Molecular characterization of cholera outbreak isolates in South Africa, 2008-2009

A thesis submitted to the Faculty of Health Science, University of the Witwatersrand, Johannesburg, South Africa, in fulfilment of the requirements for the degree, Doctorate of Philosophy in Medicine, 2009-2015Cholera is an acute diarrhoeal disease that generally presents as abrupt watery diarrhoea and vomiting. For the years 2008 to 2009, South Africa experienced two major outbreaks of cholera. The first outbreak was reported from May to July 2008 (Chapter Three) and the second outbreak from November 2008 to April 2009 (Chapter Four). Within both events, Vibrio cholerae (V. cholerae) O1 identified at peripheral laboratories displayed resistance to three or more routinely tested antimicrobial agents. The molecular epidemiology and mechanism of antimicrobial resistance of V. cholerae O1 isolates was investigated. This was achieved by using various molecular techniques, which included pulsed-field gel electrophoresis (PFGE) analysis, polymerase chain reaction (PCR), nucleotide sequencing, identification of plasmid DNA and Southern blot hybridization analysis. Methods As part of routine characterization of V. cholerae isolates at the Centre for Enteric Diseases (CED), isolates underwent serological and biochemical confirmatory identification as well as antimicrobial susceptibility testing using the Etest method. PFGE analysis was performed on V. cholerae O1 isolates digested with NotI restriction enzyme. One-hundred V. cholerae O1 isolates, ten isolates characterized in Chapter Three and 90 isolates characterized in Chapter Four were selected for further analysis to ensure that all PFGE banding patterns were represented. Three probable mechanisms of antimicrobial resistance were investigated. Firstly, PCR was used to detect for the presence of class 1 integrons (3’-CS and 5’-CS), class 2 integrons (intI2), plasmid-mediated quinolone resistance (PMQR) genes (qnrA, qnrB, qnrS, qnrC and qepA), quinolone resistance determinant (qnrVC3), ESBL producing genes (blaTEM, blaSHV and blaCTX-M), genes coding for the quinolone resistance-determining region (QRDR) of DNA gyrase (gyrA/gyrB) and topoisomerase IV (parC/parE), SXT element-integrase gene (SXTint) and associated SXT resistance genes (floR, sul2, dfrA1, dfr18, strA and strB) and the class A tetracycline resistance determinant (tetA). The presence of resistance plasmids was investigated by isolation of intact bacterial plasmid DNA. Southern blotting and DNA probing was used to investigate the location of resistance genes on the plasmids. Secondly, nucleotide sequencing was used to detect amino acid mutations in the QRDR of DNA gyrase and topoisomerase IV respectively. Thirdly, to determine the role of an active efflux pump in quinolone resistance, susceptibility testing to nalidixic acid was investigated in ten V. cholerae O1 isolates characterised in Chapter Three using agar dilution in the presence and absence of two efflux pump inhibitors, reserpine and phenylalanyl arginine- -naphthylamide. PCR analysis was used to detect for virulence determinants, which included the enzymatic A subunit of the cholera toxin (CT), ctxA and the gene encoding for the toxin co-regulated pilus (TCP), tcpA respectively. In addition, the complete coding region of the ctxAB gene was amplified and sequenced from four V. cholerae O1 isolates, two isolates characterized in Chapter Three and two isolates characterized in Chapter Four as several V. cholerae O1 atypical El Tor isolates have been described in Africa. Minimum inhibitory concentration (MIC) values for azithromycin were determined for all 100 V. cholerae O1 isolates using both the Etest and agar dilution methods (Chapter Five). PCR-analysis was used to determine the presence of seven macrolide resistance determinants (mefA, ereA, ereB, ermB, mphA, mphB and mphD) in all 100 V. cholerae O1 isolates. Results For both cholera outbreaks, a total of 751 isolates were received and available for analysis. All 31 isolates recovered from the first outbreak (Chapter Three) were characterized as V. cholerae O1 serotype Ogawa. For the second outbreak (Chapter Four) 708 isolates were characterized as serotype Ogawa, while the remaining 12 isolates were characterized as serotype Inaba. All isolates analyzed from both outbreaks were susceptible to ciprofloxacin and imipenem, but resistant to six or more antimicrobial agents tested for surveillance purposes. All V. cholerae O1 isolates were shown to be resistant to nalidixic acid, cotrimoxazole, trimethoprim, sulfamethoxazoleand streptomycin. Extended-spectrum - lactamase (ESBL) activity was observed in V. cholerae O1 isolates (MIC 64 μg/ml) from both outbreaks. In the second outbreak reduced susceptibility to ampicillin, tetracycline, kanamycin, chloramphenicol, erythromycin and furazolidone were observed. Dendrogram analysis produced two main PFGE clusters. PFGE fingerprint patterns from V. cholerae O1 isolates recovered from the first outbreak clustered away from V. cholerae O1 isolates recovered from the second outbreak (data not shown in this study). Class 1 integrons, class 2 integrons and PMQR genes were not detected by PCR.All 100 V. cholerae O1 isolates were PCR-positive for the SXTint gene and five of the six associated SXT resistance genes encoding for chloramphenicol (floR), sulfamethoxazole (sul2), trimethoprim (dfrA1) and streptomycin (strA and strB). Seventeen V. cholerae O1 isolates (ten isolates characterized in Chapter Three and seven isolates characterized in Chapter Four) were PCR-positive for the tetA resistance determinant. Nucleotide sequencing of the QRDR, showed that all nalidixic acid-resistant isolates harboured the same mutations in GyrA (S83-I) and ParC (S85-L) but none were observed in GyrB and ParE.There was no involvement of an active efflux pump in quinolone resistance in ten isolates characterised in Chapter Three. Sixteen V. cholerae O1 isolates (ten isolates characterized in Chapter Three and six isolates characterized in Chapter Four) harboured a single plasmid of approximately 140 kilobase pairs in size and showed to harbour the blaTEM gene, which produced the TEM-63 -lactamase.PCR analysis showed that all 100 V. cholerae O1 isolates were positive for the CT, and all were PCR-positive for the El Tor variant of the TCP. Nucleotide sequencing of the ctxAB gene of the four selected isolates showed that all four isolates expressed the encoded ctxB allele for the CT of the classical biotype and were defined as “altered El Tor”. A mobilome is characterized by related genome sequences that differ by combinations of genomic islands, prophages and integrative conjugative elements. All four isolates contained an identical mobilome profile pattern, profile B. Comparative analysis using both the Etest and agar dilution methods (Chapter Five) showed that all V. cholerae O1 isolates were susceptible to azithromycin provided that the tentative breakpoint of 16μg/ml is applied. All 100 isolates were PCR-negative for all seven macrolide resistance determinants, which are commonly associated in the family Enterobacteriaceae respectively. Conclusion This is the first incidence of TEM-63 -lactamase-producing, antimicrobial-resistant, toxigenic V. cholerae O1 altered El Tor isolates in South Africa. This study highlights the need to further analyze antimicrobial resistance and track emerging epidemic isolates of V cholerae O1. The MIC values and PCR results reported in this study for azithromycin provides a foundation for the surveillance of azithromycin susceptibility and to determine MIC breakpoints in V. cholerae O1 isolates circulating in South Africa

