Virus-induced infections of the central nervous system (CNS) are among the most serious problems in public health and can be associated with high rates of morbidity and mortality, mainly in low- and middle-income countries, where these manifestations have been neglected. Typically, herpes simplex virus 1 and 2, varicella-zoster, and enterovirus are responsible for a high number of cases in immunocompetent hosts, whereas other herpesviruses  are the most common in immunocompromised individuals. Arboviruses have also been associated with outbreaks with a high burden of neurological disorders, such as the Zika virus epidemic in Brazil. There is a current lack of understanding in Brazil about the most common viruses involved in CNS infections. In this review, we briefly summarize the most recent studies and findings associated with the CNS, in addition to epidemiological data that provide extensive information on the circulation and diversity of the most common neuro-invasive viruses in Brazil. We also highlight important aspects of the prion-associated diseases. This review provides readers with better knowledge of virus-associated CNS infections. A deeper understanding of these infections will support the improvement of the current surveillance strategies to allow the timely monitoring of the emergence/re-emergence of neurotropic viruses. Globally, central nervous system (CNS) infections present a high rate of morbidity and mortality, mainly in children and the elderly, resulting in serious public health problems and having a major impact in developing countries ,2. IndeeMany CNS infections are caused by bacteria, fungi, parasites, prions, and viruses and are an ongoing concern due to increased migration, tourism travel, and the ease with which people are displaced. The ongoing pandemic caused by the SARS-CoV-2 virus is a devastating example . RegardiAdditionally, the vast majority of viral infections present at least one neurological symptom during the acute phase, which can lead to neurological, behavioral, and cognitive sequelae months after the initial infection ,6. A recThere is increasing interest in the characteristics and mechanisms associated with viral CNS infections to understand how a wide variety of viruses were selected throughout evolution to gain access to the CNS and to induce effective infection at different stages of life. The different replication strategies adopted for each virus and the poor nucleic acid polymerase fidelity observed in RNA viruses could favor the rise of cumulative mutations and/or recombinations and the emergence or re-emergence of neurotropic viruses of unpredictable pathogenicity and clinically unknown outcomes. Interestingly, several studies have demonstrated that the role of commensal microbiota and the CNS  is associated with viral infection ,12,13. HFlaviviridae and Togaviridae families), and viruses from the Herpesviridae family  to other tissues. Some EV species  can also exploit the respiratory route and spread through the respiratory secretion. Additionally, EVs that cause a vesicular exanthem, as observed in hand, foot, and mouth disease (HFMD), can spread through direct or indirect contact with contaminated discharges or the fluid of blisters from infected people, or objects that contain an infectious virus. Exceptions are the EVs associated with acute hemorrhagic conjunctivitis (EV-D70 and CVA24) that can also be transmitted by direct or indirect eye–hand–fomite contact. Enteroviruses are distributed globally, particularly in tropical regions where poor hygiene conditions play a critical role in the efficiency of viral transmission.Enteroviruses (EVs) are nonenveloped, small, icosahedral-shaped viruses which have a capsid composed of four structural proteins (VP1–VP4), with single-stranded positive-sense RNA, belonging to the EVs enter host cells by binding to one or multiple receptors , followed by receptor-mediated endocytosis and RNA release . To gainGlobally, EVs have been responsible for the majority of aseptic meningitis/encephalitis cases, particularly in children ,20. PoliBrazil has accumulated significant information regarding WPV and vaccine-derived polioviruses (VDPV). In addition, the molecular diversity and epidemiological features of other EV types related to CNS infections circulating in the country have also been highlighted in recent years ,24,25,26Another remarkable observation concerns EV types involved with AFP cases in Brazil. Although polioviruses are the main causative agent of AFP, a significant proportion of NPEVs has been associated with this neurological disorder . The lasAlthough the major causative agents of encephalitis/meningoencephalitis are herpesviruses and arboviruses, particularly in developing countries, EVs have also been found to be among the most frequent pathogens associated with encephalitis globally ,17,36,37Overall, these scenarios suggest different circulation patterns of viruses associated with different neurological diseases in Brazil compared to other countries. Furthermore, it is worth noting that the cocirculation of a high number of different EV types can lead to recombination/mutation events, favoring the emergence/reemergence of novel EV types with unknown pathogenicity. EV-A71 has been identified as a major EV type associated with AFP, although it represents less than 1% of all EV types associated with aseptic meningitis in Brazil , and E6 As recommended by World Health Organization (WHO) in its guidelines for enterovirus surveillance, as support for the Polio Eradication Initiative, specimens of feces and cerebrospinal fluid (CSF) and nasopharyngeal and oropharyngeal swabs should be collected from patients for EV detection . Fecal sFlaviviridae (Dengue virus (DENV), Zika virus (ZIKV), Yellow Fever virus (YFV), Saint Louis Encephalitis virus (SLEV), West Nile virus (WNV), Japanese Encephalitis virus (JEV), Tick-Borne Encephalitis virus (TBEV), Rocio virus (ROCV), Cacipacore virus (CPCV), Ilheus virus (ILHV), Bussuquara virus (BUSV), and Iguape virus (IGUV), Togaviridae (Mayaro virus (MAYV), Chikungunya virus (CHIKV), and Eastern Equine Encephalitis virus (EEEV)), and Peribunyaviridae (Oropouche virus (OROV)). Five human arboviruses have emerged or re-emerged in both hemispheres in recent decades: The flaviviruses DENV, ZIKV, WNV, and YFV, and the alphavirus, CHIKV [Arthropod-borne viruses (arboviruses), a taxonomically diverse group of viruses transmitted between arthropod vectors and vertebrate hosts, are a significant public health problem globally. The most studied arboviruses include the viruses belonging to the families s, CHIKV ,42. MoreCommonly observed clinical manifestations of arboviruses infections in humans range from undifferentiated, moderate, or severe febrile disease; rashes; and arthralgia to neurological and hemorrhagic syndromes. The febrile illness is usually characterized by fever, headache, retro-orbital pain, and myalgia, whereas the neurological syndromes can include meningitis, myelitis, encephalitis, paralysis, paresis, seizures, problems associated with coordination, and behavioral changes .Flavivirus, Alphavirus, and Orthobunyavirus, particularly DENV, CHIKV, and ZIKV [According to the Brazilian Ministry of Health, the neuro-invasive arboviruses of major public health importance in Brazil are those belonging to the genera and ZIKV . Since 2Aedes mosquitoes in two distinct transmission cycles: A sylvatic cycle maintained between mosquitoes and nonhuman primates, and an urban cycle between humans and domestic Aedes spp., such as Aedes aegypti, the main vector [The flaviviruses addressed in this review are mainly transmitted to humans through the bites of infected n vector . Flavivin vector ,47.The flaviviruses’ evolution and epidemiology are mainly determined by the ecological needs of their invertebrate and vertebrate hosts, and they can be grouped by their genome homology, antigenicity, pathogenicity, geographic distribution, and ecological associations .Most flaviviruses infections in humans (~80%) are asymptomatic or result in mild manifestations, such as fever, arthralgia, myalgia, headache, retro-orbital pain, exanthema, and conjunctivitis. However, in some cases, they may cause systemic, hemorrhagic, and neurological syndromes once they reach the CNS. The most severe clinical features related to encephalitis and other neurological syndromes are mostly caused by JEV, WNV, TBEV, and ZIKV ,50,51, wIn Brazil, DENV, ZIKV, and YFV are the most important flaviviruses and have caused explosive and severe epidemics in recent years. Therefore, we chose to address these viruses in this review. Moreover, in endemic countries for arboviruses, such as Brazil, where individuals are highly exposed, these viruses may be involved in CNS-associated disease, although these are considered to be rare events ,44.Despite previous efforts, knowledge regarding the pathogenic mechanisms involved in the neurological syndromes caused by flaviviruses is still scarce. It is known that flaviviruses can cause neurological disease by directly attacking the nervous system or indirectly via immune-mediated processes . As prevA recent review by Mustafa and colleagues  discusseDengue is the most important mosquito-borne viral disease in humans. Dengue is caused by infection of any of four DENV serotypes (DENV-1 to DENV-4) and is associated with significant morbidity, mortality, and economic impact, particularly in developing countries. Each DENV serotype includes several genotypes, a group of DENV isolates that have no more than 6% nucleotide sequence divergence .Dengue incidence has increased 30-fold in the last 50 years, and over 50% of the global population lives in areas at risk of infection by one of the four DENV serotypes. It has been suggested that DENV originated in nonhuman primates in Africa and Asia in a sylvatic cycle about 1000 years ago ,67. DENV infections may result in a wide spectrum of clinical manifestations, from a mild to a potentially life-threatening disease. According to the WHO (2009) criteria, the disease is currently classified as Dengue, Dengue with warning signs, and severe Dengue. Symptoms usually include fever, nausea, vomiting, rash, and aches. However, if untreated, the mortality rate of the severe disease can be as high as 20% ,68.Dengue is not considered a neurotropic virus. However, neurotropism and neuro-invasion have been reported previously ,69, and Encephalitis with acute signs of cerebral involvement has been described in many studies ,74,75,76Dengue-related meningitis is rare, but it has been previously described in a few reports, including in children, for whom the acute onset of fever and symptoms such as headache, vomiting, and/or nuchal rigidity were described ,81,82,83Ischemic and hemorrhagic strokes in conjunction with hemiparesis, dysarthria, headache, vertigo, vomiting, and somnolence have been described in a few cases ,85. A caPeripheral nervous system (PNS) involvement in DENV infection occurs in 5% of neurological symptoms, and includes Guillain-Barré syndrome (GBS), which is not commonly observed; hypokalemic quadriparesis or plegia; mononeuritis multiplex; brachial plexitis; diaphragmatic