Modeling Poliovirus Transmission in Borno and Yobe, Northeast Nigeria.

Beginning in 2013, multiple local government areas (LGAs) in Borno and Yobe in northeast Nigeria and other parts of the Lake Chad basin experienced a violent insurgency that resulted in substantial numbers of isolated and displaced people. Northeast Nigeria represents the last known reservoir country of wild poliovirus (WPV) transmission in Africa, with detection of paralytic cases caused by serotype 1 WPV in 2016 in Borno and serotype 3 WPV in late 2012. Parts of Borno and Yobe are also problematic areas for transmission of serotype 2 circulating vaccine-derived polioviruses, and they continue to face challenges associated with conflict and inadequate health services in security-compromised areas that limit both immunization and surveillance activities. We model poliovirus transmission of all three serotypes for Borno and Yobe using a deterministic differential equation-based model that includes four subpopulations to account for limitations in access to immunization services and dynamic restrictions in population mixing. We find that accessibility issues and insufficient immunization allow for prolonged poliovirus transmission and potential undetected paralytic cases, although as of the end of 2019, including responsive program activities in the modeling suggest die out of indigenous serotypes 1 and 3 WPVs prior to 2020. Specifically, recent and current efforts to access isolated populations and provide oral poliovirus vaccine continue to reduce the risks of sustained and undetected transmission, although some uncertainty remains. Continued improvement in immunization and surveillance in the isolated subpopulations should minimize these risks. Stochastic modeling can build on this analysis to characterize the implications for undetected transmission and confidence about no circulation.

Progress Toward Poliomyelitis Eradication - Nigeria, January 2018-May 2019.

The number of wild poliovirus (WPV) cases in Nigeria decreased from 1,122 in 2006 to six WPV type 1 (WPV1) in 2014 (1). During August 2014-July 2016, no WPV cases were detected; during August-September 2016, four cases were reported in Borno State. An insurgency in northeastern Nigeria had resulted in 468,800 children aged <5 years deprived of health services in Borno by 2016. Military activities in mid-2016 freed isolated families to travel to camps, where the four WPV1 cases were detected. Oral poliovirus vaccine (OPV) campaigns were intensified during August 2016-December 2017; since October 2016, no WPV has been detected (2). Vaccination activities in insurgent-held areas are conducted by security forces; however, 60,000 unvaccinated children remain in unreached settlements. Since 2018, circulating vaccine-derived poliovirus type 2 (cVDPV2) has emerged and spread from Nigeria to Niger and Cameroon; outbreak responses to date have not interrupted transmission. This report describes progress in Nigeria polio eradication activities during January 2018-May 2019 and updates the previous report (2). Interruption of cVDPV2 transmission in Nigeria will need increased efforts to improve campaign quality and include insurgent-held areas. Progress in surveillance and immunization activities will continue to be reviewed, potentially allowing certification of interruption of WPV transmission in Africa in 2020.

Progress Toward Poliomyelitis Eradication - Nigeria, January-December 2017.

Nearly three decades after the World Health Assembly launched the Global Polio Eradication Initiative in 1988, four of the six World Health Organization (WHO) regions have been certified polio-free (1). Nigeria is one of three countries, including Pakistan and Afghanistan, where wild poliovirus (WPV) transmission has never been interrupted. In September 2015, after >1 year without any reported WPV cases, Nigeria was removed from WHO's list of countries with endemic WPV transmission (2); however, during August and September 2016, four type 1 WPV (WPV1) cases were reported from Borno State, a state in northeastern Nigeria experiencing a violent insurgency (3). The Nigerian government, in collaboration with partners, launched a large-scale coordinated response to the outbreak (3). This report describes progress in polio eradication activities in Nigeria during January-December 2017 and updates previous reports (3-5). No WPV cases have been reported in Nigeria since September 2016; the latest case had onset of paralysis on August 21, 2016 (3). However, polio surveillance has not been feasible in insurgent-controlled areas of Borno State. Implementation of new strategies has helped mitigate the challenges of reaching and vaccinating children living in security-compromised areas, and other strategies are planned. Despite these initiatives, however, approximately 130,000-210,000 (28%-45%) of the estimated 469,000 eligible children living in inaccessible areas in 2016 have not been vaccinated. Sustained efforts to optimize surveillance and improve immunization coverage, especially among children in inaccessible areas, are needed.