Shift & night work – the positive impact, challenges and action to deal with it

Shift work means work that takes place on a schedule of employee outside the normal hours eight at morning until at five at evening. Well, it involves day and night shifts, which is whether in an early morning shifts, and rotating shifts. In Malaysia, shift work happens usually to the health care workers, precision production and repair workers, machines operator, and aviation workers and so on. Especially in manufacturing, employees tend to make sure machines run twenty-four hours per day rather than buying more machines to operate on a day shift

Annual leave: the case of Taiwan and Malaysia

The definition of annual leave is allowing the organization’s employees to leave the number of hours or days of work within a year without any consequences. Depending on the employer’s policy, different working days may be provided and employees may be required to issue a certain amount of advance notice in advance, which may have to be coordinated with the employer to ensure adequate staffing during the absence of the employee and other requirements may have to be obtained satisfy. Today, most countries have a minimum paid annual leave in accordance with the law. This time off is paid by the company and employees can ask for time for any reason they wish to take the annual leave. Annual leave allows employees to take paid vacations so that they can rest regularly to rest and rejuvenate. Employees on regular vacations are more motivated and perform their work more effectively than those who are not on vacation. Because they have regular breaks, they are less prone to accidents and less likely to be under pressure, which means they can reduce their absence from illness

Fabrication And Performance Characterization Of PET/ITO/ZNO NANOROD/P3HT:PCBM And PET/ITO/P3HT:PCBM Solar Cell

Kerja ini memberi tumpuan kepada fabrikasi dan prestasi pencirian sel suria hibrid (NANOROD ZnO/P3HT:PCBM) dan sel suria organik (P3HT:PCBM). Bagi sel suria hibrid, bahan seperti regioregular 3 - hexylthiophene - 2 , 5 – diyl (P3HT) (bahan penderma), zink oksida (bahan penerima) dan fullerene [6,6] phenyl C61 acid butyric methyl ester (PCBM) (bahan penerima) telah digunakan. Sintesis dengan menggunakan kaedah penyinaran gelombang mikro adalah untuk mendapatkan nanorod ZnO. This work focus on the fabrication and performance characterization of hybrid (ZnO nanorod/P3HT:PCBM) and organic solar cell (P3HT:PCBM). For hybrid solar cell, material such as regioregular 3 - hexylthiophene - 2 , 5 – diyl (P3HT) (donor material), zinc oxide (ZnO) (acceptor material) and fullerene [ 6,6 ] phenyl C61 acid butyric methyl ester (PCBM) (acceptor material) was used

Hubungan Antara Kepimpinan Transformasional Dan Komitmen Organisasi Dalam Kalangan Kakitangan Awam Di Sarawak

The purpose of this study is to identify the relationship between transformational leadership and organizational commitment among civil servants in Sarawak. Data were obtained from 108 employees from a public organization in Sarawak. The data were analysed using descriptive statistics and inferential statistics. Descriptive statistics were reported, followed by reliability analysis, hypothesis testing using Pearson Correlation. Findings show that there is a strong relationship between individual consideration and affective commitment among public organization in Sarawak. Next, there is a moderate relationship between idealized influences, inspirational motivation and intellectual stimulation with affective commitment among public organization in Sarawak. This study contributes to the existing knowledge of civil servants in Sarawak. This study also has improved the knowledge and understanding of an organizational and human resources practitioners on how the leadership of a leader influences employee commitment in Sarawak. In other words, the findings of this study can help an organizations improve their quality to ensure that an organizations can achieve a level of commitment among their employee

Mediating role of psychological contract in the relationship between workplace spirituality and affective commitment

Malaysian banking sector is facing commitment challenge of their employees especially in the current highly competitive business environment. Now banks want to retain and engage their competent employees so there is a need to create or enhance their affective commitment because this affective commitment is a key to retain and engage competent employees. Previous research explains that effective commitment provide employees emotional attachment with their organisation so there is a need to explore those factors through which organisations can develop the affective commitment of their employees. Healthy amount of research has been conducted to deal with employee attitude and commitment. As time goes on new concepts are coming up with new management ideas. One such new area of research is workplace spirituality. Previous research in this field discovered numerous benefits of workplace spirituality to the organization and employee as well as organizational and employee development, and commitment. Literature review pointed out that Malaysian banking sector is facing the problem of employee commitment. Present research is an attempt to deal this problem of employee commitment with the help of workplace spirituality along with psychological contract as a mediator. Subjects for this research were 350 bank employees working in commercial banks in Malaysia. Structural equation modelling SEM – PLS was the main statistical technique utilized in this study. All the main relationships were found to have significant effect on employees affective commitment, Overall, the results indicate that the model provide good understanding of the workplace spirituality‟s influence on employees commitment in banking sector of Malaysia

The high rate of road accidents in Malaysia: What the causative factors are?

