Construction of an Attenuated Pasteurella Multocida B:2 by Mutation in the Gdha Gene

Pasteurella multocida 8:2 is a Gram negative bacteria that has been associated with haemorrhagic septicaemia in cattle and buffaloes in Asia. It has been known to produce endotoxin that leads to haemorrhages and oedema, causing deaths due to either asphyxiation and dyspnoea or septicaemia. Vaccination has been used to control the disease but with little success due to the low vaccination coverage. Therefore, an alternative live vaccine should be considered. In preparing an alternative live vaccine, an attenuated P. multocida 8:2 is created by manipulating one of the housekeeping genes of the bacteria. The selected housekeeping gene, the glutamate dehyrogenase (gdhA) gene, was successfully isolated via PCR from wild type P. multocida 8:2. The gene was then amplified using nested-PCR to determine its functional part. Both PCR products were cloned into plasmid pCR2.1, producing pSZ1 and pSZ2, respectively before being sequenced. The whole sequence of the gene is 1108 bp while the functional part of the gene was 652 bp. The functional part was 99.8% identical to the model sequence, the PM70, which is a model genome sequence of P. multocida serotype A. The pSZ 1 was subsequently digested with a unique restriction enzyme, Munl before the kanamycin cassette, isolated from plasmid pUC4K via PCR, was inserted at the centre of the housekeeping gene. The recombinant was named pSZ1K. After that, the gdhA gene that was disrupted by kanamycin cassette (GK) was isolated from the pSZ1K using restriction enzyme digestion, EcoRI. The suicide plasmid, pAKA19 was also digested with the same enzyme to achieve complimentary ligation sites. After ligation, the achieved recombinant plasmid was called pSZ19GK. All cloning products were transformed into Escherichia coli DH5a. Enroute for disruption of the gene in the host genome, both E. coli and P. multocida 8:2 were subjected to spontaneous mutation towards streptomycin. After conveying the pSZ19GK into P. multocida 8:2 via conjugation, the bacteria was incubated for five days to encourage allelic exchange to occur between disrupted gene and the host chromoso me. Subsequently, PCR of the bacteria genome proved that allelic exchange has occurred and the mutant was called P. multocida 8:2 (GK). In order to verify the characteristic of the non-pathogenic P. multocida 8:2 (GK) mutant, in vitro stability test and in vivo pathogenicity test were done. In in vitro stability test, 14 strains out of the 20 survived only up to 15 days of incubation. This proves that the mutants are unable to sustain life without glutamate supplement and therefore having a short life-span. From there, several strains were picked to be tested in vivo using mouse experimental model. Mice infected intraperitoneally or subcutaneously with different concentrations of the mutant survived throughout the 5-day study period. They were compared to the mice that were infected intraperitoneally or subcutaneously with different concentrations of the wild type organism. None of the mice infected with the mutant died but all mice infected with the wild type did not survived and were dead in less than 24 hours. P. multocida 8:2 were successfully isolated from organs of mice infected with both wild-type and mutant. This confirmed that the mutant, P. multocida 8:2 (GK) became attenuated by the disruption of the gdhA gene and has a good potential to be used as an alternative live vaccine for HS

Managing Flood in the Context of Education, Research and Partnership in Malaysia

Several structural and non-structural measures and approaches have been implemented by the Malaysian government in order to overcome and alleviate the flood disaster. To date, Education, Partnership and Research have been identified as crucial components in forming an effective Flood Management system in Malaysia. This argument has been supported by many public and private institutions worldwide. As such, the standards and plans can be developed and implemented at district, state and federal levels. The spirits of Hyogo and Sendai were used as the guiding principles. Semi-structured interviews with several responding agencies were conducted for data collection. The findings indicated that less emphasis was given to the roles of components such as Education, Partnership and Research in Flood Management. The aim of this paper is to propose an integrated system for storing, disseminating and analyzing information pertaining to Education, Partnership and Research

Comparison of analgesic efficacy between preemptive intravenous paracetamol and single shot caudal block in paediatric inguinal hernia repair

