West Nile virus: The current situation in Egypt.

Background and Aim: Due to climatic changes, arthropod-borne viruses have become a global health concern. In Egypt, West Nile virus (WNV) was initially detected in humans in 1950 and then in 1951, 1954, 1968, and 1989. Although WNV infection has been recorded in numerous Middle Eastern countries, its prevalence among the equine population in Egypt is unknown. This study aimed to investigate the current situation of vector-borne WNV in Egypt, estimate its seroprevalence, and assess the associated risk factors. Materials and Methods: We screened 1100 sera samples and nasal swabs from the same equids, 156 mosquito pools, and 336 oropharyngeal and cloacal swabs from migratory birds for WNV. The sera were investigated for the presence of immunoglobulin G (IgG) and immunoglobulin M (IgM) against WNV-prE. Real-time reverse transcription-polymerase chain reaction was used to detect WNV RNA in the nasal swab samples, mosquito pools, and migratory birds' oropharyngeal and cloacal swabs. Results: The seroprevalence showed positive IgG in sera samples collected from different districts. The data showed that horses were 1.65-fold more susceptible than donkeys, with male being 1.45 times more susceptible than females. Moreover, the tested equids samples were divided into three groups based on their age: <5 years, 5-10 years, and >10 years. The 5-10-year group was 1.1 and 1.61 times more vulnerable to infection than the <5- and >10 year groups. All the sera samples were negative for IgM. The nasal swabs from equids, oropharyngeal and cloacal swabs from migratory birds, and mosquito samples tested negative for WNV by molecular detection. Conclusion: Based on the obtained data, we recommend that effective control programs should be implemented to enable epidemiological investigations and understand the current situation of WNV in Egypt.

Double-antibody-based nano-biosensing system for the onsite monitoring of SARS-CoV-2 variants.

The fast and reliable diagnosis of COVID-19 is the foremost priority for promoting public health interventions. Therefore, double-antibody-based immunobiosensor chips were designed, constructed, and exploited for clinical diagnosis. Gold nanoparticles/tungsten oxide/carbon nanotubes (AuNPs/WO3/CNTs) were used as the active working sensor surface to support the chemical immobilization of a mixture of SARS-CoV-2 antibodies (anti-RBD-S and anti-RBD-S-anti-Llama monoclonal antibodies). The morphology and chemical functionalization of the fabricated disposable immunochips was characterized using scanning electron microscopy (SEM), Fourier transform infrared (FTIR) spectroscopy, cyclic voltammetry (CV), and electrochemical impedance spectroscopy (EIS). After full assay optimization, the immunobiosensor showed a high sensitivity to detect SARS-CoV-2-S protein with limits of detection and quantification of 1.8 and 5.6 pg/mL, respectively. On the other hand, for the SARS-CoV-2 whole virus particle analysis, the detection and quantification limits were determined to be 5.7 and 17 pg/mL, respectively. The biosensor showed a highly selective response toward SARS-CoV-2, even in the presence of influenza, nontargeting human coronaviruses, and Middle East respiratory syndrome coronavirus (MERS-CoV). The immunochips exhibited distinct responses toward the variants of concern: B.1>C.36.3>Omicron> Delta> Alpha coronavirus variants. For biosensor validation, twenty-nine clinical specimens were analyzed, and the impedimetric responses were positively detected for two Delta samples, eighteen Omicron samples, and six B.1-type samples in addition to three negative samples. Eventually, the immunobiosensor was fabricated in the form of ready-to-use chips capable of sensitive detection of virus variants, especially variants of concern (VOC) and interest, in a specimen within 15 min. The chips provided instantaneous detection with the direct application of clinical samples and are considered a point-of-care device that could be used in public places and hot spots.

Circulating foot-and-mouth disease virus serotype A African-genotype IV in Egypt during 2022.

