Determination of cytomegalovirus IgG antibodies using stable element labeling and inductively coupled plasma mass spectrometry: Further evidence of the feasibility of antibody detection.

Accuracy of rapid diagnostic tests for Strongyloides stercoralis detection: a systematic review and meta-analysis.

Inhalable DNA nano-adjuvant elicits robust lung-resident memory immunity against pneumonic plague.

Recombinant PsoSP4 confers superior 80% protection against psoroptic mange in rabbits in association with a Th1-biased immune response.

Advances in genome editing technologies in Komagataella phaffii and their applications in biomanufacturing

Multi-antigen transmission-blocking malaria vaccine elicits Th1-biased antibody responses and SMFA-confirmed oocyst reduction in Anopheles stephensi.

Recombinant production of human papillomavirus type 16 E6 and E7 vaccine antigens in Chlamydomonas reinhardtii.

Subunit vaccines rely on adjuvants to enhance immunogenicity, yet antigen-adjuvant interactions in liposomal systems remain underexplored, with unknown implications that could impact the function and stability of coformulated system designs. We report a multimodal study aimed at elucidating interactions between a recombinant protein antigen, gE, and a liposomal adjuvant system, AS01B, using a commercially available vaccine as a model. Quantitative binding assays revealed no significant binding for the antigen-adjuvant pair, with only weak, dose-dependent biolayer interferometry (BLI) signals suggesting transient contacts, in contrast to the strong responses observed in positive controls. Isothermal titration calorimetry (ITC) provided clear evidence of weak, transient interactions through dose-dependent exothermic signals and characteristic saturation behavior. Ultracentrifugation, electrophoresis, and chromatographic separation techniques confirmed independent distributions for the protein and liposome components, with minor migration shifts and trace coelution under rapid flow hinting at dynamic interactions. Structural analyses by cryogenic electron microscopy (cryo-EM) and dye labeling showed negligible changes in liposome properties upon antigen mixing. These findings suggest that vaccine formulations with little or no interaction between the protein and liposome components may be stable as premixed systems and underscore the importance of employing multiple tools for characterizing multicomponent vaccines during their development.

Diagnosis of chronic Trypanosoma cruzi infection relies on the concordant results of two serological tests based on different antigens and/or principles. Currently, the only accepted cure criterion is the switch from positive to negative serology, but anti-T. cruzi IgG may persist for years, even after treatment. Recently, a novel chemiluminescence immunoassay based on native T. cruzi excreted-secreted antigens (TESA)-Chagas TESA VirClia-was developed. This study assessed the performance of this assay for the detection of anti-T. cruzi antibodies for the diagnosis and post-treatment follow-up of chronic T. cruzi infection. A retrospective diagnostic accuracy study was conducted using 227 serum samples, including samples from individuals tested for chronic T. cruzi infection and from individuals with other parasitic or viral infections. T. cruzi infection status was established using two reference serological assays based on recombinant and native antigens. All samples were analyzed by TESA-based assay and compared with the reference tests. Concordance between the TESA-based assay and the reference tests was 93.8%, with a sensitivity and specificity of 92% and 100%, respectively. Limited cross-reactivity was observed in samples positive for Leishmania spp. antibodies, while no cross-reactivity was observed in viral infections. A decrease in antibody index over time was observed more frequently in treated patients when tested by the TESA-based assay. These findings support the use of the Chagas TESA VirClia assay as a reliable confirmatory tool within the diagnostic algorithm for chronic T. cruzi infection. Moreover, this assay was the only test showing a consistent post-treatment decline in antibody index over time.IMPORTANCEThe diagnosis and monitoring of chronic Trypanosoma cruzi infection remain challenging because no single serological test achieves 100% sensitivity and specificity, and the antibodies may persist for many years even after treatment. This study highlights the value of a novel serological assay based on native antigens excreted by the parasite, which more effectively reflects changes in antibody levels over time. Its high diagnostic accuracy and greater ability to detect declining antibody values in treated patients support its use as a reliable diagnostic tool as well as a promising method for serological follow-up.

