Friday, 21st of February 2014 Print


Our study highlights the critical role of environmental surveillance for monitoring global WPV circulation.

Abstract, introduction, discussion and references below; full text, with figures, is at 

Eurosurveillance, Volume 19, Issue 7, 20 February 2014

Surveillance and outbreak reports

Intensified environmental surveillance supporting the response to wild poliovirus type 1 silent circulation in Israel, 2013

Y Manor1, L M Shulman1,2, E Kaliner3, M Hindiyeh1, D Ram1, D Sofer1, J Moran-Gilad3,4, B Lev5, I Grotto3,6, R Gamzu2,5, E Mendelson ()1,2

  1. Central Virology Laboratory, Ministry of Health, Chaim Sheba Medical Center, Tel Hashomer, Israel
  2. School of Public Health, Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv, Israel
  3. Public Health Services, Ministry of Health, Jerusalem, Israel
  4. European Society of Clinical Microbiology and Infectious Diseases (ESCMID) Study Group for Molecular Diagnostics (ESGMD)
  5. Directorate, Ministry of Health, Jerusalem, Israel
  6. Department of Public Health, Faculty of Health Sciences, Ben Gurion University of the Negev, Beer Sheva, Israel

Citation style for this article: Manor Y, Shulman LM, Kaliner E, Hindiyeh M, Ram D, Sofer D, Moran-Gilad J, Lev B, Grotto I, Gamzu R, Mendelson E. Intensified environmental surveillance supporting the response to wild poliovirus type 1 silent circulation in Israel, 2013 . Euro Surveill. 2014;19(7):pii=20708. Available online:
Date of submission: 22 October 2013

An emergency response was triggered by recovery of wild poliovirus type 1 (WPV1) of the South Asia (SOAS) lineage from sewage in southern Israel in April 2013 during routine environmental surveillance. Public health risk assessment necessitated intensification of environmental surveillance in order to facilitate countrywide monitoring of WPV1-SOAS circulation. This involved increasing sampling frequency and broadening the geographical area, for better coverage of the population at risk, as well as modifying sewage testing algorithms to accommodate a newly developed WPV1-SOAS-specific quantitative real-time RT-PCR assay for screening of RNA extracted directly from sewage concentrates, in addition to standard virus isolation. Intensified surveillance in 74 sites across Israel between 1 February and 31 August 2013 documented a sustained high viral load of WPV1-SOAS in sewage samples from six Bedouin settlements and two cities with Jewish and Arab populations in the South district. Lower viral loads and intermittent detection were documented in sampling sites representing 14 mixed communities in three of the five health districts in central and northern Israel. Environmental surveillance plays a fundamental role in routine monitoring of WPV circulation in polio-free countries. The rapid assay specific for the circulating strain facilitated implementation of intensified surveillance and informed the public health response and decision-making.


In the drive towards global eradication of poliomyelitis, as of 2013, only three countries remained endemic for wild-type poliovirus (WPV) infection: Afghanistan, Pakistan and Nigeria, with recent introduction of the virus resulting in paralytic cases to countries with suboptimal immunisation coverage in Africa (Somalia, Kenya, Ethiopia and Cameroon) and in Asia (Syria) [1]. Only WPV type 1 (WPV1) is currently circulating, consisting of two major lineages: the South Asia (SOAS) lineage, which is indigenous to Pakistan and Afghanistan, and the West Africa (WEAF) B lineage, which is indigenous to Nigeria [2].

Poliovirus circulation in highly immune populations is far less likely to be detected via identification of clinical paralytic poliomyelitis cases, which are expected to be very rare or absent (silent circulation), and thus environmental surveillance for poliovirus has become a very useful tool for population-based alert and monitoring of WPV activity. This approach has been implemented by several countries, both for early detection of WPV introduction and transmission as well as for detection of vaccine-derived neurovirulent polioviruses (VDPVs) that emerge following the use of oral poliovirus vaccines (OPVs) [3,4].