Molecular epidemiology of hepatitis B virus infection in Tanzania.

Despite the significant public health problems associated with hepatitis B virus (HBV) in sub-Saharan Africa, many countries in this region do not have systematic HBV surveillance or genetic information on HBV circulating locally. Here, we report on the genetic characterization of 772 HBV strains from Tanzania. Phylogenetic analysis of the S-gene sequences showed prevalence of HBV genotype A (HBV/A, n=671, 86.9 %), followed by genotypes D (HBV/D, n=95, 12.3 %) and E (HBV/E, n=6, 0.8 %). All HBV/A sequences were further classified into subtype A1, while the HBV/D sequences were assigned to a new cluster. Among the Tanzanian sequences, 84 % of HBV/A1 and 94 % of HBV/D were unique. The Tanzanian and global HBV/A1 sequences were compared and were completely intermixed in the phylogenetic tree, with the Tanzanian sequences frequently generating long terminal branches, indicating a long history of HBV/A1 infections in the country. The time to the most recent common ancestor was estimated to be 188 years ago [95 % highest posterior density (HPD): 132 to 265 years] for HBV/A1 and 127 years ago (95 % HPD: 79 to 192 years) for HBV/D. The Bayesian skyline plot showed that the number of transmissions 'exploded' exponentially between 1960-1970 for HBV/A1 and 1970-1990 for HBV/D, with the effective population of HBV/A1 having expanded twice as much as that of HBV/D. The data suggest that Tanzania is at least a part of the geographic origin of the HBV/A1 subtype. A recent increase in the transmission rate and significant HBV genetic diversity should be taken into consideration when devising public health interventions to control HBV infections in Tanzania.

Accurate Genetic Detection of Hepatitis C Virus Transmissions in Outbreak Settings.

Hepatitis C is a major public health problem in the United States and worldwide. Outbreaks of hepatitis C virus (HCV) infections are associated with unsafe injection practices, drug diversion, and other exposures to blood and are difficult to detect and investigate. Here, we developed and validated a simple approach for molecular detection of HCV transmissions in outbreak settings. We obtained sequences from the HCV hypervariable region 1 (HVR1), using end-point limiting-dilution (EPLD) technique, from 127 cases involved in 32 epidemiologically defined HCV outbreaks and 193 individuals with unrelated HCV strains. We compared several types of genetic distances and calculated a threshold, using minimal Hamming distances, that identifies transmission clusters in all tested outbreaks with 100% accuracy. The approach was also validated on sequences obtained using next-generation sequencing from HCV strains recovered from 239 individuals, and findings showed the same accuracy as that for EPLD. On average, the nucleotide diversity of the intrahost population was 6.2 times greater in the source case than in any incident case, allowing the correct detection of transmission direction in 8 outbreaks for which source cases were known. A simple and accurate distance-based approach developed here for detecting HCV transmissions streamlines molecular investigation of outbreaks, thus improving the public health capacity for rapid and effective control of hepatitis C.

Notes from the Field: Hepatitis C Outbreak in a Dialysis Clinic--Tennessee, 2014.

Outbreaks of hepatitis C virus (HCV) infections can occur among hemodialysis patients when recommended infection control practices are not followed (1). On January 30, 2014, a dialysis clinic in Tennessee identified acute HCV in a patient (patient A) during routine screening and reported it to the Tennessee Department of Health. Patient A had enrolled in the dialysis clinic in March 2010 and had annually tested negative for HCV (including a last HCV test on December 19, 2012), until testing positive for HCV antibodies (anti-HCV) on December 18, 2013 (confirmed by a positive HCV nucleic acid amplification test). Patient A reported no behavioral risk factors, but did have multiple health care exposures.

High frequency of active HCV infection among seropositive cases in west Africa and evidence for multiple transmission pathways.

