Publications by authors named "Khatijah Yusoff"

103 Publications

Bioactive Compounds from Marine Sponges: Fundamentals and Applications.

Mar Drugs 2021 Apr 27;19(5). Epub 2021 Apr 27.

Aquatic Animal Health and Therapeutics Laboratory, Institute of Bioscience, Universiti Putra Malaysia, Serdang 43400, Malaysia.

Marine sponges are sessile invertebrates that can be found in temperate, polar and tropical regions. They are known to be major contributors of bioactive compounds, which are discovered in and extracted from the marine environment. The compounds extracted from these sponges are known to exhibit various bioactivities, such as antimicrobial, antitumor and general cytotoxicity. For example, various compounds isolated from have showcased various bioactivities, such as those that are antibacterial, antiviral and antifungal. In this review, we discuss bioactive compounds that have been identified from marine sponges that showcase the ability to act as antibacterial, antiviral, anti-malarial and antifungal agents against human pathogens and fish pathogens in the aquaculture industry. Moreover, the application of such compounds as antimicrobial agents in other veterinary commodities, such as poultry, cattle farming and domesticated cats, is discussed, along with a brief discussion regarding the mode of action of these compounds on the targeted sites in various pathogens. The bioactivity of the compounds discussed in this review is focused mainly on compounds that have been identified between 2000 and 2020 and includes the novel compounds discovered from 2018 to 2021.
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http://dx.doi.org/10.3390/md19050246DOI Listing
April 2021

Inhibition of Autophagy Does Not Affect Innate Cytokine Production in Human Lung Epithelial Cells During Respiratory Syncytial Virus Infection.

Viral Immunol 2021 Apr 9. Epub 2021 Apr 9.

Department of Microbiology, Faculty of Biotechnology and Biomolecular Sciences, Universiti Putra Malaysia, Serdang, Malaysia.

Human respiratory syncytial virus (RSV) is one of the major causes of childhood acute lower respiratory tract infection worldwide. Autophagy is an intracellular pathway involved in nutrient recycling. Recently, autophagy has been reported to play a role in regulating host cytokine response to several viruses, including vesicular stomatitis virus and human immunodeficiency virus. Previous studies using mouse model has shown that inhibition of autophagy reduces RSV-induced cytokine production. However, the role of autophagy in modulating RSV-induced cytokine response in human cells has not been reported. We investigated the role of autophagy in regulating the production of the cytokines C-X-C motif ligand 8 (CXCL8) and C-C motif ligand 5 (CCL5), in RSV-infected human bronchial epithelium BEAS-2B cells. Fluorescent microscopic analysis showed that RSV infection induced autophagosome formation in BEAS-2B cells. This autophagy inducing ability of RSV was further confirmed by flow cytometry. The effects of pharmacological inhibition of autophagy by SAR405 or chloroquine on cell death and cytokine release were quantified using lactate dehydrogenase assay and enzyme-linked immunosorbent assay (ELISA), respectively. We found that SAR405 or chloroquine did not cause cell death. Importantly, ELISA analysis showed that pharmacological inhibition of autophagy by SAR405 or chloroquine did not affect the productions of both CXCL5 and CXCL8. In contrast to the previous studies using mouse model, our data suggest that pharmacological inhibition of autophagy may not be a suitable strategy in controlling RSV-induced airway inflammation.
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http://dx.doi.org/10.1089/vim.2020.0217DOI Listing
April 2021

Combinatorial Antimicrobial Efficacy and Mechanism of Linalool Against Clinically Relevant .

Front Microbiol 2021 17;12:635016. Epub 2021 Mar 17.

Health Sciences Division, Abu Dhabi Women's College, Higher Colleges of Technology, Abu Dhabi, United Arab Emirates.

Antibiotic-adjuvant combinatory therapy serves as a viable treatment option in addressing antibiotic resistance in the clinical setting. This study was carried out to assess and characterize the adjuvant potential and mode of action of linalool against carbapenemase-producing (KPC-KP). Linalool exhibited bactericidal activity alone (11,250 μg/ml) and in combination with meropenem (5,625 μg/ml). Comparative proteomic analysis showed significant reduction in the number of cytoplasmic and membrane proteins, indicating membrane damage in linalool-treated KPC-KP cells. Upregulation of oxidative stress regulator proteins and downregulation of oxidative stress-sensitive proteins indicated oxidative stress. Zeta potential measurement and outer membrane permeability assay revealed that linalool increases the bacterial surface charge as well as the membrane permeability. Intracellular leakage of nucleic acid and proteins was detected upon linalool treatment. Scanning and transmission electron microscopies further revealed the breakage of bacterial membrane and loss of intracellular materials. Linalool induced oxidative stress by generating reactive oxygen species (ROS) which initiates lipid peroxidation, leading to damage of the bacterial membrane. This leads to intracellular leakage, eventually killing the KPC-KP cells. Our study demonstrated that linalool possesses great potential in future clinical applications as an adjuvant along with existing antibiotics attributed to their ability in disrupting the bacterial membrane by inducing oxidative stress. This facilitates the uptake of antibiotics into the bacterial cells, enhancing bacterial killing.
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http://dx.doi.org/10.3389/fmicb.2021.635016DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8010000PMC
March 2021

Rapid Generation of a Recombinant Genotype VIII Newcastle Disease Virus (NDV) Using Full-Length Synthetic cDNA.

Curr Microbiol 2021 Apr 4;78(4):1458-1465. Epub 2021 Mar 4.

Department of Avian Disease Research and Diagnostic, Razi Vaccine and Serum Research Institute, Agricultural Research Education and Extension Organization (AREEO), Karaj, Iran.

Rescue of (-)ssRNA viruses involves the sequential assembly and cloning of the full-length cDNA, which is often a challenging and time-consuming process. The objective of this study was to develop a novel method to rapidly clone the full-length cDNA of a very virulent NDV by only one assembly step. A completely synthetic 15 kb cDNA of a Malaysian genotype VIII NDV known as strain AF2240-I with additional flanking BsmBI sites was synthesised. However, to completely follow the rule-of-six, the additional G residues that are traditionally added after the T7 promoter transcription initiation site were not synthesised. The synthetic fragment was then cloned into low-copy number transcription vector pOLTV5-phiX between the T7 promoter and HDV Rz sequences through digestion with BbsI. The construct was co-transfected with helper plasmids into BSRT7/5 cells. A recombinant NDV called rAF was successfully rescued using transfection supernatant harvested as early as 16 h post-transfection. Virus from each passage showed an intracerebral pathogenicity index (ICPI) and a mean death time (MDT) similar to the parent strain AF2240-I. Moreover, rAF possessed an introduced mutation which was maintained for several passages. The entire rescue using the one-step assembly procedure was completed within a few weeks, which is extremely fast compared to previously used methods.
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http://dx.doi.org/10.1007/s00284-021-02421-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7929900PMC
April 2021

Development of a T7 RNA polymerase expressing cell line using lentivirus vectors for the recovery of recombinant Newcastle disease virus.