Israel has been free of poliomyelitis since the last outbreak caused by WPV1 in 1988, which resulted in 15 paralytic cases [5,6]. The outbreak strain originated in Egypt and arrived in 1987 from Gaza to Rahat and spread in 1988 from Rahat to central Israel. A routine environmental surveillance for poliovirus programme has been implemented since then in Israel, Gaza and the West Bank, which monitors sentinel sites that represent large populations (such as the Shaf-Dan, a sewage treatment facility in the metropolitan Tel Aviv area) and populations (such as Rahat) considered at high risk for introduction of WPV from other countries. A combined inactivated poliovirus vaccine (IPV)/trivalent oral poliovirus vaccine (tOPV) routine vaccination schedule was in place until 2005, when it was replaced by an IPV-only schedule [13].

Over the years, the environmental surveillance programme has detected several introductions of WPV into Gaza and Israel, but subsequent circulation in the local community occurred only once in Gaza in 1994–45 [7-9]. In addition, it has detected two lineages of highly diverged type 2 VDPV in the Tel Aviv sewage system, excreted by single individuals [9-11], demonstrating the high sensitivity of environmental surveillance for monitoring large populations. Laboratory methods for sample treatment and poliovirus isolation, including the plaque formation approach, are also a major factor in the sensitivity of the environmental surveillance, as reviewed by Hovi et al. [3]. Plaque formation allows a rough estimation of the virus circulation intensity since the number of viral plaque-forming units (PFU) present in the original sewage sample can be deduced based on spiking experiments [8].

In December 2012, WPV1-SOAS was detected in sewage collected from Cairo, Egypt [11], where systematic environmental surveillance for poliovirus has been in place since 2000. A large immunisation campaign initiated as a response led to the disappearance of the virus from the sewage and, by implication, from the population at large [12].

In April 2013, a surge in the number of plaques recovered on L20B cells from a sewage sample collected in Rahat and Beer Sheva, two major cities in southern Israel, occurred. Identification of the plaques as WPV1 suggested an importation and possible circulation of WPV1 in the region [13]. These alarming findings prompted an urgent assessment and response by the Public Health Services of the Ministry of Health. Notably, intensification of environmental surveillance for poliovirus played a key role in monitoring the spread of the virus. Here we describe the modification and enhancement of the environmental sampling and laboratory methods used in order to meet the increased demand for processing of sewage samples and generation of surveillance data that will inform public health response and incident management.

. . .


Our study highlights the critical role of environmental surveillance for monitoring global WPV circulation. In countries with high vaccination coverage (about 90–95%),  acute flaccid paralysis (AFP) surveillance might not detect virus introduction and circulation as occurred in Israel: enhanced AFP surveillance and aseptic meningitis surveillance implemented in response to the WPV1-SOAS detection in sewage did not identify any polio-associated illness from early June to late August [13] and later (data not shown). In contrast to previous introductions of WPV1 to Israel and Gaza detected by the environmental surveillance [11], after which the virus disappeared without the need for supplemental immunisation (in Israel) or following national immunisation days (in Gaza) [7], in 2013, virus importation resulted in sustained circulation [13], which eventually necessitated supplemental immunisation activity with bOPV.

The intensified environmental surveillance for polio, which was essential for management of the event, led to the development of a novel approach, implementation of a qRT-PCR assay specific to the outbreak virus. Together with modifying the testing algorithm, this allowed testing a large number of samples to be tested and result produced within a few days. The plaque and qRT-PCR assays are very precise and quantitative when used on pure viral stocks or on spiked negative sewage samples [15]. Although they are less precise when used on positive sewage samples, because of their variable contents [3], they were still very useful in assessing the intensity of virus circulation in different communities. This approach had not been used in previous outbreak investigations and allowed identification of the most affected communities and the epicentre of the silent outbreak. For example, in Rahat we continuously obtained a few hundred plaques per mL of concentrated sewage (after correcting for the dilution factor) while in Kiryat Gat the numbers ranged between less than 10 and up to 80. Thus, we estimated that the number of WPV1 excretors in Rahat may reach hundreds and may be much higher than in Kiryat Gat. These estimates were later confirmed by a stool survey that assessed the prevalence of WPV1 excretion among subpopulations (data not shown). WPV1 circulation was probably propagated by the accumulation of a large cohort of children who had been immunised only with IPV, and not with a combination of IPV and OPV, since 2005 [13]. The qRT-PCR assay replaced the plaque assay for quantification of WPV1-SOAS excretion after the beginning of the immunisation campaign since the plaque assay was unable to distinguish between WPV1-SOAS and the Sabin 1 and 3 vaccine strains found at high concentration in the sewage.