Background/Objectives: Canine papillomavirus (CPV) is an important viral pathogen associated with papillomatosis in dogs, with canine papillomavirus type 1 (CPV1) and type 2 (CPV2) among the most prevalent and clinically relevant genotypes. The L1 capsid protein is a major immunogenic antigen of papillomaviruses; however, conserved linear B-cell epitopes shared between CPV genotypes remain poorly defined. This study aimed to identify conserved cross-reactive B-cell epitopes within CPV1 and CPV2 L1 proteins and to evaluate their preliminary immunoreactivity. Methods: Conserved linear B-cell epitopes were predicted through integrated bioinformatic and structural analyses based on sequence conservation and surface accessibility. Three candidate epitopes were selected. Recombinant CPV1 and CPV2 L1 proteins were expressed in Escherichia coli (E. coli), purified, used as recombinant L1 antigens, together with BSA-conjugated synthetic epitope peptides for mouse immunization. Antigen-specific IgG responses were assessed by ELISA, antigen-associated IFN-γ responses were evaluated by ELISpot, and cross-reactive antibody recognition was assessed by Western blot. Results: Recombinant L1 proteins induced strong antigen-specific IgG responses in mice. The selected peptides induced detectable but weaker humoral responses compared with the recombinant L1 proteins. Among the three epitopes, TPSGSLV and TVVDNTR elicited antibodies that recognized both CPV1 and CPV2 L1 proteins, while the epitope VIVPKVS showed minimal or no detectable immunoreactivity. ELISpot analysis showed only modest antigen-associated IFN-γ responses, particularly in peptide-immunized groups. Conclusions: This study identified conserved cross-reactive linear B-cell epitope candidates within CPV1 and CPV2 L1 proteins and provided preliminary immunological evidence supporting their potential relevance for CPV antigen design. However, peptide-induced responses were weaker than those induced by recombinant L1 proteins, and VLP formation, antibody neutralizing activity, and protective efficacy were not evaluated. Further studies in dogs, including optimized antigen-display platforms, neutralization assays, and protection studies, are required to determine the practical value of these epitopes for CPV vaccine development.

COVID-19 vaccination with updated compositions remains important as SARS-CoV-2 continues to circulate, cause disease, and evolve. Available COVID-19 vaccines in the 2024-2025 season differed by platform, including mRNA-1273, an mRNA-based vaccine, and NVX-CoV2705, a recombinant protein-based vaccine and antigen composition (KP.2-targeted and JN.1-targeted, respectively). Limited real-world evidence exists comparing effectiveness in preventing severe COVID-19 outcomes. We compared mRNA-1273 with protein-based NVX-CoV2705 in insured US adults vaccinated during the 2024-2025 season. We conducted a retrospective matched cohort study in a large US claims database. Adults aged 18years or older who received mRNA-1273 ("exposed") or NVX-CoV2705 ("reference") between August 31, 2024 and February 28, 2025 were eligible. Recipients were matched 2:1 on key demographic and clinical factors and then weighted with stabilized inverse probability of treatment weights. Outcomes were medically attended COVID-19 and hospitalization with COVID-19 from day7 after vaccination through up to 180days of follow-up. We calculated comparative vaccine effectiveness (cVE) as 100 × (1 - hazard ratio). Of 1,156,441 mRNA-1273 recipients and 45,384 NVX-CoV2705 recipients, 69,140 and 34,570, respectively, entered the matched cohort. Median (Q1, Q3) follow-up was 180 (163, 180) days for mRNA-1273 and 180 (162, 180) for NVX-CoV2705. Medically attended COVID-19 occurred in 706 (1.02%) mRNA-1273 recipients and 512 (1.48%) NVX-CoV2705 recipients; adjusted cVE (95% confidence interval [CI]) was 31.7% (23.4%, 39.1%). Hospitalization with COVID-19 occurred in 61 (0.09%) and 49 (0.14%) recipients, respectively; adjusted cVE (95%CI) was 40.7% (13.5%, 59.4%). In the 47,754 mRNA-1273 recipients matched to 23,877 NVX-CoV2705 recipients aged ≥ 65, adjusted cVE (95%CI) was 25.7% (15.4%, 34.8%) against medically attended COVID-19 and 41.7% (14.3%, 60.4%) against hospitalization with COVID-19. In this insured US adult population, mRNA-1273 demonstrated greater effectiveness against medically attended COVID-19 and hospitalization with COVID-19 than the protein-based NVX-CoV2705. These findings highlight the potential public-health importance of considering vaccine platform and variant selection when planning for upcoming seasons.

