ABSTRACT
Melioidosis is a particularly dangerous infection with endemic distribution caused by the Gram-negative microorganism from the pathogenicity group II Burkholderia pseudomallei. In endemic countries, melioidosis holds one of the leading places in mortality rate after HIV, tuberculosis and, in recent years, COVID-19. The natural ecological pathogen niches are located in tropical and subtropical climate zones, primarily in Southeast Asia and Australia, where its existence as a species is maintained in moist soil and water in a certain temperature environmental range. However, at present, more and more often cases of melioidosis are registered outside endemic territories, which emphasizes the relevance of improving the means and methods of laboratory diagnostics of this disease both for countries located in the zone of natural foci as well as for local healthcare of the countries after importation of this poorly known infection into their territory. In such countries, including the Russian Federation, the population has no natural immunity to the pathogen, and therefore this infection acquires even greater clinical and epidemiological significance. In the Volgograd Plague Control Research Institute, an erythrocyte antigenic melioidosis diagnostic agent for IHA was designed allowing to detect the presence of serum melioidosis antibodies. The diagnostic agent was obtained on the basis of a biological carrier - ram erythrocytes sensitized with isolated protein antigenic complexes of B. pseudomallei. The high analytical characteristics of the diagnostic agent were confirmed on sera models of immunized and recovering experimental animals. Using the obtained set of reagents, the level of serum antibodies against the causative agent of melioidosis was studied in residents from the 3 provinces of Vietnam (Ha Giang, Lang Son and Quang Ninh), as well as in control group composed of residents of the Volgograd region. In samples obtained from a non-endemic region, not more than 25% of cases contained IHA titers at lower than 1:10 dilution, which is apparently due to cross-reactivity of serum immunoglobulins. Positive serum samples from clinically healthy residents of Ha Giang, Lang Son and Quang Ninh provinces were at a titer of 1:10 detected in 71.5%, in dilutions of 1:20-1:80 - in 28.5% of cases. Thus, we believe that serum antibody titer of 1:80 established in the IHA results, has a diagnostic significance, reflecting the intensity of the anti-melioidosis populational immunity.
ABSTRACT
In resource-limited conditions such as the COVID-19 pandemic, on-site detection of diseases using the Point-of-care testing (POCT) technique is becoming a key factor in overcoming crises and saving lives. For practical POCT in the field, affordable, sensitive, and rapid medical testing should be performed on simple and portable platforms, instead of laboratory facilities. In this review, we introduce recent approaches to the detection of respiratory virus targets, analysis trends, and prospects. Respiratory viruses occur everywhere and are one of the most common and widely spreading infectious diseases in the human global society. Seasonal influenza, avian influenza, coronavirus, and COVID-19 are examples of such diseases. On-site detection and POCT for respiratory viruses are state-of-the-art technologies in this field and are commercially valuable global healthcare topics. Cutting-edge POCT techniques have focused on the detection of respiratory viruses for early diagnosis, prevention, and monitoring to protect against the spread of COVID-19. In particular, we highlight the application of sensing techniques to each platform to reveal the challenges of the development stage. Recent POCT approaches have been summarized in terms of principle, sensitivity, analysis time, and convenience for field applications. Based on the analysis of current states, we also suggest the remaining challenges and prospects for the use of the POCT technique for respiratory virus detection to improve our protection ability and prevent the next pandemic.
Subject(s)
COVID-19 , Viruses , Humans , Point-of-Care Testing , PandemicsABSTRACT
Melioidosis is a particularly dangerous infection with endemic distribution caused by the Gram-negative microorganism from the pathogenicity group II Burkholderia pseudomallei. In endemic countries, melioidosis holds one of the leading places in mortality rate after HIV, tuberculosis and, in recent years, COVID-19. The natural ecological pathogen niches are located in tropical and subtropical climate zones, primarily in Southeast Asia and Australia, where its existence as a species is maintained in moist soil and water in a certain temperature environmental range. However, at present, more and more often cases of melioidosis are registered outside endemic territories, which emphasizes the relevance of improving the means and methods of laboratory diagnostics of this disease both for countries located in the zone of natural foci as well as for local healthcare of the countries after importation of this poorly known infection into their territory. In such countries, including the Russian Federation, the population has no natural immunity to the pathogen, and therefore this infection acquires even greater clinical and epidemiological significance. In the Volgograd Plague Control Research Institute, an erythrocyte antigenic melioidosis diagnostic agent for IHA was designed allowing to detect the presence of serum melioidosis antibodies. The diagnostic agent was obtained on the basis of a biological carrier - ram erythrocytes sensitized with isolated protein antigenic complexes of B. pseudomallei. The high analytical characteristics of the diagnostic agent were confirmed on sera models of immunized and recovering experimental animals. Using the obtained set of reagents, the level of serum antibodies against the causative agent of melioidosis was studied in residents from the 3 provinces of Vietnam (Ha Giang, Lang Son and Quang Ninh), as well as in control group composed of residents of the Volgograd region. In samples obtained from a non-endemic region, not more than 25% of cases contained IHA titers at lower than 1:10 dilution, which is apparently due to cross-reactivity of serum immunoglobulins. Positive serum samples from clinically healthy residents of Ha Giang, Lang Son and Quang Ninh provinces were at a titer of 1:10 detected in 71.5%, in dilutions of 1:20-1:80 - in 28.5% of cases. Thus, we believe that serum antibody titer of 1:80 established in the IHA results, has a diagnostic significance, reflecting the intensity of the anti-melioidosis populational immunity.Copyright © 2022 Saint Petersburg Pasteur Institute. All rights reserved.
