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In vitro demonstration of the hemolytic, cytotoxic activities and induction of apoptosis in Orientia tsutsugamushi infected L929 mouse fibroblast cells

Tay ST, Rohani MY, Ho TM, Devi S

Southeast Asian J Trop Med Public Health, 2003 Jun;34(2):352-6.
PMID: 12971561

Using cultured mouse fibroblast L929 cells, this study demonstrated the hemolytic and cytotoxic activities and induction of apoptosis in cells infected with Orientia tsutsugamushi. Low levels of hemolytic activity were detected using heavily infected cells. No hemolysin or cytotoxin were detected in the infected culture fluid regardless of the pathogenicity of the O. tsutsugamushi strains in mice. Using propidium iodide uptake assay, acridine orange/ethidium bromide staining and terminal deoxynucleotide transferase-mediated dUTP-digoxigenin nick-end labeling assay, apoptosis was observed in L929 cells infected with Karp and Gilliam strains.
Antibodies to Orientia tsutsugamushi, Rickettsia typhi and spotted fever group rickettsiae among febrile patients in rural areas of Malaysia

Tay ST, Ho TM, Rohani MY, Devi S

Trans R Soc Trop Med Hyg, 2000 9 7;94(3):280-4.
PMID: 10974999

A serosurvey was conducted in 1995-97 among 1596 febrile patients from 8 health centres in Malaysia for antibodies against Orientia tsutsugamushi (OT), Rickettsia typhi (RT) and TT118 spotted fever group rickettsiae (SFGR) by using an indirect immunoperoxidase assay. A total of 51.4% patients had antibody against at least 1 of those rickettsiae. Antibody to SFGR was most prevalent (42.5%), followed by RT (28.1%) and OT (24.9%). The seroprevalences of antibodies to SFGR, RT or OT alone were 12.4, 3.6 and 4.3%, respectively. Antibodies against more than 1 species of rickettsiae were presence in 31.1% of the patients, suggesting the possibility of co-infection, previous exposures or serological cross-reactivities. Seroprevalence of the various rickettsiae varied according to locality, with SFGR antibodies being the most prevalent in most areas. There was no significant association of prevalence of rickettsial antibody with gender. The seroprevalence of OT, SFGR and RT increased with patient age but an increase of antibody titre with age was not significant. Those working in the agricultural sectors had significantly higher seroprevalence of OT, SFGR and RT than those not related with agricultural activities. Scrub typhus remains a public health problem with an estimated annual attack rate of 18.5%. Tick typhus and murine typhus as shown in this serosurvey appear much more widespread than scrub typhus in this country.
Detection of haemolytic activity of campylobacters by agarose haemolysis and microplate assay

Tay ST, Devi S, Puthucheary SD, Kautner IM

J Med Microbiol, 1995 Mar;42(3):175-80.
PMID: 7884798

There are several methods for the detection of haemolytic activity in campylobacters. However, we found the haemolytic effect of campylobacters on conventional blood agar plates to be variable, inconsistent and difficult to interpret. Blood agarose plates showed campylobacter haemolytic activity more clearly. The incubation conditions (temperature and gaseous) appear to be important for the expression of this activity. Ninety four percent of the Campylobacter isolates examined were found to be haemolytic by the microplate assay with minimal haemolytic units that ranged from 1 to 64. Haemolytic activity was detected only from live bacterial cultures and not from any of the 50 bacterial culture supernates, which suggests that campylobacters may possess a cell-associated haemolysin. The identification of such haemolytic activity in a large number of campylobacters (94%) suggests its potential role as a virulence factor in campylobacter gastroenteritis.
Fulltext Exotoxin profiles of Campylobacters isolated in Malaysia

