Andrographolide effect on both Plasmodium falciparum infected and non infected RBCs membranes

Objective: To explore whether its antiplasmodium effect of andrographolide is attributed to its plausible effect on the plasma membrane of both Plasmodium falciparum infected and non-infected RBCs. Methods: Anti-plasmodium effect of andrographolide against Plasmodium falciparum strains was screened using the conventional malaria drug sensitivity assay. The drug was incubated with uninfected RBCs to monitor its effect on their morphology, integrity and osmotic fragility. It was incubated with the plasmodium infected RBCs to monitor its effect on the parasite induced permeation pathways. Its effect on the potential of merozoites to invade new RBCs was tested using merozoite invasion assay. Results: It showed that at andrographolide was innocuous to RBCs at concentrations approach its therapeutic level against plasmodia. Nevertheless, this inertness was dwindled at higher concentrations. Conclusions: In spite of its success to inhibit plasmodium induced permeation pathway and the potential of merozoites to invade new RBCs, its anti-plasmodium effect can't be attributed to these functions as they were attained at concentrations higher than what is required to eradicate the parasite. Consequently, other mechanisms may be associated with its claimed actions

Monoclonal antibodies: A review of therapeutic applications and future prospects

The increasing demand for monoclonal antibodies (mAbs) used for diagnostic and therapeutic applications has led to the development of large scale manufacturing processes, with improvements in production achieved through continuous  optimization of the inherent systems. The number of monoclonal antibodies (mAbs) that have already been approved for therapeutic applications and for use in clinical trials have significantly increased in the past few years. In view of the side effects and limitations of mAbs, several improvements and modifications to monoclonal antibodies have been developed. These modifications have facilitated the use of mAbs in various forms of therapeutic applications such as treatment of infectious diseases caused by bacterial, viral, fungal and parasitic organisms. Monoclonal antibodies have also been applied in the treatment of non-infectious diseases such as cancer, immune diseases, arthritis and other disorders resulting from organ transplantation. This review highlights mAbs applications in biomedicine, and discusses state-of-the-art technologies related to their potential uses.Keywords: Monoclonal antibodies, Therapeutic application, Infectious diseases, Cancer, Auto-immune diseases, Metabolic disorder

Molecular detection of Strongyloides ratti in faecal samples from wild rats in Serdang, Malaysia

Purpose: To detect Strongyloides ratti in faecal samples using conventional methods and to confirm the identification using a sensitive and specific method, namely, polymerase chain reaction (PCR). Methods: A PCR method targeting the small subunit of the rRNA gene was performed in this study for the detection of DNA from Strongyloides ratti (an animal model of S. stercoralis) in faecal samples of wild Brown rats, Rattus norvegicus. Results: Strongyloides ratti was detected in 34.2 % of collected rats by different conventional techniques and confirmed by PCR. The essay presented 100 % sensitivity with Strongyloides universal primer. Conclusion: The findings of this study suggest that the application of PCR with universal primer is a very sensitive methodology to detect S. ratti in faecal material of wild rats infected even with very low parasite burden

Histopathological confirmation of disseminated larvae (iL3) of Strongyloides ratti in an immunosuppressed Wistar rat

Human strongyloidiasis research requires a large supply of Strongyloides stercoralis.This can be achieved through in vivo maintenance of Strongyloides stercoralis in Merionesunguiculatus, but isolating a large quantity of Strongyloides stercoralis to establish thecolony from an infected patient is too difficult to achieve. Hence, Strongyloides ratti havebeen used as a model in human strongyloidiasis research. This study describes a successfulestablishment and maintenance of Strongyloides ratti infection in experimentallyimmunosuppressed Wistar rats. Large quantities of filariform (iL3) larvae of Strongyloidesratti for research related to human strongyloidiasis have been harvested following thisprotocol. Molecular detection method based on PCR using species specific primers was usedto confirm the species of the harvested infective larvae (iL3). Additionally, the identificationof histopathological lesions confirmed the presence of infective larvae (iL3) in the liver andlungs as a result of an increased parasite burden due to hyperinfection and disseminateddisease. This pathological presentation was found to be similar to that reported inStrongyloides stercoralis-infected immunocompromised human subjects