Background and Aim: Foot-and-mouth disease (FMD) virus causes continuous outbreaks, leading to serious economic consequences that affect animal productivity and restrict trade movement. The potential influence of the disease was due to the emergence of new strains or re-emergence of local strains with major antigenic variations due to genetic mutations. This study aims to evaluate circulating virus in samples collected from infected animals during an outbreak using antigenic characterization and identify whether there is an emergence of a new strain or mutation. Materials and Methods: Reverse-transcription polymerase chain reaction (RT-PCR) was used to screen 86 samples. Viral protein 1 (VP1) codon sequencing was performed. The virus was isolated from the samples inoculated on the baby-hamster kidney cell line and Enzyme-linked immunosorbent assay was performed for serotyping and antigen detection. Results: Based on the RT-PCR screening results, 10 positive samples were selected for sequencing. The sequences belonged to the FMD serotype A African topotype originating from the ancestor prototype Sudan/77, with which it shared 98.48% ± 1.2% similarity. The divergence with local isolates from 2020 was 9.3%. In addition, the sequences were 96.84% ± 1.01% and 95.84% ± 0.79% related to Egyptian-Damietta type 2016 and Sudanese-2018, respectively. Divergence with vaccinal strains ranged from 10% to 17%. Amino acid sequence analysis revealed that the isolates had variation in the most prominent antigenic regions (residues 35-75) and the immunogenic determinants of the G-H loop of VP1 (residues 100-146 and 161-175). Conclusion: The current isolates should be included in the locally produced vaccine to provide broader immunogenic coverage against serotype A African topotypes.

Designing and fabrication of electrochemical nano-biosensor for the fast detection of SARS-CoV-2-RNA.

SARS-CoV-2 caused a global panic among populations. Rapid diagnostic procedures for the virus are crucial for disease control. Thus, the designed signature probe from a highly conserved region of the virus was chemically immobilized onto the nanostructured-AuNPs/WO3-screen printed electrodes. Different concentrations of the matched oligonucleotides were spiked to test the specificity of the hybridization affinity whereas the electrochemical impedance spectroscopy was used for tracking the electrochemical performance. After a full assay optimization, limits of detection and quantification were calculated based on linear regression and were valued at 298 and 994 fM, respectively. Further, the high performance of the fabricated RNA-sensor chips was confirmed after testing the interference status in the presence of the mismatched oligos in one nucleotide and completely one. Worthy to mention that the single-stranded matched oligos can be hybridized to the immobilized probe in 5 min at room temperature. The designed disposable sensor chips are capable of detecting the virus genome directly. Therefore, the chips are a rapid tool for SARS-CoV-2 detection.

Molecular detection and phylogenetic analysis of newly emerging foot-and-mouth disease virus type A, Lineage EURO-SA in Egypt in 2022.

A newly emerging and exotic foot-and-mouth disease virus (FMDV) caused a recent outbreak of serotype A in Egypt in 2022, which affected cattle and water buffalo. Previous phylogenetic studies on FMDV circulating in Egypt have mainly focused on genomic regions encoding the structural proteins which determine FMDV serotype. No study has yet determined structural proteins sequences of the newly emerging Europe-South America (EURO-SA) lineage which was recently isolated from Egypt during a routine surveillance in 2022. The objective of the current study was to analyze the structural proteins of the Venezuelan type which belongs to EURO-SA. The new isolate was related to serotype A lineage Euro-South America. Phylogentic analyses have reveled that the newly isolated lineage samples were closely related to reported sequences that have been identified in Venzuela and Colombia. Analysis of structural protein sequences revealed the recent isolates belong to prototype strain A24 Cruzeiro. Notably, nucleotide sequences of the Egyptian isolate was related to Venezuelan, Brazilian, and Colombian strains with identity not exceeding 90%. The divergence which appears in the genetic identity of the Egyptian A/EURO-SA lineage from other related strains may be attributed to the absence of Euro-SA lineage sequence from Egypt. The present study is the first report on the detection of EURO-SA lineage in Egypt. The recent detection of the EURO-SA lineage samples may be explained due to imported animals from Colombia or Brazil which share geographical borders with Venezuela. The findings of the present study highlight the significance of continuous monitoring of FMDV in Egypt for newly emerging FMDVs.

Assessment of intralesional injection of botulinum toxin type A in hypertrophic scars and keloids: Clinical and pathological study.

