Evaluation and validation of an RT-PCR assay for specific detection of monkeypox virus (MPXV)

Alberto Paniz-Mondolfi; Susana Guerra; Marina Muñoz; Nicolas Luna; Matthew M. Hernandez; Luz H. Patino; Jason Reidy; Radhika Banu; Paras Shrestha; Bernadette Liggayu; Audrey Umeaku; Feng Chen; Liyong Cao; Armi Patel; Ayman Hanna; Sunny Li; Andy Look; Nina Pagani; Randy Albrecht; Rebecca Pearl; Adolfo Garcia-Sastre; Dusan Bogunovic; Gustavo Palacios; Lucia Bonnier; Freddy Cera; Heidi Lopez; Yvette Calderon; Erick Eiting; Karr Mullen; Sangyoon Jason Shin; Luz Amarilis Lugo; Antonio E. Urbina; Carlotta Starks; Tonny Koo; Patricia Uychiat; Avery Look; Harm van Bakel; Ana Gonzalez-Reiche; Adolfo Firpo Betancourt; David Reich; Carlos Cordon-Cardo; Viviana Simon; Emilia M. Sordillo; Juan David Ramírez

doi:10.1002/jmv.28247

Evaluation and validation of an RT-PCR assay for specific detection of monkeypox virus (MPXV)

Alberto Paniz-Mondolfi, Susana Guerra, Marina Muñoz, Nicolas Luna, Matthew M. Hernandez, Luz H. Patino, Jason Reidy, Radhika Banu, Paras Shrestha, Bernadette Liggayu, Audrey Umeaku, Feng Chen, Liyong Cao, Armi Patel, Ayman Hanna, Sunny Li, Andy Look, Nina Pagani, Randy Albrecht, Rebecca PearlAdolfo Garcia-Sastre, Dusan Bogunovic, Gustavo Palacios, Lucia Bonnier, Freddy Cera, Heidi Lopez, Yvette Calderon, Erick Eiting, Karr Mullen, Sangyoon Jason Shin, Luz Amarilis Lugo, Antonio E. Urbina, Carlotta Starks, Tonny Koo, Patricia Uychiat, Avery Look, Harm van Bakel, Ana Gonzalez-Reiche, Adolfo Firpo Betancourt, David Reich, Carlos Cordon-Cardo, Viviana Simon, Emilia M. Sordillo, Juan David Ramírez

Research output: Contribution to journal › Article › peer-review

36 Scopus citations

Abstract

Monkeypox virus (MPXV) is a zoonotic orthopoxvirus within the Poxviridae family. MPXV is endemic to Central and West Africa. However, the world is currently witnessing an international outbreak with no clear epidemiological links to travel or animal exposure and with ever-increasing numbers of reported cases worldwide. Here, we evaluated and validated a new, sensitive, and specific real-time PCR-assay for MPXV diagnosis in humans and compare the performance of this novel assay against a Food & Drug Administration-cleared pan-Orthopox RT-PCR assay. We determined specificity, sensitivity, and analytic performance of the PKamp™ Monkeypox Virus RT-PCR assay targeting the viral F3L-gene. In addition, we further evaluated MPXV-PCR-positive specimens by viral culture, electron microscopy, and viral inactivation assays. The limit of detection was established at 7.2 genome copies/reaction, and MPXV was successfully identified in 20 clinical specimens with 100% correlation against the reference method with 100% sensitivity and specificity. Our results demonstrated the validity of this rapid, robust, and reliable RT-PCR assay for specific and accurate diagnosis of MPXV infection in human specimens collected both as dry swabs and in viral transport media. This assay has been approved by NYS Department of Health for clinical use.

