Genetic landscape and macro-evolution of co-circulating Coxsackieviruses A and Vaccine-derived Polioviruses in the Democratic Republic of Congo, 2008-2013.

Enteroviruses (EVs) are among the most common viruses infecting humans worldwide but only a few Non-Polio Enterovirus (NPEV) isolates have been characterized in the Democratic Republic of Congo (DR Congo). Moreover, circulating vaccine-derived polioviruses (PVs) [cVDPVs] isolated during multiple outbreaks in DR Congo from 2004 to 2018 have been characterized so far only by the sequences of their VP1 capsid coding gene. This study was carried to i) investigate the circulation and genetic diversity of NPEV and polio vaccine isolates recovered from healthy children and Acute Flaccid Paralysis (AFP) patients, ii) evaluate the occurrence of genetic recombination among EVs belonging to the Enterovirus C species (including PVs) and iii) identify the virological factors favoring multiple emergences of cVDPVs in DR Congo. The biological material considered in this study included i) a collection of 91 Sabin-like PVs, 54 cVDPVs and 150 NPEVs isolated from AFP patients between 2008 and 2012 in DR Congo and iii) a collection of 330 stool specimens collected from healthy children in 2013 in the Kasai Oriental and Maniema provinces of DR Congo. Studied virus isolates were sequenced in four distinct sub-genomic regions 5'-UTR, VP1, 2CATPase and 3Dpol. Resulting sequences were compared through comparative phylogenetic analyses. Virus isolation showed that 19.1% (63/330) healthy children were infected by EVs including 17.9% (59/330) of NPEVs and 1.2% (4/330) of type 3 Sabin-like PVs. Only one EV-C type, EV-C99 was identified among the NPEV collection from AFP patients whereas 27.5% of the 69 NPEV isolates typed in healthy children belonged to the EV-C species: CV-A13 (13/69), A20 (5/69) and A17 (1/69). Interestingly, 50 of the 54 cVDPVs featured recombinant genomes containing exogenous sequences in at least one of the targeted non-structural regions of their genomes: 5'UTR, 2CATPase and 3Dpol. Some of these non-vaccine sequences of the recombinant cVDPVs were strikingly related to homologous sequences from co-circulating CV-A17 and A20 in the 2CATPase region as well as to those from co-circulating CV-A13, A17 and A20 in the 3Dpol region. This study provided the first evidence uncovering CV-A20 strains as major recombination partners of PVs. High quality AFP surveillance, sensitive environmental surveillance and efficient vaccination activities remain essential to ensure timely detection and efficient response to recombinant cVDPVs outbreaks in DR Congo. Such needs are valid for any epidemiological setting where high frequency and genetic diversity of Coxsackieviruses A13, A17 and A20 provide a conducive viral ecosystem for the emergence of virulent recombinant cVDPVs.

Acute flaccid paralysis surveillance indicators in the Democratic Republic of Congo during 2008-2014.

INTRODUCTION: The last wild poliovirus (WPV) case in Africa was reported in July 2014, thus underscoring the tremendous progress towards polio eradication worldwide. This study aimed to analyze the results of a seven-year surveillance of Acute Flaccid Paralysis (AFP) in the Democratic Republic of Congo (DRC) and to identify potential gaps that need to be addressed. METHODS: Epidemiological and virological data obtained from AFP surveillance among AFP cases less than 15 years from January 2008 to December 2014 in DRC were retrospectively considered and analyzed in this study. RESULTS: Of the 13,749 AFP cases investigated, 58.9% received at least three doses of oral polio vaccine (OPV), 7.3% never received OPV, while the status of 18.3% was unknown. Analysis of surveillance performances showed that all, but two, indicators were below the required WHO-specified targets. Non-polio enterovirus (NPEV) isolation rate was consistently below the minimum requirement at ≥10% and the proportions of stool specimens that reached the laboratory within 72 hours of being sent were always below 15% (WHO target is ≥80%). Virus isolation and differentiation showed that 1.5% of AFP cases were infected by WPVs, 5.5% by Sabin strains, 0.5% by vaccine-derived polioviruses (VDPVs) and 7.2% by NPEVs. CONCLUSION: Our findings indicate that additional efforts are needed to address the timeliness of adequate stool specimens' arrival to the laboratory. It remains essential to maintain high polio vaccine coverage and high AFP surveillance standards to ensure rapid detection and containment of either WPV importation or VDPV re-emergence in DRC.

