Genetic Epidemiology Reveals 3 Chronic Reservoir Areas With Recurrent Population Mobility Challenging Poliovirus Eradication in Pakistan

Akhtar, Ribqa; Mahmood, Nayab; Alam, Muhammad Masroor; Naeem, Muhammad; Zaidi, Syed Sohail Zahoor; Sharif, Salman; Khattak, Zainab; Arshad, Yasir; Khurshid, Adnan; Mujtaba, Ghulam; Rehman, Lubna; Angez, Mehar; Shaukat, Shehzad; Mushtaq, Naveed; Umair, Massab; Ikram, Aamer; Salman, Muhammad

doi:10.1093/cid/ciz1037

Cited by 9 publications

(7 citation statements)

References 23 publications

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“…Immediate identification of the poliovirus sequence allows downstream phylogenetic analysis and interpretation of epidemiological significance ( 29 , 30 ). Faster generation of these results will allow earlier vaccination responses and containment of virus spread, which can have a substantially greater impact on poliomyelitis cases during an outbreak ( 31 ).…”

Section: Discussionmentioning

confidence: 99%

Rapid and Sensitive Direct Detection and Identification of Poliovirus from Stool and Environmental Surveillance Samples by Use of Nanopore Sequencing

et al. 2020

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Global poliovirus surveillance involves virus isolation from stool and environmental samples, intratypic differential (ITD) by PCR and sequencing of the VP1 region to distinguish vaccine (Sabin), vaccine-derived and wild-type polioviruses and ensure an appropriate response. This cell-culture algorithm takes 2-3 weeks on average between sample receipt and sequencing. Direct detection of viral RNA using PCR allows faster detection but has traditionally faced challenges related to poor sensitivity and difficulties in sequencing common samples containing poliovirus and enterovirus mixtures. We present a nested PCR and nanopore sequencing protocol that allows rapid (<3 days) and sensitive direct detection and sequencing of polioviruses in stool and environmental samples. We developed barcoded primers and a real-time analysis platform that generate accurate VP1 consensus sequences from multiplexed samples. The sensitivity and specificity compared with cell-culture were 90.9% (95% Confidence Interval: 75.7-98.1%) and 99.2% (95.5-100.0%) for wild-type 1 poliovirus, 92.5% (79.6-98.4%) and 98.7% (95.4-99.8%) for vaccine and vaccine-derived serotype 2 poliovirus, and 88.3% (81.2-93.5%) and 93.2% (88.6-96.3%) for Sabin 1 and 3 poliovirus alone or in mixtures when tested on 155 stool samples in Pakistan. Variant analysis of sequencing reads also allowed identification of polioviruses and enteroviruses in artificial mixtures and was able to distinguish complex mixtures of polioviruses in environmental samples. The median identity of consensus nanopore sequences with Sanger or Illumina sequences from the same samples was >99.9%. This novel method shows promise as a faster and safer alternative to cell-culture for the detection and real-time sequencing of polioviruses in stool and environmental samples.

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Section: Discussionmentioning

confidence: 99%

Rapid and Sensitive Direct Detection and Identification of Poliovirus from Stool and Environmental Surveillance Samples by Use of Nanopore Sequencing

et al. 2020

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“…Qβ). 357 358 Immediate identification of the poliovirus sequence allows downstream phylogenetic analysis 359 and interpretation of epidemiological significance (30,31). Faster generation of these results will 360 allow earlier vaccination responses and containment of virus spread, which can have a 361 substantially greater impact on poliomyelitis cases during an outbreak (32).…”

Section: Real-time Visualisation Of Demultiplexed Sequencing Results 338mentioning

confidence: 99%

Rapid and sensitive direct detection and identification of poliovirus from stool and environmental surveillance samples using nanopore sequencing

