RESUMEN
Outbreaks of emerging diseases, like Mpox in 2022, pose unprecedented challenges to global healthcare systems. Although Mpox cases globally decreased since the end of 2022, numbers are still significant in the African Region, European Region, Region of the Americas, and Western Pacific Region. Rapid and efficient detection of infected individuals by precise screening assays is crucial for successful containment. In these assays, analytical and clinical performance must be assessed to ensure high quality. However, clinical studies evaluating Mpox virus (MPXV) detection kits using patient-derived samples are scarce. This study evaluated the analytical and clinical performance of a new diagnostic MPXV real-time PCR detection kit (Sansure Monkeypox Virus Nucleic Acid Diagnostic Kit) using patient-derived samples collected in Germany during the MPXV clade IIb outbreak in 2022. Our experimental approach determined the Limit of Detection (LoD) to less than 200 cp/mL using whole blood samples and samples derived from vesicles or pustules. Furthermore, we tested potentially inhibiting substances and pathogens with homologous nucleic acid sequences or similar clinical presentation and detected no cross-reactivity or interference. Following this, the assay was compared to a CE-marked test in a clinical performance study and achieved a diagnostic sensitivity of 100.00% and diagnostic specificity of 96.97%. In summary, the investigated real-time PCR assay demonstrates high analytical performance and concurs with the competitor device with high specificity and sensitivity.
Asunto(s)
Reacción en Cadena en Tiempo Real de la Polimerasa , Sensibilidad y Especificidad , Humanos , Reacción en Cadena en Tiempo Real de la Polimerasa/métodos , Alemania/epidemiología , Mpox/diagnóstico , Mpox/virología , Juego de Reactivos para Diagnóstico , Técnicas de Diagnóstico Molecular/métodos , Límite de Detección , Brotes de Enfermedades , Parapoxvirus/aislamiento & purificación , Parapoxvirus/genéticaRESUMEN
IMPORTANCE: Enteric bacterial infections caused by Salmonella, Shigella, pathogenic Escherichia coli, and Campylobacter represent one of the most common causes of infectious enteritis worldwide. The timely and accurate diagnosis of pathogens causing gastroenteritis is crucial for patient care, public health, and disease surveillance. While stool culture has long been the standard and highly specific method for detecting enteric pathogens, it is labor-intensive and time-consuming with limited sensitivity. To improve patient outcomes, there is a need to implement new cost-effective approaches for the detection of bacterial enteric pathogens with higher sensitivity and faster time to result. This study shows that multiplex real-time polymerase chain reaction-based tests, such as the TAQPATH Enteric Bacterial Select Panel, are accurate and cost-effective diagnostic alternatives for the detection and differentiation of the most common enteric bacterial pathogens, offering quicker time to result and higher sensitivity compared to routine stool culture.
Asunto(s)
Infecciones por Enterobacteriaceae , Enterobacteriaceae , Humanos , Enterobacteriaceae/genética , Salmonella , Infecciones por Enterobacteriaceae/diagnóstico , Infecciones por Enterobacteriaceae/microbiología , Bacterias/genética , Escherichia coli , Heces/microbiología , Pruebas Diagnósticas de Rutina , Diarrea/microbiologíaRESUMEN
BACKGROUND: The cobas® omni Utility Channel enables users to integrate lab-developed tests (LDTs) on the cobas® 6800 System to perform molecular diagnostics with high-throughput capacity and full automation. At present, there are no CE- or FDA-approved tests for stool pathogens on this system. To assess the performance of stool as a matrix, we evaluated the analytical and clinical performance of an LDT for detection of Clostridioides difficile (C. difficile) toxin B using the Utility Channel (C.diff_UTC). METHODS: A 10% stool suspension prepared from liquid stool samples diluted in phosphate buffered saline was used for analysis. Limit of detection (LoD) was determined in six dilutions with 126 replicates/dilution. Clinical evaluation was performed using 514 predetermined patient stool samples from two study sites in Germany. The C.diff_UTC was compared with LC 480 amplification and an LDT or the R-BioPharm C. difficile assay. Discrepant results were further analyzed using the GeneXpert C. difficile assay. RESULTS: Limit of detection was 23.48 cfu/mL (95% Confidence Interval [CI]: 19.14-31.01) with inter-run variation of <2 cycle thresholds at 3 × and 10 × LoD. No cross-reactivity was observed with a panel of fecal organisms and pathogens. Bioinformatic analysis showed coverage of the major C. difficile toxinotypes by the primer/probe set. Clinical evaluation revealed sensitivity of 96.7% (95% CI: 88.7-99.6) and specificity of 99.3% (95% CI: 98.0-99.9) compared with the reference method; inhibition rate was 3.5% (18/514). CONCLUSION: Using a predesigned primer/probe set, the C.diff_UTC assay features analytical performance and clinical sensitivity and specificity comparable to currently available nucleic acid amplification tests (NAATs) and is suitable for high-throughput testing. This was a proof-of-concept study, indicating the cobas Utility Channel could likely be adapted for other clinically relevant stool pathogens in outbreak scenarios.
Asunto(s)
Proteínas Bacterianas/aislamiento & purificación , Toxinas Bacterianas/aislamiento & purificación , Clostridioides difficile/genética , Clostridioides difficile/aislamiento & purificación , Infecciones por Clostridium/diagnóstico , Heces/química , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa/métodos , Proteínas Bacterianas/genética , Toxinas Bacterianas/genética , Infecciones por Clostridium/microbiología , Alemania , Humanos , Técnicas de Amplificación de Ácido Nucleico , Sensibilidad y EspecificidadRESUMEN
BACKGROUND: Lower respiratory tract infections are a major threat to public health systems worldwide, with RSV and influenza being the main agents causing hospitalization. In outbreak situations, high-volume respiratory testing is needed. In this study, we evaluated the analytical and clinical performance of a pre-designed primer/probe set for the simultaneous multiplex detection of both viruses on a high-throughput platform, the cobas® 6800, using the "open channel" of the system for integration of lab-developed assays for the detection of influenza and RSV. RESULTS: Using the influenza/RSV qPCR Assay with swabs, LoD (95%) in TCID50/mL for influenza-A was 0.009, influenza-B 0.003, RSV-A 0.202, and RSV-B 0.009. Inter-run variability (3xLoD) was low (<1 Ct for all targets). Of 371 clinical respiratory specimens analyzed, results were concordant for 358 samples. The calculated sensitivity and specificity of the assay were 98.3% and 98.4% for Flu-A, 100% and 98.5% for Flu-B, and 98.6% and 99.7% for RSV. All quality assessment panel specimens (N = 63, including avian influenza strains) were correctly identified. None of the tested microorganisms showed cross-reactivity. CONCLUSION: Compared with CE-IVD assays, the assay evaluated here showed good analytical and clinical sensitivity and specificity with broad coverage of different virus strains. It offers high-throughput capacity with low hands-on time, facilitating the laboratory management of large respiratory outbreaks.