Detection Of Antibodies Research Articles

Development and Bayesian validation of a competitive inhibition ELISA for detection of antibodies against Brucella abortus in cattle

Bovine brucellosis is a zoonotic disease caused by Brucella abortus, responsible for abortions in cows. It is endemic in low- and middle-income countries, where the brucellosis control and eradication programs are based on compulsory vaccination, detection of infected cattle through serologic assays, and culling of infected animals at slaughterhouses. The development of high sensitivity and specificity, and low-cost serologic assays guarantee their implementation in countries where the disease is endemic. The aim of the present study was to develop and validate a competitive inhibition enzyme-linked immune assay (ciELISA) to detect anti-B. abortus antibodies in sera from cattle. The developed ciELISA was validated using 2833 serum samples from dairy and beef cattle. From these, 1515 sera were from uninfected cows that belonged to free of brucellosis herds and 1318 were from infected cows that belonged positive to brucellosis herds. Sera were analyzed with the developed ciELISA, the buffer plate antigen (BPA) test, and the complement fixation test (CFT). The brucellosis status of the herds was officially established according to the country legislation and consistent for at least 5 years and was defined for each cow using the CFT as gold standard. The cutoff for the ciELISA was calculated using a ROC curve and its sensitivity and specificity were analyzed using the Bayesian Latent Class Model (BLCM) approach. The agreement among tests was calculated using the kappa (κ) value. In addition, 15 calves were vaccinated with 3 × 1010 viable cells of B. abortus Strain 19 vaccine, and the dynamics of antibodies were measured by CFT, buffered plate antigen (BPA) test, and the developed ciELISA. The obtained cutoff for ciELISA was ≥ 47 percentage of inhibition (% I), at the BLCM approach the sensitivity was 99.01 % (95 % CI: 97.55–100) and the specificity 98.74 % (95 % CI: 97.68–99.8). The κ between the ciELISA and BPA was κ = 0.88 and between the ciELISA and CFT κ = 0.95. Antibodies against B. abortus were detected in all the vaccinated calves 7 days after vaccination (AV) by the three assays, at day 135 AV all the calves were negative to CFT (15/15), 93.3 % (14/15) to ciELISA and 73.3 % (11/15) to BPA, and at day 190 AV all the calves were negative to the three assays. The developed ciELISA showed a very good performance, could detect the majority of vaccinated animals as negative after 135 days and could be used for the detection of anti-B. abortus antibodies in serum samples for the brucellosis control and eradication program.

Detection and Serological Evidence of European Bat Lyssavirus 1 in Belgian Bats between 2016 and 2018.

Lyssaviruses are neurotropic viruses capable of inducing fatal encephalitis. While rabies virus has been successfully eradicated in Belgium, the prevalence of other lyssaviruses remains uncertain. In this study, we conducted a survey on live animals and passive surveillance to investigate the presence of lyssaviruses in Belgium. In 2018, a total of 113 saliva samples and 87 blood samples were collected from bats. Saliva was subjected to RT-qPCR to identify lyssavirus infections. Additionally, an adapted lyssavirus neutralisation assay was set up for the detection of antibodies neutralising EBLV-1 in blood samples. Furthermore, we examined 124 brain tissue samples obtained from deceased bats during passive surveillance between 2016 and 2018. All saliva samples tested negative for lyssaviruses. Analysis of the blood samples uncovered the presence of lyssavirus-neutralising antibodies in five bat species and 32% of samples with a wide range depending on bat species, suggesting past exposure to a lyssavirus. Notably, EBLV-1 was detected in brain tissue samples from two Eptesicus serotinus specimens collected in 2016 near Bertrix and 2017 near Étalle, confirming for the first time the presence of EBLV-1 in Belgium and raising awareness of the potential risks associated with this species of bats as reservoirs of the virus.

