Epidemiological Surveillance of Norovirus and Rotavirus in Sewage (2016–2017) in Valencia (Spain)
Abstract
:1. Introduction
2. Materials and Methods
2.1. Sample Collection and Treatment
2.2. RNA Extraction and Molecular Detection of Noroviruses and Rotaviruses
2.3. Sequencing and Phylogenetic Analysis
3. Results
3.1. Prevalence of Rotaviruses and Noroviruses in Sewage Samples from Valencia
3.2. Rotavirus and Norovirus Genotypes from Sewage Samples
3.3. Phylogenetic Analyses of Sewage Detected Norovirus VP1 Coding Gene
3.4. Phylogenetic Analyses of Sewage Detected Rotavirus VP4 and VP7 Coding Genes
4. Discussion
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
References
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Target | Purpose | Primer or Probe Name | Sequence | Nucleotide Positions |
---|---|---|---|---|
Norovirus GI | qPCR | QNIF4 | 5′-CGC TGG ATG CGN TTC CAT-3′ | 5291–5308 |
Norovirus GI | qPCR | NV1LCR | 5′-CCT TAG ACG CCA TCA TCA TTT AC-3′ | 5354–5376 |
Norovirus GI | qPCR | NVGG1p | VIC-5′-TGG ACA GGA GAY CGC RAT CT-3′-Quencher | 5321–5340 |
Norovirus GII | qPCR | QNIF2 | 5′-ATG TTC AGR TGG ATG AGR TTC TCW GA-3′ | 5012–5037 |
Norovirus GII | qPCR | COG2R | 5′-TCG ACG CCA TCT TCA TTC ACA-3′ | 5080–5100 |
Norovirus GII | qPCR | QNIFs | FAM-5′-AGC ACG TGG GAG GGC GAT CG-3′-Quencher | 5042–5061 |
Mengovirus | qPCR | Mengo 110 | 5′-GCG GGT CCT GCC GAA AGT-3′ | 110–127 |
Mengovirus | qPCR | Mengo 209 | 5′-GAA GTA ACA TAT AGA CAG ACG CAC AC-3′ | 245–270 |
Mengovirus | qPCR | Mengo 147 | FAM-5′-ATC ACA TTA CTG GCC GAA GC-3′-Quencher | 209–227 |
Norovirus GI | 1st PCR | COG1F | 5′-CGY TGG ATG CGN TTY CAT GA-3′ | 5291–5311 |
Norovirus GI | 1st PCR/2nd PCR | * G1SKR | 5′-CCA ACC CAR CCA TTR TAC A-3′ | 5653–5671 |
Norovirus GI | 2nd PCR | * G1SKF | 5′-CTG CCC GAA TTY GTA AAT GA-3′ | 5342–5361 |
Norovirus GII | 1st PCR | COG2F | 5′-CAR GAR BCN ATG TTY AGR TGG ATG AG-3′ | 5003–5029 |
Norovirus GII | 1st PCR/2nd PCR | * G2SKR | 5′-CCR CCN GCA TRH CCR TTR TAC AT-3′ | 5379–5401 |
Norovirus GII | 2nd PCR | * G2SKF | 5′-CNT GGG AGG GCG ATC GCA A-3′ | 5058–5076 |
Rotavirus NSP3 | qPCR | JVKF | 5′-CAG TGG TTG ATG CTC AAG ATG GA-3′ | 17–39 |
Rotavirus NSP3 | qPCR | JVKR | 5′-TCA TTG TAA TCA TAT TGA ATA CCC A-3′ | 123–147 |
Rotavirus NSP3 | qPCR | JVKP | FAM-5′-ACA ACT GCA GCT TCA AAA GAA GWG T-3-Quencher | 72–96 |
Rotavirus VP7 | 1st PCR | VP7-F | 5′-ATG TAT GGT ATT GAA TAT ACC AC-3′ | 51–71 |
