Outbreaks of monkeypox (mpox) have historically resulted from zoonotic spillover of clade I monkeypox virus (MPXV) in Central Africa and clade II MPXV in West Africa. In 2022, subclade IIb caused a global epidemic linked to transmission through sexual contact. Here we describe the epidemiological and genomic features of an mpox outbreak in a mining region in eastern Democratic Republic of the Congo, caused by clade I MPXV. Surveillance data collected between September 2023 and January 2024 identified 241 suspected cases. Genomic analysis demonstrates a distinct clade I lineage divergent from previously circulating strains in the Democratic Republic of the Congo. Of the 108 polymerase chain reaction-confirmed mpox cases, the median age of individuals was 22 years, 51.9% were female and 29% were sex workers, suggesting a potential role for sexual transmission. The predominance of APOBEC3-type mutations and the estimated emergence time around mid-September 2023 imply recent sustained human-to-human transmission.

Infection with Zika virus (ZIKV) is associated with human congenital fetal anomalies. To model fetal outcomes in nonhuman primates, we administered Asian-lineage ZIKV subcutaneously to four pregnant rhesus macaques. While non-pregnant animals in a previous study contemporary with the current report clear viremia within 10–12 days, maternal viremia was prolonged in 3 of 4 pregnancies. Fetal head growth velocity in the last month of gestation determined by ultrasound assessment of head circumference was decreased in comparison with biparietal diameter and femur length within each fetus, both within normal range. ZIKV RNA was detected in tissues from all four fetuses at term cesarean section. In all pregnancies, neutrophilic infiltration was present at the maternal-fetal interface (decidua, placenta, fetal membranes), in various fetal tissues, and in fetal retina, choroid, and optic nerve (first trimester infection only). Consistent vertical transmission in this primate model may provide a platform to assess risk factors and test therapeutic interventions for interruption of fetal infection. The results may also suggest that maternal-fetal ZIKV transmission in human pregnancy may be more frequent than currently appreciated. Maternal ZIKV infection in pregnancy is associated with severe fetal anomalies, including microcephaly. It has been shown that infection manifests differently in pregnancy than in the non-pregnant state, with prolonged maternal viremia. ZIKV is spread by mosquitos and through sexual contact and since its first detection in early 2015, has become endemic to the Americas. While much has been learned from studying infected human pregnancies, there are still many questions concerning transmission of ZIKV from mother to fetus. Investigating ZIKV infection in non-human primates could help answer these questions due to similarities in the immune system, and the tissues separating the fetus from the mother during pregnancy. Our study serves to model ZIKV transmission in early and late pregnancy, as well as study the effects of this infection on the fetus and mother at these different times in pregnancy. The data collected provides an important insight on ZIKV in pregnancy where the pregnancies have been monitored throughout the entire infection period until term, and suggests that vertical transmission may be very efficient, although severe fetal outcomes are uncommon.

Congenital Zika virus (ZIKV) infection impacts fetal development and pregnancy outcomes. We infected a pregnant rhesus macaque with a Puerto Rican ZIKV isolate in the first trimester. The pregnancy was complicated by preterm premature rupture of membranes (PPROM), intraamniotic bacterial infection and fetal demise 49 days post infection (gestational day 95). Significant pathology at the maternal-fetal interface included acute chorioamnionitis, placental infarcts, and leukocytoclastic vasculitis of the myometrial radial arteries. ZIKV RNA was disseminated throughout fetal tissues and maternal immune system tissues at necropsy, as assessed by quantitative RT-PCR for viral RNA. Replicating ZIKV was identified in fetal tissues, maternal uterus, and maternal spleen by fluorescent in situ hybridization for viral replication intermediates. Fetal ocular pathology included a choroidal coloboma, suspected anterior segment dysgenesis, and a dysplastic retina. This is the first report of ocular pathology and prolonged viral replication in both maternal and fetal tissues following congenital ZIKV infection in a rhesus macaque. PPROM followed by fetal demise and severe pathology of the visual system have not been described in macaque congenital ZIKV infection previously. While this case of ZIKV infection during pregnancy was complicated by bacterial infection with PPROM, the role of ZIKV on this outcome cannot be precisely defined, and further nonhuman primate studies will determine if increased risk for PPROM or other adverse pregnancy outcomes are associated with congenital ZIKV infection.

