Background: Sustaining progress toward polio eradication requires not only effective vaccination but also a resilient laboratory network capable of timely poliovirus detection, confirmation, and genetic characterization. In the WHO African region, the African polio laboratory network provides essential virological support for eradication efforts; however, persistent circulating Vaccine-derived Polioviruses (cVDPVs) outbreaks and increasing demands of genomic surveillance continue to challenge laboratory capacity. Despite its critical role, evidence describing the operational performance, adaptive capacity, and evolution of regional polio laboratory network remains limited. This study evaluates the performance and operational experience of the African polio laboratory network in 2025, highlighting contributions to poliovirus detection, genomic surveillance, quality assurance, and outbreak response, while sharing lessons to inform regional public health systems. Methods: A retrospective secondary data analysis was conducted using laboratory surveillance and outbreak databases, supplemented by reviews of programmatic reports. Data were cleaned and harmonized prior to analysis. Descriptive analyses were performed to assess laboratory workload, timeliness indicators, genomic capacity, quality assurance performance, and operational challenges. Qualitative review of programmatic documents was conducted to identify contextual factors influencing performance. Results: In 2025, 77,230 Acute Flaccid Paralysis (AFP) specimens were analyzed across 16 laboratories. Ninety-two percent of specimens met the timeliness target for virus isolation, and 94% met the target for PCR-ITD. Genomic sequencing confirmed 20 cVDPV outbreaks in 10 countries. The establishment of new sequencing laboratories in Uganda and Nigeria reduced turnaround times, and a 2025 expansion plan aimed to build sequencing capacity in 12 additional countries. All laboratories assessed in 2024 for virus isolation, PCR-ITD, and environmental surveillance exceeded quality assurance accreditation thresholds and were rated “Pass”. The pilot implementation of Direct Detection Nanopore Sequencing (DDNS) techniques is expected to further strengthen genomic surveillance. Challenges remain, including stock out of essential supplies, uneven workloads, and infrastructure gaps. Conclusions: The African polio laboratory network has demonstrated sustained capacity, adaptability, and innovation in supporting polio eradication activities in the WHO African region. This study provides evidence on the importance of strengthening laboratory systems, expanding genomic capacity, and maintaining quality assurance mechanisms to support the final phase of polio eradication and broader public health surveillance.

Background: Mumps is a highly infectious viral disease that poses a significant public health concern despite presence of safe and effective vaccine. This study aimed to determine seroprevalence of mumps among patients attending selected hospitals in Coast and Central regions of Kenya, and to evaluate the demographic factors associated with mumps seropositivity. Methodology: This was a cross-sectional study where serum samples collected from patients attending Kiambu, Murang’a and Kilifi County hospitals were quantitatively tested for anti-mumps IgG antibodies using commercial enzyme linked immunosorbent assay (ELISA) kits. Sample size estimation was calculated using Cochran formula. A total of 451 volunteers were enrolled between October 2020 and November 2021. Ethical approval was obtained from Institutional Review Board of Kenya Medical Research Institute Scientific and Ethics Review Unit. Statistical analyses were conducted using Statistical Package for the Social Sciences (SPSS) version 23.0 software. Results: The overall mumps seropositivity rate was 90.5% and geometric mean of mumps IgG titers (GMT) was 401.3 U/ml. Both seroprevalence and GMT were not significantly influenced by the demographic factors of the population under study. Nonetheless, there were trends of higher prevalence related to certain demographic groups. Seroprevalence increased with increasing age from 85% for less than 10-year-olds to 100% among those aged between 60 and 69 years (P=.779). Similarly, females had a higher prevalence at 91.5% compared to 88.6% among males (P=.063). Also, the population from the central region of Kenya had a seroprevalence of 92% compared to 87.3% at the coast region (P=0.249) and rural residents reported 91.3% compared to 88.5% in urban residents (P=.607). Conclusions: The high mumps seroprevalence in this Kenyan population reflects widespread natural infection in the absence of vaccination. Age-related increases in antibody prevalence and geometric mean titres indicate cumulative exposure over time. These findings provide essential pre-vaccine baseline data and support the need for strengthened mumps surveillance and consideration of mumps vaccine inclusion in Kenya’s national immunization program.

