The kidney in hantavirus infection-epidemiology, virology, pathophysiology, clinical presentation, diagnosis and management.

Hantavirus-induced diseases are emerging zoonoses with endemic appearances and frequent outbreaks in different parts of the world. In humans, hantaviral pathology is characterized by the disruption of the endothelial cell barrier followed by increased capillary permeability, thrombocytopenia due to platelet activation/depletion and an overactive immune response. Genetic vulnerability due to certain human leukocyte antigen haplotypes is associated with disease severity. Typically, two different hantavirus-caused clinical syndromes have been reported: hemorrhagic fever with renal syndrome (HFRS) and hantavirus cardiopulmonary syndrome (HCPS). The primarily affected vascular beds differ in these two entities: renal medullary capillaries in HFRS caused by Old World hantaviruses and pulmonary capillaries in HCPS caused by New World hantaviruses. Disease severity in HFRS ranges from mild, e.g. Puumala virus-associated nephropathia epidemica, to moderate, e.g. Hantaan or Dobrava virus infections. HCPS leads to a severe acute respiratory distress syndrome with high mortality rates. Due to novel insights into organ tropism, hantavirus-associated pathophysiology and overlapping clinical features, HFRS and HCPS are believed to be interconnected syndromes frequently involving the kidneys. As there are no specific antiviral treatments or vaccines approved in Europe or the USA, only preventive measures and public awareness may minimize the risk of hantavirus infection. Treatment remains primarily supportive and, depending on disease severity, more invasive measures (e.g., renal replacement therapy, mechanical ventilation and extracorporeal membrane oxygenation) are needed.

INTRODUCTION

Hantavirus-associated diseases are emerging zoonoses that remain a clinical challenge with increasing incidence and multiple serious outbreak situations (Figure 1) [1-4]. Hantavirus infections were first recognized during the Korean War, when more than 3000 United Nation’s troops developed a syndrome typically comprising hemorrhagic fever and kidney failure. This clinical picture was eventually referred to as hemorrhagic fever with renal syndrome (HFRS). The causative viral pathogen, Hantaan virus (HTNV), was first discovered in its natural reservoir, the striped field mouse (Apodemus agrarius), in the late 1970s (Figure 2) [5]. Puumala virus (PUUV), causing a milder course of HFRS also named nephropathia epidemica, was identified in a different mouse strain, bank voles (Myodes glareolus), in the 1980s, although the disease had been known in Scandinavia since the 1930s [6]. Dobrava–Belgrade virus (DOBV) was first isolated from a field mouse (Apodemus flavicollis) in Slovenia in the 1990s and was recognized as a frequent hazard for soldiers during the Yugoslav wars [7-11]. Mortality due to HFRS ranges from <1% in PUUV infections to 10–15% in HTNV or DOBV infections [12-14].

FIGURE 1:

Epidemiology of hantavirus infections in Europe. Incidence for hantavirus infection in 2019 as recorded by the European Centre for Disease Prevention and Control (ECDC). More than 4000 cases of hantavirus disease were reported in Europe (0.8 cases per 100,000 population), with detection of PUUV as the causative pathogen in 98% of cases. Finland and Germany accounted for 69% of all reported cases. Distribution of PUUV, Dobrava virus (DOBV), HTNV and Tula virus (TULV) across Europe are depicted by colour. Recent outbreak situations as reported to the ECDC from 2011 to 2021 are indicated with approximately affected cases and year of the outbreak in parenthesis. European countries that do not report hantaviral infections to the ECDC are depicted in grey (Belarus, Denmark, Moldavia, Montenegro, Kosovo, Ukraine).

FIGURE 2:

European hosts for pathogenic hantaviruses. Source: Images were provided by Shutterstock.com: (A) Holger Kirk/Shutterstock.com, (B) Monika Surzin/Shutterstock.com, (C) Stephan Morris/Shutterstock.com, (D) corlaffra/Shutterstock.com and (E) Ryzhkov Sergey/Shutterstock.com.

Apart from HFRS, there is the hantavirus cardiopulmonary syndrome (HCPS), with a high mortality rate of up to 40% in America [3, 15]. The causative hantavirus of HCPS, Sin Nombre (SNV), was identified within months after its first outbreak in the Four Corners Region in the USA [16, 17]. Other hantaviruses causing HCPS have been found in North and South America [18-21] over the years. Based on the geographical evolution, hantaviruses are separated into Old World viruses (e.g., HTNV, DOBV, PUUV) that commonly elicit HFRS and New World viruses [e.g., SNV, Andes virus (ANDV)] that commonly elicit HCPS (Table 1).

Table 1.

Human pathogenic and medically important hantaviruses according to the 2019 classification of hantavirids [264]

Disease
HFRS	HCPS	Virus	Geographic distribution	Reservoir host	Commercial serological test	Comments
		Old World hantaviruses
X		SEOV	Worldwide	Rat (Rattus)	X	Metropolitan distribution
NEa		PUUV	Europe, Russia, Americas	Bank vole (Myodes glareolus)	X	Main European virus
X		DOBV	Balkans	Yellow-necked mouse (Apodemus flavicollis)	X
X		TULV	Europe, Russia	Common vole (Microtus arvalis)
X		AMRV/SOOV	Far-East Russia	Korean field mouse (Apodemus peninsulae)
X		HTNV	China, Russia, Korea, Central Europe	Striped field mouse (Apodemus agrarius)	X	Main Asian virus
X		Luxi virus (LUXV)	China	Yunnan red-backed vole (Eothenomys miletus)
X		THAIVb	Southeast Asia	Greater bandicoot rat (Bandicota indica)
X		SANGV	Africa	African Wood Mouse (Hylomyscus simus)		First African virus, spillover infections to bats
		New World hantaviruses
	X	Bayou virus (BAYV)	North America	Marsh rice rat (Oryzomys palustris)
	X	Black Creek Canal virus (BCCV)	North America	Hispid cotton rat (Sigmodon hispidus)
	X	New York virus (NYV)	North America	White-footed mouse (Peromyscus leucopus)
	X	SNV	North America	Eastern deer mouse (Peromyscus maniculatus)	X	Main North American virus
	X	Choclo virus (CHOV)	Panama	Fulvous colilargo (Oligoryzomys fulvescens)
	X	Araraquara virus (ARQV)	Brazil	Hairy-tailed bolo mouse (Bolomys lasiurus)
	X	Anajatuba virus (ANJV)	South America	Fornes' colilargo (Oligoryzomys fornesi)
	X	Castelo dos Sonhos virus (CASV)	South America	Brazilian colilargo (Oligoryzomys eliurus)
	X	ANDV	Argentina, Chile	Long-tailed colilargo (Oligoryzomys longicaudatus)	X	Main South American virus, human-to-human transmission
	X	Bermejo virus (BMJV)	Argentina	Hairy-tailed bolo mouse (Bolomys Lasiurus)
	X	Laguna Negra virus (LANV)	Argentina, Bolivia, Paraguay	Small vesper mouse (Calomys laucha)
	X	Lechiguanas virus (LECV)	Argentina	Flavescent colilargo (Oligoryzomys flavescens)
	X	Oran virus (ORNV)	Argentina	Long-tailed colilargo (Oligoryzomys longicaudatus)

PUUV causes nephropathia epidemica, a mild form of HFRS. PUUV is the main European hantavirus, whereas HTNV is the main Asian hantavirus. ANDV and SNV are the major South and North American hantaviruses causing HCPS. Novel New World hantaviruses were detected recently with unknown pathogenicity.

