How the Orthodox Features of Orthopoxviruses Led to an Unorthodox Mpox Outbreak: What We've Learned, and What We Still Need to Understand.

Orthopoxviruses are complex, large-genome DNA viruses that have repeatedly confounded expectations in terms of the clinical illness they cause and their patterns of spread. Monkeypox virus (MPXV) was originally characterized during outbreaks among captive primates in the late 1950's. Human disease (mpox) has been observed since the 1970's and inter-human spread has largely been associated with non-sexual, close physical contact in endemic areas of west and central Africa. In May 2022, a focus of Clade IIb MPXV transmission was detected, spreading largely by sexual contact through international networks of gay, bisexual, and other men who have sex with men. Despite decades of preparedness for the potential biothreat risk posed by smallpox, the outbreak grew in both size and geographic scope, testing the strength of smallpox preparedness tools and public health science alike. In this article we consider what was known about mpox prior to the 2022 outbreak, what we have learned about mpox and Clade IIb virus during the outbreak, and what outbreak response actions and continued research are needed to ensure the global public health community is equipped to detect and halt the further spread of this disease threat. We focus on how epidemiologic characterization and investigation together with laboratory studies have advanced our understanding of the transmission and pathogenesis of mpox, and describe what work remains to be done to optimize diagnostics, therapeutics, and vaccines. Persistent health inequities challenge our capacity to fully eliminate circulation of the 2022 outbreak strain of MPXV currently in the United States.

Clinical characterization and placental pathology of mpox infection in hospitalized patients in the Democratic Republic of the Congo.

We describe the results of a prospective observational study of the clinical natural history of human monkeypox (mpox) virus (MPXV) infections at the remote L'Hopital General de Reference de Kole (Kole hospital), the rainforest of the Congo River basin of the Democratic Republic of the Congo (DRC) from March 2007 until August 2011. The research was conducted jointly by the Institute National de Recherche Biomedical (INRB) and the US Army Medical Research Institute of Infectious Diseases (USAMRIID). The Kole hospital was one of the two previous WHO Mpox study sites (1981-1986). The hospital is staffed by a Spanish Order of Catholic Nuns from La Congregation Des Soeurs Missionnaires Du Christ Jesus including two Spanish physicians, who were members of the Order as well, were part of the WHO study on human mpox. Of 244 patients admitted with a clinical diagnosis of MPXV infection, 216 were positive in both the Pan-Orthopox and MPXV specific PCR. The cardinal observations of these 216 patients are summarized in this report. There were three deaths (3/216) among these hospitalized patients; fetal death occurred in 3 of 4 patients who were pregnant at admission, with the placenta of one fetus demonstrating prominent MPXV infection of the chorionic villi. The most common complaints were rash (96.8%), malaise (85.2%), sore throat (78.2%), and lymphadenopathy/adenopathy (57.4%). The most common physical exam findings were mpox rash (99.5%) and lymphadenopathy (98.6%). The single patient without the classic mpox rash had been previously vaccinated against smallpox. Age group of less than 5 years had the highest lesion count. Primary household cases tended to have higher lesion counts than secondary or later same household cases. Of the 216 patients, 200 were tested for IgM & IgG antibodies (Abs) to Orthopoxviruses. All 200 patients had anti-orthopoxvirus IgG Abs; whereas 189/200 were positive for IgM. Patients with hypoalbuminemia had a high risk of severe disease. Patients with fatal disease had higher maximum geometric mean values than survivors for the following variables, respectively: viral DNA in blood (DNAemia); maximum lesion count; day of admission mean AST and ALT.

Design and Optimization of a Monkeypox virus Specific Serological Assay.