ABSTRACT
Melioidosis is a particularly dangerous infection with endemic distribution caused by the Gram-negative microorganism from the pathogenicity group II Burkholderia pseudomallei. In endemic countries, melioidosis holds one of the leading places in mortality rate after HIV, tuberculosis and, in recent years, COVID-19. The natural ecological pathogen niches are located in tropical and subtropical climate zones, primarily in Southeast Asia and Australia, where its existence as a species is maintained in moist soil and water in a certain temperature environmental range. However, at present, more and more often cases of melioidosis are registered outside endemic territories, which emphasizes the relevance of improving the means and methods of laboratory diagnostics of this disease both for countries located in the zone of natural foci as well as for local healthcare of the countries after importation of this poorly known infection into their territory. In such countries, including the Russian Federation, the population has no natural immunity to the pathogen, and therefore this infection acquires even greater clinical and epidemiological significance. In the Volgograd Plague Control Research Institute, an erythrocyte antigenic melioidosis diagnostic agent for IHA was designed allowing to detect the presence of serum melioidosis antibodies. The diagnostic agent was obtained on the basis of a biological carrier - ram erythrocytes sensitized with isolated protein antigenic complexes of B. pseudomallei. The high analytical characteristics of the diagnostic agent were confirmed on sera models of immunized and recovering experimental animals. Using the obtained set of reagents, the level of serum antibodies against the causative agent of melioidosis was studied in residents from the 3 provinces of Vietnam (Ha Giang, Lang Son and Quang Ninh), as well as in control group composed of residents of the Volgograd region. In samples obtained from a non-endemic region, not more than 25% of cases contained IHA titers at lower than 1:10 dilution, which is apparently due to cross-reactivity of serum immunoglobulins. Positive serum samples from clinically healthy residents of Ha Giang, Lang Son and Quang Ninh provinces were at a titer of 1:10 detected in 71.5%, in dilutions of 1:20-1:80 - in 28.5% of cases. Thus, we believe that serum antibody titer of 1:80 established in the IHA results, has a diagnostic significance, reflecting the intensity of the anti-melioidosis populational immunity. Copyright © 2022 Saint Petersburg Pasteur Institute. All rights reserved.
ABSTRACT
Melioidosis is a particularly dangerous infection with endemic distribution caused by the Gram-negative microorganism from the pathogenicity group II Burkholderia pseudomallei. In endemic countries, melioidosis holds one of the leading places in mortality rate after HIV, tuberculosis and, in recent years, COVID-19. The natural ecological pathogen niches are located in tropical and subtropical climate zones, primarily in Southeast Asia and Australia, where its existence as a species is maintained in moist soil and water in a certain temperature environmental range. However, at present, more and more often cases of melioidosis are registered outside endemic territories, which emphasizes the relevance of improving the means and methods of laboratory diagnostics of this disease both for countries located in the zone of natural foci as well as for local healthcare of the countries after importation of this poorly known infection into their territory. In such countries, including the Russian Federation, the population has no natural immunity to the pathogen, and therefore this infection acquires even greater clinical and epidemiological significance. In the Volgograd Plague Control Research Institute, an erythrocyte antigenic melioidosis diagnostic agent for IHA was designed allowing to detect the presence of serum melioidosis antibodies. The diagnostic agent was obtained on the basis of a biological carrier - ram erythrocytes sensitized with isolated protein antigenic complexes of B. pseudomallei. The high analytical characteristics of the diagnostic agent were confirmed on sera models of immunized and recovering experimental animals. Using the obtained set of reagents, the level of serum antibodies against the causative agent of melioidosis was studied in residents from the 3 provinces of Vietnam (Ha Giang, Lang Son and Quang Ninh), as well as in control group composed of residents of the Volgograd region. In samples obtained from a non-endemic region, not more than 25% of cases contained IHA titers at lower than 1:10 dilution, which is apparently due to cross-reactivity of serum immunoglobulins. Positive serum samples from clinically healthy residents of Ha Giang, Lang Son and Quang Ninh provinces were at a titer of 1:10 detected in 71.5%, in dilutions of 1:20-1:80 - in 28.5% of cases. Thus, we believe that serum antibody titer of 1:80 established in the IHA results, has a diagnostic significance, reflecting the intensity of the anti-melioidosis populational immunity. Copyright © 2022 Saint Petersburg Pasteur Institute. All rights reserved.