So far only serological evidence reported.

NE: nephropathia epidemica.

Hantavirus infections are rare infectious diseases. However, hantavirus diseases emerge progressively, in particular in Europe, and new hantaviruses with yet unknown pathogenic impact are being discovered in different parts of the world, calling for joint efforts at the crossroads of nephrology, infectious diseases and public health [1, 2, 14, 22].

Hantavirus ecology and epidemiology

Hantaviruses are carried and transmitted to humans by persistently infected rodents, insectivore hosts and bats. The geographic distribution and ecology of hantaviruses are therefore closely connected to their hosts. Myodes, Ratus and Apodemus are the primary rodent reservoirs of Old World hantaviruses, whereas New World hantaviruses are transmitted via Sigmodontines, a rodent subfamily of New World rats and mice [1]. Hantavirus species have been discovered in Africa, Asia and Europe, as well as North America and South America (Table 1) [1, 23, 24]. The evolution and distribution of hantavirus populations and the subsequent rate of infections are associated with climate change and disturbed rodent habitats caused by excessive agriculture and deforestation [25-27]. Increases in temperature, humidity and rainfall are directly associated with the incidence of hantaviral infections in different parts of the world [28-30]. For example, warm ambient temperature and abundant rainfall directly affect vegetation growth, boosting rodent populations [31, 32]. In Europe, rodent food availability and higher ambient temperatures in cold seasons have been reported to cause more frequent and severe outbreaks of hantavirus infections [30, 33]. Higher temperature is also correlated with increased rodent reproduction, sexual maturation and survival [34, 35]. The extent of rodent habitat growth and subsequent hantavirus distribution by long-term climate change is unclear (Table 2) [24]. Nevertheless, migration of humans due to climate change to areas less vulnerable to extreme weather events will increase rodent density and may even contribute to hantavirus disease in new regions [36]. Of note, the increase in atmospheric moisture as well as the increase in air temperature facilitates aerosolization and thus augments hantaviral infectivity [29, 37].

Table 2.

Research needs in hantavirus diseases

Hantavirus ecology and epidemiology• Extension of host reservoir habitats due to climate change• Impact of human migration on hantaviral spread• Characterization of spillover infections, especially to bats, facilitating hantaviral reassortment and spread• Role of human-to-human transmission of ANDV and SNV in public health

Hantavirus structure and life cycle• Role of integrins for cell entry in pathogenic and apothegenic hantaviruses• Integrin gene polymorphisms in hantaviral attachment and the susceptibility for hantaviral infection

Hantavirus pathogenesis and immunopathology• Hantavirus cell tropism and organ-specific dysfunction in HRFS and HCPS• Generation of HFRS animal models reflecting human hantavirus disease• Local and systemic tissue damage caused by pathogenic mediators in hantavirus disease in in vivo models• Assessing causal attribution of involved pathogenic mediators using targeted approaches, i.e. antibodies• Pathomechanisms of AKI in hantavirus disease

Clinical presentation• Characterization of hantavirus disease beyond the dichotomous denominations of HFRS and HCPS• Revised taxonomy beyond HFRS and HCPS• Long-term kidney sequelae in DOBV and HTNV• Prognostic impact of pre-existing CKD, RRT, kidney transplantation and immune suppression in hantavirus disease• Phenotype, frequency and sequelae of kidney involvement in ANDV- and SNV-caused disease• Systematic analyses of overlapping features in pathogenesis, phenotype and treatment approaches between emerging viruses, i.e. SARS-CoV-2, Ebola virus and hantaviruses

Diagnosis• Definition of clinical and diagnostic criteria for hantavirus diseases• Prognosis-indicating scores facilitating risk stratification for the ER and ICU

Prevention• Development of EMA- and FDA-approved vaccines, i.e. on the basis of recent RNA vaccine technology• Development for preventive/precaution measures for public health (especially in endemic areas)

Therapy• Development of targeted antiviral pharmacological approaches examined in randomized controlled clinical trials

AKI: acute kidney injury; CKD: chronic kidney disease; EMA: European Medicines Agency; ER: emergency room; FDA: US Food and Drug Administration; ICU: intensive care unit; RNA: ribonucleic acid.

Hantaviruses are circulating between chronically infected host reservoirs, whose infection is usually inapparent. Transmission to humans is caused by inhalation of contaminated aerosolized rodent excreta, leading to so-called spillover infections [3]. These spillover infections to humans or other animals, such as red fox, moose or domestic cat and dog, are a concern for public health, as they facilitate the evolution of new hantavirus species by natural reassortment (Table 2) [33, 38, 39]. The risk of hantavirus infections in humans mainly depends on the closeness to and frequency of contact with rodents. Therefore people with close exposure to rodent habitats, such as farmers, forestry workers, military personnel and zoologists are subject to a greater risk [40-42]. However, many cases are observed after sporadic cleansing of environments that are a habitat for rodents, like basements or attics, without a specific professional exposure.

The epidemiology of hantavirus-associated diseases depends on their host reservoirs’ distribution. In Europe, Tula virus (TULV), DOBV and PUUV are detected most frequently in their rodent reservoirs (Figure 2). PUUV is the most widely distributed hantavirus in Northern and Central Europe and usually causes mild disease (Figure 1, Table 1) [43]. However, PUUV still poses a threat due to its increasing incidence, with recurring outbreak situations and thousands of patients in epidemic years [2, 44, 45]. In contrast, Southeastern Europe is dominated by DOBV, resulting in a moderate–severe disease form of HFRS (Figure 2) [2]. Here, the Balkan region is primarily affected by DOBV, although its host distribution is greater [3, 46, 47]. Of note, DOBV has been emerging in Central and Eastern Europe in recent years [48-51]. TULV is dominantly circulating in the Tula region in Russia and leads to a mild disease course [52]. However, patient cases are found throughout Central Europe and the Baltic states (Figure 2) [52-55].

Asia is the continent with the longest history of hantavirus infections. HTNV and the related Amur/Soochong virus (AMRV/SOOV) are found in China, Far East Russia and Korea, causing moderate forms of HFRS [1, 5, 56, 57]. China reports >10 000 cases and southern Korea ∼300–500 cases/year [58]. Further species with unknown pathogenicity, such as Thailand virus (THAIV), Imjin virus (MJNV) and Jeju virus (JJUV) have been recognized in different parts of Asia (Table 1) [1, 2, 59, 60].