paralysis; and myositis ,94,95,96Although first isolated in 1947 , ZIKV beA. aegypti mosquitoes. However, vertical transmission, from mother to fetus, can also occur. Vertical transmission been associated not only with spontaneous abortions and stillbirth but also with congenital malformations in the fetuses and/or newborns, for which symptoms are not limited to microcephaly. Congenital Zika virus syndrome (CZS) is mainly characterized by signs and symptoms such as microcephaly, parenchymal or cerebellar calcifications, ventriculomegaly, CNS hypoplasia or atrophy, arthrogryposis, focal pigmentary retina mottling, chorioretinal atrophy and/or coloboma, pallor, atrophy, increased excavation, hypoplasia and/or coloboma of the optic nerve and abnormal visual function, and low birthweight for gestational age, as reviewed in Freitas and colleagues [As an arbovirus, ZIKV is mainly transmitted by the bites of infected lleagues .It is estimated that 80% of ZIKV infections are asymptomatic. When symptomatic, the disease is similar to but usually milder than Dengue, and is characterized by fever, rash, arthralgia, and conjunctival hyperemia . The cliIn a single-center Brazilian study, ZIKV infection was associated with an increased incidence of a wide spectrum of serious neurologic syndromes, some of which lead to death, such as GBS, encephalitis, transverse myelitis, and chronic inflammatory demyelinating polyneuropathy .The mature adult brain appears to be less susceptible to the possible neuro-invasive effect of ZIKV. Although CZS has been associated with encephalitis and microcephaly in fetuses and newborns, a few cases of CNS involvement, such as encephalopathy, encephalitis, meningitis, myelitis, and/or seizures, have been reported in adults. The ZIKV- related CNS complications appear to have a predominantly neuroinflammatory profile ,116,117.YFV is the etiological agent of yellow fever, an acute hemorrhagic disease with a significant impact on public health that is endemic in tropical regions in Africa and South America. Approximately 200,000 YFV infections and 30,000 deaths occur annually, mainly (90%) in Africa. Despite the availability of a safe and efficient vaccine for more than 85 years, an increased number of YF outbreaks have been reported recently in Africa and Brazil .Haemogogus and Sabethes genera) and nonhuman primates and where humans are incidental hosts [In Brazil, since the last urban cycle transmission was reported in 1942, YFV infections have occurred due to the sylvatic cycle. YFV infections have mainly been confined to the Amazon region, where transmission is maintained between sylvatic mosquitoes  or present as a mild undifferentiated febrile illness characterized by an abrupt onset of fever, headache, photophobia, lumbosacral pain, nausea, prostration, myalgias, facial flushing, and conjunctivitis. In severe cases, clinical manifestations such as jaundice, albuminuria, oliguria, bradycardia, delirium, stupor, and metabolic acidosis, leading to shock and hemorrhage, may occur. About 15% of cases do not recover and progress to an intoxication phase with high fever, multisystem organ failure, and fatality .Although rare, YFV infection may induce encephalitis. However, neurological complications after wild-type YFV infection, signs of CNS involvement, and generalized seizures have been reported in the past . Adverse vaccine events have been reported and associated with viscerotropic disease, which presents signs and symptoms that are not distinguished from the natural infection, and neurological disease ,134,135.A study on neurological adverse events following administration of the 17DD yellow fever vaccine in Brazil found the highest rate of neurological adverse events in children aged 5 to 9 years old, with a rate of 0.83 per 100,000 doses in national analysis . Among tOverall, the diagnosis of CNS viral infection involves a combination of clinical findings, molecular and serological assays in CSF, neuroimaging, and chemocytological analysis , as reviIgM ELISA is commonly used to diagnose flaviviruses in the CNS, because virus-specific IgM in CSF represents intrathecal synthesis as IgM is a pentamer and is too large to cross the blood–brain barrier . TherefoViruses can reach the brain through different mechanisms, including peripheral nervous system and axonal transport and the hematogenous route. As described here and based on several reports, it has been shown that, in general, flaviviruses can be neurotropic and neurovirulent. However, the pathways by which these viruses reach and invade the CNS are yet to be fully understood. Significant research has been conducted so far, but more investigations are needed to elucidate the neuropathogenic mechanisms and the resulting outcomes.Alphavirus belongs to the Togaviridae family, which currently comprises 31 recognized species divided into 8 complexes based on antigenic relationships. Alphaviruses are enveloped with icosahedral capsid and single-stranded positive-sense RNA genomes of about 12 kilobases with 2 open reading frames (ORFs), which encode the nonstructural (ns) or replicase polyprotein and the structural polyprotein [The genus yprotein .CHIKV, Mayaro virus (MAYV), Ross River virus (RRV), and Sindbis virus (SINV), which are collectively known as Old World alphaviruses, tend to cause a clinical syndrome characterized by fever, rash, and arthritis, whereas the New World alphaviruses—Venezuelan equine encephalitis virus (VEEV), eastern equine encephalitis virus (EEEV), and western equine encephalitis virus (WEEV) —are geneCHIKV was initially identified in 1952 in Tanzania but spread throughout the Indian Ocean region in the mid-2000s and was introduced into the Caribbean in 2013 . There iAedes aegypti and A. albopictus [The local transmission of CHIKV in Brazil was first detected in 2014 in Oiapoque, Amapá . Since tbopictus .CHIKV infection typically induces a mild disease in humans, characterized by fever, arthralgia, myalgia, and rash. Moreover, long-term sequelae can occur after infection. During the CHIKV epidemic in Brazil, atypical and severe cases were reported, including neuronal impairment in adults and newborns (mother-to-child infection) ,152,153.CHIKV-associated neurological disorders were first described in the 1960s in Asian countries ,155. TheIt has been shown that neurological disease associated with CHIKV infections has the highest mortality rate of potential atypical disease manifestations ,160, andSeveral reports have described the occurrence of neurological complications associated with CHIKV infection in Brazil . The freNeuro-invasive CHIKV infections do not have specific symptoms, which can cause difficulty in clinical diagnosis . The neuThe diagnosis of CHIKV is based on molecular (RT-PCR) or serological  methods from blood or CSF samples. Neurological impairment can be detected with the support of CSF laboratory analyses (pleocytosis is a strong indication) in addition to brain imaging and an electroencephalogram ,168,169.Management for neurological complications depends on the symptoms presented. However, antivirals have been used, mainly acyclovir, in addition to intravenous immunoglobulin, plasmapheresis, and corticoids ,170,171.Other alphaviruses, such as Western equine encephalitis (WEEV) and Madariaga virus, also known as South American eastern equine encephalitis virus, are enzootic pathogens in Brazil with significant potential for neurological disease in both humans and horses. However, outbreaks of equine encephalitis in Brazil are mostly epizootic, with isolated human cases in recent decades. A single case of encephalitis by EEEV was reported in Bahia, Northeast Brazil, in the 1950s . No humaThe implementation of arbovirus neuro-invasive disease surveillance programs has contributed to improving the notification and investigation of cases, but most laboratories still do not routinely diagnose those alphaviruses.Herpes simplex virus (HSV)-1, herpes simplex virus (HSV)- 2 (HSV-2), and varicella-zoster virus (VZV) are members of the alphaherpesvirus subfamily of viruses commonly identified as causing encephalitis . All of Herpesviridae family. HSV-1 and HSV-2 are important causes of CNS infection in the setting of neonatal herpes [Infection with herpes simplex virus, commonly known as herpes, can be due to either herpes simplex virus type 1 (HSV-1) or herpes simplex virus type 2 (HSV-2). HSV-1 is mainly transmitted by oral-to-oral contact, causing infection in or around the mouth . However, HSV-1 can also be transmitted through oral–genital contact to cause infection in or around the genital area . HSV-2 is almost exclusively transmitted through genital-to-genital contact during sex, causing infection in the genital or anal area . Although HSV encephalitis (HSE) is a rare disease with an estimated global incidence of 1 case per 250,000–500,000, it is the most common cause of encephalitis among the l herpes ,177. Appl herpes . Howeverl herpes .It is believed that the reactivation of HSV in the peripheral ganglia with axonal transport to the temporal lobe and the reactivation of the virus in the brain is a possible cause of invasion in the CNS after an episode of recurrent HSV infection with subsequent spread . About 3The varicella-zoster virus (VZV) is also responsible for a wide range of CNS manifestations. VZV can cause a wide variety of symptoms. After the primary infection (chickenpox), the virus lies dormant in the nerves, including the cranial nerve ganglia, dorsal root ganglia, and autonomic ganglia. Many years after the person has recovered from chickenpox, VZV can reactivate to cause neurological conditions . VZV reaThe Alphaherpesviruses have a high prevalence in developing countries, with prevalence higher than 60% in adults. Encephalitis caused by HSV is the most common cause of sporadic fatal encephalitis globally, and CNS involvement is observed in approximately one-third of neonatal HSV infections. Most patients with encephalitis associated with HSV-1 or HSV-2 in primary infection are younger than 18 years of age. The infection associated with VZV usually occurs in patients of older ages after reactivation episodes ,177,179.In Brazil, a limited number of studies have reported the detection of HSV-1, HSV-2, and VZV in infections associated with the CNS. A study conducted in the Western Brazilian Amazon that investigated CSF samples from 165 patients with suspected CNS viral infection through polymerase chain reaction (PCR) found herpes simplex virus (HSV-1) (4.1% of patients), HSV-2 (4.1%), and VZV (20.4%) . In RibeWhole-genome sequencing has been successfully applied for genotyping of HSV-1 ,185,186,In recent years, single-gene inborn errors of innate immunity have been shown to be associated with susceptibility to HSV encephalitis. Regarding the clinical manifestation, HSV-1 causes more than 90% of cases of HSE in adults, whereas HSV-2 infection generally causes aseptic meningitis. In addition, HSV-2 is a common cause of generalized acute encephalitis in neonates . AlthougWhereas diagnostic studies are pending, the best approach to improving long-term outcomes is to include HSE in the differential diagnosis of every