Proteomic and transcriptomic analyses of protein biomarkers during early hyperinfection in strongyloidiasis

Strongyloides stercoralis is the most common intestinal nematode in humans, and it infects millions of people worldwide. This infection results in asymptomatic chronic disease that may remain undetectable. Its unique ability to proliferate within the host can cause a hyperinfection syndrome and dissemination of infective larvae in individuals with impaired cell-mediated immunity, thus increasing the mortality rate up to 87%. The diagnosis of hyperinfection syndrome is difficult to establish and entails a high level of suspicion. The main objective of the current study was to identify specific protein biomarkers from the excretory/secretory (ES) products of the infective filariform larva and from the serum proteins of infected Wistar rats that can be used as diagnostic indicators for early hyperinfection syndrome in strongyloidiasis. In this study, S. ratti, which is an animal parasite similar to S. stercoralis and commonly used in research related to S. stercoralis, was used as a model. Seventy wild rats, Rattus norvegicus, were trapped from different locations in the Serdang areas and were brought back to the Animal Experimental Unit and Medical Parasitology and Entomology Lab in UPM for further examination. S. ratti was detected in 34.2% of the trapped wild rats by using different conventional parasitological techniques and this was then reconfirmed by using Polymerase chain reaction (PCR) methods. A PCR method targeting the rRNA gene of the species of Strongyloides was conducted based on the published universal primers for the detection of S. ratti in faecal samples of wild rats. Strongyloides ratti was then isolated from the wild rats and new colonies were established and maintained in laboratory bred Wistar rats for the continuous supply of S. ratti. Faecal pellets from infected Wistar rats were collected and cultured using Modified Faecal Filtration Culture technique (MFFC) for the harvesting of high quality and quantity of infective filariform larvae (L3) used for this research. An experimental study was performed to induce hyperinfection syndrome and dissemination of L3 larvae of S. ratti in experimentally immunosuppressed Wistar rats using prednisolone, a corticosteroid immunosuppressive drug to validate that the pathological changes that occurred were similar between the Wistar rat and human strongyloidiasis. Infected Wistar rats were sacrificed and tissue samples were collected for histopathology study. Prednisolone treatment resulted in a dramatic increase in the infection intensities as proved by the increased in eggs and larval output, and adult recovery that exceeded the inoculated doses. The results of the histopathology study showed dissemination of infective filariform larvae mainly in the tissues of the lungs and liver because of an increased parasite burden during hyperinfection and disseminated strongyloidiasis. These observations were similar to human strongyloidiasis under immunosuppressive or anti-inflammatory regimens, including corticosteroid therapies. In this study, the Wistar rats were divided into two main groups, the first group was infected with S. ratti but without giving prednisolone (non-treated group) and the second group was infected with S. ratti and treated with prednisolone (treated group). Excretory/secretory (ES) products from the filariform larvae and blood serum from the non-treated Wistar rats and the Wistar rats treated with 4.5 mg/kg prednisolone were analysed in one-dimensional and two-dimensional gel electrophoresis (1D, 2D), LC-MS/MS and MALDI-TOF/TOF MS. A total of 10 protein biomarkers were detected as overexpressed from a treated ES product with molecular weights ranging from 30-90 kDa and isoelectric points ranging from 3-11, as well as 8 overexpressed protein biomarkers which were detected from treated Wistar rat sera. Relative semi-quantitative real-time PCR (qPCR) with SYBR Green was performed using a Mastercycler Realplex to compare the expression level of the detected biokmaker’s genes between treated and non-treated groups. These genes were selected based on proteomics results and are representing respective protein that have identified earlier, in order to validate the related expressed protein biomarkers. Nine related genes were identified which showed significantly higher expression levels to the related identified biomarkers, whereas one gene (Arg) recorded down regulation in its expression. The study concluded that, 18 protein biomarkers were successfully identified and could be used as diagnostic biomarkers during early hyperinfection syndrome in strongyloidiasis

Quantification of C-type lectin gene expression during hyperinfection in strongyloidiasis