Monkeypox virus (MPXV), a member of the Orthopoxvirus (OPXV) genus, is a zoonotic virus, endemic to central and western Africa that can cause smallpox-like symptoms in humans with fatal outcomes in up to 15% of patients. The incidence of MPXV infections in the Democratic Republic of the Congo, where the majority of cases have occurred historically, has been estimated to have increased as much as 20-fold since the end of smallpox vaccination in 1980. Considering the risk global travel carries for future disease outbreaks, accurate epidemiological surveillance of MPXV is warranted as demonstrated by the recent Mpox outbreak, where the majority of cases were occurring in non-endemic areas. Serological differentiation between childhood vaccination and recent infection with MPXV or other OPXVs is difficult due to the high level of conservation within OPXV proteins. Here, a peptide-based serological assay was developed to specifically detect exposure to MPXV. A comparative analysis of immunogenic proteins across human OPXVs identified a large subset of proteins that could potentially be specifically recognized in response to a MPXV infection. Peptides were chosen based upon MPXV sequence specificity and predicted immunogenicity. Peptides individually and combined were screened in an ELISA against serum from well-characterized Mpox outbreaks, vaccinee sera, and smallpox sera collected prior to eradication. One peptide combination was successful with ~86% sensitivity and ~90% specificity. The performance of the assay was assessed against the OPXV IgG ELISA in the context of a serosurvey by retrospectively screening a set of serum specimens from the region in Ghana believed to have harbored the MPXV-infected rodents involved in the 2003 United States outbreak.

Mpox respiratory transmission: the state of the evidence.

The relative contribution of the respiratory route to transmission of mpox (formerly known as monkeypox) is unclear. We review the evidence for respiratory transmission of monkeypox virus (MPXV), examining key works from animal models, human outbreaks and case reports, and environmental studies. Laboratory experiments have initiated MPXV infection in animals via respiratory routes. Some animal-to-animal respiratory transmission has been shown in controlled studies, and environmental sampling studies have detected airborne MPXV. Reports from real-life outbreaks demonstrate that transmission is associated with close contact, and although it is difficult to infer the route of MPXV acquisition in individual case reports, so far respiratory transmission has not been specifically implicated. Based on the available evidence, the likelihood of human-to-human MPXV respiratory transmission appears to be low; however, studies should continue to assess this possibility.

Exportation of Monkeypox Virus From the African Continent.

BACKGROUND: The largest West African monkeypox outbreak began September 2017, in Nigeria. Four individuals traveling from Nigeria to the United Kingdom (n = 2), Israel (n = 1), and Singapore (n = 1) became the first human monkeypox cases exported from Africa, and a related nosocomial transmission event in the United Kingdom became the first confirmed human-to-human monkeypox transmission event outside of Africa. METHODS: Epidemiological and molecular data for exported and Nigerian cases were analyzed jointly to better understand the exportations in the temporal and geographic context of the outbreak. RESULTS: Isolates from all travelers and a Bayelsa case shared a most recent common ancestor and traveled to Bayelsa, Delta, or Rivers states. Genetic variation for this cluster was lower than would be expected from a random sampling of genomes from this outbreak, but data did not support direct links between travelers. CONCLUSIONS: Monophyly of exportation cases and the Bayelsa sample, along with the intermediate levels of genetic variation, suggest a small pool of related isolates is the likely source for the exported infections. This may be the result of the level of genetic variation present in monkeypox isolates circulating within the contiguous region of Bayelsa, Delta, and Rivers states, or another more restricted, yet unidentified source pool.

Educational Approach to Prevent the Burden of Vaccinia Virus Infections in a Bovine Vaccinia Endemic Area in Brazil.

Bovine vaccinia (BV), caused by Vaccinia virus (VACV), is a zoonotic disease characterized by exanthematous lesions on the teats of dairy cows and the hands of milkers, and is an important public health issue in Brazil and South America. BV also results in economic losses to the dairy industry, being a burden to the regions involved in milk production. In the past 20 years, much effort has been made to increase the knowledge regarding BV epidemiology, etiologic agents, and interactions with the hosts and the environment. In the present study, we evaluated milking practices that could be associated with VACV infections in an endemic area in Brazil and proposed an educational tool to help prevent VACV infections. In our survey, 124 individuals (51.7%) from a total of 240 had previously heard of BV, 94 of which knew about it through BV outbreaks. Although most individuals involved in dairy activities (n = 85/91) reported having good hygiene practices, only 29.7% used adequate disinfecting products to clean their hands and 39.5% disinfected cows' teats before and after milking. Furthermore, 46.7% of individuals reported having contact with other farm and domestic animals besides dairy cattle. We also evaluated the presence of IgG and IgM antibodies in the surveyed population. Overall, 6.1% of likely unvaccinated individuals were positive for anti-Orthopoxvirus IgG antibodies, and 1.7% of all individuals were positive for IgM antibodies. Based on our findings, we proposed educational materials which target individuals with permanent residence in rural areas (mainly farmers and milkers), providing an overview and basic information about preventive measures against VACV infections that could enhance BV control and prevention efforts, especially for vulnerable populations located in endemic areas.