ABSTRACT
Since its emergence in late 2019, the coronavirus disease 2019 (COVID-19) pandemic has caused severe disruption to key aspects of human life globally and highlighted the need for timely, adaptive, and accessible pandemic response strategies. Here, we introduce the cell-free dot blot (CFDB) method, a practical and ultra-low-cost immune diagnostic platform capable of rapid response and mass immunity screening for the current and future pandemics. Similar in mechanism to the widely used enzyme-linked immunosorbent assays (ELISAs), our method is novel and advantageous in that (i) it uses linear DNA to produce the target viral antigen fused to a SpyTag peptide in a cell-free expression system without the need for traditional cloning and antigen purification, (ii) it uses SpyCatcher2-Apex2, an Escherichia coli-produced peroxidase conjugate as a universal secondary detection reagent, obviating the need for commercial or sophisticated enzyme conjugates, and (iii) sera are spotted directly on a nitrocellulose membrane, enabling a simple "dipping" mechanism for downstream incubation and washing steps, as opposed to individual processing of wells in a multiwell plate. To demonstrate the utility of our method, we performed CFDB to detect anti-severe acute respiratory syndrome coronavirus 2 nucleocapsid protein antibodies in precharacterized human sera (23 negative and 36 positive for COVID-19) and hamster sera (16 negative and 36 positive for COVID-19), including independent testing at a collaborating laboratory, and we show assay performance comparable to that of conventional ELISAs. At a similar capacity to 96-well plate ELISA kits, one CFDB assay costs only ~$3 USD. We believe that CFDB can become a valuable pandemic response tool for adaptive and accessible sero-surveillance in human and animal populations. IMPORTANCE The recent COVID-19 pandemic has highlighted the need for diagnostic platforms that are rapidly adaptable, affordable, and accessible globally, especially for low-resource settings. To address this need, we describe the development and functional validation of a novel immunoassay technique termed the cell-free dot blot (CFDB) method. Based on the principles of cell-free synthetic biology and alternative dot blotting procedures, our CFDB immunoassay is designed to provide for timely, practical, and low-cost responses to existing and emerging public health threats, such as the COVID-19 pandemic, at a similar throughput and comparable performance as conventional ELISAs. Notably, the molecular detection reagents used in CFDB can be produced rapidly in-house, using established protocols and basic laboratory infrastructure, minimizing reliance on strained commercial reagents. In addition, the materials and imaging instruments required for CFDB are the same as those used for common Western blotting experiments, further expanding the reach of CFDB in decentralized facilities.
ABSTRACT
The novel coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has severely impacted human health and the health management system globally. The ongoing pandemic has required the development of more effective diagnostic strategies for restricting deadly disease. For appropriate disease management, accurate and rapid screening and isolation of the affected population is an efficient means of containment and the decimation of the disease. Therefore, considerable efforts are being directed toward the development of rapid and robust diagnostic techniques for respiratory infections, including SARS-CoV-2. In this article, we have summarized the origin, transmission, and various diagnostic techniques utilized for the detection of the SARS-CoV-2 virus. These higher-end techniques can also detect the virus copy number in asymptomatic samples. Furthermore, emerging rapid, cost-effective, and point-of-care diagnostic devices capable of large-scale population screening for COVID-19 are discussed. Finally, some breakthrough developments based on spectroscopic diagnosis that could revolutionize the field of rapid diagnosis are discussed.
ABSTRACT
The mosquito-borne disease caused by the Rocio virus is a neglected threat, and new immune inputs for serological testing are urgently required for diagnosis in low-resource settings and epidemiological surveillance. We used in silico approaches to identify a specific antigenic peptide (p_ROCV2) in the NS1 protein of the Rocio virus that was theoretically predicted to be stable and exposed on its surface, where it demonstrated key properties allowing it to interact with antibodies. These findings related to the molecular dynamics of this peptide provide important insights for advancing diagnostic platforms and investigating therapeutic alternatives.