patient presenting with fever and neurologic findings and to rapidly initiate empiric intravenous acyclovir therapy. Detection of HSV DNA in the cerebrospinal fluid by PCR is the gold standard for the diagnosis of HSV encephalitis and neonatal meningoencephalitis. Temporal lobe abnormalities detected by brain imaging are considered strong evidence of HSE. PCR has overall sensitivity and specificity of >95% for diagnosis of HSE compared to brain biopsy . MagnetiAcyclovir (ACV) is currently the antiviral drug approved by the FDA for the treatment of HSE with a reduction in mortality ,200. ACVHuman Betaherpesvirus 5 or Human Cytomegalovirus (HCMV) belongs to the human herpesvirus family and the betaherpesvirus subfamily and is among the largest viruses known to cause clinical diseases. It is a double-stranded DNA virus with a genome size of around 235 kb and an icosahedral shape measuring from 150 nm to 200 nm in diameter . HCMV waTransmission can occur through direct or indirect contact from person to person with infectious body fluids, including semen, cervical or vaginal secretions, saliva, urine, and blood products. The excretion of HCMV can last for months or years and can be continuous or intermittent. The two main sources of primary HCMV infection during pregnancy are through sexual activity and contact with young children. HCMV can be transmitted from mother to child in utero, intrapartum, or during breastfeeding. History of sexually transmitted diseases, younger sexual onset, and having multiple sexual partners have been associated with seropositivity for HCMV. Horizontal transmission from child to child or from child to adult, particularly in group daycare centers, probably occurs through the transmission of the virus through saliva on hands and toys .CNS cytomegalovirus infection occurs most commonly in patients with severe immunosuppression, such as those with advanced HIV infection  and those who have undergone bone marrow or solid organ transplantation. Immunocompetent patients are affected very rarely. CNS infection can affect the brain  or the spinal cord . HCMV infection of the CNS is less frequent compared to other HCMV infections, both in AIDS patients and in those who have had a solid organ transplant. Studies performed at autopsy have revealed evidence of CNS infection in almost one-third of patients with AIDS .The prevalence of HCMV increases with age. In children aged 1 to 5 years, prevalence can be as low as 20.7% , while iHCMV is most commonly acquired early in life, from childhood to adulthood, through exposure to saliva, tears, urine, feces, breast milk, semen, and other bodily secretions from infected individuals. It can also be transmitted efficiently through organ and tissue transplantation and blood transfusions. Leukoreduction of blood products significantly reduces the risk of transfusion transmitted HCMV infections .The prevalence of HCMV in Brazil is high, but few studies have shown the viral infections of the central nervous system. A study conducted in 2007 showed that among 43 CSF samples, HCMV was detected in 6% . In 2013The genetic polymorphism of envelope glycoproteins, such as the UL55 gene that encodes gB, the UL73 gene that encodes N glycoprotein (gN), and the UL75 gene that encodes H glycoprotein (gH), are used to classify strains of HCMV. Genetic variability has been evaluated in clinical isolates due to its significant role in tropism and tissue virulence that can influence HCMV infectivity or pathogenicity. Currently, there is no consensus on the classification of HCMV strains based on genotype, evolutionary relationship, or clinical relevance . Some auDiagnosis of HCMV-related encephalitis cannot be made in the clinic without the help of images or virological markers. Because patients with CNS HCMV infection have highly variable symptoms, the clinical manifestations suggest brain or spinal cord involvement with rapid or subacute onset, with signs of meningitis in some cases. Infection of the CNS can occur simultaneously in the brain, meninges, spinal cord, and radius as encephalomyelomeningoradiculitis . The maiDue to the absence of specific clinical symptomatology, clinical and laboratorial diagnoses via imaging and detection of the virus are important to confirm HCMV infection of the CNS. The method of choice is the detection of HCMV in CSF by qPCR or PCR, and genotypic assays have been evolved to rapidly diagnose drug-resistant viruses ,216,217.C), which is highly abundant in the CNS. PrPC can undergo a conformational change into a form that has the ability to self-associate and become infectious, and is known as prion scrapie protein (PrPSc), the PrP pathogenic form [Transmissible spongiform encephalopathies (TSEs) are a group of fatal diseases that cause neurodegeneration . Cognitinic form .The history of prion disease began in the 18th century when Merino sheep were found to display symptoms that included gait changes and excessive licking and itching. With time, it was hypothesized to be an infectious disease of viral or bacterial origin . The term prion is derived from the acronym for Proteinaceous Infectious Particles and was postulated by Stanley Prusiner in 1982 to define a protein, not a viral particle, that was able to promote infection in the prion diseases. Although these diseases are caused by a single protein without a nucleic acid, their characteristics are similar to those of viral diseases .Currently, it is known that prions cause neurodegenerative damage to humans and several other animals. The prion diseases that affect humans are Kuru, Creutzfeldt-Jakob Disease (CJD), Gerstmann-Straussler-Scheinker disease (GSS), fatal familial insomnia (FFI), and variably protease-sensitive prionopathy (VPSPr) . Bovine Sc, through contact with contaminated surgical instruments (iatrogenic), or through using grafts or hormones from cadavers. The acquired forms are considered to account for about 1% of cases in humans [TSEs can be transmitted in different ways, including through environmental, intravenous, oral, and ocular routes. Thus, the etiology of prion diseases is classified into sporadic (idiopathic), acquired (infectious and transmitted), and genetic  . The acqn humans .An example of the orally acquired form is the epidemic of Kuru, which occurred in a tribe in Papua New Guinea that had cannibalistic rituals . In addiSc administration, prions migrate and accumulate in the lymphoid tissues associated with the intestine. Subsequently, other tissues are affected, such as the spleen, tonsils, and lymph nodes. Access to the enteric nervous system (ENS) occurs when nerve cells approach prion-containing lymphoid tissues. After reaching the ENS, there are two pathways that might explain CNS propagation: Through the splanchnic nerve circuit, which involves the celiac ganglion and mesenteric complex, and the parasympathetic vagus nerve circuit  is the most common TSE in humans, constituting about 85–90% of cases . The gento PrPSc .C is a glycoprotein widely expressed in the CNS, neurons, astrocytes, microglia, and oligodendrocytes, and can also be found in other tissues such as the heart, skeletal muscle, retina, intestine, lungs, testes, and immune cells. PrPC is encoded by the PRNP gene and is highly conserved in several species [PrP species . C to PrPSc (Sc has a greater contribution of secondary structure in the β-sheet (43%) and less in the α-helix (30%) than PrPC [Sc, such as resistance to proteases, insolubility in nonionic detergents, and the tendency to aggregate [The pathogenesis of prion disease occurs through the conversion of PrPto PrPSc . These phan PrPC . This stggregate . Sc is autocatalytic, and the conversion of PrPC produces new PrPSc if there is substrate. PrP aggregates sequentially, forming oligomers, protofibrils, and amyloid fibers [The replication of PrPd fibers . Some mod fibers ,231. The toxicity in neural cells is caused not only by the presence of mature amyloid fibers but also by prefibrillar oligomers that interact in an aberrant manner with different molecules and cellular structures. Among other effects, this leads to activation of apoptosis, mitochondrial dysfunction, oxidative stress, damage and permeabilization of the cell membrane . The maiSc can accumulate in different regions of the brain depending on the type of strain, generating different lesions and clinical features [PrPfeatures . The spefeatures .Sc which are known as prion strains. The different strains are not necessarily related to changes in PrP sequence, because the same sequence can fold differently. Strains are mainly characterized according to differences in disease severity, distribution of PrP aggregates throughout the brain, and incubation times, and are related to heterogeneous structures and physicochemical properties of the aggregates [As mentioned above, there are different infectious forms of PrPgregates .PRNP genotype at the polymorphic codon 129, encoding for a methionine (M) or a valine (V), plays an important role in the disease phenotype. For example, the allele V has been correlated with a plaque-like deposition pattern of PrPSc [Prion strains have been mostly characterized in patients with sCJD, the most common prion disease, but very similar patterns are observed in genetic CJD (gCJD), vCJD, iatrogenic CJD (iCJD), sporadic fatal insomnia (sFI), and Kuru. iCJD and Kuru are caused by infection with a sCJD strain . The PRNof PrPSc .CJD and its variants have been included on the list of compulsorily reported diseases in Brazil since 2005. The notification organization chart progresses from the identification of the suspected case by the health care services, through its confirmation, until its notification to the local epidemiological surveillance system and registration in the Notifiable Diseases Information System (Sinan) . From 20PRNP gene was reported, representing seven cases in total: Four with 129M/V and two with 129M/M (molecular data was not obtained from one patient) [Few case reports exist of Brazilian patients with prion diseases, and most consist of genetic and iatrogenic forms. The frequency of allele 129 is 46% MM, 48% MV, and 6% VV in the Brazilian population , suggestpatient) .Although prion diseases have a significant social and economic impact, thus justifying the compulsory reporting in the country, the differential diagnosis among other neurodegenerative diseases is relatively difficult, requiring professional experience and adequate equipment and materials. Thus, the number and geographical distribution of cases in Brazil are underestimated and restricted to major health care centers with the necessary resources and expertise.Neurodegeneration often leads to cognitive impairment, a very common symptom of diseases such as Alzheimer’s disease (AD), Parkinson’s disease (PD), Huntington’s disease (HD), and CJD. Protein aggregation inside and outside the cell, observed in these pathologies, is thought to be related to the generation of toxic species, leading to death. Classically, prion diseases are called spongiform encephalopathies precisely because the patient’s central nervous tissue displays spongiform change, neuronal loss, and gliosis, all of which