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Figure 1. Measles assay development and validation. (A) Alignment of the M gene of measles virus genotypes D8, B3, and <t>Edmonston</t> vaccine (VA) strains. Assays for distinguishing between wild type (WT) strains (D8 and B3) and vaccine strains used the same forward and reverse primers and different probe sequences, shown above the aligned sequences. (B) Validation of the WT2 and vaccine assays using ATCC standards spiked into wastewater. ddPCR results for the WT2 probe (channel 2, top panel) showed a cluster of positive droplets between 5,000 and 6,000 amplitude, corresponding to presence of the WT standard (genotype B3), and a cluster with a 3,000 amplitude corresponding to the vaccine standard. The VA probe (channel 3, bottom panel) showed a positive cluster at 7,000 amplitude corresponding to the VA standard and a positive cluster at 4,000 corresponding to the WT standard. The quantitative results (concentrations and positive droplet counts) are shown in Table SI.12. (C) Validation of the WT2 and vaccine assays using wastewater samples from a facility in Chicago, IL, with an active measles outbreak. All samples showed positive detections of WT strain, as indicated by the amplitude of the positive droplets in both the WT assay (channel 2, top panel) and VA assay (channel 3, bottom panel).
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edmonston strain measles virus  (ATCC)
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ATCC edmonston strain measles virus
Figure 1. Measles assay development and validation. (A) Alignment of the M gene of measles virus genotypes D8, B3, and <t>Edmonston</t> vaccine (VA) strains. Assays for distinguishing between wild type (WT) strains (D8 and B3) and vaccine strains used the same forward and reverse primers and different probe sequences, shown above the aligned sequences. (B) Validation of the WT2 and vaccine assays using ATCC standards spiked into wastewater. ddPCR results for the WT2 probe (channel 2, top panel) showed a cluster of positive droplets between 5,000 and 6,000 amplitude, corresponding to presence of the WT standard (genotype B3), and a cluster with a 3,000 amplitude corresponding to the vaccine standard. The VA probe (channel 3, bottom panel) showed a positive cluster at 7,000 amplitude corresponding to the VA standard and a positive cluster at 4,000 corresponding to the WT standard. The quantitative results (concentrations and positive droplet counts) are shown in Table SI.12. (C) Validation of the WT2 and vaccine assays using wastewater samples from a facility in Chicago, IL, with an active measles outbreak. All samples showed positive detections of WT strain, as indicated by the amplitude of the positive droplets in both the WT assay (channel 2, top panel) and VA assay (channel 3, bottom panel).
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Figure 1. Measles assay development and validation. (A) Alignment of the M gene of measles virus genotypes D8, B3, and Edmonston vaccine (VA) strains. Assays for distinguishing between wild type (WT) strains (D8 and B3) and vaccine strains used the same forward and reverse primers and different probe sequences, shown above the aligned sequences. (B) Validation of the WT2 and vaccine assays using ATCC standards spiked into wastewater. ddPCR results for the WT2 probe (channel 2, top panel) showed a cluster of positive droplets between 5,000 and 6,000 amplitude, corresponding to presence of the WT standard (genotype B3), and a cluster with a 3,000 amplitude corresponding to the vaccine standard. The VA probe (channel 3, bottom panel) showed a positive cluster at 7,000 amplitude corresponding to the VA standard and a positive cluster at 4,000 corresponding to the WT standard. The quantitative results (concentrations and positive droplet counts) are shown in Table SI.12. (C) Validation of the WT2 and vaccine assays using wastewater samples from a facility in Chicago, IL, with an active measles outbreak. All samples showed positive detections of WT strain, as indicated by the amplitude of the positive droplets in both the WT assay (channel 2, top panel) and VA assay (channel 3, bottom panel).

