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Articles by Mariana N. Shamsudin
Total Records ( 10 ) for Mariana N. Shamsudin
  Nagi A. AL-Haj , Mariana N. Shamsudin , Raha A. Rahim and Zamri Ishak
  Although, PCR methods aimed on the detection of genes associated with the pathogenicity of Escherichia coli have been reported, tests allowing the direct identification of this serotype are rare. In this study the Random Amplified Polymorphic DNA (RAPD) fingerprinting technique allowed genetic diversity assessment of 25 E. coli isolates of various sources. A highly significant finding from the DNA fingerprinting is the display of a predominant band at a size of 308 bp when arbitrary OPAE-10 primer was used. After sequencing this fragment primer called secD was designed to be used as PCR primer. secD primer pairs was highly specific to detect all isolates including E. coli O157: H7.
  Nagi A. Al-Haj , Mariana N. Shamsudin , Raha A. Rahim and Zamri Ishak
  Quantitation assay of Escherichia coli, Samonell sp. and Vibrio cholerae cells investigated by exploiting the component consistently present on the outer surface of Gram-negative bacteria. In this study, a simple marine biolysate-based method for simultaneous detection of the gram negative pathogenic bacteria based on lipopolysaccharide component was optimized. The detection technique focused on the surface of these bacterial species, which is covered by polysaccharides and has high affinity to marine biolysate. E. coli, Samonell sp. and V. cholerae with similar initial cell count per mL have different but consistent absorbance readings by using the spectrophotometer and turbidity meter. This revealed that different genera of Gram Negative bacteria can be directly differentiated through standard curve that is plotted from the carbohydrate and marine biolysate assays absorbance readings. Both the assays elucidated a qualitative and quantitative detection of the pure culture pathogens.
  Nagi A. Al-Haj , Mariana N. Shamsudin , Raha A. Rahim and Zamri Ishak
  Recent advances in molecular techniques have revolutionized the detection of microorganism. The development of a molecular-based technique for detection of the three different targets of Enterbacteriacae was undertaken. Primer and probe were designed based on specific pepted of novel hemolymph protein of horseshoe crabs (Factor C anti-LPS) Tachypleus tridentatus that is believed to be involved in the binds to the lipopolysaccharide of Escherichia coli, Salmonella and Vibrio cholerae. The aim of our study the exploit part of cell wall polysaccharide in the development of improved detection method based on molecular approaches. In the gene detection assay, Lipopolysaccharide gene of Salmonella, V. cholera and E. coil were hybridized to anti-LPS factor gene found in the biolysate of the marine animals. The wzm and wzt genes encoding O-polysaccharide genes were amplified in these pathogens and the LPS factor C were amplified from the marine lysate. Development of a PCR-based technique for detection of the food-borne pathogens particularly Sa Salmonella, V. cholera and E. coil were achieved. Thus rapid, sensitive and reliable techniques for the detection of food-borne pathogens developed.
  E. Amghalia , Nagi AL-Haj , Mariana N. Shamsudin , V. Neela and W.M.Z. Somarny
  Staphylococcus aureus is among the most prominent pathogen in both community acquired and nosocomial infection. The epidemiology analysis of Staphylococcus aureus isolates will be required to ascertain the incidence, prevalence and diversity of strains. To investigate the epidemiological of S. aureus in Malaysia, a highly reliable typing method-Randomly amplified polymorphic DNA was applied to 50 S. aureus isolates obtained from 3 different hospitals in Malaysia namely Hospital Tunku Ampuan Afzan Kuantan, Hospital Besar Seremban and Hospital Miri Malaysia. The results obtained from this study showed that the isolates can be clustered into 8 different clones. All members of the respective clones are of the same origin. In addition, there were 2 clonal grouping of isolates for each hospital. However, the clonal groupings are not in accordance to the geographical distribution. To understand the epidemiology of these isolates in depth it is very important to have information about the patient’s history. The Nei and Li’s genetic distances obtained from this study ranged from 0.0803922-0.11111. Two genetic markers a band of size 500 bp when amplified with primer OPAE-14 and another marker band of size 750 bp amplified with primer OPAE-15 was identified and this band can be used as diagnostic marker for the rapid identification of S. aureus. Apart from the genetic markers, an epidemiological marker of 1200 bp was also identified for the Miri isolates. This marker can be used as the epidemiological marker for the identification of the isolates from Miri in the future outbreaks. From this study, RAPD has proved to be an useful aid to epidemiological investigations of S. aureus.
