Zeehaida Mohamed

Application of Real-Time Polymerase Chain Reaction in Detection of Entamoeba histolytica in Pus Aspirates of Liver Abscess Patients

Entamoeba histolytica is the second major cause of liver abscess disease in humans, particularly in developing countries. Recently, DNA molecular-based methods have been employed to enhance the detection of E. histolytica in either pus or stool specimens. In this study, the results of real-time polymerase chain reaction (PCR) to detect E. histolytica DNA in pus from liver abscess cases were compared with those of indirect hemagglutination assay on the corresponding serum samples. Bacterial cultures were also performed on the pus samples for the diagnosis of pyogenic liver abscess. The real-time PCR detected E. histolytica DNA in 23 of 30 (76.7%) pus samples, when compared with 14 of 30 (46.7%) serum samples in which anti-Entamoeba antibodies were detected by indirect hemagglutination assay and 4 of 30 (13.3%) pus samples that showed bacterial infection by culture. The use of real-time PCR is a promising detection method for diagnosis and epidemiology assessment of amoebic liver abscess.

Segmentation based approach for detection of malaria parasites using moving k-means clustering

Recent progress based on microscopic imaging has given significant contribution in diagnosis of malaria infection based on blood images. Due to the requirement of prompt and accurate diagnosis of malaria, the current study has proposed an unsupervised colour image segmentation of malaria parasites using moving k-means (MKM) clustering algorithm. It has been applied on malaria images of P. vivax species. The proposed segmentation method provides a basic step for detection of the presence of malaria parasites in thin blood smears. With the aim of obtaining the fully segmented red blood cells infected with malaria parasites, the malaria images will firstly enhanced by using the partial contrast stretching technique. Then, the MKM clustering algorithm has been applied on the saturation and intensity components of HSI (hue, saturation, intensity) colour space for segmenting the infected cell from the background. After that, the segmented images have been processed using median filter and seeded region growing area extraction algorithms for smoothing the image and removing any unwanted regions from the image, respectively. Finally, the holes inside the infected cell are filled by applying region filling based on morphological reconstruction algorithm. The proposed segmentation method has been analyzed using 100 malaria images which consist of the trophozoite and gametocyte stages. Overall, the results indicate that MKM clustering that has been performed on saturation component image has produced the best segmentation performance with segmentation accuracy of 99.49% compared to the intensity component image with segmentation accuracy of 98.89%.

Toxoplasma gondii excretory secretory antigenic proteins of diagnostic potential.

Saadatnia G, Mohamed Z, Ghaffarifar F, Osman E, Kazemi Moghadam Z, Noordin R. Toxoplasma gondii excretory secretory antigenic proteins of diagnostic potential. APMIS 2012; 120: 47–55.

Analysis of Entamoeba histolytica Excretory-Secretory Antigen and Identification of a New Potential Diagnostic Marker

ABSTRACT Serodiagnosis of amoebiasis remains the preferred method for diagnosis of amoebic liver abscess (ALA). However, the commercially available kits are problematic in areas of endemicity due to the persistently high background antibody titers. Human serum samples (n = 38) from patients with ALA who live in areas of endemicity were collected from Hospital Universiti Sains Malaysia during the period of 2008 to 2010. Western blots using excretory-secretory antigen (ESA) collected from axenically grown Entamoeba histolytica were probed with the above serum samples. Seven antigenic proteins of ESA with various reactivities were identified, i.e., 152 kDa, 131 kDa, 123 kDa, 110 kDa, 100 kDa, 82 kDa, and 76 kDa. However, only the 152-kDa and 110-kDa proteins showed sensitivities above 80% in the Western blot analysis. All the antigenic proteins showed undetectable cross-reactivity when probed with healthy human serum samples (n = 30) and serum samples from other infections (n = 33). From the matrix-assisted laser desorption ionization–two-stage time of flight (MALDI-TOF/TOF) analysis, the proteins were identified as heavy subunits of E. histolytica lectin and E. histolytica pyruvate phosphate dikinase, respectively. Use of the E. histolytica lectin for diagnosis of ALA has been well reported by researchers and is being used in commercialized kits. However, this is the first report on the potential use of pyruvate phosphate dikinase for diagnosis of ALA; thus, this molecule merits further evaluation on its diagnostic value using a larger panel of serum samples.