J Virol Methods 2021 05 13;291:114099. Epub 2021 Feb 13.

Department of Microbiology, Faculty of Biotechnology and Biomolecular Sciences, Universiti Putra Malaysia, 43400 UPM Serdang, Selangor Darul Ehsan, Malaysia; UPM-MAKNA Cancer Research Laboratory, Institute of Bioscience, Universiti Putra Malaysia, 43400 UPM Serdang, Selangor Darul Ehsan, Malaysia. Electronic address:

The development of a T7 RNA polymerase (T7 RNAP) expressing cell line i.e. BSR T7/5 cells marks an improvement of reverse genetics for the recovery of recombinant Newcastle disease virus (rNDV). BSR T7/5 is developed by transient transfection of plasmid encoding T7 RNAP gene for rNDV rescue. However, the gene expression decreases gradually over multiple passages and eventually hinders the rescue of rNDV. To address this issue, lentiviral vector was used to develop T7 RNAP-expressing HEK293-TA (HEK293-TA-Lv-T7) and SW620 (SW620-Lv-T7) cell lines, evidenced by the expression of T7 RNAP after subsequent 20 passages. rNDV was rescued successfully using HEK293-TA-Lv-T7 clones (R1D3, R1D8, R5B9) and SW620-Lv-T7 clones (R1C11, R3C5) by reverse transfection, yielding comparable virus rescue efficiency and virus titres to that of BSR T7/5. This study provides new tools for rNDV rescue and insights into cell line development and virology by reverse genetics.
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http://dx.doi.org/10.1016/j.jviromet.2021.114099DOI Listing
May 2021

Klebsiella virus UPM2146 lyses multiple drug-resistant Klebsiella pneumoniae in vitro and in vivo.

PLoS One 2021 8;16(1):e0245354. Epub 2021 Jan 8.

Faculty of Biotechnology and Biomolecular Sciences, Department of Microbiology, Universiti Putra Malaysia, Serdang, Selangor, Malaysia.

Klebsiella pneumoniae are opportunistic bacteria found in the gut. In recent years they have been associated with nosocomial infections. The increased incidence of multiple drug-resistant K. pneumoniae makes it necessary to find new alternatives to treat the disease. In this study, phage UPM2146 was isolated from a polluted lake which can lyse its host K. pneumoniae ATCC BAA-2146. Observation from TEM shows that UPM2146 belongs to Caudoviriales (Order) based on morphological appearance. Whole genome analysis of UPM2146 showed that its genome comprises 160,795 bp encoding for 214 putative open reading frames (ORFs). Phylogenetic analysis revealed that the phage belongs to Ackermannviridae (Family) under the Caudoviriales. UPM2146 produces clear plaques with high titers of 1010 PFU/ml. The phage has an adsorption period of 4 min, latent period of 20 min, rise period of 5 min, and releases approximately 20 PFU/ bacteria at Multiplicity of Infection (MOI) of 0.001. UPM2146 has a narrow host-range and can lyse 5 out of 22 K. pneumoniae isolates (22.72%) based on spot test and efficiency of plating (EOP). The zebrafish larvae model was used to test the efficacy of UPM2146 in lysing its host. Based on colony forming unit counts, UPM2146 was able to completely lyse its host at 10 hours onwards. Moreover, we show that the phage is safe to be used in the treatment against K. pneumoniae infections in the zebrafish model.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0245354PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7794032PMC
January 2021

Oncolytic effects of the recombinant Newcastle disease virus, rAF-IL12, against colon cancer cells in vitro and in tumor-challenged NCr-Foxn1nu nude mice.

PeerJ 2020 8;8:e9761. Epub 2020 Dec 8.

Universiti Putra Malaysia, Serdang, Malaysia.

Colon cancer remains one of the main cancers causing death in men and women worldwide as certain colon cancer subtypes are resistant to conventional treatments and the development of new cancer therapies remains elusive. Alternative modalities such as the use of viral-based therapeutic cancer vaccine is still limited, with only the herpes simplex virus (HSV) expressing granulocyte-macrophage colony- stimulating factor (GM-CSF) or talimogene laherparepvec (T-Vec) being approved in the USA and Europe so far. Therefore, it is imperative to continue the search for a new treatment modality. This current study evaluates a combinatorial therapy between the oncolytic Newcastle disease virus (NDV) and interleukin-12 (IL-12) cytokine as a potential therapeutic vaccine to the current anti-cancer drugs. Several in vitro analyses such as MTT assay, Annexin V/FITC flow cytometry, and cell cycle assay were performed to evaluate the cytotoxicity effect of recombinant NDV, rAF-IL12. Meanwhile, serum cytokine, serum biochemical, histopathology of organs and TUNEL assay were carried out to assess the anti-tumoral effects of rAF-IL12 in HT29 tumor-challenged nude mice. The apoptosis mechanism underlying the effect of rAF-IL12 treatment was also investigated using NanoString Gene expression analysis. The recombinant NDV, rAF-IL12 replicated in HT29 colon cancer cells as did its parental virus, AF2240-i. The rAF-IL12 treatment had slightly better cytotoxicity effects towards HT29 cancer cells when compared to the AF2240-i as revealed by the MTT, Annexin V FITC and cell cycle assay. Meanwhile, the 28-day treatment with rAF-IL12 had significantly ( < 0.05) perturbed the growth and progression of HT29 tumor in NCr-Foxn1nu nude mice when compared to the untreated and parental wild-type NDV strain AF2240-i. The rAF-IL12 also modulated the immune system in nude mice by significantly ( < 0.05) increased the level of IL-2, IL-12, and IFN-γ cytokines. Treatment with rAF-IL12 had also significantly ( < 0.05) increased the expression level of apoptosis-related genes such as Fas, caspase-8, BID, BAX, Smad3 and granzyme B in vitro and in vivo. Besides, rAF-IL12 intra-tumoral delivery was considered safe and was not hazardous to the host as evidenced in pathophysiology of the normal tissues and organs of the mice as well as from the serum biochemistry profile of liver and kidney. Therefore, this study proves that rAF-IL12 had better cytotoxicity effects than its parental AF2240-i and could potentially be an ideal treatment for colon cancer in the near future.
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http://dx.doi.org/10.7717/peerj.9761DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7731658PMC
December 2020

Recombinant Enterovirus 71 Viral Protein 1 Fused to a Truncated Newcastle Disease Virus NP (NPt) Carrier Protein.

Vaccines (Basel) 2020 Dec 7;8(4). Epub 2020 Dec 7.

Department of Microbiology, Universiti Putra Malaysia, Serdang 43400, Malaysia.