Tee TS, Devi S, Puthucheary SD, Kautner IM

Southeast Asian J Trop Med Public Health, 1994 Sep;25(3):443-8.
PMID: 7777904

Approximately 57% of clinical and 33% of poultry isolates examined produced a cytotoxin. Cytotoxic activity was detected in 25 (50%) isolates of Campylobacter of which 12 were isolated from bloody diarrhea and 9 from watery stools. The cytotoxin titers were low, ranging from 2 to 16. The crude filtrates from 50 Campylobacter isolates showed no cytotoxic effect in Vero cells, no fluid accumulation in suckling mice and no hemolytic activity.
Epitope mapping of the Sta58 major outer membrane protein of Rickettsia tsutsugamushi

Lachumanan R, Devi S, Cheong YM, Rodda SJ, Pang T

Infect Immun, 1993 Oct;61(10):4527-31.
PMID: 7691753

Binding studies of 160 overlapping, synthetic octapeptides from the hydrophilic regions of the Sta58 major outer membrane protein of Rickettsia tsutsugamushi with sera from patients with scrub typhus revealed 15 immunodominant peptides which are recognized by all the sera tested. Further analysis of the specificity of peptide binding with five of these peptides indicated that the peptides showed significantly stronger binding to scrub typhus patients' sera than they did to sera from patients with other febrile illnesses common in the region, i.e., malaria, dengue fever, typhoid fever, and leptospirosis. The main antibody class binding to these peptides appears to be immunoglobulin M, and there appears to be little correlation between reactivity with peptides and antibody titers measured by the indirect immunoperoxidase test.
Isolation and PCR detection of rickettsiae from clinical and rodent samples in Malaysia

Tay ST, Rohani MY, Ho TM, Devi S

Southeast Asian J Trop Med Public Health, 2002 Dec;33(4):772-9.
PMID: 12757225

Isolation of rickettsiae from patients' blood samples and organ samples of wild rodents from areas with high seroprevalence of rickettsial infections was attempted using cell culture assay and animal passages. L929 mouse fibroblast cells grown in 24 well tissue culture plate were inoculated with buffy coat of febrile patients and examined for the growth of rickettsiae by Giemsa, Gimenez staining and direct immunofluorescence assay. No rickettsiae were isolated from 48 patients' blood samples. No symptomatic infections were noted in mice or guinea pigs infected with 50 organ samples of wild rodents. There was no rickettsial DNA amplified from these samples using various PCR detection systems for Orientia tsutsugamushi, typhus and spotted fever group rickettsiae.
Immunogenic epitopes of Salmonella typhi GroEL heat shock protein reactive with both monoclonal antibody and patients sera

Panchanathan V, Naidu BR, Devi S, Di Pasquale A, Mason T, Pang T

Immunol Lett, 1998 Jun;62(2):105-9.
PMID: 9698106

A series of 122, 9-mer overlapping peptides based on the sequence of the Salmonella typhi GroEL gene was synthesized on the surfaces of polyethylene pins and screened with monoclonal antibody to GroEL and with human sera from patients with typhoid fever and normal healthy blood donors. Three immunogenic epitopes corresponding to peptides EGQDRGYSY, YSYNKETGE and GKGTEEKEK were identified upon screening with the human sera. In addition, screening of the peptides with a monoclonal antibody to GroEL detected binding to a third peptide, KGGKGTEEK, which contains a common overlapping sequence to peptide GKGTEEKEK. Identification and definition of these epitopes will be important in delineating the biological and immunological functions of this protein and in designing better diagnostic tests and vaccines.
Fulltext Serological evidence of natural infection of wild rodents (Rattus spp and Tupaia glis) with rickettsiae in Malaysia

Tay ST, Kaewanee S, Ho TM, Rohani MY, Devi S

Southeast Asian J Trop Med Public Health, 1998 Sep;29(3):560-2.
PMID: 10437957
Expression of recombinant proteins of Orientia tsutsugamushi and their applications in the serodiagnosis of scrub typhus