Original language	English
Article number	e28247
Journal	Journal of Medical Virology
Volume	95
Issue number	1
DOIs	https://doi.org/10.1002/jmv.28247
State	Published - Jan 2023

Keywords

diagnosis
monkeypox virus
orthopoxvirus
real-time PCR

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Paniz-Mondolfi, A., Guerra, S., Muñoz, M., Luna, N., Hernandez, M. M., Patino, L. H., Reidy, J., Banu, R., Shrestha, P., Liggayu, B., Umeaku, A., Chen, F., Cao, L., Patel, A., Hanna, A., Li, S., Look, A., Pagani, N., Albrecht, R., ... Ramírez, J. D. (2023). Evaluation and validation of an RT-PCR assay for specific detection of monkeypox virus (MPXV). Journal of Medical Virology, 95(1), Article e28247. https://doi.org/10.1002/jmv.28247

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abstract = "Monkeypox virus (MPXV) is a zoonotic orthopoxvirus within the Poxviridae family. MPXV is endemic to Central and West Africa. However, the world is currently witnessing an international outbreak with no clear epidemiological links to travel or animal exposure and with ever-increasing numbers of reported cases worldwide. Here, we evaluated and validated a new, sensitive, and specific real-time PCR-assay for MPXV diagnosis in humans and compare the performance of this novel assay against a Food & Drug Administration-cleared pan-Orthopox RT-PCR assay. We determined specificity, sensitivity, and analytic performance of the PKamp{\texttrademark} Monkeypox Virus RT-PCR assay targeting the viral F3L-gene. In addition, we further evaluated MPXV-PCR-positive specimens by viral culture, electron microscopy, and viral inactivation assays. The limit of detection was established at 7.2 genome copies/reaction, and MPXV was successfully identified in 20 clinical specimens with 100% correlation against the reference method with 100% sensitivity and specificity. Our results demonstrated the validity of this rapid, robust, and reliable RT-PCR assay for specific and accurate diagnosis of MPXV infection in human specimens collected both as dry swabs and in viral transport media. This assay has been approved by NYS Department of Health for clinical use.",

keywords = "diagnosis, monkeypox virus, orthopoxvirus, real-time PCR",

author = "Alberto Paniz-Mondolfi and Susana Guerra and Marina Mu{\~n}oz and Nicolas Luna and Hernandez, {Matthew M.} and Patino, {Luz H.} and Jason Reidy and Radhika Banu and Paras Shrestha and Bernadette Liggayu and Audrey Umeaku and Feng Chen and Liyong Cao and Armi Patel and Ayman Hanna and Sunny Li and Andy Look and Nina Pagani and Randy Albrecht and Rebecca Pearl and Adolfo Garcia-Sastre and Dusan Bogunovic and Gustavo Palacios and Lucia Bonnier and Freddy Cera and Heidi Lopez and Yvette Calderon and Erick Eiting and Karr Mullen and Shin, {Sangyoon Jason} and Lugo, {Luz Amarilis} and Urbina, {Antonio E.} and Carlotta Starks and Tonny Koo and Patricia Uychiat and Avery Look and {van Bakel}, Harm and Ana Gonzalez-Reiche and Betancourt, {Adolfo Firpo} and David Reich and Carlos Cordon-Cardo and Viviana Simon and Sordillo, {Emilia M.} and Ram{\'i}rez, {Juan David}",

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year = "2023",

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doi = "10.1002/jmv.28247",

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Paniz-Mondolfi, A, Guerra, S, Muñoz, M, Luna, N, Hernandez, MM, Patino, LH, Reidy, J, Banu, R, Shrestha, P, Liggayu, B, Umeaku, A, Chen, F, Cao, L, Patel, A, Hanna, A, Li, S, Look, A, Pagani, N, Albrecht, R, Pearl, R, Garcia-Sastre, A, Bogunovic, D, Palacios, G, Bonnier, L, Cera, F, Lopez, H, Calderon, Y, Eiting, E, Mullen, K, Shin, SJ, Lugo, LA, Urbina, AE, Starks, C, Koo, T, Uychiat, P, Look, A, van Bakel, H, Gonzalez-Reiche, A, Betancourt, AF, Reich, D, Cordon-Cardo, C, Simon, V , Sordillo, EM & Ramírez, JD 2023, 'Evaluation and validation of an RT-PCR assay for specific detection of monkeypox virus (MPXV)', Journal of Medical Virology, vol. 95, no. 1, e28247. https://doi.org/10.1002/jmv.28247

TY - JOUR

T1 - Evaluation and validation of an RT-PCR assay for specific detection of monkeypox virus (MPXV)

AU - Paniz-Mondolfi, Alberto

AU - Guerra, Susana

AU - Muñoz, Marina

AU - Luna, Nicolas

AU - Hernandez, Matthew M.

AU - Patino, Luz H.