Molecular Characterization of Lineage-IV Peste Des Petits Ruminants Virus and the Development of In-House Indirect Enzyme-Linked Immunosorbent Assay (IELISA) for its Rapid Detection”

Peste des petits ruminants (PPRV), a highly contagious viral disease, causes significant economic losses concerning sheep and goats. Recently, PPR viruses (PPRVs), have adopted new hosts and lineage IV of PPRVs represents genetic diversity within the same lineage. 350 samples, including blood, swabs, and tissues from sheep/goats, were collected during the 2020–2021 disease outbreaks in Pakistan. These samples were analysed through RT-PCR and three isolates of PPRV with accession numbers, MW600920, MW600921, and MW600922, were submitted to GenBank, based on the partial N-gene sequencing. This analysis provides a better understanding of genetic characterizations and a targeted RT-PCR approach for rapid PPRV diagnosis. An IELISA test was developed using the semi-purified antigen MW600922 isolate grown in Vero cells. The PPRV isolates currently present high divergence with the Turkish strain; conversely, similarities equivalent to 99.73% were observed for isolates collected from Pakistan. The developed indirect ELISA (IELISA) test demonstrated antibody detection rates at dilutions of 1:200 for antibodies (serum) and 1:32 for antigens. In comparison to cELISA, high specificity (85.23%) and sensitivity (90.60%) rates were observed. In contrast to the virus neutralization test (VNT), IELISA was observed to be 100% specific and 82.14% sensitive in its results. Based on these results, serological surveys conducted for PPR antibodies using IELISA can be a more effective strategy on a larger scale. Furthermore, our results demonstrate a significant breakthrough in the research in terms of cost-effectiveness and storage efficiency, and the developed IELISA test is highly recommended for use in developing countries.

Diagnosis of Human Cystic Echinococcosis by detecting Antigen B in Serum and Urine using Nanomagnetic Beads-Enzyme Linked Immunosorbent Assay

Abstract Cystic echinococcosis (CE) is a zoonotic parasitic disease caused by the larval stage; hydatid cyst, of E. granulosus. The disease is usually asymptomatic, and mostly presents by pressure symptoms according to its location. The diagnosis of CE is currently based on imaging techniques, which may not be available in some areas and antibody detection in serum, with several known drawbacks. Conversely, antigen detection has proven its efficacy in serodiagnosis of CE. The study aimed to test the diagnostic efficacy of Nanomagnetic beads-enzyme linked immunosorbent assay (NMB-sandwich ELISA) for diagnosis of CE by detection of E. granulosus AgB in serum and urine samples in comparison to conventional sandwich ELISA. The study included 42 individuals classified into 3 groups; CE infection group (Gp I), other parasitic infections group (Gp II) and healthy control group (Gp III). E. granulosus AgB was prepared from human hydatid cysts and used to produce anti- E. granulosus AgB-IgG-pAbs in rabbit which was used to detect AgB level in serum and urine samples by sandwich ELISA and NMB-sandwich ELISA. Using sandwich ELISA, AgB was detected in serum samples of 88.9% of CE patients with sensitivity and specificity of 88.9% and 91.7%, respectively. In urine samples, sandwich ELISA detected AgB in 83.3% of CE patients and the sensitivity and specificity were 83.3% and 87.5%, respectively. NMB-sandwich ELISA detected AgB in serum samples of 94.4% of CE patients with a sensitivity and specificity of 94.4% and 95.8%, respectively, whereas in urine samples it was detected in 88.9% of CE cases and the sensitivity and specificity were 88.9% and 95.8%, respectively. There was no statistically significant difference (p > 0.05) between the use of serum and urine samples for detection of AgB when examined by either sandwich ELISA or NMB- sandwich ELISA. The best test accuracy was obtained with NMB-sandwich ELISA detecting AgB in serum samples (95.2%), whereas, it showed a slightly lower accuracy of 92.9% when using urine samples. It was concluded that NMB-sandwich ELISA is efficient for detection of AgB in serum samples. Moreover, urine samples might be a good alternative to serum samples for the detection of AgB showing reasonable sensitivity, specificity and accuracy.

Seroprevalence of Arboviruses in a Malaria Hyperendemic Area in Southern Mali.