BACKGROUND: Sub-Saharan Africa (SSA) has one of the highest global hepatitis C virus (HCV) prevalence estimates. However, reports that suggest high rates of serologic false positives and low levels of viremia have led to uncertainty regarding the burden of active infection in this region. Additionally, little is known about the predominant transmission risk factors in SSA. METHODS: We prospectively recalled 363 past blood donors (180 who were rapid screen assay [RSA] positive and 183 who were RSA negative at time of donation) to identify the level of active infection and risk factors for infection at a teaching hospital in Kumasi, Ghana. Participants had repeat blood testing and were administered a questionnaire on risk factors. RESULTS: The frequency of HCV active infection ranged from 74.4% to 88% depending on the criteria used to define serologically positive cases. Individuals with active disease had biochemical evidence of liver inflammation and median viral loads of 5.7 log copies/mL. Individuals from the northern and upper regions of Ghana had greater risks of infection compared with participants from other areas. Additional risk factors included traditional circumcision, home birth, tribal scarring, and hepatitis B virus coinfection. CONCLUSIONS: Viremic infection was common among serologically confirmed cases. Attention to testing algorithms is needed in order to define the true HCV burden in SSA. These data also suggest that several transmission modes are likely contributing to the current HCV epidemic in Ghana and that the distribution of these practices may result in substantial regional variation in prevalence.

A re-evaluation of the origin of hepatitis C virus genotype 2 in West Africa.

Hepatitis C virus (HCV) is classified into seven genotypes based on genetic diversity, and most genotypes have been found in Africa. Infections with HCV genotype 2 (HCV2) are most prevalent in West Africa and it was suggested that HCV2 originated in West Africa. To better understand the evolutionary epidemiology of HCV2 in Africa, we examined new NS5B sequences of HCV2 strains obtained from Côte d'Ivoire, Ghana and Nigeria sequenced at the Centers for Disease Control and Prevention with those available from West, North and Central Africa. Bayesian phylogeographic analysis using a discrete trait model showed that Ghana was the most likely geographical region for the origin of HCV2. Spread of HCV2 from Ghana did not appear to be through diffusion to adjacent countries along the coast. Rather, it was transmitted from Ghana to many distant countries in Africa, suggesting that certain routes of geographical dissemination were historically more efficient than mere proximity and that the HCV2 epidemic history in West Africa is extremely complex.

Next-generation sequencing reveals large connected networks of intra-host HCV variants.

BACKGROUND: Next-generation sequencing (NGS) allows for sampling numerous viral variants from infected patients. This provides a novel opportunity to represent and study the mutational landscape of Hepatitis C Virus (HCV) within a single host. RESULTS: Intra-host variants of the HCV E1/E2 region were extensively sampled from 58 chronically infected patients. After NGS error correction, the average number of reads and variants obtained from each sample were 3202 and 464, respectively. The distance between each pair of variants was calculated and networks were created for each patient, where each node is a variant and two nodes are connected by a link if the nucleotide distance between them is 1. The work focused on large components having > 5% of all reads, which in average account for 93.7% of all reads found in a patient. CONCLUSIONS: Most intra-host variants are organized into distinct single-mutation components that are: well separated from each other, represent genetic distances between viral variants, robust to sampling, reproducible and likely seeded during transmission events. Facilitated by NGS, large components offer a novel evolutionary framework for genetic analysis of intra-host viral populations and understanding transmission, immune escape and drug resistance.

Characterization of hepatitis delta virus in sub-Saharan Africa.

Hepatitis D virus (HDV) is a satellite of hepatitis B virus (HBV), and infection with this virus aggravates acute and chronic liver disease. While HBV seroprevalence is very high across sub-Saharan Africa, much less is known about HDV in the region. In this study, almost 2,300 blood serum samples from Burkina Faso (n=1,131), Nigeria (n=974), Chad (n=50), and the Central African Republic (n = 118) were screened for HBV and HDV. Among 743 HBsAg-positive serum samples, 74 were positive for HDV antibodies and/or HDV RNA, with considerable differences in prevalence, ranging from <2% (pregnant women from Burkina Faso) to 50% (liver patients from Central African Republic). HDV seems to be much more common in chronic liver disease patients in the Central African Republic (CAR) than in similar cohorts in Nigeria. In a large nested mother-child cohort in Burkina Faso, the prevalence of HDV antibodies was 10 times higher in the children than in their mothers, despite similar HBsAg prevalences, excluding vertical transmission as an important route of infection. The genotyping of 16 full-length and 8 partial HDV strains revealed clade 1 (17/24) in three of the four countries, while clades 5 (5/24) and 6 (2/24) were, at least in this study, confined to Central Nigeria. On the amino acid level, almost all our clade 1 strains exhibited a serine at position 202 in the hepatitis D antigen, supporting the hypothesis of an ancient African HDV-1 subgroup. Further studies are required to understand the public health significance of the highly varied HDV prevalences in different cohorts and countries in sub-Saharan Africa.