Road accidents become one of the crucial social problems over the world. The number of accidental death worldwide became the third largest after HIV/AIDS and TB, approximately with 0.8 million of deaths per year.In Malaysia, road accident is become one of major problem with high rate of road accident occurrences.This paper attempts to review the present status of road accidents in Malaysia and what the causative factors are.This paper uses an overview of previous road accidents statistic in Malaysia, the total numbers of casualties, and data of registered vehicle

Nutritional Status and Factors affecting Food Intake among Hospitalised Patients in Hospital Al-Sultan Abdullah

Hospital food intake can impact patients' nutritional status, resulting in a lengthier hospital stay or a higher mortality rate. This study aimed to investigate the nutritional status and the factors influencing the food intake of patients at Hospital Al-Sultan Abdullah (HASA). Malnutrition risk was assessed using Nutritional Risk Screening 2002, and factors affecting food intake were investigated using a questionnaire. Participants' weight and height were estimated using the anthropometry assessment formula. The body mass index was 25.3 ± 8.1 kg/m2 and 70.6% (113) posed no risk of malnutrition. Food tasting differently (48.8%) was the highest affecting factor in food intake. Keywords: food intake; hospitalised patients; nutritional status; malnutrition eISSN: 2398-4287 © 2023. The Authors. Published for AMER & cE-Bs by e-International Publishing House, Ltd., UK. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/). Peer–review under responsibility of AMER (Association of Malaysian Environment-Behaviour Researchers), and cE-Bs (Centre for Environment-Behaviour Studies), College of Built Environment, Universiti Teknologi MARA, Malaysia DOI: https://doi.org/10.21834/ebpj.v8i24.4674

Pendidikan Agama Islam Di Sekolah Inklusi: Strategi Pembelajaran Bagi Anak Penyandang Tunalaras

Every child has the same right to get a quality education. It also emphasizes that every child with special needs, such as children with special needs, mentally unstable, has physical and psychological deficiencies, is also entitled to a proper education. In this regard, the implementation of education for children with special needs must also be considered. Children with emotional limitations are called Tunalaras. Research related to the learning of mentally retarded children has previously been carried out and has been carried out and it can be seen that the ability of self-control of mentally retarded children can be increased through better behavior improvement. So that this research is intended to find information about the process of receiving education for children with disabilities, especially Islamic religious education. This study wants to know how the process and method of learning Islamic Religious Education in inclusive schools and how Islamic Religious Education learning is carried out to be able to help change behavior for children with mental retardation. The research method used is qualitative with interviews to get as much information as possible in the field

Effects of Different Drying Methods and Hydrocolloids on Quality Properties of Semi-dried Catfish Jerky

This study was carried out to evaluate the effects of different drying methods (sun drying, cabinet drying and convection oven) and hydrocolloids (carrageenan and alginate) onphysicochemical properties of semi-dried catfish jerky. The concentration of hydrocolloids used was 1% and 2%. Samples without the addition of hydrocolloid served as the control group. The water activity of semi-dried catfish jerky decreased with the addition of hydrocolloids. For colour properties, lightness (L*) value of semi-dried catfish jerky increased with the increased concentration of hydrocolloids. The addition of 2% alginate (2%A) and 2% carrageenan (2%C) showed higher lightness (L*) than the controlled group for all drying methods, except for sun drying with carrageenan. Both carrageenan and alginate added into semi-dried catfish jerky increased the processing yields. The addition of 2% carrageenan (2%C) and 1% alginate (1%A) improved the product yields for all drying methods. This paper argues that the application of cabinet dryer gives better shelf stability due to the lower range of water activity than other drying methods while preserving colour quality and product yields