Diagnostics for febrile illnesses other than malaria are not readily available in rural sub-Saharan Africa. This study assessed exposure to three mosquito-borne arboviruses-dengue virus (DENV), Zika virus (ZIKV), and chikungunya virus (CHIKV)-in southern Mali. Seroprevalence for DENV, CHIKV, and ZIKV was analyzed by detection of IgG antibodies and determined to be 77.2%, 31.2%, and 25.8%, respectively. Among study participants, 11.3% were IgG-positive for all three arboviruses. DENV had the highest seroprevalence rate at all sites; the highest seroprevalence of CHIKV and ZIKV was observed in Bamba. The seroprevalence for all three arboviruses increased with age, and the highest seroprevalence was observed among adults older than 50 years. The prevalence of Plasmodium spp. in the cohort was analyzed by microscopy and determined to be 44.5% (N = 600) with Plasmodium falciparum representing 95.1% of all infections. This study demonstrates the co-circulation of arboviruses in a region hyperendemic for malaria and highlights the needs for arbovirus diagnostics in rural sub-Saharan Africa.

Biofunctional photoelectrochemical/electrochemical immunosensor based on BiVO4/BiOI-MWCNTs and Au@PdPt for alpha-fetoprotein detection

A biofunctional immunosensor combining photoelectrochemical (PEC) and electrochemical (EC) was proposed for the quantitative detection of the liver cancer marker alpha-fetoprotein (AFP) in human blood. BiVO4/BiOI-MWCNTs photoactive materials were first prepared on conductive glass FTO, and the photoelectrode was functionalized by chitosan and glutaraldehyde. Then, the AFP capture antibody (Ab1) was successfully modified on the photoelectrode, and the label-free rapid detection of AFP antigen was achieved by PEC. In addition, Au@PdPt nanospheres were also used as a marker for binding to AFP detection antibody (Ab2). Due to the excellent catalytic properties of Au@PdPt in EC reaction, a signal increase in the EC response can be achieved when Ab2 binds to the AFP antigen, which ensures high sensitivity for the detection of AFP. The detection limits of PEC and EC are 0.050 pg/mL and 0.014 pg/mL, respectively. The sensor also possesses good specificity, stability and reproducibility, shows excellent performance in the detection of clinical samples and has good clinical applicability.

Prevention of vertical transmission of hepatitis B: A retrospective review of a 5-year maternal-infant cohort in London.

The World Health Organization (WHO) estimates that 3.5% of the population live with hepatitis B virus (HBV); migrants to Europe are disproportionately affected. UK birth dose HBV vaccination is limited to infants born to those living with HBV (LWHBV). High-risk infants (high maternal infectivity, low birthweight) also receive HBV immunoglobulin (HBIG). The Family Hepatitis Clinic follows infants and those LWHBV working towards WHO goals of combating viral hepatitis by 2030. A trust-wide electronic note review of outcomes for infants born to those LWHBV (2016-2020). Two hundred and eighty-three infants, 134 (47%) females, born to those LWHBV were referred. Two hundred and thirty-one (82%) attended follow-up with a vertical transmission rate of 0%. Twenty (7%) individuals LWHBV received tenofovir disoproxil fumerate in pregnancy; median viral load (VL) at initiation 125 416 376 DNA IU/mL, one having birth VL. Twenty-eight (10%) infants were stratified as high risk and all received HBIG and birth dose vaccination with 9 (32%) subsequently lost to follow-up, compared to 48 (19%) low-risk infants. 267/283 (94%) had birth dose vaccination documented and 206/283 (73%) received at least four vaccine doses. 215/283 (76%) infants had serology by 24 months; 17 (6%) with suboptimal vaccine responses: hepatitis B surface antibody <100 IU/mL. Serology before 18 months resulted in higher rates of maternal hepatitis B core antibody detection (15% vs. 3%). Prevention of vertical transmission of HBV was universal in those attending, although high-risk infants were more likely lost to follow up. HBV post-vaccine serological protection was comparable with national data from 2021 (77% >4 doses, 77% HBsAb >100).

Identification of circulating miRNA biomarkers in leptospirosis for early detection: A promising diagnostic approach