Tay ST, Rohani MY, Ho TM, Devi S

Diagn Microbiol Infect Dis, 2002 Oct;44(2):137-42.
PMID: 12458119

In this study, recombinant proteins that encompassed the AD I-AD III regions of 56 kDa immunodominant gene of 2 Orientia tsutsugamushi (OT) serotypes; Gilliam and TA763 were expressed in Escherichia coli. Both recombinant proteins exhibited serologic cross-reactivity with the rabbit antisera against various OT serotypes, as evaluated by enzyme-linked immunosorbent assay (ELISA), but not against other rickettsial species, including Rickettsia typhi, R. prowazekii and TT118 SFG rickettsiae. The feasibility of using the recombinant proteins as a diagnostic reagent was further evaluated by ELISA using sera from blood donors and scrub typhus patients. The results suggested a higher affinity of the antihuman IgM than IgG with both recombinant proteins. The IgM ELISA findings were agreeable with the results of indirect immunoperoxidase (IIP) assay especially with sera of high antibody (1:1600). However, more than one antigen are probably needed for development of an effective assay for serodiagnosis of scrub typhus in endemic areas.
Helicobacter pylori protein HP0986 (TieA) interacts with mouse TNFR1 and triggers proinflammatory and proapoptotic signaling pathways in cultured macrophage cells (RAW 264.7)

Ansari SA, Devi S, Tenguria S, Kumar A, Ahmed N

Cytokine, 2014 Aug;68(2):110-7.
PMID: 24767863 DOI: 10.1016/j.cyto.2014.03.006

HP0986 protein of Helicobacter pylori has been shown to trigger induction of proinflammatory cytokines (IL-8 and TNF-α) through the activation of NF-κB and also to induce Fas mediated apoptosis of human macrophage cells (THP-1). In this study, we unravel mechanistic details of the biological effects of this protein in a murine macrophage environment. Up regulation of MCP-1 and TNF-α in HP0986-induced RAW 264.7 cells occurred subsequent to the activation and translocation of NF-κB to the cell nucleus. Further, HP0986 induced apoptosis of RAW 264.7 cells through Fas activation and this was in agreement with previous observations made with THP-1 cells. Our studies indicated activation of TNFR1 through interaction with HP0986 and this elicited the aforementioned responses independent of TLR2, TLR4 or TNFR2. We found that mouse TNFR1 activation by HP0986 facilitates formation of a complex comprising of TNFR1, TRADD and TRAF2, and this occurs upstream of NF-κB activation. Furthermore, FADD also forms a second complex, at a later stage, together with TNFR1 and TRADD, resulting in caspase-8 activation and thereby the apoptosis of RAW 264.7 cells. In summary, our observations reveal finer details of the functional activity of HP0986 protein in relation to its behavior in a murine macrophage cell environment. These findings reconfirm the proinflammatory and apoptotic role of HP0986 signifying it to be an important trigger of innate responses. These observations form much needed baseline data entailing future in vivo studies of the functions of HP0986 in a murine model.
Dengue infection in pregnancy: prevalence, vertical transmission, and pregnancy outcome

Tan PC, Rajasingam G, Devi S, Omar SZ

Obstet Gynecol, 2008 May;111(5):1111-7.
PMID: 18448743 DOI: 10.1097/AOG.0b013e31816a49fc

To estimate prevalence rate of recent dengue infection in parturients, as well as the vertical transmission rate, and to compare pregnancy outcomes among infected women.
Fulltext Formaldehyde-inactivated whole-virus vaccine protects a murine model of enterovirus 71 encephalomyelitis against disease

