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Fulltext Crystal structure and immunological properties of the first annexin from Schistosoma mansoni: insights into the structural integrity of the schistosomal tegument

Leow CY, Willis C, Osman A, Mason L, Simon A, Smith BJ, et al.

FEBS J, 2014 Feb;281(4):1209-25.
PMID: 24428567 DOI: 10.1111/febs.12700

Schistosomiasis is a major parasitic disease of humans, second only to malaria in its global impact. The disease is caused by digenean trematodes that infest the vasculature of their human hosts. These flukes are limited externally by a body wall composed of a syncytial epithelium, the apical surface membrane of which is a parasitism-adapted dual membrane complex. Annexins are thought to be of integral importance for the stability of this apical membrane system. Here, we present the first structural and immunobiochemical characterization of an annexin from Schistosoma mansoni. The crystal structure of annexin B22 confirms the presence of the previously predicted α-helical segment in the II/III linker and reveals a covalently linked head-to-head dimer. From the calcium-bound crystal structure of this protein, canonical type II, type III and B site positions are occupied, and a novel binding site has been identified. The dimer arrangement observed in the crystal structure suggests the presence of two prominent features, a potential non-canonical membrane binding site and a potential binding groove opposite to the former. Results from transcriptional profiling during development show that annexin B22 expression is correlated with life stages of the parasite that possess the syncytial tegument layer, and ultrastructural localization by immuno-electron microscopy confirms the occurrence of annexins in the tegument of S. mansoni. Data from membrane binding and aggregation assays indicate the presence of differential molecular mechanisms and support the hypothesis of annexin B22 providing structural integrity in the tegument.
Fulltext Structure-function analysis of apical membrane-associated molecules of the tegument of schistosome parasites of humans: prospects for identification of novel targets for parasite control

Leow CY, Willis C, Hofmann A, Jones MK

Br J Pharmacol, 2015 Apr;172(7):1653-63.
PMID: 25176442 DOI: 10.1111/bph.12898

Neglected tropical diseases are a group of some 17 diseases that afflict poor and predominantly rural people in developing nations. One significant disease that contributes to substantial morbidity in endemic areas is schistosomiasis, caused by infection with one of five species of blood fluke belonging to the trematode genus Schistosoma. Although there is one drug available for treatment of affected individuals in clinics, or for mass administration in endemic regions, there is a need for new therapies. A prominent target organ of schistosomes, either for drug or vaccine development, is the peculiar epithelial syncytium that forms the body wall (tegument) of this parasite. This dynamic layer is maintained and organized by concerted activity of a range of proteins, among which are the abundant tegumentary annexins. In this review, we will outline advances in structure-function analyses of these annexins, as a means to understanding tegument cell biology in host-parasite interaction and their potential exploitation as targets for anti-schistosomiasis therapies.
Fulltext Performance Evaluation of Localization Accuracy for a Log-Normal Shadow Fading Wireless Sensor Network under Physical Barrier Attacks

Hussein AA, Rahman TA, Leow CY

Sensors (Basel), 2015;15(12):30545-70.
PMID: 26690159 DOI: 10.3390/s151229817

Localization is an apparent aspect of a wireless sensor network, which is the focus of much interesting research. One of the severe conditions that needs to be taken into consideration is localizing a mobile target through a dispersed sensor network in the presence of physical barrier attacks. These attacks confuse the localization process and cause location estimation errors. Range-based methods, like the received signal strength indication (RSSI), face the major influence of this kind of attack. This paper proposes a solution based on a combination of multi-frequency multi-power localization (C-MFMPL) and step function multi-frequency multi-power localization (SF-MFMPL), including the fingerprint matching technique and lateration, to provide a robust and accurate localization technique. In addition, this paper proposes a grid coloring algorithm to detect the signal hole map in the network, which refers to the attack-prone regions, in order to carry out corrective actions. The simulation results show the enhancement and robustness of RSS localization performance in the face of log normal shadow fading effects, besides the presence of physical barrier attacks, through detecting, filtering and eliminating the effect of these attacks.
Fulltext Physical Layer Security Using Two-Path Successive Relaying