Leptospirosis is a zoonotic disease of global significance, contributing to morbidity and mortality worldwide. It is endemic to tropical regions, with outbreaks during monsoons. The disease manifestations are similar to that of other febrile illness such as dengue, malaria hence often misdiagnosed and underreported. The zoonoses if undetected, progresses to cause severe life-threatening complications also known as Weil's disease. Routine diagnostic tests are based on the detection of antibodies in patient serum and are not accurate during the initial phase of the infection. Therefore, it is necessary to detect novel biomarkers that can be used in early detection of leptospirosis. Circulating miRNAs are known to be promising biomarkers for various diseases including cancer, tuberculosis, influenza; hence in this study the potential of miRNAs as biomarkers for leptospirosis was evaluated. A total of 30 leptospirosis cases were screened for the differential expression of 10 miRNA by RT-qPCR assay. The differential expression was calculated by relative quantification using healthy individuals as controls. Among the 10 miRNA,3 miRNA, miR-28–5p, miR-302c-3p and miR-302a-3p were reported to exhibit a significant trend of upregulation. Further their role in immune pathways and biological processes was investigated by KEGG analysis and Gene Ontology. The 3 miRNAs were observed to target various immune response pathways, thus confirming their role in host immune response. Based on the results obtained in this study, miR-28–5p, miR-302c-3p and miR-302a-3p can be considered as potential biomarkers for the detection of leptospirosis.

A Novel Electrochemiluminescence (ECL) Immunoassay for the Quantitation of Monoclonal Antibody (mAb) PYX-106 in Human Serum

Background:: PYX-106 is a novel monoclonal antibody (mAb), targeting the sialic acidbinding immunoglobulin-like lectin 15 (Siglec-15) in the Tumor Microenvironment (TME). Precise measurement of PYX-106 is essential for the thorough assessment of PYX-106 pharmacokinetics in clinical investigations. Methods:: A novel Electrochemiluminescence (ECL) immunoassay for the quantitation of PYX- 106 in human serum was developed and validated. Biotinylated anti-PYX-106 antibody Bio-A1A1 was employed as the capture antibody, and ruthenylated anti-PYX-106 antibody Ru-A3G10 was utilized as the detection antibody in the ECL immunoassay on Meso Scale Discovery (MSD) platform. Results:: This assay was fully validated in terms of selectivity, accuracy, precision, hook effect, stability, etc., with a dynamic range from 50.0 to 2,500 ng/mL in human serum under the 2018 U.S. Food and Drug Administration (FDA) guidance and the 2022 U.S. FDA ICH M10 guidance. Conclusion:: PYX-106 bioanalytical assay validation was reported for the first time in a biological matrix, and this assay has been successfully applied to support a clinical trial PYX-106-101.

Clinical Issue of Myasthenia Gravis Related to Immune Checkpoint Inhibitors.

Immune-related adverse events (irAEs) are pivotal in the management of immune checkpoint inhibitors (ICIs) across various human neoplasms. While common irAEs are manageable by oncologists, the detailed features of rare complications related to ICI therapy remain elusive. Among these, immune-related myasthenia gravis (irMG) stands out as a life-threatening disease. Research articles published in English between 2017 and 2023 were identified using the PubMed database. Forty-six relevant research studies were examined to collate information for this review. The incidence of ICI-induced MG was found to be less than 1.0%, with approximately 20-30% of irMG patients presenting with overlap syndrome involving myocarditis and myositis. The detection of acetylcholine receptor antibodies (AChR-Ab) and elevated creatinine kinase (CK) levels proved useful in identifying 50-70% and 60-80% of cases, respectively. However, the utility of muscle-specific kinase antibodies (MuSK-Ab) in detecting irMG was limited due to a low positivity rate (0-5.3%). Ptosis emerged as the most common initial symptom of irMG, with an approximate positivity rate of 80%. Recommended treatment for irMG involves high-dose steroids in conjunction with plasmapheresis or immunoglobulins to mitigate the increased mortality associated with irMG. Early initiation of immunosuppressive therapy is imperative to prevent the worsening of irMG. Furthermore, facilitating a fulfilling social life post-hospitalization is crucial. This review sheds light on the clinical aspects and management strategies pertaining to irMG.

Detection of Anti-Strongyloides Antibodies in the Serum of Liver Transplant Recipients: Need of Screening for This Neglected Helminthiasis.

Immunosuppressed patients, particularly transplant recipients, can develop severe strongyloidiasis. This study aimed to detect anti-Strongyloides IgG antibodies in a panel of sera from liver transplant patients. Two techniques were used: ELISA as the initial screening test and Western blotting as a confirmatory test. ELISA reactivity of 10.9% (32/294) was observed. The 40-30 kDa fraction was recognised in 93.7% (30/32) of the patients, resulting in a positivity rate of 10.2%. These data highlight the importance of serological screening for Strongyloides stercoralis infection in liver transplant recipients.

