PCR Diagnostic Test Research Articles

Common Practice Patterns in the Diagnosis and Management of Acute Retinal Necrosis: A Survey Study of Uveitis Specialists

ABSTRACT Purpose Due to lack of large randomized clinical trials to determine best practices in treating acute retinal necrosis (ARN), there is not a clear consensus amongst ophthalmologists on how to best manage this potentially blinding condition. The aim of this study is to survey common practice patterns and analyze the factors that affect ophthalmologists’ management of ARN. Methods An anonymous survey was distributed to uveitis and retina specialists who are members of the American Uveitis Society (AUS) via email to query practice patterns regarding ARN. The survey included 22 questions with an additional 10 questions based on response. Survey question topics included demographic information, diagnostic testing, antiviral therapy, corticosteroids, and surgical procedures. Results 67 surveys were included for analysis. Most respondents (87%) always or frequently obtain intraocular aqueous fluid for diagnostic PCR testing. The majority of respondents would administer intravitreal antiviral injections to a unilateral immunocompetent ARN patient (67%), but would be even more likely to do so for a bilateral immunosuppressed ARN patient (87%). Respondents tend to treat ARN with systemic rather than local corticosteroids, with the majority (63%) of respondents initiating corticosteroid treatment 48 hours after treatment. Most respondents (79%) never perform a vitrectomy to manage ARN unless the patient has a retinal detachment or tear. The majority (63%) rarely or never perform prophylactic laser barricade, but may consider laser treatment if there is extensive retinal involvement. Conclusion Current practice patterns for diagnosis and management of ARN among AUS members generally align with the suggested practices outlined by the American Academy of Ophthalmology.

Shift in SARS-CoV-2 variants of concern from Delta to Omicron was associated with reduced hospitalizations, increased risk of breakthrough infections but lesser disease severity

BackgroundCOVID-19 epidemiology changed with the emergence of SARS-CoV-2 variants of concern (VOC). Pakistan administered mostly inactivated vaccines. We investigated the association between VOC and breakthrough infections in a mixed-vaccination-status population of Karachi. MethodsWe investigated SARS-CoV-2 VOC tested in 392 respiratory specimens collected between May and December 2021. Data for age, sex, hospital admission, vaccinations, together with CT values of the diagnostic PCR test were analyzed. ResultsThe median age of COVID-19 cases tested was 40 (27−57) years and 43.4% were female. Delta variants were most common (56.4%) followed by Alpha (15.9%), Omicron (12.2%), Beta/Gamma (11.3%), and others (4.3%). Eighteen percent of cases were hospitalized whereby, predominant VOC were Beta/Gamma (40.8%), Alpha (35.2%) and Delta (22.5%). Overall, 55.4% of individuals were fully vaccinated, 7.4% were partially vaccinated and 37.2% were unvaccinated. Most (74.6%) inpatients were unvaccinated. Vaccines comprised inactivated (85.34%), single-shot vector (8.62%), two-shot vector (3.02%) and mRNA (3.02%) types. Omicron variants showed lower viral loads as compared to Alpha, Beta/Gamma, and Delta (p = 0.017). The risk of infection with Delta and Omicron variants was higher, 8 weeks after vaccination. The majority of those with breakthrough infections after receiving inactivated vaccines acquired COVID-19 within 4 months of vaccination. ConclusionOur data highlights the shifting of VOC from Delta to Omicron during 2021 and that COVID-19 vaccinations reduced both hospitalizations and viral transmission. It informs on the increased risk of breakthrough infection within 8 weeks of vaccination, indicating the need for booster vaccinations.

Comparative evaluation of RT-PCR and antigen-based rapid diagnostic tests (Ag-RDTs) for SARS-CoV-2 detection: performance, variant specificity, and clinical implications.