Liau QY, Leow CY, Ding Z

Sensors (Basel), 2016 Jun 09;16(6).
PMID: 27294924 DOI: 10.3390/s16060846

Relaying is one of the useful techniques to enhance wireless physical-layer security. Existing literature shows that employing full-duplex relay instead of conventional half-duplex relay improves secrecy capacity and secrecy outage probability, but this is at the price of sophisticated implementation. As an alternative, two-path successive relaying has been proposed to emulate operation of full-duplex relay by scheduling a pair of half-duplex relays to assist the source transmission alternately. However, the performance of two-path successive relaying in secrecy communication remains unexplored. This paper proposes a secrecy two-path successive relaying protocol for a scenario with one source, one destination and two half-duplex relays. The relays operate alternately in a time division mode to forward messages continuously from source to destination in the presence of an eavesdropper. Analytical results reveal that the use of two half-duplex relays in the proposed scheme contributes towards a quadratically lower probability of interception compared to full-duplex relaying. Numerical simulations show that the proposed protocol achieves the ergodic achievable secrecy rate of full-duplex relaying while delivering the lowest probability of interception and secrecy outage probability compared to the existing half duplex relaying, full duplex relaying and full duplex jamming schemes.
Fulltext Two-Stage Design Method for Enhanced Inductive Energy Transmission with Q-Constrained Planar Square Loops

Eteng AA, Abdul Rahim SK, Leow CY, Chew BW, Vandenbosch GA

PLoS One, 2016;11(2):e0148808.
PMID: 26890878 DOI: 10.1371/journal.pone.0148808

Q-factor constraints are usually imposed on conductor loops employed as proximity range High Frequency Radio Frequency Identification (HF-RFID) reader antennas to ensure adequate data bandwidth. However, pairing such low Q-factor loops in inductive energy transmission links restricts the link transmission performance. The contribution of this paper is to assess the improvement that is reached with a two-stage design method, concerning the transmission performance of a planar square loop relative to an initial design, without compromise to a Q-factor constraint. The first stage of the synthesis flow is analytical in approach, and determines the number and spacing of turns by which coupling between similar paired square loops can be enhanced with low deviation from the Q-factor limit presented by an initial design. The second stage applies full-wave electromagnetic simulations to determine more appropriate turn spacing and widths to match the Q-factor constraint, and achieve improved coupling relative to the initial design. Evaluating the design method in a test scenario yielded a more than 5% increase in link transmission efficiency, as well as an improvement in the link fractional bandwidth by more than 3%, without violating the loop Q-factor limit. These transmission performance enhancements are indicative of a potential for modifying proximity HF-RFID reader antennas for efficient inductive energy transfer and data telemetry links.
Fulltext Sidelobe reduction and capacity improvement of open-loop collaborative beamforming in wireless sensor networks

Jayaprakasam S, Abdul Rahim SK, Leow CY, Ting TO

PLoS One, 2017;12(5):e0175510.
PMID: 28464000 DOI: 10.1371/journal.pone.0175510

Collaborative beamforming (CBF) with a finite number of collaborating nodes (CNs) produces sidelobes that are highly dependent on the collaborating nodes' locations. The sidelobes cause interference and affect the communication rate of unintended receivers located within the transmission range. Nulling is not possible in an open-loop CBF since the collaborating nodes are unable to receive feedback from the receivers. Hence, the overall sidelobe reduction is required to avoid interference in the directions of the unintended receivers. However, the impact of sidelobe reduction on the capacity improvement at the unintended receiver has never been reported in previous works. In this paper, the effect of peak sidelobe (PSL) reduction in CBF on the capacity of an unintended receiver is analyzed. Three meta-heuristic optimization methods are applied to perform PSL minimization, namely genetic algorithm (GA), particle swarm algorithm (PSO) and a simplified version of the PSO called the weightless swarm algorithm (WSA). An average reduction of 20 dB in PSL alongside 162% capacity improvement is achieved in the worst case scenario with the WSA optimization. It is discovered that the PSL minimization in the CBF provides capacity improvement at an unintended receiver only if the CBF cluster is small and dense.
Fulltext Compact circularly polarized truncated square ring slot antenna with suppressed higher resonances