Keloids and hypertrophic scars are cosmetic problems with significant morbidity. Many clinical modalities were tried in order to modulate the disfigurement related to these pathologic scars. To evaluate the clinical and histopathological effects of Botulinum toxin type A (BTX-A) injection on keloids and hypertrophic scars. Twelve patients with keloids and 8 with hypertrophic scars were enrolled in this study. Botulinum toxin type A was injected intralesional (1 session/month) for three sessions. Clinical outcome was assessed via Vancouver Scar Scale (VSS), Observer Scar Assessment Scale (OSAS), and the Patient Scar Assessment Scale (PSAS). Histologic grading scores were used to assess the changes in the quality of collagen and elastic tissues and image analysis was used to detect their quantitative morphometric changes. This study showed a high statistically significant difference between baseline and the result after each of the three sessions of injection and 3, 6 months after the last session regarding VSS, OSAS, and PSAS with p value ≤0.001 for each. The study also showed that there was a statistically significant difference between the histopathologic findings before injection of BTX and 1 month after the third session regarding all parameters used. Botulinum toxin type A can be a good therapeutic maneuver for management of keloid and hypertrophic scars with significant clinical and histologic improvement.

Molecular characterization and pathological identification of a novel strain of delta papillomavirus-4 (bovine papillomavirus-2) in Egypt.

BACKGROUND AND AIM: Bovine papillomaviruses (BPV) are a heterogeneous group of oncoviruses, distributed globally, which produce major economic losses. In the current study, we compared the results of different diagnostic approaches and compared the strains identified in this study with previously characterized strains at local and international levels. MATERIALS AND METHODS: Samples of skin warts were collected from five bovines with generalized papillomatosis from two Egyptian provinces, Menya and Ismailia, in 2020. Electron microscopy, molecular characterization, histopathological, and immunohistochemical examination were performed. RESULTS: BPV was detected using electron microscopy in the collected samples. Using molecular characterization, BPV-2 was successfully identified for 1st time in Egypt. The strain has 99.6% identity with the BPV-2 reference strains obtained from GenBank. These results were supported by histopathology and immunohistochemistry examination. Partial nucleotide sequences of the L1 gene were submitted to GenBank with accession numbers MW289843 and MW289844. CONCLUSION: BPV-2 was reported for 1st time in the current study. The strain was identified grossly, microscopically, and pathologically and confirmed using molecular approaches. All results were consistent. The sequence analysis revealed that this strain has high sequence similarity to the reference Deltapapillomavirus-4, BPV-2 strains from Brazil and China.

SARS-CoV-2-Impedimetric Biosensor: Virus-Imprinted Chips for Early and Rapid Diagnosis.

Due to the current global SARS-CoV-2 pandemic, rapid and accurate diagnostic tools are needed to prevent the spread of COVID-19 across the globe. An electrochemical sensing platform was constructed using CNTs/WO3-screen printed electrodes for imprinting the complete virus particles (SARS-CoV-2 particles) within the polymeric matrix to create viral complementary binding sites. The sensor provided high selectivity toward the target virus over other tested human corona and influenza respiratory interference viruses. The sensitivity performance of the sensor chips was evaluated using different viral concentrations, while the limits of detection and quantification were 57 and 175 pg/mL, respectively. Reaching this satisfied low detection limit (almost 27-fold more sensitive than the RT-PCR), the sensor was applied in clinical specimens obtained from SARS-CoV-2 suspected cases. Thus, dealing directly with clinical samples on the chip could be provided as a portable device for instantaneous and simple point of care in hospitals, airports, and hotspots.

High selectivity detection of FMDV- SAT-2 using a newly-developed electrochemical nanosensors.

