Detection Of Virus Research Articles

Co-circulation of seasonal influenza A(H1N1)pdm09, A(H3N2) and B/Victoria lineage viruses with further genetic diversification, EU/EEA, 2022/23 influenza season.

BackgroundInfluenza viruses can cause large seasonal epidemics with high healthcare impact and severity as they continually change their virological properties such as genetic makeup over time.AimWe aimed to monitor the characteristics of circulating influenza viruses over the 2022/23 influenza season in the EU/EEA countries. In addition, we wanted to compare how closely the circulating viruses resemble the viral components selected for seasonal influenza vaccines, and whether the circulating viruses had acquired resistance to commonly used antiviral drugs.MethodsWe performed a descriptive analysis of the influenza virus detections and characterisations reported by National Influenza Centres (NIC) from the 30 EU/EEA countries from week 40/2022 to week 39/2023 to The European Surveillance System (TESSy) as part of the Global Influenza Surveillance and Response System (GISRS).ResultsIn the EU/EEA countries, the 2022/23 influenza season was characterised by co-circulation of A(H1N1)pdm09, A(H3N2) and B/Victoria-lineage viruses. The genetic evolution of these viruses continued and clade 6B.1A.5a.2a of A(H1N1)pdm09, 3C.2a1b.2a.2b of A(H3N2) and V1A.3a.2 of B/Victoria viruses dominated. Influenza B/Yamagata-lineage viruses were not reported.DiscussionThe World Health Organization (WHO) vaccine composition recommendation for the northern hemisphere 2023/24 season reflects the European virus evolution, with a change of the A(H1N1)pdm09 component, while keeping the A(H3N2) and B/Victoria-lineage components unchanged.

The association between infectious agents and breast cancer: a review of the epidemiologic evidence.

Several viruses have been casually linked to human cancers, including cervical, nasopharyngeal, liver, sarcoma, and Merkel cell carcinomas. However, the etiologic contribution of viral infections to breast cancer, the number one incident cancer among women worldwide, is not well established. Among studies exploring associations of viruses with breast cancer, potential linkages have been identified between breast cancer and five viruses: beta retrovirus, (i.e., mouse mammary tumor virus), human papillomavirus, Epstein Barr virus. bovine leukemia virus, and human cytomegalovirus. In this review, we provide a comprehensive evaluation of epidemiological ecologic, case-control, case-only, and cohort studies investigating these associations. Wediscuss results from several existing reviews and meta-analyses, evaluate epidemiological studies published in the past five years, and assess the relationship between these viruses and breast tumor clinicopathological factors. The strongest epidemiological evidence for a viral role in breast cancer exists for MMTV and HPV, though limitations include lack of prospective studies for MMTV and potential detection bias in HPV studies. Viral detection challenges have limited studies of EBV and HCMV. Fewer studies have evaluated BLV, and though it has been associated with higher risk of breast cancer, sample sizes are quite small. CONCLUSION: While epidemiologic evidence exists for an association between these five viruses and breast cancer, various methodological issues and lack of prospective studies preclude robustconclusions. Future research should prioritize establishing a temporal relationship between infection and disease, minimizing misclassification of detection assays, and further exploring the influence of co-infections.

Development of simplex and quintuplex RT-PCR for simultaneous detection of soybean viruses

Five simplex and a multiplex-RT-PCR (m-RT-PCR) protocols were developed for detection and differentiation of bean pod mottle virus (BPMV), cherry leaf roll virus (CLRV), raspberry ringspot virus (RpRSV), soybean mosaic virus (SMV) and tomato ringspot virus (ToRSV) infecting soybean. The simplex RT-PCR protocols produced virus-specific amplicons of 538 bp for BPMV, 139 bp for CLRV, 298 bp for RpRSV, 403 bp for SMV, and 282 bp for ToRSV, with sensitivity down to 10−4 diluted cDNA. Further, to detect all the five viruses simultaneously in a single tube a quintuplex RT-PCR protocol was optimized with as low as 10−3 diluted cDNA and 0.05 µM primer. To validate the reliability of the simplex RT-PCR protocol, imported soybean samples were tested by ELISA as well as RT-PCR. The results revealed that the developed protocol could detect the viruses in imported soybean, and found to be efficient than ELISA in resolving ambiguity in detection of seed borne viruses. The developed simplex and quintuplex RT-PCR protocol will be quite helpful for the diagnosis of soybean germplasm co-infected with viruses during the quarantine processing for ensuring virus free long term seed conservation in the National Gene Bank as well as for quarantine certification.