Enterovirus 71 (EV71) is the major causative agent in hand, foot, and mouth disease (HFMD), and it mainly infects children worldwide. Despite the risk, there is no effective vaccine available for this disease. Hence, a recombinant protein construct of truncated nucleocapsid protein viral protein 1 (NPt-VP1), which is capable of inducing neutralizing antibody against EV71, was evaluated in a mouse model. Truncated nucleocapsid protein Newcastle disease virus that was used as immunological carrier fused to VP1 of EV71 as antigen. The recombinant plasmid carrying corresponding genes was constructed by recombinant DNA technology and the corresponding protein was produced in expression system. The recombinant NPt-VP1 protein had elicited neutralizing antibodies against EV71 with the titer of 1:16, and this result is higher than the titer that is elicited by VP1 protein alone (1:8). It was shown that NPt containing immunogenic epitope(s) of VP1 was capable of inducing a greater functional immune response when compared to full-length VP1 protein alone. It was capable to carry larger polypeptide compared to full-length NP protein. The current study also proved that NPt-VP1 protein can be abundantly produced in recombinant protein form by expression system. The findings from this study support the importance of neutralizing antibodies in EV71 infection and highlight the potential of the recombinant NPt-VP1 protein as EV71 vaccine.
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http://dx.doi.org/10.3390/vaccines8040742DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7762238PMC
December 2020

Palm Kernel Cake Oligosaccharides Acute Toxicity and Effects on Nitric Oxide Levels Using a Zebrafish Larvae Model.

Front Physiol 2020 24;11:555122. Epub 2020 Sep 24.

Institute of Tropical Agriculture and Food Security, Universiti Putra Malaysia, Kuala Lumpur, Malaysia.

One of the beneficial effects of non-digestible oligosaccharides (NDOs) is their anti-inflammatory effects on host animals. While conventional animal studies require that analysis be done after samples have been taken from the host, zebrafish larvae are optically transparent upon hatching and this provides an opportunity for observations to be made within the living zebrafish larvae. This study aimed to take advantage of the optical transparency of zebrafish larvae to study the nitric oxide (NO) reducing effects of NDOs through the use of lipopolysaccharide (LPS) from serovar (ser.) Enteritidis (. Enteritidis) to induce cardiac NO production. Prior to running the above experiment, an acute toxicity assay was conducted in order to determine the appropriate concentration of oligosaccharides to be used. The oligosaccharides tested consisted of oligosaccharides which were extracted from palm kernel cake with a degree of polymerization (DP) equal to or less than six (OligoPKC), commercial mannanoligosaccharide (MOS) and commercial fructooligosaccharide (FOS). Acute toxicity test results revealed that the OligoPKC has a LC of 488.1 μg/ml while both MOS and FOS were non-toxic up to 1,000 μg/ml. Results of the NO measurements revealed that all three NDOs were capable of significantly reducing NO levels in LPS stimulated zebrafish embryos. In summary, at 250 μg/ml, OligoPKC was comparable to MOS and better than FOS at lowering NO in LPS induced zebrafish larvae. However, at higher doses, OligoPKC appears toxic to zebrafish larvae. This implies that the therapeutic potential of OligoPKC is limited by its toxicity.
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http://dx.doi.org/10.3389/fphys.2020.555122DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7541901PMC
September 2020

In-Vitro and In-Silico Evaluations of Heterocyclic-Containing Diarylpentanoids as Bcl-2 Inhibitors Against LoVo Colorectal Cancer Cells.

Molecules 2020 Aug 26;25(17). Epub 2020 Aug 26.

Department of Microbiology, Faculty of Biotechnology and Biomolecular Sciences, Universiti Putra Malaysia, UPM Serdang 43400, Selangor Darul Ehsan, Malaysia.

In the present study, we investigated the in-vitro anti-cancer potential of six diarylpentanoids against a panel of BRAF- and KRAS-mutated colorectal cancer cell lines including T84, SW620, LoVo, HT29, NCI-H508, RKO, and LS411N cells. Structure-activity relationship study suggested that the insertions of tetrahydro-4-thiopyran-4-one and brominated phenyl moieties are essential for better cytotoxicity. Among the evaluated analogs, has been identified as the lead compound due to its low IC values of approximately 1 µM across all cancer cell lines and high chemotherapeutic index of 7.1. Anti-proliferative studies on LoVo cells showed that could inhibit cell proliferation and colony formations by inducing G2/M cell cycle arrest. Subsequent cell apoptosis assay confirmed that is a Bcl-2 inhibitor that could induce intrinsic cell apoptosis by creating a cellular redox imbalance through its direct inhibition on the Bcl-2 protein. Further molecular docking studies revealed that the bromophenyl moieties of could interact with the Bcl-2 surface pocket through hydrophobic interaction, while the tetrahydro-4-thiopyran-4-one fragment could form additional Pi-sulfur and Pi-alkyl interactions in the same binding site. In all, the present results suggest that could be a potent lead that deserves further modification and investigation in the development of a new Bcl-2 inhibitor.
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http://dx.doi.org/10.3390/molecules25173877DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7504466PMC
August 2020

Effect of different cloning strategies in pET-28a on solubility and functionality of a staphylococcal phage endolysin.

Biotechniques 2020 09 13;69(3):161-170. Epub 2020 Aug 13.

Institute of Bioscience, Universiti Putra Malaysia, 43400, Serdang, Selangor, Malaysia.

Endolysins have been studied intensively as an alternative to antibiotics. In this study, endolysin derived from a phage which infects methicillin-resistant (MRSA) was cloned and expressed in pET28a. Initially, the endolysin was cloned using HI/I, resulting in expression of a recombinant endolysin which was expressed in inclusion bodies. While solubilization was successful, the protein remained nonfunctional. Recloning the endolysin using I/I resulted in expression of soluble and functional proteins at 18°C. The endolysin was able to form halo zones on MRSA plates and showed a reduction in turbidity of MRSA growth. Therefore, cloning strategies should be chosen carefully even in an established expression system as they could greatly affect the functionality of the expressed protein.
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http://dx.doi.org/10.2144/btn-2020-0034DOI Listing
September 2020

Cytotoxicity study of the interleukin-12-expressing recombinant Newcastle disease virus strain, rAF-IL12, towards CT26 colon cancer cells in vitro and in vivo.

Cancer Cell Int 2020 29;20:278. Epub 2020 Jun 29.

Institute of Bioscience, Universiti Putra Malaysia, Serdang, Selangor Darul Ehsan 43400 Malaysia.

Background: Oncolytic viruses have emerged as an alternative therapeutic modality for cancer as they can replicate specifically in tumour cells and induce toxic effects leading to apoptosis. Despite the great potentials and promising results shown in multiple studies, it appears that their efficacy is still moderate and deemed as not sufficient in clinical studies. In addressing this issue, genetic/molecular engineering approach has paved its way to improve the therapeutic efficacy as observed in the case of herpes simplex virus (HSV) expressing granulocyte-macrophage colony-stimulating factor (GM-CSF). This study aimed to explore the cytotoxicity effects of recombinant NDV strain AF2240-i expressing interleukin-12 (rAF-IL12) against CT26 colon cancer cells.