ABSTRACT In the Global Polio Laboratory Network (GPLN), poliovirus (PV) screening results from acute flaccid paralysis (AFP) surveillance is based on virus isolation (VI) through cell culture, entailing long turnaround times and the amplification of live poliovirus. An alternative Direct Detection strategy (DD-ITD) for screening viral nucleic acid from stools, bypassing the need for virus culture, has been developed and extensively validated by GPLN partners. A multi-laboratory demonstration project was conceived to field-test the DD-ITD method by GPLN laboratories from the WHO African, Western Pacific, and Eastern Mediterranean regions, where wild serotype 1 or vaccine-derived polioviruses still circulate. Strategically selected laboratories were tasked to simultaneously process stool suspensions with the current gold-standard VI method and the new DD-ITD strategy. Results from 12 laboratories were compiled and analyzed to assess the quality of each RNA extraction and rRT-PCR run. Matched results for both methods of over 10,500 specimens showed an overall method agreement of 91%. All laboratories detected more PV presumptive positive samples with the DD-ITD strategy than with VI, but a large proportion of DD-ITD positive results (72%) were inconclusive or non-typeable, requiring confirmation through sequencing. A total of 298 (2.8%) samples were PV positive using both methods, 828 (7.9%) positive only for DD-ITD, and 62 (0.6%) positive only with VI. The DD-ITD overall performance, quality of results, and agreement between method results varied significantly across participating laboratories. DD-ITD implementation would entail building proficiency in advanced molecular laboratory techniques and data analysis, and increased demand for confirmatory sequencing. IMPORTANCE Surveillance of acute flaccid paralysis (AFP) and sensitive poliovirus detection are key components of the WHO Global Polio Eradication Strategy. This work summarizes the results of a multi-laboratory evaluation designed to field-test the performance and applicability of a molecular Direct Detection strategy (DD-ITD) that does not require amplification of live poliovirus. AFP samples were processed in parallel with both the DD-ITD and the current gold-standard PV detection methodology, based on virus isolation (VI) through cell culture. All participating laboratories detected more PV presumptive positive samples using the DD-ITD strategy than with virus isolation methodology, although a higher proportion of DD-ITD results required confirmatory sequencing. Significant variability among laboratories was observed in the quality of the results and overall DD-ITD performance. Implementing DD-ITD would entail building proficiency in advanced molecular laboratory techniques and strengthening data analysis skills.

Wastewater surveillance has been applied in various parts of the world to monitor the introduction and transmissions of SARS-CoV-2 variants in a population.The knowledge of SARS-CoV-2 variants circulating in a population is critical to COVID-19 management and timing of the application of public health countermeasures.Contrary to the routine clinical surveillance of SARS-CoV-2 where cases from asymptomatic patients are often underreported, wastewater surveillance offers an unbiased tool for monitoring the extent of SARS-CoV-2 transmissions in a community.The present study aimed to characterize SARS-CoV-2 variants that circulated in Nairobi County, Kenya, between April 2021 and August 2021 utilizing wastewater samples.Viral RNA was extracted from wastewater samples, followed by SARS-CoV-2 screening by real-time RT-qPCR before targeted sequencing of the Spike gene.Forty samples were analyzed, of which 50% (n = 20) tested positive for SARS-CoV-2 by real-time RT-qPCR.Of these, 45% (n = 9) were successfully amplified by RT-PCR and sequenced.The majority (78%, 7/9) of the viruses belonged to the Delta (B.1.617.2) lineage of SARS-CoV-2, while a minority (22%) belonged to the Alpha (B.1.1.7)and Alpha-Delta lineages.Phylogenetic analysis of the SARS-COV-2 delta lineage strains revealed scattered clustering of the Kenyan viruses among the global strains included in the analysis, suggesting different introductory routes into the country.On the whole, our re-