The COVID-19 pandemic has highlighted the critical need for accurate and efficient diagnostic tools for detecting severe acute respiratory coronavirus 2 (SARS-CoV-2) infections. This study presents a comparison of two diagnostic tests: RT-PCR and antigen detection rapid diagnostic tests (Ag-RDTs). This study focused on their performance, variant specificity, and their clinical implications. A simultaneous testing of 268 samples was carried out for SARS-CoV-2 using RT-PCR and Ag-RDTs [flourescence immunoassay (FIA) and lateral flow immunoassay (LFIA)]. Viral load was quantified, and variant identification was performed using a PCR-based assay. The prevalence was found to be 30.2% using reverse transcription PCR (RT-PCR), 26.5% using FIA, and 25% using LFIA. When comparing the FIA and LFIA, the overall diagnostic performance was found to be 80.25% vs 76.54%, 96.79% vs 97.33%, 91.55% vs 90.51%, and 91.88% vs 92.56% for sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV), respectively. Both Ag-RDTs showed a strong agreement with RT-PCR (κ = 0.78-0.80). The overall accuracies of the FIA and LFIA were 92.41% and 92.13%, respectively. The FIA showed higher sensitivity (73.68%) and PPV (92.08%) than the LFIA (65.79% and 90.56%, respectively) in asymptomatic patients. At low Ct values (<25), both Ag-RDTs had 100% sensitivity, but the sensitivity reduced to 31.82% for FIA and 27.27% for LFIA at Ct values > 30. The diagnostic sensitivity of FIA compared to LFIA for detecting the Alpha variant was 78.85% vs. 69.23% and 72.22% vs. 83.33% for the Delta variant. Both Ag-RDTs had 100% sensitivity for detecting Omicron. Both Ag-RDTs performed well in patients with high viral loads and Omicron variant infections compared to those infected with Alpha and Delta variants. This study confirms the comparable performance of RT-PCR and Ag-RDTs, specifically FIA and LFIA, for SARS-CoV-2 detection. The FIA showed higher sensitivity and PPV in asymptomatic cases, while both Ag-RDTs exhibited strong agreement with RT-PCR results. Notably, Ag-RDTs, particularly FIA, proved effective in detecting the Omicron variant and cases with high viral loads, highlighting their potential clinical utility in managing the COVID-19 pandemic.IMPORTANCEThis study is of utmost importance in providing effective responses to manage the COVID-19 pandemic. It rigorously compares the diagnostic accuracy, variant specificity, and practical considerations of reverse transcription PCR (RT-PCR) and antigen detection rapid diagnostic tests (Ag-RDTs) for severe acute respiratory coronavirus 2 (SARS-CoV-2), answering critical questions. The results of this study will help healthcare professionals choose the appropriate testing methods, allocate resources effectively, and enhance public health strategies. Given the evolution of the virus, understanding the performance of these diagnostic tools is crucial to adapting to emerging variants. Additionally, the study provides insights into logistical challenges and accessibility issues, which will contribute to refining testing workflows, particularly in resource-limited settings. Ultimately, the study's impact extends to global healthcare, providing valuable information for policymakers, clinicians, and public health officials as they work together for mitigating the impact of the pandemic.

Inactivation of highly transmissible livestock and avian viruses including influenza A and Newcastle disease virus for molecular diagnostics.

There is a critical need for an inactivation method that completely inactivates pathogens at the time of sample collection while maintaining the nucleic acid quality required for diagnostic PCR testing. This inactivation method is required to alleviate concerns about transmission potential, minimize shipping complications and cost, and enable testing in lower containment laboratories, thereby enhancing disease diagnostics through improved turn-around time. This study evaluated a panel of 10 surrogate viruses that represent highly pathogenic animal diseases. These results showed that a commercial PrimeStore® molecular transport media (PSMTM) completely inactivated all viruses tested by >99.99%, as determined by infectivity and serial passage assays. However, the detection of viral nucleic acid by qRT-PCR was comparable in PSMTM and control-treated conditions. These results were consistent when viruses were evaluated in the presence of biological material such as sera and cloacal swabs to mimic diagnostic sample conditions for non-avian and avian viruses, respectively. The results of this study may be utilized by diagnostic testing laboratories for highly pathogenic agents affecting animal and human populations. These results may be used to revise guidance for select agent diagnostic testing and the shipment of infectious substances.

