Evaluation of the focus reduction neutralization and ELISA tests compared to the plaque reduction neutralization test for the detection of antibodies against measles virus.

Measles is an infectious febrile sickness caused by the measles virus (MeV). Despite an effective vaccine, regional elimination of measles remains a global priority and still faces challenges. To estimate community protection against measles, sensitive tests are needed to identify measles-specific antibodies. The enzyme-linked immunosorbent assay (ELISA) is the standard test for assessing immunity but may fail to detect weak antibody responses in vaccinated populations. The plaque reduction neutralization test (PRNT), is the gold standard test for the assessment of protective antibody levels, however, it is not suitable for routine use. This study validated the focus reduction neutralization test (FRNT) as an alternative. In eight assay runs, fifty serum samples were analyzed in triplicate using PRNT, FRNT, and ELISA. Data analysis revealed that 38 samples were positive by PRNT, 37 by FRNT, and 19 by ELISA. The results showed that ELISA was not sensitive enough to identify low levels of anti-measles antibodies and showed weak agreement with neutralization assays. In contrast, the two neutralization assays had a perfect correlation and similar sensitivity. FRNT appears to be a suitable alternative to PRNT for characterizing immunological responses and vaccination efficacy. Our results highlight the necessity of validating negative and equivocal ELISA results through neutralization methods, during the elimination phases.

Intrathecal immune reactivity against Measles-, Rubella-, and Varicella Zoster viruses is associated with cerebrospinal fluid inflammation in multiple sclerosis.

BACKGROUND/OBJECTIVES: We aimed to determine in multiple sclerosis (MS) whether intrathecal immunoglobulin G (IgG) production against measles- (M), rubella- (R), and varicella zoster (Z) viruses, which is called MRZ reaction (MRZR) and considered the most specific soluble biomarker for MS, is associated with demographic and basic cerebrospinal fluid (CSF) parameters reflecting inflammation. METHODS: We analyzed the presence of positive MRZR and associations with demographic and clinical routine CSF parameters in 513 patients with MS and 182 non-MS patients. RESULTS: Comparing MS patients versus non-MS patients, positive MRZR (38.8% versus 2.2%; specificity 97.8%; positive likelihood ratio, PLR 17.7) had a better specificity and PLR for MS than CSF-specific OCB (89.5% versus 22.0%; specificity 78.0%; PLR 4.1). A positive MRZR in MS patients was associated with female sex (p = 0.0001), pleocytosis (p < 0.0001), higher frequency of presence of plasma cells in CSF (p = 0.0248), normal CSF/serum albumin ratio (p = 0.0005), and intrathecal production of total IgG or CSF-specific OCB (both p < 0.0001), but not with intrathecal production of total IgA or IgM. CONCLUSIONS: This study confirms the MRZR as a highly specific marker of MS and shows that MRZR-positive MS patients more frequently are female and show inflammatory changes of basic CSF parameters than MRZR-negative MS patients.

A measles virus collective infectious unit that caused lethal human brain disease includes many locally restricted and few widespread copy-back defective genomes.

During virus replication in cultured cells, copy-back defective viral genomes (cbDVGs) can arise. CbDVGs are powerful inducers of innate immune responses in vitro, but their occurrence and impact on natural infections of human hosts remain poorly defined. We asked whether cbDVGs were generated in the brain of a patient who succumbed to subacute sclerosing panencephalitis (SSPE) about 20 years after acute measles virus (MeV) infection. Previous analyses of 13 brain specimens of this patient indicated that a collective infectious unit (CIU) drove lethal MeV spread. In this study, we identified 276 replication-competent cbDVG species, each present in over 100 copies in the brain. Six species were detected in multiple forebrain locations, implying that they travelled long-distance with the CIU. The cbDVG to full-length genomes ratio was often close to 1 (0.6-1.74). Most cbDVGs were 324-2,000 bases in length, corresponding to 2%-12% of the full-length genome; all are predicted to have complementary terminal sequences. If improperly encapsidated, these sequences have the potential to form double-stranded structures that can induce innate immune responses. To assess this, we examined the transcriptome of all brain specimens. Several interferon and inflammatory response genes were upregulated, but upregulation levels did not correlate with cbDVG levels in the specimens. Thus, the CIU that drove MeV pathogenesis in this brain includes, in addition to two complementary full-length genome populations, many locally restricted and few widespread cbDVG species. The widespread cbDVG species may have been positively selected but how they impacted pathogenesis remains to be determined.IMPORTANCECopy-back defective viral genomes (cbDVGs) can drive virus-host interactions. They can suppress virus replication directly, by competing with full-length genomes, or indirectly by stimulating antiviral immunity. In vitro, cbDVG can slow down infections and promote persistence, but there is limited documentation of their presence in human hosts or of their impact on disease. We had the unique opportunity to analyze the brain of a patient who succumbed to subacute sclerosing panencephalitis, a rare but lethal consequence of measles. We detected more than 270 distinct cbDVG species; most were restricted to one specimen, but several reached all lobes of the forebrain, suggesting positive selection. Our analyses provide the missing knowledge of the diversity of cbDVG in a natural infection of a human host. They also reveal that a collective infectious unit that caused lethal human brain disease includes few widespread cbDVG, in addition to two ubiquitous complementary full-length genome populations.