Foot-and-mouth disease virus serotype South-Africa territories-2 (FMDV-SAT-2) is the most fastidious known type in Aphthovirus which is subsequently reflected in the diagnosis regime. Rapid and early diagnostic actions are usually taken in response to the FMDV outbreak to prevent the dramatic spread of the disease. Virus imprinted sensor (VIP sensor) is gathering huge attention for the selective detection of pathogens. Thus, the whole virus particles of SAT-2 together with an electropolymerized film of poly(o-phenylenediamine) (PoPD) on gold-copper modified screen-printed electrode were applied to fabricate SAT-2-virus imprinted polymer (SAT-2-VIP). The SAT-2-VIPs were fully characterized using cyclic voltammetry (CV), linear sweep voltammetry (LSV), Atomic force microscopy (AFM), Scanning electron microscope (SEM), and Fourier transform Infra-Red (FTIR) spectroscopy. Excellent selective binding affinity towards the targeted virus particle was achieved with limits of detection and quantification of 0.1 ng/mL and 0.4 ng/mL, respectively. In terms of viral interference, the sensor did not show cross-reactivity towards other animal viruses including FMDV serotype A, O, or even SAT-2 subtype Libya and the un-related virus Lumpy skin disease virus (LSDV). This high selectivity provides a sensible platform with 70 folds more sensitivity than the reference RT-PCR as revealed from the application of SAT-2-VIP sensor for rapid analysis of clinical samples with no need for treatment or equipped labs. Thus, as diagnostic and surveillance technologies, on-site point of care diagnostics for SAT-2 virus are supported.

Point-of-Care Diagnostics of COVID-19: From Current Work to Future Perspectives.

Coronaviruses have received global concern since 2003, when an outbreak caused by SARS-CoV emerged in China. Later on, in 2012, the Middle-East respiratory syndrome spread in Saudi Arabia, caused by MERS-CoV. Currently, the global crisis is caused by the pandemic SARS-CoV-2, which belongs to the same lineage of SARS-CoV. In response to the urgent need of diagnostic tools, several lab-based and biosensing techniques have been proposed so far. Five main areas have been individuated and discussed in terms of their strengths and weaknesses. The cell-culture detection and the microneutralization tests are still considered highly reliable methods. The genetic screening, featuring the well-established Real-time polymerase chain reaction (RT-PCR), represents the gold standard for virus detection in nasopharyngeal swabs. On the other side, immunoassays were developed, either by screening/antigen recognition of IgM/IgG or by detecting the whole virus, in blood and sera. Next, proteomic mass-spectrometry (MS)-based methodologies have also been proposed for the analysis of swab samples. Finally, virus-biosensing devices were efficiently designed. Both electrochemical immunosensors and eye-based technologies have been described, showing detection times lower than 10 min after swab introduction. Alternative to swab-based techniques, lateral flow point-of-care immunoassays are already commercially available for the analysis of blood samples. Such biosensing devices hold the advantage of being portable for on-site testing in hospitals, airports, and hotspots, virtually without any sample treatment or complicated lab precautions.

Designing and fabrication of new VIP biosensor for the rapid and selective detection of foot-and-mouth disease virus (FMDV).

Foot and mouth disease virus (FMDV), is a highly contagious virus due to its ease of transmission. FMDV has seven genetically distinguished serotypes with many subtypes within each serotype. The traditional diagnostic methods of FMDV have demonstrated many drawbacks related to sensitivity, specificity, and cross-reactivity. In the current study, a new viral imprinted polymer (VIP)-based biosensor was designed and fabricated for the rapid and selective detection of the FMDV. The bio-recognition components were formed via electrochemical polymerization of the oxidized O-aminophenol (O-AP) film imprinted with FMDV serotype O on a gold screen-printed electrode (SPE). The overall changes in the design template have been investigated using cyclic voltammetry (CV), atomic force microscopy (AFM), Field emission-scanning electron microscopy (FE-SEM), and Fourier-transform infrared spectroscopy (FT-IR). Optimal conditions were achieved through investigating the capturing efficiency, binding stability, selectivity and life-time of the developed biosensor. The results depicted a high selectivity of the biosensor to the serotype O over all other genus serotypes A, SAT2 and Lumpy skin disease virus (LSDV), as well as, the inactivated serotype O. The limits of detection (LOD) and quantification (LOQ) were around 2 ng/mL and 6 ng/mL, respectively, in addition to the tested repeatability and reproducibility values with a variance coefficient of 1.0% and 3.6%, respectively. In comparison with the reference methods (ELISA and PCR), the analysis of saliva real samples using the developed affordable biosensor offered 50 folds lower LOD with the possibility of an on-line monitoring in the field with no prior sample treatment.