Using a Systems Engineering Approach to Build a PCR Testing System at a Medical School During the COVID-19 Pandemic.

During the COVID-19 pandemic, there was an increasing need to expand diagnostic testing in hospitals. At Keio University Hospital (KUH), clinical staff were concerned that the demand for PCR testing might exceed the capacity of the Clinical Laboratory. In response, basic researchers at Keio University School of Medicine (KUSM) set out to build a new, collaborative, PCR testing system. To be authorized to perform such diagnostic PCR testing, KUSM registered its core laboratory as an external clinical laboratory (ECL). In the pandemic, there was a pressure to build the PCR system quickly. Speed required discussions that developed a shared understanding of the unprecedented, new KUH/KUSM PCR system. To design, construct, and archive the new PCR testing system, we used a systems engineering (SE) approach. This included diagram visualization of functional flows and application of the Unified Architecture Framework (UAF), both of which are often used in system building. We considered daily demand for PCR testing at KUH and KUSM, and daily COVID-19 infections in Japan. We operated the collaborative PCR testing system from August 2020 to June 2022. Given public health insurance reimbursement policies, KUH focused on individuals with suspicious symptoms, while the ECL at KUSM screened samples from asymptomatic individuals. KUSM performed about half as many tests as KUH. Interviewing KUH staff revealed that diagrams helped promote a better understanding of the KUH/KUSM PCR testing system. When designing temporary systems that may be repurposed in the future, we suggest using an SE approach with diagrams and UAF perspectives. This approach will enable stakeholders to understand what is being proposed to be built, and facilitate achieving an informed consensus on the proposed system. We suggest that SE approaches should be widely used in projects that involve building and operating complex, collaborative systems, and documenting the process.

Genótipos de Circovírus Suíno Tipo 2 e PCV3 em Amostras Clínicas de Suínos no Brasil

Abstract Porcine circovirus type 2 (PCV2), an important pathogen in swine, causes PCV disease (PCVD). Although PCVD is effectively controlled using commercial vaccines, its clinical presentation is changing. Moreover, PCV2 is genetically evolving, with new genotypes emerging in vaccinated or unvaccinated pigs. In this study, we aimed to verify the presence of the PCV2a, PCV2b, and PCV2d genotypes in PCV-positive porcine samples. Furthermore, to identify coinfections between the PCV2 genotypes and/or PCV3, which can also induce disease in pigs, we employed a quick, effective, and low-cost PCR diagnostic test. In this study, 333 PCV2 PCR and clinically positive samples from various production stages and herds across Brazil were analyzed. Among these, 266 samples were genotyped, with PCV2b emerging as the most predominant genotype (56.77% of the positive samples), mainly observed in nursery pigs. PCV2d was also identified in 33.10% of the samples, primarily from finishing pigs and breeding sows. The employed PCR test was compared with a commercial kit, proving effective in PCV2 genotyping. This study demonstrates the significance of PCV2 genotyping, showing PCV2b as the most predominant genotype responsible for disease in pig farms in Brazil. PCV2a, the prevalent genotype used in commercial vaccines, was not detected in any of the analyzed samples. While pigs infected with other PCV2 genotypes may receive some heterologous protection from PCV2a vaccines, adequate diagnosis and vaccine monitoring for updates must be considered.

Designing a sustainable-resilient-responsive supply chain network considering uncertainty in the COVID-19 era.

Effective supply chain management is crucial for economic growth, and sustainability is becoming a key consideration for large companies. COVID-19 has presented significant challenges to supply chains, making PCR testing a vital product during the pandemic. It detects the presence of the virus if you are infected at the time and detects fragments of the virus even after you are no longer infected. This paper proposes a multi-objective mathematical linear model to optimize a sustainable, resilient, and responsive supply chain for PCR diagnostic tests. The model aims to minimize costs, negative societal impact caused by shortages, and environmental impact, using a scenario-based approach with stochastic programming. The model is validated by investigating a real-life case study in one of Iran's high-risk supply chain areas. The proposed model is solved using the revised multi-choice goal programming method. Lastly, sensitivity analyses based on effective parameters are conducted to analyze the behavior of the developed Mixed-Integer Linear Programming. According to the results, not only is the model capable of balancing three objective functions, but it is also capable of providing resilient and responsive networks. To enhance the design of the supply chain network, this paper has considered various COVID-19 variants and their infectious rates, in contrast to prior studies that did not consider the variations in demand and societal impact exhibited by different virus variants.