Terdapat pelbagai kaedah untuk mencapai tahap keberkesanan analgesik yang mencukupi selepas pembedahan angin pasang di kalangan kanak-kanak. Pembiusan setempat ‘caudal block’ biasanya digemari dalam pembedahan bahagian bawah abdomen melibatkan kanak-kanak. Sebaliknya, paracetamol yang diberi melalui vena telah digunakan secara meluas kerana tahap keselamatan dan keberkesanan analgesiknya yang sangat baik. Kajian ini bertujuan untuk membandingkan tahap keberkesanan analgesic paracetamol yang diberi melalui vena dengan pembiusan setempat ‘caudal block’ selepas pembedahan angina pasang di kalangan kanak-kanak. Ini adalah satu kajian prospektif rawak melibatkan 40 orang kanak-kanak yang menjalani pembedahan elektif sebelah angin pasang di bawah pembiusan am di Hospital Universiti Sains Malaysia. Pesakit yang telah mendapat keizinan dari penjaga mereka dibahagi secara rawak kepada dua kumpulam yang menerima paracetamol melalui vena (kumpulan A) dan pembiusan setempat ‘caudal block’ (kumpulan B). Skor kesakitan diukur menggunakan skala FLACC pada 10 minit, 30 minit, 1 jam, 2 jam dan 6 jam selepas pembedahan dan dibandingkan. Keputusan lain yang diukur and dibandingkan adalah purata tekanan darah dan denyutan nadi semasa pembedahan, purata masa pertama untuk permintaan ubat tahan sakit, purata jumlah fentaniyl yang digunakan semasa tempoh pembedahan dan kadar kesan sampingan antara kedua-dua kumpulan Purata umur pesakit adalah 30.48 bulan. Pesakit dalam kumpulan B mendapat keputusan skor kesakitan yang lebih rendah pada 10 minit (p=0.018), 30 minit (p=0.013) dan 1 jam (p=0.05) dibandingkan dengan kumpulan A. Pada 2 jam dan 6 jam, tiada perbezaan yang signifikan direkodkan. Tiada ketidakstabilan hemodinamik direkodkan. Tiada perbezaan signifikan dalam masa pertama untuk permintaan ubat tahan sakit (p=0.079) dan juga jumlah total penggunaan fentanyl (p=0.090) semasa tempoh pembedahan. Tiada komplikasi besar yang signifikan diperhatikan pada kedua-dua kumpulan. Pembiusan setempat ‘caudal block’ merupakan pilihan yang lebih baik daripada paracetamol yang diberi melalui vena pada jam pertama selepas pembedahan namun tidak memberi perubahan yang signifikan selepas tempoh tersebut. Keberkesanan analgesik paracetamol yang diberi melaui vena adalah sama dengan pembiusan setempat ‘caudal block’ jika dikira dari perspektif masa pertama permintaan ubat tahan sakit dan juga jumlah penggunaan fentanyl

A study on unsafe act, unsafe condition and communication barrier for fall from height accident in construction industry

Malaysian construction industry growth is expected to fluctuate since it was announced by the Finance Minister as part of the infrastructure project for Malaysian Budget 2020. In Malaysia, construction industry is one of the sectors which contribute to fall from height accidents. It is one of the important issues that need to be addressed by every organization in order to understand the impacts on the organizations including the workers. Currently, fall from height accidents is a challenge that many organizations struggle to overcome. Thus, to gain better understanding, this study identifies the factors that influence fall from height accidents. The main purpose of this study is to investigate the safety problem when working at height in construction site, to identify the factors that cause fall from height accidents and to analyze the factors that influence the accident and the prevention. For data collection, 264 set of questionnaires were distributed to 12 different construction sites with two (2) target levels which is supervisors and site workers. The data collected were analyze using SPSS Version 26 and the result gain through analysis of Pearson correlation and Multiple regression indicated that unsafe act, unsafe condition and communication barrier have positive relationship with fall from height accident. These finding provides useful information to the organization regarding their employees’ well-being. Besides that, it helps to enhance the underpinning theory in this study which is Accident Causation Model

In vitro treatment of lipopolysaccharide increases invasion of Pasteurella multocida serotype B:2 into bovine aortic endothelial cells

Pasteurella multocida serotype B:2 causes hemorrhagic septicemia in cattle and buffalo. The invasion mechanism of the bacterium when invading the bloodstream is unclear. This study aimed to characterize the effects of immunomodulatory molecules, namely dexamethasone and lipopolysaccharide, on the invasion efficiency of P. multocida serotype B:2 toward bovine aortic endothelial cells (BAECs) and the involvement of actin microfilaments in the invasion mechanism. The results imply that treatment of BAECs with lipopolysaccharide at 100 ng/mL for 24 h significantly increases the intracellular bacteria number per cell (p < 0.01) compared with those in untreated and dexamethasone-treated cells. The lipopolysaccharide-treated cells showed a significant decrease in F-actin expression and an increase in G-actin expression (p < 0.001), indicating actin depolymerization of BAECs. However, no significant differences were detected in the invasion efficiency and actin filament reorganization between the dexamethasone-treated and untreated cells. Transmission electron microscopy showed that P. multocida B:2 resided in a vacuolar compartment of dexamethasone-treated and untreated cells, whereas the bacteria resided in cellular membrane of lipopolysaccharide-treated cells. The results suggest that lipopolysaccharide destabilizes the actin filaments of BAECs, which could facilitate the invasion of P. multocida B:2 into BAECs