Pediatric Case of Autoimmune Cerebellitis with Positive Anti-GAD Antibodies and Accidental Finding of Roth Spots

Abstract We present a pediatric case of autoimmune cerebellitis, which was distinguished by the presence of positive anti-glutamic acid decarboxylase (anti-GAD) antibodies. The patient, a young child, presented with a four-day history of progressively worsening headaches, coupled with persistent vomiting. These symptoms prompted immediate medical evaluation. Initial computed tomography (CT) imaging revealed a hypodense area in the left cerebellum and mild hydrocephalus, indicating fluid accumulation in the brain. Subsequent magnetic resonance imaging (MRI) provided more detailed visualization, confirming the diagnosis of acute cerebellitis, which was characterized by bilateral enlargement of the cerebellum and an associated mass effect that exerted pressure on the adjacent brain structures. The medical team initiated a comprehensive treatment regimen that included antibiotics to address any potential bacterial infection, antivirals to counter possible viral etiologies, immunoglobulin therapy to modulate the immune response, and corticosteroids to reduce inflammation. The patient responded well to this treatment protocol, with follow-up imaging showing a significant reduction in cerebellar swelling and overall improvement in clinical symptoms. The detection of elevated anti-GAD antibodies, along with anti-thyroglobulin antibodies, indicated an autoimmune origin of the cerebellitis. This autoimmune response can lead to the body’s immune system mistakenly attacking healthy brain tissue. Moreover, the incidental finding of a Roth spot, a retinal hemorrhage with a white center, during a routine eye examination suggested potential systemic involvement, possibly indicating a broader immune-mediated condition.

Autoimmune Thyroiditis in Patients With Diabetes Mellitus: Retrospective Study of 91 Cases.

Background Detection and quantification of anti-thyroid antibodies make it possible to confirm the diagnosis of thyroid dysfunction as well as its autoimmune origin and monitor thyroid damage in diabetic patients. The aim of this study is to determinethe seroprevalence of anti-thyroid antibodies in hospitalized diabetic patients. Materials and methods This retrospective study focused on 91 diabetic patients hospitalized in the endocrinology department of Ibn Sina Hospital, Rabat, Morocco, between January 1 and December 31, 2022. The study population was divided into two groups: 19 patients with type 1 diabetes (13 females and six males, with an age range of 20-70 years) and 72 patients with type 2 diabetes (52 females and 20 males, with an age range of 40-71 years). Hemoglobin (HbA1c) levels were determined with high-performance liquid chromatography (Hb-HPLC) analyzer from blood samples collected in EDTA tubes, and anti-thyroid antibodies (anti-TPO and/or anti-TG) were measured by chemiluminescent microparticle immunoassays (CMIA) in human serum using the ALINITY analyzer. Results Among type 1 diabetic patients, 42.1% (n = 8)were positive for anti-TPO and anti-TG antibodies, while 31.5% (n = 6)were positive only for anti-TPO antibodies. Among type 2 diabetic patients, 15.2% (n = 11) were positive only for anti-TPO antibodies, while 20.8% (n = 15) were positive for anti-TPO and/or anti-TG antibodies. The prevalence of anti-thyroid antibodies was higher in females, consistent with other studies. This could be linked to the involvement of autoimmune processes in the development of thyroid dysfunction in type 2 diabetics. Conclusions Testing for anti-thyroid antibodies in diabetic patients and their relatives helps detect subclinical conditions, which could later manifest as biological and clinical deficiencies, guiding monitoring parameters.

Domestic Dog Infection with Trypanosoma cruzi from Northern and Southern Regions of Mexico.

