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Fulltext A rapid and sensitive system for recovery of nucleic acids from Mycobacteria sp. on archived glass slides

A Talip B, Snelling WJ, Sleator RD, Lowery C, Dooley JSG

BMC Microbiol, 2018 11 26;18(1):196.
PMID: 30477427 DOI: 10.1186/s12866-018-1335-0

BACKGROUND: The field of diagnostics continues to advance rapidly with a variety of novel approaches, mainly dependent upon high technology platforms. Nonetheless much diagnosis, particularly in developing countries, still relies upon traditional methods such as microscopy. Biological material, particularly nucleic acids, on archived glass slides is a potential source of useful information both for diagnostic and epidemiological purposes. There are significant challenges faced when examining archived samples in order that an adequate amount of amplifiable DNA can be obtained. Herein, we describe a model system to detect low numbers of bacterial cells isolated from glass slides using (laser capture microscopy) LCM coupled with PCR amplification of a suitable target.
RESULTS: Mycobacterium smegmatis was used as a model organism to provide a proof of principle for a method to recover bacteria from a stained sample on a glass slide using a laser capture system. Ziehl-Neelsen (ZN) stained cells were excised and catapulted into tubes. Recovered cells were subjected to DNA extraction and pre-amplified with multiple displacement amplification (MDA). This system allowed a minimum of 30 catapulted cells to be detected following a nested real-time PCR assay, using rpoB specific primers. The combination of MDA and nested real-time PCR resulted in a 30-fold increase in sensitivity for the detection of low numbers of cells isolated using LCM.
CONCLUSIONS: This study highlights the potential of LCM coupled with MDA as a tool to improve the recovery of amplifiable nucleic acids from archived glass slides. The inclusion of the MDA step was essential to enable downstream amplification. This platform should be broadly applicable to a variety of diagnostic applications and we have used it as a proof of principle with a Mycobacterium sp. model system.
Cephalexin removal by a novel Cu-Zn bionanocomposite biosynthesized in secondary metabolic products of Aspergillus arenarioides EAN603 with pumpkin peels medium: Optimization, kinetic and artificial neural network models

Al-Gheethi A, Noman E, Saphira Radin Mohamed RM, Talip B, Vo DN, Algaifi HA

J Hazard Mater, 2021 10 05;419:126500.
PMID: 34214856 DOI: 10.1016/j.jhazmat.2021.126500

The present study aimed to investigate the removal efficiency of cephalexin (CFX) by a novel Cu-Zn bionanocomposite biosynthesized in the secondary metabolic products of Aspergillus arenarioides EAN603 with pumpkin peels medium (CZ-BNC-APP). The optimization study was performed based on CFX concentrations (1, 10.5 and 20 ppm); CZ-BNC-APP dosage (10, 55 and 100 mg/L); time (10, 55 and 100 min), temperature (20, 32.5 and 45 °C). The artificial neural network (ANN) model was used to understand the CFX behavior for the factors affecting removal process. The CZ-BNC-APP showed an irregular shape with porous structure and size between 20 and 80 nm. The FTIR detected CC, C-O and OH groups. ANN model revealed that CZ-BNC-APP dosage exhibited the vital role in the removal process, while the removal process having a thermodynamic nature. The CFX removal was optimized with 12.41 ppm CFX, 60.60 mg/L of CZ-BNC-APP, after 97.55 min and at 35 °C, the real maximum removal was 95.53% with 100.52 mg g-1 of the maximum adsorption capacity and 99.5% of the coefficient. The adsorption of CFX on CZ-BNC-APP was fitted with pseudo-second-order model and both Langmuir and Freundlich isotherms models. These findings revealed that CZ-BNC-APP exhibited high potential to remove CFX.
Quantitative microbiological risk assessment of complex microbial community in Prawn farm wastewater and applicability of nanoparticles and probiotics for eliminating of antibiotic-resistant bacteria

Noman E, Al-Gheethi A, Radin Mohamed RMS, Talip B, Al-Sahari M, Al-Shaibani M

J Hazard Mater, 2021 10 05;419:126418.
PMID: 34171673 DOI: 10.1016/j.jhazmat.2021.126418

The current review highlighted the quantitative microbiological risk assessment of Vibrio parahaemolyticus in Prawn farm wastewaters (PFWWs) and the applicability of nanoparticles for eliminating antibiotic-resistant bacteria (ARB). The high availability of the antibiotics in the environment and their transmission into human through the food-chain might cause unknown health effects. The aquaculture environments are considered as a reservoir for the antibiotic resistance genes (ARGs) and contributed effectively in the increasing of ABR. The metagenomic analysis is used to explore ARGs in the non-clinical environment. V. parahaemolyticus is among the pathogenic bacteria which are transmitted through sea food causing human acute gastroenteritis due to available thermostable direct hemolysin (tdh), adhesins, TDH related hemolysin (trh). The inactivation of pathogenic bacteria using nanoparticles act by disturbing the cell membrane, interrupting the transport system, DNA and mitochondria damage, and oxidizing the cellular component by reactive oxygen species (ROS). The chloramphenicol, nitrofurans, and nitroimidazole are among the prohibited drugs in fish and fishery product. The utilization of probiotics is the most effective and safe alternative for antibiotics in Prawn aquaculture. This review will ensure public understanding among the readers on how they can decrease the risk of the antimicrobial resistance distribution in the environment.
Inactivation of fungal spores from clinical environment by silver bio-nanoparticles; optimization, artificial neural network model and mechanism

Noman E, Al-Gheethi A, Saphira Radin Mohamed RM, Talip B, Othman N, Hossain S, et al.

Environ Res, 2022 03;204(Pt A):111926.
PMID: 34461120 DOI: 10.1016/j.envres.2021.111926

The present study aimed to assess the efficiency of silver bio-nanoparticles (Ag-NPs) in inactivating of the Aspergillus fumigatus, A. parasiticus and A. flavus var. columnaris and A. aculeatus spores. The AgNPs were synthesized in secondary metabolic products of Penicillium pedernalens 604 EAN. The inactivation process was optimized by response surface methodology (RSM) as a function of Ag NPs volume (1-10 μL/mL); time (10-120 min); pH (5-8); initial fungal concentrations (log10) (3-6). The artificial neural network (ANN) model was used to understand the behavior of spores for the factors affecting inactivation process. The best conditions to achieved SAL 10-6 of the fungal spores were recorded with 3.46 μl/mL of AgNPs, after 120 min at pH 5 and with 6 log of initial fungal spore concentrations, at which 5.99 vs. 6.09 (SAL 10-6) log reduction was recorded in actual and predicted results respectively with coefficient of 87.00%. The ANN revealed that the timehas major contribution in the inactivation process compare to Ag NPs volume. The fungal spores were totally inactivated (SAL 10-6, 6 log reduction with 99.9999%) after 110 min of the inactivation process, 10 min more was required to insure the irreversible inactivation of the fungal spores. The absence of protease and cellulase enzymes production confirm the total inactivation of the fungal spores. FESEM analysis revealed that the AgNPs which penetrated the fungal spores leading to damage and deform the fungal spore morphology. The AFM analysis confirmed the total spore surface damage. The bands in the range of the Raman spectroscopy from 1300 to 1600 cm-1 in the inactivated spores indicate the presence of CH3, CH2 and the deformation of lipids released outside the spore cytoplasm. These finding indicate that the AgNPs has high potential as a green alternative inactivation process for the airborne fungal spores.