Ong KC, Devi S, Cardosa MJ, Wong KT

J Virol, 2010 Jan;84(1):661-5.
PMID: 19864378 DOI: 10.1128/JVI.00999-09

Enterovirus 71 (EV71) causes childhood hand, foot, and mouth disease and neurological complications, and no vaccines or therapeutic drugs are currently available. Formaldehyde-inactivated whole-virus vaccines derived from EV71 clinical isolates and a mouse-adapted virus (MAV) were tested in a mouse model of EV71 encephalomyelitis. After only two immunizations, given to mice at 1 and 7 days of age, the MAV vaccine protected mice at 14 days of age from disease. Tissues from immunized mice were negative for virus by viral culture, reverse transcriptase PCR, immunohistochemistry analysis, and in situ hybridization. Cross-neutralizing EV71 antibodies to strains with genotypes B3, B4, and C1 to C5 generated in immunized adult mice were able to passively protect 14-day-old mice from disease.
Rapid detection, serotyping and quantitation of dengue viruses by TaqMan real-time one-step RT-PCR

Kong YY, Thay CH, Tin TC, Devi S

J Virol Methods, 2006 Dec;138(1-2):123-30.
PMID: 17000012 DOI: 10.1016/j.jviromet.2006.08.003

The use of the polymerase chain reaction (PCR) in molecular diagnosis is now accepted worldwide and has become an essential tool in the research laboratory. In the laboratory, a rapid detection, serotyping and quantitation, one-step real-time RT-PCR assay was developed for dengue virus using TaqMan probes. In this assay, a set of forward and reverse primers were designed targeting the serotype conserved region at the NS5 gene, at the same time flanking a variable region for all four serotypes which were used to design the serotype-specific TaqMan probes. This multiplex one-step RT-PCR assay was evaluated using 376 samples collected during the year 2003. These groups included RNA from prototype dengue virus (1-4), RNA from acute serum from which dengue virus was isolated, RNA from tissue culture supernatants of dengue virus isolated, RNA from seronegative acute samples (which were culture and IgM negative) and RNA from samples of dengue IgM positive sera. The specificity of this assay was also evaluated using a panel of sera which were positive for other common tropical disease agents including herpes simplex virus, cytomegalovirus, measles virus, varicella-zoster virus, rubella virus, mumps virus, WWF, West Nile virus, Japanese encephalitis virus, S. typhi, Legionella, Leptospira, Chlamydia, and Mycoplasma. The sensitivity, specificity and real-time PCR efficiency of this assay were 89.54%, 100% and 91.5%, respectively.
Peptide mimotopes of complex carbohydrates in Salmonella enterica serovar typhi which react with both carbohydrate-specific monoclonal antibody and polyclonal sera from typhoid patients

Thong KL, Tang SS, Tan WS, Devi S

Microbiol. Immunol., 2007;51(11):1045-52.
PMID: 18037781

Polyclonal sera from typhoid patients and a monoclonal antibody, mAb ATVi, which recognizes the capsular polysaccharide Vi antigen (ViCPS), were used to select for peptides that mimic the ViCPS by using a phage-displayed random 12-mer peptide library. Two major common mimotopes selected from the library carried the amino acid sequences TSHHDSHGLHRV and ENHSPVNIAHKL. Enzyme-linked immunosorbent assays (ELISAs) showed that these peptides carry mimotopes to ViCPS. Phage clones that contained the 12-mer peptides were also tested against pooled/individual typhoid patients' sera and found to have 3 to 5 times higher binding compared to normal sera. By using Phage-ELISA assays, the derived synthetic peptides, TSHHDSHGLHRV and ENHSPVNIAHKL, were tested against a monoclonal antibody mAb ATVi and over 2-fold difference in binding was found between these peptides and a control unrelated peptide, CTLTTKLYC. Inhibition of the mAb's binding to ViCPS indicated that the synthetic peptides successfully competed with the capsular polysaccharide for antibody binding.
Fulltext Development and evaluation of a one-step SYBR-Green I-based real-time RT-PCR assay for the detection and quantification of Chikungunya virus in human, monkey and mosquito samples