Chemical-physical investigations of nine types of nasopharyngeal swabs for the PCR analyses

During the SARS-Cov-2 pandemic, a test of samples collected through naso/oropharyngeal swab was set up; for this aim new swabs were developed. Materials and methods: The laboratory New Nanodiagnostics srl (Modena - Italy) analyzed nine different types of swabs used to collect human organic material for PCR diagnostic test for the SARS-CoV-2 infection. The swabs were observed under Optical Microscope and analyzed by Field Emission Gun Environmental Scanning Electron Microscope (FEGE-SEM) coupled with an Energy Dispersive System (EDS), to verify their morphology and chemical composition. Three different morphologies and nanostructures, along with their chemical composition were identified. Surprisingly enough, the presence of identified chemical contaminants like Titanium-Silicon-Aluminum or Silicon- Aluminium-Chromium-Manganese does not be understandable nor is it explained in the data sheet. Some fibers also present a nanostructured coating of Silicon-Zirconium. This might be of concern, as their presence could invalidate the accuracy of the PCR testing. In addition to that, the biocompatibility of the medical devices is discussed since a particular tendency of the glassy fibers to break has been verified. The presence of broken fibers in the nasal and oropharyngeal mucosa can cause irreversible damages. Finally, after their use, the swabs must be incinerated and their fumes contribute to the increase of the environmental pollution.

Staphylococcus epidermidis induced toxic shock syndrome (TSS) secondary to influenza infection

BackgroundTo date, few cases of TSS caused by coagulase negative (CoN) staphylococci have been reported in the literature. Recent data show that CoN staphylococci are capable of secreting a number of enterotoxins and cytotoxins, normally produced by S. aureus. Herewith, we describe a case of TSS caused by Staphylococcus epidermidis with a favorable outcome.Case presentationWe report a case of a 46-year-old man who developed TSS from S. epidermidis. The patient was admitted for a 7-day history of general malaise and headache following a recent influenza infection and a 3-day history of vomiting, diarrhea, diffuse erythroderma, and fever. The main laboratory findings on admission were leukopenia (WBC 800/mm3), thrombocytopenia (Plt count 78.000/mm3), elevated urea, creatine levels and increased inflammatory markers (CRP 368 mg/ml). The patient had clinical and radiological evidence of pneumonia with chest computed tomography (CT) showing diffuse bilateral airspace opacifications with air bronchogram. On the second day, a methicillin resistant S. epidermidis (MRSE) strain was detected in both sets of blood cultures, but the organism was unavailable for toxin testing. All other cultures and diagnostic PCR tests were negative. His clinical signs and symptoms fulfilled at that stage four out of five clinical criteria of TSS with a fever of 39 °C, diffuse erythroderma, multisystem involvement and hypotension. On the same day the patient was admitted to the ICU due to acute respiratory failure. The initial treatment was meropenem, vancomycin, levofloxacin, clindamycin, IVIG and steroids. Finger desquamation appeared on the 9th day of hospitalization, fulfilling all five clinical criteria for TSS.ConclusionsTo our knowledge, this is the first adult case with TSS induced by CoNS (MRSE) secondary to an influenza type B infection, who had favorable progression and outcome. Further research is warranted to determine how TSS is induced by the CoNS infections.