Methods: The cytotoxicity effect of rAF-IL12 against CT26 colon cancer cell line was determined by MTT assay. Based on the IC value from the anti-proliferative assay, further downward assays such as Annexin V FITC and cell cycle progression were carried out and measured by flow cytometry. Then, the in vivo study was conducted where the rAF-IL12 viral injections were given at the intra-tumoral site of the CT26 tumour-burden mice. At the end of the experiment, serum biochemical, T cell immunophenotyping, serum cytokine, histopathology of tumour and organ section, TUNEL assay, and Nanostring gene expression analysis were performed.

Results: The rAF-IL12 induced apoptosis of CT26 colon cancer cells in vitro as revealed in the Annexin V FITC analysis and also arrested the cancer cells progression at G phase of the cell cycle analysis. On the other hand, the rAF-IL12 significantly (p < 0.05) inhibited the growth of CT26 tumour in Balb/c mice and had regulated the immune system by increasing the level of CD4 + , CD8 + , IL-2, IL-12, and IFN-γ. Furthermore, the expression level of apoptosis-related genes (bax and p53) was up-regulated as a result of the rAF-IL12 treatment. Additionally, the rAF-IL12 had also down-regulated the expression level of KRAS, BRAF, MAPK1, Notch1, CCL2, and VEGF oncogenes. Besides, rAF-IL12 intra-tumoral delivery was considered safe and not hazardous to the host as evidenced in pathophysiology of the normal tissues and organs of the mice as well as from the serum biochemistry profile of liver and kidney.

Conclusions: These results indicated that rAF-IL12 had better anti-tumoral and cytotoxicity effects compared to its parental wild-type, AF2240-i in combatting the CT26 colon cancer model.
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http://dx.doi.org/10.1186/s12935-020-01372-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7325054PMC
June 2020

Development of an Effective and Stable Genotype-Matched Live Attenuated Newcastle Disease Virus Vaccine Based on a Novel Naturally Recombinant Malaysian Isolate Using Reverse Genetics.

Vaccines (Basel) 2020 Jun 2;8(2). Epub 2020 Jun 2.

Laboratory of Vaccines and Immunotherapeutics, Institute of Bioscience, University Putra Malaysia, Serdang, Selangor 43400, Malaysia.

Genotype VII Newcastle disease viruses are associated with huge economic losses in the global poultry industry. Despite the intensive applications of vaccines, disease outbreaks caused by those viruses continue to occur frequently even among the vaccinated poultry farms. An important factor in the suboptimal protective efficacy of the current vaccines is the genetic mismatch between the prevalent strains and the vaccine strains. Therefore, in the present study, an effective and stable genotype-matched live attenuated Newcastle disease virus (NDV) vaccine was developed using reverse genetics, based on a recently isolated virulent naturally recombinant NDV IBS025/13 Malaysian strain. First of all, the sequence encoding the fusion protein (F) cleavage site of the virus was modified in silico from virulent polybasic (RRQKRF) to avirulent monobasic (GRQGRL) motif. The entire modified sequence was then chemically synthesized and inserted into pOLTV5 transcription vector for virus rescue. A recombinant virus termed mIBS025 was successfully recovered and shown to be highly attenuated based on OIE recommended pathogenicity assessment indices. Furthermore, the virus was shown to remain stably attenuated and retain the avirulent monobasic F cleavage site after 15 consecutive passages in specific-pathogen-free embryonated eggs and 12 passages in one-day-old chicks. More so, the recombinant virus induced a significantly higher hemagglutination inhibition antibody titre than LaSota although both vaccines fully protected chicken against genotype VII NDV induced mortality and morbidity. Finally, mIBS025 was shown to significantly reduce both the duration and quantity of cloacal and oropharyngeal shedding of the challenged genotype VII virus compared to the LaSota vaccine. These findings collectively indicate that mIBS025 provides a better protective efficacy than LaSota and therefore can be used as a promising vaccine candidate against genotype VII NDV strains.
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http://dx.doi.org/10.3390/vaccines8020270DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7349954PMC
June 2020

Secretion of recombinant xylanase in Lactococcus lactis using signal peptides Usp45 and Spk1.

Biotechnol Lett 2020 Sep 25;42(9):1727-1733. Epub 2020 Apr 25.

Department of Cell and Molecular Biology, Faculty of Biotechnology and Biomolecular Sciences, Universiti Putra Malaysia, 43400, Serdang, Selangor, Malaysia.

Objective: The effect of two signal peptides, namely Usp45 and Spk1 on the secretion of xylanase in Lactococcus lactis was analysed.

Results: Xylanase was successfully expressed in Lactococcus lactis. Recombinant xylanase fused to either signal peptide Usp45 or Spk1 showed halo zone on Remazol Brilliant Blue-Xylan plates. This indicated that the xylanase was successfully secreted from the cell. The culture supernatants of strains secreting the xylanase with help of the Spk1 and Usp45 signal peptides contained 49.7 U/ml and 34.4 U/ml of xylanase activity, respectively.

Conclusion: Although Usp45 is the most commonly used signal peptide when secreting heterologous proteins in Lactococcus lactis, this study shows that Spk1 isolated from Pediococcus pentosaceus was superior to Usp45 in regard to xylanase protein secretion.
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http://dx.doi.org/10.1007/s10529-020-02894-1DOI Listing
September 2020

Exploring the Prospects of Engineered Newcastle Disease Virus in Modern Vaccinology.

Viruses 2020 04 16;12(4). Epub 2020 Apr 16.

Laboratory of Vaccines and Immunotherapeutics, Institute of Bioscience, Universiti Putra Malaysia, Serdang, Selangor 43400, Malaysia.

Many traditional vaccines have proven to be incapable of controlling newly emerging infectious diseases. They have also achieved limited success in the fight against a variety of human cancers. Thus, innovative vaccine strategies are highly needed to overcome the global burden of these diseases. Advances in molecular biology and reverse genetics have completely restructured the concept of vaccinology, leading to the emergence of state-of-the-art technologies for vaccine design, development and delivery. Among these modern vaccine technologies are the recombinant viral vectored vaccines, which are known for their incredible specificity in antigen delivery as well as the induction of robust immune responses in the vaccinated hosts. Although a number of viruses have been used as vaccine vectors, genetically engineered Newcastle disease virus (NDV) possesses some useful attributes that make it a preferable candidate for vectoring vaccine antigens. Here, we review the molecular biology of NDV and discuss the reverse genetics approaches used to engineer the virus into an efficient vaccine vector. We then discuss the prospects of the engineered virus as an efficient vehicle of vaccines against cancer and several infectious diseases of man and animals.
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http://dx.doi.org/10.3390/v12040451DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7232247PMC
April 2020

Synthesis and in-vitro anti-cancer evaluations of multi-methoxylated asymmetrical diarylpentanoids as intrinsic apoptosis inducer against colorectal cancer.

Bioorg Med Chem Lett 2020 04 26;30(8):127065. Epub 2020 Feb 26.

Department of Microbiology, Faculty of Biotechnology and Biomolecular Sciences, Universiti Putra Malaysia, 43400 UPM Serdang, Selangor Darul Ehsan, Malaysia; Malaysia Genome Institute (MGI), National Institute of Biotechnology Malaysia (NIBM), Jalan Bangi, 43000 Kajang, Selangor Darul Ehsan, Malaysia.