Due to the cosmopolitan distribution of rats as its natural host, Seoul virus (SEOV) is considered to circulate worldwide [1]. SEOV is primarily reported from urban China; however, patient cases have also been observed occasionally in the USA and Europe [61-63]. Surveillance studies indicate that HFRS caused by rat-borne SEOV usually occurs in metropolitan areas and current trends in urbanization may further influence SEOV epidemiology [64].

New World hantaviruses were first discovered after an outbreak of an acute respiratory disease named HCPS in the USA in 1993 and SNV was found as a first causative pathogen [15, 17]. SNV is the main pathogen circulating in North America [2]. In contrast, ANDV is the dominant hantavirus in Latin America [65]. Of note, ANDV is the only known hantavirus that can be transmitted from human to human by superspreading events, further threatening public health and making pandemics a potential scenario [18–21, 66]. Recently, novel hantaviruses with unclear taxonomic status were discovered in their natural reservoirs in America, but little is known about their pathogenicity (Table 1) [1, 22].

Africa is the continent with the most recent scientific progress in hantavirus ecology and epidemiology. Fifteen years ago the first pathogenic hantavirus in Africa, Sangassou virus (SANGV), phylogenetically related to Old World hantaviruses, was isolated in Guinea [67, 68]. Novel hantavirus species have been discovered not only in rodents, but also in shrews and bats in Africa [2, 69, 70]. In particular, the role of bats as novel hosts facilitating easier spillover infections to humans reflects an additional concern for public health (Table 2) [3].

Taken together, the ecology and epidemiology of hantaviruses and their associated diseases are mainly reflected by their host reservoirs’ distribution. Hence HFRS caused by DOBV, HTNV and PUUV dominates Eurasia, whereas SNV- and ANDV-associated HCPS is primarily found in America (Table 1).

Hantavirus structure and life cycle

Hantaviruses, members of the Bunyaviridae family, are enveloped, single-stranded ribonucleic acid (ssRNA) viruses [71]. The total size of the viral RNA (vRNA) ranges from 11 845 nucleotides for HTNV to 12 317 nucleotides for SNV [1]. The viral genome is separated into three segments referred to as S (small), M (medium) and L (large), which share a 3′ terminal sequence and encode the nucleocapsid (N) protein, the glycoprotein precursors (GPCs) and the RNA-dependent RNA polymerase (RdRp) [23, 72]. The vRNA itself is encapsulated by the N protein, forming circular ribonucleoproteins (RNPs) [73]. The virion's surface consists of the two glycoproteins Gn and Gc (formerly G1 and G2, respectively) that mature from GPCs. The RdRp amplifies and transcribes vRNA and is essential for hantavirus replication (Figure 3). Hantaviruses can survive for >10 days at room temperature and for >18 days at +4°C outside cells, facilitating transmission and spillover infection [74].

FIGURE 3:

Hantavirus structure and life cycle. Hantavirus virions have a trisegmented single-stranded ribonucleid acid (ssRNA) genome, referred to as L (large), M (medium) and S (small) segments. The particle's surface consists of the glycoproteins Gn and Gc and viral RNA-dependent RNA polymerase (RdRp) is essential for hantavirus replication and transcription. The hantavirus life cycle consists of eight essential steps. (1) Hantaviruses attach to the host cell's surface by binding to surface receptors with their glycoproteins. (2) The virion particle enters the host cell either by Clathrin-dependent (Old World hantavirus) or -independent endocytosis (New World hantavirus). (3) Hantaviruses are uncoated in the host cell's endosomes and lysosomes, facilitating the release of the viral genome and proteins. (4) Viral RNA is transcribed by the RdRp and (5) viral mRNA is translated into viral proteins, hijacking the host cell's machinery. (6) vRNA is replicated by RdRp and (7) all viral components are put together at the Golgi apparatus (Old World hantavirus) or directly at the cell membrane (New World hantavirus). (8) Mature virion particles egress the host cell by fusion of the Golgi apparatus (Old World hantavirus) or the viral vesicle (New World hantavirus) with the cell membrane. ER: endoplasmic reticulum; Gc: C-glycoprotein; Gn: N-glycoprotein; L: large segment; M: medium segment; S: small segment; vRNA: viral RNA. Source: Figure created with Biorender.com.

Hantaviruses not only infect primarily endothelial cells, but also replicate in the epithelium—including podocytes and tubular cells in the kidney as well as alveolar cells in the lungs—macrophages, dendritic cells and lymphocytes [21, 75–79]. Figure 3 highlights the hantavirus life cycle. In vitro studies suggest that hantaviruses attach to integrins (αVβ3, α5β1, αMβ2 and αXβ2) on host cells through binding with their larger glycoprotein Gn [76, 80, 81]. β3- and β1-containing integrins are proposed as the major entry receptors for both virulent and avirulent Old and New World hantaviruses [80-82]. There is evidence that binding to α5β1 integrin instead of αVβ3 is associated with reduced virulence—as can be seen with the apathogenic Prospect Hill virus (PHV). On the other side, the African SANGV is the only pathogenic hantavirus known to bind to α5β1 and to cause hantavirus disease [83, 84]. Different nucleotide polymorphisms have been implicated in infection susceptibility in humans, calling for further research in hantaviral attachment (Table 2) [85, 86]. Of note, protocadherin-1 (PCDH1) was reported recently to be another critical determinant of attachment, entry and infection of New World but not Old World hantaviruses [87].

After cellular attachment, Old World hantaviruses enter host cells by clathrin-dependent endocytosis, whereas New World viruses employ different mechanisms of cell invasion simultaneously, including micropinocytosis, clathrin-independent receptor-mediated endocytosis and cholesterol- or caveolae-dependent endocytosis (Figure 3) [23, 88, 89]. After cell entry, viral particles are processed in endosomes or lysosomes, where they detach from their cellular receptors and uncoat as a consequence of decreasing pH. Afterwards, hantavirus RNPs are liberated into the cytoplasm by fusion of viral and endolysosomal membranes [88]. In the cytoplasm, vRNA is then transcribed to L, M and S messenger RNA (mRNA), enabling their translation into viral proteins that hijack the host machinery [1]. Additionally, vRNA is transcribed to complementary RNA (cRNA) using host-derived primers facilitating amplification and replication of vRNA [1]. Both transcription and replication processes of hantaviruses are performed by viral RdRP [23]. After replication of the hantavirus genome, the vRNA is encapsulated by the N protein. Then, further N proteins bind, resulting in larger RNPs [90, 91]. The different hantavirus components are finally assembled at the Golgi apparatus, or alternatively, as currently suggested for New World hantavirus, at the plasma membrane itself [92]. By fusion with the host's cell membrane, hantaviruses finally egress [93].

Hantavirus pathogenesis and immunopathology

Despite different causative pathogens and disease patterns, HFRS and HCPS have an overlapping pathophysiology consisting of increased vascular permeability, platelet activation and an overreacting host immune response (Figure 4, Table 3) [23, 46]. Differences in disease are reflected by the different vascular beds that are primarily infected—renal medulla or pulmonary capillaries. However, the mechanisms mediating cell tropism and organ-specific dysfunction during HFRS and HCPS remain unknown (Table 2) [94].