Comparison of antimicrobial resistance in neonatal and adult intensive care units in a tertiary teaching hospital

Intrahospital variations in antimicrobial profiles may be related to many factors. This study compared causative agents of nosocomial bloodstream infections between a neonatal intensive care unit (NICU) that adopted a ward-tailored antibiotic policy and adult intensive care units (ICUs). Data on organisms from blood cultures obtained from the respective wards between 2005 and 2009 were analyzed. Compared with the adult ICUs, the NICU had a higher frequency of Enterobacteriacae and lower frequencies of typical hospital-acquired pathogens (eg, Klebsiella pneumoniae, 17.4% vs 10.0% [P < .001]; Acinetobacter baumannii, 3.9% vs 11.6% [P < .001]). Antibiotic resistance of gram-negative organisms was also significantly lower in the NICU, including resistance to imipenem (5.7% vs 32.1%; P < .001), amikacin (8.8% vs 30.3%), and ceftriaxone (36.1% vs 74.6%; P < .001). This could possibly be due to the ward-tailored antibiotic policy adopted by the NICU but not by the other ICUs.

The prevalence and preventive measures of the respiratory illness among Malaysian pilgrims in 2013 hajj season

Abstract Background. Respiratory illness continues to exert a burden on hajj pilgrims in Makkah. The purpose of this study is to determine the prevalence of respiratory illness and its associated factors among Malaysian hajj pilgrims in 2013 and to describe its preventive measures. Methods. A cross-sectional study was conducted in Makkah and Malaysia during the 2013 hajj season. A self-administered proforma on social demographics, previous experience of hajj or umrah, smoking habits, co-morbid illness and practices of preventive measures against respiratory illness were obtained. Results. A total of 468 proforma were analysed. The prevalence of the respiratory illness was 93.4% with a subset of 78.2% fulfilled the criteria for influenza-like illness (ILI). Most of them (77.8%) had a respiratory illness of <2 weeks duration. Approximately 61.8% were administered antibiotics but only 2.1% of them had been hospitalized. Most of them acquired the infection after a brief stay at Arafat (81.2%). Vaccination coverages for influenza virus and pneumococcal disease were quite high, 65.2% and 59.4%, respectively. For other preventive measures practices, only 31.8% of them practiced good hand hygiene, ∼82.9% of pilgrims used surgical face masks, N95 face masks, dry towels, wet towels or veils as their face masks. Nearly one-half of the respondents (44.4%) took vitamins as their food supplement. Malaysian hajj pilgrims with previous experience of hajj (OR 0.24; 95% CI 0.10–0.56) or umrah (OR 0.19; 95% CI 0.07–0.52) and those who have practiced good hand hygiene (OR 0.35; 95% CI 0.16–0.79) were found to be significantly associated with lower risk of having respiratory illness. Otherwise, pilgrims who had contact with those with respiratory illness (OR 2.61; 95% CI 1.12–6.09) was associated with higher risk. Conclusions. The prevalence of respiratory illness remains high among Malaysian hajj pilgrims despite having some practices of preventive measures. All preventive measures which include hand hygiene, wearing face masks and influenza vaccination must be practiced together as bundle of care to reduce respiratory illness effectively.

Modified Global and Modified Linear Contrast Stretching Algorithms: New Colour Contrast Enhancement Techniques for Microscopic Analysis of Malaria Slide Images

Malaria is one of the serious global health problem, causing widespread sufferings and deaths in various parts of the world. With the large number of cases diagnosed over the year, early detection and accurate diagnosis which facilitates prompt treatment is an essential requirement to control malaria. For centuries now, manual microscopic examination of blood slide remains the gold standard for malaria diagnosis. However, low contrast of the malaria and variable smears quality are some factors that may influence the accuracy of interpretation by microbiologists. In order to reduce this problem, this paper aims to investigate the performance of the proposed contrast enhancement techniques namely, modified global and modified linear contrast stretching as well as the conventional global and linear contrast stretching that have been applied on malaria images of P. vivax species. The results show that the proposed modified global and modified linear contrast stretching techniques have successfully increased the contrast of the parasites and the infected red blood cells compared to the conventional global and linear contrast stretching. Hence, the resultant images would become useful to microbiologists for identification of various stages and species of malaria.