Rotavirus VP7 | 1st PCR/2nd PCR | * VP7-R | 5′-AAC TTG CCA CCA TTT TTT CC-3′ | 914–932 |
Rotavirus VP7 | 2nd PCR | * G1 | 5′-CAA GTA CTC AAA TCA ATG ATG G-3′ | 314–335 |
Rotavirus VP7 | 2nd PCR | * G2 | 5′-CAA TGA TAT TAA CAC ATT TTC TGT G-3′ | 411–435 |
Rotavirus VP7 | 2nd PCR | * G3 | 5′-ACG AAC TCA ACA CGA GAG G-3′ | 250–269 |
Rotavirus VP7 | 2nd PCR | G4 | 5′-CGT TTC TGG TGA GGA GTT G-3′ | 480–499 |
Rotavirus VP7 | 2nd PCR | G8 | 5′-GTC ACA CCA TTT GTA AAT TCG-3′ | 178–198 |
Rotavirus VP7 | 2nd PCR | * G9 | 5′-CTT GAT GTG ACT AYA AAT AC-3′ | 757–776 |
Rotavirus VP7 | 2nd PCR | G10 | 5′-ATG TCA GAC TAC ARA TAC TGG-3′ | 666–687 |
Rotavirus VP7 | 2nd PCR | * G12 | 5′-GGT TAT GTA ATC CGA TGG ACG-3′ | 548–567 |
Rotavirus VP4 | 1st PCR/2nd PCR | * VP4-F | 5′-TAT GCT CCA GTN AAT TGG-3′ | 132–149 |
Rotavirus VP4 | 1st PCR | VP4-R | 5′-ATT GCA TTT CTT TCC ATA ATG-3′ | 775–795 |
Rotavirus VP4 | 2nd PCR | P[4]-R | 5′-CTA TTG TTA GAG GTT AGA GTC-3′ | 474–494 |
Rotavirus VP4 | 2nd PCR | P[6]-R | 5′-TGT TGA TTA GTT GGA TTC AA-3′ | 259–278 |
Rotavirus VP4 | 2nd PCR | * P[8]-R | 5′-TCT ACT GGR TTR CAN TGC-3′ | 339–356 |
Rotavirus VP4 | 2nd PCR | P[9]-R | 5′-TGA GAC ATG CAA TTG GAC-3′ | 385–402 |
Rotavirus VP4 | 2nd PCR | P[10]-R | 5′-ATC ATA GTT AGT AGT CGG-3′ | 575–594 |
Rotavirus VP4 | 2nd PCR | P[11]-R | 5′-GTA AAC ATC CAG AAT GTG-3′ | 305–323 |
Sample | Sample | Sampling Date | Norovirus | Rotavirus | Recovery Efficiency | ||||
---|---|---|---|---|---|---|---|---|---|
GI Norovirus | GII Norovirus | ||||||||
Genotype | log PCRU/L ± SD | Genotype | log PCRU/L ± SD | Genotype | log PCRU/L ± SD | % | |||
R1 | #1 | 29.9.16 | ND | 4.98 ± 0.43 | ND | 6.13 ± 0.05 | GxP[8] | 5.21 ± 0.08 | 1.47 |
R3 | #2 | 18.10.16 | GI.4 | 4.74 ± 0.32 | GII.2 | 6.12 ± 0.35 | GxP[8] | 5.46 ± 0.14 | 5.11 |
R4 | #3 | 20.10.16 | ND | 5.01 ± 0.25 | GII.2 | 6.60 ± 0.01 | GxP[8] | 4.62 ± 0.18 | 2.22 |
R5 | #4 | 27.10.16 | ND | 4.92 ± 0.06 | GII.2 | 6.13 ± 0.12 | G12P[8] | 4.14 ± 0.10 | 4.37 |
R9 | #5 | 17.11.2016 | ND | 4.66 ± 0.64 | GII.2 | 6.39 ± 0.00 | G2P[8] | 5.03 ± 0.06 | 7.73 |
R10 | #6 | 24.11.2016 | ND | 5.04 ± 0.28 | GII.2 | 6.46 ± 0.11 | ND | 5.77 ± 0.22 | 6.39 |
R11 | #7 | 2.12.2016 | ND | 4.89 ± 0.06 | GII.2 | 6.24 ± 0.15 | G9P[8] | 5.26 | 1.37 |
R12A | #8 | 6.12.2016 | ND | 5.47 ± 0.43 | GII.2 | 6.95 ± 0.10 | G2P[8] | <LOD | 7.62 |
R13 | #9 | 21.12.2016 | ND | 5.09 ± 0.13 | GII.2 | 6.02 ± 0.00 | G3P[x] | 5.97 | 6.73 |