Background. Genomic data is key in understanding the spread and evolution of SARS-CoV-2 pandemic and informing the design and evaluation of interventions. However, SARS-CoV-2 genomic data remains scarce across Africa, with no reports yet from the Indian Ocean islands. Methods. We genome sequenced six SARS-CoV-2 positive samples from the first major infection wave in the Union of Comoros in January 2021 and undertook detailed phylogenetic analysis. Results. All the recovered six genomes classified within the 501Y.V2 variant of concern (also known as lineage B.1.351) and appeared to be from 2 sub-clusters with the most recent common ancestor dated 30 th Oct-2020 (95% Credibility Interval: 06 th Sep-2020 to 10 th Dec-2020). Comparison of the Comoros genomes with those of 501Y.V2 variant of concern from other countries deposited into the GISAID database revealed their close association with viruses identified in France and Mayotte (part of the Comoros archipelago and a France, Overseas Department). Conclusions. The recovered genomes, albeit few, confirmed local transmission following probably multiple introductions of the SARS-CoV-2 501Y.V2 variant of concern during the Comoros's first major COVID-19 wave. These findings demonstrate the importance of genomic surveillance and have implications for ongoing control strategies on the islands.

Background. Genomic data is key in understanding the spread and evolution of SARS-CoV-2 pandemic and informing the design and evaluation of interventions. However, SARS-CoV-2 genomic data remains scarce across Africa, with no reports yet from the Indian Ocean islands. Methods. We genome sequenced six SARS-CoV-2 positive samples from the first major infection wave in the Union of Comoros in January 2021 and undertook detailed phylogenetic analysis. Results. All the recovered six genomes classified within the 501Y.V2 variant of concern (also known as lineage B.1.351) and appeared to be from 2 sub-clusters with the most recent common ancestor dated 30th Oct-2020 (95% Credibility Interval: 06th Sep-2020 to 10th Dec-2020). Comparison of the Comoros genomes with those of 501Y.V2 variant of concern from other countries deposited into the GISAID database revealed their close association with viruses identified in France and Mayotte (part of the Comoros archipelago and a France, Overseas Department). Conclusions. The recovered genomes, albeit few, confirmed local transmission following probably multiple introductions of the SARS-CoV-2 501Y.V2 variant of concern during the Comoros’s first major COVID-19 wave. These findings demonstrate the importance of genomic surveillance and have implications for ongoing control strategies on the islands.

Phylogenetic analysis of six SARS-CoV-2 genomes collected from the Comoros islands confirmed local circulation of the 501Y.V2 variant of concern during the country’s first major SARS-CoV-2 wave in January 2021. These findings demonstrate the importance of SARS-CoV-2 genomic surveillance and have implications for ongoing COVID-19 control strategies on the islands. Article summary line Circulation of SARS-CoV-2 501Y.V2 variant of concern in the Comoros Islands during a major COVID-19 infection wave in January 2021 ### Competing Interest Statement The authors have declared no competing interest. ### Funding Statement This work was supported two National Institute for Health Research (NIHR) grants (project references 17/63/82 and 16/136/33) using UK aid from the UK Government to support global health research, the UK Foreign, Commonwealth and Development Office and Wellcome Trust (grant# 220985). The views expressed in this publication are those of the authors and not necessarily those of NIHR, the Department of Health and Social Care, Foreign Commonwealth and Development Office, Wellcome Trust or the UK government. ### Author Declarations I confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained. Yes The details of the IRB/oversight body that provided approval or exemption for the research described are given below: The genomes generated were part of a regional collaborative COVID-19 public health rapid response. This manuscript therefore provides context on the genome data already available on GISAID. The whole genome sequencing study protocol was reviewed and approved by the Scientific and Ethics Review Committee (SERU) that sits at the Kenya Medical Research Institute (KEMRI) headquarters in Nairobi (SERU # 4035). All necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived. Yes I understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance). Yes I have followed all appropriate research reporting guidelines and uploaded the relevant EQUATOR Network research reporting checklist(s) and other pertinent material as supplementary files, if applicable. Yes The six genomes have been deposited in GISAID (Accession numbers EPI\_ISL\_1323658 - EPI\_ISL\_1323663).