Preliminary studies on the detection and presence of Lymphocystis disease virus (LCDV) in sea breams (Sparus aurata) raised in the Aegean Sea

Preliminary studies on the detection and presence of Lymphocystis disease virus (LCDV) in sea breams (Sparus aurata) raised in the Aegean Sea

Unveiling the Impact of Epstein-Barr Virus on the Risk of Prostate Cancer: A Mendelian Randomization Study

Given the consistent detection of Epstein-Barr virus (EBV) in prostate tissues and the clinical evidence suggesting its involvement in prostate cancer (PCa), the potential association between EBV infection and PCa warrants further investigation. This study aimed to assess the causal relationship between EBV infection and PCa using Mendelian randomization (MR). We utilized data from a publicly available genome-wide association study (GWAS) on PCa, alongside data on five serum anti-EBV virus-related antibodies. Our findings indicate a potential causal link between serum EBV EA-D antibody levels and an increased risk of PCa. These results highlight the need for additional research to elucidate the mechanisms by which EBV may contribute to the progression of PCa, potentially offering new insights into its pathogenesis and therapeutic targets.

Investigating a Detection Method for Viruses and Pathogens Using a Dual-Microcantilever Sensor.

Piezoresistive microcantilever sensors for the detection of viruses, pathogens, and trace chemical gasses, with appropriate measurement and signal processing methods, can be a powerful instrument with high speed and sensitivity, with in situ and real-time capabilities. This paper discusses a novel method for mass sensing on the order of a few femtograms, using a dual-microcantilever piezoresistive sensor with a vibrating common base. The two microcantilevers have controllably shifted natural frequencies with only one of them being active. Two active piezoresistors are located on the surfaces of each of the two flexures, which are specifically connected in a Wheatstone bridge with two more equivalent passive resistors located on the sensor base. A dedicated experimental system measures the voltages of the two half-bridges and, after determining their amplitude-frequency responses, finds the modulus of their differences. The modified amplitude-frequency response possesses a cusp point which is a function of the natural frequencies of the microcantilevers. The signal processing theory is derived, and experiments are carried out on the temperature variation in the natural frequency of the active microcantilever. Theoretical and experimental data of the temperature-frequency influence and equivalent mass with the same impact are obtained. The results confirm the sensor's applicability for the detection of ultra-small objects, including early diagnosis and prediction in microbiology, for example, for the presence of SARS-CoV-2 virus, other viruses, and pathogens. The versatile nature of the method makes it applicable to other fields such as medicine, chemistry, and ecology.

Cluster of Influenza A(H5) Cases Associated with Poultry Exposure at Two Facilities - Colorado, July 2024.

Persons who work in close contact with dairy cattle and poultry that are infected with highly pathogenic avian influenza (HPAI) A(H5N1) virus are at increased risk for infection. In July 2024, the Colorado Department of Public Health & Environment responded to two poultry facilities with HPAI A(H5N1) virus detections in poultry. Across the two facilities, 663 workers assisting with poultry depopulation (i.e., euthanasia) received screening for illness; 109 (16.4%) reported symptoms and consented to testing. Among those who received testing, nine (8.3%) received a positive influenza A(H5) virus test result, and 19 (17.4%) received a positive SARS-CoV-2 test result. All nine workers who received positive influenza A(H5) test results had conjunctivitis, experienced mild illness, and received oseltamivir. This poultry exposure-associated cluster of human cases of influenza A(H5) is the first reported in the United States. The identification of these cases highlights the ongoing risk to persons who work in close contact with infected animals. Early response to each facility using multidisciplinary, multilingual teams facilitated case-finding, worker screening, and treatment. As the prevalence of HPAI A(H5N1) virus clade 2.3.4.4b genotype B3.13 increases, U.S. public health agencies should prepare to rapidly investigate and respond to illness in agricultural workers, including workers with limited access to health care.