R14 | #10 | 30.12.2016 | ND | 5.36 ± 0.19 | GII.2 | 6.95 ± 0.05 | GxP[8] | 6.20 ± 0.12 | 7.38 |
R51 | #11 | 15.12.2016 | ND | 5.19 ± 0.32 | ND | 6.22 ± 0.13 | G2P[x] | <LOD | 8.54 |
R15 | #12 | 4.01.2017 | ND | 5.62 ± 0.19 | GII.2 | 6.44 ± 0.10 | G12P[8] | <LOQ | 7.73 |
R16 | #13 | 11.01.2017 | ND | 5.57 ± 0.08 | GII.2 | 6.56 ± 0.06 | G2P[8] | 6.00 ± 0.10 | 7.44 |
R17 | #14 | 18.01.2017 | ND | <LOD | ND | 6.54 ± 0.18 | ND | <LOD | 8.38 |
R18 | #15 | 15.02.2017 | ND | <LOD | ND | <LOD | GxP[8] | 6.24 ± 0.05 | 3.68 |
R19 | #16 | 16.02.2017 | ND | 4.70 ± 0.21 | GII.2 | 6.14 ± 0.09 | G9P[x] | 5.34 ± 0.04 | 4.79 |
R20 | #17 | 2.02.2017 | ND | 5.55 ± 0.04 | GII.2 | 6.53 ± 0.34 | GxP[8] | 5.05 ± 0.09 | 10.82 |
R21 | #18 | 22.02.2017 | ND | 5.47 ± 0.08 | GII.2 | 6.19 ± 0.76 | ND | 4.08 ± 0.23 | 4.39 |
R22 | #19 | 02.03.2017 | ND | <LOD | GII.6 | 5.38 ± 0.21 | GxP[8] | 6.32 ± 0.81 | 18.39 |
R23 | #20 | 08.03.2017 | GI.2 | <LOD | ND | 6.25 | ND | <LOD | 20.50 |
R24 | #21 | 15.03.2017 | GI.2 | 5.62 ± 0.05 | ND | 5.01 ± 1.71 | GxP[8] | 6.55 ± 0.25 | 12.50 |
R25 | #22 | 23.03.2017 | ND | 4.80 ± 0.02 | ND | 5.82 ± 0.05 | GxP[8] | 6.85 ± 0.22 | 11.39 |
R26 | #23 | 30.03.2017 | ND | <LOD | ND | <LOD | ND | 6.19 ± 0.08 | 1.18 |
R27 | #24 | 06.04.2017 | ND | <LOD | ND | <LOD | ND | <LOD | 1.97 |
R28 | #25 | 12.04.2017 | ND | 4.87 ± 0.10 | ND | 5.34 ± 0.00 | ND | 6.92 ± 0.04 | 2.82 |
R29 | #26 | 27.04.2017 | ND | <LOD | ND | <LOD | G2P[8] | 4.59 | 4.31 |
R30 | #27 | 04.05.2017 | ND | <LOD | ND | <LOD | ND | 6.31 ± 0.04 | 4.75 |
R31 | #28 | 11.05.2017 | ND | <LOD | ND | <LOD | G3P[8] | <LOD | 11.03 |
R32 | #29 | 18.05.2017 | ND | <LOD | GII.6 | <LOD | GxP[8] | 6.13 ± 0.01 | 9.05 |
R33 | #30 | 25.02.2017 | ND | <LOD | ND | <LOD | GxP[8] | 6.80 | 9.34 |
R34 | #31 | 01.06.2017 | ND | <LOD | ND | 5.17 | ND | <LOD | 6.73 |
R35 | #32 | 08.06.2017 | ND | <LOD | ND | <LOD | ND | 4.99 | 2.02 |
R36 | #33 | 15.06.2017 | ND | 4.33 ± 0.45 | ND | <LOD | ND | 5.28 | 7.10 |
R37 | #34 | 22.06.2017 | ND | 4.81 ± 0.57 | ND | 4.69 ± 0.00 | ND | <LOD | 9.40 |
R38 | #35 | 28.06.2017 | ND | 5.41 ± 0.01 | GII.17 | 5.04 ± 0.27 | ND | 5.98 | 2.48 |
R39 | #36 | 05.07.2017 | ND | 5.45 ± 0.26 | GII.6 | 5.43 ± 1.58 | ND | 5.65 | 1.53 |
R40 | #37 | 06.07.2017 | ND | 5.03 ± 0.02 | GII.17 | <LOD | ND | 4.95 | 3.53 |
R41 | #38 | 13.07.2017 | GI.4 | 5.73 ± 0.54 | ND | 5.50 ± 0.15 | ND | <LOD | 8.04 |
R42 | #39 | 20.07.2017 | ND | 5.75 ± 0.31 | ND | 5.25 ± 0.00 | ND | 4.73 ± 0.10 | 2.34 |