Ummul Haninah A, Vasan SS, Ravindran T, Chandru A, Lee HL, Shamala Devi S

Trop Biomed, 2010 Dec;27(3):611-23.
PMID: 21399603 MyJurnal

This paper reports the development of a one-step SYBR-Green I-based realtime RT-PCR assay for the detection and quantification of Chikungunya virus (CHIKV) in human, monkey and mosquito samples by targeting the E1 structural gene. A preliminary evaluation of this assay has been successfully completed using 71 samples, consisting of a panel of negative control sera, sera from healthy individuals, sera from patients with acute disease from which CHIKV had been isolated, as well as monkey sera and adult mosquito samples obtained during the chikungunya fever outbreak in Malaysia in 2008. The assay was found to be 100-fold more sensitive than the conventional RT-PCR with a detection limit of 4.12x10(0) RNA copies/μl. The specificity of the assay was tested against other related viruses such as Dengue (serotypes 1-4), Japanese encephalitis, Herpes Simplex, Parainfluenza, Sindbis, Ross River, Yellow fever and West Nile viruses. The sensitivity, specificity and efficiency of this assay were 100%, 100% and 96.8% respectively. This study on early diagnostics is of importance to all endemic countries, especially Malaysia, which has been facing increasingly frequent and bigger outbreaks due to this virus since 1999.
Fulltext Exploring the impact of emotionalized learning experiences on the affective domain: A comprehensive analysis

Kuo YK, Batool S, Devi S, Tahir T, Yu J

Heliyon, 2024 Jan 15;10(1):e23263.
PMID: 38163212 DOI: 10.1016/j.heliyon.2023.e23263

The central aim of this research endeavor was to delve into the profound influence of affective learning experiences on the cognitive and psychomotor domains. Concurrently, the study sought to discern the effects of these experiences on students' academic accomplishments within the three domains. This differentiation was predicated upon the interplay between pedagogical resources and the multifaceted dimensions of cognitive, affective, and social contexts for learning. Over the course of four consecutive semesters, data were meticulously collected from a cohort of 140 undergraduates enrolled at a private-sector university. The experimental cohorts 1, 2 and 3, comprising 35, 46, and 31 students, respectively, were instructed through distinct methodologies - namely, activity learning, reflective learning, and collaborative learning. In parallel, the control group, composed of 28 students, received instruction via the discussion method. The findings eloquently established a robust and affirmative linear correlation between affective experiences and achievements within the cognitive and psychomotor domains. This correlation notably underscored the far-reaching influence of the affective domain upon its cognitive and psychomotor counterparts. Furthermore, the variable of emotional context for learning emerged as a conspicuously noteworthy determinant of students' achievements across all three domains. In contrast, the remaining independent variables - namely, pedagogical resources, cognitive learning context, and social learning context - did not exhibit a substantial contribution. However, it was observed that the amalgamation of all four independent variables yielded a statistically significant relationship with achievements within the cognitive, affective, and psychomotor domains. This underscored the imperative nature of accounting for all pertinent factors when formulating instructional approaches that yield optimal outcomes.
Fulltext Detection of Adherence of Enterococcus faecalis in Infected Dentin of Extracted Human Teeth Using Confocal Laser Scanning Microscope: An In vitro Study

Nair VS, Nayak M, Ramya MK, Sivadas G, Ganesh C, Devi SL, et al.

J Pharm Bioallied Sci, 2017 Nov;9(Suppl 1):S41-S44.
PMID: 29284934 DOI: 10.4103/jpbs.JPBS_92_17