Background: Chagas disease or American trypanosomiasis, caused by Trypanosoma cruzi and vectored by triatomines, affects millions of people worldwide. In endemic countries including Mexico, infections in domestic animals, such as dogs, may affect the risk of human disease when they serve as a source of infection to vectors that subsequently infect humans. Materials and Methods: We conducted a cross-sectional study of 296 dogs from two cities near the northern and southern borders of Mexico: Reynosa, Tamaulipas, and Tuxtla Gutierrez, Chiapas. Infection was measured based on testing of blood using T. cruzi quantitative PCR (qPCR) and up to three antibody detection assays. The StatPak immunochromatographic assay was used to screen samples and the indirect fluorescent antibody (IFA) and multiplex microsphere immunoassay (MIA) tests were used as secondary tests on all samples that screened positive and a subset of negatives. Serologic positivity was defined based on reactivity on at least two independent tests. Results: Of the 280 samples tested for parasite DNA, two (0.7%) were positive, one of which (0.4%) was confirmed as T. cruzi discrete typing unit TcIV. Overall, 72 (24.3%) samples were reactive for T. cruzi antibodies via StatPak of which 8 were also positive using MIA and 2 were also positive using IFA (including one of the PCR-positive dogs). Overall, nine dogs (3.4%) met study criteria of positivity based on either/both serology or PCR tests. Positive dogs were found in both regions of Mexico; five (2.7%) from Reynosa and four (3.6%) from Tuxtla Gutierrez. We found no association between infection status and state of origin, sex, age group, breed group, neighborhood, and whether other pets lived in the home. Conclusion: Our results re-emphasize dogs' utility as sentinels for T. cruzi in Mexico and underscore the need for improved veterinary diagnostic tests and parasite surveillance at the household level in endemic countries.

Anti-SARS-CoV-2 total immunoglobulin and neutralising antibody responses in healthy blood donors throughout the COVID-19 pandemic: a longitudinal observational study.

Quantifying antibodies against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and neutralising antibodies may help to understand protection at the individual and population levels. Determination of neutralising antibodies using classical virus neutralisation tests (VNT) is considered the gold standard, but they are costly and time-intensive. Enzyme-linked immunosorbent assay (ELISA)-based surrogate VNTs (sVNT) or anti-SARS-CoV-2 spike protein receptor binding domain immunoglobulins (anti-S-RBD Ig) may be suitable alternatives to VNTs. We aimed to (a) explore the correlations between anti-S-RBD Ig, VNT, and sVNT measurements and (b) describe humoral immunity against SARS-CoV-2 after vaccination, natural infection, and vaccine breakthrough infection in healthy blood donors. We measured total anti-SARS-CoV-2 Ig in 5714 serum samples from 2748 healthy individuals visiting the Swiss Red Cross Blood Donation Centre in Basel from 03/2020 to 04/2022. We used the Elecsys®Anti-SARS-CoV-2 immunoassay (Roche) against the N- and S-receptor binding domain (RBD) proteins. In a subset of 548 samples from 123 donors, we conducted sVNTs against the Wuhan wild-type SARS-CoV-2 (SARS-CoV-2 Neutralizing Antibodies Detection Kit; Adipogen™). In 100 samples from 40 donors, we correlated sVNT and VNTs against the wild-type (D614G WU1) virus. Surveys were sent to the blood donors to collect data on their SARS-CoV-2 infection and vaccination status. Using this data, donors were categorised as "vaccination only", "infection before vaccination", "post-vaccine breakthrough infection", and "natural infection only". Our longitudinal observation study cohort consisted of 50.7% males with a median age of 31 years (range 18-75 y). Anti-SARS-CoV-2 N protein positivity rates per month indicate 57.1% (88/154) of the cohort was infected up to 04/2022. No differences in seropositivity were found between sexes, age groups, blood types (AB0 or RhD), and cytomegalovirus serostatus. We observed a high correlation between anti-S-RBD Ig and inhibition percentage (Spearman'sρ= 0.92, Kendall'sτ= 0.77, p <0.0001). We determined the sensitivity and specificity for the manufacturers' thresholds for detecting virus-neutralising effects and computed the "best" cut-off based on our real-world data. We categorised 722/1138 (63.5%) donors as vaccination only (82.3%), post-vaccine breakthrough infection (7.8%), infection before vaccination (5.8%), and natural infection only (4.2%). We observed a lower inhibition percentage in the natural infection-only group than in all other vaccinated groups. The infection before vaccination group had higher anti-S-RBD Ig titres after the first vaccine dose than the other vaccinated groups. In total, 57.1% of healthy blood donors were infected with SARS-CoV-2, but natural infection without evidence of vaccination seems to result in substantially lower neutralising antibody levels. An estimate of antibody neutralisation may be helpful to assess reinfection risk. Total anti-S-RBD Ig correlates with surrogate virus neutralisation test results, a surrogate for neutralisation; therefore, we suggest that total anti-S-RBD Ig may estimate the level of neutralising antibodies. The threshold for protection from an unfavourable clinical outcome must be evaluated in prospective clinical cohorts.