While no paralytic poliomyelitis cases were identified, the epidemiological picture that unfolded by the intensified environmental surveillance, including sample collection from sewage upstream lines, was very detailed with regard to virus circulation in different communities. This included persistently positive sites (mostly with a high number of plaques and corresponding low Ct values in the qRT-PCR), with sustained virus transmission rates and intermittently positive sites (lower number of plaques and higher Ct values), which indicated either a lower rate of virus transmission or occasional importations by visitors or dayworkers from Rahat (involved in various types of work), who we assume were a possible source of the virus found in the sewage. On the basis of intensive sewage surveillance using semi-quantitative methods, we speculated that the epicentre of the outbreak was in the Bedouin communities in southern Israel, in which sustained transmission has occurred. These findings have prompted an IPV catch-up campaign among Bedouin communities to minimise the already low risk for clinical poliomyelitis and to initiate bOPV supplemental immunisation to communities in southern Israel before introducing the campaign to the rest of the country.

Our report highlights the importance of environmental surveillance, which is the most sensitive and efficient approach for detection of WPV introduction and circulation in highly immunised populations. It requires systematic composite sample collection and experienced laboratories, but under these conditions, it has the highest population sensitivity, compared with other methods [3]. The routine environmental surveillance programme implemented in Israel since 1989 included monthly sampling of large cities and populations at high risk of virus penetration, covering around 40–50% of the Israeli population, which now counts around 8 million people [19]. During the silent WPV1 outbreak, coverage of the programme was increased, to around 70% of the population, focusing on communities with circulating virus. Other supplementary surveillance approaches addressing the general population rather than only AFP cases, which are currently in use in countries in Europe and elsewhere and which can detect subclinical circulation, are stool surveys or general testing of enterovirus PCR-positive stools from patients without poliomyelitis symptoms for the presence of poliovirus by culture on L20B cells. France, the Netherlands and Australia, for example, implement enterovirus surveillance that includes testing for poliovirus [20-22]. However, none of the surveillance methods used is comparable to environmental surveillance in efficiently covering large populations. For example, in France 192,598 samples were tested over five years, between 2000 and 2004 [20], which on an average yearly basis represent roughly less than 0.3% of the French population [23], while in Israel, routine surveillance before 2013 covered 30–40% of the population. Environmental surveillance could be very useful for monitoring and detecting WPV introduction and silent circulation in countries that use only IPV.

The WPV1-SOAS specific qRT-PCR assay is limited to the detection of this particular virus lineage and may lose its sensitivity even if there are minor mutations, which may occur naturally during WPV evolution. It cannot detect other poliovirus types or strains such as the WEAF lineage or type 2 vaccine-derived polioviruses (VDPV2), which circulate in Africa. Therefore, adoption of this method should take these limitations into consideration.

In conclusion, our study provides a proof of concept for the rapid implementation of qRT-PCR in the framework of an outbreak in which short turnaround times and high-throughput testing are essential for incident management, while maintaining confirmatory culture-based methods. In addition, molecular assays capable of directly detecting a wider range of wild polioviruses and VDPVs currently in global circulation should be developed for routine surveillance and emergency response.

The authors are grateful to the dedicated staff of the Central Virology Laboratory, particularly Tova Halmout, Irena Agobaiev and Yuri Perepliotchikov (sample processing and virus culturing), Roberto Azar (sequencing of viral isolates), Jacklyn Alfandary and Ilana Zilberstein (molecular confirmation). We also wish to thank Michal Tepperberg, Michal Mandelboim, Liora Regev, Hilda Shaharbany, Virginia Levi and Irena Jornist for helping in sample processing and testing at times of heavy workload. We also wish to thank the South Health District Office and Environmental Health Department, particularly Yotvat Bar El (sample acquisition) and the Israel Centre for Disease Control, particularly Zalman Kaufman and Tamar Shohat (GIS maps).