An Unconventional Case Study of Neoadjuvant Oncolytic Virotherapy for Recurrent Breast Cancer.

Intratumoural oncolytic virotherapy may have promise as a means to debulk and downstage inoperable tumours in preparation for successful surgery. Here, we describe the unique case of a 50-year-old self-experimenting female virologist with locally recurrent muscle-invasive breast cancer who was able to proceed to simple, non-invasive tumour resection after receiving multiple intratumoural injections of research-grade virus preparations, which first included an Edmonston-Zagreb measles vaccine strain (MeV) and then a vesicular stomatitis virus Indiana strain (VSV), both prepared in her own laboratory. The intratumoural virus therapy was well tolerated. Frequent imaging studies and regular clinical observations documenting size, consistency and mobility of the injected tumour demonstrate that both the MeV- and VSV-containing parts of the protocol contributed to the overall favourable response. Two months after the start of the virus injections, the shrunken tumour was no longer invading the skin or underlying muscle and was surgically excised. The excised tumour showed strong lymphocytic infiltration, with an increase in CD20-positive B cells, CD8-positive T cells and macrophages. PD-L1 expression was detected in contrast to the baseline PD-L1-negative phenotype. The patient completed one-year trastuzumab adjuvant therapy and remains well and recurrence-free 45 months post-surgery. Although an isolated case, it encourages consideration of oncolytic virotherapy as a neoadjuvant treatment modality.

Epidemiology of measles outbreaks, incidence and associated risk factors in Ethiopia from 2000 to 2023: a systematic review and meta-analysis.

BACKGROUND: Although a universal vaccine is available and Ethiopia is working outstandingly towards measles elimination, a recurrent measles outbreak has occurred each year in different parts of the country. Therefore, understanding the epidemiology of measles cases, the incidence of confirmed measles virus cases and related risk factors is crucial. Here, we conducted a systematic review and meta-analysis to summarize information regarding the epidemiology, measles incidence rate and risk factors for national measles infections occurring in the past two decades, from 2000 to 2023. METHODS: Data from electronic databases, including PubMed, African Journal Online, WHO databases and Google Scholars, were searched to identify studies describing measles outbreaks, incidence rates and associated factors in Ethiopia that occurred between 2000 and 2023. Important basic information was extracted in an Excel spreadsheet and imported into Comprehensive Meta-analysis Software version 3 to evaluate the associations between measles outbreaks and different risk factors. We pooled the odds ratios (ORs) and 95% confidence intervals (CIs) for every included risk factor to evaluate the associations with measles outbreaks. RESULTS: We included 36 studies involving 132,502 patients with confirmed measles cases in Ethiopia. The results of this systematic review and meta-analysis revealed that measles outbreaks were more frequently reported in the Oromia region (73,310 (33.1%)), followed by the Southern Nation Nationalities of Ethiopia region (29,057 (13.4%)). The overall pooled analysis indicated that the prevalence of measles susceptibility was 67.5% (95% CI: 67.3-67.8%), with an I2 of 99.86% and a p value for heterogeneity < 0.0001. The non-vaccinated status of the children, their contact history with measles cases, their travel history, the presence of cases in family or neighbors, and malnourished patients were identified as factors associated with the high prevalence and recurrent measles infections in Ethiopia. CONCLUSION: The results of this systematic review and meta-analysis indicated that the pooled prevalence of measles infection was high, which is a public health concern in Ethiopia. Thus, strengthening healthcare services, regular vaccination campaigns, and the integration of health education activities with other services may decrease the incidence rate.