FIGURE 4:

Hantavirus-caused pathogenesis is characterized by vascular leakage and platelet activation. Hantaviruses primarily infect endothelial cells, reducing their barrier function while increasing vascular permeability. Endothelial cell–cell contacts are disturbed by the downregulation of VE-cadherin in adherens junctions caused by vascular endothelial growth factor A (VEGFA) or bradykinin. Platelets are activated after hantavirus infection by either the direct interaction of viral glycoproteins and platelet integrin αIIβ3 or by endothelial cell damage releasing adhesive factors such as fibrinogen, fibronectin and von Willebrand factor. Hantavirus can additionally cause intravascular coagulation. Both activated platelets and coagulation contribute to thrombocytopenia. Source: Figure created with Biorender.com and figure concept adapted from Vaheri et al. [23].

Table 3.

Pathophysiology in response to hantavirus infection as discovered in cell culture, in vivo models and human biosamples

	Evidence reported in
Pathogenic mechanisms in hantavirus disease	Cell culture	In vivo models	Humans	Comments	References
Increased vascular permeability
VEGF-induced endothelial hyperpermeability	X	X	X	Orchestrated by a decrease in VE-cadherin and inactivation of the αVβ3-integrin–VEGFR2 complex	[98, 99, 105–107]
Bradykinin-induced capillary leakage	X		X		[109–111]
Cytokine-mediated hyperpermeability	X		X		[113–119]
Platelet activation
Direct viral-caused platelet consumption	X			Interaction of viral glycoproteins and integrin αIIβ3 on platelets	[124, 125]
Endothelial cell injury causing platelet activation	X		X	Release of adhesive agents, such as fibrinogen, fibronectin, extracellular vesicle tissue factor and von Willebrand factor after endothelial infection	[23, 126, 127, 131, 132]
Overreacting host immune response
Reverse CD4+:CD8+ T-cell ratio	X		X	Causes further activation of pro-inflammatory cytokines	[1, 3, 151, 152, 156]
Triggered T-cell immune response by HLA haplotypes			X	May explain interpersonal and regional differences in susceptibility and vulnerability	[153, 156–163]
Cytokine-mediated activation of innate and adaptive immune responses causing tissue damage			X	Distinct cytokine profiles in HFRS and HCPS; cytokine storm is a common central component in response in hemorrhagic fevers	[116, 118, 119, 138, 142–145]

Increased vascular permeability, platelet activation and an overreacting host immune response are the central pathomechanisms in human disease caused by pathogenic Old World and New World hantaviruses.

The pathophysiology of capillary leakage is characterized by the disruption of endothelial cell–cell contacts. Hantaviruses primarily infect and replicate in endothelial cells of capillaries; however, there are few to no direct cytopathic effects to the endothelium as detected in histological samples [23, 95]. In contrast, it is hypothesized that the breakdown of endothelial cell-to-cell contact is mainly caused by the release of vasoactive factors, including vascular endothelial growth factors (VEGF), bradykinin and cytokines, rather than endothelial cell death [96, 97]. Mechanistically, released VEGF results in a decrease of VE-cadherin, a major component of adherens junctions in endothelial cells, in vitro [98, 99]. VEGF-induced capillary hyperpermeability may additionally be a direct consequence of hantavirus-caused inactivation of αVβ3 integrins followed by a decrease of VEGF receptor 2 (VEGFR2) on the cell surface [75, 100]. VEGFR2 and αVβ3 integrins normally form a complex affecting VEGFR2-directed permeability in response to VEGF [101]. Hantaviruses functionally block the VEGFR2/αVβ3 integrins complex that causes VEGF-orchestrated cellular permeability in vitro and in vivo [102-105]. In line with these findings, levels of VEGF and soluble VEGFR2 in serum and urine correlate with disease severity, indicated by low urine output and haemorrhages in PUUV- and DOBV-infected patients [106]. Moreover, as hypoxia additionally challenges VEGF production, elevated levels of VEGF are detected in pulmonary edema fluids of HCPS patients and correlate with HCPS disease severity [99]. Of note, vandetanib, a small-molecule antagonist of VEGFR2, leads to modest survival benefits in vivo as examined in the Syrian hamster model of ANDV-caused HCPS [107]. However, transfer of this treatment approach to the patient setting has not been successful due to the reported frequent severe adverse events of vandetanib in large-scale cancer trials on the one hand and the fragile, critically ill HCPS patients on the other [107, 108].

Furthermore, in vitro studies revealed activation of the plasma kallikrein–kinin system leading to an increase in bradykinin as another potential mechanism causing vascular permeability in hantavirus diseases [109, 110]. Successful treatment with the bradykinin receptor antagonist icatibant in case reports of severe PUUV infections underline the potential involvement of bradykinin in human disease [110-112]. Breakdown of cell-to-cell contacts is additionally caused by cytokines in vitro and in vivo [113-116]. For example, tumour necrosis factor (TNF)-α contributes to vascular permeability through the production of nitric oxide [117]. In particular, capillary leakage and extracellular matrix degradation are linked to certain cytokine profiles in serum samples in both HFRS and HCPS patients and may even predict disease severity [118, 119].

Interestingly, the ability of hantaviruses to infect endothelial cells is shared with many (emerging) viruses, i.e. severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), flaviruses (including dengue virus) and filoviruses (i.e., Ebola and Marburg viruses), although endothelial receptor, entry mechanism, mediators of disease and the extent of endothelial damage differ widely [100, 120–123]. Hence dysregulation in vascular function contributes significantly to pathophysiology in many life-threatening viral infections and our growing knowledge due to the current coronavirus disease 2019 (COVID-19) pandemic may promote research advances for hantavirus disease (Table 2) [100].

A common feature of hantavirus infection is thrombocytopenia; however, the exact pathomechanisms remain unknown [3, 23, 78]. Currently it is hypothesized that the adhesion of hantaviruses mediated by viral glycoproteins and integrin αIIβ3 on the surface of platelets contributes to platelet depletion [124, 125]. Also, hantavirus infection–derived endothelial cell injury may directly cause platelet activation through the release of adhesive agents, such as fibrinogen, fibronectin and von Willebrand factor [23, 126]. Moreover, hantaviruses promote intravascular coagulation, leading to an increase in thrombin and fibrinolysis that additionally may cause thrombocytopenia due to increased platelet consumption [127]. Of note, clinical criteria of disseminated intravascular coagulation (DIC) are fulfilled frequently in hantavirus patients, posing risks for bleeding and thromboembolic complications [128-130]. Platelet activation and elevated levels of tissue factor and extravesicular tissue factor expressed by both endothelial and immune cells are reported to promote this systemic procoagulant state [131-133]. Although the clinical picture consisting of renal injury and thrombocytopenia might suggest thrombotic microangiopathy, no histological or laboratory evidence for this entity has been reported. Hemolysis as well as schistocytes are usually not present.