Sabran MI, Abdul Rahim SK, Leow CY, Soh PJ, Chew BW, Vandenbosch GA

PLoS One, 2017;12(2):e0172162.
PMID: 28192504 DOI: 10.1371/journal.pone.0172162

This paper presents a compact circularly polarized (CP) antenna with an integrated higher order harmonic rejection filter. The proposed design operates within the ISM band of 2.32 GHz- 2.63 GHz and is suitable for example for wireless power transfer applications. Asymmetrical truncated edges on a square ring create a defected ground structure to excite the CP property, simultaneously realizing compactness. It offers a 50.5% reduced patch area compared to a conventional design. Novel stubs and slot shapes are integrated in the transmission line to reduce higher (up to the third) order harmonics. The proposed prototype yields a -10 dB reflection coefficient (S11) impedance bandwidth of 12.53%, a 3 dB axial ratio bandwidth of 3.27%, and a gain of 5.64 dBi. Measurements also show good agreement with simulations.
Fulltext Robust multiple frequency multiple power localization schemes in the presence of multiple jamming attacks

Hussein AA, Leow CY, Rahman TA

PLoS One, 2017;12(5):e0177326.
PMID: 28493977 DOI: 10.1371/journal.pone.0177326

Localization of the wireless sensor network is a vital area acquiring an impressive research concern and called upon to expand more with the rising of its applications. As localization is gaining prominence in wireless sensor network, it is vulnerable to jamming attacks. Jamming attacks disrupt communication opportunity among the sender and receiver and deeply impact the localization process, leading to a huge error of the estimated sensor node position. Therefore, detection and elimination of jamming influence are absolutely indispensable. Range-based techniques especially Received Signal Strength (RSS) is facing severe impact of these attacks. This paper proposes algorithms based on Combination Multiple Frequency Multiple Power Localization (C-MFMPL) and Step Function Multiple Frequency Multiple Power Localization (SF-MFMPL). The algorithms have been tested in the presence of multiple types of jamming attacks including capture and replay, random and constant jammers over a log normal shadow fading propagation model. In order to overcome the impact of random and constant jammers, the proposed method uses two sets of frequencies shared by the implemented anchor nodes to obtain the averaged RSS readings all over the transmitted frequencies successfully. In addition, three stages of filters have been used to cope with the replayed beacons caused by the capture and replay jammers. In this paper the localization performance of the proposed algorithms for the ideal case which is defined by without the existence of the jamming attack are compared with the case of jamming attacks. The main contribution of this paper is to achieve robust localization performance in the presence of multiple jamming attacks under log normal shadow fading environment with a different simulation conditions and scenarios.
Fulltext The role of REST and HDAC2 in epigenetic dysregulation of Nav1.5 and nNav1.5 expression in breast cancer