Serological Detection of West Nile Virus in Single-Humped Camels in Northern Nigeria

Serological Detection of West Nile Virus in Single-Humped Camels in Northern Nigeria

Multiplex qPCR development for the simultaneous and rapid detection of largemouth bass virus and infectious spleen and kidney necrosis virus in aquaculture

Largemouth bass virus (LMBV) and infectious spleen and kidney necrosis virus (ISKNV) are both belong to Iridoviridae that cause considerable economic losses in the fish industry. There is no reported literature that can detect these two viruses simultaneously. In this study, we established a multiplex quantitative polymerase chain reaction (qPCR) assay that can specifically and simultaneously detect both LMBV and ISKNV in fish samples. The specificity experiment showed that the method only amplified LMBV and ISKNV but not the other 10 common fish viruses. The slope (m), efficiency (E) and linearity (R2) determined from the generated standard curve were all within the optimal range of qPCR values. The detection limit of the multiplex qPCR assay was as low as 4 copies/μL for LMBV DNA and 7 copies/μL for ISKNV DNA, respectively. The established method exhibited adequate repeatability and reproducibility, and the intra- and inter-assay coefficients of variation were both less than 3 %. The accuracy of the multiplex qPCR method was validated using 229 fish samples and was more precise than that of the conventional PCR assay. In summary, the established multiplex qPCR assay can simultaneously detect LMBV and ISKNV to monitor the risk of infection LMBV and ISKNV and control the disease early.

An automated syringe-based PoC RT-LAMP LFB platform for infectious disease detection from saliva

Decentralized Point-of-Care (PoC) diagnostics hold momentous potential for rapid and accessible viral infection disease detection. Presented is a unique design application of an easy-to-use (plug-and-play) platform for viral detection. The platform leverages a simplified multiplex Reverse-Transcription Loop-mediated Isothermal Amplification (RT-LAMP) Lateral Flow Biosensor (LFB) assay with a lyophilized master mix, eliminating the need for RNA isolation or special reporting equipment. A user-friendly Saliva Measuring Tube (SMT) ensures accurate saliva volume self-collection, and a Syringe-based PoC (SPoC) platform automates sample treatment, reagent mixing, and temperature control using readily available components and consumables. The platform’s performance was evaluated by multiplexed detection of the SARS-CoV-2 N2 target gene and human ACTB gene from saliva samples. The SPoC platform achieved a detection limit of spiked 500 copies/mL for SARS-CoV-2 and consistent internal control readout. The presented PoC system offers a promising initial step for further development toward a decentralized solution for viral infection testing.

Comparative analysis of metagenomic and targeted next-generation sequencing for pathogens diagnosis in bronchoalveolar lavage fluid specimens.

Although the emerging NGS-based assays, metagenomic next-generation sequencing (mNGS) and targeted next-generation sequencing (tNGS), have been extensively utilized for the identification of pathogens in pulmonary infections, there have been limited studies systematically evaluating differences in the efficacy of mNGS and multiplex PCR-based tNGS in bronchoalveolar lavage fluid (BALF) specimens. In this study, 85 suspected infectious BALF specimens were collected. Parallel mNGS and tNGS workflows to each sample were performed; then, we comparatively compared their consistency in detecting pathogens. The differential results for clinically key pathogens were confirmed using PCR. The microbial detection rates of BALF specimens by the mNGS and tNGS workflows were 95.18% (79/83) and 92.77% (77/83), respectively, with no significant difference. mNGS identified 55 different microorganisms, whereas tNGS detected 49 pathogens. The comparative analysis of mNGS and tNGS revealed that 86.75% (72/83) of the specimens were complete or partial concordance. Particularly, mNGS and tNGS differed significantly in detection rates for some of the human herpesviruses only, including Human gammaherpesvirus 4 (P<0.001), Human betaherpesvirus 7 (P<0.001), Human betaherpesvirus 5 (P<0.05) and Human betaherpesvirus 6 (P<0.01), in which tNGS always had higher detection rates. Orthogonal testing of clinically critical pathogens showed a total coincidence rate of 50% for mNGS and PCR, as well as for tNGS and PCR. Overall, the performance of mNGS and multiplex PCR-based tNGS assays was similar for bacteria and fungi, and tNGS may be superior to mNGS for the detection of DNA viruses. No significant differences were seen between the two NGS assays compared to PCR.