Structural basis of paramyxo- and pneumovirus polymerase inhibition by non-nucleoside small-molecule antivirals.

Small-molecule antivirals can be used as chemical probes to stabilize transitory conformational stages of viral target proteins, facilitating structural analyses. Here, we evaluate allosteric pneumo- and paramyxovirus polymerase inhibitors that have the potential to serve as chemical probes and aid the structural characterization of short-lived intermediate conformations of the polymerase complex. Of multiple inhibitor classes evaluated, we discuss in-depth distinct scaffolds that were selected based on well-understood structure-activity relationships, insight into resistance profiles, biochemical characterization of the mechanism of action, and photoaffinity-based target mapping. Each class is thought to block structural rearrangements of polymerase domains albeit target sites and docking poses are distinct. This review highlights validated druggable targets in the paramyxo- and pneumovirus polymerase proteins and discusses discrete structural stages of the polymerase complexes required for bioactivity.

Host WD repeat-containing protein 5 inhibits protein kinase R-mediated integrated stress response during measles virus infection.

Some negative-sense RNA viruses, including measles virus (MeV), share the characteristic that during their infection cycle, cytoplasmic inclusion bodies (IBs) are formed where components of the viral replication machinery are concentrated. As a foci of viral replication, how IBs act to enhance the efficiency of infection by affecting virus-host interactions remains an important topic of investigation. We previously established that upon MeV infection, the epigenetic host protein, WD repeat-containing protein 5 (WDR5), translocates to cytoplasmic viral IBs and facilitates MeV replication. We now show that WDR5 is recruited to IBs by forming a complex with IB-associated MeV phosphoprotein via a conserved binding motif located on the surface of WDR5. Furthermore, we provide evidence that WDR5 promotes viral replication by suppressing a major innate immune response pathway, the double-stranded RNA-mediated activation of protein kinase R and integrated stress response. IMPORTANCE: MeV is a pathogen that remains a global concern, with an estimated 9 million measles cases and 128,000 measles deaths in 2022 according to the World Health Organization. A large population of the world still has inadequate access to the effective vaccine against the exceptionally transmissible MeV. Measles disease is characterized by a high morbidity in children and in immunocompromised individuals. An important area of research for negative-sense RNA viruses, including MeV, is the characterization of the complex interactome between virus and host occurring at cytoplasmic IBs where viral replication occurs. Despite the progress made in understanding IB structures, little is known regarding the virus-host interactions within IBs and the role of these interactions in promoting viral replication and antagonizing host innate immunity. Herein we provide evidence suggesting a model by which MeV IBs utilize the host protein WDR5 to suppress the protein kinase R-integrated stress response pathway.

Antiviral Properties of Moringa oleifera Leaf Extracts against Respiratory Viruses.

Moringa oleifera (M. oleifera) is a plant widely used for its beneficial properties both in medical and non-medical fields. Because they produce bioactive metabolites, plants are a major resource for drug discovery. In this study, two different cultivars of leaves of M. oleifera (Salento and Barletta) were obtained by maceration or microwave-assisted extraction (MAE). We demonstrated that extracts obtained by MAE exhibited a lower cytotoxic profile compared to those obtained by maceration at concentrations ranged from 25 to 400 µg/mL, on both Vero CCL-81 and Vero/SLAM cells. We examined their antiviral properties against two viruses, i.e., the human coronavirus 229E (HCoV-229E) and measles virus (MeV), which are both responsible for respiratory infections. The extracts were able to inhibit the infection of both viruses and strongly prevented their attack and entry into the cells in a range of concentrations from 50 to 12 µg/mL. Particularly active was the variety of Salento that registered a 50% inhibitory concentration (IC50) at 21 µg/mL for HCoV-229E and at 6 µg/mL for MeV. We identified the presence of several compounds through high performance liquid chromatography (HPLC); in particular, chlorogenic and neochlorogenic acids, quercetin 3-O-ß-d-glucopyranoside (QGP), and glucomoringin (GM) were mainly observed. In the end, M. oleifera can be considered a promising candidate for combating viral infections with a very strong action in the early stages of viral life cycle, probably by destructuring the viral particles blocking the virus-cell fusion.