Subject(s)
Flavivirus , Molecular Dynamics Simulation , Animals , Immunologic Tests , Molecular Docking Simulation , Peptides , Viral Nonstructural Proteins/chemistryABSTRACT
BACKGROUND: The emergence of novel viruses poses severe challenges to global public health highlighting the crucial necessity for new antivirals. MAIN BODY: Monoclonal antibodies (mAbs) are immunoglobulins that bind to a single epitope. Mouse mAbs are generated by classic hybridoma technology and are mainly used for immunodiagnostics. For immunotherapy, it is critical to use monoclonal antibodies in their human form to minimize adverse reactions. They have been successfully used to treat numerous illnesses, accordingly, an increasing number of mAbs, with high potency against emerging viruses is the target of every biopharmaceutical company. The diagnostic and therapeutic mAbs market grows rapidly into a multi-billion-dollar business. Biopharmaceuticals are innovative resolutions which revolutionized the treatment of significant chronic diseases and malignancies. Currently, a variety of therapeutic options that include antiviral medications, monoclonal antibodies, and immunomodulatory agents are available for the management of COVID-19. SHORT CONCLUSION: The invasion of mAbs in new medical sectors will increase the market magnitude as it is expected to generate revenue of about 300 billion $ by 2025. In the current mini-review, the applications of monoclonal antibodies in immune-diagnosis and immunotherapy will be demonstrated, particularly for COVID-19 infection and will focus mainly on monoclonal antibodies in the market.
Subject(s)
Antibodies, Monoclonal , COVID-19 , Animals , Antibodies, Viral/therapeutic use , Epitopes , Humans , Hybridomas , Mice , SARS-CoV-2ABSTRACT
The quickly emerged global COVID-19 pandemic raised a desperate need in the development of pro-tecting vaccines targeting this disease. Therefore, a generation of effective producers of recombinant SARS-CoV-2 proteins became an urgent task. Its resolving contributes to the study of functional SarS-CoV-2 properties, as well as will allow developing the domestic COVID-19 vaccine in Ukraine, thus playing an important strategic role in tackling the pandemics. The aim of the study was to generate prokaryotic and eukaryotic producers of recombinant SarS-CoV-2 proteins and to isolate nucleocapsid (N) protein, receptor-binding do-main (RBD) of spike (S) protein, as well as RBD fused to the carrier – diphtheria toxoid CRM197. For this purpose, appropriate genetic constructs, in particular, replication deficient recombinant AdvC5-based adenoviral vectors expressing the SarS-CoV-2 proteins and CRM197-fused conjugate were created through methods of molecular biology and genetic engineering. Restriction analysis and/or dna sequencing confirmed that we created the correct constructs. Immobilized metal affinity chromatography was used to purify the recombinant proteins. Compliance of their properties was confirmed by the results from polyacrylamide gel electrophoresis, Western blotting, immunoenzymatic assay and MALDI-TOF mass spectrometry. As a result, we generated E. coli Rosetta (de3) bacterial strain and HEK293 cell line producing recombinant SARS-CoV-2 proteins and CRM197-based fusion. In addition, pure N protein, RBD of S protein and RBD-crm197 fusion protein were isolated. The obtained recombinant SarS-CoV-2 proteins can be used to study immunogenic and antigenic properties of the SarS-CoV-2 proteins. Cells producing recombinant SarS-CoV-2 proteins and RBD-crm197 fusion protein are able to provide cheap and safe synthesis of the antigenic substances for domestic development and production of immunodiagnostics for COVID-19 and COVID-19 vaccines in Ukraine. © 2021 Krynina O. I. et al.
ABSTRACT
Vaccine bears hope to bring COVID-19 pandemic under control. With limited supply, vaccines must be utilized efficiently to provide protection to those who need it most. Currently, no practical framework has been proposed to ensure fair vaccine allocation at individual level, which is a recognized problem. We propose here an evidence-based decision-making framework for COVID-19 vaccine appropriation that prioritizes vaccine doses to individuals based on their immunological status, or immuno-triaging. To ensure successful implementation of the proposed framework, point-of-care (POC) immunodiagnostic testing is needed to quickly ramp up the testing capability. Considerations for deploying POC immunodiagnostic testing at such a large scale are discussed. We hope that the proposed immunological decision-making framework for evidence-based COVID-19 vaccine appropriation provides an objective approach to ensure fair and efficient utilization of the scarce vaccine resource at the individual level that also maximizes the collective societal benefit.
