Biology subjects

Bell-Sakyi, L.

Publications and source records attributed to Bell-Sakyi, L..

12 recordsLinked to original sources

bioRxiv2026

A near-complete chromosome from a bacterial pathogen integrated into the genome of its arthropod vector

Lateral gene transfer (LGT) from organellar to eukaryotic genomes is ubiquitous, resulting in the presence of widespread nuclear mitochondrial DNA segments (NUMTs) and/or plastid gene transfers. While LGT from bacterial associates of eukaryotes is generally less common, many arthropod genomes are littered with LGT originating from Wolbachia (order Rickettsiales) - a vertically transmitted, obligate intracellular symbiont of invertebrates. This contrasts with Rickettsia, a related genus including many important human pathogens, which has not been shown to promulgate LGT within its arthropod vectors. Here, we present the 8.6 Gb genome of a cell line derived from A. variegatum (the tropical bont tick), which is the primary vector of Rickettsia africae (agent of African tick-bite fever). In addition to numerous NUMTs and diverse families of transposable elements, an almost-complete chromosome of R. africae origin was identified in the cell line genome, localised to the putative sex chromosome. Sequencing of field-collected A. variegatum confirmed the presence of this large LGT in wild ticks, although the absence of transferred genes from the R. africae plasmid provided a means to differentiate LGT from genuine rickettsial infections. These findings highlight the imperative to consider the possibility of LGT when screening vectors for pathogens by PCR.

genomics↗

bioRxiv2026

Functional divergence of the NSs protein defines interferon antagonism and viral fitness across Bhanja virus lineages.

Bhanja virus (BHAV) is a tick-borne bandavirus (Family Phenuiviridae) with a broad geographic distribution and documented neuroinvasive capacity, yet its molecular biology remains poorly understood. Within the genus Bandavirus, the non-structural protein NSs functions as the primary antagonist of innate immune responses, although mechanistic details for BHAV remain unknown. We established a reverse genetics platform for BHAV by combining virion RNA sequencing with terminal untranslated region mapping, enabling generation of recombinant viruses and systematic investigation of viral determinants. Using this system, we characterized recombinant BHAV replication in mammalian and arthropod cell lines and demonstrated that interferon competence is a critical determinant of viral replication in mammalian cells. Notably, we observed substantial amino acid divergence within the NSs protein across geographically distinct BHAV isolates, despite higher conservation of other viral proteins. To investigate the functional significance of this variation, we generated recombinant viruses expressing heterologous NSs proteins from African and European isolates. Viruses expressing NSs from ibAr2709 or R1819 isolates exhibited enhanced capacity to suppress interferon-beta induction compared to the prototype IG690 strain, correlating with increased viral protein accumulation in interferon-competent cells. Mechanistic studies revealed that BHAV NSs proteins inhibit interferon induction upstream of IRF3, with ibAr2709 and R1819 NSs showing selective inhibition of TBK1 phosphorylation. However, unlike highly pathogenic bandaviruses such as severe fever with thrombocytopenia syndrome virus (SFTSV), BHAV NSs proteins exhibit comparatively weak antagonism of downstream interferon signalling. In vivo studies using interferon alpha/beta receptor-deficient mice demonstrated that NSs sequence variation influences viral replication in splenic tissue without substantially altering disease phenotype. Together, these findings establish BHAV reverse genetics tools for future investigation and reveal how naturally-occurring NSs divergence modulates innate immune antagonism and viral fitness while maintaining an overall attenuated disease phenotype. Author SummaryBhanja virus is a tick-borne virus distributed across Asia, Africa, and Europe that causes neurological disease, yet little is known about how it replicates or evades immunity. We developed the laboratory tools to create recombinant Bhanja viruses and investigate viral functions. We discovered that the virus replicates efficiently in cells lacking interferon responses but is severely restricted in normal immune-competent cells, revealing interferon as a major barrier to viral growth. We observed that Bhanja virus isolates from distinct geographic regions have different versions of a viral protein called NSs that suppresses interferon responses. Isolates from Africa and Europe were better at blocking interferon production than the original Asian strain, correlating with increased viral growth in immune-competent cells. However, unlike related viruses that cause more serious disease such as SFTSV, BHAV NSs proteins only partially antagonise interferon. While this weak antagonism likely contributes to the attenuated phenotype observed in our experimental systems, the natural pathogenesis of Bhanja virus in human hosts and tick vectors remains to be determined. This study provides tools for examining Bhanja virus biology and demonstrates that small viral differences in viral sequence can meaningfully influence evasion of innate immunity and viral fitness without triggering severe disease.