Geographic distribution and genetic characterization of poxviruses from human infections in Georgia, 2009-2014.

Anthrax is endemic in Georgia, as are multiple zoonotic poxviruses. Poxvirus-associated infections share some clinical manifestations and exposure risks with anthrax, and so it is important to distinguish between the two. With this in mind, an archived collection of anthrax-negative DNA samples was retrospectively screened for poxviruses, and of the 148 human samples tested, 64 were positive. Sequence analysis confirmed the presence of orf virus, bovine papular stomatitis virus, and pseudocowpox virus. This study provides evidence of previously unrecognized poxvirus infections in Georgia and highlights the benefit of the timely identification of such infections by improving laboratory capacity.

Clinical and Epidemiological Findings from Enhanced Monkeypox Surveillance in Tshuapa Province, Democratic Republic of the Congo During 2011-2015.

BACKGROUND: Monkeypox is a poorly described emerging zoonosis endemic to Central and Western Africa. METHODS: Using surveillance data from Tshuapa Province, Democratic Republic of the Congo during 2011-2015, we evaluated differences in incidence, exposures, and clinical presentation of polymerase chain reaction-confirmed cases by sex and age. RESULTS: We report 1057 confirmed cases. The average annual incidence was 14.1 per 100 000 (95% confidence interval, 13.3-15.0). The incidence was higher in male patients (incidence rate ratio comparing males to females, 1.21; 95% confidence interval, 1.07-1.37), except among those 20-29 years old (0.70; .51-.95). Females aged 20-29 years also reported a high frequency of exposures (26.2%) to people with monkeypox-like symptoms.The highest incidence was among 10-19-year-old males, the cohort reporting the highest proportion of animal exposures (37.5%). The incidence was lower among those presumed to have received smallpox vaccination than among those presumed unvaccinated. No differences were observed by age group in lesion count or lesion severity score. CONCLUSIONS: Monkeypox incidence was twice that reported during 1980-1985, an increase possibly linked to declining immunity provided by smallpox vaccination. The high proportion of cases attributed to human exposures suggests changing exposure patterns. Cases were distributed across age and sex, suggesting frequent exposures that follow sociocultural norms.

A Tale of Two Viruses: Coinfections of Monkeypox and Varicella Zoster Virus in the Democratic Republic of Congo.

Recent enhanced monkeypox (MPX) surveillance in the Democratic Republic of Congo, where MPX is endemic, has uncovered multiple cases of MPX and varicella zoster virus (VZV) coinfections. The purpose of this study was to verify if coinfections occur and to characterize the clinical nature of these cases. Clinical, epidemiological, and laboratory results were used to investigate MPX/VZV coinfections. A coinfection was defined as a patient with at least one Orthopoxvirus/MPX-positive sample and at least one VZV-positive sample within the same disease event. Between September 2009 and April 2014, 134 of the 1,107 (12.1%) suspected MPX cases were confirmed as MPX/VZV coinfections. Coinfections were more likely to report symptoms than VZV-alone cases and less likely than MPX-alone cases. Significantly higher lesion counts were observed for coinfection cases than for VZV-alone but less than MPX-alone cases. Discernible differences in symptom and rash severity were detected for coinfection cases compared with those with MPX or VZV alone. Findings indicate infection with both MPX and VZV could modulate infection severity. Collection of multiple lesion samples allows for the opportunity to detect coinfections. As this program continues, it will be important to continue these procedures to assess variations in the proportion of coinfected cases over time.

Asymptomatic Orthopoxvirus Circulation in Humans in the Wake of a Monkeypox Outbreak among Chimpanzees in Cameroon.