Kamarulzaman NS, Dewadas HD, Leow CY, Yaacob NS, Mokhtar NF

Cancer Cell Int, 2017;17:74.
PMID: 28785170 DOI: 10.1186/s12935-017-0442-6

BACKGROUND: Increased expression of voltage-gated sodium channels (VGSCs) have been implicated with strong metastatic potential of human breast cancer in vitro and in vivo where the main culprits are cardiac isoform Nav1.5 and its 'neonatal' splice variant, nNav1.5. Several factors have been associated with Nav1.5 and nNav1.5 gain of expression in breast cancer mainly hormones, and growth factors.
AIM: This study aimed to investigate the role of epigenetics via transcription repressor, repressor element silencing transcription factor (REST) and histone deacetylases (HDACs) in enhancing Nav1.5 and nNav1.5 expression in human breast cancer by assessing the effect of HDAC inhibitor, trichostatin A (TSA).
METHODS: The less aggressive human breast cancer cell line, MCF-7 cells which lack Nav1.5 and nNav1.5 expression was treated with TSA at a concentration range 10-10,000 ng/ml for 24 h whilst the aggressive MDA-MB-231 cells was used as control. The effect of TSA on Nav1.5, nNav1.5, REST, HDAC1, HDAC2, HDAC3, MMP2 and N-cadherin gene expression level was analysed by real-time PCR. Cell growth (MTT assay) and metastatic behaviors (lateral motility and migration assays) were also measured.
RESULTS: mRNA expression level of Nav1.5 and nNav1.5 were initially very low in MCF-7 compared to MDA-MB-231 cells. Inversely, mRNA expression level of REST, HDAC1, HDAC2, and HDAC3 were all greater in MCF-7 compared to MDA-MB-231 cells. Treatment with TSA significantly increased the mRNA expression level of Nav1.5 and nNav1.5 in MCF-7 cells. On the contrary, TSA significantly reduced the mRNA expression level of REST and HDAC2 in this cell line. Remarkably, despite cell growth inhibition by TSA, motility and migration of MCF-7 cells were enhanced after TSA treatment, confirmed with the up-regulation of metastatic markers, MMP2 and N-cadherin.
CONCLUSIONS: This study identified epigenetics as another factor that regulate the expression level of Nav1.5 and nNav1.5 in breast cancer where REST and HDAC2 play important role as epigenetic regulators that when lacking enhances the expression of Nav1.5 and nNav1.5 thus promotes motility and migration of breast cancer. Elucidation of the regulatory mechanisms for gain of Nav1.5 and nNav1.5 expression may be helpful for seeking effective strategies for the management of metastatic diseases.
Fulltext Current progress of immunoinformatics approach harnessed for cellular- and antibody-dependent vaccine design

Kazi A, Chuah C, Majeed ABA, Leow CH, Lim BH, Leow CY

Pathog Glob Health, 2018 05;112(3):123-131.
PMID: 29528265 DOI: 10.1080/20477724.2018.1446773

Immunoinformatics plays a pivotal role in vaccine design, immunodiagnostic development, and antibody production. In the past, antibody design and vaccine development depended exclusively on immunological experiments which are relatively expensive and time-consuming. However, recent advances in the field of immunological bioinformatics have provided feasible tools which can be used to lessen the time and cost required for vaccine and antibody development. This approach allows the selection of immunogenic regions from the pathogen genomes. The ideal regions could be developed as potential vaccine candidates to trigger protective immune responses in the hosts. At present, epitope-based vaccines are attractive concepts which have been successfully trailed to develop vaccines which target rapidly mutating pathogens. In this article, we provide an overview of the current progress of immunoinformatics and their applications in the vaccine design, immune system modeling and therapeutics.
Isolation and characterization of a novel anti-salbutamol chicken scFv for human doping urinalysis

