Experimental Rnomics : : Towards The Identification And Characterization Of Non-Protein-Coding Ribonucleic Acids In Pathogenic Agents, Salmonella Typhi

This thesis focused on the experimental identification of small npcRNAs from Salmonella enterica serovar Typhi (S. Typhi), the aetiological agent of typhoid fever. Tesis ini memberikan tumpuan ke atas pengenalpastian npcRNA secara eksperimental daripada bakteria patogenik Salmonella enterica serovar Typhi (S. Typhi), penyebab penyakit demam kepialu

Experimental Rnomics: Towards The Identification And Characterization Of Non-Protein-Coding Ribonucleic Acids In Pathogenic Agent, Salmonella Typhi

RNA bukan-pengkod-protein (npcRNA) merupakan satu kelas pengawalatur-ribo yang bertindak di dalam bentuk kompleks RNA-protein (sebagai RNPs) didalam pelbagai laluan pengawalaturan. Tesis ini memberikan tumpuan ke atas_ pengenalpastian npcRNA secara eksperimental daripada bakteria patogenik Salmonella enterica serovar Typhi (S. Typhi), penyebab penyakit demam kepialu. Melalui pendekatan RNomiks Eksperimental, 82 calon novel npcRNAdaripada perpustakaan cDNA S. Typhi telah dikenalpasti dan dicirikan. Daripada jumlah ini, 28 telah ditranskrip daripada IGR, 29 ditranskrip di dalam arah antisense kepada ORF dan 18 dikenalpasti bertindihan dengan ORF. Sementara 7 calon yang lain telah ditranskIlp daripada kawasan repititif dan beberapa kedudukan bukan repitatif yang lain. Sebelas npcRNA merupakan npcRNAs yang telahpun dilaporkan. Non-protein-coding RNA (npcRNA) is a large class of riboregulators that act in complex with proteins (as RNPs) in diverse regulatory pathways. This thesis focused on the experimental identification of small npcRNAs from Salmonella enterica serovar Typhi (s. Typhi), the aetiological agent of typhoid fever. By an Experimental RNomics approach, 82 species of uncharacterized novel npcRNA candidates were identified from library generated from different growth phases of a clinically isolated S. Typhi. From this, 28 were transcribed from the IGRs, 29 were transcribed in the antisense orientation of the ORFs and 18 were identified to overlap the ORFs. Another 7 candidates were transcribed from repetitive regions and several non-repetitive locations. Eleven known npcRNAs were also detected

Occurrence of intestinal parasitic contamination in selected consumed local raw vegetables and fruits in Kuantan, Pahang

