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The prodromal phase of HPS is indistinguishable clinically from numerous other viral infections. Often the only guide to the etiology of the patient's illness is the blood picture, which may show circulating immunoblasts, which appear as large atypical lymphocytes, and thrombocytopenia. However, unlike other viral infections, HPS patients usually have concurrent left-shifted neutrophilia with circulating myelocytes.

In the cardiopulmonary stage of the disease, the patients have a diffuse pulmonary edema. The most frequent cause for such a picture is silent myocardial infarction so it is important to obtain an ECG and echocardiogram early to aid in the assessment. Intensivists at the University of New Mexico, where many of the patients have been managed, have found that a echocardiogram also helps to distinguish these patients from patients with ARDS as cardiac function is depressed to a much greater degree in the HPS patients and cardiac output does not respond to fluid challenge as it tends to with ARDS.

Infections in the immunocompetent which might present with a non-specific prodrome leading to acute cardiopulmonary deterioration as in HPS include leptospirosis, Legionnaire's disease, mycoplasma, Q fever, chlamydia, and in regions where the organisms are present, septicemic plague, tularemia, coccidioidomycosis and histoplasmosis. Non-infectious conditions such as Goodpasture's syndrome should also be considered. Lack of coryza aids the clinical distinction between HPS and Influenza A infection.

It must be remembered that HPS is relatively uncommon and in the immunocompromised PCP, CMV, cryptococcus, aspergillus and graft vs. host disease are far more likely to be the cause of diffuse pulmonary infiltrates than a hantaviral infection.

Microbiology

Like other members of the bunyavirus family, hantaviruses are enveloped viruses with a genome that consists of three single-stranded RNA segments designated S (small), M (medium), and L (large). All hantaviral genes are encoded in the negative (genome complementary) sense. The S RNA encodes the nucleocapsid (N) protein. The M RNA encodes a polyprotein that is cotranslationally cleaved to yield the envelope glycoproteins G1 and G2. The L RNA encodes the L protein, which functions as the viral transcriptase/replicase. Within virions, the genomic RNAs of hantaviruses are thought to complex with the N protein to form helical nucleocapsids, which circularize due to sequence complementarity between the 5' and 3' terminal sequences of each genomic segment.

Hantaviruses replicate exclusively in the host cell cytoplasm. Entry into host cells is thought to occur by attachment of virions to cellular receptors and subsequent endocytosis. Nucleocapsids are introduced into the cytoplasm by pH-dependent fusion of the virion with the endosomal membrane. Transcription of viral genes is initiated by association of the L protein with the three nucleocapsid species. In addition to transcriptase and replicase functions, the viral L protein is also thought to have an endonuclease activity that cleaves cellular messenger RNAs (mRNAs) for the production of capped primers used to initiate transcription of viral mRNAs. As a result of this "cap snatching," the mRNAs of hantaviruses are believed to be capped and contain nontemplated 5' terminal extensions. The viral N and L mRNAs are thought to undergo translation at free ribosomes, whereas the M mRNA is translated in the endoplasmic reticulum. G1 and G2 glycoproteins form heterodimers and are then transported from the endoplasmic reticulum to the Golgi complex, where glycosylation is completed. The L protein produces nascent genomes by replication via a positive-sense RNA intermediate. Hantavirions are believed to form by association of nucleocapsids with glycoproteins embedded in the membranes of the Golgi, followed by budding into the Golgi cisternae. Nascent virions are then transported in secretory vesicles to the plasma membrane and released by exocytosis.

Other Hantaviruses

Several members of the hantavirus genus cause different forms of hemorrhagic fever with renal syndrome (HFRS), an ancient disease first described in Russia in 1913. The four viruses that are associated with HFRS, each named for the region from where they were first isolated, have different primary rodent hosts: Apodemus agrarius (the striped field mouse) for Hantaan virus, Rattus norvegicus (the Norway rat) and Rattus rattus (the black rat) for Seoul virus, Clethrionomys glareolus (the bank vole) for Puumala virus, and Apodemus flavicollis (the yellow-necked field mouse) for Dobrava virus. Hantaan virus from Korea and Dobrava virus from Slovenia are associated with a severe form of HFRS characterized by renal failure that can precede pulmonary edema and disseminated intravascular coagulation (DIC), with estimated mortality rates of 5% to 15%. A moderate form of HFRS caused by Seoul virus (which, along with its host, is distributed worldwide) is responsible for thousands of Eurasian cases annually. Serologic evidence for infection with Seoul-like hantaviruses has been found in rodents in major cities of the United States, and this virus was recently implicated in human cases of HFRS in Baltimore. One report has also associated Seoul virus with chronic renal disease. A mild form of HFRS, caused by Puumala virus, is responsible for nephropathia epidemica in Scandinavia, with an estimated mortality rate of 1% to 3%.

Genetics

Sin nombre virus sequencing

Polymerase chain reaction (PCR) flow chart

The entire genomic sequence of SNV has subsequently been determined by using RNA extracted from autopsy material as well as RNA extracted from cell culture-adapted virus. The L RNA is 6562 nucleotides (nt) in length; the M RNA is 3696 nt long; and the S RNA is 2059 to 2060 nt long. Interestingly, when the prototype sequence (NMH10) of SNV detected in tissues from an HPS case was compared with the sequence of the SNV isolate (NMR11; isolated in Vero E6 cells from P. maniculatus trapped in the residence of the same case), only 16 nucleotide changes were found, and none of these changes resulted in alterations in amino acid sequences of viral proteins. It had been assumed that in the process of adaptation to cell culture, selection of SNV variants which grow optimally in cell culture would occur, and selected variants would differ genetically from the parental virus. Though NMH10 and NMR11 are identical in protein sequence, nucleotide substitutions in nontranslated regions of the genome could be responsible for altered viral phenotypes, as could changes in protein glycosylation or virus membrane components.

