Generation and Selection of Specific Aptamers Targeting Brucella Species through an Enhanced Cell-SELEX Methodology.

Brucellae are Gram-negative, aerobic, non-motile coccobacilli causing brucellosis in man and animals. The disease is one of the most significant yet neglected global zoonoses. Especially in developing countries, brucellosis is causing public health problems and economic losses to private animal owners and national revenues. Composed of oligonucleotides, aptamers are chemical analogues of antibodies that are promising components for developing aptamer-based rapid, sensitive, and specific tests to identify the Brucella group of bacteria. For this purpose, aptamers were generated and selected by an enhanced protocol of cell systematic evolution of ligands by exponential enrichment (cell-SELEX). This enhanced cell-SELEX procedure involved the combination of both conventional and toggle cell-SELEX to boost the specificity and binding affinity to whole Brucella cells. This procedure, combined with high-throughput sequencing of the resulting aptamer pools, comprehensive bioinformatics analysis, and wet lab validation assays, led to the selection of a highly sensitive and specific aptamer for those Brucella species known to circulate in Egypt. The isolated candidate aptamer showed dissociation constant (KD) values of 43.5 ± 11, 61.5 ± 8, and 56 ± 10.8 nM for B. melitensis, B. abortus, and B. suis, respectively. This is the first development of a Brucella-specific aptamer using an enhanced combination of conventional and toggle cell-SELEX to the authors' best knowledge.

Application of gold nanoparticle-assisted PCR for equine herpesvirus 1 diagnosis in field samples.

Equine herpesvirus 1 (EHV-1) is one of the most significant pathogens that affects equine species worldwide, causing sporadic abortion, neonatal deaths, chorioretinopathy, as well as neurological and upper respiratory tract diseases. Currently, conventional PCR targeting different genes is used widely for the molecular detection of EHV-1, but the low viral titer in some clinical samples can lead to false negative results. In this study, we aimed to assess gold nanoparticle (GNP)-assisted PCR as an inexpensive, highly efficient, and sensitive method for the detection of EHV-1, and to compare its results with conventional PCR and real-time quantitative PCR (qPCR). Out of 83 field samples, 28.9%, 26.5%, and 15.6% were EHV-1-positive by qPCR, GNP-assisted PCR and conventional PCR, respectively. All three techniques specifically target the viral glycoprotein B gene. The optimized GNP-assisted PCR showed no cross-reactivity with EHV-1-negative samples (diagnosed by qPCR). GNP-assisted PCR is a powerful new tool for EHV-1 detection and surveillance, because of its simplicity, sensitivity and specificity. It can be used as an alternative to qPCR in laboratories that cannot afford the expense of a qPCR system.

Removal of urea from dilute streams using RVC/nano-NiOx-modified electrode.

Reticulated vitreous carbon (RVC), a high surface area electrode (40 cm2/cm3), has been modified with nickel oxide nanoparticles (nano-NiOx) and used for electrochemical oxidation of urea from alkaline solution. For the cyclic voltammetry measurements, the used dimensions are 0.8 cm × 0.8 cm × 0.3 cm. The purpose was to offer high specific surface area using a porous open network structure to accelerate the electrochemical conversion. NiOx nanoparticles have been synthesized via an electrochemical route at some experimental conditions. The morphological, structural, and electrochemical properties of the RVC/nano-NiOx are characterized by using scanning electron microscopy (SEM), energy dispersive X-ray spectroscopy (EDX), cyclic voltammetry (CV), and potentiostatic measurements. The fabricated electrode, RVC/nano-NiOx, demonstrates high electrocatalytic activity towards urea oxidation in an alkaline electrolyte. The onset potential of the RVC/nano-NiOx compared to that of the planar GC/NiOx is shifted to more negative value with higher specific activity. The different loadings of the NiOx have a substantial influence on the conversion of urea which has been evaluated from concentration-time curves. The urea concentration decreases with time to a limit dependent on the loading extent. Maximum conversion is obtained at 0.86 mg of NiOx per cm3 of the RVC matrix.

Colorimetric Detection of Unamplified Rift Valley Fever Virus Genetic Material Using Unmodified Gold Nanoparticles.

Rift Valley fever virus (RVFV) is considered an enzootic virus in Africa. RVFV has caused several outbreaks in Egypt, sub-Saharan Africa and the Arabian Peninsula and is responsible for high mortality in ruminants and haemorrhagic fever in severe human cases. Although there are several molecular and serological diagnostic techniques used to detect this arthropod-borne virus with high sensitivity and efficiency, there is a need for a fast and reliable field screening test for rapid outbreak recording and containment. In this study, we developed a prototype point-of-care diagnostic test specific for RVFV detection using unmodified gold nanoparticles (AuNPs) that change colour in the presence of RVFV RNA, resulting in a simple but sensitive assay. The nanogold assay provides qualitative results showing the presence of the RVFV RNA in different sample types. The assay showed high accuracy and specificity, with a detection limit of 10 RNA copies/reaction, comparable with quantitative reverse transcription polymerase chain reaction. The assay result could be determined within 30 min with no need for specific detection instruments. To our knowledge, this is the first field test prototype to directly detect the RNA of RVFV without amplification using AuNPs.