Phylogeny of imported and reestablished wild polioviruses in theDemocratic Republic of the Congo from 2006 to 2011.

BACKGROUND: The last case of polio associated with wild poliovirus (WPV) indigenous to the Democratic Republic of the Congo (DRC) was reported in 2001, marking a major milestone toward polio eradication in Africa. However, during 2006-2011, outbreaks associated with WPV type 1 (WPV1) were widespread in the DRC, with >250 reported cases. METHODS: WPV1 isolates obtained from patients with acute flaccid paralysis (AFP) were compared by nucleotide sequencing of the VP1 capsid region (906 nucleotides). VP1 sequence relationships among isolates from the DRC and other countries were visualized in phylogenetic trees, and isolates representing distinct lineage groups were mapped. RESULTS: Phylogenetic analysis indicated that WPV1 was imported twice in 2004-2005 and once in approximately 2006 from Uttar Pradesh, India (a major reservoir of endemicity for WPV1 and WPV3 until 2010-2011), into Angola. WPV1 from the first importation spread to the DRC in 2006, sparking a series of outbreaks that continued into 2011. WPV1 from the second importation was widely disseminated in the DRC and spread to the Congo in 2010-2011. VP1 sequence relationships revealed frequent transmission of WPV1 across the borders of Angola, the DRC, and the Congo. Long branches on the phylogenetic tree signaled prolonged gaps in AFP surveillance and a likely underreporting of polio cases. CONCLUSIONS: The reestablishment of widespread and protracted WPV1 transmission in the DRC and Angola following long-range importations highlights the continuing risks of WPV spread until global eradication is achieved, and it further underscores the need for all countries to maintain high levels of poliovirus vaccine coverage and sensitive surveillance to protect their polio-free status.

Emergence of vaccine-derived polioviruses, Democratic Republic of Congo, 2004-2011.

Polioviruses isolated from 70 acute flaccid paralysis patients from the Democratic Republic of Congo (DRC) during 2004-2011 were characterized and found to be vaccine-derived type 2 polioviruses (VDPV2s). Partial genomic sequencing of the isolates revealed nucleotide sequence divergence of up to 3.5% in the viral protein 1 capsid region of the viral genome relative to the Sabin vaccine strain. Genetic analysis identified at least 7 circulating lineages localized to specific geographic regions. Multiple independent events of VDPV2 emergence occurred throughout DRC during this 7-year period. During 2010-2011, VDPV2 circulation in eastern DRC occurred in an area distinct from that of wild poliovirus circulation, whereas VDPV2 circulation in the southwestern part of DRC (in Kasai Occidental) occurred within the larger region of wild poliovirus circulation.

An outbreak of wild poliovirus in the Republic of Congo, 2010-2011.

BACKGROUND: The Republic of Congo has had no cases of wild poliovirus type 1 (WPV1) since 2000. In October 2010, a neurologist noted an abnormal number of cases of acute flaccid paralysis (AFP) among adults, which were later confirmed to be caused by WPV1. METHODS: Those presenting with AFP underwent clinical history, physical examination, and clinical specimen collection to determine if they had polio. AFP cases were classified as laboratory-confirmed, clinical, or nonpolio AFP. Epidemiologic features of the outbreak were analyzed. RESULTS: From 19 September 2010 to 22 January 2011, 445 cases of WPV1 were reported in the Republic of Congo; 390 cases were from Pointe Noire. Overall, 331 cases were among adults; 378 cases were clinically confirmed, and 64 cases were laboratory confirmed. The case-fatality ratio (CFR) was 43%. Epidemiologic characteristics differed among polio cases reported in Pointe Noire and cases reported in the rest of the Republic of Congo, including age distribution and CFR. The outbreak stopped after multiple vaccination rounds with oral poliovirus vaccine, which targeted the entire population. CONCLUSIONS: This outbreak underscores the need to maintain high vaccination coverage to prevent outbreaks, the need to maintain timely high-quality surveillance to rapidly identify and respond to any potential cases before an outbreak escalates, and the need to perform ongoing risk assessments of immunity gaps in polio-free countries.