Comparison of Real-time PCR to ELISA for the detection of human cytomegalovirus infection in renal transplant patients in the Sudan

Background

This study was carried out to detect human cytomegalovirus (HCMV) IgG and IgM antibodies using an Enzyme-linked immunosorbent assay (ELISA) in renal transplant patients in Khartoum state, Sudan and to improve the diagnosis of HCMV through the introduction of Real-time Polymerase Chain Reaction (PCR) testing. A total of 98 plasma samples were collected randomly from renal transplant patients at Ibin Sina Hospital and Salma Centre for Transplantation and Haemodialysis during the period from August to September 2006.

Results

Among the 98 renal transplant patients, 65 were males and 33 females. The results revealed that HCMV IgG was present in all patients' plasma 98/98 (100%), while only 6/98 (6.1%) had IgM antibodies in their plasma. HCMV DNA viral loads were detected in 32 patients 32/98 (32.7%) using Real-time PCR.

Conclusions

The HCMV IgG results indicate a high prevalence of past HCMV infection in all tested groups, while the finding of IgM may reflect a recent infection or reactivation. HCMV detection by real-time PCR in the present study indicated a high prevalence among renal transplant patients in Khartoum. In conclusion, the prevalence of HCMV in Khartoum State was documented through detection of HCMV-specific antibodies. Further study using various diagnostic methods should be considered to determine the prevalence of HCMV disease at the national level.

     

Validation of the ALS assay in adult patients with culture confirmed pulmonary tuberculosis

Background

We have earlier shown that Bacille Calmette-Guérin (BCG) vaccine-specific IgG Antibodies in Lymphocyte Supernatant (ALS) can be used for diagnosis of active tuberculosis (TB) in adults and children.

Methodology/Principal Findings

The ALS method was validated in a larger cohort (n = 212) of patients with suspicion of pulmonary TB using multiple antigens (BCG, LAM, TB15.3, TB51A, CFP10-ESAT6-A, CFP, CW) from Mycobacterium tuberculosis. The sensitivity and specificity of the ALS assay was calculated using non-TB patients as controls. The sensitivity and the specificity were highest with BCG vaccine (90% and 88% respectively) followed by LAM (89% and 87% respectively). Simultaneous assessment of multiple antigen-specific antibodies increased sensitivity (91%) and specificity (88%). Using higher lymphocyte count in smaller volume of culture media increased detection and reduced the assay duration to ∼30 hrs. Twenty one patients with clinical findings strongly suggestive of TB finally diagnosed as non-TB patients were positive by the ALS assay, of which 9 (43%) were positive for 7 antigens and 19 (90%) for at least 3 antigens.

Conclusions/Significance

Our findings show that simultaneous detection of antigens improves the diagnostic potential of the ALS assay; the modified method increases sensitivity and can provide results in <48 hours, and enable detection of some cases of pulmonary TB that are not detectable by standard methods.     

In silico and in vitro comparative activity of novel experimental derivatives against Leishmania major and Leishmania infantum promastigotes

This in silico and in vitro comparative study was designed to evaluate the effectiveness of some biurets (K1 to K8) and glucantime against Leishmania major and Leishmania infantum promastigotes. Overall, eight experimental ligands and glucantime were docked using AutoDock 4.3 program into the active sites of Leishmania major and Leishmania infantum pteridine reductase 1, which were modeled using homology modeling programs. The colorimetric MTT assay was used to find L. major and L. infantum promastigotes viability at different concentrations of biuret derivatives in a concentration and time-dependent manner and the obtained results were expressed as 50% and 90% of inhibitory concentration (IC₅₀ and IC₉₀). In silico method showed that out of eight experimental ligands, four compounds were more active on pteridine reductase 1. K3 was the most active against L. major promastigotes with an IC₅₀ of 6.8 μM and an IC₉₀ of 40.2 μM, whereas for L. infantum promastigotes was K8 with IC₅₀ of 7.8 μM. The phenylethyl derivative (K7) showed less toxicity (IC₅₀s > 60 μM) in both Leishmania strains. Glucantime displayed less growth inhibition in concentration of about 20 μM. In silico and especially docking results in a recent study were in accordance with the in vitro activity of these compounds in presented study and compound K3, K2 and K8 showed reasonable levels of selectivity for the Leishmania pteridine reductase 1.     

Mapping of the quantitative trait locus (QTL) conferring partial resistance to rice leaf blast disease

Malaysian rice, Pongsu Seribu 2, has wide-spectrum resistance against blast disease. Chromosomal locations conferring quantitative resistance were detected by linkage mapping with SSRs and quantitative trait locus (QTL) analysis. For the mapping population, 188 F₃ families were derived from a cross between the susceptible cultivar, Mahsuri, and a resistant variety, Pongsu Seribu 2. Partial resistance to leaf blast in the mapping population was assessed. A linkage map covering ten chromosomes and consisting of 63 SSR markers was constructed. 13 QTLs, including 6 putative and 7 putative QTLs, were detected on chromosomes 1, 2, 3, 5, 6, 10, 11 and 12. The resulting phenotypic variation due to a single QTL ranged from 2 to 13 %. These QTLs accounted for approx. 80 % of the total phenotypic variation within the F₃ population. Therefore, partial resistance to blast in Pongsu Seribu 2 is due to combined effects of multiple loci with major and minor effects.     