In human kidneys, hantaviruses primarily replicate in the endothelial cells of the renal medulla [134]. However, hantaviruses may additionally infect other cell types in the kidney, i.e. tubular cells, podocytes and the glomerular endothelium as detected in kidney biopsy specimens of infected patients [135]. Of note, the barrier function in all these cell types can be altered due to the breakdown of cell–cell contacts, which may be an explanation for the observed proteinuria during hantavirus infection [135, 136]. The detection of viral antigen along with inflammatory cell infiltrations in the peritubular area indicates that viral replication and the host's immune response together cause tissue damage in human disease [137, 138]. The renal immunopathology is primarily characterized by an upregulation of pro-inflammatory cytokines, such as type I interferons (IFNs) and interleukin-8 (IL-8) in the distal nephron activating both innate and adaptive immune cells [139-141]. The further systemic immune response is driven by macrophages and antigen-specific cells, including CD4+ and CD8+ T cells, as well as antibody-producing B cells [1, 3, 23]. Inflammatory cytokines and chemokines, i.e. transforming growth factor (TGF)-β1, tumour necrosis factor (TNF)-α, IL-6 and IL-10, play double-sided roles in human hantavirus infection. On the one hand, immunosuppressive TGF-β1 acts protectively in the late phase of PUUV infection by limiting tissue damage caused by the immune system [142]. On the other hand, TNF-α, IL-1 and IL-6 are associated with fever and septic shock as well as the further induction of acute-phase proteins and increased levels of those cytokines can be detected in urine, plasma and tissue samples of severely infected patients [115, 116, 142, 143].

In line with this notion, disease severity in PUUV and DOBV infections is associated with elevated serum levels of TNF-α, IL-6 and IL-10 [115, 116, 143, 144]. Elevated urinary IL-6 levels in PUUV-induced HFRS indicate local renal production of IL-6 in addition to the systemic immune response [115]. With regard to HCPS, elevated serum IL-6 levels are directly linked to fatal outcomes in humans [145]. Interestingly, IL-6 production is an overlapping characteristic of viral hemorrhagic fevers, as high levels of IL-6 have been linked to disease severity in Ebola virus infection, dengue virus disease and Crimean–Congo hemorrhagic fever [146-148].

Another remarkable parallel can be drawn to the cytokine release syndrome (CRS), a toxic immune reaction upon cancer treatment with chimeric antigen receptor–modified T cells that leads to a systemic increase in IL-6 and IL-10 [149]. Symptoms of CRS and hantavirus disease highly overlap—in mild cases, flu-like symptoms; in severe cases, vascular leakage, coagulopathy, hypotension, acute kidney injury (AKI) and pulmonary edema—and blockage of the IL-6 receptor with tocilizumab reverses CRS [150]. This raises the question whether hantavirus-infected patients could benefit from similar treatment. However in vivo animal and human data in hantavirus disease are lacking and the full causal roles of the highlighted mediators on local tissue damage remain largely unaddressed [145].

With regard to the adaptive immune system, hantaviruses increase the number of CD8+ T cells and reverse the CD4+:CD8+ T-cell ratio [3]. As a consequence, further pro-inflammatory cytokines are produced while regulatory cytokines are downregulated, enhancing the harmful effect of the immune system [151, 152]. Of interest, certain human leukocyte antigen (HLA) haplotypes are associated with severe disease courses in both HFRS and HCPS by triggering T-cell-mediated immune responses, especially of CD8+ T-cells [153-156]. This genetic vulnerability may explain interpersonal and regional differences in disease severity of both HFRS and HCPS in endemic areas [136]. For example, HLA alleles HLA-B*08, HLA-DRB1*0301 and HLA-DRB1*15 are associated with a severe form of PUUV infection, whereas HLA-B*27 has a benign prognosis [153, 157, 158]. In HTNV-caused HFRS, individuals with HLA-B*46 and HLA-B*46-DRB1*09 or HLA-B*51-DRB1*09 haplotypes are at a greater risk, whereas HLA-DRB1*12 is protective [159, 160]. With regard to HCPS, HLA-B*3501, HLA-DRB1*1402 and HLA-B*08 are negative prognostic factors in SNV- and ANDV-infected patients, respectively [156, 161]. In contrast, HLA-DRB1*15 and HLA-B*35-restricted memory T-cell responses are reported to be protective in ANDV-caused HCPS [161, 162]. Of note, different hantaviruses may be processed differently through the same HLA molecules, leading to either a mild or severe disease course as shown for HLA-DRB1*13 and HLA-B*35 in PUUV- and DOBV-caused HFRS [160, 163].

Small animal models recapitulating human hantavirus disease are scarce, as human pathogenic hantaviruses are maintained in nature by persisting rodent infection. For example, mice, rats and hamsters infected with HTNV, DOBV and PUUV fail to develop a human HFRS phenotype, whereas new born rodents that are infected within 3 days after birth die due to neurologic complications not recapitulating any disease characteristics resembling human disease [164]. The generation of HFRS animal models reflecting human hantavirus disease remains an urgent scientific need to fully understand hantaviral pathogenesis (Table 2). In contrast, Syrian hamsters infected with ANDV and SNV mimic human disease with regard to incubation time, rapid-progressing respiratory failure and pathological lung findings, but biosafety measurements make experimental design complex [165-167].

Clinical presentation

Common early symptoms in the prodromal phase of both HFRS and HCPS include myalgia, fatigue, abdominal pain, headache, high fever and other flu-like symptoms [168]. Although similar in pathogenesis, further observed clinical differences between HFRS and HCPS are the consequence of the primarily affected organs [3].

HFRS

HFRS is a generalized infection and its clinical course and outcome are dependent on the causative (Old World) hantavirus, although disease courses vary individually from subclinical to lethal.