Conflict of interest
None declared.

Authors contributions
Yosef Manor: responsible for conducting the polio sewage surveillance, virus isolation and data analysis. Planned conducted and analysed results of the sewage surveillance; participated in writing of the manuscript.
Lester M Shulman: .planned the laboratory algorithm, analysed results of the sewage surveillance, supervised and conducted typing and molecular analyses, participated in writing of the manuscript.
Ehud Kaliner: was in charge of supervising the environmental surveillance activities, sampling programme and participated in data analysis.
Musa Hindiyeh: in charge of assay development and performance of clinical testing. Developed and validated the real-time RT-PCR assays for the wild poliovirus type 1 (SOAS) and for Sabin 1 and Sabin 3 strains. Conducted all the real-time RT-PCR testing and analysed the results.
Daniela Ram: participated in development and validation of the real-time RT-PCR specific assay for SOAS, Sabin 1 and Sabin 3 and, participated in testing and data analysis.
Jacob Moran Gilad: led and guided the validation of the real-time RT-PCR specific assays for SOAS, Sabin 1 and Sabin 3. Participated in data analysis and in drafting of the manuscript.
Danit Sofer: conducted virus isolations in tube cultures and participated in the validation of the sensitivity and specificity of the real-time RT-PCR specific assays and in data analysis.
Boaz Lev: participated in supervision and evaluation of the surveillance activities and results on a national level.
Itamar Grotto: participated and supervised the environmental surveillance activity in all districts, and in data analysis and manuscript preparation.
Roni Gamzu: was involved in evaluation and in routine consultations regarding the environmental surveillance sampling programme and results.
Ella Mendelson: coordinated and supervised the laboratory groups work, participated in planning of the environmental surveillance, development and validation of the real-time RT-PCR assay, and data analysis, wrote the manuscript.


