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Fulltext FabV/Triclosan Is an Antibiotic-Free and Cost-Effective Selection System for Efficient Maintenance of High and Medium-Copy Number Plasmids in Escherichia coli

Ali SA, Chew YW

PLoS One, 2015;10(6):e0129547.
PMID: 26057251 DOI: 10.1371/journal.pone.0129547

Antibiotic resistance genes and antibiotics are frequently used to maintain plasmid vectors in bacterial hosts such as Escherichia coli. Due to the risk of spread of antibiotic resistance, the regulatory authorities discourage the use of antibiotic resistance genes/antibiotics for the maintenance of plasmid vectors in certain biotechnology applications. Overexpression of E. coli endogenous fabI gene and subsequent selection on Triclosan has been proposed as a practical alternative to traditional antibiotic selection systems. Unfortunately, overexpression of fabI cannot be used to select medium-copy number plasmids, typically used for the expression of heterologous proteins in E. coli. Here we report that Vibrio cholera FabV, a functional homologue of E. coli FabI, can be used as a suitable marker for the selection and maintenance of both high and medium-copy number plasmid vectors in E. coli.
Fulltext Use of FabV-Triclosan Plasmid Selection System for Efficient Expression and Production of Recombinant Proteins in Escherichia coli

Ali SA, Chew YW, Omar TC, Azman N

PLoS One, 2015;10(12):e0144189.
PMID: 26642325 DOI: 10.1371/journal.pone.0144189

Maintenance of recombinant plasmid vectors in host bacteria relies on the presence of selection antibiotics in the growth media to suppress plasmid -free segregants. However, presence of antibiotic resistance genes and antibiotics themselves is not acceptable in several applications of biotechnology. Previously, we have shown that FabV-Triclosan selection system can be used to select high and medium copy number plasmid vectors in E. coli. Here, we have extended our previous work and demonstrated that expression vectors containing FabV can be used efficiently to express heterologous recombinant proteins in similar or better amounts in E. coli host when compared with expression vectors containing β-lactamase. Use of small amount of non-antibiotic Triclosan as selection agent in growth medium, enhanced plasmid stability, applicability in various culture media, and compatibility with other selection systems for multiple plasmid maintenance are noteworthy features of FabV-Triclosan selection system.
Fulltext Medical students' choice of specialty and factors determining their choice: a cross-sectional questionnaire survey in Melaka-Manipal Medical College, Malaysia

Chew YW, Rajakrishnan S, Low CA, Jayapalan PK, Sreeramareddy CT

Biosci Trends, 2011;5(2):69-76.
PMID: 21572250

Information about medical students' choice of specialty can be helpful for planning health manpower. However, such information from medical students in Malaysian medical schools is lacking. We carried out a cross-sectional questionnaire survey among fourth- and fifth-year medical undergraduate students at Melaka-Manipal Medical College. A total of 425 students responded to the survey questionnaire. Nearly a quarter of the students indicated internal medicine as their choice of specialty. Other choices were general surgery (13.2%), pediatrics (11.3%), orthopedics (12.7%) and obstetrics & gynecology (Ob/Gyn) (12.1%). Female students (OR 1.91; 95% CI 1.18-3.08), fourth-year students (OR 1.9; 95% CI 1.15-3.12), and students who reported a higher self-rated knowledge of their subject of choice were more likely to choose internal medicine and allied specialties (OR 1.53; 95% CI 1.07-2.19). The influence of teaching faculty and consultants at the teaching hospitals (74.4%) and inspiration obtained during clinical postings (71.9%) were the factors which were rated by the most students as 'important' for choosing a specialty. About half of the students intended to pursue their postgraduate studies in Malaysia, most of the rest in the United Kingdom or Australia. While internal medicine and surgical subspecialties were preferred, students were not inclined towards primary care or diagnostic subspecialties. Incentives should be provided and other measures should be taken to make these branches more attractive.
Fulltext Engineering and Validation of a Vector for Concomitant Expression of Rare Transfer RNA (tRNA) and HIV-1 nef Genes in Escherichia coli