Monkeypox virus is a zoonotic Orthopoxvirus (OPXV) that causes smallpox-like illness in humans. In Cameroon, human monkeypox cases were confirmed in 2018, and outbreaks in captive chimpanzees occurred in 2014 and 2016. We investigated the OPXV serological status among staff at a primate sanctuary (where the 2016 chimpanzee outbreak occurred) and residents from nearby villages, and describe contact with possible monkeypox reservoirs. We focused specifically on Gambian rats (Cricetomys spp.) because they are recognized possible reservoirs and because contact with Gambian rats was common enough to render sufficient statistical power. We collected one 5-mL whole blood specimen from each participant to perform a generic anti-OPXV ELISA test for IgG and IgM antibodies and administered a questionnaire about prior symptoms of monkeypox-like illness and contact with possible reservoirs. Our results showed evidence of OPXV exposures (IgG positive, 6.3%; IgM positive, 1.6%) among some of those too young to have received smallpox vaccination (born after 1980, n = 63). No participants reported prior symptoms consistent with monkeypox. After adjusting for education level, participants who frequently visited the forest were more likely to have recently eaten Gambian rats (OR: 3.36, 95% CI: 1.91-5.92, P < 0.001) and primate sanctuary staff were less likely to have touched or sold Gambian rats (OR: 0.23, 95% CI: 0.19-0.28, P < 0.001). The asymptomatic or undetected circulation of OPXVs in humans in Cameroon is likely, and contact with monkeypox reservoirs is common, raising the need for continued surveillance for human and animal disease.

Do Monkeypox Exposures Vary by Ethnicity? Comparison of Aka and Bantu Suspected Monkeypox Cases.

In 2017, a monkeypox outbreak occurred in Likouala Department, Republic of the Congo. Many of the affected individuals were of Aka ethnicity, hunter-gatherers indigenous to Central Africa who have worse health outcomes in comparison with other forest-dwelling peoples. To test the hypothesis that Aka people have different risk factors for monkeypox, we analyzed questionnaire data for 39 suspected cases, comparing Aka and Bantu groups. Aka people were more likely to touch animal urine/feces, find dead animals in/around the home, eat an animal that was found dead, or to have been scratched or bitten by an animal (P < 0.05, all variables). They were also more likely to visit the forest ≥ once/week, sleep outside, or sleep on the ground (P < 0.001, all variables), providing opportunities for contact with monkeypox reservoirs during the night. The Aka and possibly other vulnerable groups may warrant special attention during educational and health promotion programs.

Genome of Alaskapox Virus, A Novel Orthopoxvirus Isolated from Alaska.

Since the eradication of smallpox, there have been increases in poxvirus infections and the emergence of several novel poxviruses that can infect humans and domestic animals. In 2015, a novel poxvirus was isolated from a resident of Alaska. Diagnostic testing and limited sequence analysis suggested this isolate was a member of the Orthopoxvirus (OPXV) genus but was highly diverged from currently known species, including Akhmeta virus. Here, we present the complete 210,797 bp genome sequence of the Alaska poxvirus isolate, containing 206 predicted open reading frames. Phylogenetic analysis of the conserved central region of the genome suggested the Alaska isolate shares a common ancestor with Old World OPXVs and is diverged from New World OPXVs. We propose this isolate as a member of a new OPXV species, Alaskapox virus (AKPV). The AKPV genome contained host range and virulence genes typical of OPXVs but lacked homologs of C4L and B7R, and the hemagglutinin gene contained a unique 120 amino acid insertion. Seven predicted AKPV proteins were most similar to proteins in non-OPXV Murmansk or NY_014 poxviruses. Genomic analysis revealed evidence suggestive of recombination with Ectromelia virus in two putative regions that contain seven predicted coding sequences, including the A-type inclusion protein.

Varicella in Tshuapa Province, Democratic Republic of Congo, 2009-2014.