Utility of Extraction-Free SARS-CoV-2 Detection by RT-qPCR for COVID-19 Testing in a Resource-Limited Setting.

The COVID-19 epidemic had a profound impact on global health and the economy and Ghana was no exception to its far-reaching consequences. Regarding detection of the causative agent-the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), reverse-transcription-qPCR (RT-qPCR) is widely recognized as a very sensitive and reliable diagnostic technique used globally. There are, however, high operational costs in acquiring test kits, equipment, and accessories for RT-qPCR testing, which pose significant challenges in resource-limited settings. Hence, this proof-of-concept study set out to develop a more affordable COVID-19 protocol for use in low or lower-middle-income settings, such as Ghana, that would bypass the traditional extraction process using inexpensive reagents and evaluate the possibility of processing samples collected using wooden shaft swabs. Several less expensive media were used for the extraction-free process. Results demonstrated that direct RT-qPCR assay after 5 min heat inactivation of virus at 95 °C in 0.1× PBS or molecular grade water resulted in viral detection with quantification cycle (Cq) values that are comparable to results obtained following the extraction process. Also, wooden shaft swabs could be used for sampling if incubation times are kept to less than 6 h. The study demonstrates that extraction-free protocols are one way to minimize the cost of COVID-19 testing by RT-qPCR.

Inter-laboratory comparison of eleven quantitative or digital PCR assays for detection of proviral bovine leukemia virus in blood samples

Bovine leukemia virus (BLV) is the etiological agent of enzootic bovine leukosis and causes a persistent infection that can leave cattle with no symptoms. Many countries have been able to successfully eradicate BLV through improved detection and management methods. However, with the increasing novel molecular detection methods there have been few efforts to standardize these results at global scale. This study aimed to determine the interlaboratory accuracy and agreement of 11 molecular tests in detecting BLV. Each qPCR/ddPCR method varied by target gene, primer design, DNA input and chemistries. DNA samples were extracted from blood of BLV-seropositive cattle and lyophilized to grant a better preservation during shipping to all participants around the globe. Twenty nine out of 44 samples were correctly identified by the 11 labs and all methods exhibited a diagnostic sensitivity between 74 and 100%. Agreement amongst different assays was linked to BLV copy numbers present in samples and the characteristics of each assay (i.e., BLV target sequence). Finally, the mean correlation value for all assays was within the range of strong correlation. This study highlights the importance of continuous need for standardization and harmonization amongst assays and the different participants. The results underscore the need of an international calibrator to estimate the efficiency (standard curve) of the different assays and improve quantitation accuracy. Additionally, this will inform future participants about the variability associated with emerging chemistries, methods, and technologies used to study BLV. Altogether, by improving tests performance worldwide it will positively aid in the eradication efforts.