Immunogenicity of a peptide-based vaccine for measles: a pilot evaluation in a mouse model.

Although neutralizing antibody is an established correlate of protection for measles, T cell-mediated responses play at least two critical roles in immunity to measles: first, through provision of 'help' enabling robust humoral immune responses; and second, through clearance of measles virus-infected cells. Previously, we identified 13 measles-derived peptides that bound to human leukocyte antigen (HLA) molecules in Priess cells infected with measles virus. In this study, we evaluated the immunogenicity of these peptides in a transgenic mouse model. Our results demonstrated that these peptides induced Th1-biased immune responses at varying levels. Of the 13 peptides, the top four immunogenic peptides were further selected for a viral challenge study in mice. A vaccine based on a combination of these four peptides reduced morbidity and weight loss after viral challenge compared to placebo. Our results emphasize the potential of T cell-mediated, peptide-based vaccines against measles.

Predicting the Airborne Transmission of Measles: Impact of Indoor Carbon Dioxide (CO2) Levels and Mitigation Strategies.

BACKGROUND: Measles is a highly contagious cause of febrile illness typically seen in young children. It is transmitted primarily through respiratory droplets and small-particle aerosols and can remain viable in the air. Despite the availability of an effective vaccine, measles remains a major global issue, particularly in regions with low vaccination rates. AIM: This study aimed to quantify the airborne transmission risk of the measles virus in various indoor environments. METHODS: Using indoor carbon dioxide (CO2) levels, we estimated the probability of airborne transmission and the basic reproduction number (Ro) in four hypothetical indoor scenarios, including restaurants, mass gathering events, homes, and business meetings, based on the modified Wells-Riley model. RESULTS: The relationship between airborne transmission rates and indoor CO2 concentrations was visualized, with and without mask usage. Without masks, at an indoor CO2 concentration of 1,000 ppm, the airborne transmission rates were high in homes (100.0%) and business meetings (100.0%) and moderate in restaurants (45.6%) and live events (30.6%). By contrast, the Ro was high in audience-participatory live events (60.9%) and restaurants (13.2%), indicating a higher risk of cluster infections. DISCUSSION AND CONCLUSION: In all indoor environmental scenarios, a positive linear relationship was found between the risk of airborne transmission and indoor CO2 levels. The risk of airborne transmission varied significantly across scenarios, which was influenced by various parameters, such as mask usage, quality of ventilation, conversation, and exposure duration. This model suggests that the risk of airborne transmission of measles can be easily predicted using a CO2 meter.

A Serological Survey of Measles and Rubella Antibodies among Different Age Groups in Eastern China.

BACKGROUND: Measles and rubella are vaccine-preventable diseases targeted for elimination in most World Health Organization regions, and China is considered to have momentum towards measles elimination. Therefore, this study aimed to assess the population immunity levels against measles and rubella in Zhejiang Province in China in order to provide valuable insights for informing future public health measures and contributing to the ongoing global campaign against these diseases. MATERIALS AND METHODS: A cross-sectional serological survey was conducted in 2022. A total of 2740 blood samples were collected from healthy individuals spanning the age range of 0-59 years, representing diverse demographic strata across 11 prefectures in Zhejiang Province in China. The sera were tested for measles and rubella IgG antibodies to determine positivity rates and geometric mean concentrations (GMCs). RESULTS: The overall positivity rate for the measles IgG antibody was 85.3%, with a GMC of 588.30 mIU/mL. The positivity rate for the rubella IgG antibody was 70.9%, and the GMC was 35.30 IU/mL. Measles IgG antibody positivity rates across the 0-11 months, 12-23 months, 24-35 months, 3-5 years, 6-9 years, 10-14 years, 15-19 years, 20-29 years, and 30-59 years age groups were 63.1%, 92.5%, 97.0%, 94.0%, 85.8%, 77.3%, 86.9%, 84.9%, and 88.7%, respectively (trend χ2 = 118.34, p < 0.001). Correspondingly, rubella antibody positivity rates for these same age brackets were 55.9%, 87.9%, 94.7%, 88.2%, 69.9%, 54.2%, 72.6%, 67.5%, and 74.3% (trend χ2 = 199.18, p < 0.001). Both univariate and multivariate analyses consistently demonstrated that age, immunization history, and differing economic levels were significant factors contributing to variations in antibody levels. CONCLUSIONS: The seroprevalence of measles and rubella was lower than that required for herd immunity. Periodic vaccination campaigns should be launched to increase immunity.

