This month’s May 2025 publication roundup showcases how researchers are using PacBio HiFi sequencing to generate deeper insight into persistent scientific challenges. From uncovering hidden HIV-1 reservoirs, to revealing overlooked pathways of antimicrobial resistance, to refining population-specific references for global pangenome initiatives.
The studies span infectious disease, antimicrobial resistance, population genomics, and aging biology, each illustrating how long-read data can reduce workflow complexity and increase clarity where short reads often fall short.
Featured approaches include targeted enrichment with SMRTcap, phased de novo assembly with methylation and isoform integration, and single-molecule methylation profiling without cell sorting.
Together, they show how HiFi sequencing supports more complete and interpretable datasets for discovery-driven research across diverse fields.
Jump to topic:
Infectious disease | Antimicrobial resistance | Pangenomes | Translational research
Infectious disease
In this preprint, researchers from the University of Louisville and 20 collaborating institutions introduce a new HIV-1 reservoir analysis method using HiFi sequencing. They describe it as “a highly accurate, single-molecule HIV-1 reservoir sequencing and characterization pipeline” that enables “direct and simultaneous resolution of integration site and proviral integrity in an HIV-1 subtype-agnostic manner … with high accuracy and lower costs than comparable methods.”
Key highlights:
- As authors note, “The primary obstacle to achieving a … cure is … a persistent HIV-1 reservoir, arising from HIV-1 DNA integration into target host cell genomes.”
- However, “technical limitations associated with short-read, next-generation sequencing (NGS) have made it challenging to directly associate integration sites with proviral integrity and reactivation potential within the intact proportion of the HIV-1 reservoir.”
- HIV SMRTcap addresses this by “directly and simultaneously [identifying] HIV-1 integration sites, [defining] proviral integrity, and [characterizing] clonal expansion of HIV-1 provirus-containing cells with single molecule resolution.”
- This method is broadly applicable, performing “robustly across the major global subtypes (HIV-1 subtype A, B, C, D and A/D recombinant viruses), and can use both cell- and tissue-derived inputs, including samples from antiretroviral therapy (ART) treated individuals with undetectable viral loads.”
- By reducing the number of required assays and avoiding indirect measurements, “HIV SMRTcap decreases the variety of assays needed and reliance on indirect estimations to define HIV-1 reservoir composition”, thereby “providing critical information needed to inform HIV-1 cure.”
Conclusion:
HIV SMRTcap delivers a streamlined, all-in-one approach to HIV reservoir analysis, replacing multiple assays with a single workflow that cuts time, cost, and complexity. The result is a scalable, clinically relevant alternative to traditional short-read methods. This end-to-end solution enables accurate reservoir characterization, even from limited samples. With high-precision enrichment panels, the Ampli-Fi protocol for low-input samples, and Revio sequencing powered by SPRQ chemistry, HIV SMRTcap brings together everything needed for a cost-effective and high-performance workflow.
Antimicrobial resistance
In this study, researchers from Germany use PacBio technology to show that “more than a decade of bacterial genomic surveillance missed at least one-third of all AMR transmission events due to plasmids.”
Key highlights:
- Over an 18-month period, the team collected “540 multidrug-resistant gram-negative bacterial (MDRB) isolates through routine surveillance” in a tertiary care hospital, aiming to “identify relevant AMR plasmid transmission pathways.” These samples were sequenced using PacBio long-read whole genome sequencing.
- The analysis uncovered 1,539 plasmids in total. According to the authors, “not only the yet unknown plasmid transmissions within healthcare facilities or within the community but also resulted, in addition to the clonal transmissions, in at least a third more transmissions due to AMR plasmids.”
- They demonstrate that “intra-host and patient-to-patient transmissions of AMR plasmids can be identified by combining long-read sequencing with real-time applicable tools during routine molecular surveillance.”
Conclusion:
HiFi sequencing revealed over a third of AMR transmission events missed by short reads, capturing both clonal and plasmid-driven spread across patients, within individuals, and between unrelated bacteria. With the Vega system, HiFi plex prep kit 96, and SMRT Link v3.0, labs get a scalable, cost-effective workflow for real-time AMR surveillance that reduces delays, potentially improves containment, and empowers infection control teams with information that could help support action before outbreaks spread.
Pangenomes
HiFi sequencing is powering the next wave of population-scale genomics, and this month there are three strong examples of this coming to life:
Studies and work to highlight:
- Human Pangenome Reference Consortium (HPRC) released Data Release 2, featuring open-access “high-quality phased genomes from over 200 individuals”—a nearly fivefold increase over Release 1.
- Produced with 60x HiFi (Revio) and includes 5mC methylation calls
- “Additional resources … will soon follow, including … gene annotations that integrate full-length RNA isoform data (PacBio Kinnex) for most samples.”
- Genome diversity and signatures of natural selection in mainland Southeast Asia (led by Kunming, China, with 43 contributing institutions) used Revio-based assemblies to analyze 40 individuals from 23 ethnic groups (17 indigenous, 6 non-indigenous).
- Included pangenome and structural variant calling.
- Revealed a “large number of novel genomic variants,” and provides “a dataset for understanding historical migration patterns, trait and disease associations, and sequences that are impacted by natural selection.”
- Near-complete Middle Eastern genomes refine autozygosity and enhance disease-causing and population-specific variant discovery from the Qatar Genome Program, UW, and NHGRI produced HiFi-based “highly accurate, near-complete and phased genomes” from six family trios (n=18) with neurodevelopmental conditions, representing ancestries from Sudan, Jordan, Syria, Qatar, and Afghanistan.
- Found 2 Mb of new sequence, with 13.8% impacting known genes, and 75 new HLA/KIR alleles.
- Detected strong signals of inbreeding, with ROH covering up to one-third of chromosomes 6 and 12 in one individual.
- Identified 23 de novo and recessive variants as strong candidates for previously unresolved symptoms.
- Showed unique variation compared to existing references, improving mappability and variant calling.
- “These results underscore the value of de novo assembly for disease variant discovery and the need for sampled ME-specific references to better characterize population-relevant variation.”
Conclusion:
HiFi sequencing is driving the era of high-quality human pangenomes, enabling discovery of population-specific variation, supporting rare disease research, and delivering comprehensive insight with less complexity and lower cost. With the Revio system and SPRQ chemistry, PacBio offers one platform for phased genomes, methylation profiling, and full-length RNA isoforms in a single, automated workflow, making it the go-to solution for global, scalable, and equitable genomics.
Translational research
Profiling epigenetic aging at cell-type resolution through long-read sequencing
In this preprint, researchers from Harvard “evaluate cell type-specific aging using long-read sequencing data, without the need for cell sorting”.
Key highlights:
- Authors note that “most studies profile [methylation] at the bulk tissue level, which masks cell type-specific alterations that may follow distinct aging trajectories.”
- In this study, the team leverages cell type-specific methylation pattern to “…map long-read fragments to individual cell types and generate cell type-specific methylation profiles.”
- They applied this approach “to track aging of myeloid cells and lymphocytes [two fundamental hematopoietic cell types that display distinct features during aging] … as well as circulating cell-free DNA, demonstrating robust performance in predicting age.”
- Overall, they found that “This approach provides a novel method for profiling the dynamics of epigenetic aging at cell-type resolution.”
Conclusion:
HiFi sequencing delivers a streamlined, cost-effective solution for cell-type-specific epigenetic aging analysis, without the need for cell sorting or multiple assays. Using a single HiFi workflow, researchers gain both high-accuracy sequence and DNA methylation data from individual DNA fragments, enabling precise cell-of-origin mapping and aging predictions even from challenging samples like cfDNA.
Ready to make discoveries of your own?
The research featured this month highlights how scientists are turning to HiFi sequencing to answer questions that demand more than standard approaches can offer. Whether it’s identifying intact HIV-1 proviruses, tracking the silent spread of AMR plasmids, or assembling high-resolution references for underrepresented populations, these studies show the value of richer, more complete data in moving discovery forward.
With support for phasing, methylation, and full-length RNA isoforms – all in a single workflow – PacBio technology is helping researchers streamline their efforts and focus on what matters: asking better questions, and finding clearer answers.
Stay tuned for next month’s update featuring new research powered by HiFi sequencing.
Ready to see how you can use HiFi sequencing for your next project? Let’s get started.