A portable label-free electrochemical DNA biosensor for rapid detection of Salmonella Typhi.

A highly accurate, rapid, portable, and robust platform for detecting Salmonella enterica serovar Typhi (S. Typhi) is crucial for early-stage diagnosis of typhoid to avert and control the outbreaks of this pathogen, which threaten global public health. This study presents a proof-of-concept for our developed label-free electrochemical DNA biosensor system for S. Typhi detection, which employs a printed circuit board gold electrode (PCBGE), integrated with a portable potentiostat reader. Initially, the functionalized DNA biosensor and target detection were characterized using cyclic voltammetry (CV), differential pulse voltammetry (DPV), and electrochemical impedance spectroscopy (EIS) methods using a benchtop potentiostat. Interestingly, the newly developed DNA biosensor can identify target single-stranded DNA concentrations ranging from 10 nM to 20 µM, achieving a detection limit of 7.6 nM within a brief 5 minute timeframe. Under optimal detection conditions, the DNA biosensor exhibits remarkable selectivity, capable of distinguishing a single mismatch base pair from the target single-stranded DNA sequence. We then evaluated the feasibility of the developed DNA biosensor system as a diagnostic tool by detecting S. Typhi in 50 clinical samples using a portable potentiostat reader based on the DPV technique. Remarkably, the developed biosensor can distinctly distinguish between positive and negative samples, indicating that the miniaturised DNA biosensor system is practical for detecting S. Typhi in real biological samples. The developed DNA biosensor device in this work proves to be a promising point-of-care (POC) device for Salmonella detection due to its swift detection time, uncomplicated design, and streamlined workflow detection system.

Screening biosurfactant-producing actinomycetes: Identification of Streptomyces sp. RP1 as a potent species for bioremediation.

This study aimed to isolate biosurfactant-producing and hydrocarbon-degrading actinomycetes from different soils using glycerol-asparagine and starch-casein media with an antifungal agent. The glycerol-asparagine agar exhibited the highest number of actinomycetes, with a white, low-opacity medium supporting pigment production and high growth. Biosurfactant analyses, such as drop collapse, oil displacement, emulsification, tributyrin agar test, and surface tension measurement, were conducted. Out of 25 positive isolates, seven could utilize both olive oil and black oil for biosurfactant production, and only isolate RP1 could produce biosurfactant when grown in constrained conditions with black oil as the sole carbon source and inducer, demonstrating in situ bioremediation potential. Isolate RP1 from oil-spilled garden soil is Gram-staining-positive with a distinct earthy odor, melanin formation, and white filamentous colonies. It has a molecular size of ~621 bp and 100% sequence similarity to many Streptomyces spp. Morphological, biochemical, and 16 S rRNA analysis confirmed it as Streptomyces sp. RP1, showing positive results in all screenings, including high emulsification activity against kerosene (27.2%) and engine oil (95.8%), oil displacement efficiency against crude oil (7.45 cm), and a significant reduction in surface tension (56.7 dynes/cm). Streptomyces sp. RP1 can utilize citrate as a carbon source, tolerate sodium chloride, resist lysozyme, degrade petroleum hydrocarbons, and produce biosurfactant at 37°C in a 15 mL medium culture, indicating great potential for bioremediation and various downstream industrial applications with optimization.

A label-free electrochemical DNA biosensor used a printed circuit board gold electrode (PCBGE) to detect SARS-CoV-2 without amplification.