Blast resistance in rice: a review of conventional breeding to molecular approaches

Blast disease caused by the fungal pathogen Magnaporthe oryzae is the most severe diseases of rice. Using classical plant breeding techniques, breeders have developed a number of blast resistant cultivars adapted to different rice growing regions worldwide. However, the rice industry remains threatened by blast disease due to the instability of blast fungus. Recent advances in rice genomics provide additional tools for plant breeders to improve rice production systems that would be environmentally friendly. This article outlines the application of conventional breeding, tissue culture and DNA-based markers that are used for accelerating the development of blast resistant rice cultivars. The best way for controlling the disease is to incorporate both qualitative and quantitative genes in resistant variety. Through conventional and molecular breeding many blast-resistant varieties have been developed. Conventional breeding for disease resistance is tedious, time consuming and mostly dependent on environment as compare to molecular breeding particularly marker assisted selection, which is easier, highly efficient and precise. For effective management of blast disease, breeding work should be focused on utilizing the broad spectrum of resistance genes and pyramiding genes and quantitative trait loci. Marker assisted selection provides potential solution to some of the problems that conventional breeding cannot resolve. In recent years, blast resistant genes have introgressed into Luhui 17, G46B, Zhenshan 97B, Jin 23B, CO39, IR50, Pusa1602 and Pusa1603 lines through marker assisted selection. Introduction of exotic genes for resistance induced the occurrence of new races of blast fungus, therefore breeding work should be concentrated in local resistance genes. This review focuses on the conventional breeding to the latest molecular progress in blast disease resistance in rice. This update information will be helpful guidance for rice breeders to develop durable blast resistant rice variety through marker assisted selection.     

Platelet-derived growth factor oncoprotein detection using three-dimensional carbon microarrays

The potential of aptamers as ligand binding molecule has opened new avenues in the development of biosensors for cancer oncoproteins. In this paper, a label-free detection strategy using signaling aptamer/protein binding complex for platelet-derived growth factor (PDGF-BB) oncoprotein detection is reported. The detection mechanism is based on the release of fluorophore (TOTO intercalating dye) from the target binding aptamer's stem structure when it captures PDGF. Amino-terminated three-dimensional carbon microarrays fabricated by pyrolyzing patterned photoresist were used as a detection platform. The sensor showed near linear relationship between the relative fluorescence difference and protein concentration even in the sub-nanomolar range with an excellent detection limit of 5pmol. This detection strategy is promising in a wide range of applications in the detection of cancer biomarkers and other proteins.     

Deoxyribonucleic acid repair gene X-ray repair cross-complementing group 1 polymorphisms and non-carcinogenic disease risk in different populations: A meta-analysis

PURPOSE:

This study aims to assess a meta-analysis of the association of X-ray repair cross-complementing group 1 (XRCC1) polymorphisms with the risk of various non-carcinogenic diseases in different population.

MATERIALS AND METHODS:

This meta-analysis was performed by critically reviewing reveals 38 studies involving 10043 cases and 11037 controls. Among all the eligible studies, 14 focused on Arg194Trp polymorphism, 33 described the Arg399Gln and three articles investigated on Arg280His. Populations were divided into three different ethnic subgroups include Caucasians, Asians and other (Turkish and Iranian).

RESULTS:

Pooled results showed no correlation between Arg194Trp and non-carcinogenic disease. There was only weak relation in the recessive (odds ratio [OR] =1.11, 95% confidence interval [CI]: 0.86-1.44) model in Asian population and dominant (OR = 1.04, 95% CI: 0.66-1.63) model of other populations. In Arg399Gln polymorphism, there was no relation with diseases of interest generally. In the pooled analysis, there were weak relation in the dominant (OR = 1.08, 95% CI: 0.86-1.35) model of Asian population and quite well-correlation with recessive (OR = 1.49, 95% CI: 1.19-1.88), dominant (OR = 1.23, 95% CI: 0.94-1.62), and additive (OR = 1.23, 95% CI: 0.94-1.62) models of other subgroup. For Arg280His, there was a weak relation only in the dominant model (OR = 1.06, 95% CI: 0.74-1.51).

CONCLUSION:

The present meta-analysis correspondingly shows that Arg399Gln variant to be associated with increased non-carcinogenic diseases risk through dominant and recessive modes among Iranian and Turkish population. It also suggests a trend of dominant and recessive effect of Arg280His variant in all population and its possible protective effect on non-carcinogenic diseases.     

Horizontal gene and chromosome transfer in plant pathogenic fungi affecting host range

Plant pathogenic fungi adapt quickly to changing environments including overcoming plant disease resistance genes. This is usually achieved by mutations in single effector genes of the pathogens, enabling them to avoid recognition by the host plant. In addition, horizontal gene transfer (HGT) and horizontal chromosome transfer (HCT) provide a means for pathogens to broaden their host range. Recently, several reports have appeared in the literature on HGT, HCT and hybridization between plant pathogenic fungi that affect their host range, including species of Stagonospora/Pyrenophora, Fusarium and Alternaria. Evidence is given that HGT of the ToxA gene from Stagonospora nodorum to Pyrenophora tritici-repentis enabled the latter fungus to cause a serious disease in wheat. A nonpathogenic Fusarium species can become pathogenic on tomato by HCT of a pathogenicity chromosome from Fusarium oxysporum f.sp lycopersici, a well-known pathogen of tomato. Similarly, Alternaria species can broaden their host range by HCT of a single chromosome carrying a cluster of genes encoding host-specific toxins that enabled them to become pathogenic on new hosts such as apple, Japanese pear, strawberry and tomato, respectively. The mechanisms HGT and HCT and their impact on potential emergence of fungal plant pathogens adapted to new host plants will be discussed.