  E. Amghalia , Nagi A. AL-Haj , Mariana N. Shamsudin , Son Radu , Rozita Rosli , V. Neela and Raha A. Rahim
  Multiple drug resistant Staphylococcus aureus is one the most common nosocomial pathogen worldwide. The timely identification of this hospital acquired pathogen and detection of the various antibiotic resistant genes harbored is one of the most important function of the microbiology laboratory. In this study, we report the development of a multiplex PCR system for the diagnosis of S. aureus and the detection of clinically relevant antibiotic resistance genes harbored by some isolates. This system was designed to identify S. aureus at species level and to detect methicillin, gentamycin, erythromycin, vancomycin and mupirocin resistant genes, respectively from a single colony in a single tube reaction. All isolates amplified a 108 bp fragment (conserved in S. aureus) confirming the identity of S. aureus, 23 isolates produced a band at the position of 533 bp, 28 isolates at 139 bp and 30 isolates at 174 bp evidencing the presence of mecA (methicillin or oxacillin resistance), ermA (erythromycin resistance), aac (6`)-aph (2``) (gentamycin resistance) genes. None of the isolates amplified van A (vancomycin resistance) and ileS-2 (mupirocin resistance) genes showing the absence of their resistance in the isolates studied. These genotypic results when compared with classical antibiotic susceptibility tests showed less correlation. Overall, we found a correlation between phenotypic and genotypic methods of 60% for methicillin, 36.7% for gentamycin, 43.3% for erythromycin, 100% for vancomycin and mupirocin. This suggests that classical antibiotic sensitivity test is not accurate, but need to be supplemented with other methods to be applied in a clinical laboratory. The system developed in this study offers a rapid, simple specific and accurate detection of multiple antibiotic resistant genes in clinical S. aureus isolates and thus could be systematically applied as a diagnostic test in clinical microbiology laboratories, facilitating the design and use of antibiotic therapy.
  E. Amghalia , Nagi A. AL-Haj , Mariana N. Shamsudin , Nurmas I. Mashan , V. Neela and Zamberi Sekawi
  Methicilin Resistant Staphylococcus aureus (MRSA) implicated in many post-surgical and cancer treatment as well hospital and community fatalities need to be treated with an effective alternative antimicrobial agent. In the search for anti-MRSA agent, 2 types of natural products were investigated for inhibitory activity against MRSA. In addition to the bioassay, the activity of the anti-MRSA agent was elucidated based on the effect of both natural products on nucleotide changes of chromosome-encoded genes. In this study, the methanol extract of the red marine algae and the natural pure honey were studied for its antibacterial property based on disc diffusion test and Minimum Inhibitory Concentration (MIC). The effects of both natural products on selected gene sequences of S. aureus’s were determined by RT-PCR analysis. The genes of interest, which have been chosen in this study, are genes that are involved in the antibacterial mechanism including inhibition of cell wall synthesis, protein synthesis and nucleic acid synthesis. Five genes of interest chosen in this study include mecA gene, mecR1 gene, mecI gene, adaB gene and sav1017 gene. The results for antibacterial property showed the methanol extract of a red seaweed and the pure honey, inhibited growth of S. aureus strain according to the inhibition zones around discs saturated with the seaweed extract and pure honey, respectively. The MIC test showed decrease in growth of MRSA isolates after growing in broth incorporated with the extract and honey, respectively. The effect of the inhibitory activity of the natural products on selected gene sequences showed that several nucleotide changes occurred in the sequences of certain genes of interest based on the gene sequences of the cDNA after RT-PCR was carried out on the mRNA of S. aureus treated with the natural products. This research underlined that the inhibition effect of the natural products may be chromosome mediated evidenced by the changes of chromosome-encoded genes. The significant findings on activities of the seaweed extract and pure honey may become very useful in the process to find a better treatment for S. aureus infection especially, for the multiple drug resistant isolates. In addition, it is also, a new finding for natural product discovery through gene-expression analysis.