A novel syphilis Treponema pallidum lipoprotein peptide antigen diagnostic assay using red cell kodecytes in routine blood centre column agglutination testing platforms.

The detection of treponemal antibodies, which are used to make a diagnosis of syphilis, is important both for diagnostic purposes and as a mandatory blood donor test in most countries. We evaluated the feasibility of using Kode Technology to make syphilis peptide red cell kodecytes for use in column agglutination serologic platforms. Candidate Kode Technology function-spacer-lipid (FSL) constructs were made for the Treponema pallidum lipoprotein (TmpA) of T. pallidum, using the peptide and FSL selection algorithms, and then used to make kodecytes. Developmental kodecytes were evaluated against a large range of syphilis antibody reactive and non-reactive samples in column agglutination platforms and compared against established methodologies. Overall, 150 reactive and 2072 non-reactive Syphicheck assay (a modified T. pallidum particle agglutination) blood donor samples were used to evaluate the agreement rate of the developed kodecyte assay. From three FSL-peptide candidate constructs, one was found to be the most suitable for diagnostics. Of 150 Syphicheck assay reactive samples, 146 were TmpA-kodecyte reactive (97.3% agreement), compared with 58.0% with the rapid plasmin reagin (RPR) assay for the same samples. Against the 2072 expected syphilis non-reactive samples the agreement rate for TmpA-kodecytes was 98.8%. TmpA-kodecytes are viable for use as cost-effective serologic reagent red cells for the detection of treponemal antibodies to diagnose syphilis with a high level of specificity in blood centres. This kodecyte methodology also potentially allows for introduction of the reverse-algorithm testing into low-volume laboratories, by utilizing existing transfusion laboratory infrastructure.

SARS-CoV-2 detection in COVID-19 patients' sample using Wooden quoit conformation structural aptamer (WQCSA)-Based electronic bio-sensing system

The COVID-19 epidemic and its continuous spread pose a serious threat to public health. Coronavirus strains known as SARS-CoV-2 (Severe acute respiratory syndrome coronavirus 2) variants have undergone genomic changes. The severity of the symptoms, the efficiency of vaccinations, and the transmission capacity of the virus can be impacted by these alterations. Point-of-care diagnostic assays can identify particular genetic or protein sequences that are exclusive to each variety. Currently, ultrafast, responsive, and accurate antibody detection faces several challenges. Here, we outline the fabrication, implementation, and sensing performance benchmarking of an ultrafast (5 s) and inexpensive (0.15 USD) assay with label-free sensing of SARS-CoV-2 S (Spike)/N (Nucleocapsid) protein and other variants in real patient samples. A label-free DNA aptameric capacitive bio-sensing device was used to detect SARS-CoV-2 variants. Our novel, cutting-edge bio-sensing device contains a Wooden quoits conformation structural aptamer (WQCSA)-based inter-digitated capacitor electronic (WQCSA-IDCE) system. WQCSA-aptamer was used as a switch-turn on response to achieve ultrasensitivity in the variable area of the SARS-CoV-2. The molecular beacon (MB) method was also used to measure the fluorescently colored SARS-CoV-2 S/N protein. These sensors can be used with several types of label-free DNA aptamers to act as rapid, affordable, and label-free biosensors for a variety of critical acute respiratory virus syndrome disorders.

Investigation of the Seroprevalence of Brucella Antibodies and Characterization of Field Strains in Immunized Dairy Cows by B. abortus A19.