AU - Reidy, Jason

AU - Banu, Radhika

AU - Shrestha, Paras

AU - Liggayu, Bernadette

AU - Umeaku, Audrey

AU - Chen, Feng

AU - Cao, Liyong

AU - Patel, Armi

AU - Hanna, Ayman

AU - Li, Sunny

AU - Look, Andy

AU - Pagani, Nina

AU - Albrecht, Randy

AU - Pearl, Rebecca

AU - Garcia-Sastre, Adolfo

AU - Bogunovic, Dusan

AU - Palacios, Gustavo

AU - Bonnier, Lucia

AU - Cera, Freddy

AU - Lopez, Heidi

AU - Calderon, Yvette

AU - Eiting, Erick

AU - Mullen, Karr

AU - Shin, Sangyoon Jason

AU - Lugo, Luz Amarilis

AU - Urbina, Antonio E.

AU - Starks, Carlotta

AU - Koo, Tonny

AU - Uychiat, Patricia

AU - Look, Avery

AU - van Bakel, Harm

AU - Gonzalez-Reiche, Ana

AU - Betancourt, Adolfo Firpo

AU - Reich, David

AU - Cordon-Cardo, Carlos

AU - Simon, Viviana

AU - Sordillo, Emilia M.

AU - Ramírez, Juan David

PY - 2023/1

Y1 - 2023/1

N2 - Monkeypox virus (MPXV) is a zoonotic orthopoxvirus within the Poxviridae family. MPXV is endemic to Central and West Africa. However, the world is currently witnessing an international outbreak with no clear epidemiological links to travel or animal exposure and with ever-increasing numbers of reported cases worldwide. Here, we evaluated and validated a new, sensitive, and specific real-time PCR-assay for MPXV diagnosis in humans and compare the performance of this novel assay against a Food & Drug Administration-cleared pan-Orthopox RT-PCR assay. We determined specificity, sensitivity, and analytic performance of the PKamp™ Monkeypox Virus RT-PCR assay targeting the viral F3L-gene. In addition, we further evaluated MPXV-PCR-positive specimens by viral culture, electron microscopy, and viral inactivation assays. The limit of detection was established at 7.2 genome copies/reaction, and MPXV was successfully identified in 20 clinical specimens with 100% correlation against the reference method with 100% sensitivity and specificity. Our results demonstrated the validity of this rapid, robust, and reliable RT-PCR assay for specific and accurate diagnosis of MPXV infection in human specimens collected both as dry swabs and in viral transport media. This assay has been approved by NYS Department of Health for clinical use.

AB - Monkeypox virus (MPXV) is a zoonotic orthopoxvirus within the Poxviridae family. MPXV is endemic to Central and West Africa. However, the world is currently witnessing an international outbreak with no clear epidemiological links to travel or animal exposure and with ever-increasing numbers of reported cases worldwide. Here, we evaluated and validated a new, sensitive, and specific real-time PCR-assay for MPXV diagnosis in humans and compare the performance of this novel assay against a Food & Drug Administration-cleared pan-Orthopox RT-PCR assay. We determined specificity, sensitivity, and analytic performance of the PKamp™ Monkeypox Virus RT-PCR assay targeting the viral F3L-gene. In addition, we further evaluated MPXV-PCR-positive specimens by viral culture, electron microscopy, and viral inactivation assays. The limit of detection was established at 7.2 genome copies/reaction, and MPXV was successfully identified in 20 clinical specimens with 100% correlation against the reference method with 100% sensitivity and specificity. Our results demonstrated the validity of this rapid, robust, and reliable RT-PCR assay for specific and accurate diagnosis of MPXV infection in human specimens collected both as dry swabs and in viral transport media. This assay has been approved by NYS Department of Health for clinical use.

KW - diagnosis

KW - monkeypox virus

KW - orthopoxvirus

KW - real-time PCR

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U2 - 10.1002/jmv.28247

DO - 10.1002/jmv.28247

M3 - Article

C2 - 36271493

AN - SCOPUS:85141377699

SN - 0146-6615

VL - 95

JO - Journal of Medical Virology

JF - Journal of Medical Virology

IS - 1

M1 - e28247

ER -

Evaluation and validation of an RT-PCR assay for specific detection of monkeypox virus (MPXV)

Abstract

Keywords

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