Brucellosis is a globally prevalent zoonotic disease that imposes considerable economic burdens on the livestock industry and remains a significant threat to public health. Although lipopolysaccharide (LPS)-based serological assays are widely used in routine diagnosis, their inherent limitations-particularly cross-reactivity with other Gram-negative bacteria-underscore the need for the development of diagnostic approaches based on non-LPS antigens. In this study, we developed a trivalent nanobody-horseradish peroxidase (3Nbs-HRP) fusion protein targeting Brucella OMP16 and established a cELISA for the serological detection of bovine brucellosis. The diagnostic performance of the assay was assessed using 204 Brucella antibody-negative and 123 Brucella antibody-positive bovine serum samples. ROC curve analysis yielded a sensitivity of 87.7% and a specificity of 89.4%, with no significant cross-reactivity observed. By employing a recombinant antigen and a 3Nbs-HRP probe, this assay enhances biosafety and demonstrates strong potential for standardization and large-scale application, serving as a complementary tool to conventional LPS-based assays for the surveillance, diagnosis, and control of bovine brucellosis.

Toxoplasmosis, a globally prevalent zoonosis caused by Toxoplasma gondii (T. gondii), poses major threats to both human and animal health, leading to reproductive losses in livestock and severe disease in immunocompromised individuals. Although enzyme‐linked immunosorbent assay (ELISA) and PCR are widely used for diagnosis and surveillance, they may be limited by turnaround time, laboratory instrumentation, and, in the case of serological assays, the need for species‐specific reagents. To enable rapid, equipment‐minimal detection applicable to multiple host species, we developed a point‐of‐care (POC) double‐antigen sandwich colloidal gold immunochromatographic assay (GICA) based on recombinant surface antigen 2 (rSAG2) of T. gondii. In this assay, rSAG2 served both as the capture antigen immobilized on the test line and as the colloidal gold‐conjugated detection probe. Key parameters, including conjugation pH, antigen loading, and buffer composition, were systematically optimized. The resulting strip showed a detection limit of a 1:40 serum dilution, and no cross‐reactivity was observed with sera positive for 23 common pathogens from different host species. Good repeatability and storage stability for 4 months at 4°C were also observed. For field evaluation, 409 clinical serum samples from six animal groups (100 chickens, 68 dogs, 30 cats, 81 pigs, 80 yaks, and 50 sheep) were tested, and seropositivity rates ranged from 2.9% to 31.3% across the sampled groups. In a subset of chicken (n = 46) and dog (n = 44) sera tested in parallel with the corresponding commercial ELISA kits, the rSAG2‐GICA showed good preliminary agreement, with overall agreement rates of 91.3% and 97.7%, respectively. Collectively, this rSAG2‐based GICA shows potential as a rapid and practical tool for on‐site serological screening of T. gondii antibodies across multiple host species and may support epidemiological surveillance in diverse animal populations.

Canine seroepidemiology of Trypanosoma cruzi in socially vulnerable urban communities of Salvador, Brazil.

Immunocastration, a nonsurgical strategy based on active immunization against gonadotropin-releasing hormone (GnRH), effectively suppresses steroidogenesis and spermatogenesis. However, peptide vaccines targeting poorly immunogenic antigens such as GnRH often fail to elicit robust adaptive immune responses, requiring adjuvants or carrier proteins. Previously, we introduced Coated Bacterial Vaccines (CBVs), a platform that uses chemically inactivated Gram-positive bacteria to display recombinant antigens fused to the SlpA carboxy-terminal domain (dSLPA) on their surface. This system leverages natural pathogen-associated molecular patterns (PAMPs) to enhance immunogenicity without additional adjuvants. In this work, we extended the application of the CBVs platform to enhance the immune response against a poorly immunogenic GnRH-based peptide vaccine. GnRH-CBVs were formulated using inactivated Bacillus subtilis var. natto coated with a recombinant GnRH tandem-repeat-dSLPA fusion protein and administered to male BALB/c mice. A chitosan-adjuvanted GnRH-dSLPA formulation served as a positive control. Immunization with GnRH-CBVs could favor a strong Th2-biased humoral response, characterized by predominant IgG1 levels comparable to those achieved with chitosan. The resulting antibodies effectively neutralized endogenous GnRH, reducing steroidogenesis and spermatogenesis and inducing marked testicular histological alterations. These findings support CBVs as a promising strategy to enhance peptide vaccine immunogenicity for veterinary immunocastration.