can be observed through histological analysis and light microscopy.CJD evolves rapidly following the onset of symptoms, with a mean survival of about 6 months. The global incidence is 1.5–2 cases per million per year in 70- to 79-year-old people. CJD is classically characterized by symptoms such as ataxia, myoclonus, and cognitive impairment . gCJD anTSEs are diagnosed according to criteria based on clinical evaluation, clinical tests, and biomarkers. Thus, they are classified as possible, probable, and definitive/confirmed. The European Centre for Disease Prevention and Control  and the Centers for Disease Control and Prevention  have the most updated clinical diagnostic criteria. World Health Organization criteria date from 2003. Therefore, they do not include analyses that have improved and developed the criteria after that date . The vast majority of definite cases are diagnosed after death, because analyses are made from brain tissue and biopsies are invasive. To classify as a probable or possible TSE, evaluation of symptoms is highly important, but is not sufficient in isolation. Paraclinical tests are essential for the classification of cases and the distinction of other treatable pathologies with similar symptoms . Table 2Two paraclinical tests have gained importance in recent years for improving sensitivity and specificity in diagnosis: The MRI (with sequences such as diffusion-weighted imaging (DWI), fluid-attenuated inversion recovery (FLAIR), apparent diffusion coefficient (ADC)) and the real-time quaking-induced conversion assay (RT-QuIC). MRI shows 80% sensitivity and specificity for detecting CJD . RT-QuICSc. A positive RT-QuIC test, together with neuropsychiatric symptoms, is considered sufficient to classify the case as probable CJD [In addition to its high sensitivity and specificity, the RT-QuIC assay has several advantages, not only for sCJD but also for other TSEs . It is aable CJD . In the Brazilian guidelines for TSE diagnosis are still based on the WHO protocol . RecentlPrion diseases are still untreatable today. There is no curative or preventive treatment for these disorders, and only palliative treatments exist to attempt to mitigate some symptoms and provide supportive care . The rarity of the disease and the absence of pre-symptomatic diagnosis also hinder the development of efficient therapies.Some molecules with interesting effects in vitro and in animal models have been tested in human clinical trials, but without success. Observational studies were clinically tested with pentosan polysulfate, quinacrine, flupirtine, and doxycycline without significant effects . These sC expression, leading to a decrease in substrate for conversion. Antisense oligonucleotides (ASOs) are molecules able to specifically bind in a complementary manner, degrade the target messenger RNA, and, consequently, decrease protein production. This approach is now in preclinical development for prion diseases [Another therapeutic approach is to decrease PrPdiseases . Sc and/or PrPC, hindering the conversion. Protective effects of active immunization studies have not proven sufficient to date [C monoclonal antibody, called PRN100, is being tested in six patients with sCJD, but the results of this trial are not yet known. Some antibodies show an important effect in prolonging the survival of prion-infected mice models [Immunotherapy is also a promising approach against prion disease because it may block PrP to date . Passivee models . Sc infection and replication [The use of cell therapies has been suggested as a strategy to recover the damaged areas of the affected brain. Grafted neural stem cells (NSCs) may support the survival of existing neurons  and stimlication . Moreovelication . These sIn the viewpoint of the authors, it is worth mentioning the recently recognized infection caused by SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2) in the CNS infection context. Coronavirus disease 2019 (COVID-19) is caused by SARS-CoV-2 infection, and the clinical symptoms range from mild to severe, for which acute respiratory distress is observed. Although several studies have demonstrated that SARS-CoV-2 can be associated with CNS infections such as meningitis, encephalitis, and Guillain-Barre syndrome , furtherViral CNS infections have been identified as a significant cause of morbidity affecting millions of children and adults, particularly in low- and middle-income countries. These infections may be associated not only with motor problems, but also with cognitive, behavioral, and mental sequelae. Additionally, viral CNS infections can trigger neurodegenerative disorders such as Parkinson’s disease, multiple sclerosis, and Alzheimer’s disease ,8,9,10. Based on this scenario, we can suggest several ways to reduce the impact of these infections, especially in developing countries: (i) Strengthening surveillance systems to identify the emergence of novel neuro-invasive viruses; (ii) ameliorating the tools for early diagnosis from radiographic evidence, and molecular tests to avoid diagnostic mistakes and delays in specific therapy; (iii) performing long-term epidemiological studies to evaluate the incidence and prevalence of the most common CNS infections; and, finally, (iv) implementing effective rehabilitation programs to support patients with sequelae.


WHO revealed that morbidity and mortality from non-communicable diseases (NCDs) are on the increase and NCDs accounted for approximately 29% of all deaths in Nigeria in 2016. This study was conducted to estimate the economic cost of selected NCDs—lung cancer, liver cancer and liver cirrhosis. These diseases are known to be associated with key modifiable health risk behaviours , which are prevalent in Nigeria and often commence during the adolescent years.Data were obtained between 2016 and 2017, from mortality records of patients managed for the selected diseases in the University College Hospital, a major referral centre in Nigeria. Information on costs of treatment, clinic visits, admission and transportation was obtained. Average costs of terminal in-patient care and transportation costs (in 2020 prices) were computed per patient. Costs were converted to the US dollar equivalent using the current official rate of US$1: ₦360.50.Twenty-two (out of 90 cases recorded) could be retrieved and all the patients had been diagnosed in the terminal stages of the disease. The average direct costs were ₦510 152.62 (US$1415.13) for an average of 49.2 days of terminal care for lung cancer; ₦308 950.27 (US$857.00) and ₦238 121.83 (US$660.53) for an average of 16.6 and 21.7 days of terminal care for patients managed for liver cancer and liver cirrhosis, respectively.The economic costs of each of the diseases were very high. Findings emphasise the need for aggressive efforts to promote primary prevention, improve early diagnosis and provide affordable treatment in view of the fact that the monthly minimum wage is less than US$85.00 and treatment costs are borne out-of-pocket by the generality of the population in Nigeria. This is a novel study providing direct costs of terminal care for selected non-communicable diseases (NCDs) in Nigeria from a healthcare perspective.The data collection methods used were robust; lending credence to the strength of the findings.Costs are based on data retrieved from a Federal government tertiary hospital in Nigeria which are lower than equivalent costs obtainable in private facilities.Costs could only be computed for the 22 records retrieved .In spite of the small sample, the findings are reasonable estimates of direct costs of terminal care for the selected NCDs in the study setting and are an important first step to further enquiry.Globally, there has been an increase in the prevalence of non-communicable diseases (NCDs).et al in their review of records of patients admitted with liver diseases into a tertiary hospital in Ile–Ife, Nigeria, from 2013 to 2017 found that hepatocellular carcinoma and liver cirrhosis accounted for 52.8% and 27.2% of all cases of liver disease admitted during the review period.10In Nigeria, the disease burden from communicable diseases is still very high, however, available data indicate that the burden from NCDs is significant. WHO estimated that morbidity and mortality from NCDs accounted for approximately 29% of all deaths in the country in 2016.Cost-of-illness studies determine the burden of diseases to society in monetary terms. In addition, they provide an estimate of the amount that could potentially be saved if a disease were to be eradicated.13 14et al in their review of the global impact of NCDs on healthcare found that globally, cancer and CVDs had the highest reported mean annual total direct costs per patient of approximately US$197 772 and US$81 096, respectively.et al reported national direct costs of diabetes ranging from 3.5 to 4.5 billion international dollars per annum.There has been an increase in the numbers of studies documenting the cost of various NCDs, however, the overwhelming majority of these studies have been conducted in developed countries or in low-income and middle-income countries in Asia.This prevalence-based study adopted a healthcare system perspective and thus focused on the medical costs of the illnesses. The estimation of the cost of care for the long-term illnesses in this study was guided by the cost-of-illness approach described by Rice15 25Data on the illness costs were collected between September 2016 and January 2017 from the University College Hospital (UCH), Ibadan. Established in 1952, it is the premier teaching hospital in Nigeria and is located in Ibadan, South west Nigeria. The hospital also houses the first cancer registry in Nigeria—the Ibadan Cancer Registry which was established in 1960 within the Pathology department of the University of Ibadan.We obtained data retrospectively from a review of case records of patients who had been diagnosed and received treatment for the illnesses investigated. The Case note numbers of patients who died from the diseases of interest—lung cancer and liver disease (liver cancer and liver cirrhosis) in 2014 and 2015 were obtained from the Medical records department. There were a total of 12 reported deaths from lung cancer, of these, 5 case notes could be retrieved. In all, 78 patients were reported to have died from liver diseases  in 2014 and 2015. Of these, nine case notes of patients with liver cancer and eight for patients with liver cirrhosis could be retrieved. Nine casefiles of patients with chronic liver disease were retrieved but information from these cases notes were not used because there was no definitive diagnosis in the case note . Ladep and Taylor-Robinson had noted that some challenges to a definitive diagnosis of liver cirrhosis in Nigeria were reluctance to give consent for liver biopsy and inadequate numbers of trained doctors to conduct the procedure.Direct medical  costs obtained included the following: cost of hospital admission, clinic visits, investigations, medical procedures, medications and consumables. The time from presentation with symptoms to demise for the patients whose case records were obtained ranged from zero to