Detection of immunogenic parasite and host-specific proteins in the sera of active and chronic individuals infected with Toxoplasma gondii

Toxoplasma gondii infection in pregnant women may result in abortion and foetal abnormalities, and may be life‐threatening in immunocompromised hosts. To identify the potential infection markers of this disease, 2‐DE and Western blot methods were employed to study the parasite circulating antigens and host‐specific proteins in the sera of T. gondii‐infected individuals. The comparisons were made between serum protein profiles of infected (n=31) and normal (n=10) subjects. Antigenic proteins were identified by immunoblotting using pooled sera and monoclonal anti‐human IgM‐HRP. Selected protein spots were characterised using mass spectrometry. Prominent differences were observed when serum samples of T. gondii‐infected individuals and normal controls were compared. A significant up‐regulation of host‐specific proteins, α2‐HS glycoprotein and α1‐B glycoprotein, was also observed in the silver‐stained gels of both active and chronic infections. However, only α2‐HS glycoprotein and α1‐B glycoprotein in the active infection showed immunoreactivity in Western blots. In addition, three spots of T. gondii proteins were detected, namely (i) hypothetical protein chrXII: 3984434‐3 TGME 49, (ii) dual specificity protein phosphatase, catalytic domain TGME 49 and (iii) NADPH‐cytochrome p450 reductase TGME 49. Thus, 2‐DE approach followed by Western blotting has enabled the identification of five potential infection markers for the diagnosis of toxoplasmosis: three are parasite‐specific proteins and two are host‐specific proteins.

Evaluation of Entamoeba histolytica recombinant phosphoglucomutase protein for serodiagnosis of amoebic liver abscess

BackgroundAmoebic liver abscess (ALA) is the most frequent clinical presentation of extra-intestinal amoebiasis. The diagnosis of ALA is typically based on the developing clinical symptoms, characteristic changes on radiological imaging and serology. Numerous serological tests have been introduced for the diagnosis of ALA, either detecting circulating amoebic antigens or antibodies. However those tests show some pitfalls in their efficacy and/or the preparation of the tests are costly and tedious. The commercial IHA kit that used crude antigen was reported to be useful in diagnosis of ALA, however high antibody background in endemic areas may cause problems in its interpretation. Thus, discovery of well-defined antigen(s) is urgently needed to improve the weaknesses of current serodiagnostic tests.MethodsCrude antigen of E. histolytica was analysed by 2-DE and Western blot to identify a protein of diagnostic potential for ALA. The corresponding gene of the antigenic protein was then cloned, expressed and the purified recombinant protein was subsequently evaluated for serodiagnosis of ALA in an indirect ELISA format.ResultsAnalysis of crude antigen showed that phosphoglucomutase (PGM) has the diagnostic potential. Recombinant PGM (rPGM) showed 79.17% (19/24) sensitivity and 86.67% (195/225) specificity in diagnosis of ALA based on the COV of mean +1SD. There was no significant difference between rPGM-ELISA and IHA diagnostic kit in the diagnosis of ALA in terms of sensitivity and specificity at p-value < 0.05.ConclusionIn conclusion, rPGM-ELISA is found to be useful for serodiagnosis of ALA. Future studies will determine whether rPGM-ELISA also detects antibodies produced in amoebic dysentery and asymptomatic cases.

Ocular surface infections in northeastern state of malaysia: a 10-year review of bacterial isolates and antimicrobial susceptibility.

Objective: Ocular surface infections that include infections of conjunctiva, adnexa, and cornea have the potential risk of causing blindness within a given population. Empirical antibiotic therapy is usually initiated based on epidemiological data of common causative agents. Thus, the aims of this study were to determine the bacterial agents and their susceptibility patterns of isolates from ocular surface specimens in our hospital. Methods: This is a retrospective analysis and records of bacterial isolates from ocular surface specimens in Hospital Universiti Sains Malaysia from January 2001 to December 2010 were examined. Specimens were processed according to standard laboratory procedures. Antimicrobial susceptibility testing was conducted based on Clinical and Laboratory Standards Institute recommendations. Only single, nonrepetitive isolates were included in the analysis. Results: A total of 1,267 isolates were obtained during the study period, which comprised Staphylococcus aureus (n = 299, 23.6%), Pseudomonas aeruginosa (n = 194, 15.3%), Streptococcus pneumoniae (n = 108, 8.5%), Haemophilus influenzae (n = 100, 7.9%), Haemophilus parainfluenzae (n = 84, 6.6%), and Enterobacter spp. (n = 81, 6.4%). Fungi contributed to 4.4% of the total isolates. The antimicrobial susceptibility testing demonstrated that gram-positive bacteria were generally resistant to gentamicin (19%–57%), whereas gram-negative bacteria were resistant to chloramphenicol (27%–58%). Conclusions: Based on the above results, knowledge of the initial Gram stain findings is imperative before the commencement of empirical antibiotic therapy. Therefore, a simple Gram staining for all eye specimens is highly recommended.