The nested RT-PCR assay developed during the initial HPS outbreak provided a rapid method for the genetic characterization of novel hantaviruses that did not require a virus isolate. Numerous new hantaviruses have been detected by RT-PCR in rodent tissues but have yet to be associated with human disease. These include El Moro Canyon virus associated with the western harvest mouse (Reithrodontomys megalotis), Tula virus with Microtus arvalis and M. rossiaemeridionalis, Rio Segundo virus with the Mexican harvest mouse (R. mexicanus), Isla Vista virus with the California vole (M. californicus), and Prospect Hill-like viruses in Microtus species.

Phylogenetic Analysis

Phylogenetic analysis of Old World and American hantaviruses indicates that the relationship among hantaviruses corresponds with the phylogeny of their rodent hosts. Viruses of rodents belonging to the subfamily Murinae are monophyletic as are hantaviruses of arvicoline and sigmodontine rodents, suggesting that long-term virus-rodent coevolution is taking place. Hantavirus evolution is best understood as co-evolution within specific lineages in the rodent family Muridae. The apparent coupling between hantaviruses and their rodent hosts suggests that viruses of sigmodontine rodents share a common ancestor, as do viruses of the subfamily Arvicolinae and Murinae. This coupling also has a geographic and clinical correlate: viruses found in Old World murine rodents, including Hantaan virus (HTNV), Seoul virus (SEOV) and Dobrava virus, are associated with HFRS in Eurasia. By contrast, viruses carried by New World sigmodontine rodents, including SNV Black Creek Canal virus (BCCV) and Bayou virus (BAYV), are associated with HPS in the Americas. This distinction can narrow the search for a rodent host for newly discovered HPS-like diseases and suggest disease implications for the various new viruses being genetically amplified from rodents.

Sequence Divergence

Detection and characterization of Sin Nombre-like viruses in P. maniculatus and P. leucopus populations have shown that multiple phylogenetic lineages of SNV exist in North America, and in some cases similar viruses are detected in both Peromyscus species. Sequence divergence among SNV genes has been shown to be as high as 23% nucleotide dissimilarity and 7% amino acid dissimilarity. Comparison of SNV sequences with those of other hantaviruses provides no obvious explanation as to why SNV and related viruses cause HPS while other hantaviruses are associated with HFRS. The development of a reverse genetics system for manipulation of virus genomes and an animal model for studying pathogenesis will be necessary to define the molecular mechanism(s) of SNV pathogenicity.

Genomic reassortment by RNA viruses with segmented genomes is well documented and has the potential to produce viruses with altered biological activity, host range, and disease potential. For example, segment reassortment among influenza virus strains (antigenic shift) is thought to be responsible for influenza pandemics. Genomic reassortment among SNV variants is known to occur in nature, but the precise role of genomic reassortment in the epidemiology of HPS and HFRS is unknown.

Pathogenesis

Following inhalation exposure to hantavirus in rodent excreta, there is an incubation period of 9 to 33 days (median of 14-17 days, and up to 3 weeks after rodent bite), [47] [48] during which the virus replicates in pulmonary macrophages and dendritic cells without causing cell death and is distributed to lymphoid organs. [49] A viremia is produced that infects target endothelial cells and stimulates T cells. Host neutralizing antibody is produced that may mitigate the severity of the infection. [50] Immune-stimulated T cells are distributed to areas of hantavirus concentration, particularly the lung and heart interstitium. [51] A nonspecific viral syndrome with predominantly severe myalgias and GI symptoms follows that is often not recognized. Rarely, the illness does not progress; [52] however, usually patients are admitted to the hospital within 5 days having progressive respiratory failure with pulmonary edema, metabolic acidosis, and cardiogenic shock.

Progressive disease results from infected endothelial cell function being disturbed and local T-cell cytokine production, which causes capillary leak in target organs. [53] [54] The capillary leak may be the result of pathogenic hantaviruses using an alpha-variable, beta-3 integrin ligand on the endothelial cell and platelet surfaces, resulting in disturbed endothelial cell migration and endothelial cell barrier function. [49] [55]

Recent studies in a Syrian hamster HPS model have suggested a direct effect of Hantavirus on resting platelets through binding to the platelet beta-3 integrin with resultant platelet cross-linking and clumping on endothelial surfaces. There is also binding to the beta-3 integrin on the resting endothelial cells with disruption of vascular endothelial growth factor receptor activity, increased phosphorylation and internalization of vascular endothelial-cadherin, and loss of cellular tight junction competence. [56]

All pathogenic hantaviruses have also been demonstrated to have an immunoreceptor tyrosine-based activation motif (ITAM) on their G1 envelope glycoproteins, which may modulate cellular downstream signaling and endothelial and immune cell function. [57] Sensitized mononuclear cells infiltrate the lung, myocardial interstitium, and spleen to produce cytokines, particularly TNF-alpha and interferon-gamma, resulting in pulmonary edema and myocarditis. [58] [59] Hantaviruses may also attach to beta-2 integrin receptors on neutrophils and induce the release of neutrophil extracellular traps. [60]