The incubation phase of hantavirus ranges from 2 to 6 weeks [1]. Afterwards, the disease can be divided in five distinct phases: fever, hypotension, oliguria, polyuria and convalescence (Figure 5) [1, 3]. HFRS is characterized by the sudden onset of prodromal symptoms, such as myalgia, fatigue, abdominal pain, headache, high fever, blurred vision and other flu-like symptoms lasting 3–6 days. Somnolence is reported less frequently. The beginning of the hypotensive phase is characterized by hemorrhage (i.e. conjunctival suffusion, skin and mucosal petechiae, hematemesis, epistaxis, melaena, hematuria or even intracranial bleeding) due to thrombocytopenia, as well as hypotension or shock caused by vascular leakage. Of note, approximately one-third of case fatalities are related to an irreversible and fulminant shock [3]. Hemorrhage is also facilitated by thrombocytopenia, which is usually present in this stage and is the direct consequence of hantavirus-caused activation and depletion of platelets. The nadir in platelet count ranges from ∼70 × 109/L in PUUV to <30 × 109/L in DOBV- and HTNV-caused HFRS [46, 169, 170]. Bleeding events, typically reported as gastrointestinal bleeding, hematuria or hemoptysis are reported in ∼10% of HFRS cases. However, the incidence of bleeding events ranges from <5% in PUUV to 20% in DOBV and 35% in HTNV infections [171-173]. Microscopic hematuria is present in the vast majority of HFRS patients, whereas macroscopic hematuria is more frequently reported in DOBV and HTNV infections [174]. Hematuria usually derives from medullary or interstitial hemorrhages, i.e. is of intrarenal origin [175, 176]. DIC occurs in ∼30% of DOBV- or HTNV-infected patients as a consequence of an activated coagulation system and is a negative prognostic factor [127, 128, 177]. DIC is reported in ∼20% of PUUV patients according to the modified scoring system of the International Society of Thrombosis and Haemostasis; however, DIC is less severe in PUUV-caused disease [128, 177]. The hypotensive stage typically lasts 1–2 days and is followed by the often rapidly progressing oliguric AKI. With hemodynamic stabilization, renal function is eventually restored over a period of 3–7 days (Figure 5). The decrease in systemic blood pressure does not fully explain AKI, as a decline in kidney function may even occur without hypotension and measured blood pressures do not correlate with disease severity [96]. Intrarenal events—especially hantaviral-caused acute interstitial nephritis with cortical and peritubular capillaritis—are currently believed to contribute to the development of AKI; however, the exact mechanisms remain elusive and call for further research (Table 2) [96, 97, 176].

FIGURE 5:

The typical disease course in HFRS can be divided into five distinct phases: fever, hypotension, oliguria, polyuria and convalescence. After human infection, viral load peaks after 5–10 days and prodromal symptoms such as flu-like symptoms, myalgia, backache, abdominal pain and blurred vision occur. In parallel, platelets as well as urine output and kidney function decrease, leading to the hallmark triad of AKI, hypotension and hemorrhages in HFRS. With the onset of clinical symptoms, antibodies increase, leading to viral clearance and convalescence. Source: Figure created with Biorender.com and figure concept adapted from Avsic-Zupanc et al. [3].

Due to the intrarenal inflammatory processes, swelling of the kidneys is a hallmark of hantavirus-caused AKI, which is typically accompanied by—sometimes very severe—abdominal pain or backache. Proteinuria, sometimes in the nephrotic range, is also a typical finding in hantavirus-caused AKI and reflects the involvement of podocytes. The magnitude of proteinuria is associated with disease severity [178-180]. Renal replacement therapy (RRT) is needed in ∼5% of PUUV and 30% of both DOBV and HTNV infections, and half of HFRS deaths occur in this oliguric phase [3, 181–185]. Approximately 10% of DOBV- and HTNV-infected patients require mechanical ventilation due to the development of acute respiratory distress syndrome (ARDS) caused by acute progressive noncardiac pulmonary edema, whereas mechanical ventilation is rarely needed in PUUV-caused disease [168, 172, 186–188]. With the onset of the polyuric stage, kidney function gradually improves and urinary output may be increased for several weeks (Figure 5). Finally, convalescence, which is characterized by full recovery of kidney function, is usually prolonged and takes up to 3–6 months [189]. Of note, the risk for acute myocardial infarction, stroke and venous thromboembolisms is significantly elevated in this phase as a consequence of enhanced coagulation and inflammation after hantavirus disease [129, 130]. Although kidney function is usually completely restored, chronic kidney disease (CKD) and hypertension have been reported as rare long-term sequelae—robust data on the frequency and relative risk of CKD and RRT in HFRS (especially in DOBV- and HTNV-infected patients) are missing (Table 2) [172, 190–192].

PUUV-caused nephropathia epidemica is a mild form of HFRS that is the most common hantavirus disease in Europe. The typical disease stages are often not clearly distinguishable, as most patients suffer from less severe kidney failure that rarely requires RRT and hemorrhagic manifestations are usually mild [177, 181, 182]. PUUV-caused HFRS is often misdiagnosed due to its rather uncharacteristic clinical phenotype that resembles a febrile disease with abdominal pain [3, 193]. Different from other hantaviruses, SEOV is characterized by accompanying liver dysfunction during HFRS [194].

Children have a similar clinical presentation of HFRS as adults; however, the disease course is usually milder, with abdominal manifestations of hantavirus infection being more frequent [195, 196].

HCPS

The clinical phenotype of HCPS is characterized by ARDS, with case fatality rates of 30–50% [15, 197]. However, disease severity ranges from mild hypoxemia to fatal respiratory failure with precapillary cardiogenic shock [198].

The prodromal phase observed in HCPS is similar to HFRS and includes unspecific flu-like symptoms, including fever, chills, myalgia, nausea and vomiting. The disease course is characterized by progress to a cardiopulmonary stage with tachycardia, arrhythmia, shortness of breath and cough. Due to the vascular leakage in lung capillaries, patients rapidly develop noncardiac pulmonary edemas, oxygenation impairment and hypotension. Additionally, pulmonary infiltrates and pleural effusion may be detected. Patients with severe HCPS frequently require mechanical ventilation; often the disease course is further complicated by cardiogenic shock, lactic acidosis and hemoconcentration [3]. Therefore patients often die within hours after hospitalization when entering the fulminant cardiopulmonary phase [3]. When pulmonary edema resolves, patients develop polyuria and full recovery is achieved only after months [199].

With regard to kidney involvement in HCPS, proteinuria is frequently seen and is associated with mortality [200-202]. Moreover, AKI occurs in ∼15–20% of HCPS patients and RRT is needed in ∼10% of all HCPS patients [200, 203, 204]. The pathogenesis of kidney involvement in HCPS is currently unclear, as—due to the profound thrombocytopenia and DIC in the critically ill HCPS patient cohort—kidney biopsy samples are not available. A recent but small Brazilian autopsy study reported marked acute tubular necrosis. This notwithstanding, more pathological characterization of kidney phenotype in HCPS is needed [205]. Long-term kidney outcome in HCPS has been scarcely examined, mainly due to the low incidence and high mortality of the disease (Table 2). A small prospective study from the USA reported kidney sequelae after HCPS consisting of proteinuria and development of CKD in half of the reported cases [206].

Hantavirus disease—an interconnected syndrome beyond HFRS and HCPS

Renal and pulmonary diseases are often assigned to either Old World or New World hantaviruses in a dichotomous manner. However, HFRS and HCPS are defined and named after symptoms that also may be partly absent or incomplete. Recent advances in characterizing and understanding pathogenesis, organ tropism and clinical phenotype have confirmed common features of HFRS and HCPS and have therefore led to the perception of an interconnected disease [200, 207]. To this end, various reports indicate a large overlap of renal and pulmonary impairment in Old World and New World hantaviruses [168, 200, 206, 208–210]. For example, frequent development of ARDS is observed in HFRS caused by DOBV and HTNV [186–188, 211]. In PUUV infections, there are case reports that the human lung may be the primarily organ system affected, with PUUV being additionally detected in bronchoalveolar lavage (BAL) fluid [168, 208, 209, 212–215]. In HCPS-assigned SNV and ANDV, RRT is often needed for acute treatment [198, 203, 216]. In order to refine our knowledge on epidemiologic and clinical aspects of hantavirus disease and to determine disease progression beyond the dichotomous denominations of HFRS and HCPS—i.e., to define diagnostic criteria from large cohorts rather than case reports—a novel platform, the Hantavirus Registry (NCT04323904), was recently developed [217].