microbiology↗

bioRxiv2026

Transcriptomic profiling of embryo-derived cell lines from the Chagas disease insect vector Rhodnius prolixus

Rhodnius prolixus is a primary insect vector of Trypanosoma cruzi, the causative agent of Chagas disease, a neglected parasitosis endemic in Latin American countries. It has been estimated that Chagas disease affects 7-8 million people worldwide and is responsible for approximately 1000 deaths per year. Genetic and molecular studies in this species remain challenging due to its life cycle and feeding habits, thus hindering the development of new strategies to control their populations and reduce the diffusion of Chagas disease. Recently, two stable cell lines - RPE/LULS53 and RPE/LULS57 - were derived from Rhodnius embryos, which represent promising new tools to investigate the genetics of this insect vector. Here, we describe their gene expression landscapes through transcriptomic approaches. We show that 8,968 expressed genes are shared between the two cell lines, whereas 391 and 1,088 genes are uniquely expressed in RPE/LULS53 and RPE/LULS57, respectively. Although key components of primary developmental, immune and redox signaling pathways are expressed in both cell lines, some genes such as Frizzled-10-a-like and catalase show marked differences in expression. Our results strongly suggest that RPE/LULS53 and RPE/LULS57 likely represent two different cell phenotypes. Consistent with this, gene ontology analysis reveals that RPE/LULS53 is enriched for animal organ morphogenesis and stress response, while RPE/LULS57 for DNA-directed RNA polymerase activity, among others. Despite these differences, both cell lines express comparable levels of transcripts from resident transposable elements, including the highly abundant Mariner and LINE/I elements, as well as horizontally transferred transposons. Our findings shed light on the nature of the RPE/LULS53 and RPE/LULS57 embryo-derived cell lines and provide valuable transcriptomic resources for future genetic and functional studies in Rhodnius and other triatomine insect vectors. Author summaryRhodnius prolixus is a blood-feeding insect and a major vector of Chagas disease, a parasitosis endemic in Latin America and affecting millions of people worldwide. In the absence of effective drugs and vaccines, the control of the insect population represents a promising strategy to reduce the diffusion of the disease. Yet, genetic and functional studies in Rhodnius are extremely challenging due to its feeding habit and life cycle. To overcome these limitations, researchers have previously developed two stable cell lines derived from Rhodnius embryos. In this study, we provide the first characterization of the genes expressed in these cell lines. We found that, while the two cell lines share many expressed genes, each of them also has distinct gene expression patterns pointing to two different cell types with specialized functions. These differences likely affect the way they respond to stress and regulate biological processes. Our findings provide an important resource for researchers studying Rhodnius prolixus and other insect vectors, helping advance our understanding of the genetic and molecular mechanisms that control the insect development and mediate the interactions between insect vectors and the parasites they transmit

genetics↗

bioRxiv2026

Host-derived lipids license proliferation of sterol-auxotrophic tick cells and promote flavivirus output