(1) Background: One method of eradicating brucellosis is to cull cattle that test positive for antibodies 12 months after being vaccinated with the 19-strain vaccine. Variations in immunization regimens and feeding practices may contribute to differences in the rate of persistent antibodies. We conducted this study to investigate the real positive rate of Brucella antibody in field strains of Brucella spp. after immunization over 12 months in dairy cows. This research aims to provide data to support the development of strategies for preventing, controlling, and eradicating brucellosis. (2) Method: We employed the baseline sampling method to collect samples from cows immunized with the A19 vaccine for over 12 months in Lingwu City from 2021 to 2023. Serological detection was conducted using the RBPT method. An established PCR method that could distinguish between 19 and non-19 strains of Brucella was utilized to investigate the field strains of Brucella on 10 dairy farms based on six samples mixed into one using the Mathematical Expectation strategy. (3) Results: We analyzed the rates of individual seropositivity and herd seropositive rates in dairy cattle in Lingwu City from 2021 to 2023 and revealed that antibodies induced by the Brucella abortus strain A19 vaccine persist in dairy herds for more than 12 months. We established a PCR method for identifying both Brucella A19 and non-A19 strains, resulting in the detection of 10 field strains of Brucella abortus from 1537 dairy cows. By employing a Mathematical Expectation strategy, we completed testing of 1537 samples after conducting only 306 tests, thereby reducing the workload by 80.1%. (4) Conclusions: There was a certain proportion of cows with a persistent antibody titer, but there was no evidence that all of these cattle were naturally infected with Brucella. The established PCR method for distinguishing between Brucella abortus strain 19 and non-19 strains can be specifically utilized for detecting natural Brucella infection in immunized cattle. We propose that relying solely on the detection of antibodies in cattle immunized with the A19 vaccine more than 12 months previously should not be solely relied upon as a diagnostic basis for brucellosis, and it is essential to complement this approach with PCR analysis to specifically identify field Brucella spp. Brucella abortus was the predominant strain identified in the field during this study. Detection based on the Mathematical Expectation strategy can significantly enhance detection efficiency.

The epizootic situation of bovine leukemia in the Republic of Dagestan in 2023

Bovine enzootic leukemia (EBLV) remains an acute problem for animal husbandry in many regions of Russia. Based on this, the goal was set: to study the epizootic situation of bovine leukemia in the Republic of Dagestan. The material for the study was taken from the official source of the Veterinary Committee of the Republic, as well as supplemented with our own serological blood tests for bovine leukemia virus (bovine leukemia). Laboratory tests for the detection of antibodies to BLV in RID were carried out according to the "Guidelines for the diagnosis of bovine leukemia" (2000). Research results: the spread of BLV has been established in some municipal districts and urban districts throughout the republic. In 2023, from all veterinary laboratories (stations), serological studies were conducted in the amount of 1101,689 animal blood samples for BLV. Of the animals studied, antibodies to the BLV antigen in RID were obtained in 2,778 (0.25%) samples. A high percentage of animal HCV infection was detected in the following districts and cities: Kizlyarsky – 2% (388), Tlaratinsky – 1.3% (341), Botlikhsky – 1.2% (450), Babayurt – 1.1% (188), Gunibsky – 0.8% (228) and in Makhachkala – 0.9% (180). In 16 districts and in Khasavyurt, the percentage of infection of animals with BLV was at the level of 0.3% or lower. We obtained 440 animal blood samples from sample farms, which were examined in the RID. 37 (8.4%) animal blood samples gave a positive result for BLV. Thus, the general epizootic situation in EBLV remains relevant, but the prevalence of BLV among animals is decreasing in the republic.

Prevalence of Hepatitis D in People Living with HIV: A National Cross-Sectional Pilot Study.

This study assesses the prevalence of hepatitis D virus (HDV) in people living with HIV (PLWHIV) in Greece. Given the compounding effects of HDV and hepatitis B (HBV) on liver disease progression, as well as the emergence of new therapeutic options such as bulevirtide, understanding regional disparities and the epidemiological impact of such co-infections is vital. A cross-sectional analysis was conducted utilizing 696 serum samples from PLWHIV attending five major university hospitals. The methodology included HDV antibody detection by ELISA and HDV RNA confirmation. Of the 30 HBsAg-positive samples analyzed, the study population was primarily male (93%), with a median age of 54 years. Participants had been on antiretroviral therapy for a median of 10 years, and the median CD4 count was 738 (539-1006) copies/mL. Additional serological findings revealed a 7% prevalence of hepatitis C virus (HCV) IgG antibodies and a 55% prevalence of hepatitis A virus (HAV) IgG antibodies. Seroreactivity for syphilis (RPR/VDRL/TPHA positive) was identified in 33% of the participants. The results indicated a low HDV prevalence, with only one individual (3%) testing positive for anti-HDV IgG antibodies and none for HDV RNA. This indicates a lower prevalence of HDV among PLWHIV with chronic HBV in Greece compared to global data.