Lee W, Syed A A, Leow CY, Tan SC, Leow CH

Anal Biochem, 2018 08 15;555:81-93.
PMID: 29775561 DOI: 10.1016/j.ab.2018.05.009

Anti-salbutamol antibodies remain as important tools for the detection of salbutamol abuse in athletic doping. This study evaluated the feasibility and efficiency of the chicken (Gallus gallus domesticus) as an immunization host to generate anti-salbutamol scFv antibodies by phage display. A phage display antibody library was constructed from a single chicken immunized against salbutamol-KLH conjugate. After a stringent biopanning strategy, a novel scFv clone which was inhibited by free salbutamol recorded the highest affinity. This scFv was expressed as soluble and functional protein in Escherichia coli T7 SHuffle Express B (DE3) strain. Cross-reactivity studies of the scFv towards other relevant β2-agonists revealed that the scFv cross-reacted significantly towards clenbuterol. The determined IC50 of the scFv towards the two β2-agonists were; IC50 salbutamol = ∼0.310 μg/ml, IC50 clenbuterol = ∼0.076 μg/ml. The generated scFv demonstrated poor stability based on accelerated stability studies. The scFv was used to develop an competitive indirect ELISA (LOD = 0.125 μg/ml) for detection of parent salbutamol in spiked human urine (n = 18) with ∼83.4% reliability at the cut-off of 1 μg/ml currently implemented by WADA and may be of potential use in human doping urinalysis.
Fulltext Isolation and characterization of malaria PfHRP2 specific VNAR antibody fragments from immunized shark phage display library

Leow CH, Fischer K, Leow CY, Braet K, Cheng Q, McCarthy J

Malar J, 2018 Oct 24;17(1):383.
PMID: 30355309 DOI: 10.1186/s12936-018-2531-y

BACKGROUND: Malaria rapid diagnostic tests (RDTs) represent an important antibody based immunoassay platform. Unfortunately, conventional monoclonal antibodies are subject to degradation shortening shelf lives of RDTs. The variable region of the receptor (VNAR) from shark has a potential as alternative to monoclonal antibodies in RDTs due to high thermal stability.
METHODS: In this study, new binders derived from shark VNAR domains library were investigated. Following immunization of a wobbegong shark (Orectolobus ornatus) with three recombinant malaria biomarker proteins (PfHRP2, PfpLDH and Pvaldolase), a single domain antibody (sdAb) library was constructed from splenocytes. Target-specific VNAR phage were isolated by panning. One specific clone was selected for expression in Escherichia coli expression system, and study of binding reactivity undertaken.
RESULTS: The primary VNAR domain library possessed a titre of 1.16 × 106 pfu/mL. DNA sequence analysis showed 82.5% of isolated fragments appearing to contain an in-frame sequence. After multiple rounds of biopanning, a highly dominant clone specific to PfHRP2 was identified and selected for protein production in an E. coli expression system. Biological characterization showed the recombinant protein expressed in periplasmic has better detection sensitivity than that of cytoplasmic proteins. Assays of binding activity indicated that its reactivity was inferior to the positive control mAb C1-13.
CONCLUSIONS: Target-specific bacteriophage VNARs were successfully isolated after a series of immunization, demonstrating that phage display technology is a useful tool for selection of antigen binders. Generation of new binding reagents such as VNAR antibodies that specifically recognize the malaria biomarkers represents an appealing approach to improve the performance of RDTs.
Schistosomiasis in Malaysia: A review

Chuah C, Gobert GN, Latif B, Heo CC, Leow CY

Acta Trop, 2019 Feb;190:137-143.
PMID: 30448471 DOI: 10.1016/j.actatropica.2018.11.012

Schistosomiasis, a neglected tropical parasitic disease caused by the trematode flatworms of the genus Schistosoma, affects approximately 207 million people worldwide. Among the five main species infecting humans, Schistosoma mansoni and S. japonicum are responsible for the majority of hepatointestinal schistosomiasis. Human settlements near fresh water sites that lack proper sanitary systems often contribute to the transmission of disease. This risk particularly impacts on travellers or immigrants who come into contact with larvae-contaminated water. This review discusses the central features of schistosomiasis; including clinical manifestations, diagnosis, treatments, and the preventive measures available for the control of this disease. The description of the Malaysian schistosome species Schistosoma malayensis and the current status of schistosomiasis in Malaysia including the compilation of cases diagnosed from 1904 to 2015 are also discussed in this paper.
Flavivirus infection-A review of immunopathogenesis, immunological response, and immunodiagnosis