Shaw¹,

Majumdar²,

Troman³

et al. 2020

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25Global poliovirus surveillance involves virus isolation from stool and environmental samples, 26 intratypic differential (ITD) by PCR and sequencing of the VP1 region to distinguish vaccine 27 (Sabin), vaccine-derived and wild-type polioviruses and ensure an appropriate response. This 28 cell-culture algorithm takes 2-3 weeks on average between sample receipt and sequencing. 29Direct detection of viral RNA using PCR allows faster detection but has traditionally faced 30 challenges related to poor sensitivity and difficulties in sequencing common samples containing 31 poliovirus and enterovirus mixtures. We present a nested PCR and nanopore sequencing protocol 32 that allows rapid (<3 days) and sensitive direct detection and sequencing of polioviruses in stool 33 and environmental samples. We developed barcoded primers and a real-time analysis platform 34 that generate accurate VP1 consensus sequences from multiplexed samples. The sensitivity and 35 specificity compared with cell-culture were 90.9% (95% Confidence Interval: 75.7-98.1%) and 36 99.2% (95.5-100.0%) for wild-type 1 poliovirus, 92.5% (79.6-98.4%) and 98.7% (95.4-99.8%) 37 for vaccine and vaccine-derived serotype 2 poliovirus, and 88.3% (81.2-93.5%) and 93.2% 38 (88.6-96.3%) for Sabin 1 and 3 poliovirus alone or in mixtures when tested on 155 stool samples 39 in Pakistan. Variant analysis of sequencing reads also allowed identification of polioviruses and 40 enteroviruses in artificial mixtures and was able to distinguish complex mixtures of polioviruses 41 in environmental samples. The median identity of consensus nanopore sequences with Sanger or 42 Illumina sequences from the same samples was >99.9%. This novel method shows promise as a 43 faster and safer alternative to cell-culture for the detection and real-time sequencing of 44 polioviruses in stool and environmental samples. 45 46

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“…They also highlight repeated introductions of virus from the tribal regions (between Islamabad and Kabul) into the Punjab province of Pakistan and show uninterrupted virus circulation within these tribal regions. Analysis of the distribution of clusters of transmission without formal DTA analysis also highlights these three, key, high-burden regions but does not construct probable ancestral locations or virus movements ( Akhtar et al. 2019 ).…”

Section: Investigation Of Transmissionmentioning

confidence: 99%

The role of genetic sequencing and analysis in the polio eradication programme

et al. 2020

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Genetic sequencing of polioviruses detected through clinical and environmental surveillance is used to confirm detection, identify their likely origin, track geographic patterns of spread and determine the appropriate vaccination response. The critical importance of genetic sequencing and analysis to the Global Polio Eradication Initiative has grown with the increasing incidence of vaccine-derived poliovirus infections in Africa specifically (470 reported cases in 2019), and globally, alongside persistent transmission of serotype 1 wild-type poliovirus in Pakistan and Afghanistan (197 reported cases in 2019). Adapting what has been learned about the virus genetics and evolution to address these threats has been a major focus of recent work. Here we review how phylogenetic and phylogeographic methods have been used to trace the spread of wild-type polioviruses and identify the likely origins of vaccine-derived polioviruses. We highlight the analysis methods and sequencing technology currently used and the potential for new technologies to speed up poliovirus detection and the interpretation of genetic data. At a pivotal point in the eradication campaign with the threat of anti-vaccine sentiment and donor and public fatigue, innovation is critical to maintain drive and overcome the last remaining circulating virus.

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Genetic Epidemiology Reveals 3 Chronic Reservoir Areas With Recurrent Population Mobility Challenging Poliovirus Eradication in Pakistan

Cited by 9 publications

References 23 publications

Rapid and Sensitive Direct Detection and Identification of Poliovirus from Stool and Environmental Surveillance Samples by Use of Nanopore Sequencing

Rapid and Sensitive Direct Detection and Identification of Poliovirus from Stool and Environmental Surveillance Samples by Use of Nanopore Sequencing

Rapid and sensitive direct detection and identification of poliovirus from stool and environmental surveillance samples using nanopore sequencing

The role of genetic sequencing and analysis in the polio eradication programme

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