R43 | #40 | 27.07.2017 | ND | 5.49 ± 0.22 | ND | 5.58 ± 0.13 | ND | <LOD | 26.30 |
R44 | #41 | 10.08.2017 | ND | 4.75 ± 0.37 | ND | 5.23 ± 0.37 | ND | <LOD | 13.44 |
R45 | #42 | 17.08.2017 | ND | 5.51 ± 0.00 | ND | 5.65 ± 0.11 | G1G2P[x] | <LOD | 25.72 |
R46 | #43 | 30.08.2017 | ND | <LOD | ND | 5.13 ± 0.18 | ND | 5.20 ± 0.07 | 10.05 |
R48 | #44 | 7.09.2017 | ND | 5.02 ± 0.17 | ND | 5.16 ± 0.00 | ND | <LOD | 9.86 |
R49 | #45 | 13.09.2017 | ND | 5.67 ± 0.06 | ND | 5.64 ± 0.09 | ND | <LOD | 4.24 |
R50 | #46 | 20.09.2017 | ND | 5.28 ± 0.08 | ND | 6.19 ± 0.09 | ND | 4.72 ± 0.13 | 13.27 |
Norovirus | Rotavirus | ||||
---|---|---|---|---|---|
Genotype | GI (%) | GII (%) | Genotype | VP7 (%) | VP4 (%) |
ND | 87.5 (28/32) | 45.7 (16/35) | ND | 63.5 (21/33) | 42.3 (14/33) |
GI.2 | 6.25 (2/32) | - | P[8] | - | 57.7 (19/33) |
GI.4 | 6.25 (2/32) | - | G1+G2 | 3 (1/33) | - |
GII.2 | - | 40 (14/35) | G2 | 15.2 (5/33) | - |
GII.6 | - | 8.6 (3/35) | G3 | 6.1 (2/33) | - |
GII.17 | - | 5.7 (2/35) | G9 | 6.1 (2/33) | - |
G12 | 6.1 (2/33) | - |
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Santiso-Bellón, C.; Randazzo, W.; Pérez-Cataluña, A.; Vila-Vicent, S.; Gozalbo-Rovira, R.; Muñoz, C.; Buesa, J.; Sanchez, G.; Rodríguez Díaz, J. Epidemiological Surveillance of Norovirus and Rotavirus in Sewage (2016–2017) in Valencia (Spain). Microorganisms 2020, 8, 458. https://doi.org/10.3390/microorganisms8030458
Santiso-Bellón C, Randazzo W, Pérez-Cataluña A, Vila-Vicent S, Gozalbo-Rovira R, Muñoz C, Buesa J, Sanchez G, Rodríguez Díaz J. Epidemiological Surveillance of Norovirus and Rotavirus in Sewage (2016–2017) in Valencia (Spain). Microorganisms. 2020; 8(3):458. https://doi.org/10.3390/microorganisms8030458
Chicago/Turabian StyleSantiso-Bellón, Cristina, Walter Randazzo, Alba Pérez-Cataluña, Susana Vila-Vicent, Roberto Gozalbo-Rovira, Carlos Muñoz, Javier Buesa, Gloria Sanchez, and Jesús Rodríguez Díaz. 2020. "Epidemiological Surveillance of Norovirus and Rotavirus in Sewage (2016–2017) in Valencia (Spain)" Microorganisms 8, no. 3: 458. https://doi.org/10.3390/microorganisms8030458
APA StyleSantiso-Bellón, C., Randazzo, W., Pérez-Cataluña, A., Vila-Vicent, S., Gozalbo-Rovira, R., Muñoz, C., Buesa, J., Sanchez, G., & Rodríguez Díaz, J. (2020). Epidemiological Surveillance of Norovirus and Rotavirus in Sewage (2016–2017) in Valencia (Spain). Microorganisms, 8(3), 458. https://doi.org/10.3390/microorganisms8030458