Chong HY, Leow CY, Abdul Majeed AB, Leow CH

Virus Res, 2019 12;274:197770.
PMID: 31626874 DOI: 10.1016/j.virusres.2019.197770

Flaviviruses are group of single stranded RNA viruses that cause severe endemic infection and epidemics on a global scale. It presents a significant health impact worldwide and the viruses have the potential to emerge and outbreak in a non-endemic geographical region. Effective vaccines for prophylaxis are only available for several flaviviruses such as Yellow Fever virus, Tick-borne Encephalitis Virus, Dengue Virus and Japanese Encephalitis Virus and there is no antiflaviviral agent being marketed. This review discusses the flavivirus genome, replication cycle, epidemiology, clinical presentation and pathogenesis upon infection. Effective humoral response is critical to confer protective immunity against flaviviruses. Hence, we have also highlighted the immune responses elicited upon infection, various diagnostic facilities available for flaviviral disease and monoclonal antibodies available to date against flavivirus infection.
Molecular characterization of Schistosoma mansoni tegument annexins and comparative analysis of antibody responses following parasite infection

Leow CY, Willis C, Leow CH, Hofmann A, Jones M

Mol Biochem Parasitol, 2019 12;234:111231.
PMID: 31628972 DOI: 10.1016/j.molbiopara.2019.111231

Schistosomes are parasitic blood flukes that infect approximately 250 million people worldwide. The disease known as schistosomiasis, is the second most significant tropical parasitic disease after malaria. Praziquantel is the only effective drug currently licensed for schistosomiasis and there are concerns about resistance to the drug. There has been much effort to develop vaccines against schistosomiasis to produce long-term protection in endemic regions. Surface-associated proteins, and in particular, those expressed in the body wall, or tegument, have been proposed as potential vaccine targets. Of these, annexins are thought to be of integral importance for the stability of this apical membrane system. Here, we present the structural and immunobiochemical characterization of four homologous annexins namely annexin B30, annexin B5a, annexin B7a and annexin B5b from S. mansoni. Bioinformatics analysis showed that there was no signal peptide predicted for any annexin in this study. Further analysis showed that each of all four annexin protein possesses a primary structure consisting of a short but variable N-terminal region and a long C-terminal core containing four homologous annexin repeats (I-IV), which contain five alpha-helices. The life cycle expression profile of each annexin was assessed using quantitative PCR. The results showed that the overall transcript levels of the each of four homologous annexins were relatively low in the egg stage, but increased gradually after the transition of cercariae (the invasive schistosome larvae) to schistosomula (the post-invasive larvae). Circular dichroism (CD) demonstrated that rAnnexin B30, rAnnexin B5a and rAnnexin 7a were folded, showing a secondary structure content rich in alpha-helices. The membrane binding affinity was enhanced when rAnnexin B30, rAnnexin B5a and rAnnexin 7a was incubated in the presence of Ca2+. All annexin members evaluated in this study were immunolocalized to the tegument, with immunoreactivity also occurring in cells and in muscle of adult parasites. All four recombinant annexins were immunoreactive and they were recognized by the sera of mice infected with S. mansoni. In conclusion, the overall results present the molecular characterization of annexin B30, annexin B5a, annexin B7a and annexin B5b from S. mansoni in host-parasite interactions and strongly suggest that the molecules could be useful candidates for vaccine or diagnostic development.
Fulltext Reverse vaccinology approach for the identification and characterization of outer membrane proteins of Shigella flexneri as potential cellular- and antibody-dependent vaccine candidates

Leow CY, Kazi A, Hisyam Ismail CMK, Chuah C, Lim BH, Leow CH, et al.