Central Nervous System Disorders of Marine Mammals: Models for Human Disease?

This article deals with Central Nervous System (CNS) disorders of marine mammals as putative neuropathology and neuropathogenesis models for their human and, to some extent, their animal "counterparts" in a dual "One Health" and "Translational Medicine" perspective. Within this challenging context, special emphasis is placed upon Alzheimer's disease (AD), provided that AD-like pathological changes have been reported in the brain tissue of stranded cetacean specimens belonging to different Odontocete species. Further examples of potential comparative pathology interest are represented by viral infections and, in particular, by "Subacute Sclerosing Panencephalitis" (SSPE), a rare neurologic sequela in patients infected with Measles virus (MeV). Indeed, Cetacean morbillivirus (CeMV)-infected striped dolphins (Stenella coeruleoalba) may also develop a "brain-only" form of CeMV infection, sharing neuropathological similarities with SSPE. Within this framework, the global threat of the A(H5N1) avian influenza virus is another major concern issue, with a severe meningoencephalitis occurring in affected pinnipeds and cetaceans, similarly to what is seen in human beings. Finally, the role of Brucella ceti-infected, neurobrucellosis-affected cetaceans as putative neuropathology and neuropathogenesis models for their human disease counterparts is also analyzed and discussed. Notwithstanding the above, much more work is needed before drawing the conclusion marine mammal CNS disorders mirror their human "analogues".

Interplay between oncolytic measles virus, macrophages and cancer cells induces a proinflammatory tumor microenvironment.

Attenuated measles virus (MV) exerts its oncolytic activity in malignant pleural mesothelioma (MPM) cells that lack type-I interferon (IFN-I) production or responsiveness. However, other cells in the tumor microenvironment (TME), such as myeloid cells, possess functional antiviral pathways. In this study, we aimed to characterize the interplay between MV and the myeloid cells in human MPM. We cocultured MPM cell lines with monocytes or macrophages and infected them with MV. We analyzed the transcriptome of each cell type and studied their secretion and phenotypes by high-dimensional flow cytometry. We also measured transgene expression using an MV encoding GFP (MV-GFP). We show that MPM cells drive the differentiation of monocytes into M2-like macrophages. These macrophages inhibit GFP expression in tumor cells harboring a defect in IFN-I production and a functional signaling downstream of the IFN-I receptor, while having minimal effects on GFP expression in tumor cells with defect of responsiveness to IFN-I. Interestingly, inhibition of the IFN-I signaling by ruxolitinib restores GFP expression in tumor cells. Upon MV infection, cocultured macrophages express antiviral pro-inflammatory genes and induce the expression of IFN-stimulated genes in tumor cells. MV also increases the expression of HLA and costimulatory molecules on macrophages and their phagocytic activity. Finally, MV induces the secretion of inflammatory cytokines, especially IFN-I, and PD-L1 expression in tumor cells and macrophages. These results show that macrophages reduce viral proteins expression in some MPM cell lines through their IFN-I production and generate a pro-inflammatory interplay that may stimulate the patient's anti-tumor immune response.

Glycan-shielded homodimer structure and dynamical features of the canine distemper virus hemagglutinin relevant for viral entry and efficient vaccination.

Canine distemper virus (CDV) belongs to morbillivirus, including measles virus (MeV) and rinderpest virus, which causes serious immunological and neurological disorders in carnivores, including dogs and rhesus monkeys, as recently reported, but their vaccines are highly effective. The attachment glycoprotein hemagglutinin (CDV-H) at the CDV surface utilizes signaling lymphocyte activation molecule (SLAM) and Nectin-4 (also called poliovirus-receptor-like-4; PVRL4) as entry receptors. Although fusion models have been proposed, the molecular mechanism of morbillivirus fusion entry is poorly understood. Here, we determined the crystal structure of the globular head domain of CDV-H vaccine strain at 3.2 Å resolution, revealing that CDV-H exhibits a highly tilted homodimeric form with a six-bladed ß-propeller fold. While the predicted Nectin-4-binding site is well conserved with that of MeV-H, that of SLAM is similar but partially different, which is expected to contribute to host specificity. Five N-linked sugars covered a broad area of the CDV-H surface to expose receptor-binding sites only, supporting the effective production of neutralizing antibodies. These features are common to MeV-H, although the glycosylation sites are completely different. Furthermore, real-time observation using high-speed atomic force microscopy revealed highly mobile features of the CDV-H dimeric head via the connector region. These results suggest that sugar-shielded tilted homodimeric structure and dynamic conformational changes are common characteristics of morbilliviruses and ensure effective fusion entry and vaccination.