In the present study, a series of nine stable 3,4,5-methoxylphenyl-containing asymmetrical diarylpentanoids, derivatives of curcuminoids, have been synthesized, characterized and evaluated for their in-vitro anti-cancer potential against a panel of BRAF- and KRAS-mutated colorectal cancer cell lines including T84, LoVo and SW620, HT29, RKO and NCI-H508, respectively. Structure-activity relationship study on cytotoxicity of tested compounds suggested that the presence of meta-hydroxyl and adjacent dimethoxyl groups are crucial for enhanced cytotoxicity of diarylpentanoids. Among the evaluated analogs, 8 has been identified as the lead compound due to its highest chemotherapeutic index of 9.9 and nano molar scale cytotoxicity against SW620 and RKO. Colonies formation and cell cycle analyses on 8-treated RKO cells showed that 8 exhibits strong anti-proliferative activity by inducing G2/M-phase cell arrest. Subsequent flow cytometry based annexin-V and DCFHDA studies suggested that 8 could induce apoptosis through intracellular ROS-dependent pathway. Further Western blot studies confirmed that 8 has induced intrinsic apoptosis in RKO cells through the up-regulations of Bad and Bax pro-apoptotic proteins and down-regulations of Bcl-2 and Bcl-xL pro-survival proteins. In all, the present results suggest that 8 could be a potent lead which deserves further modification and investigation in the development of small molecule-based anti-colorectal cancer agents.
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http://dx.doi.org/10.1016/j.bmcl.2020.127065DOI Listing
April 2020

Lavender essential oil induces oxidative stress which modifies the bacterial membrane permeability of carbapenemase producing Klebsiella pneumoniae.

Sci Rep 2020 01 21;10(1):819. Epub 2020 Jan 21.

Health Sciences Division, Abu Dhabi Women's College, Higher Colleges of Technology, 41012, Abu Dhabi, United Arab Emirates.

Misuse of antibiotics in the clinical and agricultural sectors has caused the emergence of multidrug-resistant (MDR) Klebsiella pneumoniae which contributes a threat to human health. In this study, we assessed the feasibility of lavender essential oil (LVO) as an antimicrobial agent in combinatory therapy with meropenem in suppressing the growth of carbapenemase-producing K. pneumoniae (KPC-KP). Synergistic interactions between LVO and meropenem were detected, which significantly reduce the inhibitory concentration of both LVO and meropenem by 15 and 4-fold respectively. Comparative proteomic profiling identified a disruption in the bacterial membrane via oxidative stress that was indicated by loss of membrane and cytoplasmic proteins and the upregulation of oxidative regulators. As a proof of concept, zeta potential measurements showed a change in cell surface charge while outer membrane permeability measurement indicated an increase in membrane permeability following exposure to LVO. This was indicative of a disrupted outer membrane. Ethidium bromide influx/efflux assays demonstrated no significant efflux pump inhibition by LVO, and scanning electron microscopy revealed irregularities on the cell surface after exposure to LVO. Oxidative stress was also detected with increased level of ROS and lipid peroxidation in LVO-treated cells. In conclusion, our data suggest that LVO induced oxidative stress in K. pneumoniae which oxidizes the outer membrane, enabling the influx of generated ROS, LVO and meropenem into the bacterial cells, causing damage to the cells and eventually death.
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http://dx.doi.org/10.1038/s41598-019-55601-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6972767PMC
January 2020

An improved method for the rescue of recombinant Newcastle disease virus.

Biotechniques 2020 02 15;68(2):96-100. Epub 2020 Jan 15.

Department of Microbiology, Faculty of Biotechnology & Biomolecular Sciences, Universiti Putra Malaysia, 43400 UPM Serdang, Selangor, Malaysia.

Reverse genetics has been used to generate recombinant Newcastle disease virus with enhanced immunogenic properties for vaccine development. The system, which involves co-transfecting the viral antigenomic plasmid with three helper plasmids into a T7 RNA polymerase-expressing cell to produce viral progenies, poses a great challenge. We have modified the standard transfection method to improve the transfection efficiency of the plasmids, resulting in a higher titer of virus progeny production. Two transfection reagents (i.e., lipofectamine and polyethylenimine) were used to compare the transfection efficiency of the four plasmids. The virus progenies produced were quantitated with flow cytometry analysis of the infectious virus unit. The modified transfection method increased the titer of virus progenies compared with that of the standard transfection method.
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http://dx.doi.org/10.2144/btn-2019-0110DOI Listing
February 2020

Evaluation of Gene as a Potential Biomarker for Bladder Cancer.

Dis Markers 2019 15;2019:3875147. Epub 2019 Sep 15.

Medical Genetics Laboratory, Faculty of Medicine and Health Sciences, Universiti Putra Malaysia, Selangor Darul Ehsan, Malaysia.

The ceramide synthase 2 () gene has been linked to tumour recurrence and invasion in many different types of cancers including bladder cancer. In this study, the expression levels of in bladder cancer cell lines were analysed using qRT-PCR and the protein expression in clinical bladder cancer histopathological specimens were examined via immunohistochemistry. The potential utility of as a predictive biomarker of response to oncolytic virotherapy was assessed by correlating the mRNA expression to IC values of cells treated with the Newcastle disease virus (NDV), AF2240 strain. This study demonstrates that is differentially expressed in different types of bladder cancer cell lines and that the siRNA-mediated downregulation of the expression of reduces the migratory potential of UMUC1 bladder cancer cells. However, there were no significant correlations between the expression levels of the CERS2 protein with bladder cancer grade/stage or between the IC50 values of cells treated with NDV and expression. Although the utility of expression may be limited, its potential as an antimigration cancer therapeutic should be further examined.
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http://dx.doi.org/10.1155/2019/3875147DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6766133PMC
April 2020

Evaluation of a Recombinant Newcastle Disease Virus Expressing Human IL12 against Human Breast Cancer.

Sci Rep 2019 Sep 30;9(1):13999. Epub 2019 Sep 30.

Department of Microbiology, Faculty of Biotechnology and Biomolecular Sciences, Universiti Putra Malaysia, 43400, UPM Serdang, Selangor Darul Ehsan, Malaysia.

The Newcastle disease virus (NDV) strain AF2240 is an avian avulavirus that has been demonstrated to possess oncolytic activity against cancer cells. However, to illicit a greater anti-cancer immune response, it is believed that the incorporation of immunostimulatory genes such as IL12 into a recombinant NDV backbone will enhance its oncolytic effect. In this study, a newly developed recombinant NDV that expresses IL12 (rAF-IL12) was tested for its safety, stability and cytotoxicity. The stability of rAF-IL12 was maintained when passaged in specific pathogen free (SPF) chicken eggs from passage 1 to passage 10; with an HA titer of 2. Based on the results obtained from the MTT cytotoxic assay, rAF-IL12 was determined to be safe as it only induced cytotoxic effects against normal chicken cell lines and human breast cancer cells while sparing normal cells. Significant tumor growth inhibition (52%) was observed in the rAF-IL12-treated mice. The in vivo safety profile of rAF-IL12 was confirmed through histological observation and viral load titer assay. The concentration and presence of the expressed IL12 was quantified and verified via ELISA assay. In summary, rAF-IL12 was proven to be safe, selectively replicating in chicken and cancer cells and was able to maintain its stability throughout several passages; thus enhancing its potential as an anti-breast cancer vaccine.
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http://dx.doi.org/10.1038/s41598-019-50222-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6768883PMC
September 2019

Increased ROS Scavenging and Antioxidant Efficiency of Chlorogenic Acid Compound Delivered via a Chitosan Nanoparticulate System for Efficient In Vitro Visualization and Accumulation in Human Renal Adenocarcinoma Cells.