Polymorphism analysis of drug resistance markers in Plasmodium falciparum isolates from Benin

Like most countries in sub-Saharan African countries, Benin continues to bear a heavy malaria burden. In 2014, the National Malaria Control Programme (NMCP) changed its treatment policy, and recommended the use of artemisinin-based combination therapy (ACT) as first-line treatment for uncomplicated Plasmodium falciparum cases. The study presented here was conducted to investigate the impact of current antimalarial drug resistance on the country. Molecular surveillance targeting the Pfcrt, Pfmdr1, Pfkelch13, dhfr, and dhps genes was carried out on samples from patients positive for P. falciparum malaria by microscopy, LAMP and PCR diagnostic test. Molecular analysis was performed using targeted amplicon deep sequencing (TADS). In addition, the frequency of parasites with dual deletion of the histidine-rich protein 2 and 3 genes (pfhrp2 and pfhrp3), known to be responsible of the performance of HRP-based malaria rapid diagnostic tests (HRP-RDT), was estimated. Fifty-three falciparum samples collected at the Saint Jean de Dieu hospital in Tanguiéta, Benin, were tested. No Pfkelch13 validated or candidate artemisinin partial resistant variants were identified. A marked prevalence of Asn51Ile (N51I), Cys59Arg (C59R), and Ser108Asn (S108N) mutant alleles was found in the dhfr gene, representing the most frequent genotype (64%). Five-point mutations were detected in dhps, Ile431Val (I431V), Ser436Ala (S436A), Ala437Gly (A437G), Ala581Gly (A581G), Ala613Ser (A613S) of which the third was the most common (92%). No mutation was identified in dhps Lys540Glu (K540E). The quintuple mutant genotype resulting from the combination of the dhfr triple mutant (51I/59R/108N) with the dhps double mutant 436A/437G was detected at a frequency of 30%. Low levels of mutations in Pfcrt and no mutation at codon 86 in the Pfmdr1 DNA fragment were observed, whereas a high level of Tyr184Phe (Y184F) polymorphism in the Pfmdr1 gene was found. These results could be indicative, over a decade after the implementation of ACT therapy, of the return of chloroquine-sensitive but artemether-lumefantrine resistant falciparum genotypes in Benin. There was no evidence of HRP2 and HRP3 deletions. Data from the present study support the need for routine monitoring of molecular markers of antimalarial drug resistance as part of surveillance activities aimed to make informed treatment policy decisions at the national level.

High-throughput isolation of SARS-CoV-2 nucleocapsid antibodies for improved antigen detection

SARS-CoV-2 nucleocapsid protein (NP) is the main target for COVID-19-diagnostic PCR and antigen rapid diagnostic tests (Ag-RDTs). Ag-RDTs are more convenient than PCR tests for point-of-care testing or self-testing to identify the SARS-CoV-2 antigen. The sensitivity and specificity of this method depends mainly on the affinity and specificity of NP-binding antibodies; therefore, antigen-antibody binding is key elements for the Ag-RDTs. Here, we applied the high-throughput antibody isolation platform that has been utilized to isolate therapeutic antibodies against rare epitopes. Two NP antibodies were identified to recognize non-overlapping epitopes with high affinity. One antibody specifically binds to SARS-CoV-2 NP, and the other rapidly and tightly binds to SARS-CoV-2 NP with cross-reactivity to SARS-CoV NP. Furthermore, these antibodies were compatible with a sandwich enzyme-linked immunosorbent assay that exhibited enhanced sensitivity for NP detection compared to the previously isolated NP antibodies. Thus, the NP antibody pair is applicable to more sensitive and specific Ag-RDTs, highlighting the utility of a high-throughput antibody isolation platform for diagnostics development.

Correlation of C-Reactive Protein and Erythrocyte Sedimentation Rate in Covid-19 Cases