Diagnosis

The diagnosis of hantavirus-associated diseases is based on clinical findings, local epidemiology and laboratory methods. Hallmark laboratory findings of hantavirus infection are thrombocytopenia, leukocytosis, hemoconcentration, elevated serum creatinine levels, hematuria and proteinuria. Further laboratory markers depend on the other preferentially targeted organs [2].

Hantavirus-specific immunoglobulin M (IgM) and IgG antibodies are usually present in patients at the onset of symptoms. In contrast to IgG antibodies that persist lifelong, IgM titers are detectable in the acute phase of infection and decline over a period of 2–6 months (Figure 5) (3, 218–220). Therefore an IgM capture test is combined with an IgG test to detect acute infection. In Europe, hantavirus-specific diagnostic strategies should at least cover DOBV and PUUV. The serodiagnosis of hantavirus infection can be performed by enzyme-linked immunosorbent assay (ELISA) or immunoblot, predominantly detecting antibodies against N proteins [3, 221, 222]. ELISA is used in most commercially available serologic tests and employs recombinant protein antigens taken from DOBV, HTNV, PUUV, SEOV, SNV and ANDV (Table 4) [223]. The sensitivity and specificity for ELISA-based serodiagnosis are both ∼95% [223-225]. Also, immunoblots for DOBV, HTNV, PUUV and SEOV are commercially available and the reported sensitivity and specificity are 96% and 100%, respectively [223, 226]. Immunofluorescence assays (IFAs) examine the reaction of patient serum samples to uninfected or hantavirus-infected cells. IFAs have been most widely used in Europe and facilitate the detection of DOBV/HTNV and PUUV. The reported sensitivity and specificity are 98% and 91%, respectively [227]. Of note, results obtained from ELISAs should be confirmed by independent tests and in Europe both immunoblot and IFA are used for confirmation. Rapid immunochromatographic IgM antibody tests allow for point-of-care diagnosis but have not yet entered widespread use; they are also limited in sensitivity [228, 229].

Table 4.

Comparison of frequently used commercially available diagnostic approaches for hantavirus disease

Diagnostic test	Antigen type/hantavirus detected	Sensitivity (%)	Specificity (%)	Comments	References
ELISA	DOBVHTNVPUUVSEOVANDVSNV	95–97	94–99	Combination of IgG test and IgM capture test is recommended	[224, 225]
IFA	DOBVHTNVPUUV	98	91	Used as a confirmation test in Europe	[227]
Immunoblot assay	DOBVHTNVPUUVSEOV	96	100	Used as a confirmation test in Europe	[226]
Rapid immunochromatographic IgM antibody tests	DOBVHTNVPUUVSNV	80–93	96	Point-of-care test	[228, 229]
(Real-time)-PCR	Facilitates sequencing of the viral genome and the detection of novel hantaviruses	92–98	80–98	Time to test positivity <24 h Only useful in early viremic stage of infection	[265, 266]

The hantavirus genome can be detected by both traditional and real-time polymerase chain reaction (PCR)-based diagnostics in blood, saliva, BAL fluids and tissue samples. Of note, PCR-based diagnostics facilitate postmortem diagnosis and organ involvement. The viral load detected in blood samples at the onset of infection may even predict disease severity in both HFRS and HCPS; however, the viremic phase is limited to the very early stage of the disease (Figure 5) [230-233]. Another drawback of PCR-based diagnostics is the possibility of obtaining false-negative results in low-viremic infections often seen in, for example, PUUV or in patients tested late in disease [2]. Moreover, PCR-based diagnostics are prone to cross-contamination. Major advantages of molecular-based diagnostics are rapid test results obtained within 24 h, especially important in critically ill patients, and the ability to sequence the viral genome for phylogenetic analysis [223].

Prevention

Exposure to infected rodents is the main risk factor for hantavirus-associated diseases and, due to a lack of targeted antiviral therapy, preventive measures such as rodent control and avoiding contact with potentially infected areas are recommended to minimize risk of transmission [234-236]. A history of possible exposure to rodent excreta can be obtained in ∼50% of cases. Thus, identifying typical scenarios (e.g., sweeping out areas that are likely inhabited by rodents) is helpful but can certainly not be a prerequisite for making the diagnosis. Especially in endemic areas, hantavirus diseases should be suspected based on the clinical picture and laboratory findings rather than a history of exposure [236].

Vaccinations still remain controversial: in Korea, Hantavax is in use, a vaccine derived from formalin-inactivated HTNV-infected suckling mouse brain; however, frequent booster doses are mandatory for achieving immunity [237, 238]. Several formalin-inactivated vaccines have been used in China with unknown protective efficacies [238, 239]. No vaccines are currently approved in Europe or the USA. DNA vaccines for hantaviruses that may facilitate construction of multivalent vaccines and long-lasting immunity are being tested in clinical trials [240, 241]. Based on the recent experiences in the COVID-19 pandemic, RNA vaccines may actually be a very promising strategy to extend to hantaviruses.

Therapy

Currently treatment of both HFRS and HCPS is mainly supportive, as there is no specific therapy available [242]. For close monitoring, patients with a severe disease course should be admitted to the intensive care unit to maintain euvolemia and electrolyte balance. Especially in anuric patients, the volume status should be closely monitored to avoid extensive fluid retention and pulmonary edema due to leaky capillaries. RRT is needed in 4–6% of the hospital-treated patients in PUUV, in 10–20% in DOBV and up to 40–50% in HTNV infections [170, 179, 182, 183, 193, 243–245]. In the case of severe thrombocytopenia and major bleeding events due to DIC, platelet transfusion and/or fresh frozen plasma may be necessary to improve coagulation and control the bleeding event [3, 246]. Of note, there are hints that the extent of thrombocytopenia at the onset of infection predicts AKI severity as well as the clinical course in HFRS patients [170, 247]. In HCPS patients, supportive therapy consists of oxygen supplementation, mechanical ventilation when necessary, inotropic support and maintenance of euvolemia [199]. Additionally, extracorporeal membrane oxygenation improves outcome in patients with refractory shock and ARDS [248, 249].