Bongkrekic acid (BA) is a highly toxic, heat-stable bacterial toxin associated with severe foodborne outbreaks, particularly in Southeast Asia, with reported mortality rates of up to 100%. Despite its clinical relevance, current detection methods rely on complex, laboratory-based analytical techniques, thereby limiting the ability to reach a rapid diagnosis. Here, we present a novel strategy for BA detection that employs a recombinant antigen mimic generated by fusing a phage-display-selected BA epitope to maltose-binding protein (MBP), enabling safe, reproducible, and scalable immunoassay development without handling the native toxin. The BA-MBP fusion protein was used as a coating antigen in a competitive immunoassay (ic-ELISA), demonstrating sensitive antibody recognition with an assay IC50 of 6.24 ng/mL. This approach was further translated into a nanophotonic label-free biosensing platform to demonstrate the potential for rapid diagnostics in human samples. A single-step competitive biosensor assay provided an IC50 of 0.64 ng/mL and a limit of detection of 0.12 ng/mL, representing a 10-fold improvement over the ic-ELISA using conventional conjugates and providing a performance comparable to LC-MS/MS. The method was successfully validated in spiked blind urine samples, highlighting its applicability in clinically relevant matrices. The combination of BA-MBP fusion conjugate with photonic biosensing establishes a robust, safe, and scalable approach for rapid, reliable BA detection and quantification, with strong potential for point-of-care diagnostics and decentralized toxin monitoring.

Background: Modified Vaccinia Ankara (MVA) vectors are highly immunogenic vaccine platforms for the delivery of recombinant antigens. Efficient downstream processing is still challenging, particularly because substantial fractions of the virus remain intracellular. While chemical cell lysis that releases MVA particles into the supernatant before clarification can greatly enhance process efficiency and scalability, this step remains insufficiently characterized. Methods: This study assessed the compatibility of ionic, non-ionic, and zwitterionic detergents with the virus as purification target. Polysorbate 20 (Tween 20) was selected as a candidate detergent and evaluated across harvest times of 48-72 h post-infection (hpi) at concentrations of 0.01-0.5% (v/v). Results: The addition of 0.01% to 0.05% Tween 20 at 48 hpi resulted in a twofold increase in supernatant virus within one hour of application. Extended exposure to Tween 20, combined with a 650 mM mixture of NaCl, NaBr, and KCl, promoted virus particle release. However, Tween 20 concentrations above 0.1% reduced MVA infectivity. A filtration cascade using pore sizes of 5 µm and 1.2 µm achieved product yields of 77-83% at 48 hpi and 41-69% at 72 hpi, respectively. Host-cell DNA is an important contaminant during viral vector processing. However, the application of 0.05% (v/v) Tween 20 resulted in a 35% reduction of dsDNA released into the culture supernatant; the nuclei could not be preserved intact under high-salt conditions to avoid the release of cellular DNA. Conclusions: In summary, this comprehensive data demonstrated that non-ionic detergents can be used to induce cell lysis while maintaining infectious activity of enveloped MVA.