In December 2019, a mysterious pneumonia case was first reported in Wuhan, Hubei Province known as COVID-19, and its etiologic agent, SARS-Cov-2 virus. C-Reactive Protein (CRP) is a homopentameric protein that appears in inflammatory conditions. Erythrocyte sedimentation rate (ESR) increases in both acute and chronic inflammations. This study aimed to determine the correlation between CRP and ESR concentrations with the status of PCR diagnostic test results in patients. It was a correlation study with a cross-sectional approach. The data was taken from results from the first PCR and laboratory examination of the patients for COVID-19. The population of this study was patients under monitoring (PDP) for COVID-19 who were treated at a hospital in East Bekasi, West Java. The sample for this study was 65 people, consisting of 28 (43.1%) negative COVID-19 patients and 37 (56.9%) positive COVID-19 patients. The results showed that at the start of the PCR test for COVID-19, there was a weakly significant increase in CRP (r=0.311), and there was a significant relationship between the COVID-19 PCR results and the patient's CRP levels (p=0.040) in the age group 1-40 years. In addition, there was an increase in the respiratory rate in patients with high CRP with a weak significance (r=0.366), and a significant relationship between the CRP test results and the patients’ respiratory rate (p=0.026). In addition, there was a strong relationship between CRP increase and LED increase (p=0.000).

Emerging and re-emerging viral diseases, predisposing risk factors, and implications of international travel: a call for action for increasing vigilance and imposing restrictions under the current threats of recently emerging multiple Omicron subvariants.

Emerging and re-emerging viral diseases, predisposing risk factors, and implications of international travel: a call for action for increasing vigilance and imposing restrictions under the current threats of recently emerging multiple Omicron subvariants.

Изучение генетической вариабельности выделенных изолятов вируса лейкоза крупного рогатого скота в Белгородской области

Abstract. The purpose of the research Was to study genetic diversity of bovine leukemia virus isolates obtained in the Belgorod Region using restriction fragment length polymorphism method (RFLP). Scientific novelty. Bovine leukosis is one of the most common chronic infectious diseases of cattle in many countries of the world, which causes significant economic losses in the livestock industry. The typing of bovine leukemia virus (BLV), the study of its genetic structure, the evaluation of the mutation vector and a more detailed disclosure of the biological properties of the pathogen represent fundamental and applied value. Methods. The object of the research was 3-4-year-old cows infected with leukemia virus (n = 10), identified by serological methods in disadvantaged dairy farms. The immunodiffusion (ID) test, hematologic studies, PCR, genotyping, statistical processing of obtained data were conducted. Results. Conducted hematological studies determined the stage of the leukemic process in each animal. The target env fragment of the BLV gene (444 bp) was amplificated by 2-stage nested PCR, and this region was genotyped for all studied leukemia virus isolates using the restriction fragment length polymorphism (RFLP) method. In the course of the work, specific regions of the BLV env (gp51) gene, 970 bp long, were also obtained. We have given a primary assessment of the genetic diversity of BLV with the establishment of a genetic group (Belgian genotype according to RFLP). In the course of the work, specific regions of the env gp51 BLV gene, 444 bp long, were obtained. These fragments will be used for further DNA sequencing followed by phylogenetic analysis and determination of amino acid changes in the structure of the surface glycoprotein (gp51) of the bovine leukemia virus. Monitoring studies of BLV genotypes and the study of antigenic changes in the pathogen will allow timely development of the latest means of controlling and restricting the spread of bovine leukosis and improvement of diagnostic serological and PCR test systems.

Development and Validation of ScriptTaq COVID PCR: An In-House Multiplex rRT-PCR for Low-Cost Detection.

The COVID-19 pandemic necessitated an extensive testing for active SARS-CoV-2 infection. However, securing affordable diagnostic tests is a struggle for low-resource settings. We report herein the development and validation of an in-house multiplex real-time RT-PCR diagnostic test for the detection of active COVID-19 infection (ScriptTaq COVID PCR). Furthermore, we describe two methods for RNA extraction using either an in-house silica column or silica-coated magnetic beads to replace commercial RNA extraction kits. Different buffer formulations for silica column and silica-coated magnetic beads were tested and used for RNA isolation. Taq polymerase enzyme and thermostable reverse transcriptase enzyme were purified from bacterial clones. Primers/probes sequences published by the WHO and CDC were used for the qualitative detection of the RNA-dependent RNA polymerase (RdRp) and nucleocapsid (N) genes, respectively. ScriptTaq COVID PCR assay was able to detect up to 100 copies per reaction of the viral RdRP and N genes. The test demonstrated an overall agreement of 95.4%, a positive percent agreement (PPA) of 90.2%, and a negative percent agreement (NPA) of 100.0% when compared with two commercially available kits. ScriptTaq COVID PCR diagnostic test is a specific, sensitive, and low-cost alternative for low-resource settings.