Int J Mol Sci 2019 Sep 20;20(19). Epub 2019 Sep 20.

Department of Cell and Molecular Biology, Faculty of Biotechnology and Biomolecular Sciences, University Putra Malaysia, 43400 Serdang, Selangor, Malaysia.

Naturally existing Chlorogenic acid (CGA) is an antioxidant-rich compound reported to act a chemopreventive agent by scavenging free radicals and suppressing cancer-causing mechanisms. Conversely, the compound's poor thermal and pH (neutral and basic) stability, poor solubility, and low cellular permeability have been a huge hindrance for it to exhibit its efficacy as a nutraceutical compound. Supposedly, encapsulation of CGA in chitosan nanoparticles (CNP), nano-sized colloidal delivery vector, could possibly assist in enhancing its antioxidant properties, in vitro cellular accumulation, and increase chemopreventive efficacy at a lower concentration. Hence, in this study, a stable, monodispersed, non-toxic CNP synthesized via ionic gelation method at an optimum parameter (600 µL of 0.5 mg/mL of chitosan and 200 µL of 0.7 mg/mL of tripolyphosphate), denoted as CNP°, was used to encapsulate CGA. Sequence of physicochemical analyses and morphological studies were performed to discern the successful formation of the CNP°-CGA hybrid. Antioxidant property (studied via DPPH (1,1-diphenyl-2-picrylhydrazyl) assay), in vitro antiproliferative activity of CNP°-CGA, and in vitro accumulation of fluorescently labeled (FITC) CNP°-CGA in cancer cells were evaluated. Findings revealed that successful formation of CNP°-CGA hybrid was reveled through an increase in particle size 134.44 ± 18.29 nm (polydispersity index (PDI) 0.29 ± 0.03) as compared to empty CNP°, 80.89 ± 5.16 nm (PDI 0.26 ± 0.01) with a maximal of 12.04 μM CGA loaded per unit weight of CNP° using 20 µM of CGA. This result correlated with Fourier-Transform Infrared (FTIR) spectroscopic analysis, transmission Electron Microscopy (TEM) and field emission scanning (FESEM) electron microscopy, and ImageJ evaluation. The scavenging activity of CNP°-CGA (IC 5.2 ± 0.10 µM) were conserved and slightly higher than CNP° (IC 6.4±0.78 µM). An enhanced cellular accumulation of fluorescently labeled CNP°-CGA in the human renal cancer cells (786-O) as early as 30 min and increased time-dependently were observed through fluorescent microscopic visualization and flow cytometric assessment. A significant concentration-dependent antiproliferation activity of encapsulated CGA was achieved at IC of 16.20 µM as compared to CGA itself (unable to determine from the cell proliferative assay), implying that the competent delivery vector, chitosan nanoparticle, is able to enhance the intracellular accumulation, antiproliferative activity, and antioxidant properties of CGA at lower concentration as compared to CGA alone.
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http://dx.doi.org/10.3390/ijms20194667DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6801874PMC
September 2019

Virotherapy: Current Trends and Future Prospects for Treatment of Colon and Rectal Malignancies.

Cancer Invest 2019 10;37(8):393-414. Epub 2019 Sep 10.

Department of Microbiology, Faculty of Biotechnology and Biomolecular Sciences, Universit Putra Malaysia , Serdang , Malaysia.

Colorectal cancer (CRC) is one of the most common malignancies. In recent decades, early diagnosis and conventional therapies have resulted in a significant reduction in mortality. However, late stage metastatic disease still has very limited effective treatment options. There is a growing interest in using viruses to help target therapies to tumour sites. In recent years the evolution of immunotherapy has emphasised the importance of directing the immune system to eliminate tumour cells; we aim to give a state-of-the-art over-view of the diverse viruses that have been investigated as potential oncolytic agents for the treatment of CRC.
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http://dx.doi.org/10.1080/07357907.2019.1660887DOI Listing
October 2019

Postbiotic metabolites produced by Lactobacillus plantarum strains exert selective cytotoxicity effects on cancer cells.

BMC Complement Altern Med 2019 Jun 3;19(1):114. Epub 2019 Jun 3.

Institute of Bioscience, Universiti Putra Malaysia, 43400 UPM, Serdang, Selangor, Malaysia.

Background: Lactobacillus plantarum, a major species of Lactic Acid Bacteria (LAB), are capable of producing postbiotic metabolites (PM) with prominent probiotic effects that have been documented extensively for rats, poultry and pigs. Despite the emerging evidence of anticancer properties of LAB, very limited information is available on cytotoxic and antiproliferative activity of PM produced by L. plantarum. Therefore, the cytotoxicity of PM produced by six strains of L. plantarum on various cancer and normal cells are yet to be evaluated.

Methods: Postbiotic metabolites (PM) produced by six strains of L. plantarum were determined for their antiproliferative and cytotoxic effects on normal human primary cells, breast, colorectal, cervical, liver and leukemia cancer cell lines via MTT assay, trypan blue exclusion method and BrdU assay. The toxicity of PM was determined for human and various animal red blood cells via haemolytic assay. The cytotoxicity mode was subsequently determined for selected UL4 PM on MCF-7 cells due to its pronounced cytotoxic effect by fluorescent microscopic observation using AO/PI dye reagents and flow cytometric analyses.

Results: UL4 PM exhibited the lowest IC value on MCF-7, RG14 PM on HT29 and RG11 and RI11 PM on HL60 cell lines, respectively from MTT assay. Moreover, all tested PM did not cause haemolysis of human, dog, rabbit and chicken red blood cells and demonstrated no cytotoxicity on normal breast MCF-10A cells and primary cultured cells including human peripheral blood mononuclear cells, mice splenocytes and thymocytes. Antiproliferation of MCF-7 and HT-29 cells was potently induced by UL4 and RG 14 PM respectively after 72 h of incubation at the concentration of 30% (v/v). Fluorescent microscopic observation and flow cytometric analyses showed that the pronounced cytotoxic effect of UL4 PM on MCF-7 cells was mediated through apoptosis.