Intestinal parasitic infections are one of the most common human diseases that cause serious health and economic issues in many developing and developed countries. Raw vegetables and fruits play an important role in transmitting parasites into humans. Hence, the aim of this study is to investigate the parasitological contamination of the commonly consumed selected local leafy vegetables and fruits in Kuantan, Malaysia. One kilogram of local consumed-raw vegetables and fruits were collected randomly from Kuantan wet market (Pasar Tani) during monsoon seasons (November 2014-January 2014) and dry seasons (February 2015- April 2015). Standard wet mount procedure and modified Ziehl-Neelsen stain were used for the detection of parasites. In the present study, the examination of vegetables revealed five different species of parasites. The vegetables samples collected from Kuantan’s wet market were positive for both helminths and protozoa. However, the fruits samples were negative for parasitic contamination. Pegaga was the most contaminated leafy vegetables in this study and Strongyloides was the most frequently found parasite. Furthermore, there was a high diversity in the type of observed parasites during the dry season as compared to monsoon season. Therefore, further action should be taken to reduce the occurrence of parasitic contamination in vegetables by implementing the principles of good agriculture practices and improving the water treatment efficacy. Jangkitan parasit usus adalah salah satu penyakit manusia yang lazimnya menyebabkan masalah kesihatan yang serius dan mengakibatkan isu-isu ekonomi di negara-negara maju dan membangun. Sayur-sayuran mentah dan buah-buahan memainkan peranan yang penting sebagai medium jangkitan parasit ke dalam manusia. Oleh itu, tujuan kajian ini adalah untuk menyiasat pencemaran parasitologi daripada sayur-sayuran berdaun dan buah-buahan yang biasa dimakan serta dipilih penduduk tempatan di Kuantan, Malaysia. Sebanyak satu kilogram sayur-sayuran tempatan yang dimakan mentah dan buah-buahan telah dikumpulkan secara rawak dari Kuantan pasar basah (Pasar Tani) semasa musim tengkujuh (November 2014-Januari 2014) dan musim kering (Februari 2015- April 2015). Prosedur lazim ‘wet mount’ dan dimodifikasi perwarnaan Ziehl-Neelsen telah digunakan untuk mengesan parasit. Dalam kajian ini, pemeriksaan sayur-sayuran mendedahkan lima spesies parasite yang berbeza. Sampel sayur-sayuran sampel yang diambil daripada pasar basah Kuantan adalah positif untuk kedua-dua helmin dan protozoa. Walau bagaimanapun, sampel buah-buahan adalah negatif untuk pencemaran parasit. Pegaga merupakan sayuran yang paling tercemar dalam kajian ini dan Strongyloides adalah parasit yang paling kerap ditemui. Tambahan pula, terdapat kepelbagaian yang tinggi dalam jenis parasit telah diperhatikan semasa musim kering berbanding musim tengkujuh. Oleh itu, tindakan lanjut perlu diambil untuk mengurangkan berlakunya pencemaran parasit didalam sayur-sayuran dengan melaksanakan prinsip-prinsip amalan pertanian yang baik serta meningkatkan keberkesanan rawatan air perlu dipertingkat

Short Report: Differences In Dihydrofolate Reductase But Not Dihydropteroate Synthase Alleles In Plasmodium Falciparum Isolates From Geographically Distinct Areas In Malaysia

Dihydropteroate synthase (dhps) and dihydrofolate reductase (dhfr) alleles were typed in 67 Malaysian Plasmodium falciparum isolates. The isolates were collected from two geographically distinct locations: 51 from Sabah, Malaysian Borneo, where sulfadoxine/pyrimethamine (SDX/PYR) is used to treat uncomplicated malaria and 16 from Peninsular Malaysia where in vivo resistance to SDX/PYR has been reported. A total of seven dhps alleles were identified with no significant difference in allele frequency between the 2 populations. Two of the dhps alleles described here have not been previously reported. Four dhfr alleles were detected in 67 P. falciparum isolates. Eightyseven percent of the isolates from the Peninsula, where clinical SDX/PYR failure has been reported, had dhfr alleles with triple point mutations while all of the isolates from Sabah had dhfr alleles with 2 or less point mutations. The difference in dhfr allele frequency between the two populations was highly significant. There was no correlation between in vitro PYR response and accumulation of dhfr point mutations

Experimental identification and characterization of 97 novel npcRNA candidates in Salmonella enterica serovar Typhi

We experimentally identified and characterized 97 novel, non-protein-coding RNA candidates (npcRNAs) from the human pathogen Salmonella enterica serovar Typhi (hereafter referred to as S. typhi). Three were specific to S. typhi, 22 were restricted to Salmonella species and 33 were differentially expressed during S. typhi growth. We also identified Salmonella Pathogenicity Island-derived npcRNAs that might be involved in regulatory mechanisms of virulence, antibiotic resistance and pathogenic specificity of S. typhi. An in-depth characterization of S. typhi StyR-3 npcRNA showed that it specifically interacts with RamR, the transcriptional repressor of the ramA gene, which is involved in the multidrug resistance (MDR) of Salmonella. StyR-3 interfered with RamR–DNA binding activity and thus potentially plays a role in regulating ramA gene expression, resulting in the MDR phenotype. Our study also revealed a large number of cis-encoded antisense npcRNA candidates, supporting previous observations of global sense–antisense regulatory networks in bacteria. Finally, at least six of the npcRNA candidates interacted with the S. typhi Hfq protein, supporting an important role of Hfq in npcRNA networks. This study points to novel functional npcRNA candidates potentially involved in various regulatory roles including the pathogenicity of S. typhi

