Cloning and characterization of bovine growth hormone (bGH) gene from Malaysian gaur (Bos gaurus hubbacla)

Total genomic DNA from Malaysian gaur (Bos gaurus hubbacla) was isolated from whole blood. The genomic DNA was then subjected to PCR amplification using specific primers for bovine growth hormone gene (bGH). This PCR product which represents part of the bGH gene was cloned into pCR® 2.1 TOPO® cloning vector before sent for sequencing analysis. The nucleotide sequence of the gene coding regions was found to be identical with that of GenBank database sequence of Bos gaurus growth hormone mRNA sequence except for two nucleotides position (position 101 and 128). The current database of Bos gaurus growth hormone mRNA sequence at GenBank showed one unknown nucleotide at position 128 which denoted as N in the sequence. The finding of this study determined that the nucleotide base at position 128 is cytosine (C). The entire cloned gene sequence is 665 bp in length and contains 333 bp coding regions that code for 110 amino acid residue. The in-silico restriction mapping site analysis carried out in this study revealed that 70 enzymes produce unique site when they cut the sequence. A comparison of the cloned Malaysian gaur bGH gene with Bos taurus (local cattle) growth hormone mRNA sequence also showed a high homology between both sequences

Avidez do IgG pelo Western Blot usando o r GRA-7 do Toxoplasma gondii clonado de nucleotídeos 39-711 para sorodiagnóstico de toxoplasmose aguda

Toxoplasmosis is an important cause of congenital infection. The present study was performed to evaluate the usefulness of recombinant (r) GRA-7 cloned from nucleotides (n) 39-711 in discriminating between acute and chronic toxoplasmosis. First, commercial IgM, IgG and IgG avidity ELISAs were used to determine the serological profile of the sera. Serum samples were from 20 symptomatic patients with acute infection (low IgG avidity, IgM positive), 10 with chronic infection (high IgG avidity, IgM negative) and 10 with indeterminate IgG avidity (IgM positive) which were tested for IgG avidity status with an in-house developed IgG avidity Western blot using the rGRA-7 recombinant antigen. All 20 sera from cases of probable acute infection showed bands which either faded out completely or reduced significantly in intensity after treatment with 8 M urea, whereas the band intensities of the 10 serum samples from chronic cases remained the same. Of the 10 sera with indeterminate IgG avidity status, after treatment with 8 M urea the band intensities with six sera remained the same, two sera had completely faded bands and another two sera had significantly reduced band intensities. Discrimination between acute and chronic toxoplasmosis was successfully performed by the in-house IgG avidity Western blot.Toxoplasmose é uma causa importante de infecção congênita. O presente estudo foi feito para avaliar o uso do recombinante (r) GRA-7 clonado de nucleotídeos (n) 30-711 para discriminar entre toxoplasmose aguda e crônica. Inicialmente IgM, IgG e ELISA avidez IgG comerciais foram usados para determinar o perfil sorológico do soro. Amostras de soro de 20 pacientes sintomáticos com infecção aguda (IgG avidez baixa, IgM positivo), 10 com infecção crônica (alta avidez IgG, IgM negativo) e 10 com avidez IgG indeterminada (IgM positivo) que foram testados para o status de avidez IgG com um doméstico Western Blot desenvolvendo avidez IgG usando o rGRA-7 antígeno recombinante. Todos os 20 soros de provável infecção aguda mostraram bandas que ou se apagaram completamente ou tiveram a sua intensidade significantemente reduzida após tratamento com uréia 8 M, enquanto as intensidades das bandas das 10 amostras de soros de casos crônicos permaneceram iguais. Dos 10 soros com status indeterminado de avidez de IgG, após tratamento com uréia 8 M a intensidade das bandas em seis soros permaneceram iguais, dois soros tiveram bandas apagadas completamente e dois outros tiveram significante redução da intensidade das bandas. Discriminação entre toxoplasmose aguda e crônica foi feita com sucesso através do IgG avidez Western blot doméstico

Simultaneous amperometric aptasensor based on diazonium grafted screen-printed carbon electrode for detection of CFP10 and MPT64 biomarkers for early tuberculosis diagnosis

Early diagnosis is highly crucial for life-saving and transmission management of tuberculosis (TB). Despite the low sensitivity and time-consuming issues, TB antigen detection still relies on conventional smear microscopy and culture techniques. To address this limitation, we report the development of the first amperometric dual aptasensor for the simultaneous detection of Mycobacterium tuberculosis secreted antigens CFP10 and MPT64 for better diagnosis and control of TB. The developed sensor was based on the aptamers–antibodies sandwich assay and detected by chronoamperometry through the electrocatalytic reaction between peroxidase-conjugated antibodies, H2O2, and hydroquinone. The CFP10 and MPT64 aptamers were immobilized via carbodiimide covalent chemistry over the disposable dual screen-printed carbon electrodes modified with a 4-carboxyphenyl diazonium salt. Under optimized conditions, the aptasensor achieved a detection limit of 1.68 ng mL−1 and 1.82 ng mL−1 for CFP10 and MPT64 antigens, respectively. The developed assay requires a small sample amount (5 µL) and can be easily performed within 2.5 h. Finally, the dual aptasensor was successfully applied to clinical sputum samples with the obtained diagnostic sensitivity (n = 24) and specificity (n = 13) of 100%, respectively, suggesting the readiness of the developed assay to be used for TB clinical application