Chikungunya virus (family Togavirdae and genus Alphavirus) is an emerging and reemerging virus of public health importance both regionally and globally. In Kenya, about 50-60% of the suspected measles cases remain undiagnosed once measles and rubella is ruled out by immunoglobulin M (IgM) ELISA thus prompted the need to do differential diagnosis on the measles/rubella negative samples. Nothing is known about the role played by chikungunya infection among these suspected measles cases. Febrile rash illness is a common clinical presentation of arboviruses, including chikungunya. In this study, we conducted a serosurvey to explore the possible role of chikungunya infections among suspected measles cases in Kenya that had tested negative for measles and rubella. Sera were tested by commercially available ELISA for the presence of IgG and IgM antibodies against the chikungunya virus. All positive samples for chikungunya by ELISA were confirmed by plaque reduction neutralization test (PRNT), and to rule out cross-reactivity with other alphaviruses a panel of viruses was used, namely o' nyong' nyong, Semliki Forest, and Sindbis viruses. Of the 392 serum samples screened, 0.3% (n = 1) tested positive for IgM antibodies, while 4.6% (n = 18) tested positive for IgG antibodies against the chikungunya virus. PRNT results indicated 2 (11%) chikungunya positives and 7 (38.9%) o' nyong' nyong positives. We recommend awareness among health care providers and improved surveillance for these arboviruses by both serology and molecular testing. Testing for other pathogens should also be done to improve disease detection and diagnosis.

The bag-mediated filtration system (BMFS) was developed to facilitate poliovirus (PV) environmental surveillance, a supplement to acute flaccid paralysis surveillance in PV eradication efforts. From April to September 2015, environmental samples were collected from four sites in Nairobi, Kenya, and processed using two collection/concentration methodologies: BMFS (> 3 L filtered) and grab sample (1 L collected; 0.5 L concentrated) with two-phase separation. BMFS and two-phase samples were analyzed for PV by the standard World Health Organization poliovirus isolation algorithm followed by intratypic differentiation. BMFS samples were also analyzed by a cell culture independent real-time reverse transcription polymerase chain reaction (rRT-PCR) and an alternative cell culture method (integrated cell culture-rRT-PCR with PLC/PRF/5, L20B, and BGM cell lines). Sabin polioviruses were detected in a majority of samples using BMFS (37/42) and two-phase separation (32/42). There was statistically more frequent detection of Sabin-like PV type 3 in samples concentrated with BMFS (22/42) than by two-phase separation (14/42, p = 0.035), possibly due to greater effective volume assayed (870 mL vs. 150 mL). Despite this effective volume assayed, there was no statistical difference in Sabin-like PV type 1 and Sabin-like PV type 2 detection between these methods (9/42 vs. 8/42, p = 0.80 and 27/42 vs. 32/42, p = 0.18, respectively). This study demonstrated that BMFS can be used for PV environmental surveillance and established a feasible study design for future research.