Conclusion: In conclusion, PM produced by the six strains of L. plantarum exhibited selective cytotoxic via antiproliferative effect and induction of apoptosis against malignant cancer cells in a strain-specific and cancer cell type-specific manner whilst sparing the normal cells. This reveals the vast potentials of PM from L. plantarum as functional supplement and as an adjunctive treatment for cancer.
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http://dx.doi.org/10.1186/s12906-019-2528-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6547513PMC
June 2019

Genomic analyses of two novel biofilm-degrading methicillin-resistant Staphylococcus aureus phages.

BMC Microbiol 2019 05 28;19(1):114. Epub 2019 May 28.

Department of Microbiology, Faculty of Biotechnology and Biomolecular Sciences, Universiti Putra Malaysia, 43400, Serdang, Selangor Darul Ehsan, Malaysia.

Background: Methicillin-resistant Staphylococcus aureus (MRSA) biofilm producers represent an important etiological agent of many chronic human infections. Antibiotics and host immune responses are largely ineffective against bacteria within biofilms. Alternative actions and novel antimicrobials should be considered. In this context, the use of phages to destroy MRSA biofilms presents an innovative alternative mechanism.

Results: Twenty-five MRSA biofilm producers were used as substrates to isolate MRSA-specific phages. Despite the difficulties in obtaining an isolate of this phage, two phages (UPMK_1 and UPMK_2) were isolated. Both phages varied in their ability to produce halos around their plaques, host infectivity, one-step growth curves, and electron microscopy features. Furthermore, both phages demonstrated antagonistic infectivity on planktonic cultures. This was validated in an in vitro static biofilm assay (in microtiter-plates), followed by the visualization of the biofilm architecture in situ via confocal laser scanning microscopy before and after phage infection, and further supported by phages genome analysis. The UPMK_1 genome comprised 152,788 bp coding for 155 putative open reading frames (ORFs), and its genome characteristics were between the Myoviridae and Siphoviridae family, though the morphological features confined it more to the Siphoviridae family. The UPMK_2 has 40,955 bp with 62 putative ORFs; morphologically, it presented the features of the Podoviridae though its genome did not show similarity with any of the S. aureus in the Podoviridae family. Both phages possess lytic enzymes that were associated with a high ability to degrade biofilms as shown in the microtiter plate and CLSM analyses.

Conclusions: The present work addressed the possibility of using phages as potential biocontrol agents for biofilm-producing MRSA.
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http://dx.doi.org/10.1186/s12866-019-1484-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6540549PMC
May 2019

Recovery of recombinant Mycobacterium tuberculosis antigens fused with cell wall-anchoring motif (LysM) from inclusion bodies using non-denaturing reagent (N-laurylsarcosine).

BMC Biotechnol 2019 05 14;19(1):27. Epub 2019 May 14.

Department of Microbiology, Faculty of Biotechnology and Biomolecular Sciences, Universiti Putra Malaysia, 43400, Serdang, Selangor, Malaysia.

Background: The current limitations of conventional BCG vaccines highlights the importance in developing novel and effective vaccines against tuberculosis (TB). The utilization of probiotics such as Lactobacillus plantarum for the delivery of TB antigens through in-trans surface display provides an effective and safe vaccine approach against TB. Such non-recombinant probiotic surface display strategy involves the fusion of candidate proteins with cell wall binding domain such as LysM, which enables the fusion protein to anchor the L. plantarum cell wall externally, without the need for vector genetic modification. This approach requires sufficient production of these recombinant fusion proteins in cell factory such as Escherichia coli which has been shown to be effective in heterologous protein production for decades. However, overexpression in E. coli expression system resulted in limited amount of soluble heterologous TB-LysM fusion protein, since most of it are accumulated as insoluble aggregates in inclusion bodies (IBs). Conventional methods of denaturation and renaturation for solubilizing IBs are costly, time-consuming and tedious. Thus, in this study, an alternative method for TB antigen-LysM protein solubilization from IBs based on the use of non-denaturating reagent N-lauroylsarcosine (NLS) was investigated.

Results: Expression of TB antigen-LysM fusion genes was conducted in Escherichia coli, but this resulted in IBs deposition in contrast to the expression of TB antigens only. This suggested that LysM fusion significantly altered solubility of the TB antigens produced in E. coli. The non-denaturing NLS technique was used and optimized to successfully solubilize and purify ~ 55% of the recombinant cell wall-anchoring TB antigen from the IBs. Functionality of the recovered protein was analyzed via immunofluorescence microscopy and whole cell ELISA which showed successful and stable cell wall binding to L. plantarum (up to 5 days).

Conclusion: The presented NLS purification strategy enables an efficient and rapid method for obtaining higher yields of soluble cell wall-anchoring Mycobacterium tuberculosis antigens-LysM fusion proteins from IBs in E. coli.
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http://dx.doi.org/10.1186/s12896-019-0522-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6518676PMC
May 2019

Disruption of KPC-producing Klebsiella pneumoniae membrane via induction of oxidative stress by cinnamon bark (Cinnamomum verum J. Presl) essential oil.

PLoS One 2019 2;14(4):e0214326. Epub 2019 Apr 2.

Department of Cell and Molecular Biology, Faculty of Biotechnology and Biomolecular Sciences, Universiti Putra Malaysia, Serdang, Selangor, Malaysia.

Klebsiella pneumoniae (KP) remains the most prevalent nosocomial pathogen and carries the carbapenemase (KPC) gene which confers resistance towards carbapenem. Thus, it is necessary to discover novel antimicrobials to address the issue of antimicrobial resistance in such pathogens. Natural products such as essential oils are a promising source due to their complex composition. Essential oils have been shown to be effective against pathogens, but the overall mechanisms have yet to be fully explained. Understanding the molecular mechanisms of essential oil towards KPC-KP cells would provide a deeper understanding of their potential use in clinical settings. Therefore, we aimed to investigate the mode of action of essential oil against KPC-KP cells from a proteomic perspective by comparing the overall proteome profile of KPC-KP cells treated with cinnamon bark (Cinnamomum verum J. Presl) essential oil (CBO) at their sub-inhibitory concentration of 0.08% (v/v). A total of 384 proteins were successfully identified from the non-treated cells, whereas only 242 proteins were identified from the CBO-treated cells. Proteins were then categorized based on their biological processes, cellular components and molecular function prior to pathway analysis. Pathway analysis showed that CBO induced oxidative stress in the KPC-KP cells as indicated by the abundance of oxidative stress regulator proteins such as glycyl radical cofactor, catalase peroxidase and DNA mismatch repair protein. Oxidative stress is likely to oxidize and disrupt the bacterial membrane as shown by the loss of major membrane proteins. Several genes selected for qRT-PCR analysis validated the proteomic profile and were congruent with the proteomic abundance profiles. In conclusion, KPC-KP cells exposed to CBO undergo oxidative stress that eventually disrupts the bacterial membrane possibly via interaction with the phospholipid bilayer. Interestingly, several pathways involved in the bacterial membrane repair system were also affected by oxidative stress, contributing to the loss of cells viability.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0214326PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6445408PMC
December 2019

Evaluation of Ultra-Microscopic Changes and Proliferation of Apoptotic Glioblastoma Multiforme Cells Induced by Velogenic Strain of Newcastle Disease Virus AF2240

Asian Pac J Cancer Prev 2019 Mar 26;20(3):757-765. Epub 2019 Mar 26.