about four and a half months indicating that most patients were admitted at the point when the illnesses were at the terminal stages. Costs were obtained from the relevant departments in the hospital. In addition, costs of investigations and drugs were also obtained from a registered private laboratory and a pharmacy within Ibadan. This was because anecdotal reports indicated that some patients procure their drugs or get some of their investigations done outside the hospital. The costs of drugs and investigations from these private sources were compared with hospital costs, summed and the average for each item was documented. For each illness investigated , the total medical costs for each patient was summed and the average costs incurred from the time of diagnosis or presentation with symptoms to demise was estimated for each disease based on information obtained from the patient’s case note.The direct non-medical cost of illness was limited to transportation costs only. Transportation costs were computed for each patient using the home address provided in the case note. A list of all the addresses was compiled and information on transportation costs from these neighbourhoods to the hospital was obtained from informal interviews with commercial transport drivers in the relevant motor parks. The cost of regular public transportation and the cost of car hire services were obtained. For public transportation, the costs of a return trip for two people was calculated as it was assumed that at least one person would accompany the patient to hospital at every visit because information obtained from the case notes indicated that the patients presented quite late in the course of the disease. For car hire, the cost of hiring the vehicle for a return trip was computed. The average costs (public transportation and car hire) were computed for each patient and the average transportation costs per patient calculated for each disease  between 2017 and May 2020 which was 13.23%.Costs were computed using the formulae below:Total direct cost of care (DC) for all patients for duration of treatment in days=direct medical (DM) + direct non-medical costs (DN-M)Average direct cost for all the patients for the duration of treatment in days (ADC)=DC/number of patients (n).Patients or the public were not involved in the design, or conduct, or reporting, or dissemination plans of our research.The mean age (and SD) of the five patients  with a diagnosis of lung cancer was, 59.8 (±10.6) years. The patients  with liver cancer had an average age of 52.6 years (SD=7.9). There were eight male patients with liver cirrhosis and they had a mean age of 44.3 years (SD=13.1).Records retrieved revealed that on the average, the patients were managed for 49.2 days (from presentation with symptoms to demise) and the average number of days in total spent on admission in hospital was 26.7 days (range: 1–90 days). Patients made an average of 1.6 trips to the hospital (range=1–3 trips). The average direct medical costs of managing one patient for 49.2 days in 2020 prices was ₦484 445. 46 (US$1343.82) and transportation cost was ₦25 707.16 (US$71.31). Average direct  cost of managing one patient was about ₦510 152.62 (US$1415.13); .The average number of days in total spent in hospital across all of the patients’ admissions was 16.6 days (1– 77). Details of the average direct costs for managing a patient with liver cancer (from presentation with symptoms to demise) are presented in Details of the average direct costs for managing a patient with liver cirrhosis in 2020 prices are presented in et al in a tertiary hospital in South East Nigeriaet al in a tertiary facility in North Central Nigeria.et al in their review of records of patients with liver disease (including liver cancer and liver cirrhosis) admitted between 2005 and 2010 in a tertiary hospital in South East Nigeria.et al (2019) similarly documented a mean age of 46.7 (SD=15.3) years, mean duration of admission of 33.4 (SD=26.5) days and a mortality of 47.6% in their retrospective study of patients with liver disease , conducted between 2013 and 2017in a teaching hospital in Southwest Nigeria.This study on the economic costs of NCDs found that all patients for whom records were found were diagnosed in the terminal stages of disease and the average direct medical costs of care were very high. The mean age of the patients who died from lung cancer was 59.8 SD=10.6) years. This is comparable with a mean age of 56.6 (SD=21.6) years and range of 30–81 years reported by Ezemba .6 years.In our study, all the patients for whom data were available presented in the late stages of disease, hence time between presentation and demise was very short. This would have resulted in a reduction in healthcare costs from hospital admission and transportation for follow-up clinic visits and poorer health outcome. Existing literature has highlighted that access and utilisation of healthcare services is still suboptimal in many low-income and middle-income countries and patients often present late leading to poor disease prognosis.36 37et al reported an average annual direct cost of RMB63 675;  per patient with lung cancer in China. Patients in their study had been diagnosed up to 2 years prior to the study compared with about 49.2 days in our study. Cipriano et al reported that monthly costs in the first 6 months of care for a 72-year-old diagnosed with lung cancer in the USA in the year 2000, ranged from US$2687 (no active treatment) to US$9360 (chemoradiotherapy) in 2006 prices, although these costs varied by stage at diagnosis and histologic type.et al in a more recent study reported an estimate of US$21 603 for a month of care in the terminal phase, for a patient who died of lung cancer at age 70 in 2017 in the USA.18The average direct costs  for terminal care for each of the diseases studied was very high. We estimated an average direct cost of more than ₦500 000  for managing a patient with lung cancer. Similarly, other studies have reported high medical costs of care for patients with lung cancer. For example, Edis and Karlıkaya reported that among patients in Turkey, the mean total direct costs per lung cancer patient was more than US$14 000 with costs per patient ranging from US$771.00 to US$104 079.et al estimated that the annual cost of HCC in the USA was approximately US$31 641 per patient.et al estimated that the median cost of treatment per session in 2015 was VND7 439 527.40±VND52 514 030.97, that is, approximately US$329.90±US$232.88 (US$1 in VND:US$22 550.18 for 31 December 2015)20In the current study, the average direct cost of managing one patient with liver cancer in Ibadan, Nigeria for approximately 16.6 days in 2020 was estimated to be ₦308 950.27 (US$857.00). Gondek Although comparisons between our estimates and those provided by the studies cited above need to be made with caution because of our small sample, variations in the disease stage, exact types and duration of treatment provided (among other variables), our findings confirm that irrespective of the setting, the amount expended to treat each of the diseases considered is significant. Boutayeb and Boutayeb in their review of the burden of NCDs in low-income and middle-income countries highlighted the huge difference in costs expended in managing NCDs between patients in developed and low-income and middle-income countries and between affluent and poor patients within developed countries noting that this reflected inequity in access to treatment. In developed countries, a significant proportion of the treatment costs are covered by health insurance and availability of free or largely subsidised care promotes early presentation of patients. This is unlike the situation in Nigeria where the monthly minimum wage for government civil servants was only recently increased to ₦30 000.00—less than US$85.00 in 2019.Our study has demonstrated that direct medical costs of lung and liver cancer and liver cirrhosis in Ibadan, Nigeria are very high, especially when viewed against the prevailing economic environment of the country. Furthermore, many patients in this setting present late resulting in poor prognosis. The substantial costs of care further emphasises the need for greater efforts at prevention of these diseases. Major risk factors for each of the diseases investigated have been identified and interventions exist to reduce engagement in these risky practices. Early diagnosis which can only occur if patients present early and are diagnosed on time needs to be urgently addressed. This would improve disease outcome in the patients and possibly reduce medical costs.The results are based on a relatively small sample, however, they are an important first step to further enquiry.Use of hospital records would have missed other economic costs including costs incurred in other health facilities while trying to obtain a diagnosis, costs of modifications to housing to make homes conducive for patients, indirect and intangible costs. We acknowledge this constraint and limited costs estimates to direct medical and non-medical (transportation) costs only.The hospital data used to compute the average costs of long-term illness were obtained from patients’ records in their case notes. However, all the case notes could not be retrieved. This was because patient records are yet to be fully computerised, hence the case notes had to be manually retrieved. However, all case notes were not found because some were yet to be returned to the central records office as they were still being processed administratively. In addition, records found were for patients who had an average survival of less than a month indicating late stage disease. This could have resulted in an under-estimation of the overall costs incurred.Transportation costs were based on the initial addresses documented in the patients’ case notes and it was not possible to determine if the patients had relocated closer to the hospital during the course of the illness. This could have led to an overestimation of the costs for the patients who relocated closer to the hospital although costs of relocating to a closer location could offset this difference. The costs, however, still provide a reasonable estimation of the transportation costs.The hospital is a federal government tertiary facility, treatment costs could therefore differ from costs in other federal and private facilities within the country. Information obtained from expert opinion revealed that, medical costs in the private wing of the study hospital are typically at least three times more than the costs in the main stream hospital and consultation fees and admission fees are considerable higher.Some information such as costs for drug supplements, dietary modifications, costs of seeking for additional medical care from other hospitals, costs of repeat outpatient visits, costs of investigations conducted to arrive at a definitive diagnosis and costs of cancer management received  would have been missed.Inspite of these limitations, our findings are still very useful and represent estimates of costs of terminal care for lung cancer and liver cancer and cirrhosis in our setting.