Sterol molecules play indispensable roles in the biology of cells and organisms. Yet, arthropods, including ticks, lack the capacity for de novo sterol biosynthesis and must rely on exogenous sources. How these organisms sense and utilise exogenous lipids at the cellular level remains poorly understood. Here, we show that host cholesterol and linoleic acid act as essential external signals that license proliferation of Ixodes ricinus tick cells. In lipid-depleted conditions, cells fail to proliferate despite retaining intracellular cholesterol, indicating that exogenous lipids function as regulatory cues rather than merely structural components. We further demonstrate that tick cells exhibit a non-canonical intracellular distribution of cholesterol, with predominant accumulation in lysosome-associated compartments, and that lipid availability induces a coordinated transcriptional program involving lipid uptake and trafficking machinery. Importantly, we find that limiting host lipid availability significantly reduces extracellular RNA output of tick-borne encephalitis virus (TBEV) from infected tick cells, indicating that viral production is affected by the metabolic state of the host cell. Together, our findings establish host-derived lipids as key regulators of cellular state in a sterol-auxotrophic organism and identify lipid availability as a determinant of flavivirus output. This work provides a conceptual framework linking nutrient sensing, cell proliferation, and vector-pathogen interactions.

microbiology↗

bioRxiv2026

Molecular dialogue between Orthonairovirus and tick: RNA-protein interactome of Hazara virus, a BSL2 model of Crimean-Congo Hemorrhagic Fever virus, in Hyalomma cells

Climate change and ecosystem collapse promote geographic expansion of vector-borne diseases, as witnessed by the recent incursions into Spain of the virus responsible for Crimean-Congo hemorrhagic fever (CCHFV). CCHFV is maintained in a tick-vertebrate cycle, principally involving ticks of the genus Hyalomma. Faced with the spread of Hyalomma ticks, and therefore the threat of a natural introduction of CCHFV into Western Europe, appropriate surveillance tools and control measures need to be implemented. It is both within and by the tick that CCHFV is maintained and spread in the environment. Despite prolonged portage of the virus, the tick is not overtly affected by CHFV infection. One of the prerequisites in conceiving control strategies is to understand the molecular mechanisms that intimately link the virus to its arthropod host. Despite the central role of the tick in the biology of CCHFV, these mechanisms are ill-defined, owing in part to the constraints associated with handling CCHFV-infected ticks in biosafety level 4 containment. In this study, we established the network of interactions between the S segment of the RNA genome Hazara virus (HAZV), a BSL-2 model of CCHFV, and Hyalomma proteins using ChIRP-MS technique. We identified 166 tick proteins, 21 of which have been described as RNA-binding proteins. Gene ontology and pathway enrichment analyses revealed that the S segment RNA interacts predominantly with mitochondrial proteins that belong to various mitochondrial metabolic pathways.

microbiology↗

bioRxiv2025

Morphological and molecular characterisation of Orientia tsutsugamushi grown in tick cells

Orientia tsutsugamushi (Ot), the causative agent of scrub typhus, is an obligate intracellular bacterium naturally maintained in Leptotrombidium mites, yet its interactions within arthropod hosts remain poorly understood. Here, we developed two tick cell lines, Ixodes scapularis ISE6 and Rhipicephalus microplus BME/CTVM23, as arthropod models to investigate the intracellular lifecycle of Ot strains TA686 and Karp. Both strains efficiently infected and replicated within tick cells, with ISE6 supporting more robust growth. Electron microscopy images revealed that Ot maintains its characteristic cytoplasmic, non-vacuolar location in tick cells and exits infected cells by budding off the surface in a membrane-encased structure. Time-course immunofluorescence imaging demonstrated progressive intracellular replication and dynamic expression of Ot outer membrane autotransporters ScaA and ScaC, with ScaC enriched early in infection and ScaA at later stages. Metabolic labelling using a clickable methionine analog L-homopropargylglycine showed that high ScaA abundance correlated with reduced translational activity, suggesting a link between ScaA abundance and late-stage or extracellular-like developmental states. The subcellular location of Ot in tick cells differs from the characteristic dynein-driven perinuclear clustering observed in mammalian cells, and was not sensitive to disruption of microtubules, indicating a distinct mode of distribution. Together, these findings identify tick cell lines as tractable and biologically relevant arthropod models for studying Ot and dissecting host-microbe interactions.