The emergence of coronavirus disease 2019 (COVID-19) motivates continuous efforts to develop robust and accurate diagnostic tests to detect severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). Detection of viral nucleic acids provides the highest sensitivity and selectivity for diagnosing early and asymptomatic infection because the human immune system may not be active at this stage. Therefore, this work aims to develop a label-free electrochemical DNA biosensor for SARS-CoV-2 detection using a printed circuit board-based gold substrate (PCBGE). The developed sensor used the nucleocapsid phosphoprotein (N) gene as a biomarker. The DNA sensor-based PCBGE was fabricated by self-assembling a thiolated single-stranded DNA (ssDNA) probe onto an Au surface, which performed as the working electrode (WE). The Au surface was then treated with 6-mercapto-1-hexanol (MCH) before detecting the target N gene to produce a well-oriented arrangement of the immobilized ssDNA chains. The successful fabrication of the biosensor was characterized using cyclic voltammetry (CV), electrochemical impedance spectroscopy (EIS), and atomic force microscopy (AFM). The DNA biosensor performances were evaluated using a synthetic SARS-CoV-2 genome and 20 clinical RNA samples from healthy and infected individuals through EIS. The developed DNA biosensor can detect as low as 1 copy per µL of the N gene within 5 minutes with a LOD of 0.50 µM. Interestingly, the proposed DNA sensor could distinguish the expression of SARS-CoV-2 RNA in a patient diagnosed with COVID-19 without any amplification technique. We believe that the proposed DNA sensor platform is a promising point-of-care (POC) device for COVID-19 viral infection since it offers a rapid detection time with a simple design and workflow detection system, as well as an affordable diagnostic assay.

Aptamer-Based Electrochemical Biosensors for the Detection of Salmonella: A Scoping Review.

The development of rapid, accurate, and efficient detection methods for Salmonella can significantly control the outbreak of salmonellosis that threatens global public health. Despite the high sensitivity and specificity of the microbiological, nucleic-acid, and immunological-based methods, they are impractical for detecting samples outside of the laboratory due to the requirement for skilled individuals and sophisticated bench-top equipment. Ideally, an electrochemical biosensor could overcome the limitations of these detection methods since it offers simplicity for the detection process, on-site quantitative analysis, rapid detection time, high sensitivity, and portability. The present scoping review aims to assess the current trends in electrochemical aptasensors to detect and quantify Salmonella. This review was conducted according to the latest Preferred Reporting Items for Systematic review and Meta-Analyses extension for Scoping Reviews (PRISMA-ScR) guidelines. A literature search was performed using aptamer and Salmonella keywords in three databases: PubMed, Scopus, and Springer. Studies on electrochemical aptasensors for detecting Salmonella published between January 2014 and January 2022 were retrieved. Of the 787 studies recorded in the search, 29 studies were screened for eligibility, and 15 studies that met the inclusion criteria were retrieved for this review. Information on the Salmonella serovars, targets, samples, sensor specification, platform technologies for fabrication, electrochemical detection methods, limit of detection (LoD), and detection time was discussed to evaluate the effectiveness and limitations of the developed electrochemical aptasensor platform for the detection of Salmonella. The reported electrochemical aptasensors were mainly developed to detect Salmonella enterica Typhimurium in chicken meat samples. Most of the developed electrochemical aptasensors were fabricated using conventional electrodes (13 studies) rather than screen-printed electrodes (SPEs) (two studies). The developed aptasensors showed LoD ranges from 550 CFU/mL to as low as 1 CFU/mL within 5 min to 240 min of detection time. The promising detection performance of the electrochemical aptasensor highlights its potential as an excellent alternative to the existing detection methods. Nonetheless, more research is required to determine the sensitivity and specificity of the electrochemical sensing platform for Salmonella detection, particularly in human clinical samples, to enable their future use in clinical practice.

Utilizing Electrochemical-Based Sensing Approaches for the Detection of SARS-CoV-2 in Clinical Samples: A Review.

The development of precise and efficient diagnostic tools enables early treatment and proper isolation of infected individuals, hence limiting the spread of coronavirus disease 2019 (COVID-19). The standard diagnostic tests used by healthcare workers to diagnose severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection have some limitations, including longer detection time, the need for qualified individuals, and the use of sophisticated bench-top equipment, which limit their use for rapid SARS-CoV-2 assessment. Advances in sensor technology have renewed the interest in electrochemical biosensors miniaturization, which provide improved diagnostic qualities such as rapid response, simplicity of operation, portability, and readiness for on-site screening of infection. This review gives a condensed overview of the current electrochemical sensing platform strategies for SARS-CoV-2 detection in clinical samples. The fundamentals of fabricating electrochemical biosensors, such as the chosen electrode materials, electrochemical transducing techniques, and sensitive biorecognition molecules, are thoroughly discussed in this paper. Furthermore, we summarised electrochemical biosensors detection strategies and their analytical performance on diverse clinical samples, including saliva, blood, and nasopharyngeal swab. Finally, we address the employment of miniaturized electrochemical biosensors integrated with microfluidic technology in viral electrochemical biosensors, emphasizing its potential for on-site diagnostics applications.