  Nurul H. Idris , Nagi A. Al-Haj , Mariana N. Shamsudin and Raha A. Rahim
  Safe attenuation has been done on marine pathogen Vibrio alginolyticus using naturally acidified fructose against vibriosis. Attenuation was confirmed by injecting the attenuated bacterium into fish where the survival rate was 100% compared to 50% survival in fish injected with non-attenuated bacteria. The attenuated bacterium was then evaluated for oral vaccination of Lates calcarifer (Asian seabass). Fish were fed with fish pellet incorporated with attenuated and non-attenuated bacterium of V. alginolyticus for 30 days. They were measured for serum antibody production by conventional agglutination titer and also monitored for the fish weight gain to observe the health improvement. Vaccinated fish showed comparable increased in weight gain, 90% survival after challenge and significantly high antibody titer compared to other treatment and control.
  Nagi A. Al-Haj , Nurmas I. Mashan , Mariana N. Shamsudin , Habsah Mohamad , Charles S. Vairappan and Zamberi Sekawi
  The in vitro antibacterial activities of seaweed belong to Euchema denticulatum extract showed inhibitory activity only on gram positive organisms tested including Staphylococcus aureus and Streptococcus pyogenes, which were expressed in terms of minimum inhibitory concentration and minimum bactericidal concentration test. Thus, gram negative pathogens tested including Escherichia coli, Klebsiella pneumoniae and Pseudomonas aeruginosa showed resistant phenotypic pattern to both extracts. Results of the present study confirmed the potential use of seaweed extract as a source of antibacterial compounds.
  Nagi A. AL-Haj , Nurmas I. Mashan , Mariana N. Shamsudin , Habsah Mohamad , Charles S. Vairappan and Zamberi Sekawi
  Gram-positive bacteria are more sensitive to antibiotics than gram-negative bacilli because of the lack of outer membrane which prevent easy access of the drug into the bacterial cells. However, there are many gram-positive organisms with natural, intrinsic resistance to antimicrobials. In addition, these bacteria are able to acquire resistance to frequently used drugs rapidly through selective pressure of the environment and also via the genetic evolution of bacteria. The resistance of those bacteria to antibacterial agents is mediated by antibiotic resistant genes. Therefore, the current study was designed to explore the effect of seaweed extracts on several antibiotic resistant genes in Staphylococcus aureus mec genes mecA, mecR1 and mecI that regulate the expression of methicillin resistance was investigated by PCR.
  Nagi A. AL-Haj , Lai L. Suang , Mariana N. Shamsudin , Rasedee Abdullah , Rahmah Mohamed and Zamberi Sekawi
  Total of 23 putative Open Read Farms from B. pseudomallei strain D286 was successfully cloned and the nucleotide sequence analysis of the putative genes showed the homologue (98-100%) to strain K96243. The high similarity in gene sequences between these strains is confirmed for presence of the necessary ORF for LPS biosynthesis through PCR amplification the application of the ORFs in the PCR amplification and expression method. The findings of this study have contributed to some information on the molecular bases of the LPS biosynthesis genes in B. pseudomallei specifically for strain D286. PCR amplification, a specific pair of primer for each ORFs was proving specific for amplification of genes in B. pseudomallei strain D286. The PCR mixture with addition of DMSO, formamide and glycerol could ease the PCR optimization where different pairs of primers were involved. The specific primer pairs with the PCR mixture could be used in developing a PCR diagnosis of melioidosis.
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