microbiology↗

bioRxiv2025

SFTSV NSs protein is a novel tick antiviral RNAi response suppressor

Severe fever with thrombocytopenia syndrome virus (SFTSV) is an emerging tick-borne phenuivirus causing high mortality in humans. While the non-structural protein NSs is dispensable for replication in interferon-deficient mammalian cells, we demonstrate for the first time that NSs is essential for viral replication in tick cells. SFTSV infection triggers canonical Dicer-2-mediated antiviral RNA interference (RNAi) in tick cells, producing virus-derived small interfering RNAs (siRNAs) that target viral transcripts for degradation. We show that NSs functions as a viral suppressor of RNAi by selectively engaging and depleting single-stranded RNAs derived from 22-nucleotide siRNAs, likely limiting their incorporation into RNA-induced silencing complexes (RISC). Complementation with a heterologous RNAi suppressor (p19 protein) partially rescues replication of NSs-deficient virus, validating the RNAi suppressive function of NSs. These findings reveal that successful tick-borne viral replication requires host-specific immune evasion strategies and establish NSs-mediated RNAi suppression as essential for SFTSV persistence in arthropod vectors. Significance StatementThis study reveals a critical requirement for SFTSV infection that differs between hosts. While the viral NSs protein is dispensable in interferon deficient mammalian cells, it is essential for replication in tick cells, the natural vectors of the virus. We show that NSs functions as a viral RNA silencing suppressor by associating with virus derived small RNAs and limiting their availability to the tick antiviral RNA interference machinery. This provides mechanistic insight into how tick-borne viruses evade arthropod immune defences through engagement of functional siRNAs rather than broad inhibition of RNA interference pathways. Together, these findings demonstrate that SFTSV employs distinct immune evasion strategies in different hosts and identify NSs as a key determinant of vector cell infection.

microbiology↗

bioRxiv2024

Uukuniemi virus infection causes a pervasive remodelling of the RNA-binding proteome in tick cell cultures

Cellular RNA-binding proteins (RBPs) are pivotal for the viral lifecycle, mediating key host-virus interactions that promote or repress virus infection. While these interactions have been largely studied in the vertebrate host, no comprehensive analyses of protein-RNA interactions occurring in cells of arbovirus vectors, in particular ticks, have been performed to date. Here we systematically identified the responses of the RNA-binding proteome (RBPome) to infection with a prototype bunyavirus (Uukuniemi virus; UUKV) in tick cells and discovered changes in RNA-binding activity for 283 proteins. In an orthogonal approach, we analysed the composition of the viral ribonucleoprotein by immunoprecipitation of UUKV nucleocapsid protein (N) in infected cells. We found many tick RBPs that are regulated by UUKV infection and associate with viral nucleocapsid protein complexes. We confirmed experimentally that these RBPs impact UUKV infection. This includes the tick homolog of topoisomerase 3B (TOP3B), a protein able to manipulate the topology of RNA, which showed an effect on viral particle production. Our data thus reveals the first protein-RNA interaction map for infected tick cells. Research highlightsO_LIUUKV RNAs interact with nearly three hundred tick cell RBPs. C_LIO_LIDemonstrated an enrichment of N protein interactors within the upregulated RIC data suggesting a direct involvement in viral RNA metabolism and translation. C_LIO_LIDeveloped a robust methodology to silence gene expression in tick cell cultures. C_LIO_LIThe TOP3B complex facilitates efficient packaging of UUKV virions. C_LI

microbiology↗

bioRxiv2023

The phenuivirus Toscana virus makes an atypical use of vacuolar acidity to enter host cells

Toscana virus is a major cause of arboviral disease in humans in the Mediterranean basin during summer. However, early virus-host cell interactions and entry mechanisms remain poorly characterized. Investigating iPSC-derived human neurons and cell lines, we found that virus binding to the cell surface was specific but inefficient, and 50% of bound virions were endocytosed within 10 min. Virions entered Rab5a+ early endosomes and, subsequently, Rab7a+ and LAMP-1+ late endosomal compartments. Penetration required intact late endosomes and occurred within 30 min following internalization. Virus entry relied on vacuolar acidification, with an optimal pH for viral membrane fusion at pH 5.5. The pH threshold increased to 5.8 with longer pre-exposure of virions to the slightly acidic pH in early endosomes. Strikingly, the particles remained infectious after entering late endosomes with a pH below the fusion threshold. Overall, our study establishes Toscana virus as a late-penetrating virus and reveals an atypical use of vacuolar acidity by this virus to enter host cells.