Widespread use of live attenuated (Sabin) oral poliovirus vaccine (OPV) has resulted in marked progress toward global poliomyelitis eradication (1). However, in underimmunized populations, extensive person-to-person transmission of Sabin poliovirus can result in genetic reversion to neurovirulence and paralytic vaccine-derived poliovirus (VDPV) disease (1). This report updates (as of February 26, 2019) previous reports on circulating VDPV type 2 (cVDPV2) outbreaks during 2017-2018 in the Democratic Republic of the Congo (DRC) and in Somalia, which experienced a concurrent cVDPV type 3 (cVDPV3) outbreak* (2,3). In DRC, 42 cases have been reported in four cVDPV2 outbreaks; paralysis onset in the most recent case was October 7, 2018 (2). Challenges to interrupting transmission have included delays in outbreak-response supplementary immunization activities (SIAs) and difficulty reaching children in all areas. In Somalia, cVDPV2 and cVDPV3 were detected in sewage before the detection of paralytic cases (3). Twelve type 2 and type 3 cVDPV cases have been confirmed; the most recent paralysis onset dates were September 2 (cVDPV2) and September 7, 2018 (cVDPV3). The primary challenge to interrupting transmission is the residence of >300,000 children in areas that are inaccessible for vaccination activities. For both countries, longer periods of surveillance are needed before interruption of cVDPV transmission can be inferred.

Enteric virus environmental surveillance via a highly sensitive method is critical, as many enteric viruses have low infectious doses and can persist in the environment for extended periods. This study determined the potential of the novel bag-mediated filtration system (BMFS) to recover human enteric viruses and pepper mild mottle virus (PMMoV) from wastewater and wastewater-impacted surface waters, examined PMMoV use as a fecal contamination indicator in Kenya, and identified potential BMFS process controls. From April 2015 to April 2016, BMFS samples were collected from seven sites in Kenya (n = 59). Enteroviruses and PMMoV were detected in 100% of samples, and human adenovirus, human astrovirus, hepatitis A virus, norovirus GI, norovirus GII, sapovirus, and human rotavirus were detected in the majority of samples. The consistent detection of enteroviruses and PMMoV suggests that these viruses could be used as indicators in similarly fecally contaminated sites and BMFS process controls. As contamination of surface water sources remains a global issue, enteric virus environmental surveillance is necessary. This study demonstrates an effective way to sample large volumes of wastewater and wastewater-impacted surface waters for the detection of multiple enteric viruses simultaneously.

Historically, poliovirus infection has been an important cause of Acute Flaccid Paralysis (AFP) worldwide [1]. Human artistic manifestations in form of painted or carved objects traced acute flaccid paralysis back to Ancient Egypt. The 1500 BC Egyptian Wall stele of a priest called Ruma with a shorter leg and helping himself with a stick. This art shows a withered leg of the priest, and his foot in the equinus position. This deformities characteristic of an attack of paralytic polio (Copenhagen, Ny Carlsberg Museum ÆIN 0134) is considered to be one of the first representations of a polio victim. This piece of art represents a polio victim in the 18th Dynasty (1403-1365 BC) (Figure 1) [2].

Introduction Measles is targeted for elimination in the World Health Organization African Region by the year 2020. In 2011, Kenya was off track in attaining the 2012 pre-elimination goal. We describe the epidemiology of measles in Kenya and assess progress made towards elimination. Methods We reviewed national case-based measles surveillance and immunization data from January 2003 to December 2016. A case was confirmed if serum was positive for anti-measles IgM antibody, was epidemiologically linked to a laboratory-confirmed case or clinically compatible. Data on case-patient demographics, vaccination status, and clinical outcome and measles containing vaccine (MCV) coverage were analyzed. We calculated measles surveillance indicators and incidence, using population estimates for the respective years. Results The coverage of first dose MCV (MCV1) increased from 65% to 86% from 2003-2012, then declined to 75% in 2016. Coverage of second dose MCV (MCV2) remained < 50% since introduction in 2013. During 2003-2016, there were 26,188 suspected measles cases were reported, with 9043(35%) confirmed cases, and 165 deaths (case fatality rate, 1.8%). The non-measles febrile rash illness rate was consistently > 2/100,000 population, and “80% of the sub-national level investigated a case in 11 of the 14 years. National incidence ranged from 4 to 62/million in 2003-2006 and decreased to 3/million in 2016. The age specific incidence ranged from 1 to 364/million population and was highest among children aged < 1 year. Conclusion Kenya has made progress towards measles elimination. However, this progress remains at risk and the recent declines in MCV1 coverage and the low uptake in MCV2 could reverse these gains.