An evaluation of a recombinant multiepitope based antigen for detection of Toxoplasma gondii specific antibodies

Abstract Background The inefficiency of the current tachyzoite antigen-based serological assays for the serodiagnosis of Toxoplasma gondii infection mandates the need for acquirement of reliable and standard diagnostic reagents. Recently, epitope-based antigens have emerged as an alternative diagnostic marker for the achievement of highly sensitive and specific capture antigens. In this study, the diagnostic utility of a recombinant multiepitope antigen (USM.TOXO1) for the serodiagnosis of human toxoplasmosis was evaluated. Methods An indirect enzyme-linked immunosorbent assay (ELISA) was developed to evaluate the usefulness of USM.TOXO1 antigen for the detection of IgG antibodies against Toxoplasma gondii in human sera. Whereas the reactivity of the developed antigen against IgM antibody was evaluated by western blot and Dot enzyme immunoassay (dot-EIA) analysis. Results The diagnostic performance of the new antigens in IgG ELISA was achieved at the maximum values of 85.43% and 81.25% for diagnostic sensitivity and specificity respectively. The USM.TOXO1 was also proven to be reactive with anti- T. gondii IgM antibody. Conclusions This finding makes the USM.TOXO1 antigen an attractive candidate for improving the toxoplasmosis serodiagnosis and demonstrates that multiepitope antigens could be a potential and promising diagnostic marker for the development of high sensitive and accurate assays

Immunogenicity of Multi-epitope Vaccine Candidate against Toxoplasma gondii Infection in BALB/c Mice

Background: Toxoplasma gondii is a widely prevalent intracellular protozoan parasite which causes serious clinical and veterinary problems. Development of an effective vaccine for controlling toxoplasmosis is an extremely important aim. In the present study, the protective efficacy of recombinant multiepitope antigen (USM.TOXO1) expressing nine potential epitopes identified from SAG1, GRA2, and GRA7 of Toxoplasma gondii was evaluated in BALB/c mice. Methods: Mice were immunized subcutaneously with three doses of USM.TOXO1 antigen (10 µg/ml). Following the immunization, the IgG antibody, IgG subclass, IFN-γ and IL-4 production were evaluated using ELISA, the study was conducted at Animal Research and Service Center (ARASC), USM Health Campus in 2016. Results: Mice immunized with USM.TOXO1 significantly induced a mixed Th1/Th2 response polarized toward the IgG1 antibody isotype. While the cytokine analysis revealed a significant release of IFN-γ cytokines. Conclusion: USM.TOXO1 is a potential vaccine candidate that elicits strong immunity in BALB/c mice. The proven immunogenicity of the generated antigen can serve as a premise for further use of epitope-based vaccine in the immunoprevention of human and animal toxoplasmosis

Antimicrobial effect of Baccaurea angulata fruit extracts against human pathogenic microorganisms

The research application for drugs and food supplements derived from plants extracts have increased in recent years. Plants extract and their constituents are recognized to be safe, either because of their traditional use without any documented detrimental impact or because of dedicated toxicological studies. The potential of higher plants as a source for new drugs is largely unexplored. Thus, a systematic investigation was undertaken to screen for antibacterial activity from Baccaurea angulata. Plant that belongs to the family is used as food as well as treatment of infectious diseases such as diarrhoea, skin infections and gonorrhoea. The anti-microbial activity of the B. angulata fruit extracts have revealed different antimicrobial properties, that vary between three parts (whole fruit, fruit skin, and berry), three solvents (methanol, ethanol and aqueous), using agar well diffusion, and microdilution methods and different pathogens (Streptococcus pneumoniae, Staphylococcus epidermidis, Klebsiella pneumoniae and Pseudomonas aeruginosa). The highest observed antimicrobial activity was in ethanol extract of fruit skin using agar well diffusion against S. pneumoniae. Among tested Gram negative bacteria, K. pneumoniae was the most susceptible bacterium which showed the highest bacteriostatic and bactericidal activity using microdilution method. All parts of fruit extracts poses the antimicrobial activities against all human pathogenic microorganisms used in this study