Journal: Environmental science & technology

Article Title: Multiplexed Detection, Partitioning, and Persistence of Wild-Type and Vaccine Strains of Measles, Mumps, and Rubella Viruses in Wastewater.

doi: 10.1021/acs.est.4c05344

Figure Lengend Snippet: Figure 1. Measles assay development and validation. (A) Alignment of the M gene of measles virus genotypes D8, B3, and Edmonston vaccine (VA) strains. Assays for distinguishing between wild type (WT) strains (D8 and B3) and vaccine strains used the same forward and reverse primers and different probe sequences, shown above the aligned sequences. (B) Validation of the WT2 and vaccine assays using ATCC standards spiked into wastewater. ddPCR results for the WT2 probe (channel 2, top panel) showed a cluster of positive droplets between 5,000 and 6,000 amplitude, corresponding to presence of the WT standard (genotype B3), and a cluster with a 3,000 amplitude corresponding to the vaccine standard. The VA probe (channel 3, bottom panel) showed a positive cluster at 7,000 amplitude corresponding to the VA standard and a positive cluster at 4,000 corresponding to the WT standard. The quantitative results (concentrations and positive droplet counts) are shown in Table SI.12. (C) Validation of the WT2 and vaccine assays using wastewater samples from a facility in Chicago, IL, with an active measles outbreak. All samples showed positive detections of WT strain, as indicated by the amplitude of the positive droplets in both the WT assay (channel 2, top panel) and VA assay (channel 3, bottom panel).

Article Snippet: Next, we validated the multiplexed measles WT2, VA, mumps, and rubella assays using synthetic gblock DNA gene fragments (IDT, Coralville, IA, USA), ATCC viral standards, and wastewater samples spiked with B3 and Edmonston measles strains, mumps, and rubella ATCC standards (Table SI.9 multiplexed assay 3).

Techniques: Biomarker Discovery, Virus

Figure 1. Measles assay development and validation. (A) Alignment of the M gene of measles virus genotypes D8, B3, and Edmonston vaccine (VA) strains. Assays for distinguishing between wild type (WT) strains (D8 and B3) and vaccine strains used the same forward and reverse primers and different probe sequences, shown above the aligned sequences. (B) Validation of the WT2 and vaccine assays using ATCC standards spiked into wastewater. ddPCR results for the WT2 probe (channel 2, top panel) showed a cluster of positive droplets between 5,000 and 6,000 amplitude, corresponding to presence of the WT standard (genotype B3), and a cluster with a 3,000 amplitude corresponding to the vaccine standard. The VA probe (channel 3, bottom panel) showed a positive cluster at 7,000 amplitude corresponding to the VA standard and a positive cluster at 4,000 corresponding to the WT standard. The quantitative results (concentrations and positive droplet counts) are shown in Table SI.12. (C) Validation of the WT2 and vaccine assays using wastewater samples from a facility in Chicago, IL, with an active measles outbreak. All samples showed positive detections of WT strain, as indicated by the amplitude of the positive droplets in both the WT assay (channel 2, top panel) and VA assay (channel 3, bottom panel).

Journal: Environmental science & technology

Article Title: Multiplexed Detection, Partitioning, and Persistence of Wild-Type and Vaccine Strains of Measles, Mumps, and Rubella Viruses in Wastewater.

doi: 10.1021/acs.est.4c05344

Figure Lengend Snippet: Figure 1. Measles assay development and validation. (A) Alignment of the M gene of measles virus genotypes D8, B3, and Edmonston vaccine (VA) strains. Assays for distinguishing between wild type (WT) strains (D8 and B3) and vaccine strains used the same forward and reverse primers and different probe sequences, shown above the aligned sequences. (B) Validation of the WT2 and vaccine assays using ATCC standards spiked into wastewater. ddPCR results for the WT2 probe (channel 2, top panel) showed a cluster of positive droplets between 5,000 and 6,000 amplitude, corresponding to presence of the WT standard (genotype B3), and a cluster with a 3,000 amplitude corresponding to the vaccine standard. The VA probe (channel 3, bottom panel) showed a positive cluster at 7,000 amplitude corresponding to the VA standard and a positive cluster at 4,000 corresponding to the WT standard. The quantitative results (concentrations and positive droplet counts) are shown in Table SI.12. (C) Validation of the WT2 and vaccine assays using wastewater samples from a facility in Chicago, IL, with an active measles outbreak. All samples showed positive detections of WT strain, as indicated by the amplitude of the positive droplets in both the WT assay (channel 2, top panel) and VA assay (channel 3, bottom panel).

Article Snippet: Persistence experiments were performed by spiking the Edmonston strain measles virus (VR-24, ATCC), mumps virus (VR-106, ATCC), and rubella virus (VR-1359, ATCC) into 400 mL of influent wastewater (details in SI 1.5).

Techniques: Biomarker Discovery, Virus