microbiology↗

bioRxiv2023

Metagenomics of culture isolates and insect tissue illuminate the evolution of Wolbachia, Rickettsia and Bartonella symbionts in Ctenocephalides spp. fleas

While fleas are often perceived simply as a biting nuisance and cause of allergic dermatitis, they represent important disease vectors worldwide, especially for bacterial zoonoses such as plague (transmitted by rodent fleas) and some of the rickettsioses and bartonelloses. The cosmopolitan cat (Ctenocephalides felis) and dog (C. canis) fleas, as well as C. orientis (restricted to tropical and subtropical Asia), breed in human dwellings and are vectors of cat-scratch fever (caused by Bartonella spp.) and Rickettsia spp. of the so-called "transitional group". The latter includes R. felis (agent of flea-borne spotted fever) and R. asembonensis, an emerging pathogen. The relatively depauperate flea microbiome can also contain arthropod-specific endosymbionts, including a diverse range of Wolbachia strains. Here, we present circularized genome assemblies for two C. orientis-associated pathogens (Bartonella clarridgeiae and R. asembonensis) from Malaysia, a novel Wolbachia strain (wCori), and the C. orientis mitochondrion; all obtained by direct metagenomic sequencing of flea tissues. Moreover, we isolated two Wolbachia strains from Malaysian C. felis into tick cell culture and recovered circularized genome assemblies for both, one of which (wCfeF) is newly sequenced. We demonstrate that the three Wolbachia strains are representatives of different major clades ("supergroups"), two of which appear to be flea-specific. These Wolbachia genomes exhibit unique combinations of features associated with reproductive parasitism or mutualism, including prophage WO, cytoplasmic incompatibility factors, and the biotin operon of obligate intracellular microbes. The first circularized assembly for R. asembonensis includes a plasmid with a markedly different structure and gene content compared to the published plasmid; moreover, this novel plasmid was also detected in cat flea metagenomes from the US. Analysis of loci under positive selection in the transitional group revealed genes involved in host-pathogen interactions that may facilitate host switching. Finally, the first B. clarridgeiae genome from Asia exhibited largescale genome stability compared to isolates from other continents, except for SNPs in regions predicted to mediate interactions with the vertebrate host. These findings highlight the paucity of data on the genomic diversity of Ctenocephalides-associated bacteria and raise questions regarding how interactions between members of the flea microbiome might influence vector competence. Impact statementWhile fleas are vectors for several globally-distributed bacterial zoonoses, including plague, murine typhus, cat-scratch fever, and flea-borne spotted fever, data on the genomic diversity of the etiological agents of these diseases and other members of the flea microbiome are scant. Here, we focused on Ctenocephalides spp. fleas associated with dogs and cats in Malaysia, a region where flea microbiome data are particularly sparse. Using a combination of direct metagenomic sequencing of flea tissues coupled with culture isolation and sequencing of infected cell lines, we obtained the first genomes for a cat-scratch agent (Bartonella clarridgeiae) and an emerging human pathogen (Rickettsia asembonensis) from Asia, as well as genomes for three different Wolbachia endosymbiont strains and the first mitochondrial genome for Ctenocephalides orientis. The pathogen genomes reveal evidence of selection in genes involved in host-pathogen interactions and in the case of R. asembonensis, a novel plasmid genome was obtained. Two of the Wolbachia genomes are entirely novel and exhibit highly unusual combinations of features associated with potential reproductive manipulations and mutualistic roles. Our findings highlight major gaps in our understanding of flea microbiome diversity worldwide, with implications for the emergence of bacterial pathogens transmitted by these ubiquitous ectoparasites associated with human dwellings. Data summarySequencing reads and genome assemblies produced in study have been deposited in NCBI Genbank under BioProject accession PRJNA909216. Accession numbers for symbiont genomes: wCfeF (CP116767), wCfeJ (CP116768), wCori (CP116769), Rickettsia asembonensis str. Perak (CP116496), pRAS02 (CP116495) and Bartonella clarridgeiae str. Perak (CP116497). Ctenocephalides orientis mitochondrial assembly was deposited with the accession number OQ383237. The authors confirm all supporting data, code and protocols have been provided within 86 the article or through supplementary data files.