Serosurveys are useful for assessing population susceptibility to vaccine-preventable disease outbreaks. Although at-risk populations in remote areas could benefit from this type of information, they face several logistical barriers to implementation, such as lack of access to centralized laboratories, cold storage, and transport of samples. We describe a potential solution: a compact and portable, field-deployable, point-of-care system relying on digital microfluidics that can rapidly test a small volume of capillary blood for disease-specific antibodies. This system uses inexpensive, inkjet-printed digital microfluidic cartridges together with an integrated instrument to perform enzyme-linked immunosorbent assays (ELISAs). We performed a field validation of the system’s analytical performance at Kakuma refugee camp, a remote setting in northwestern Kenya, where we tested children aged 9 to 59 months and caregivers for measles and rubella immunoglobulin G (IgG). The IgG assays were determined to have sensitivities of 86% [95% confidence interval (CI), 79 to 91% (measles)] and 81% [95% CI, 73 to 88% (rubella)] and specificities of 80% [95% CI, 49 to 94% (measles)] and 91% [95% CI, 76 to 97% (rubella)] (measles, n = 140; rubella, n = 135) compared with reference tests (measles IgG and rubella IgG ELISAs from Siemens Enzygnost) conducted in a centralized laboratory. These results demonstrate a potential role for this point-of-care system in global serological surveillance, particularly in remote areas with limited access to centralized laboratories.

Widespread Transmission of Circulating VaccineDerived Poliovirus Identified by Environmental Surveillance and Immunization Response — Horn of Africa, 2017–2018 Victor A. Eboh, MBBS1; Jeevan K. Makam, MBBS1; Rohit A. Chitale, PhD1; Chukwuma Mbaeyi, DDS1; Jaume Jorba, PhD2; Derek Ehrhardt, MSN, MPH1; Elias Durry, MD1; Tracie Gardner, PhD3; Kamil Mohamed, MD4; Christopher Kamugisha, MScPH5; Peter Borus, DrPH6; Eltayeb Ahmed Elsayed, MD7

Poliomyelitis is an acute viral infection caused by poliovirus and transmitted via the fecal–oral route. The causative agent is one of the three serotypes of poliovirus (serotypes 1, 2, 3) that differ slightly in capsid protein. Prolonged vaccine-related poliovirus shedding in human immunodeficiency virus (HIV) positive individuals has been linked to possible reservoir for reintroduction of polioviruses after eradication. The study therefore aimed at estimating the duration for vaccine-related poliovirus shedding among potentially and HIV-infected persons.

Environmental surveillance of poliovirus (PV) plays an important role in the global program for eradication of wild PV. The bag-mediated filtration system (BMFS) was first developed in 2014 and enhances PV surveillance when compared to the two-phase grab method currently recommended by the World Health Organization (WHO). In this study, the BMFS design was improved and tested for its usability in wastewater and wastewater-impacted surface waters in Nairobi, Kenya. Modifications made to the BMFS included the size, color, and shape of the collection bags, the filter housing used, and the device used to elute the samples from the filters. The modified BMFS concentrated 3-10 L down to 10 mL, which resulted in an effective volume assayed (900-3000 mL) that was 6-20 times greater than the effective volume assayed for samples processed by the WHO algorithm (150 mL). The system developed allows for sampling and in-field virus concentration, followed by transportation of the filter for further analysis with simpler logistics than the current methods. This may ultimately reduce the likelihood of false-negative samples by increasing the effective volume assayed compared to samples processed by the WHO algorithm, making the BMFS a valuable sampling system for wastewater and wastewater-impacted surface waters.