Evaluation of the sensitivity of a measles diagnostic real-time RT-PCR assay incorporating recently observed priming mismatch variants, 2024.

We investigated a variant of measles virus that encodes three mismatches to the reverse priming site for a widely used diagnostic real-time RT-PCR assay; reduction of sensitivity was hypothesised. We examined performance of the assay in context of the variant using in silico data, synthetic RNA templates and clinical specimens. Sensitivity was reduced observed at low copy numbers for templates encoding the variant sequence. We designed and tested an alternate priming strategy, rescuing the sensitivity of the assay.

M-F noncoding region sequences of H1 genotype measles virus provide higher resolution for virus transmission tracing.

As countries and regions move toward measles elimination, extended sequence window including noncoding region located between the matrix and fusion protein genes (M - F NCR) was considered to be used in molecular surveillance. The molecular resolution of M - F NCR was evaluated with 192 genotype H1 strains circulating during 2011-2018 in China. Phylogenetic analyses of the N450 and M - F NCR targets indicated that both two targets could confirm epi-linked outbreak, while M - F NCR target could further improve resolution of the molecular characterization: (1) it could differentiate the strains with identical N450 circulated in one county within one month of disease onset; (2) different transmission chains could be distinguished for strains with identical N450; (3) better spatial-temporal consistency with topology could be provided among sporadic cases with inconsistent N450. Accordingly, M - F NCR could be used to complement the information from N450 to address the specific questions in tracking the virus transmission chains.

A Semiquantitative Protein-Fragment Complementation Assay to Study Protein-Protein Interactions of the Polymerase Complex in Cellula.

Protein-fragment complementation assays (PCAs) are powerful tools to investigate protein-protein interactions in a cellular context. These are especially useful to study unstable proteins and weak interactions that may not resist protein isolation or purification. The PCA based on the reconstitution of the Gaussia princeps luciferase (split-luc) is a sensitive approach allowing the mapping of protein-protein interactions and the semiquantitative measurement of binding affinity. Here, we describe the split-luc protocol we used to map the viral interactome of measles virus polymerase complex.

Optical Control of Mononegavirus Gene Expression and Replication.

Mononegaviruses are promising tools as oncolytic and transgene vectors for gene therapy and regenerative medicine. However, when mononegaviruses are used for therapeutic applications, the viral activity must be strictly controlled due to concerns about toxicity and severe side effects. With this technology, mononegavirus vectors can be grown where they are intended and can be easily removed when they are no longer needed. In particular, a photoswitch protein called Magnet (consisting of two magnet domains) is incorporated into the hinge region between the connector and methyltransferase domains of the mononegavirus polymerase protein (L protein) to disrupt the L protein functions. Blue light (470 ± 20 nm) irradiation causes the dimerization of the two magnet domains, and the L protein is restored to activity, allowing viral gene expression and virus replication. Since the magnet domains' dimerization is reversible, viral gene expression and replication cease when blue light irradiation is stopped.

Investigating Liquid-Liquid Phase Separation in Virus-Generated Inclusion Bodies Using Fluorescence Recovery After Photobleaching of Fluorescently Labeled Host Proteins.

Many negative-sense single-stranded RNA viruses within the order Mononegavirales harm humans. A common feature shared among cells infected by these viruses is the formation of subcellular membraneless structures called biomolecular condensates, also known as inclusion bodies (IBs), that form through a process called liquid-liquid phase separation (LLPS). Like many other membraneless organelles, viral IBs enrich a specific subset of viral and host proteins involved in the formation of viral particles. Elucidation of the properties and regulation of these IBs as they mature throughout the viral replication process are important for our understanding of viral replication, which may also lead to the development of alternative antiviral treatments. The protocol outlined in this chapter aims to characterize the intrinsic properties of LLPS within the measles virus (MeV, a member of Mononegavirales) IBs by using an imaging approach that fluorescently tags an IB-associated host protein. This method uses common laboratory techniques and is generalizable to any host factors as well as other viral systems.