Atypical presentation of herpes simplex virus 2 primary infection: a case report

BackgroundCervicitis, an infectious or noninfectious inflammation of the cervix, encompasses a wide range of clinical conditions, from asymptomatic infections to severe lesions, making its diagnosis difficult. Acute cervicitis may develop into pelvic inflammatory disease. In patients with cervicitis, current guidelines recommend testing for herpes simplex virus when external genital lesions are present. Here, we present the case of a patient with an atypical primary herpes simplex virus 2 infection manifesting as cervicitis without genital lesions.Case presentationA 29-year-old Caucasian woman was hospitalized for pelvic inflammatory disease. The patient complained of severe suprapubic pain, fever, and heavy vaginal discharge. The external genitalia were unremarkable, so empirical antibiotic treatment was initiated. Despite 48 hours of well-administered antibiotic therapy, her complaints persisted. Polymerase chain reaction for possible microbial causes was negative for Chlamydia trachomatis and Neisseria gonorrhoeae. There was no bacterial vaginosis. Repeat gynecological examinations with endovaginal ultrasound revealed an enlarged cervix, and pelvic magnetic resonance imaging supported a diagnosis of cervicitis. At this point, additional screening for other sexually transmitted infections and infectious disease-related etiologies of cervicitis was performed, and the polymerase chain reaction analysis of newly isolated samples was positive for herpes simplex virus 2. No antiviral treatment was initiated given the delay in diagnosing herpes simplex virus 2 infection and the slow but spontaneous abatement of symptoms.ConclusionHerpes simplex virus infection should be considered as a possible cause of cervicitis, even in the absence of typical genital lesions. Early detection of herpes simplex virus allows early treatment, helping to reduce the duration and severity of symptoms and therefore potentially reducing recurrences and improving disease control. These data and data from future cases might spur changes in the guidelines on cervicitis testing and treatment.

The First Isolation of Insect-Specific Alphavirus (Agua Salud alphavirus) in Culex (Melanoconion) Mosquitoes in the Brazilian Amazon.

Advances in diagnostic techniques coupled with ongoing environmental changes have resulted in intensified surveillance and monitoring of arbovirus circulation in the Amazon. This increased effort has resulted in increased detection of insect-specific viruses among hematophagous arthropods collected in the field. This study aimed to document the first isolation of Agua Salud alphavirus in mosquitoes collected within the Brazilian Amazon. Arthropods belonging to the family Culicidae were collected within a forest fragment located in the Environmental Protection Area of the metropolitan region of Belem. Subsequently, these specimens were meticulously identified to the species level. Afterward, the collected batches were macerated, and the resulting supernatant was then inoculated into C6/36 and Vero cell cultures to facilitate viral isolation. The presence of arboviruses within the inoculated cell cultures was determined through indirect immunofluorescence analysis. Furthermore, positive supernatant samples underwent nucleotide sequencing to precisely identify the viral strains present. Notably, a batch containing Culex (Melanoconion) mosquitoes was identified to be positive for the genus Alphavirus via indirect immunofluorescence. This study is the first report on insect-specific alphavirus isolation in Brazil and the first-ever description of Agua Salud alphavirus isolation within Amazon Forest remnants.

Star-shaped flexible arm multivalent aptamers for largely improved virus binding affinity

Here we explore the click chemistry-based synthesis of high affinity multivalent aptamers and their application for virus detection. In contrast to attempts based on rigid constructs, such as aptamer-modified nanoparticles or hybridization-connected constructs, we propose highly flexible “star-shaped” multivalent aptamers to facilitate the cooperative binding to randomly distributed viral proteins on the surface of pleomorphic viruses such as the respiratory syncytial virus (RSV). Polyethylene glycols (PEGs) with 4 azide terminated flexible arms spanning from a pentaerythritol core were conjugated to aptamer strands with dibenzocyclooctyne and a fluorescent label at their, 3’ and 5’ terminus, respectively. By controlling the aptamer - PEG ratio and following electrophoretic separation all the different valency aptamer conjugates could be prepared. For proof of concept, we used an aptamer selected for the G protein of RSV. We show that significantly higher apparent affinities can be obtained by using multivalent aptamers for both the G protein and RSV virus than with the single stranded aptamer, i.e. with up to ca. 2 orders of magnitude. The inherent convenience of synthesizing fluorescently labelled multivalent aptamers leads to a versatile class of affinity ligands for sensing as demonstrated through the improved detection sensitivity of tetravalent aptamer tagged RSV by fluorescence nanoparticle tracking analysis.

