The study was structured as a descriptive cross-sectional investigation of every patient who met the World Health Organization’s suspected-case criteria for mpox and was reported through the Khyber Pakhtunkhwa Integrated Disease Surveillance and Response System, known as KP-IDSRS. Over the study period, 137 suspected cases were flagged by the surveillance network. Of these, 29 were laboratory-confirmed using real-time polymerase chain reaction, the standard molecular diagnostic for orthopoxvirus infection. Samples with sufficiently low cycle threshold values, at or below 23, indicating high viral loads, were selected for whole-genome sequencing on an Illumina iSeq platform, allowing the researchers to reconstruct near-complete viral genomes for phylogenetic analysis.

The demographic profile of the 29 confirmed cases illustrates that mpox in this setting is not confined to any single population group. The median age of patients was 31 years, with ages ranging from an infant of 0.6 years to an adult of 80 years. Twenty patients, or 68.9 percent, were male, eight, or 27.6 percent, were female, and one patient, 3.4 percent, was transgender. This breadth of age and gender stands in contrast to the epidemiology seen in many high-income countries during the 2022 global outbreak, where cases were heavily concentrated among men who have sex with men, and suggests that in Pakistan the routes of introduction and transmission are more varied, driven largely by travel and household contact rather than by a single transmission network.

Classifying each case by likely origin, the investigators found that 13 infections, or 44.8 percent, were imported, meaning the patients acquired the virus abroad and brought it back to Pakistan. Five cases, 17.2 percent, were secondary transmission cases, representing people infected through contact with an imported case. Six cases, 20.7 percent, were classified as locally acquired with no direct epidemiological link to a known imported infection, and a further five, 17.2 percent, were deemed probable local infections. Taken together, the locally acquired and probable local categories account for more than a third of confirmed cases, providing clear evidence that imported mpox viruses have seeded chains of transmission within the province rather than simply arriving and dying out.

Clinically, the illness observed was characteristic of mpox but generally mild. A vesiculopustular rash, the hallmark dermal manifestation of orthopoxvirus infection in which fluid-filled vesicles evolve into pus-filled pustules before crusting over, was present in all 29 confirmed cases, a 100 percent frequency. Fever was recorded in 82.8 percent of patients, musculoskeletal pain in 37.9 percent, genital or perianal lesions in 34.5 percent, and sore throat in 24.1 percent. The presence of genital and perianal lesions in roughly a third of cases is notable because such lesions can be mistaken for other sexually transmitted infections, delaying diagnosis and isolation. Most patients recovered completely, but the study also records two deaths among the confirmed cases, a somber reminder that even in a largely mild outbreak, mpox carries real mortality risk, particularly for vulnerable patients.

The genomic component of the study is where its significance is most pronounced. Whole-genome sequencing of four representative cases identified both Clade IIb viruses, specifically of lineage A.2.1, and Clade Ib viruses. Mpox virus, an enveloped double-stranded DNA virus of the genus Orthopoxvirus, was long divided into two major clades, with the former Congo Basin or Clade I lineage generally associated with more severe disease and the West African or Clade II lineage with milder illness. The nomenclature has since been refined: Clade IIb denotes the lineage responsible for the sustained human-to-human global spread that began in 2022, while Clade Ib is a recently emerged branch of Clade I that has driven outbreaks in the Democratic Republic of the Congo and neighboring countries and prompted the WHO to declare a second mpox Public Health Emergency of International Concern in 2024. Detecting Clade Ib in a non-endemic country is therefore a marker of how far this newer lineage has traveled.

Phylogenetic analysis of the Pakistani genomes revealed close genetic relationships with sequences reported from within Pakistan itself and from the United States, Nepal, and Thailand. This pattern of clustering across widely separated countries is consistent with the well-established role of international travel in moving mpox virus between continents. Pakistan’s particular vulnerability stems from extensive labor migration to Gulf countries, which creates a constant flow of travelers between South Asia and regions where mpox introductions occur. Each returning traveler represents a potential introduction event, and with limited decentralized diagnostic capacity, infections can go unrecognized until secondary cases appear, exactly the pattern the epidemiological classifications in this study document.

The laboratory work underpinning these findings reflects a coordinated public health infrastructure operating under real constraints. Specimens were processed at the Khyber Pakhtunkhwa Public Health Reference Laboratory at Khyber Medical University in Peshawar, with sequencing support from the National Institutes of Health in Islamabad. The workflow, from standardized surveillance forms through real-time PCR confirmation to genome assembly and phylogenetic comparison against international databases, demonstrates that a provincial reference laboratory in a resource-limited setting can generate genomic data of direct global relevance. The authors acknowledge the WHO and the Khyber Pakhtunkhwa Health Department for supplying diagnostic kits and consumables, and note that the study received no formal external funding, underscoring how much of this surveillance was sustained through institutional commitment rather than dedicated program budgets.

The public health implications of co-circulating clades are substantial. When two genetically distinct lineages circulate simultaneously, each introduction event must be traced independently, and genomic surveillance becomes essential for distinguishing imported cases from established local transmission chains. A locally acquired Clade Ib case, in particular, signals that the newer lineage has achieved at least limited autonomous transmission in the province. The authors argue that this evidence makes a case for strengthened genomic surveillance, decentralized diagnostics that can confirm cases outside a single reference laboratory, and improved access to vaccines and therapeutics. Vaccines such as the Modified Vaccinia Ankara Bavarian Nordic product, which is used for mpox prevention in many countries, remain limited in availability across much of South Asia, and expanding access is a policy challenge that extends well beyond laboratory walls.

For the broader scientific community, the study adds an important data point to the global map of the ongoing mpox emergency. It shows that Clade Ib has reached South Asia and established detectable local transmission there, that Clade IIb lineage A.2.1 continues to circulate in the same region, and that a surveillance system built on WHO case criteria, rapid response teams, and provincial laboratory capacity can detect and characterize this diversity. As international travel continues to connect endemic and non-endemic regions, the Pakistani experience illustrates a pattern likely to recur elsewhere: imported cases are inevitable, but whether they fade out or ignite sustained transmission depends on how quickly diagnostics, contact tracing, vaccination, and genomic sequencing can be brought to bear. The findings from Khyber Pakhtunkhwa suggest that in much of the world, that capacity still needs deliberate investment before the next introduction arrives.

Subject of Research: Emergence and co-circulation of Clade Ib and Clade IIb mpox virus in a non-endemic region of Pakistan

Article Title: Emergence and co-circulation of Clade Ib and Clade IIb Mpox viruses in Khyber Pakhtunkhwa Pakistan, 2024–2026: clinical, genomic, and public health insights from a non-endemic region

Article References: Zia, A., Islam, A., Jamal, Z., Khan, M., Khattak, A. A., Manzoor, M., Umair, M., Muhammad, A., Mehmood Noor, S., Rehman, K., Zohaib, A., Khan, M. A., & Yousafzai, Y. M. (2026). Emergence and co-circulation of Clade Ib and Clade IIb Mpox viruses in Khyber Pakhtunkhwa Pakistan, 2024–2026: clinical, genomic, and public health insights from a non-endemic region. BMC Infectious Diseases. https://doi.org/10.1186/s12879-026-14542-6

Image Credits: AI Generated

DOI: 10.1186/s12879-026-14542-6

Keywords: mpox, Clade Ib, Clade IIb, orthopoxvirus, whole-genome sequencing, genomic surveillance, Pakistan, Khyber Pakhtunkhwa, travel-associated infections, public health, epidemiology, emerging infectious diseases