Clin Exp Vaccine Res, 2020 Jan;9(1):15-25.
PMID: 32095437 DOI: 10.7774/cevr.2020.9.1.15

Purpose: In the developing world, bacillary dysentery is one of the most common communicable diarrheal infections. There are approximately 169 million cases of shigellosis reported worldwide. The disease is transmitted by a group of Gram-negative intracellular enterobacteria known as Shigella flexneri, S. sonnei, S. dysenteriae, and S. boydii. Conventional treatment regimens for Shigella have been less effective due to the development of resistant strains against antibiotics. Therefore, an effective vaccine for the long term control of Shigella transmission is urgently needed.
Materials and Methods: In this study, a reverse vaccinology approach was employed to identify most conserved and immunogenic outer membrane proteins (OMPs) of S. flexneri 2a.
Results: Five OMPs including fepA, ompC, nlpD_1, tolC, and nlpD_2 were identified as potential vaccine candidates. Protein-protein interactions analysis using STRING software (https://string-db.org/) revealed that five of these OMPs may potentially interact with other intracellular proteins which are involved in beta-lactam resistance pathway. B- and T-cell epitopes of the selected OMPs were predicted using BCPred as well as Propred I and Propred (http://crdd.osdd.net/raghava/propred/), respectively. Each of these OMPs contains regions which are capable to induce B- and T-cell immune responses.
Conclusion: Analysis acquired from this study showed that five selected OMPs have great potential for vaccine development against S. flexneri infection. The predicted immunogenic epitopes can also be used for development of peptide vaccines or multi-epitope vaccines against human shigellosis. Reverse vaccinology is a promising strategy for the discovery of potential vaccine candidates which can be used for future vaccine development against global persistent infections.
Immunogenicity, antibody responses and vaccine efficacy of recombinant annexin B30 against Schistosoma mansoni

Leow CY, Willis C, Chuah C, Leow CH, Jones M

Parasite Immunol., 2020 03;42(3):e12693.
PMID: 31880816 DOI: 10.1111/pim.12693

AIMS: Schistosomes infect approximately 250 million people worldwide. To date, there is no effective vaccine available for the prevention of schistosome infection in endemic regions. There remains a need to develop means to confer long-term protection of individuals against reinfection. In this study, an annexin, namely annexin B30, which is highly expressed in the tegument of Schistosoma mansoni was selected to evaluate its immunogenicity and protective efficacy in a mouse model.
METHODS AND RESULTS: Bioinformatics analysis showed that there were three potential linear B-cell epitopes and four conformational B-cell epitopes predicted from annexin B30, respectively. Full-length annexin B30 was cloned and expressed in Escherichia coli BL21(DE3). In the presence of adjuvants, the soluble recombinant protein was evaluated for its protective efficacy in two independent vaccine trials. Immunization of CBA mice with recombinant annexin B30 formulated either in alum only or alum/CpG induced a mixed Th1/Th2 cytokine profile but no significant protection against schistosome infection was detected.
CONCLUSION: Recombinant annexin B30 did not confer significant protection against the parasite. The molecule may not be suitable for vaccine development. However, it could be an ideal biomarker recommended for immunodiagnostics development.
Fulltext Diagnostic and therapeutic potential of shark variable new antigen receptor (VNAR) single domain antibody

Cheong WS, Leow CY, Abdul Majeed AB, Leow CH

Int J Biol Macromol, 2020 Mar 15;147:369-375.
PMID: 31926922 DOI: 10.1016/j.ijbiomac.2020.01.039

Conventional monoclonal antibodies (mAbs) have been widely used in research and diagnostic applications due to their high affinity and specificity. However, multiple limitations, such as large size, complex structure and sensitivity to extreme ambient temperature potentially weaken the performance of mAbs in certain applications. To address this problem, the exploration of new antigen binders is extensively required in relation to improve the quality of current diagnostic platforms. In recent years, a new immunoglobulin-based protein, namely variable domain of new antigen receptor (VNAR) was discovered in sharks. Unlike conventional mAbs, several advantages of VNARs, include small size, better thermostability and peculiar paratope structure have attracted interest of researchers to further explore on it. This article aims to first present an overview of the shark VNARs and outline the characteristics as an outstanding new reagent for diagnostic and therapeutic applications.
Designing and evaluation of an antibody-targeted chimeric recombinant vaccine encoding Shigella flexneri outer membrane antigens

Kazi A, Hisyam Ismail CMK, Anthony AA, Chuah C, Leow CH, Lim BH, et al.