  1. Global Polio Eradication Initiative. Polio this week. Geneva: Global Polio Eradication Initiative. [Accessed 20 Feb 2014]. Available from:
  2. Centers for Disease Control and Prevention (CDC). Laboratory surveillance for wild and vaccine-derived polioviruses - worldwide, January 2008-June 2009. MMWR Morb Mortal Wkly Rep. 2009 Sep 4;58(34):950-4.
  3. Hovi T, Shulman LM, van der Avoort H, Deshpande J, Roivainen M, de Gourville EM. Role of environmental poliovirus surveilllance in global polio eradication and beyond. Epidemiol Infect. 2012;140(1):1-13.
  4. Blomqvist S, El Bassioni L, El Maamoon Nasr EM, Paananen A, Kaijalainen S, Asghar H, et al. Detection of imported wild polioviruses and of vaccine-derived polioviruses by environmental surveillance in Egypt. Appl Environ Microbiol. 2012;78(15):5406-9.
  5. Slater PE, Orenstein WA, Morag A, Avni A, Handsher R, Green MS, et al. Poliomyelitis outbreak in Israel in 1988: a report with two commentaries. Lancet. 1990;335(8699):1192-5; discussion 1196-8.
  6. Shulman LM, Handsher R, Yang CF, Yang SJ, Manor J, Vonsover A, et al. Resolution of the pathways of poliovirus type 1 transmission during an outbreak. J Clin Microbiol. 2000;38(3):945-52.
  7. Manor Y, Handsher R, Halmut T, Neuman M, Bobrov A, Rudich H, et al. Detection of poliovirus circulation by environmental surveillance in the absence of clinical cases in Israel and the Palestinian authority. J Clin Microbiol. 1999;37(6):1670-5.
  8. Manor Y, Blomqvist S, Sofer D, Alfandari J, Halmut T, Abramovitz B, et al. Advanced environmental surveillance and molecular analyses indicate separate importations rather than endemic circulation of wild type 1 poliovirus in Gaza district in 2002. Appl Environ Microbiol. 2007;73(18):5954-8.
  9. Shulman LM, Manor Y, Sofer D, Mendelson E. Bioterrorism and surveillance for infectious diseases - lessons from poliovirus and enteric virus surveillance J Bioterr Biodef. 2012;S4.
  10. Shulman LM, Manor Y, Handsher R, Delpeyroux F, McDonough MJ, Halmut T, et al. Molecular and antigenic characterization of a highly evolved derivative of the type 2 oral poliovaccine strain isolated from sewage in Israel. J Clin Microbiol. 2000;38(10):3729-34.
  11. Shulman LM, Manor Y, Sofer D, Handsher R, Swartz T, Delpeyroux F, et al. Neurovirulent vaccine-derived polioviruses in sewage from highly immune populations. PLoS One. 2006;1:e69.
  12. World Health Organization (WHO). Poliovirus detected from environmental samples in Egypt. Geneva: WHO; 11 February 2013. Available from:
  13. Anis E, Kopel E, Singer SR, Kaliner E, Moerman L, Moran-Gilad J, et al. Insidious reintroduction of wild poliovirus into Israel, 2013. Euro Surveill. 2013;18(38):pii=20586.
  14. Shulman LM, Hindiyeh M, Muhsen K, Cohen D, Mendelson E, Sofer D. Evaluation of four different systems for extraction of RNA from stool suspensions using MS-2 coliphage as an exogenous control for RT-PCR Inhibition. PLoS One. 2012;7(7):e39455.
  15. Hindiyeh MY, Moran-Gilad J, Manor Y, Ram D, Shulman LM, Sofer D, Mendelson E. Development and validation of a real time quantitative reverse transcription-polymerase chain reaction (qRT-PCR) assay for investigation of wild poliovirus type 1-South Asian (SOAS)strain reintroduced into Israel, 2013 to 2014 . Euro Surveill. 2014;19(7):pii=20710.
  16. Kilpatrick DR, Yang CF, Ching K, Vincent A, Iber J, Campagnoli R, et al. Rapid group-, serotype-, and vaccine strain-specific identification of poliovirus isolates by real-time reverse transcription-PCR using degenerate primers and probes containing deoxyinosine residues. J Clin Microbiol. 2009;47(6):1939-41.
  17. Sofer D, Weil M, Hindiyeh M, Ram D, Shulman LM, Mendelson E. Human Nonpolio enteroviruses. In: Liu D, editor. Molecular detection of human viral pathogens. Boca Raton, FL: CRC Press; 2011. p. 37-52.
  18. Shulman LM, Gavrilin E, Jorba J, Martin J, Burns CC, Manor Y, Moran-Gilad J, Sofer D, Hindiyeh MY, Gamzu R, Mendelson E, Grotto I, for the Genotype - Phenotype Identification (GPI) group. Molecular epidemiology of silent introduction and sustained transmission of wild poliovirus type 1, Israel, 2013. Euro Surveill. 2014;19(7):pii=20709.
  19. Central Bureau of Statistics. 65th Independence Day - more than 8 million residents in the State of Israel. Jerusalem: Central Bureau of Statistics; 14 April 2013. Press release. 097/2013. Available from:
  20. Antona D, Lévêque N, Chomel JJ, Dubrou S, Lévy-Bruhl D, Lina B. Surveillance of enteroviruses in France, 2000-2004.Eur J Clin Microbiol Infect Dis. 2007;26(6):403-12.
  21. van der Sanden SM, Koopmans MP, van der Avoort HG. Detection of human enteroviruses and parechoviruses as part of the national enterovirus surveillance in the Netherlands, 1996-2011. Eur J Clin Microbiol Infect Dis. 2013;32(12):1525-31.
  22. Roberts J, Hobday L, Ibrahim A, Aitken T, Thorley B. Annual report of the Australian National Enterovirus Reference Laboratory 2012. Commun Dis Intell Q Rep. 2013;37(2):E97-E104.
  23. World Population Statistics. France Population 2013. World Population Statistics; 21 Oct 2013. [Accessed 8 Feb 2014]. Available from: