Viral Diversity Research Articles

Diversity and impact of single-stranded RNA viruses in Czech Heterobasidion populations.

Heterobasidion annosum sensu lato comprises some of the most devastating pathogens of conifers. Exploring virocontrol as a potential strategy to mitigate economic losses caused by these fungi holds promise for the future. In this study, we conducted a comprehensive screening for viruses in 98 H. annosum s.l. specimens from different regions of Czechia aiming to identify viruses inducing hypovirulence. Initial examination for dsRNA presence was followed by RNA-seq analyses using pooled RNA libraries constructed from H. annosum and Heterobasidion parviporum, with diverse bioinformatic pipelines employed for virus discovery. Our study uncovered 25 distinct ssRNA viruses, including two ourmia-like viruses, one mitovirus, one fusarivirus, one tobamo-like virus, one cogu-like virus, one bisegmented narna-like virus and one segment of another narna-like virus, and 17 ambi-like viruses, for which hairpin and hammerhead ribozymes were detected. Coinfections of up to 10 viruses were observed in six Heterobasidion isolates, whereas another six harbored a single virus. Seventy-three percent of the isolates analyzed by RNA-seq were virus-free. These findings show that the virome of Heterobasidion populations in Czechia is highly diverse and differs from that in the boreal region. We further investigated the host effects of certain identified viruses through comparisons of the mycelial growth rate and proteomic analyses and found that certain tested viruses caused growth reductions of up to 22% and significant alterations in the host proteome profile. Their intraspecific transmission rates ranged from 0% to 33%. Further studies are needed to fully understand the biocontrol potential of these viruses in planta.IMPORTANCEHeterobasidion annosum sensu lato is a major pathogen causing significant damage to conifer forests, resulting in substantial economic losses. This study is significant as it explores the potential of using viruses (virocontrol) to combat these fungal pathogens. By identifying and characterizing a diverse array of viruses in H. annosum populations from Czechia, the research opens new avenues for biocontrol strategies. The discovery of 25 distinct ssRNA viruses, some of which reduce fungal growth and alter proteome profiles, suggests that these viruses could be harnessed to mitigate the impact of Heterobasidion. Understanding the interactions between these viruses and their fungal hosts is crucial for developing effective, environmentally friendly methods to protect conifer forests and maintain ecosystem health. This study lays the groundwork for future research on the application of mycoviruses in forest disease management.

Characterization and genetic diversity of grapevine Pinot gris virus in Serbian vineyards

Sixty-five samples of grapevine from commercial vineyards in the Rasina district of Serbia were tested for the presence of grapevine Pinot gris virus (GPGV), using the reverse transcription-polymerase chain reaction. Fourteen samples of the grapevine varieties ‘Red Globe’, ‘Victoria’ and ‘Preobraženje’ were infected with GPGV. All the infected plants showed symptoms of leaf chlorotic mottling, puckering, and deformation, stunting, and reduced yields. The coding regions of the movement and coat protein (MP/CP) and a region of the RNA-dependent RNA polymerase (RdRP) domains of eight virus isolates were sequenced. Phylogenetic analyses of these genomic regions showed high nucleotide similarity among the Serbian GPGV isolates. This study is the first to describe genetic diversity of GPGV isolates in Serbia.

A new lineage nomenclature to aid genomic surveillance of dengue virus.

Dengue virus (DENV) is currently causing epidemics of unprecedented scope in endemic settings and expanding to new geographical areas. It is therefore critical to track this virus using genomic surveillance. However, the complex patterns of viral genomic diversity make it challenging to use the existing genotype classification system. Here, we propose adding 2 sub-genotypic levels of virus classification, named major and minor lineages. These lineages have high thresholds for phylogenetic distance and clade size, rendering them stable between phylogenetic studies. We present assignment tools to show that the proposed lineages are useful for regional, national, and subnational discussions of relevant DENV diversity. Moreover, the proposed lineages are robust to classification using partial genome sequences. We provide a standardized neutral descriptor of DENV diversity with which we can identify and track lineages of potential epidemiological and/or clinical importance. Information about our lineage system, including methods to assign lineages to sequence data and propose new lineages, can be found at: dengue-lineages.org.

Structure-guided assembly of an influenza spike nanobicelle vaccine provides pan H1 intranasal protection.

Development of intranasal vaccines for respiratory viruses has gained popularity. However, currently only a live-attenuated influenza vaccine is FDA-approved for intranasal administration. Here, we focused on influenza virus as it circulates seasonally, has pandemic potential, and has vaccine formulations that present hemagglutinin (HA) in different structural arrangements. These display differences have not been correlated with induction of pan-H1 antibodies or shown to provide intranasal protection. Using electron microscopy, biochemistry and animal studies, we identified HA complexes arranged as lipid discs with multiple trimeric HAs displayed along the perimeter, termed spike nanobicelles (SNB). We utilized a structure-guided approach to synthesize in vitro assembled spiked nanobicelles (IA-SNB) from a classical 1934 H1N1 influenza virus. IA-SNBs elicited pan-H1 antibodies and provided protection against antigenically divergent H1N1 viruses via intranasal immunizations. Viral glycoprotein spikes displayed as SNBs could aid in combating antigenic variation and provide innovative intranasal vaccines to aid universal influenza vaccine development.

Mapping the heterogeneous removal landscape of wastewater virome in effluents of different advanced wastewater treatment systems of swine farm

In advanced wastewater treatment plants on pig farms, meticulous design aims to eliminate intrinsic pollutants such as organic matter, heavy metals, and biological contaminants. In our field survey across Southern China, a notable disparity in wastewater treatment procedures among various farming facilities lies in the utilization of terminal chemical oxidation post-sedimentation tank. However, recent focus in wastewater surveillance has predominantly centered on antibiotic resistance genes, leaving the efficacy of virus removal in different effluent systems largely unexplored. To profile virus composition at the effluent, assess the virus elimination efficiency of chemical oxidation at the effluent end, and the potential environmental driver of virus abundance, we deployed a meta-transcriptomics approach to first determine the total virome in effluent specimens of terminal clean water tank system (CWT) and terminal chemical oxidation system (TCO) in Southern China pig farms, respectively. From these data, 172 viruses were identified, with a median reads per million (RPM) of 27,789 in CWT and 19,982 in TCO. Through the integration of analyses encompassing the co-occurrence patterns within viral communities, the ecology of viral diversity, and a comparative assessment of average variation degrees, we have empirically demonstrated that the procedure of TCO may perturb viral communities and diminish their abundance, particularly impacting RNA viral communities. However, despite the diminished abundance, pathogenic viruses such as PEDV and PRRSV persisted in the effluent following chemical deoxidation at a moderate RPM value, indicating a substantial in situ presence at effluent. Our environmental driver modeling, employing GLM and mantel tests, substantiated the intricate nature of virus community variation within the effluent, influenced heterogeneously by diverse factors. Notably, pond temperature emerged as the foremost determinant, while fishing farming exhibited a positive correlation with virus diversity (p < 0.05). This revelation of the cryptic persistence of virus communities in wastewater effluent expands our understanding of the varied responses of different virus categories to oxidation. Such insights transcend mere virus characterization, offering valuable implications for enhancing biosafety measures in farming practices and informing wastewater-based epidemiological surveillance.

Viral diversity within marine biofilms and interactions with corrosive microbes

In marine environments, a wide variety of microbes like bacteria, and archaea influence on the corrosion of materials. Viruses are widely distributed in biofilms among these microbes and may affect the corrosion process through interactions with key corrosive prokaryotes. However, understanding of the viral communities within biofilms and their interactions with corrosive microbes remains is limited. To improve this knowledge gap, 53 metagenomes were utilized to investigate the diversity of viruses within biofilms on 8 different materials and their interactions with corrosive microbes. Notably, the viruses within biofilms predominantly belonged to Caudoviricetes, and phylogenetic analysis of Caudoviricetes and protein-sharing networks with other environments revealed the presence of numerous novel viral clades in biofilms. The virus‒host linkages revealed a close association between viruses and corrosive microbes in biofilms. This means that viruses may modulate host corrosion-related metabolism through auxiliary metabolic genes. It was observed that the virus could enhance host resistance to metals and antibiotics via horizontal gene transfer. Interestingly, viruses could protect themselves from host antiviral systems through anti-defense systems. This study illustrates the diversity of viruses within biofilms formed on materials and the intricate interactions between viruses and corrosive microbes, showing the potential roles of viruses in corrosive biofilms.

Discovery and biological confirmation of a highly divergent Tacaribe virus in metatranscriptomic data from neotropical bats.

Clade B New World arenaviruses (NWA) include rodent-borne lethal hemorrhagic fever viruses, whereas Tacaribe virus (TCRV) stands out because of its detection in bats and its presumably low zoonotic potential. However, the bat association of TCRV was put into question by lethal experimental neotropical fruit bat infections and rare TCRV detection in bats. Scarce genomic data include near-identical viruses from Caribbean bats and ticks from the US sampled 50 years later. The prototype TCRV isolate used for experimental risk assessments has an extensive passage history in suckling mouse brains. Exploring the true genetic diversity, geographic distribution, and host range of bat-borne NWA is pivotal to assess their zoonotic potential and transmission cycles. We analyzed metatranscriptomic data for evidence of NWA identifying a highly divergent TCRV in bats and confirmed virus detection in original biological materials, supporting the association of TCRV with neotropical bats and warranting investigation of strain-associated TCRV pathogenicity.

Antarctic lake viromes reveal potential virus associated influences on nutrient cycling in ice-covered lakes.

The McMurdo Dry Valleys (MDVs) of Antarctica are a mosaic of extreme habitats which are dominated by microbial life. The MDVs include glacial melt holes, streams, lakes, and soils, which are interconnected through the transfer of energy and flux of inorganic and organic material via wind and hydrology. For the first time, we provide new data on the viral community structure and function in the MDVs through metagenomics of the planktonic and benthic mat communities of Lakes Bonney and Fryxell. Viral taxonomic diversity was compared across lakes and ecological function was investigated by characterizing auxiliary metabolic genes (AMGs) and predicting viral hosts. Our data suggest that viral communities differed between the lakes and among sites: these differences were connected to microbial host communities. AMGs were associated with the potential augmentation of multiple biogeochemical processes in host, most notably with phosphorus acquisition, organic nitrogen acquisition, sulfur oxidation, and photosynthesis. Viral genome abundances containing AMGs differed between the lakes and microbial mats, indicating site specialization. Using procrustes analysis, we also identified significant coupling between viral and bacterial communities (p = 0.001). Finally, host predictions indicate viral host preference among the assembled viromes. Collectively, our data show that: (i) viruses are uniquely distributed through the McMurdo Dry Valley lakes, (ii) their AMGs can contribute to overcoming host nutrient limitation and, (iii) viral and bacterial MDV communities are tightly coupled.

HIV-1 residual risk and pre-treatment drug resistance among blood donors: A sentinel surveillance from Gabon.

Surveillance of HIV-1 pre-treatment drug resistance (PDR) is essential for ensuring the success of first-line antiretroviral therapy (ART). Beside population-based surveys, sentinel surveillance of PDR and circulating HIV-1 clades in specific populations such as blood donors could efficiently inform decision-making on ART program. We therefore sought to ascertain HIV-1 residual infection, the threshold of PDR and viral diversity among recently-diagnosed blood donors in Gabon. A sentinel surveillance was conducted among 381 consenting blood donors at the National Blood Transfusion Center (NBTC) in Gabon from August 3,2020 to August, 31, 2021. In order to determine the residual risk of HIV transmission, viral load and HIV-1 Sanger-sequencing were performed at the Chantal BIYA International Reference Center (CIRCB)-Cameroon on HIV samples previously tested seronegative with ELISA in Gabon. Phylogeny was performed using MEGA X, PDR threshold>10% was considered high and data were analysed using p≤0.05 for statistical significance. Five HIV-negative blood donors had a detectable viral load indicating a high residual risk of HIV transmission. Among the samples successfully sequenced, four participants had major drug resistance mutations (DRMs), giving a threshold of PDR of 25% (4/16). By drug class, major DRMs targeting NNRTI (K103N, E138G), NRTIs (L210W) and PI/r (M46L). The most representative viral clades were CRF02_AG and subtype A1. The genetic diversity of HIV-1 had no significant effect on the residual risk in blood transfusion (CRF02_AG, P = 0.3 and Recombinants, P = 0.5). This sentinel surveillance indicates a high residual risk of HIV-1 transfusion in Gabon, thereby underscoring the need for optimal screening strategy for blood safety. Moreover, HIV-1 transmission goes with high-risk of PDR, suggesting suboptimal efficacy of ART. Nonetheless, the genetic diversity has limited (if any effect) on the residual risk of infection and PDR in blood donors.

SARS-CoV-2 variant replacement constrains vaccine-specific viral diversification.

Coronavirus disease 2019 (COVID-19) vaccine breakthrough infections have been important for all circulating severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variant periods, but the contribution of vaccine-specific SARS-CoV-2 viral diversification to vaccine failure remains unclear. This study analyzed 595 SARS-CoV-2 sequences collected from the Military Health System beneficiaries between December 2020 and April 2022 to investigate the impact of vaccination on viral diversity. By comparing sequences based on the vaccination status of the participant, we found limited evidence indicating that vaccination was associated with increased viral diversity in the SARS-CoV-2 spike, and we show little to no evidence of a substantial sieve effect within major variants; rather, we show that rapid variant replacement constrained intragenotype COVID-19 vaccine strain immune escape. These data suggest that, during past and perhaps future periods of rapid SARS-CoV-2 variant replacement, vaccine-mediated effects were subsumed with other drivers of viral diversity due to the massive scale of infections and vaccinations that occurred in a short time frame. However, our results also highlight some limitations of using sieve analysis methods outside of placebo-controlled clinical trials.

Correction to Distribution and Diversity of Emergent Banana Bunchy Top Virus Infecting Banana and Plantain in Cameroon, Central Africa

Correction to Distribution and Diversity of Emergent Banana Bunchy Top Virus Infecting Banana and Plantain in Cameroon, Central Africa

The Effect of Mask Use on Seasonal Virus Diversity in SARS CoV-2 Negative Patients Treated as Inpatients During the 2021-2022 and 2022-2023 Seasonal Flu Period.

The study aimed to explore the protective effect of mask use against respiratory tract viral agents during the pandemic. The study included patients with a COVID-19 negative test who were hospitalized in the pulmonary disease clinic with the diagnoses of asthma attack, chronic obstructive pulmonary disease (COPD) exacerbation, and pneumonia in two periods: during mandatory mask use (October 2021 - May 2022) and after the mask mandate was lifted (October 2022 - May 2023). Combined nose and throat swab samples taken from the patients were evaluated for viral agents by using the PCR test method. Viral agents isolated from the patients in the two periods were compared based on hospitalization diagnoses and periods. The study enrolled 1,335 patients, 483 female and 852 male. It was found that viral agents significantly increased during the period without a mask mandate compared to the period when the mask mandate was in effect (41.6% vs. 23.4%) (p < 0.001). During the period without mask mandate, influenza A, H1N1, and RSV/AB viruses significantly increased (p = 0.019, p = 0.003, p < 0.001, respectively). Our results indicated that mask use during the pandemic is protective against the transmission of respiratory tract viruses. Thus, it can be concluded that mask use is important not only in the coronavirus pandemic but also especially in influenza and RSV epidemics.

Intrahost evolution leading to distinct lineages in the upper and lower respiratory tracts during SARS-CoV-2 prolonged infection.

Accumulating evidence points to persistent severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections in immunocompromised individuals as a source of novel lineages. While intrahost evolution of the virus in chronically infected patients has previously been reported, existing knowledge is primarily based on samples from the nasopharynx. In this study, we investigate the intrahost evolution and genetic diversity that accumulated during a prolonged SARS-CoV-2 infection with the Omicron BF.7 sublineage, which is estimated to have persisted for >1 year in an immunosuppressed patient. Based on the sequencing of eight samples collected at six time points, we identified 87 intrahost single-nucleotide variants, 2 indels, and a 362-bp deletion. Our analysis revealed distinct viral genotypes in the nasopharyngeal (NP), endotracheal aspirate, and bronchoalveolar lavage samples. This suggests that NP samples may not offer a comprehensive representation of the overall intrahost viral diversity. Our findings not only demonstrate that the Omicron BF.7 sublineage can further diverge from its already exceptionally mutated state but also highlight that patients chronically infected with SARS-CoV-2 can develop genetically specific viral populations across distinct anatomic compartments. This provides novel insights into the intricate nature of viral diversity and evolution dynamics in persistent infections.

Viral plasticity facilitates host diversity in challenging environments

The antagonistic coevolution of microbes and viruses influences fundamentally the diversity of microbial communities. Information on how environmental variables interact with emergent defense-counterdefense strategies and community composition is, however, still scarce. Following biological intuition, diversity should increase with improved growth conditions, which offset evolutionary costs; however, laboratory and regional data suggest that microbial diversity decreases in nutrient-rich conditions. Moreover, global oceanic data show that microbial and viral diversity decline for high latitudes, although the underlying mechanisms are unknown. This article addresses these gaps by introducing an eco-evolutionary model for bacteria-virus antagonistic coevolution. The theory presented here harmonizes the observations above and identifies negative density dependence and viral plasticity (dependence of virus performance on host physiological state) as key drivers: environmental conditions selecting for slow host growth also limit viral performance, facilitating the survival of a diverse host community; host diversity, in turn, enables viral portfolio effects and bet-hedging strategies that sustain viral diversity. From marine microbes to phage therapy against antibiotic-resistant bacteria or cancer cells, the ubiquity of antagonistic coevolution highlights the need to consider eco-evolutionary interactions across a gradient of growth conditions.

Comparative Analysis of HPV16 Variants in the Untranslated Regulatory Region, L1, and E6 Genes among Vaccinated and Unvaccinated Young Women: Assessing Vaccine Efficacy and Viral Diversity.

HPV16 is occasionally detected in vaccinated women who received the bivalent HPV16/18 vaccine, usually at low viral loads. This study explored potential differences in HPV16 variants between vaccinated and unvaccinated women. HPV16-postive viral loads were detected in 1.9% (17/875) and 13% (162/760) of vaccinated and unvaccinated women, respectively, showcasing the vaccine's high efficacy. The L1, E6, and URR regions of HPV16 were sequenced from genital swabs from 16 vaccinated and 25 unvaccinated women in the HAVANA (HPV Among Vaccinated And Non-vaccinated Adolescents) study. The majority of HPV16 variants from vaccinated and unvaccinated women clustered similarly with sub-lineages A1 and A2. Additionally, a separate cluster within lineage A was found, with the variants sharing the L1-located SNP A753G (synonymous) and the URR-located SNP T340C, which did not occur in the other variants. Furthermore, four variants from vaccinated women had relatively long branches, but were not characterized by specific SNPs. The frequency of G712A in the URR was the only SNP observed to be marginally higher among vaccinated women than unvaccinated women. Non-synonymous SNPs T266A in the FG-loop of L1 and L83V in E6 were common among variants from vaccinated and unvaccinated women, but present in similar frequencies. In conclusion, the detection of HPV16 in vaccinated (and unvaccinated) women seemed to be the result of random circulation within this study population.

Massive expansion of the pig gut virome based on global metagenomic mining

The pig gut virome plays a vital role in the gut microbial ecosystem of pigs. However, a comprehensive understanding of their diversity and a reference database for the virome are currently lacking. To address this gap, we established a Pig Virome Database (PVD) that comprised of 5,566,804 viral contig sequences from 4650 publicly available gut metagenomic samples using a pipeline designated “metav”. By clustering sequences, we identified 48,299 viral operational taxonomic units (vOTUs) genomes of at least medium quality, of which 92.83% of which were not found in existing major databases. The majority of vOTUs were identified as Caudoviricetes (72.21%). The PVD database contained a total of 2,362,631 protein-coding genes across the above medium-quality vOTUs genomes that can be used to explore the functional potential of the pig gut virome. These findings highlight the extensive diversity of viruses in the pig gut and provide a pivotal reference dataset for forthcoming research concerning the pig gut virome.

Diverse RNA viruses discovered in multiple seagrass species.

Seagrasses are marine angiosperms that form highly productive and diverse ecosystems. These ecosystems, however, are declining worldwide. Plant-associated microbes affect critical functions like nutrient uptake and pathogen resistance, which has led to an interest in the seagrass microbiome. However, despite their significant role in plant ecology, viruses have only recently garnered attention in seagrass species. In this study, we produced original data and mined publicly available transcriptomes to advance our understanding of RNA viral diversity in Zostera marina, Zostera muelleri, Zostera japonica, and Cymodocea nodosa. In Z. marina, we present evidence for additional Zostera marina amalgavirus 1 and 2 genotypes, and a complete genome for an alphaendornavirus previously evidenced by an RNA-dependent RNA polymerase gene fragment. In Z. muelleri, we present evidence for a second complete alphaendornavirus and near complete furovirus. Both are novel, and, to the best of our knowledge, this marks the first report of a furovirus infection naturally occurring outside of cereal grasses. In Z. japonica, we discovered genome fragments that belong to a novel strain of cucumber mosaic virus, a prolific pathogen that depends largely on aphid vectoring for host-to-host transmission. Lastly, in C. nodosa, we discovered two contigs that belong to a novel virus in the family Betaflexiviridae. These findings expand our knowledge of viral diversity in seagrasses and provide insight into seagrass viral ecology.

New lineages of RNA viruses from clinical isolates of Rhizopus microsporus revealed by fragmented and primer-ligated dsRNA sequencing (FLDS) analysis.

Rhizopus microsporus is a species in the order Mucorales that is known to cause mucormycosis, but it is poorly understood as a host of viruses. Here, we examined 25 clinical strains of R. microsporus for viral infection with a conventional double-stranded RNA (dsRNA) assay using agarose gel electrophoresis (AGE) and the recently established fragmented and primer-ligated dsRNA sequencing (FLDS) protocol. By AGE, five virus-infected strains were detected. Then, full-length genomic sequences of 12 novel RNA viruses were revealed by FLDS, which were related to the families Mitoviridae, Narnaviridae, and Endornaviridae, ill-defined groups of single-stranded RNA (ssRNA) viruses with similarity to the established families Virgaviridae and Phasmaviridae, and the proposed family "Ambiguiviridae." All the characterized viruses, except a potential phasmavirid with a negative-sense RNA genome, had positive-sense RNA genomes. One virus belonged to a previously established species within the family Mitoviridae, whereas the other 11 viruses represented new species or even new genera. These results show that the fungal pathogen R. microsporus harbors diverse RNA viruses and extend our understanding of the diversity of RNA viruses in the fungal order Mucorales, division Mucoromycota. Identifying RNA viruses from clinical isolates of R. microsporus may expand the repertoire of natural therapeutic agents for mucormycosis in the future.IMPORTANCEThe diversity of mycoviruses in fungal hosts in the division Mucoromycota has been underestimated, mainly within the species Rhizopus microsporus. Only five positive-sense RNA genomes had previously been discovered in this species. Because current sequencing methods poorly complete the termini of genomes, we used fragmented and primer-ligated double-stranded RNA sequencing to acquire the full-length genomes. Eleven novel mycoviruses were detected in this study, including the first negative-sense RNA genome reported in R. microsporus. Our findings extend the understanding of the viral diversity in clinical strains of Mucoromycota, may provide insights into the pathogenesis and ecology of this fungus, and may offer therapeutic options.

Viral diversity and co-evolutionary dynamics across the ant phylogeny.

Knowledge of viral biodiversity within insects, particularly within ants, is extremely limited with only a few environmental viruses from invasive ant species identified to date. This study documents and explores the viral communities in ants. We comprehensively profile the metagenomes of a phylogenetically broad group of 35 ant species with varied ecological traits and report the discovery of 3710 novel and unique ant-associated viral genomes. These previously unknown viruses discovered within this study constitute over 95% of all currently described ant viruses, significantly increasing our knowledge of the ant virosphere. The identified RNA and DNA viruses fill gaps in insect-associated viral phylogenies and uncover evolutionary histories characterized by both frequent host switching and co-divergence. Many ants also host diverse bacterial communities, and we discovered that approximately one-third of these new ant-associated viruses are bacteriophages. Two ecological categories, bacterial abundance in the host and habitat degradation are both correlated with ant viral diversity and help to structure viral communities within ants. These data demonstrate that the ant virosphere is remarkably diverse phylogenetically and genomically and provide a substantial foundation for studies in virus ecology and evolution within eukaryotes. We highlight the importance of studying insect-associated viruses in natural ecosystems in order to more thoroughly and effectively understand host-microbe evolutionary dynamics.

A retrospective study revealing complex viral diversity and a substantial burden of HPV infection in SARS-CoV-2 positive individuals, Sierra Leone

BackgroundThe COVID-19 pandemic has underscored the critical role of sequencing technology in disease control and outbreak response. However, resource limitations and challenging environments often impede such efforts in low and middle-income countries. This study aimed to investigate the spectrum of viral co-infections, particularly with human viral pathogens, in SARS-CoV-2 positive individuals in Sierra Leone using metagenomic sequencing, evaluating the feasibility of utilizing this technology for epidemiological and evolutionary surveillance of pathogens related to public health in low-income environments.MethodsWe retrospectively collected and analyzed 98 nasopharyngeal swab specimens from SARS-CoV-2 positive individuals in Sierra Leone. Samples were pre-processed locally and transferred to China via FTA cards for metagenomic sequencing, which was performed using the Novaseq platform. The study focused on the identification of nasopharyngeal viruses co-infecting with SARS-CoV-2, with a deeper analysis of significant human viral pathogens such as HPV.ResultsThe study identified 22 viral taxa from 20 families, including 4 human viruses. Notably, 19.4% of samples showed HPV co-infection with 34 distinct types, predominantly beta and gamma HPVs. Multiple HPV types were found in individual samples, indicating a high complexity of viral co-infections.ConclusionsThe identification of a wide range of co-infecting viruses, particularly multiple HPV genotypes, highlights the complexity of viral interactions and their potential implications for public health. These findings enhance our understanding of viral co-infections and provide valuable insights for public health interventions in Sierra Leone. Further research is needed to explore the clinical significance of these findings and their impact on disease outcomes.