Performance of Immunodiagnostic Tests for Typhoid Fever: A Systematic Review and Meta-Analysis.

Typhoid fever, also known as typhoid, is a life-threatening bacterial infection that remains a global health concern. The infection is associated with a significant morbidity and mortality rate, resulting in an urgent need for specific and rapid detection tests to aid prevention and management of the disease. The present review aims to assess the specificity and sensitivity of the available literature on the immunodiagnostics of typhoid fever. A literature search was conducted using three databases (PubMed, ProQuest and Scopus) and manual searches through the references of identified full texts to retrieve relevant literature published between 1 January 2011 and 31 December 2020. Of the 577 studies identified in our search, 12 were included in further analysis. Lipopolysaccharides (LPS) and hemolysin E (HlyE) were the most frequently studied antigens. The specimens examined in these studies included serum and saliva. Using blood culture as the gold standard, anti-LPS IgA gave the highest sensitivity of 96% (95% CI: 93-99) and specificity of 96% (95% CI: 93-99) for distinguishing between typhoid cases and healthy controls, whereas the combination of anti-LPS and anti-flagellin total IgGAM gave the highest sensitivity of 93% (95% CI: 86-99) and specificity of 95% (95% CI: 89-100) for distinguishing typhoid cases and other febrile infections. A comparably high sensitivity of 92% (95% CI: 86-98) and specificity of 89% (95% CI: 78-100) were shown in testing based on detection of the combination of anti-LPS (IgA and IgM) and anti-HlyE IgG as well as a slightly lower sensitivity of 91% (95% CI: 74-100) in the case of anti-50kDa IgA. Anti-50kDa IgM had the lowest sensitivity of 36% (95% CI: 6-65) against both healthy and febrile controls. The development of a rapid diagnostic test targeting antibodies against lipopolysaccharides combined with flagellin appeared to be a suitable approach for the rapid detection test of typhoid fever. Saliva is added benefit for rapid typhoid diagnosis since it is less invasive. As a result, further studies could be done to develop additional approaches for adopting such samples.

Advancement in Salmonella Detection Methods: From Conventional to Electrochemical-Based Sensing Detection.

Large-scale food-borne outbreaks caused by Salmonella are rarely seen nowadays, thanks to the advanced nature of the medical system. However, small, localised outbreaks in certain regions still exist and could possess a huge threat to the public health if eradication measure is not initiated. This review discusses the progress of Salmonella detection approaches covering their basic principles, characteristics, applications, and performances. Conventional Salmonella detection is usually performed using a culture-based method, which is time-consuming, labour intensive, and unsuitable for on-site testing and high-throughput analysis. To date, there are many detection methods with a unique detection system available for Salmonella detection utilising immunological-based techniques, molecular-based techniques, mass spectrometry, spectroscopy, optical phenotyping, and biosensor methods. The electrochemical biosensor has growing interest in Salmonella detection mainly due to its excellent sensitivity, rapidity, and portability. The use of a highly specific bioreceptor, such as aptamers, and the application of nanomaterials are contributing factors to these excellent characteristics. Furthermore, insight on the types of biorecognition elements, the principles of electrochemical transduction elements, and the miniaturisation potential of electrochemical biosensors are discussed.

Production of lipopeptide biosurfactant in batch and fed-batch Streptomyces sp. PBD-410L cultures growing on palm oil.