Subject(s)
COVID-19 Vaccines , COVID-19 , Humans , Pandemics , Point-of-Care Systems , SARS-CoV-2 , VaccinationABSTRACT
Humoral immunity has emerged as a vital immune component against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Nevertheless, a subset of recovered Coronavirus Disease-2019 (COVID-19) paucisymptomatic/asymptomatic individuals do not generate an antibody response, constituting a paradox. We assumed that immunodiagnostic assays may operate under a competitive format within the context of antigenemia, potentially explaining this phenomenon. We present a case where persistent antigenemia/viremia was documented for at least 73 days post-symptom onset using 'in-house' methodology, and as it progressively declined, seroconversion took place late, around day 55, supporting our hypothesis. Thus, prolonged SARS-CoV-2 antigenemia/viremia could mask humoral responses, rendering, in certain cases, the phenomenon of 'non-responders' a misnomer.
Subject(s)
Antibodies, Viral/blood , Antigens, Viral/blood , Antigens, Viral/immunology , COVID-19 Serological Testing/standards , COVID-19/diagnosis , SARS-CoV-2/immunology , Antibodies, Viral/metabolism , Antigens, Viral/metabolism , Binding Sites, Antibody , COVID-19/blood , COVID-19/immunology , COVID-19/virology , COVID-19 Serological Testing/statistics & numerical data , Humans , Immunity, Humoral/immunology , Immunoglobulin G/blood , Male , Sensitivity and Specificity , Seroconversion , Young AdultABSTRACT
Accurate diagnosis of acute severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection is critical for appropriate management of patients with this disease. We examined the possible complementary role of laboratory-developed class-specific clinical serology in assessing SARS-CoV-2 infection in hospitalized patients. Serological tests for immunoglobulin G (IgG), IgA, and IgM antibodies against the receptor binding domain (RBD) of SARS-CoV-2 were evaluated using samples from real-time reverse transcription-quantitative PCR (qRT-PCR)-confirmed inpatient coronavirus disease 2019 (COVID-19) cases. We analyzed the influence of timing and clinical severity on the diagnostic value of class-specific COVID-19 serology testing. Cross-sectional analysis revealed higher sensitivity and specificity at lower optical density cutoffs for IgA in hospitalized patients than for IgG and IgM serology (IgG area under the curve [AUC] of 0.91 [95% confidence interval {CI}, 0.89 to 0.93] versus IgA AUC of 0.97 [95% CI, 0.96 to 0.98] versus IgM AUC of 0.95 [95% CI, 0.92 to 0.97]). The enhanced performance of IgA serology was apparent in the first 2 weeks after symptom onset and the first week after PCR testing. In patients requiring intubation, all three tests exhibit enhanced sensitivity. Among PCR-negative patients under investigation for SARS-CoV-2 infection, 2 out of 61 showed clear evidence of seroconversion IgG, IgA, and IgM. Suspected false-positive results in the latter population were most frequently observed in IgG and IgM serology tests. Our findings suggest the potential utility of IgA serology in the acute setting and explore the benefits and limitations of class-specific serology as a complementary diagnostic tool to PCR for COVID-19 in the acute setting.
Subject(s)
COVID-19 , SARS-CoV-2 , Antibodies, Viral , Cross-Sectional Studies , Humans , Immunoglobulin M , Sensitivity and SpecificityABSTRACT
INTRODUCTION: COVID-19 is a recent emerging pandemic whose prognosis is still unclear. Diagnostic tools are the main players that not only indicate a possible infection but can further restrict the transmission and can determine the extent to which disease progression would occur. AREAS COVERED: In this paper, we have performed a narrative and critical review on different technology-based diagnostic strategies such as molecular approaches including real-time reverse transcriptase PCR, serological testing through enzyme-linked immunosorbent assay, laboratory and point of care devices, radiology-based detection through computed tomography and chest X-ray, and viral cell cultures on Vero E6 cell lines are discussed in detail to address COVID-19. This review further provides an overview of emergency use authorized immunodiagnostic and molecular diagnostic kits and POC devices by FDA for timely and efficient conduction of diagnostic tests. The majority of the literature cited in this paper is collected from guidelines on protocols and other considerations on diagnostic strategies of COVID-19 issued by WHO, CDC, and FDA under emergency authorization. EXPERT OPINION: Such information holds importance to the health professionals in conducting error-free diagnostic tests and researches in producing better clinical strategies by addressing the limitations associated with the available methods.