Determination of airborne influenza virus and coronavirus infectivity using capsid integrity polymerase chain reaction

Accurate airborne virus monitoring is important for preventing the spread of infectious diseases. Although standard reverse transcription-quantitative polymerase chain reaction (RT-qPCR) can efficiently detect viral ribonucleic acid (RNA), it cannot determine whether the RNA is associated with active (infectious) or inactive (non-infectious) viruses. Plaque assay is the gold standard for determining viral infectivity but is laborious and time-consuming. This study explored the viral infectivity of H1N1 influenza virus and human coronavirus (HCoV-229E) using capsid integrity RT-qPCR, where virus samples were pretreated with reagents penetrating viruses with damaged capsids, impeding amplification by binding to their RNA. Therefore, the amplified signals corresponded solely to active viruses with undamaged capsids. Propidium monoazide (PMA) and platinum (IV) chloride (PtCl4) were used to investigate the effects of reagent concentration. Feasibility tests revealed that PtCl4 was more efficient than PMA, with optimal concentrations of 125–250 μM and 250–500 μM for H1N1 influenza virus and HCoV-229E, respectively. The results of percentage of active virus showed that capsid integrity RT-qPCR provided a trend similar to that of plaque assay, indicating an accurate measure of viral infectivity. Virus sampling in the laboratory and field highlighted the precision of this methodology for determining viral infectivity. Therefore, this methodology enables rapid and accurate detection of infectious airborne H1N1 influenza virus and HCoV-229E, allowing swift response to outbreaks.

Development of a paper-based fluorescent carbon quantum dots MIPs sensor for selective detection of lumpy skin disease virus.

Lumpy skin disease (LSD) is a contagious viral disease caused by the Lumpy Skin Disease virus (LSDV), a member of the Capripoxviridae family. Traditional LSDV diagnostic procedures proved to have challenges in terms of cross reactivity as well as limited sensitivity and specificity. Herein, we combined molecularly imprinted polymers (MIPs) and quantum dots (QDs) technology to develop a paper-based turn on fluorescence sensor for rapid, sensitive and selective detection of LSDV. Under optimal conditions, the sensor showed linear enhancement in fluorescence intensity with the increase of LSDV concentration and exhibited a detection limit of 101 log10 TCID50 per ml. It also presented high specificity towards LSDV compared to other viruses viz sheep pox virus (SPV). Furthermore, the proposed sensor was successfully tested with spiked and real LSDV samples, proving its potential to serve as a sensitive selective sensor for LSDV diagnosis. Based on our knowledge, this is the first record of a paper-based diagnostic sensor for LSDV utilizing a CQDs-MIPs turn-on mechanism.

Exploring waterborne viruses in groundwater: Quantification and Virome characterization via passive sampling and targeted enrichment sequencing

Aquifers, which provide drinking water for nearly half the world's population, face significant challenges from microbial contamination, particularly from waterborne viruses such as human adenovirus (HAdV), norovirus (NoV) and enterovirus (EV). This study, conducted as part of the UPWATER project, investigates the sources of urban groundwater contamination using viral passive sampling (VPS) and target enrichment sequencing (TES). We assessed the abundance of eight viral pathogens (HAdV, EV, NoV genogroup I and II, rotavirus, influenza A virus, hepatitis E virus and SARS-CoV-2) and investigated the virome diversity of groundwater in the aquifer of the Besòs River Delta in Catalonia. Over a period of 7 months, we collected 114 samples from the aquifer using nylon and nitrocellulose membranes to adsorb viruses over a 10-day period. Human faecal contamination was detected in nearly 50 % of the groundwater samples, with mean HAdV total counts ranging from 1.23E+02 to 3.66E+03 GC, and occasional detections of EV and NoV GI and GII. In addition, deep sequencing revealed a diverse virome in the aquifer, with detection of human pathogens, including adenovirus, astrovirus, calicivirus, enterovirus, herpesvirus, papillomavirus and rotavirus. Time-integrated sampling using VPS increases the likelihood of virus detection and, when combined with TES, can provide a deeper understanding of virus prevalence in this important water compartment. This approach is expected to streamline long-term monitoring efforts and enable small communities or water managers with limited resources to effectively manage their groundwater reservoirs.

Monitoring of Graphene Properties in the Process of Viral Biosensor Manufacturing

The properties of graphene chips with low reproducibility (LR) after photolithography (PLG) and graphene functionalization have been studied. It is shown that the introduction of additional cleaning after PLG can significantly increase the reproducibility of the parameters of processed graphene in biosensors. The use of dilute PBS solutions for virus detection makes it possible to increase the relative concentration sensitivity of biosensors by several times.