosa strain MM01, Enterobacter sp. strain MM05 and Serratia marcescens strain MM06. Decolourised metabolites of consortium JR3 showed an improvement in mung bean's seed germination and shoot and root length. One-factor-at-time optimisation characterisation showed maximal of 82.9% decolourisation at 0.7 g/L ammonium sulphate, pH 8, 35 °C, and RR120 concentrations of 200 ppm. Decolourisation modelling utilising response surface methodology (RSM) successfully improved decolourisation even more. RSM resulted in maximal decolourisation of 92.79% using 0.645 g/L ammonium sulphate, pH 8.29, 34.5 °C and 200 ppm RR120.
Colorimetric detection of mercury (Hg2+) using UV-vis spectroscopy and digital image analysis based on gold nanoparticles functionalized with bromelain enzyme

Suhaidi NA, Halmi MIE, Rashidi AA, Anuar MFM, Mahmud K, Kusnin N, et al.

3 Biotech, 2023 May;13(5):121.
PMID: 37033387 DOI: 10.1007/s13205-023-03532-z

A very sensitive and selective colorimetric biosensor for the measurement of mercury ion (Hg2+) in environmental samples has been developed using functionalized gold nanoparticles with bromelain enzyme (brn-AuNPs). This work has shown that Hg2+ measurement based on spectrophotometer and digital image analysis is a very innovative and successful method for providing an effective preliminary system and has promise for the future of water quality biomonitoring. Response Surface Methodology (RSM), a Box-Behnken design-based technique, was used to identify the optimum levels of functionalization of bromelain to AuNPs. The created model's validity was confirmed, and statistical analysis revealed that the ideal functionalize conditions were 1 mM of AuNPs, functionalize with 0.59 mM bromelain concentration on 14 ℃ temperature and 72 h incubation time. The lowest colorimetric detection concentration (LOD) of brn-AuNPs of Hg2+ was 0.0092 ppm and 0.011 ppm for spectrophotometer and digital image analysis. As shown, digital image analysis had advantages based on the LOD result comparable to UV-VIS spectrophotometer. The practical application of the brn-AuNPs sensing was proven with mercury determination in water samples. The present study developed a robust sensor, which successfully implemented in a compact portable sensor kit, turning this sensor into a very potent tool for the development water quality biomonitoring system of Hg2+ application.