Henipavirus genus first identified in Malaysia in 1998. Henipaviruses have bat reservoir hosts and have been isolated from fruit bats found across Oceania, Asia, and Africa. Bat-to-human transmission is thought to be the primary mode of human NiV infection, although multiple intermediate hosts are described. Human infections with NiV were originally described as a syndrome of fever and rapid neurological decline following contact with swine. More recent outbreaks describe a syndrome with prominent respiratory symptoms and human-to-human transmission. Nearly annual outbreaks have been described since 1998 with case fatality rates reaching greater than 90%. The ubiquitous nature of the reservoir host, increasing deforestation, multiple mode of transmission, high case fatality rate, and lack of effective therapy or vaccines make NiV’s pandemic potential increasingly significant. Here we review the epidemiology and microbiology of NiV as well as the therapeutic agents and vaccines in development.Nipah virus (NiV) is a zoonotic paramyxovirus of the  Henipavirus genus of the Paramyxoviridae family and is a zoonotic virus with a high case fatality rate [Nipah virus (NiV) is a member of the ity rate . Our knoity rate ,2. Smallity rate ,4.The original human NiV infections were found to be associated with contact with swine, and it was later confirmed that NiV could be isolated from the nose and oropharynx of pigs ,2,5,6. HSimilar to lyssaviruses, filoviruses, coronaviruses, and the related Hendra virus, NiV is naturally hosted by pteropid bats ,16. FruiThe original NiV infection cluster was first identified in September 1998 in Perak, Malaysia, followed by second and third clusters in the state of Negri Sembilan, with cases occurring primarily among adult men in contact with swine ,5. In MaSporadic NiV outbreaks have occurred in multiple countries throughout South and Southeast Asia since it was first discovered , and theA new and distinct strain of NiV, with infection characterized largely by acute severe respiratory symptoms, arose in Bangladesh and India in 2000–2001. Epidemiological studies revealed clustering around household members and hospital contacts without clear animal exposure, raising suspicion for human-to-human transmission ,9,11.Contact and exposure tracing also began to point toward the consumption of raw date juice as another common behavior among infected humans, emphasized in 2013 when two individuals in Dhaka, Bangladesh developed NiV infection after consuming raw date palm juice ,12,13.Pteropus species of fruit bats, leading to the suspicion that pteropid bats may also serve as the NiV reservoir [Pteropus bat species throughout Asia as well as in countries without known human NiV infection [At the time NiV was first identified, the closely related Hendra virus (HeV) was suspected to persist in eservoir . Subsequnfection ,22,23. Infection ,25. NiV nfection ,24.In 2014, an outbreak of acute encephalitis, meningitis, and severe influenza-like illness in two villages in southern Philippines was attributed to NiV infection and resulted in a case fatality rate of 82% among those with acute encephalitis syndrome [In the recent 2018 Kerala, India outbreak, 23 confirmed and probable cases were identified, with transmission occurring in three hospitals, and primary and secondary cases all traced to a single index case [Although NiV outbreaks have not recurred in Malaysia or Singapore since the initial outbreak in 1998–1999 or in the Philippines since the single outbreak in 2014, nearly annual outbreaks have occurred in Bangladesh with recurrent sporadic outbreaks in India ,4,20.Paramyxoviridae family, now classified as the Henipavirus genus [Nipah virus belongs to the family Paramyxoviridae—the family that also comprises the human pathogenic viruses HeV, measles virus, mumps virus, respiratory syncytial virus, and human parainfluenza virus. Paramyxoviruses possess a single-stranded, non-segmented, negative sense RNA genome fully encapsulated by envelope proteins including a cell receptor binding protein—glycoprotein (G) of henipaviruses, hemagglutinin (H), or hemagglutinin/neuraminidase (HN)—and a separate fusion (F) protein. While cells infected with NiV react strongly with HeV antiserum, cross-neutralization studies have revealed a difference in neutralizing antibodies indicating that NiV and HeV are distinct viruses. Investigations comparing multiple gene regions of NiV to those of other paramyxoviruses have confirmed that HeV and NiV make up a distinct cluster within the us genus .NiV infections are characterized by endothelial syncytium formation, which results in inflammation and hemorrhage. It is postulated that the NiV-G and NiV-F proteins are physically associated and viral fusion results from conformational changes that arise following receptor binding . GuillauThe genomes of NiV-B and NiV-M are 91.8% homologous. The NiV-B genome is six nucleotides longer than that of NiV-M, and all six additional nucleotides are found on the 5′ non-translated region of the F protein gene. Among the open reading frames, only the V gene shows variation between the strains . The V gA study of clinical and autopsy findings of 32 cases of NiV infection during the initial outbreak in Malaysia and Singapore in 1998–1999 found that the mean time from fever onset to hospitalization was 3.3 days, and the mean time from fever onset to death was just 9.5 days. Clinical symptoms of this early outbreak were largely localized to the central nervous system (CNS) and included drowsiness, headache, and some degree of encephalopathy. The most common pathological features were histopathological changes in the blood vessels of multiple organs with evidence of vasculitis of the small vessels. Necrotic plaques were observed in the brain parenchyma of nearly all cases, as well as occasional syncytial or multinucleated giant endothelial cells .The strain of NiV responsible for the recurrent outbreaks in Bangladesh (NiV-B) is distinct from the strain isolated in Malaysia and Singapore (NiV-M) and has been associated with different clinical and epidemiological findings . NiV-B iSeveral antiviral therapies have been investigated for treatment or post-exposure prophylaxis against NiV infection Several nucleoside analogues have been shown to inhibit viral replication of NiV. Favipiravir (T-705) is a purine analogue, RNA-dependent RNA polymerase inhibitor with broad-spectrum anti-RNA virus activity, approved in Japan against emerging influenza strains. It has shown activity against henipaviruses in vitro and in a Syrian hamster model . Remdisi12U), an interferon inducer, has also shown some efficacy against NiV both in vitro and in a hamster model [Poly(I)-poly is a common vector for vaccine delivery and has been used to design a NiV vaccine expressing the NiV G glycoprotein with an incompatible F protein. A total of three African green monkeys challenged with NiV-M intratracheally three weeks after vaccination survived to the end of the study without clinical disease or virus isolation. Notably, two of the three control monkeys developed increased work of breathing and lethargy with oropharyngeal swabs and blood samples positive for NiV RNA, but both recovered and survived to the end of the study . On necrTo address the apparent increased pathogenicity of NiV-B over NiV-M, Mire et al. designed a NiV-B vaccine using rVSV expressing either G or F protein. All vaccinated animals developed NiV-B neutralizing antibodies within three weeks and survived challenge with NiV-B without signs of infection . RecombiA live-attenuated rabies virus-based vaccine against NiV was investigated for wildlife to take advantage of the strong humoral immune responses generated by rabies-based vaccines. NiV-B glycoprotein G encoded into the rabies virus vector induced seroconversion in test mice . The autVirus-like particles (VLPs) produce strong immunogenicity with both native F and G glycoproteins without the risks associated with using a virion containing viral genome. Walpita et al. demonstrated protection and neutralizing antibody titers among Syrian golden hamsters from a vaccine using NiV VLPs . Lastly,NiV infections in humans are sporadic, unpredictable, and often self-contained, consequently traditional clinical development pathways are not fruitful. Future studies should focus on using vaccine candidates to prevent NiV infections in exposed individuals on a compassionate basis. Non-pharmacologic interventions have dominated prevention efforts against NiV infection. Initially, this involved the bans on importation of pigs from known endemic regions and the mass culling of pigs and other animals that tested positive or were in close contact to positive animals ,2,55. LaPteropus spp. of fruit bats are the natural reservoir hosts for NiV, there is also growing evidence of rapid adaptation of virus to other hosts with varying modes of transmission. In just a few years since it was first discovered, NiV was transmitted to humans via pigs, horses, bats, and other infected humans. While infections due to singular spillover events may be limited to small, isolated outbreaks, repeated spillover events of a pathogen with the potential for human-to-human transmission can result in a much greater disease burden. Although physical barriers to prevent NiV spread between bats and humans may impart some protection, outbreaks continue to occur, and human-to-human transmission remains a threat. Continued research into antiviral drug therapies and vaccines is necessary, as are more comprehensive public health measures comprising a combination of education, hygiene, and animal husbandry practices to prevent potentially larger future outbreaks.While there is strong evidence to suggest