Currently there is no targeted antiviral therapy available in Europe or the USA. Ribavirin has shown some antiviral effects against hantaviruses in in vitro models of HTNV-caused HFRS, since it is inhibiting viral replication through targeting the RdRp [250, 251]. In humans, ribavirin has been tested in a double-blind, placebo-controlled clinical trial in China, suggesting improved outcome and reduced severity of renal insufficiency when administered within 5 days after the onset of symptoms in HTNV-caused HFRS (Table 5) [252, 253]. In contrast, a randomized, open-label Russian trial revealed that ribavirin-associated side effects such as rash, sinus bradycardia, hyperbilirubinemia and anemia were significantly increased, while showing no protective effects in PUUV-infected patients [254]. Of note, disease severity in PUUV-caused HFRS is—in contrast to HTNV —not associated with the viral load, which may be a reasonable explanation for these different outcomes (Table 5) [255-257]. Ribavirin has also been examined in HCPS patients in a prospective, double-blind, placebo-controlled trial revealing no beneficial effects. Currently there is no recommendation to use ribavirin in HFRS or HCPS [258, 259].

Table 5.

Comparison of clinical trials examining ribavirin in hantavirus disease

Characteristics	Huggins et al. [252]	Malinin et al. [254]	Mertz et al. [258]
Disease entity	HFRS	HFRS	HCPS
Study period	1985–1987	2004–2005	1996—2001
Trial design	Prospective, double-blind, placebo-controlled trial	Prospective, open-label, phase II study	Prospective, double-blind, placebo-controlled trial
Number of patients	293	73	36
Dose of ribavirin	Loading dose of 33 mg/kg, followed by a dose of 16 mg/kg every 6 h for the first 4 days and 8 mg/kg every 8 h for the subsequent 3 days	Loading dose of 33 mg/kg, followed by a dose of 16 mg/kg every 6 h for the first 4 days and 8 mg/kg every 8 h for the subsequent 3 days	Loading dose of 33 mg/kg, followed by a dose of 16 mg/kg every 6 h for the first 4 days and 8 mg/kg every 8 h for the subsequent 3 days
Timing of ribavirin with respect to onset of infection	4 days (lengthened to 6 days in 1986) after onset of symptoms	4 days after onset of symptoms	Not specified; however, patients had to be in the prodromal or cardiopulmonary stage
Hantaviruses involved	HTNV (confirmed in 82.6% by ELISA)	PUUV	SNV (confirmed in 63.9% by ELISA)
Primary endpoint	Reduction in mortality, occurrence of oliguria and hemorrhages	Change in viral load	Survival at day 28 after study entry
Results for primary endpoint	7-fold reduction of mortality in the ribavirin group (P = .01)3.7 reduction of oliguria in the ribavirin group (P = .01)	Insufficient efficacy of ribavirin	No difference in survival between the ribavirin and placebo group
Adverse events	Drug-related anemia in all male study subjects (males accounted for 75% of study subjects). Females showed similar trends that were less dramatic due to sex-related differences in hematocrit	Low hemoglobin levels in 95%, hyperbilirubinemia in 81%, sinus bradycardia in 43% and rash in 19% of the ribavirin-treated patients	No significant differences in the frequency of adverse events; however, there was trend toward a higher rate in anemia in the ribavirin group
Inclusion criteria	Age ≥14 years Fever duration ≤4 days (lengthened to 6 days in 1986) Clinical diagnosis of HFRS including fever and proteinuria History making exposure to infection likelyOR Findings consistent with early HFRS Hantaviral IgM antibodies No other evidence for an alternative diagnosis	Age 18–65 years Suspected diagnosis of HFRS within 4 days of onset of disease SOFA score = 1	Age ≥12 years Suspected or serologically confirmed acute hantavirus disease in the prodromal of cardiopulmonary stage
Exclusion criteria	Advanced renal failure manifested by oliguria or uremia Pregnancy or breast feeding Known intolerance to ribavirin Moribund on presentation or life expectancy <48 h Pre-existing non-HFRS life-threatening condition	Known intolerance to ribavirin Pregnancy or breast feeding NYHA cardiac function ≥2 History of severe chronic pulmonary or kidney disease History of autoimmune hepatitis Serum aminotransferase levels greater than two times the upper limit of normal Hemoglobin level <12 g/dL	Pregnancy or breast-feeding A likely diagnosis other than HCPS Immunocompromised status Receipt of systemic corticosteroids within 30 days prior to enrolment A mean arterial pressure of <60 mmHg for 2 h despite optimal medical management A cardiac index <2.1 L/min/m2 Arterial oxygen pressure <65 mmHg in intubated subjects receiving 100% oxygen The presence of unilateral pulmonary infiltrates that did not become bilateral within 24 h
Comments	Study showed efficacy in reducing case fatality and oliguria in HTNV-infected patients	Study revealed insufficient efficacy and safety of intravenous ribavirin in PUUV-infected patients. Severity of PUUV-caused HFRS is not associated with viral load, in contrast to HTNV, explaining the different outcomes observed [255–257]	Premature termination of the study due to the slow rate of accrual of subjects and the findings of futility analysis

NYHA: New York Hearth Association; SOFA: Sepsis-related Organ Failure Assessment score.

Apart from ribavirin, methylprednisolone has been tested in a randomized controlled clinical trial in ADNV-infected patients, failed to provide any significant benefit to patients (Table 6) [204]. Novel potential therapeutic strategies that were tested in noncontrolled clinical trials include murine and human-derived neutralizing antibodies that block viral entry. In contrast to murine-derived neutralizing antibodies, for which efficacy is unknown, human-derived neutralizing antibodies resulted in a 50% reduction of case fatalities in ANDV infections (Table 6) [260-262]. In addition, bradykinin type 2 receptor antagonists have been tested in PUUV-infected patients and may be promising candidates [107, 111, 112, 263]. However, standardized, large-scale clinical trials examining both efficacy and safety are lacking [238].

Table 6.

Summary of potential antiviral approaches for hantavirus disease tested in humans

Drug	Known /putative target	Purpose	Virus	Number of patients	Outcome/ comments	References
Murine monoclonal antibodies	Gc/Gn	Blocking viral entry	n.a.	22	Safety in a phase I trial examined, proof of efficacy lacking	[261]
Human immune plasma	Gc / Gn	Blocking viral entry	ANDV	32	Safe and efficient with a 50% reduction in case fatalities in an uncontrolled clinical trial	[267]
Icatibant	Bradykinin receptor 2	Improving vascular function	PUUV	Case reports	Clinical trials apart from case reports lacking	[110–112]
Methylprednisolone	Immunotherapy	Rebuilding immune homeostasis	ANDV	66	No beneficial effect reported	[204]
Ribavirin	RdRp	Inhibiting viral replication	ANDV, HTNV, PUUV	547	Efficient in HTNV, inefficient and unsafe in PUUV and ANDV (see Table 5)	[252, 254, 258, 259]

Gc: C-glycoprotein; Gn: N-glycoprotein; n.a.: not applicable.

CONCLUSION

Hantavirus-associated diseases are emerging zoonotic infections with increasing incidence rates in Europe. The distribution of hantaviruses is influenced by climate change and disturbed rodent habitats and new hantavirus species with unknown pathogenicity are encountered in these reservoirs. Recently discovered human-to-human transmission by superspreading events may further cause hantaviral spread and make a pandemic a possible scenario. Therefore further research in hantavirus pathogenesis, diagnosis antiviral and preventive measures, including vaccine development, remain mandatory.