OBJECTIVE: To describe varicella cases in Tshuapa Province of the Democratic Republic of the Congo identified during monkeypox surveillance. METHODS: Demographic, clinical and epidemiological data were collected from each suspected monkeypox case 2009-2014. Samples were tested by PCR for both Orthopoxviruses and varicella-zoster virus (VZV); a subset of VZV-positive samples was genotyped. We defined a varicella case as a rash illness with laboratory-confirmed VZV. RESULTS: There were 366 varicella cases were identified; 66% were ≤19 years old. Most patients had non-typical varicella rash with lesions reported as the same size and stage of evolution (86%), deep and profound (91%), on palms of hands and/or soles of feet (86%) and not itchy (49%). Many had non-typical signs and symptoms, such as lymphadenopathy (70%) and sensitivity to light (23%). A higher proportion of persons aged ≥20 years than persons aged ≤19 years had ≥50 lesions (79% vs. 65%, P = 0.007) and were bedridden (15% vs. 9%, P = 0.056). All VZV isolates genotyped from 79 varicella cases were clade 5. During the surveillance period, one possible VZV-related death occurred in a 7-year-old child. CONCLUSIONS: A large proportion of patients presented with non-typical varicella rash and clinical signs and symptoms, highlighting challenges identifying varicella in an area with endemic monkeypox. Continued surveillance and laboratory diagnosis will help in rapid identification and control of both monkeypox and varicella and improve our understanding of varicella epidemiology in Africa.

OBJECTIF: Décrire les cas de varicelle identifiés dans la province de Tshuapa en République Démocratique du Congo (RDC) au cours de la surveillance de la variole du singe (monkeypox). MÉTHODES: Des données démographiques, cliniques et épidémiologiques ont été recueillies pour chaque cas présumé de monkeypox entre 2009 et 2014. Les échantillons ont été testés par PCR pour les orthopoxvirus et le virus varicelle-zona (VZV); un sous-ensemble d'échantillons positifs au VZV a été génotypé. Nous avons défini un cas de varicelle comme une éruption cutanée avec confirmation du VZV en laboratoire. RÉSULTATS: 366 cas de varicelle ont été identifiés; 66% avaient 19 ans ou moins. La plupart des patients présentaient une éruption non typique de varicelle avec des lésions rapportées de la même taille et le même stade d'évolution (86%), profonds (91%), sur la paume des mains et/ou la plante des pieds (86%), sans démangeaisons (49%). Nombre d'entre eux présentaient des signes et des symptômes inhabituels, tels qu'une adénopathie lymphatique (70%) et une sensibilité à la lumière (23%). Une proportion plus élevée de personnes âgées de 20 ans et plus que de personnes âgées de 19 ans et moins avaient 50 lésions ou plus (79% contre 65%, p = 0,007) et étaient alitées (15% contre 9%; p = 0,056). Tous les isolats de VZV génotypés chez 79 cas de varicelle appartenaient au clade 5. Au cours de la période de surveillance, un décès possible lié au VZV est survenu chez un enfant de 7 ans. CONCLUSIONS: Une forte proportion de patients ont présenté une éruption de varicelle ainsi que des signes et symptômes cliniques non typiques, soulignant les difficultés rencontrées pour identifier la varicelle dans une zone endémique pour le monkeypox. Une surveillance continue et des diagnostics de laboratoire aideront à identifier et à contrôler rapidement le monkeypox et la varicelle et à améliorer notre compréhension sur l'épidémiologie de la varicelle en Afrique.

Human Monkeypox in Sierra Leone after 44-Year Absence of Reported Cases.

We note the reemergence of human monkeypox in Sierra Leone following a 44-year absence of reported disease. The persons affected were an 11-month-old boy and, several years later, a 35-year-old man. The reappearance of monkeypox in this country suggests a need for renewed vigilance and awareness of the disease and its manifestations.

Monkeypox re-emergence in Africa: a call to expand the concept and practice of One Health.