Intra-host diversity and evolution of hepatitis C virus endemic to Côte d'Ivoire.

Hepatitis C virus (HCV) infection presents an important, but underappreciated public health problem in Africa. In Côte d'Ivoire, very little is known about the molecular dynamics of HCV infection. Plasma samples (n = 608) from pregnant women collected in 1995 from Côte d'Ivoire were analyzed in this study. Only 18 specimens (∼3%) were found to be HCV PCR-positive. Phylogenetic analysis of the HCV NS5b sequences showed that the HCV variants belong to genotype 1 (HCV1) (n = 12, 67%) and genotype 2 (HCV2) (n = 6, 33%), with a maximum genetic diversity among HCV variants in each genotype being 20.7% and 24.0%, respectively. Although all HCV2 variants were genetically distant from each other, six HCV1 variants formed two tight sub-clusters belonging to HCV1a and HCV1b. Analysis of molecular variance (AMOVA) showed that the genetic structure of HCV isolates from West Africa with Côte d'Ivoire included were significantly different from Central African strains (P = 0.0001). Examination of intra-host viral populations using next-generation sequencing of the HCV HVR1 showed a significant variation in intra-host genetic diversity among infected individuals, with some strains composed of sub-populations as distant from each other as viral populations from different hosts. Collectively, the results indicate a complex HCV evolution in Côte d'Ivoire, similar to the rest of West Africa, and suggest a unique HCV epidemic history in the country.

Full-length genome characterization and genetic relatedness analysis of hepatitis A virus outbreak strains associated with acute liver failure among children.

Clinical infection by hepatitis A virus (HAV) is generally self-limited but in some cases can progress to liver failure. Here, an HAV outbreak investigation among children with acute liver failure in a highly endemic country is presented. In addition, a sensitive method for HAV whole genome amplification and sequencing suitable for analysis of clinical samples is described. In this setting, two fatal cases attributed to acute liver failure and two asymptomatic cases living in the same household were identified. In a second household, one HAV case was observed with jaundice which resolved spontaneously. Partial molecular characterization showed that both households were infected by HAV subtype IA; however, the infecting strains in the two households were different. The HAV outbreak strains recovered from all cases grouped together within cluster IA1, which contains closely related HAV strains from the United States commonly associated with international travelers. Full-genome HAV sequences obtained from the household with the acute liver failure cases were related (genetic distances ranging from 0.01% to 0.04%), indicating a common-source infection. Interestingly, the strain recovered from the asymptomatic household contact was nearly identical to the strain causing acute liver failure. The whole genome sequence from the case in the second household was distinctly different from the strains associated with acute liver failure. Thus, infection with almost identical HAV strains resulted in drastically different clinical outcomes.

Molecular characterization of hepatitis A virus isolates from Nigeria.

OBJECTIVE: Despite the endemicity of hepatitis A virus (HAV) in Nigeria, genetic information on the HAV genotypes/subgenotypes circulating in the country remains unknown. The objective of this study was to investigate HAV strains using molecular epidemiological and genetic analyses among apparently healthy adult Nigerian subjects. METHODS: Testing for HAV-RNA was performed on 114 serum samples by the reverse transcription-polymerase chain reaction and sequenced with primers encompassing the VP1/P2A junction. RESULTS: Twelve serum samples tested were found to be HAV-RNA positive. Phylogenetic analysis revealed that all 12 HAV isolates were classified as subgenotype IA exhibiting 98.3% nucleotide identity. Interestingly, the Nigerian HAV/IA subgenotype consisted of two distinct genomic sublineages with a unique majority (n = 11) corresponding to strains endemic in Cameroon and the other (n = 1) shows a probable link with European sequences. Predicted conserved amino acid sequences and the few deduced substitution in the VP1/P2A junction might play a role in the development of a novel Nigerian-Cameroon sublineage within the HAV/IA subgenotype and might explain the stability of HAV/IA in this subregion. CONCLUSION: This study reveals the development of a new HAV/IA sublineage in the Nigerian-Cameroon subregion. The presence of a single subgenotype indicates that this HAV strain has been predominantly circulating in Nigeria.