Validation of reduced S-gene target performance and failure for rapid surveillance of SARS-CoV-2 variants.

SARS-CoV-2, the virus that causes COVID-19, has many variants capable of rapid transmission causing serious illness. Timely surveillance of new variants is essential for an effective public health response. Ensuring availability and access to diagnostic and molecular testing is key to this type of surveillance. This study utilized reverse transcription polymerase chain reaction (RT-PCR) and whole genome sequencing results from COVID-19-positive patient samples obtained through a collaboration between Aegis Sciences Corporation and Walgreens Pharmacy that has conducted more than 8.5 million COVID-19 tests at ~5,200 locations across the United States and Puerto Rico. Viral evolution of SARS-CoV-2 can lead to mutations in the S-gene that cause reduced or failed S-gene amplification in diagnostic PCR tests. These anomalies, labeled reduced S-gene target performance (rSGTP) and S-gene target failure (SGTF), are characteristic of variants carrying the del69-70 mutation, such as Alpha and Omicron (B.1.1.529, BA.1, and BA.1.1) lineages. This observation has been validated by whole genome sequencing and can provide presumptive lineage data following completion of diagnostic PCR testing in 24-48 hours from collection. Active surveillance of trends in PCR and sequencing results is key to the identification of changes in viral transmission and emerging variants. This study shows that rSGTP and SGTF can be utilized for near real-time tracking and surveillance of SARS-CoV-2 variants, and is superior to the use of SGTF alone due to the significant proportion of Alpha and Omicron (B.1.1.529, BA.1, and BA.1.1) lineages known to carry the del69-70 mutation and observed to have S-gene amplification. Adopting new tools and techniques to both diagnose acute infections and expedite identification of emerging variants is critical to supporting public health.

The effectiveness of mRNA vaccines to prevent SARS-CoV-2 infection and hospitalisation for COVID-19 according to the time elapsed since their administration in health professionals in the Valencian Autonomous Community (Spain)

The objective was to understand the effectiveness of the BNT162b2 and mRNA-1273 vaccines against SARS-CoV-2 in health professionals(HPs) in the Valencian Autonomous Community(Spain) who had completed a full vaccination regimen, both in terms of preventing infections and avoiding hospitalisations, according to the time elapsed since the vaccine administration.Case-controlled study with negative test results. HPs who had undergone at least one PCR or antigen(Ag) active infection diagnostic test(AIDT) to rule out SARS-CoV-2 infection between 25 January and 18 July 2021 were included. HPs with positive AIDT result were considered as cases and those with a negative result controls. Adjusted vaccine effectiveness(VEa) to prevent SARS-CoV-2 infection and its 95% confidence interval(95% CI) were calculated using the formula VEa = (1 − OR) × 100.The VEa for the prevention of SARS-CoV-2 infection 12 to 120 days after completing the full two-dose vaccine regimen was 91.6%(95%CI[89.6%,93.2%]) for the BNT162b2 vaccine and 95.2%(95%CI[88.3%,98.1%]) for the mRNA-1273 vaccine. After 120 days the VEa was 71.5%(95%CI[67.0%,75.5%]) for the BNT162b2 vaccine and 88.3%(95%CI[75.7,94.4%]) for the mRNA-1273 vaccine. The VEa for prevention of hospitalisation for COVID-19 for the complete two-dose regimen of mRNA vaccines (BNT162b2 and mRNA-1273) was 96.8%(95%CI[76.1%,99.6%]).The administration of the complete regimen of the BNT162b2 and mRNA-1273 vaccine against SARS-CoV-2 was highly effective for the prevention of COVID-19 cases in HPs when 12 to 120 days had elapsed since the second dose. However, said effectiveness decreased as time from the vaccine administration elapsed, although it was maintained for the prevention of hospitalisation of HPs.