Our previous research identified a luciferase immunosorbent assay (LISA) for detecting serum antibodies against Tp17 and Tp47 as a potential tool for diagnosing neurosyphilis (NS). In this study, we developed a serological test based on a multi-antigen fusion protein (Tp15-17-47) to simplify the detection process and improve the diagnostic accuracy for NS. The Tp15, Tp17, and Tp47 genes were amplified separately by polymerase chain reaction (PCR) and assembled into a fusion gene coding for a tripartite fusion protein antigen using overlap PCR. This recombinant Tp15-17-47 antigen was used retrospectively to detect serum anti-Tp15-17-47 IgG antibodies in 434 HIV-negative patients suspected of NS from cohorts in Beijing and Guangzhou. Two diagnostic prediction models were developed using stepwise logistic regression. Serum anti-Tp15-17-47 IgG antibodies demonstrated moderate diagnostic capability for NS in the development cohort, with an area under the curve (AUC) of 0.791 (95% CI: 0.735-0.846). An optimized NS prediction model substituting anti-Tp15-17-47 antibodies for TRUST/RPR outperformed the base NS diagnostic model. The optimized model achieved an AUC of 0.877 (95% CI: 0.835-0.919) compared to 0.843 (95% CI: 0.795-0.890) for the base model in the development cohort (p = 0.035). This superior performance was confirmed in the validation cohort (AUC: 0.880, 95% CI: 0.827-0.933 vs. 0.792, 95% CI: 0.726-0.859; p < 0.001). Decision curve analysis (DCA) revealed that the optimized model provided greater net benefit than the base model across threshold probabilities ranging from 0.10 to 0.95 in both cohorts. Quantitative profiling of serum IgG targeting the recombinant treponemal antigen Tp15-17-47, along with its diagnostic model, exhibited robust discriminatory power for neurosyphilis diagnosis, which supports the viability of replacing lumbar puncture with this less invasive serological strategy for neurosyphilis ascertainment.

Toxoplasmosis is a globally prevalent zoonosis caused by Toxoplasma gondii (T. gondii), a parasite that infects nearly all warm-blooded animals, including humans, cats, and pigs. Beyond incurring substantial economic losses to the swine industry, T. gondii infection poses a severe threat to public health, as pigs, which are key intermediate hosts for T. gondii transmission, serve as a major source of human infection via the food chain. Thus, developing specific, sensitive, rapid, and easy-to-perform detection methods for porcine T. gondii is of paramount importance for the prevention, control, and eventual eradication of this pathogen in swine populations. To address this need, a novel colloidal gold immunochromatographic strip (CGIS) was developed for the rapid serological detection of T. gondii infection in swine. The strip was constructed using recombinant surface antigen 1 (SAG1) expressed in a baculovirus-insect cell system, which ensures that the protein retains native antigenic epitopes. The purified SAG1 protein was conjugated to colloidal gold nanoparticles and loaded onto a conjugate pad, while staphylococcal protein A and a polyclonal anti-SAG1 antibody were immobilized on a nitrocellulose membrane to form the test line (T line) and control line (C line), respectively. Clinical validation demonstrated that the developed CGIS exhibits high specificity, with no cross-reactivity to sera positive for 13 common porcine pathogens. The strip achieved a visual limit of detection of 1:25,600 dilution of positive serum. In parallel, a commercial enzyme-linked immunosorbent assay kit yielded positive results up to a 1:800 dilution according to the manufacturer's diagnostic cutoff. Importantly, evaluation of 300 clinical porcine serum samples demonstrated a 98.3% concordance rate with the reference microscopic agglutination test. This SAG1-based CGIS provides a practical, user-friendly tool for large-scale surveillance and on-farm screening of porcine toxoplasmosis, thereby offering potential support for the prevention and control of T. gondii infection in pigs.

Human cytomegalovirus (HCMV) is a common pathogen that poses significant health risks, particularly to immunocompromised individuals. This study aims to improve diagnostic accuracy by developing a novel enzyme-linked immunosorbent assay (ELISA) and colloidal gold immunochromatography assay (GICA) utilizing recombinant HCMV proteins (fusion of pp150 and pp28), which offer improved sensitivity and specificity. A recombinant HCMV antigen was synthesized for the detection of CMV IgG antibodies in 573 serum samples collected from Hangzhou, Zhejiang Province. All samples were analyzed using two in-house developed assays (indirect ELISA and GICA), and 528 of these samples were identified as CMV IgG-positive by a commercial ELISA kit. The diagnostic performance and reliability of the two in-house assays were subsequently compared with those of this commercial ELISA kit. The assays showed no cross-reactivity with other viruses, with a coefficient of variation of less than 10%, indicating excellent reproducibility. Compared to the commercial ELISA kit, the concordance rates of the recombinant antigen-based ELISA and GICA were 98.9% and 99.7%, respectively. The ELISA and GICA developed in this study demonstrate high sensitivity and specificity, making them suitable for screening CMV IgG antibodies particularly in immunocompromised individuals. These findings highlight the potential of recombinant antigen-based assays to advance in infectious disease diagnostics. Not applicable.