The morphology and intermolecular interaction are two of the most important factors in the design of highly efficient dye adsorbent in the industry. Millimeter-sized, bead-type, bio-based lignin/chitosan (Lig/CS) adsorbent was designed for the removal of Congo red (CR), based on the electrostatic attraction, π-π stacking, and hydrogen bonding, which were synthesized through the emulsification of the chitosan/lignin mixture followed by chemical cross-linking. The effects of the lignin/chitosan mass ratio, initial pH, temperature, concentration, and contact time on the adsorption were thoroughly investigated. The highest adsorption capacity (173 mg/g) was obtained for the 20 wt% Lig/CS beads, with a removal rate of 86.5%. To investigate the adsorption mechanism and recyclability, an evaluation of the kinetic model and an adsorption/desorption experiment were conducted. The adsorption of CR on Lig/CS beads followed the type 1 pseudo-second-order model, and the removal rate for CR was still above 90% at five cycles. With the rapid development of chemical processing textile industries, plenty of dyes were poured into the ecosystem, leading to the serious pollution of the water resource ,2. AccorThough these powder-type adsorbents have a relatively high adsorption capacity, benefiting from their large surface area and short diffusion distance, they are not suitable for the industrial application in a practical adsorption column. The powder-type adsorbents can be easily lost or blocked, which will disturb the operation process. The morphology of adsorbents is an essential parameter for the adsorption in industry; millimeter-sized spherical adsorbents are the most favorable for adsorption, according to the literature ,8, sinceExcept for the morphology of adsorbents, the intermolecular interaction between the dye and the adsorbents is another important factor in the application . The comOn the other hand, to meet the demand of sustainable development, the renewable, sustainable, economical, and ecofriendly adsorbents from waste biomass sources were developed for the treatment of dye wastewater ,22. AmonIn recent years, biomaterials have become the focus of absorbent materials, owing to their high removal efficiency, environmental friendliness, and renewability. Among these biomaterials, chitosan is the only one with a positive charge, which is beneficial for the adsorption of anionic dyes . In addiAs mentioned above, a combination of the three kinds of intermolecular interactions  was designed from a bio-based lignin/chitosan (Lig/CS) absorbent for the removal of Congo red , and a mv/v), and Congo red  were purchased from HWRK Chemical Co., Ltd. . Petroleum ether, liquid paraffin wax, and Tween-80 were supplied by Sinopharm Chemical Reagent Co., Ltd. . Distilled water was used throughout the experiments for solution preparation.All reagents were purchased from commercial suppliers of analytical grade reagents and used without further purification. Lignin , chitosan , glacial acetic acid , glutaraldehyde (50% v/v) glutaraldehyde was dropped within 30 min for the chemical cross-linking. After reacting for another hour, the Lig/CS beads were obtained with vacuum filtration and washed three times with petroleum ether and water, respectively.The bio-based lignin/chitosan beads were prepared as follows ,28 Figu: the lig−1 with a resolution of 4 cm−1. The thermogravimetry (TG) analysis was conducted via a NETZSCH TG 209F3 instrument  under an N2 atmosphere with a heating rate of 10 °C/min−1. UV-Vis absorption spectra were conducted with a S 3100 spectrophotometer .The morphologies were observed by a scanning electron microscopy . Fourier transform infrared (FTIR) spectra were recorded on an Avatar 360 Nicolet instrument  using KBr pellets, in the wave numbers ranging from 4000 to 400 cmQ (mg/g) and removal efficiency R (%) was calculated as follows:C0 (mg/L) and tC (mg/L) are the initial and final concentrations of CR in the solution, W (g) is the weight of the Lig/CS beads, and V (L) is the volume of the adsorption solution.Adsorption experiments were performed in a 100 mL Erlenmeyer flask; the Lig/CS beads (containing 30 mg adsorbent) were added to 30 mL of the Congo red (CR) solution, and then the solution was agitated under 150 rpm in an incubator shaker. The pH of the solution was adjusted with a 0.1 M NaOH and 0.1 M HCl solution. The concentration of CR in the final solution was determined by using a UV-vis spectroscopy at 497 nm. The adsorption capacity Chitosan beads and lignin beads can be easily formed in the presence of a glutaraldehyde cross-linking agent ,30. The The image of the obtained pure CS beads and 20% Lig/CS beads are shown in The surface morphology of the pure CS beads and 20% Lig/CS beads are shown in 2 groups and intermolecular hydrogen bonds were confirmed with the broad peak from 3300 to 350 cm−1. The presence of the stretching vibration (2931 cm−1) and bending vibration (1375 cm−1) of CH3 as well as the C−N stretching vibration (1650 cm−1) of amide reveal the existence of an unremoved acetyl group. In addition, the peaks at 1157 and 1092 cm−1 can be ascribed to the C−OH bending vibration and C−O−C stretching vibration [−1 formed, and an increased intensity for methylene (2861 cm−1) was observed, which confirms the cross-linking of CS with glutaraldehyde [−1 is attributed to aliphatic and phenolic -OH. The peaks at 2972 and 2920 cm−1 are ascribed to the stretch vibration of methyl and methylene; 1610 and 1461 cm−1 is the C−C stretching for the aromatic skeleton, and 1314 and 1161 cm−1 is attributed to the vibration of S=O. For the 20% Lig/CS beads, all the peaks of CS, Lig, and the characteristic peak of cross-linking can be observed, demonstrating the successful preparation of Lig/CS beads [The FTIR spectra of CS, CS beads, lignin, and 20% Lig/CS beads are shown in ibration 31 cm−1 aaldehyde . For theibration 31 cm−1 aThermalgravimetric analysis (TG) curves can describe the trend of materials degradation with an increase in temperature and reveal the thermal stability of the materials. The TG curves of CS beads with different Lig content are shown in 2)- and oxygen (-OH)-contained functional group. According to the literature [Congo red (CR) is a general commercial dye, which possesses a negative charge (sulfonate), aromatic ring (biphenyl and naphthyl), and functional group (amino) . As discterature ,18,19,20The adsorption capacity of CR by Lig/CS beads with different lignin content is shown in + concentration; the -NH2 and -OH groups on the Lig/CS beads can be easily protonated at a low pH value, which can promote the adsorption of anionic CR through electrostatic attraction. Taking the protection of the environment and equipment into account, adsorption at a very low pH value (<3) was not conducted. With the increase of the pH value, the adsorption capacity of the beads decreased gradually. This can be assigned to the suppression of protonation, which is unfavorable to the removal of CR. When the pH value exceeded 7, the adsorption capacity and removal rate were decreased further. This can be attributed to the suppression of electrostatic attraction and the competitive adsorption of -OH with anionic CR towards Lig/CS beads [The solution pH, which influences the adsorption behavior, is one of the most important factors. The effect of pH value of the CR solution on the adsorption capacity was investigated , and theCS beads .Temperature is another significant factor which can reveal whether the adsorption is exothermic or endothermic . The effK0, change in the Gibbs free energy (ΔG0), enthalpy (ΔH0), and entropy (ΔS0) and can be calculated by the following Equations (3)–(5) [eQ is the adsorption capacity (mg/g), eC is the equilibrium concentration, R is the universal gas constant (8.314 J/mol·K), and T is the absolute temperature (K). The ΔH0 and ΔS0 are calculated from the slope and intercept of the linear plot of ln K0 versus 1/T.The thermodynamic parameters, including equilibrium constant  (3)–(5) ,38,39:K0G0 was decreased from −1.31 to −4.61 kJ/mol, indicating that the high temperature is beneficial for the adsorption, which is consistent with the effect of temperature on the adsorption [G0 for adsorption indicated the adsorption was a spontaneous physical process [H0 and ΔS0 calculated from the slope and intercept was 8.84 kJ/mol and −0.169 J/mol·K, respectively, and the positive value of ΔH0 revealed an endothermic nature of adsorption again.The calculated results are listed in sorption . In addi process . The ΔH0The effect of the initial concentration  on the adsorption of CR at different times is depicted in Adsorption kinetic is one of the most important characteristics, which can reveal the adsorption efficiency and determine the potential application. In order to investigate the adsorption process and mechanism for CR adsorption onto Ling/CS beads, three kinetic models were used to evaluate the adsorption kinetic process, including the pseudo-first-order model, type 1 pseudo-second-order model, and type 2 pseudo-second-order model .2 (0.998 to 0.9998) shows that the type 1 pseudo-second-order model fits the experiment data better than that of the other two, suggesting that the type 1 pseudo-second-order mechanism is predominant in the adsorption process, and the chemisorption might have been the rate-limiting step for the adsorption process. In this mechanism, the CR molecule stuck to the adsorbents tended to find sites that maximized the coordination number with the surface [The fitting curves of the adsorption for the three models are displayed in  surface ,40.The regeneration is another significant factor in evaluating the properties of the obtained adsorbents; thus, the Lig/CS beads were evaluated by an adsorption/desorption experiment, and the CR-loaded beads were regenerated with a 20% NaOH aqueous solution in an incubator shaker . The adsTo determine the efficiency of the obtained Lig/CS adsorbent for the removal of CR, the maximum adsorption capacities, removal, regeneration, and morphology of these adsorbents were compared  11,20,3,41,42,43In summary, different from conventional powder-type, lignin/chitosan-based absorbents, the millimeter-sized, bead-type lignin/chitosan (Lig/CS) adsorbent was successfully synthesized for the removal of Congo red, which will benefit the industrial application. The introduction of lignin in chitosan can not only enhance the stability of the beads but also can improve the adsorption capacity through the strong π-π stacking interaction between the lignin component and the CR molecules. The optimum content of lignin is 20%; a higher content of lignin will hinder the diffusion of CR molecules. A lower pH value can increase the adsorption capacity, which can be ascribed to the enhanced electrostatic attraction. The experiments and corresponding calculation revealed that the adsorption of CR on Lig/CS beads followed the type 1 pseudo-second-order model. The removal rate for CR is still above 90% after five cycles, and no obvious structural difference was observed for the regenerated adsorbents. Therefore, the bio-based lignin/chitosan beads have great practical potential as an efficient adsorbent for the removal of anionic dyes in industry. Future work will focus on the selectivity, continuous column adsorption, and scale-up of this kind of bio-based bead adsorbent.