COVID-19 – GLOBAL CHALLENGE OF OUR TIME

Coronavirus disease (COVID-19) is an infectious disease caused by the SARS-CoV-2 virus. The first diagnosed case of COVID 19 in Macedonia with a PCR diagnostic test was on February 27, 2020. The first diagnosed case of COVID 19 in Bitola with a PCR diagnostic test was on April 03, 2020. The purpose of this paper is to present our experiences in the fight against the new unknown disease, with special reference to clinical expressions and post-covid conditions. A great problem and challenge for the infectologists in Clinical Hospital, Bitola, were the therapeutic possibilities, especially in treatment of patients with multiple comorbid conditions. This paper covers patients who were treated in the Infectious Diseases department’s COVID-19 center at the Clinical Hospital, over a period of 2 years.The fight against the SARS-CoV 2 continues and we are already facing the new - seventh wave of the pandemic.

Saliva-based SARS-CoV-2 serology using at-home collection kits returned via mail

Serology provides tools for epidemiologic studies, and may have a role in vaccine prioritization and selection. Automated serologic testing of saliva, especially specimens that are self-collected at home and sent to a laboratory via the mail without refrigeration, could be a highly-scalable strategy for population-wide testing. In this prospective study, non-vaccinated patients were recruited after PCR testing to self-collect saliva and return their specimens via mail. Longitudinal specimens were analyzed in order to monitor seroconversion in the weeks after a diagnostic PCR test for SARS-CoV-2. Diverse users self-collected saliva and returned specimens via mail in compliance with shipping regulations. At our pre-established threshold (0.963 AU/mL), salivary IgG reactivity to full-length spike protein achieved 95.8% sensitivity and 92.4% specificity at 2–4 weeks after diagnostic testing, which is comparable to the typical sensitivity and specificity achieved for serum testing. Reactivity to N antigen also was detected with 92.6% sensitivity and 90.7% specificity at 4–8 weeks after diagnostic testing. Moreover, serologic testing for endemic coronaviruses performed in multiplex with SARS-CoV-2 antigens has the potential to identify samples that may require retesting due to effects of pre-analytical factors. The easy-to-use saliva collection kit, coupled with thresholds for positivity and methods of flagging samples for retest, provides a framework for large-scale serosurveillance of SARS-CoV-2.

Detection of subgenomic mRNA from endemic human coronavirus OC43 and NL63 compared to viral genomic loads, single virus detection and clinical manifestations in children with respiratory tract infections.

BackgroundThe importance of endemic human coronavirus (HCoV) in children has been insufficiently elucidated upon. Our aims were to develop subgenomic (sg) mRNA tests for HCoV species OC43 and NL63, and to evaluate the relationships to HCoV genomic loads, single HCoV detections and clinical manifestations.MethodsWe have used an 11-yearlong cohort study of children admitted with respiratory tract infection (RTI) and hospital controls. Nasopharyngeal aspirates were analyzed for HCoV subtypes OC43 and NL63 with in-house diagnostic PCR. Positive samples were tested with newly developed real-time PCRs targeting sg mRNA coding for the nucleocapsid protein.ResultsOC43 sg mRNA was detected in 86% (105/122) of available OC43-positive samples in the RTI group, and in 63% (12/19) of control samples. NL63 sg mRNA was detected in 72% (71/98) and 71% (12/17) of available NL63-positive patient and control samples, respectively. In RTI samples, sg mRNA detection was strongly associated with a Ct value <32 in both diagnostic PCR tests (OC43: OR = 54, 95% CI [6.8–428]; NL63: OR = 42, 95% CI [9.0–198]) and single NL63 detections (OR = 6.9, 95% CI [1.5–32]). Comparing RTI and controls, only OC43 was associated with RTI when adjusted for age (aOR = 3.2, 95% CI [1.1–9.4]).ConclusionWe found strong associations between OC43 and NL63 sg mRNA and high viral genomic loads. sg mRNA for OC43 was associated with RTI. The association between sg mRNA and clinical manifestations needs further evaluation.