Mualif SA, Teow SY, Omar TC, Chew YW, Yusoff NM, Ali SA

PLoS One, 2015;10(7):e0130446.
PMID: 26147991 DOI: 10.1371/journal.pone.0130446

Relative ease in handling and manipulation of Escherichia coli strains make them primary candidate to express proteins heterologously. Overexpression of heterologous genes that contain codons infrequently used by E. coli is related with difficulties such as mRNA instability, early termination of transcription and/or translation, deletions and/or misincorporation, and cell growth inhibition. These codon bias -associated problems are addressed by co-expressing ColE1-compatible, rare tRNA expressing helper plasmids. However, this approach has inadequacies, which we have addressed by engineering an expression vector that concomitantly expresses the heterologous protein of interest, and rare tRNA genes in E. coli. The expression vector contains three (argU, ileY, leuW) rare tRNA genes and a useful multiple cloning site for easy in-frame cloning. To maintain the overall size of the parental plasmid vector, the rare tRNA genes replaced the non-essential DNA segments in the vector. The cloned gene is expressed under the control of T7 promoter and resulting recombinant protein has a C-terminal 6His tag for IMAC-mediated purification. We have evaluated the usefulness of this expression vector by expressing three HIV-1 genes namely HIV-1 p27 (nef), HIV-1 p24 (ca), and HIV-1 vif in NiCo21(DE3) E.coli and demonstrated the advantages of using expression vector that concomitantly expresses rare tRNA and heterologous genes.
Fulltext Pneumocephalus Following Combined Spinal Epidural Anaesthesia for Total Knee Arthroplasty: A Case Report

Chew YW, Suppan VK, Ashutosh SR, Tew MM, Jimmy-Tan JH

Malays Orthop J, 2017 Nov;11(3):42-44.
PMID: 29326765 MyJurnal DOI: 10.5704/MOJ.1711.001

The authors describe a case of pneumocephalus following epidural anaesthesia for total knee arthroplasty. Multiple attempts in locating the epidural space for the anaesthesia and the use of loss of resistance to air (LORA) technique were identified as the source of air entry. Supportive management was given including high flow oxygenation therapy and spontaneous reabsorption of air was noted five days after surgery. The presence of pneumocephalus should be kept in mind if patient develops neurological complications postoperatively following epidural anaesthesia.
Fulltext One-year follow-up of efficacy and cost of repeated doses versus single larger dose of intra-articular hyaluronic acid for knee osteoarthritis

Suppan VKL, Tew MM, Wong BC, Chan HK, Chew YW, Tan CS, et al.

J Orthop Surg (Hong Kong), 2020 3 5;28(1):2309499019895029.
PMID: 32129141 DOI: 10.1177/2309499019895029

PURPOSE: A recent 3-month randomized, open-label controlled trial found that the intra-articular hyaluronic acid injection (GO-ON®) given as a single dose of 5 mL is as effective and safe as three repeated doses of 2.5 mL in patients with knee osteoarthritis. However, the information on the long-term efficacy and economic implications of the single-dose regimen is still limited. Hence, this follow-up study was designed to compare the effectiveness and costs of the two regimens 12 months following the treatment.
METHODS: All the 127 patients, who received either three repeated doses (n = 64) or a single dose (n = 63) of GO-ON in the previous trial, were followed up in month 12 following the treatment. The effectiveness of both the regimens was assessed using the Western Ontario and McMaster Universities Osteoarthritis Index (WOMAC), and the mean WOMAC scores were compared with those recorded at the baseline and in month 3. Additionally, the total treatment costs of the two regimens, taking account of both direct and indirect costs, were computed and compared.
RESULTS: A total of 125 patients (98.4%) completed the assessment. Despite the reduction of the overall mean WOMAC score from 39.24 to 19.93 (p < 0.001) in the first 3 months following the treatment with GO-ON, no further changes were observed up to month 12 (p > 0.95). In the meantime, the two regimens did not differ in the mean WOMAC scores (p = 0.749) and in the subscale scores for pain (p = 0.970), stiffness (p = 0.526), and physical functioning (p = 0.667) in month 12. The cost for single-dose injection was found to be approximately 30% lower compared to the repeated doses.
CONCLUSION: These findings indicate that the single larger dose of GO-ON is as effective as the repeated doses over 12 months, and yet the total treatment cost is lowered.