AIM: The aim of this study is to detect in vitro adherence of Enterococcus faecalis to the infected dentinal tubules of human extracted teeth using confocal laser scanning microscope.
SUBJECTS AND METHODS: Roots from human premolar teeth (n = 40) were infected with E. faecalis strain the American Type Culture Collection 29212 in brain heart infusion for 21 days. After the experimental period, specimens were divided into two groups, Group A (n = 20), Group B (n = 20), and Group A specimens were stained with fluorescein diacetate dye for the detection of viability and adherence Group B were stained with acridine orange dye for detection of metabolic activity and adherence. Samples were washed, thoroughly sectioned and examined by confocal laser scanning microscopy. Computer-assisted determinants of fluorescence, bacterial viability, metabolic activity, and adherence were compared statistically.
RESULTS: E. faecalis was able to invade the dentinal tubules to a depth of 1-400 μm and adhere to 1-200 μm depth. Adherence (90%) was significantly higher in 1-100 μm using fluorescein diacetate and acridine orange dye.
CONCLUSION: Adherence of E. faecalis as evaluated by confocal laser scanning microscope was highest at the depth of 1-100 μm which may have an impact on the shaping and cleaning procedures on the root canal.
Fulltext Melioidosis in acute cholangitis of diabetic patient: a forgotten diagnosis

Mohamad N, Ponnusamy S, Devi S, Manikam R, Idrus II, Bakar NHA

Res Rep Trop Med, 2012;3:103-106.
PMID: 30100777 DOI: 10.2147/RRTM.S34483

Melioidosis presents with a wide range of clinical presentations, which include severe community-acquired pneumonia, septicemia, central nervous system infection, and less severe soft tissue infection. Hence, its diagnosis depends heavily on the clinical microbiology laboratory for culture. In this case report, we describe an atypical presentation of melioidosis in a 52-year-old man who had fever, right upper-abdominal pain, and jaundice for 15 days. Melioidosis caused by Burkholderia pseudomallei was subsequently diagnosed from blood culture. As a primary care physician, high suspicion index is of great importance. High suspicion index of melioidosis in a high-risk group patient, such as the patient with diabetes mellitus and diabetic foot, is crucial in view of atypical presentations of pseudomonas sepsis. A correct combination of antibiotic administration in the early phase of therapy will determine its successful outcome.
Dengue virus regulates type I interferon signalling in a strain-dependent manner in human cell lines

Umareddy I, Tang KF, Vasudevan SG, Devi S, Hibberd ML, Gu F

J Gen Virol, 2008 Dec;89(Pt 12):3052-3062.
PMID: 19008393 DOI: 10.1099/vir.0.2008/001594-0

Outbreaks of dengue disease are constant threats to tropical and subtropical populations but range widely in severity, from mild to haemorrhagic fevers, for reasons that are still elusive. We investigated the interferon (IFN) response in infected human cell lines A549 and HepG2, using two strains (NGC and TSV01) of dengue serotype 2 (DEN2) and found that the two viruses exhibited a marked difference in inducing type I IFN response. While TSV01 infection led to activation of type I antiviral genes such as EIF2AK2 (PKR), OAS, ADAR and MX, these responses were absent in NGC-infected cells. Biochemical analysis revealed that NGC but not TSV01 suppressed STAT-1 and STAT-2 activation in response to type I IFN (alpha and beta). However, these two strains did not differ in their response to type II IFN (gamma). Although unable to suppress IFN signalling, TSV01 infection caused a weaker IFN-beta induction compared with NGC, suggesting an alternative mechanism of innate immune escape. We extended our study to clinical isolates of various serotypes and found that while MY10245 (DEN2) and MY22713 (DEN4) could suppress the IFN response in a similar fashion to NGC, three other strains of dengue [EDEN167 (DEN1), MY02569 (DEN1) and MY10340 (DEN2)] were unable to suppress the IFN response, suggesting that this difference is strain-dependent but not serotype-specific. Our report indicates the existence of a strain-specific virulence factor that may impact on disease severity.
Fulltext Comparison of safety and immunogenicity of a Vi polysaccharide typhoid vaccine with a whole-cell killed vaccine in Malaysian Air Force recruits

Panchanathan V, Kumar S, Yeap W, Devi S, Ismail R, Sarijan S, et al.

Bull World Health Organ, 2001;79(9):811-7.
PMID: 11584728

To carry out a comparative study of the safety and immunogenicity of Vi polysaccharide vaccine against whole-cell killed (WCK) typhoid vaccine.

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