Preliminary assessment of the diagnostic performances of a new rapid diagnostic test for the serodiagnosis of human cystic echinococcosis

Rapid diagnostic tests for cystic echinococcosis (CE) are convenient to support ultrasound diagnosis in uncertain cases, especially in resource-limited settings. We found comparable diagnostic performances of the experimental Hyd Rapid Test and the commercial VIRapid HYDATIDOSIS Test, used in our diagnostic laboratory, using samples from well-characterized hepatic CE cases

Production of Toxocara cati TES-120 Recombinant Antigen and Comparison with its T. canis Homolog for Serodiagnosis of Toxocariasis

Photograph of a scene at Ed Galloway's Totem Pole Park

Entamoeba infections and associated risk factors among migrant workers in peninsular Malaysia

The influx of low skilled migrant workers to Malaysia from low socio-economic countries where gastrointestinal parasitic infections are prevalent has raised concerns about transmission to the local population. Three methods for detection (serology, microscopy and molecular techniques) were utilized to identify Entamoeba infections amongst the targeted cohort and determine risk factors associated with infection. Serological screening of 484 migrant workers from five working sectors in Peninsular Malaysia using IgG4 ELISA based on the rPPDK antigen showed an overall seroprevalence of 7.4% (n = 36; CL95 = 5.3–10.1%) with only one factor statistically associated with seropositivity of anti-amoebic antibodies, i.e. years of residence in Malaysia (χ21 = 4.007, p = 0.045). Microscopic examination of 388 faecal samples for protozoan cysts and trophozoites showed a slightly higher prevalence (11.6%; n=45; CL95: 8.4–14.8%). Meanwhile, amplification of the 16S rDNA gene detected two species i.e. Entamoeba dispar (23/388; 5.9%; CL95: 3.6–8.3%) and E. histolytica (11/388; 2.8%; CL95: 1.2–4.5%) and mixed infections with both parasites in only three samples (3/388; 0.8%; CL95: 0.2–2.2%). Entamoeba dispar infection was significantly associated with those employed in food and domestic services (χ24 = 12.879, p = 0.012). However, none of the factors affected the prevalence of E. histolytica infection. Despite the low prevalence of E. histolytica in faecal samples of the study cohort, the presence of this pathogenic parasite still poses potential public health risks and calls for tighter control strategies based on better availability of chemotherapeutic treatment and accessibility to appropriate health education. © 2019, Malaysian Society for Parasitology. All rights reserved

Epidemiology and immunodiagnostics of Strongyloides stercoralis infections among migrant workers in Malaysia

To investigate the status of Strongyloides(S.) stercoralis infections among migrant workers in Malaysia for the first time and identify risk factors. Methods: Four diagnostic methods were employed for the detection of S. stercoralis including microscopy, enzyme-linked immunosorbent assay (ELISA) using a commercial kit, ELISA using the rSs1a antigen and polymerase chain reaction (PCR). Low and semi-skilled workers from five working sectors (i.e. manufacturing, food service, agriculture and plantation, construction and domestic service) were tested on a voluntary basis. Results: The overall seroprevalence of S. stercoralis from 483 workers employing the ELISA commercial kit for IgG was 35.8% (n=173; 95% CI: 31.5%-40.1%) whereas seroprevalence using the rSs1a-ELISA was 13.0% (n=63; 95% CI: 10.0%-16.0%). Cross tabulation between the ELISA commercial kit and rSs1a-ELISA showed that only 6.4% (n=31; 95% CI: 4.2%-8.6%) of the samples were positive in both tests. Microscopic examination of all 388 fecal samples were negative; however subsequent testing by a nested PCR against DNA from the same samples successfully amplified DNA from three male subjects (0.8%; 3/388). Male workers, India and Myanmar nationality, food service occupation and those living in the hostel were statistically significant with seroprevalence (P<0.005). Conclusion: This is the first report on the epidemiology of S. stercoralis infections among the migrant workers in Malaysia. Our results highlight the importance of using appropriate diagnostic tools for detection. The presence of anti-S. stercoralis antibodies in the study population calls for improvements in personal hygiene and sanitation standards among migrant workers in Malaysia through control strategies including health education campaigns and programs aimed at increasing awareness and healthy behaviors. © 2019 Asian Pacific Journal of Tropical Medicine Produced by Wolters Kluwer-Medknow. All rights reserved