Efficient error correction for next-generation sequencing of viral amplicons.

BACKGROUND: Next-generation sequencing allows the analysis of an unprecedented number of viral sequence variants from infected patients, presenting a novel opportunity for understanding virus evolution, drug resistance and immune escape. However, sequencing in bulk is error prone. Thus, the generated data require error identification and correction. Most error-correction methods to date are not optimized for amplicon analysis and assume that the error rate is randomly distributed. Recent quality assessment of amplicon sequences obtained using 454-sequencing showed that the error rate is strongly linked to the presence and size of homopolymers, position in the sequence and length of the amplicon. All these parameters are strongly sequence specific and should be incorporated into the calibration of error-correction algorithms designed for amplicon sequencing. RESULTS: In this paper, we present two new efficient error correction algorithms optimized for viral amplicons: (i) k-mer-based error correction (KEC) and (ii) empirical frequency threshold (ET). Both were compared to a previously published clustering algorithm (SHORAH), in order to evaluate their relative performance on 24 experimental datasets obtained by 454-sequencing of amplicons with known sequences. All three algorithms show similar accuracy in finding true haplotypes. However, KEC and ET were significantly more efficient than SHORAH in removing false haplotypes and estimating the frequency of true ones. CONCLUSIONS: Both algorithms, KEC and ET, are highly suitable for rapid recovery of error-free haplotypes obtained by 454-sequencing of amplicons from heterogeneous viruses.The implementations of the algorithms and data sets used for their testing are available at: http://alan.cs.gsu.edu/NGS/?q=content/pyrosequencing-error-correction-algorithm.

Epidemic history of hepatitis C virus infection in two remote communities in Nigeria, West Africa.

We investigated the molecular epidemiology and population dynamics of HCV infection among indigenes of two semi-isolated communities in North-Central Nigeria. Despite remoteness and isolation, ~15% of the population had serological or molecular markers of hepatitis C virus (HCV) infection. Phylogenetic analysis of the NS5b sequences obtained from 60 HCV-infected residents showed that HCV variants belonged to genotype 1 (n=51; 85%) and genotype 2 (n=9; 15%). All sequences were unique and intermixed in the phylogenetic tree with HCV sequences from people infected from other West African countries. The high-throughput 454 pyrosequencing of the HCV hypervariable region 1 and an empirical threshold error correction algorithm were used to evaluate intra-host heterogeneity of HCV strains of genotype 1 (n=43) and genotype 2 (n=6) from residents of the communities. Analysis revealed a rare detectable intermixing of HCV intra-host variants among residents. Identification of genetically close HCV variants among all known groups of relatives suggests a common intra-familial HCV transmission in the communities. Applying Bayesian coalescent analysis to the NS5b sequences, the most recent common ancestors for genotype 1 and 2 variants were estimated to have existed 675 and 286 years ago, respectively. Bayesian skyline plots suggest that HCV lineages of both genotypes identified in the Nigerian communities experienced epidemic growth for 200-300 years until the mid-20th century. The data suggest a massive introduction of numerous HCV variants to the communities during the 20th century in the background of a dynamic evolutionary history of the hepatitis C epidemic in Nigeria over the past three centuries.

Genetic analysis of hepatitis A virus variants circulating among children presenting with acute diarrhea in Cameroon.

Molecular investigation was undertaken of circulating hepatitis A virus (HAV) associated with cases of acute diarrhea among children under 5 years of age in Kumba-Cameroon. Reverse transcription PCR, sequencing, and phylogenetic analysis of a 371 bp segment of the VP1/P2A junction of six isolates obtained from stool samples showed the exclusive emergence of genetically related HAV subgenotype IA. All the isolates clustered within a unique lineage exhibiting a 99.5% nucleotide identity suggesting infection from a common source. The Cameroonian HAV isolates did not intermix or cluster with those from other regions of Africa and the rest of the world. Tajima's neutralization tests using the six sequences suggested HAV/IA population expansion (D = -1.37; P = 0.016). This is the first description of indigenous HAV genotypes circulating in Cameroon revealing a community-wide spread and predominance of HAV/1A infection in the Kumba area. These findings stress the need for routine molecular tracking of HAV infection as a contributory cause of acute diarrhea in Cameroonian children.