INTRODUCTION: Monkeypox is a re-emerging viral zoonosis that occurs naturally in heavily forested regions of West and Central Africa. Inter-human transmission of monkeypox virus, although limited, drives outbreaks, particularly in household and health-care settings. But the available evidence suggests that without repeated zoonotic introductions, human infections would eventually cease to occur. Therefore, interrupting virus transmission from animals to humans is key to combating this disease. Areas covered: Herein we review laboratory and field studies examining the susceptibility of various animal taxa to monkeypox virus infection, and note the competence of various species to serve as reservoirs or transmission hosts. In addition, we discuss early socio-ecologic theories of monkeypox virus transmission in rural settings and review current modes of ecologic investigation - including ecologic niche modeling, and ecologic sampling - in light of their potential to identify specific animal species and features of the environment that are associated with heightened risk for human disease. Expert opinion: The role of disease ecology and scientific research in ongoing disease prevention efforts should be reinforced, particularly for wildlife-associated zoonoses such as monkeypox. Such efforts alongside those aimed at nurturing 'One Health' collaborations may ultimately hold the greatest promise for reducing human infections with this pathogen.

Epidemiologic and Ecologic Investigations of Monkeypox, Likouala Department, Republic of the Congo, 2017.

Monkeypox, caused by a zoonotic orthopoxvirus, is endemic in Central and West Africa. Monkeypox has been sporadically reported in the Republic of the Congo. During March 22-April 5, 2017, we investigated 43 suspected human monkeypox cases. We interviewed suspected case-patients and collected dried blood strips and vesicular and crust specimens (active lesions), which we tested for orthopoxvirus antibodies by ELISA and monkeypox virus and varicella zoster virus DNA by PCR. An ecologic investigation was conducted around Manfouété, and specimens from 105 small mammals were tested for anti-orthopoxvirus antibodies or DNA. Among the suspected human cases, 22 met the confirmed, probable, and possible case definitions. Only 18 patients had available dried blood strips; 100% were IgG positive, and 88.9% (16/18) were IgM positive. Among animals, only specimens from Cricetomys giant pouched rats showed presence of orthopoxvirus antibodies, adding evidence to this species' involvement in the transmission and maintenance of monkeypox virus in nature.

Vaccinating against monkeypox in the Democratic Republic of the Congo.

Healthcare-associated transmission of monkeypox has been observed on multiple occasions in areas where the disease is endemic. Data collected by the US Centers for Disease Control and Prevention (CDC) from an ongoing CDC-supported program of enhanced surveillance in the Tshuapa Province of the Democratic Republic of the Congo, where the annual incidence of human monkeypox is estimated to be 3.5-5/10,000, suggests that there is approximately one healthcare worker infection for every 100 confirmed monkeypox cases. Herein, we describe a study that commenced in February 2017, the intent of which is to evaluate the effectiveness, immunogenicity, and safety of a third-generation smallpox vaccine, IMVAMUNE®, in healthcare personnel at risk of monkeypox virus (MPXV) infection. We describe procedures for documenting exposures to monkeypox virus infection in study participants, and outline lessons learned that may be of relevance for studies of other investigational medical countermeasures in hard to reach, under-resourced populations.

Sounding the alarm: Defining thresholds to trigger a public health response to monkeypox.

Endemic to the Democratic Republic of the Congo (DRC), monkeypox is a zoonotic disease that causes smallpox-like illness in humans. Observed fluctuations in reported cases over time raises questions about when it is appropriate to mount a public health response, and what specific actions should be taken. We evaluated three different thresholds to differentiate between baseline and heightened disease incidence, and propose a novel, tiered algorithm for public health action. Monkeypox surveillance data from Tshuapa Province, 2011-2013, were used to calculate three different statistical thresholds: Cullen, c-sum, and a World Health Organization (WHO) method based on monthly incidence. When the observed cases exceeded the threshold for a given month, that month was considered to be 'aberrant'. For each approach, the number of aberrant months detected was summed by year-each method produced vastly different results. The Cullen approach generated a number of aberrant signals over the period of consideration (9/36 months). The c-sum method was the most sensitive (30/36 months), followed by the WHO method (12/24 months). We conclude that triggering public health action based on signals detected by a single method may be inefficient and overly simplistic for monkeypox. We propose instead a response algorithm that integrates an objective threshold (WHO method) with contextual information about epidemiological and spatiotemporal links between suspected cases to determine whether a response should be operating under i) routine surveillance ii) alert status, or iii) outbreak status. This framework could be modified and adopted by national and zone level health workers in monkeypox-endemic countries. Lastly, we discuss considerations for selecting thresholds for monkeypox outbreaks across gradients of endemicity and public health resources.