Defining the complex dynamics of Zika virus (ZIKV) infection in pregnancy and during transmission between vertebrate hosts and mosquito vectors is critical for a thorough understanding of viral transmission, pathogenesis, immune evasion, and potential reservoir establishment. Within-host viral diversity in ZIKV infection is low, which makes it difficult to evaluate infection dynamics. To overcome this biological hurdle, we constructed a molecularly barcoded ZIKV. This virus stock consists of a “synthetic swarm” whose members are genetically identical except for a run of eight consecutive degenerate codons, which creates approximately 64,000 theoretical nucleotide combinations that all encode the same amino acids. Deep sequencing this region of the ZIKV genome enables counting of individual barcodes to quantify the number and relative proportions of viral lineages present within a host. Here we used these molecularly barcoded ZIKV variants to study the dynamics of ZIKV infection in pregnant and non-pregnant macaques as well as during mosquito infection/transmission. The barcoded virus had no discernible fitness defects in vivo, and the proportions of individual barcoded virus templates remained stable throughout the duration of acute plasma viremia. ZIKV RNA also was detected in maternal plasma from a pregnant animal infected with barcoded virus for 67 days. The complexity of the virus population declined precipitously 8 days following infection of the dam, consistent with the timing of typical resolution of ZIKV in non-pregnant macaques and remained low for the subsequent duration of viremia. Our approach showed that synthetic swarm viruses can be used to probe the composition of ZIKV populations over time in vivo to understand vertical transmission, persistent reservoirs, bottlenecks, and evolutionary dynamics. Understanding the complex dynamics of Zika virus (ZIKV) infection during pregnancy and during transmission to and from vertebrate host and mosquito vector is critical for a thorough understanding of viral transmission, pathogenesis, immune evasion, and reservoir establishment. We sought to develop a virus model system for use in nonhuman primates and mosquitoes that allows for the genetic discrimination of molecularly cloned viruses. This “synthetic swarm” of viruses incorporates a molecular barcode that allows for tracking and monitoring individual viral lineages during infection. Here we infected rhesus macaques with this virus to study the dynamics of ZIKV infection in nonhuman primates as well as during mosquito infection/transmission. We found that the proportions of individual barcoded viruses remained relatively stable during acute infection in pregnant and nonpregnant animals. However, in a pregnant animal, the complexity of the virus population declined precipitously 8 days following infection, consistent with the timing of typical resolution of ZIKV in non-pregnant macaques and remained low for the subsequent duration of viremia.

Concerns have arisen that pre-existing immunity to dengue virus (DENV) could enhance Zika virus (ZIKV) disease, due to the homology between ZIKV and DENV and the observation of antibody-dependent enhancement (ADE) among DENV serotypes. To date, no study has examined the impact of pre-existing DENV immunity on ZIKV pathogenesis during pregnancy in a translational non-human primate model. Here we show that macaques with a prior DENV-2 exposure had a higher burden of ZIKV vRNA in maternal-fetal interface tissues as compared to DENV-naive macaques. However, pre-existing DENV immunity had no detectable impact on ZIKV replication kinetics in maternal plasma, and all pregnancies progressed to term without adverse outcomes or gross fetal abnormalities detectable at delivery. Understanding the risks of ADE to pregnant women worldwide is critical as vaccines against DENV and ZIKV are developed and licensed and as DENV and ZIKV continue to circulate.

Zika virus (ZIKV) and nonhuman primates have been inextricably linked since the virus was first discovered in a sentinel rhesus macaque in Uganda in 1947. Soon after ZIKV was epidemiologically associated with birth defects in Brazil late in 2015, researchers capitalized on the fact that rhesus macaques are commonly used to model viral immunity and pathogenesis, quickly establishing macaque models for ZIKV infection. Within months, the susceptibility of pregnant macaques to experimental ZIKV challenge and ZIKV-associated abnormalities in fetuses was confirmed. This review discusses key unanswered questions in ZIKV immunity and in the pathogenesis of thecongenital Zika virus syndrome. We focus on those questions that can be best addressed in pregnant nonhuman primates and lessons learned from developing macaque models for ZIKV amid an active epidemic.