Faculty of Agriculture and Biotechnology, University Sultan Zainalb Abidin, Malaysia.

Aim: Newcastle disease virus (NDV) is a member of genus Avulavirus within the family Paramyxoviridae. Interest of using NDV as an anticancer agent has arisen from its ability to kill tumor cells with limited toxicity to normal cells. Methods: In this investigation, the proliferation of brain tumor cell line, glioblastoma multiform (DBTRG.05MG) induced by NDV strain AF2240 was evaluated in-vitro, by using MTT proliferation assay. Furthermore, Cytological observations were studied using fluorescence microscopy and transmission electron microscopy, DNA laddering in agarose gel electrophoresis assay used to detect the mode of cell death and analysis of the cellular DNA content by flowcytometery. Results: MTT proliferation assay, Cytological observations using fluorescence microscopy and transmission electron microscopy show the anti-proliferation effect and apoptogenic features of NDV on DBTRG.05MG. Furthermore, analysis of the cellular DNA content showed that there was a loss of treated cells in all cell cycle phases (G1, S and G2/M) accompanied with increasing in sub-G1 region (apoptosis peak). Conclusion: It could be concluded that NDV strain AF2240 is a potent antitumor agent that induce apoptosis and its cytotoxicity increasing while increasing of time and virus titer.
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http://dx.doi.org/10.31557/APJCP.2019.20.3.757DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6825790PMC
March 2019

Mechanisms of Antimicrobial Resistance (AMR) and Alternative Approaches to Overcome AMR.

Curr Drug Discov Technol 2020 ;17(4):430-447

Department of Cell and Molecular Biology, Faculty of Biotechnology and Biomolecular Sciences, Universiti Putra Malaysia, 43400 Serdang, Selangor, Malaysia.

Antimicrobials are useful compounds intended to eradicate or stop the growth of harmful microorganisms. The sustained increase in the rates of antimicrobial resistance (AMR) worldwide is worrying and poses a major public health threat. The development of new antimicrobial agents is one of the critical approaches to overcome AMR. However, in the race towards developing alternative approaches to combat AMR, it appears that the scientific community is falling behind when pitched against the evolutionary capacity of multi-drug resistant (MDR) bacteria. Although the "pioneering strategy" of discovering completely new drugs is a rational approach, the time and effort taken are considerable, the process of drug development could instead be expedited if efforts were concentrated on enhancing the efficacy of existing antimicrobials through: combination therapies; bacteriophage therapy; antimicrobial adjuvants therapy or the application of nanotechnology. This review will briefly detail the causes and mechanisms of AMR as background, and then provide insights into a novel, future emerging or evolving strategies that are currently being evaluated and which may be developed in the future to tackle the progression of AMR.
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http://dx.doi.org/10.2174/1570163816666190304122219DOI Listing
January 2020

Genotype Diversity of Newcastle Disease Virus in Nigeria: Disease Control Challenges and Future Outlook.

Adv Virol 2018 2;2018:6097291. Epub 2018 Dec 2.

Laboratory of Vaccines and Immunotherapeutics, Institute of Bioscience, Universiti Putra Malaysia, 43400 Serdang, Selangor, Malaysia.

Newcastle disease (ND) is one of the most important avian diseases with considerable threat to the productivity of poultry all over the world. The disease is associated with severe respiratory, gastrointestinal, and neurological lesions in chicken leading to high mortality and several other production related losses. The aetiology of the disease is an avian paramyxovirus type-1 or Newcastle disease virus (NDV), whose isolates are serologically grouped into a single serotype but genetically classified into a total of 19 genotypes, owing to the continuous emergence and evolution of the virus. In Nigeria, molecular characterization of NDV is generally very scanty and majorly focuses on the amplification of the partial F gene for genotype assignment. However, with the introduction of the most objective NDV genotyping criteria which utilize complete fusion protein coding sequences in phylogenetic taxonomy, the enormous genetic diversity of the virus in Nigeria became very conspicuous. In this review, we examine the current ecological distribution of various NDV genotypes in Nigeria based on the available complete fusion protein nucleotide sequences (1662 bp) in the NCBI database. We then discuss the challenges of ND control as a result of the wide genetic distance between the currently circulating NDV isolates and the commonest vaccines used to combat the disease in the country. Finally, we suggest future directions in the war against the economically devastating ND in Nigeria.
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http://dx.doi.org/10.1155/2018/6097291DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6304561PMC
December 2018

Proof of concept in utilizing in-trans surface display system of Lactobacillus plantarum as mucosal tuberculosis vaccine via oral administration in mice.

BMC Biotechnol 2018 10 11;18(1):63. Epub 2018 Oct 11.

Department of Microbiology, Faculty of Biotechnology and Biomolecular Sciences, Universiti Putra Malaysia, 43400, Serdang, Selangor Darul Ehsan, Malaysia.

Background: Tuberculosis is one of the most common and deadliest infectious diseases worldwide affecting almost a third of the world's population. Although this disease is being prevented and controlled by the Bacille Calmette Guérin (BCG) vaccine, the protective efficacy is highly variable and substandard (0-80%) in adults. Therefore, novel and effective tuberculosis vaccine that can overcome the limitations from BCG vaccine need to be developed.

Results: A novel approach of utilizing an in-trans protein surface display system of Lactobacillus plantarum carrying and displaying combination of Mycobacterium tuberculosis subunit epitope antigens (Ag85B, CFP-10, ESAT-6, Rv0475 and Rv2031c) fused with LysM anchor motif designated as ACERL was constructed, cloned and expressed in Esherichia coli Rossetta expression host. Subsequently the binding capability of ACERL to the cell wall of L. plantarum was examined via the immunofluorescence microscopy and whole cell ELISA where successful attachment and consistent stability of cell wall binding up to 4 days was determined. The immunization of the developed vaccine of L. plantarum surface displaying ACERL (Lp ACERL) via the oral route was studied in mice for its immunogenicity effects. Lp ACERL immunization was able to invoke significant immune responses that favor the Th1 type cytokine response of IFN-γ, IL-12 and IL-2 as indicated by the outcome from the cytokine profiling of spleen, lung, gastrointestinal tract (GIT), and the re-stimulation of the splenocytes from the immunized mice. Co-administration of an adjuvant consisting of Lactococcus lactis secreting mouse IL-12 (LcIL-12) with Lp ACERL was also investigated. It was shown that the addition of LcIL-12 was able to further generate significant Th1 type cytokines immune responses, similar or better than that of Lp ACERL alone which can be observed from the cytokine profiling of the immunized mice's spleen, lung and GIT.

Conclusions: This study represents a proof of concept in the development of L. plantarum as a carrier for a non-genetically modified organism (GMO) tuberculosis vaccine, which may be the strategy in the future for tuberculosis vaccine development.
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http://dx.doi.org/10.1186/s12896-018-0461-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6182793PMC
October 2018