Strongyloides stercoralis is the intestinal nematode in humans, and it infects millions of people worldwide but thrives in warm countries with poor sanitation conditions. Clinical manifestations of the infection may range from asymptomatic to chronic. Strongyloides cause hyperinfection syndrome and dissemination in individuals with impaired cell-mediated immunity due to its ability to proliferate within the host that may increase the mortality rate up to 87%. The diagnosis of hyperinfection syndrome is difficult to establish and entails a high level of suspicion. The objective of the present study was to measure the expression level of C-type lectin gene coding to protein biomarker candidates from the excretory/secretory (ES) products of the infective filariform larva that can be used as diagnostic indicators for early hyperinfection syndrome in strongyloidiasis. An experimental study was carried out to induce hyperinfection of L3 larvae of S. ratti in experimentally immunosuppressed Wistar rats using prednisolone, a corticosteroid immunosuppressive drug. Prednisolone treatment resulted in a significant increase in the parasitic intensities. Relative semi-quantitative real-time PCR was performed to compare the expression level of the C-type lectin’s gene between treated and nontreated groups with this drug. C-type lectin gene showed significantly higher expression levels in the treated samples. The study concluded that C-type lectin expression level was successfully measured and could be used as a diagnostic biomarker during early hyperinfection syndrome in strongyloidiasis

Susceptibility profiles of bacterial pathogens causing community-acquired urinary tract infections

Introduction: Uncontrolled empirical treatment of urinary tract infections (UTIs) has negative aspect on predict- ing the emergence of antimicrobial resistance and knowledge of those patterns has become extremely important from time to time. Therefore, the aim of the present study was to check the prevalence and resistance patterns of uropathogens in the community acquired UTIs. Methods: A total of 7132 urine samples were combined from male 3131 (43.9%) and female 4001 (56.1%) outpatients suspected of having UTIs, respectively over a three-year period and cultured on routine culture media. The bacteria have been identified using basic biochemical tests, and sen- sitivity to various antibiotics was determined by the method of disk diffusion. Results: Of 7132 urine samples 797 (11.2%) yielded significant uropathogens. Among the bacterial species, Escherichia coli was the major causative agent of UTIs for both gender (63.7%), followed by Klebsiella spp (20.8%), Enterococcus faecalis (5.3%), Pseudo- monas spp (4.1%), Proteus spp (3.1%), Enterobacter spp (1.5%), Candida albicans (0.6%), Staphylococcus sapro- phyticus (0.5%), Providencia spp (0.1%) and Staphylococcus aureus (0.1%). The antibacterial sensitivity testing for E. coli, to commonly used antibiotics were showed variable resistant as follows: Ampicilln (78%), Amoxicillin (71%), trimethoprim sulfamethoxazole (42%), Amox/clav. (14%) gentamicin (20%), nitrofurantoin (11%), nalidixic acid (22%), ciprofloxacin (20%), Imipenem (16%),Ceftazidim (26%),Cefotaxim (25%),Ceftriaxon (21%),Cefuroxim (33%). Conclusions: The findings showed that antimicrobial resistance patterns of uropathogens in variable, and continuous monitoring of resistance patterns by using of antibiotic susceptibility testing in the laboratory is the most appropriate to treat UTIs rather than the choice of UTIs empirical treatment

Establishment of chronic Strongyloides ratti infection model and its diagnostic significance

Infection with Strongyloides stercoralis is associated with asymptomatic chronic disease involving the gut and respiratory systems and can persist for long period undetected leading to development of fatal hyperinfection syndrome. The severity is even more in immunocompromised individuals or patients undergoing immune suppressive therapy. Unfortunately the disease is difficult to diagnose due to low parasite load and the irregular larval output. Development of efficient diagnostic techniques requires large supply of infective larvae and this can be achieved through in vivo maintenance of the parasite. In this regard, S. ratti is being used as a model instead of S. stercoralis due to difficulty and risk of infection. In this study, a significant amount of previously characterized filariform (iL3) larvae of S. ratti was collected. Chronic infection was successfully established and maintained by experimentally induced immunosuppression of Wistar rats following oral administration of prednisolone. This model can serve as a good source of infective larvae for production of monoclonal antibodies and development of diagnostic assays for prevention of the disease. It can also provide room for further research in diagnostics of human strongyloidiasis