Prevalence of hepatitis B e antigen in chronic HBV carriers in North-central Nigeria.

Hepatitis B virus (HBV) is an important clinical problem due to its worldwide distribution and potential of adverse sequelae, including hepatocellular carcinoma (HCC). We studied the prevalence of hepatitis B virus e antigen (HBeAg) among individuals determined to be HBV surface antigen-positive (HBsAg+) and analyzed the gender/age category associated with more active HBV infection. A total of 572 HBsAg+ individuals, as determined by a double antibody sandwich ELISA method, participated in the study. They were tested for HbeAg, using a lateral flow chromatographic immunoassay. One hundred and ten individuals were found to be HBeAg-positive giving an overall prevalence of 19.2%. Of these 110 individuals, 20 (18.2%) were females, and 90 (81.8%) were males. Thus, the prevalence of HBeAg appears to be higher in males than in females (p < 0.05). Our data also revealed that the prevalence of HBeAg was higher in patients between the age-group of 0-10 years and 11-20 years and appeared to decrease with increase in age. Taken together, our data show that approximately 1/5 of HBV-infected individuals are HBeAg+, suggesting that the virus is actively replicating and infecting liver-cells thereby ensuring an HBV-transmission pool within the Nigerian population. We suggest strengthening of the childhood HBV vaccination programmes, massive intervention activities, and treatment programmes, especially among young people to reverse the possible devastating effect of HBV infection. The success of these efforts will depend on our resolution to make the elimination of HBV infection a top priority on the public-health agenda as we start the second decade of this new century.

Coordinated evolution among hepatitis C virus genomic sites is coupled to host factors and resistance to interferon.

Machine-learning methods in the form of Bayesian networks (BN), linear projection (LP) and self-organizing tree (SOT) models were used to explore association among polymorphic sites within the HVR1 and NS5a regions of the HCV genome, host demographic factors (ethnicity, gender and age) and response to the combined interferon (IFN) and ribavirin (RBV) therapy. The BN models predicted therapy outcomes, gender and ethnicity with accuracy of 90%, 90% and 88.9%, respectively. The LP and SOT models strongly confirmed associations of the HVR1 and NS5A structures with response to therapy and demographic host factors identified by BN. The data indicate host specificity of HCV evolution and suggest the application of these models to predict outcomes of IFN/RBV therapy.

Outbreak of hepatitis B and hepatitis C virus infections associated with a cardiology clinic, West Virginia, 2012-2014.

OBJECTIVE: To stop transmission of hepatitis B virus (HBV) and hepatitis C virus (HCV) infections in association with myocardial perfusion imaging (MPI) at a cardiology clinic. DESIGN: Outbreak investigation and quasispecies analysis of HCV hypervariable region 1 genome. SETTING: Outpatient cardiology clinic. PATIENTS: Patients undergoing MPI. METHODS: Case patients met definitions for HBV or HCV infection. Cases were identified through surveillance registry cross-matching against clinic records and serological screening. Observations of clinic practices were performed. RESULTS: During 2012-2014, 7 cases of HCV and 4 cases of HBV occurred in 4 distinct clusters among patients at a cardiology clinic. Among 3 case patients with HCV infection who had MPI on June 25, 2014, 2 had 98.48% genetic identity of HCV RNA. Among 4 case patients with HCV infection who had MPI on March 13, 2014, 3 had 96.96%-99.24% molecular identity of HCV RNA. Also, 2 clusters of 2 patients each with HBV infection had MPI on March 7, 2012, and December 4, 2014. Clinic staff reused saline vials for >1 patient. No infection control breaches were identified at the compounding pharmacy that supplied the clinic. Patients seen in clinic through March 27, 2015, were encouraged to seek testing for HBV, HCV, and human immunodeficiency virus. The clinic switched to all single-dose medications and single-use intravenous flushes on March 27, 2015, and no further cases were identified. CONCLUSIONS: This prolonged healthcare-associated outbreak of HBV and HCV was most likely related to breaches in injection safety. Providers should follow injection safety guidelines in all practice settings.