genomics↗

bioRxiv2020

African swine fever virus-like integrated elements in a soft tick genome - an ancient virus vector arms race?

BackgroundAfrican swine fever virus (ASFV) is the only known DNA-arbovirus and a most devastating suid pathogen that, originating from a sylvatic cycle in Africa, has spread to eastern Europe and recently reached western Europe and Asia, leading to a socio-economic crisis of global proportion. However, since neither closely related viruses nor integrated viral elements have yet been identified, ASFV evolution remains a mystery. ResultsHere, we show that soft ticks of the Ornithodoros moubata group, the natural arthropod vector of ASFV, harbour African swine fever virus-like integrated (ASFLI)-elements corresponding to up to 10% (over 20 kb) of the ASFV genome. Through orthologous dating and molecular clock analyses, we provide data suggesting that integration occurred over 1.47 million years ago. Furthermore, our data indicate that these elements, showing high sequence identities to modern ASFV, are maintained in the tick genome to protect the tick from infection with specific ASFV-strains through RNA interference. ConclusionWe suggest that this mechanism of protection, shaped through many years of co-evolution, is part of an evolutionary virus-vector "arms race", a finding that has not only high impact on our understanding of the co-evolution of viruses with their hosts but also provides a glimpse into the evolution of ASFV.

microbiology↗

bioRxiv2019

Alongshan virus in Russia

In recent decades, many new flavi-like viruses have been discovered predominantly in different invertebrates and, as was recently shown, some of them may cause disease in humans. The Jingmen tick virus (JMTV) group holds a special place among flavi-like viruses because, in contrast to the "classic" flaviviruses and other flavi-like viruses, they have a segmented ssRNA(+) genome. Two segments of the JMTV genome have homology with regions of the flavivirus genome encoding polymerase and helicase-protease proteins. JMTVs have several open reading frames (ORF) in segments encoding glycoprotein(s) and capsid protein and these ORF are specific only to them. JMTVs greatly differ in virion size. We isolated three strains of Alongshan virus (ALSV), which is a representative of the JMTV group, from adult Ixodes persulcatus ticks collected in two geographically-separated Russian regions in the tick cell line IRE/CTVM19. One of the strains persisted in the IRE/CTVM19 cells without cytopathic effect for three years. Most virions purified from tick cells were spherical with a diameter of approximately 40.5 nm. In addition, we found smaller particles of approximately 13.1 nm in diameter. We obtained full genome sequences of all four segments of two of the isolated ALSV strains, and partial sequences of one segment from the third strain. Phylogenetic analysis on genome segment 2 of the JMTV group clustered our novel strains with other ALSV strains. We found evidence for the existence of a novel upstream ORF in the glycoprotein-coding segment of ALSV and other members of the JMTV group. Significance StatementWe isolated three strains of Alongshan virus (ALSV) from adult Ixodes persulcatus ticks from two geographically separate areas of Russia in the Ixodes ricinus tick cell line IRE/CTVM19. One of the strains persisted in the IRE/CTVM19 cells without cytopathic effect for three years. Our study confirmed the value of tick cell lines in virus isolation and maintenance of persistent infection. The majority of virions of the ALSV strain Miass527 were enveloped spherical particles with a diameter of 40.5{+/-}3.7 nm. We found evidence for the existence of a novel upstream ORF in the glycoprotein-coding segment of ALSV and other members of the Jingmen tick virus group.

molecular biology↗