The present study focused on lipopeptide biosurfactant production by Streptomyces sp. PBD-410L in batch and fed-batch fermentation in a 3-L stirred-tank reactor (STR) using palm oil as a sole carbon source. In batch cultivation, the impact of bioprocessing parameters, namely aeration rate and agitation speed, was studied to improve biomass growth and lipopeptide biosurfactant production. The maximum oil spreading technique (OST) result (45 mm) which corresponds to 3.74 g/L of biosurfactant produced, was attained when the culture was agitated at 200 rpm and aeration rate of 0.5 vvm. The best aeration rate and agitation speed obtained from the batch cultivation was adopted in the fed-batch cultivation using DO-stat feeding strategy to further improve the lipopeptide biosurfactant production. The lipopeptide biosurfactant production was enhanced from 3.74 to 5.32 g/L via fed-batch fermentation mode at an initial feed rate of 0.6 mL/h compared to that in batch cultivation. This is the first report on the employment of fed-batch cultivation on the production of biosurfactant by genus Streptomyces.

The influence of agitation on oil substrate dispersion and oxygen transfer in Pseudomonas aeruginosa USM-AR2 fermentation producing rhamnolipid in a stirred tank bioreactor.

Water-immiscible substrate, diesel, was supplied as the main substrate in the fermentation of Pseudomonas aeruginosa USM-AR2 producing rhamnolipid biosurfactant, in a stirred tank bioreactor. In addition to the typical gas-aqueous system, this system includes gas-hydrocarbon-aqueous phases and the presence of surfactant (rhamnolipid) in the fermentation broth. The effect of diesel dispersion on volumetric oxygen transfer coefficient, k L a, and thus oxygen transfer, was evaluated at different agitations of 400, 500 and 600 rpm. The oxygen transfer in this oil-water-surfactant system was shown to be affected by different oil dispersion at those agitation rates. The highest diesel dispersion was obtained at 500 rpm or impeller tip speed of 1.31 m/s, compared to 400 and 600 rpm, which led to the highest k L a, growth and rhamnolipid production by P. aeruginosa USM-AR2. This showed the highest substrate mixing and homogenization at this agitation speed that led to the efficient substrate utilization by the cells. The oxygen uptake rate of P. aeruginosa USM-AR2 was 5.55 mmol/L/h, which showed that even the lowest k L a (48.21 h-1) and hence OTR (57.71 mmol/L/h) obtained at 400 rpm was sufficient to fulfill the oxygen demand of the cells. The effect of rhamnolipid concentration on k L a showed that k L a increased as rhamnolipid concentration increased to 0.6 g/L before reaching a plateau. This trend was similar for all agitation rates of 400, 500 and 600 rpm, which might be due to the increase in the resistance to oxygen transfer (k L decrease) and the increase in the specific interfacial area (a).

Production and partial characterization of biosurfactant produced by Streptomyces sp. R1.

The present study focused on developing a wild-type actinomycete isolate as a model for a non-pathogenic filamentous producer of biosurfactants. A total of 33 actinomycetes isolates were screened and their extracellular biosurfactants production was evaluated using olive oil as the main substrate. Out of 33 isolates, 32 showed positive results in the oil spreading technique (OST). All isolates showed good emulsification activity (E24) ranging from 84.1 to 95.8%. Based on OST and E24 values, isolate R1 was selected for further investigation in biosurfactant production in an agitated submerged fermentation. Phenotypic and genotypic analyses tentatively identified isolate R1 as a member of the Streptomyces genus. A submerged cultivation of Streptomyces sp. R1 was carried out in a 3-L stirred-tank bioreactor. The influence of impeller tip speed on volumetric oxygen transfer coefficient (k L a), growth, cell morphology and biosurfactant production was observed. It was found that the maximum biosurfactant production, indicated by the lowest surface tension measurement (40.5 ± 0.05 dynes/cm) was obtained at highest k L a value (50.94 h-1) regardless of agitation speed. The partially purified biosurfactant was obtained at a concentration of 7.19 g L-1, characterized as a lipopeptide biosurfactant and was found to be stable over a wide range of temperature (20-121 °C), pH (2-12) and salinity [5-20% (w/v) of NaCl].