Infect Genet Evol, 2020 06;80:104176.
PMID: 31923724 DOI: 10.1016/j.meegid.2020.104176

Shigellosis is one of the most common diseases found in the developing countries, especially those countries that are prone flood. The causative agent for this disease is the Shigella species. This organism is one of the third most common enteropathogens responsible for childhood diarrhea. Since Shigella can survive gastric acidity and is an intracellular pathogen, it becomes difficult to treat. Also, uncontrolled use of antibiotics has led to development of resistant strains which poses a threat to public health. Therefore, there is a need for long term control of Shigella infection which can be achieved by designing a proper and effective vaccine. In this study, emphasis was made on designing a candidate that could elicit both B-cell and T-cell immune response. Hence B- and T-cell epitopes of outer membrane channel protein (OM) and putative lipoprotein (PL) from S. flexneri 2a were computationally predicted using immunoinformatics approach and a chimeric construct (chimeric-OP) containing the immunogenic epitopes selected from OM and PL was designed, cloned and expressed in E. coli system. The immunogenicity of the recombinant chimeric-OP was assessed using Shigella antigen infected rabbit antibody. The result showed that the chimeric-OP was a synthetic peptide candidate suitable for the development of vaccine and immunodiagnostics against Shigella infection.
Differential expression of cytokine genes in THP-1-derived macrophages infected with mild and virulence strains of Shigella flexneri 2a

Shabani NRM, Mokhtar M, Leow CH, Lean QY, Chuah C, Singh KKB, et al.

Infect Genet Evol, 2020 11;85:104532.
PMID: 32911076 DOI: 10.1016/j.meegid.2020.104532

Shigella is an intracellular bacterial pathogen that causes bacterial dysentery called shigellosis. The assessment of pro- and anti-inflammatory mediators produced by immune cells against this bacteria are vital in identifying the effectiveness of the immune reaction in protecting the host. In Malaysia, Shigella is ranked as the third most common bacteria causing diarrheal disease among children below 5 years old. In the present study, we aim to examine the differential cytokine gene expressions of macrophages in response to two types of clinical strains of Shigella flexneri 2a (S. flexneri 2a) isolated from patients admitted in Hospital Universiti Sains Malaysia, Kelantan, Malaysia. THP-1-derived macrophages, as the model of human macrophages, were infected separately with S. flexneri 2a mild (SH062) and virulence (SH057) strains for 6, 12, and 24 h, respectively. The gene expression level of inflammatory mediators was identified using real-time quantitative polymerase chain reaction (RT-qPCR). The production of nitric oxide (NO) by the macrophages was measured by using a commercialized NO assay kit. The ability of macrophages to kill the intracellular bacteria was assessed by intracellular killing assay. Induction of tumor necrosis factor-alpha (TNFα), interleukin (IL)-1β, IL-6, IL-12, inducible NO synthase (iNOS), and NO, confirmed the pro-inflammatory reaction of the THP-1-derived macrophages in response to S. flexneri 2a, especially against the SH507 strain. The SH057 also induced a marked increase in the expression levels of the anti-inflammatory cytokine mRNAs at 12 h and 24 h post-infection. In the intracellular killing assay, both strains showed less viable, indicating the generation of pro-inflammatory cytokines in the presence of iNOS and NO was crucial in the stimulation of macrophages for the host defense against shigellosis. Transcription analysis of THP-1-derived macrophages in this study identifies differentially expressed cytokine genes that correlated with the virulence factor of S. flexneri 2a.

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