Biologics are recommended in Japan to treat moderate to severe Crohn’s Disease (CD). Although CD is associated with high direct costs in Japan, updated information after ustekinumab’s approval is unavailable. We aimed to evaluate the healthcare resource utilization (HRU) and associated direct costs from the payer’s perspective in Japan. Claims data (2010–2018) were retrospectively analyzed to identify patients with CD. HRU and associated costs were evaluated for 12 months before and after biologic initiation and followed-up till 36 months post-initiation. Outcomes were reported using descriptive statistics. Among the included patients , 1,783 were on biologics and 1,713 were on non-biologics. Mean (SD) age was 36.4 (13.2) years and patients were predominantly male (76.1%). Patients aged 18–39 years were affected with CD the most (55.3%). Biologic initiation was associated with a reduction in inpatient stay, length of stay, outpatient visits, and associated costs; and an increase in pharmacy costs and total costs after 12 months. Extended follow-up showed a decreasing trend in HRU and costs till 24 months but an increase after 36 months. These findings demonstrated reduction in clinical burden and slight increase in economic burden with biologics. However, indirect costs also need to be evaluated. Crohn’s disease (CD) is a chronic, progressive, inflammatory gastrointestinal disease leading to bowel damage and disability due to complications such as bowel strictures, abscesses, or fistulas , 2. The As the goal of CD treatment is prevention of progression, symptom control, and reduction in risk of complications, effective long-term management and treatment persistence is essential . The staThe introduction of the Intractable Disease Health Care Act in Japan allowed patients with rare diseases such as CD to receive financial assistance in lieu of authorizing usage of their data for research . Such suAs advanced medications become available to meet the unmet medical needs of patients with CD, there is concern whether excessive prescriptions in real-world treatment practice in Japan could have a major impact on the direct costs associated with CD. Previous Japanese studies have reported high healthcare resource utilization (HRU) and costs associated with CD , 18. WhiA mirror-image design was used for this study focusing on pre-post comparisons of medical resource used and claims cost estimate of one year before and after the biologic index date to understand the economic burden of biologics from the payer’s perspective in Japan  RetrospeInformation retrieved from the JMDC database was anonymized to protect patient privacy under the “Act on the Use of Numbers to Identify a Specific Individual in the Administrative Procedure”. This study was approved by the sponsor’s internal approval committee in accordance with Japanese ethical and legal guidelines, Inflammatory Bowel Disease Research Concept Approval Team and Health Economics Review Board. The informed consent of patients was not required as per the Ethical Guidelines for Epidemiological Research issued by the Japanese Ministry of Health, Labor, and Welfare (MHLW), since the analysis were conducted on the retrospective anonymized claims database managed by JMDC. Reporting of claims data was based on International Society for Pharmacoeconomics and Outcomes Research guidelines .th revision Clinical Modification K50.x) records within the timeframe of the evaluation period; at least one record of CD treatment  rocedure  after thEligible patients were categorized into a non-biologic group i.e. patients without a biologic treatment record  and a biologic group i.e. patients with a biologic treatment record  ; and furClaims information such as demographics (age and gender), insurance status, clinical characteristics (Charlson Comorbidity Index [CCI] and comorbid conditions), diagnosis , dispensed drugs, gastrointestinal surgical interventions, and hospitalization records of patients were extracted from the database for analysis.HRU measures such as outpatient visits, inpatient visits, surgery, enteral nutrition, and pharmacy claims, and cost claims associated with CD were obtained from the JMDC database. The medical resource used and cost estimate of one year before the biologic index date (first claim record of a biologic) and one year after the biologic index date was evaluated by the respective HRU categories. All costs were reported as Japanese Yen (¥) and were not adjusted for inflation because rates and prices have been flat for the past decade in Japan .The cost of each resource was also reported over a follow-up period of 12, 24, and 36 months from the defined biologic index date. Biologic-prescribed patients with a continuous claims record for 12, 24, or 36 months were included in this analysis. For each resource category, the sum of costs of patients were divided by the sum of their follow-up time in months. Data were obtained only if the patients were in the health plan throughout the observation period.Data for all study measures were summarized descriptively as mean values, SD, median, and range for continuous variables and frequency distribution for categorical variables. Baseline demographics and clinical characteristics were presented according to their assigned treatment group. Comorbidities were assessed with the help of CCI prior to the index date for biologic users and throughout the entire enrolment period in JMDC for non-biologic users. CCI categorizes comorbidities of patients and has an associated weight (1 to 6), on the basis of adjusted risk of mortality or resource use, sum of all the weights provides a single comorbidity score for a patient. A score of zero means no comorbidities and higher the score, the more likely the predicted outcome will result in mortality or higher resource use. Overall cost of CD includes following parameters: inpatients , outpatients  and pharmacy cost. CCI and average costs per patient per month, by the type of medical resource utilization used , were reported as mean (SD) or median .After applying the inclusion and exclusion criteria, the included patients with CD  were separated into two groups, biologic-treated  and non-biologic treated  .The mean (SD) age of the overall cohort was 36.4 (13.2) years and majority of the patients were male (76.1%). Majority of the claims were from employed patients (72.3%), followed by dependent adults (21.6%), and children (6.1%). Many patients were also affected by comorbid conditions such as peptic ulcer disease (47.9%), chronic pulmonary disease (38.3%), and mild liver disease (24.6%) .Biologic initiation was associated with a reduction in the mean (SD) number of inpatient visits per patient/month from 0.4 (1.0) to 0.2 (0.2), mean (SD) length of stay (LOS) from 5.3 (15.1) days to 2.5 (3.9) days, and mean (SD) number of outpatient visits per patient/month from 1.9 (4.3) to 1.1 (1.0). Similarly, biologic initiation was associated with a reduction in the overall number of hospitalizations and surgical procedures, but an increase in requirement of enteral nutrition .Biologic initiation was associated with a reduction in mean (SD) per patient/month inpatient costs from ¥276,164  to ¥152,073  and outpatient costs from ¥207,982  to ¥204,157 . However, biologic initiation was associated with an increase in mean (SD) per patient/month pharmacy costs from ¥145,685  to ¥245,894  which led to an increase in total costs from ¥467,483  to ¥497,289  .Biologic initiation was associated with a reduction in mean (SD) per patient/month inpatient, outpatient, pharmacy, and total costs from 12 months to 24 months. However, all the cost categories showed an increase from 24 to 36 months post-initiation .This is the first study to describe the demographics, HRU, and direct costs before and after biologic initiation in CD-affected Japanese patients, using claims data. The major strengths of this study are database capture of the entire CD treatment pathway including all hospitalizations, outpatient visits, and pharmacy dispensing, resulting in inclusive HRU and cost estimation; and evaluation of HRU and direct medical costs in patients on ustekinumab, which was approved for treatment of CD in Japan in 2017. The higher prevalence of CD in males and individuals aged 18–50 years old in the current study is consistent with previous research conducted through national surveys and hospital or claims data analyses . MurakamBiologic initiation was associated with a 11% increase in enteral nutrition. However, this could be attributed to a unique practice recommended in Japanese clinical treatment guidelines  wherein Biologic initiation was associated with a reduction in inpatient visits by 53%, LOS by 54%, surgical procedures by 40%, inpatient costs by 45%, outpatient visits by 39%, and outpatient costs by 2%. However, biologic initiation was associated with an increase in pharmacy costs by 40% and total costs by 6%. Similarly, other studies in various healthcare systems have associated biologic initiation with a reduction in inpatient and outpatient visits and their associated costs; however, an increase in pharmacy costs, most likely due to biologic prescriptions, resulted in greater total direct costs compared with pre-biologic expenses –32. AlthWe also evaluated HRU and direct costs 12, 24, and 36 months after biologic initiation. Here, compared to 12-month data, HRU and costs seemed to be similar or lower after 24 months of follow-up. However, HRU and direct costs after 36 months of follow-up were higher than the 24-month data, suggesting that some patients undergoing biologic treatment may not respond optimally after 24 months of treatment, and hence may have to switch treatment or undergo surgery to manage CD. This seems plausible since previous research has reported a reduction in persistence with biologics as time passes, with a wide range in the rate of non-persistence due to treatment discontinuation or switching .Japanese clinical guidelines recommend biologics for treatment of moderate to severe and severe CD , 40. HenPatients with inflammatory bowel disease (IBD) experience lower HRQoL; higher absenteeism, presenteeism, and work and activity impairment; higher HRU; and higher direct and indirect costs compared with the general population , 44–47. This study has several limitations. First, the JMDC database does not include clinical examination results or physician-assessed severity, which could have enabled stratification of treatment based on disease severity. Second, as the database records insurance claims for employees of medium- to large-scale companies and their dependents only, this study could not assess data of patients who are not covered in the database. Third, as the database is linked to employment status, the elderly are underrepresented in the database. Therefore, the results of this study may not be applicable to every age group. Moreover, since cost data is variable in nature, a larger sample size may be required. A limitation of assessing costs in pre- and post- study designs is that costs incurred due to failure of treatment A but before initiating treatment B, are poorly assessed. Although patients with severe disease often incur higher costs , we repoOur study demonstrated that biologic therapy in patients with CD was associated with a reduction in HRU and related costs, and a slight increase in total pharmacy costs compared with their treatment period before biologic therapy. Use of effective biologic therapies for CD treatment can gradually reduce the economic burden on government that provides financial assistance to patients with CD in Japan. Furthermore, evaluation of indirect costs in the future may reveal the true impact of biologics on payers, and may help in appropriate allocation of resources to reduce the burden on both patients and payers.S1 Table(DOCX)Click here for additional data file.S2 Table(DOCX)Click here for additional data file.S3 Table(DOCX)Click here for additional data file.


This paper showcases the development of plant virology in Croatia at the University of Zagreb, Faculty of Science, from its beginning in the 1950s until today, more than 70 years later. The main achievements of the previous and current group members are highlighted according to various research topics and fields. Expectedly, some of those accomplishments remained within the field of plant virology, but others make part of a much-extended research spectrum exploring subviral pathogens, prokaryotic plant pathogens, fungi and their viruses, as well as their interactions within ecosystems. Thus, the legacy of plant virology in Croatia continues to contribute to the state of the art of microbiology far beyond virology. Research problems pertinent for directing the future research endeavors are also proposed in this review. The origin of plant virology in Croatia is traceable back to the year 1943 and the first publication on a plant virus co-authored by Milan Panjan  and obtaUntil the early 1990s, Miličić et al. led the studies on more than 30 virus species and isolates detected in wild and cultivated plants in Croatia ,5,6,7. Thttps://www.dpvweb.net/dpv/ [accessed on 17 September 2021]). Miličić and collaborators investigated the intracellular inclusion bodies’ submicroscopic structure of Holmes’ ribgrass virus ). As phytoplasmas cannot be successfully grown in pure culture, sequencing and assembly of full genomes are still challenging [https://www.ncbi.nlm.nih.gov/genome/browse#!/overview/candidatus%20phytoplasma [accessed on 29 October 2021]), while a substantial number of genome drafts is also available. A Croatian team contributed to phytoplasma genomic research by obtaining the most complete draft of ‘Ca. P. solani’ strain of 821,322 bp, corresponding to more than 80% of the estimated genome size ) facilitated this advancement [C. scutellarioides with predominating fasciation and witches’ broom symptoms harboring a virus–viroid–phytoplasma combination [We have only started to harness the power of HTS to investigate the plant–pathogen interaction networks including viruses, their different combinations, phytoplasmas and viroids, or even other types of hosts and their viromes. The participation in the COST Action DIVAS  for each virus. Unfortunately, this collection is not curated and there are many isolates that still lack proper characterization and species, or strain, annotation.Whilst we have only remnants of the couple of hundreds of vials of the plant virus antisera collection at the Department of Biology, the current number of virus isolates in our plant virus collection is 1229. The first isolates date back to the 1970s. The first accession is TuMV from Regardless of the hardships in maintenance, contemporary and old TuMV isolates from the collection were used successfully in our research ,100. ManOld virus and viroid isolates from institutional collections and herbaria have been successfully used in contemporary research and the introduction of modern research tools, including HTS, hugely drives these changes . Therefo