NiV Pathology

FIGURE 4.2 Histopathological changes during NiV infection in human lung xenografts, (a) Human lung with focal areas of necrosis and syncytia formation (arrow) on day 3 post infection (H&E, 10x magnification), (b) Bronchi with syncytia formation (*) on day 3 post infection (H&E, 20x magnification), (c) Pulmonary vasculature with syncytial formation (*) on day 3 post infection (H&E, 40x magnification), fibrinoid necrosis of the intima, and influx of granulocytes (arrowhead). (Photo credit: Valbuena G, Halliday H, Borisevich V, Goez Y, Rockx B. PLoS Pathog 2014;10(4):e1004063.)

apoptosis, necrosis, fibrinoid necrosis, and inflammatory cell infiltration may ensue NiV infection of the endothelial cells. Perivascular cuffing is occasionally seen, with infiltration of neutrophils, monocytes, or occasionally lymphocytes. Organs with abundant expression of ephrin-B2, such as the lungs, kidneys, and the heart, show prominent vascular inflammatory changes. Spleen and other highly vascular organs may show focal perivascular necrosis.⁵⁴

The oronasal and respiratory epithelia appear to be the primary sites of NiV replication during natural infections in cats and pigs, with subsequent spread of the virus to the vascular and lymphoreticular systems. The smooth muscle cells in the tunica media of the blood vessels are permissive to NiV infection, with subsequent spread of infection occurring to organs including the brain, lungs, and the spleen by hematogenous route. The viremia is thought to be of low level, and its exact duration is unclear. NiV also specifically targets the lymphoreticular system, and lymphoid depletion and necrosis have been reported in studies of infected humans, swines, and cats.

Hematogenous dissemination of NiV often results in infection of the kidneys and urinary tract. Infected bats often show intermittent shedding of NiV in their urine.⁵⁵ In contrast, urinary shedding of NiV is not usually observed during natural infections in pigs. During the NiV outbreak in Malaysia, NiV was isolated from 3 of 20 patients with a confirmed diagnosis of NiV infection.⁵⁶ Histopathological studies in experimentally infected guinea pigs showed lymphohistiocytic vasculitis, necrosis, and ulceration of transitional epithelium and occurrence of syncytial cells, in the urinary bladder and mild vasculitis and syncytial cells in kidneys. Fogarty et al.⁵⁷ showed that NiV has a half-life of about 17.8 h in the urine (pH about 7) of Pteropus vampyrus bats at a temperature of 22°C, which dropped to 1.8 h at 37°C. In urine samples from the same species, the virus half-life decreased to less than 0.5 h at both these temperatures. The patterns of urinary shedding of NiV in naturally infected hosts and its roles in transmission of infection warrant detailed investigations.

A significant association with consumption of the sap of date palm tree (Phoenix sylvestris), a local delicacy, was observed in several recent outbreaks of NiV disease in Bangladesh and India.²⁵-⁵⁸-⁶¹ It is known that fruit bats frequently raid the clay pots used for the collection of the date palm sap in the affected regions and contaminate the contents with their saliva, urine, or feces. The near-neutral pH of date palm sap may maintain the viability of NiV, and transmission can occur if the contaminated product is consumed raw, within a few hours of collection. In addition to this, practices such as the repeated use of the same collection pot without cleaning and the pooling of sap from multiple sources increase the risk for contamination of sap with virus-laden bat secretions.⁶²

Considerable ambiguity exists about the ability of NiV to infect cells of the gastrointestinal epithelia, which lack expression of ephrin-B2 and -B3. The types of cells in the gastrointestinal tract, which are susceptible to NiV and the mechanisms of viral entry and dissemination from them, remain unclear. It is probable that the virus might enter the tissue-resident macrophages and/or dendritic cells through microscopic lesions that commonly occur in the mouth and throat. Such opportunities for entry into macrophages may also be provided by inflamed tonsils, diseased gingival tissue, etc. The role of comorbidities, such as acid reflux disease, gastric or duodenal ulceration, malnutrition, etc., in modulating the risk of NiV infection of gastrointestinal epithelia following its ingestion is also unknown.

The virus can also cross the placental barrier to infect the developing fetus in animals like bats, guinea pigs, and cats.⁵ This may contribute to the maintenance of NiV in its natural reservoir species.

NiV also shows considerable lymphoid tropism and may cause disruption in the architecture of lymphoid tissue and lymphoid necrosis. These appear to indicate that NiV infection may cause immunosuppression in the infected host.⁵

Species-specific differences are observed in the organ targeted by NiV in natural infections. Brain is invariably involved in NiV infections in several species and immunohistochemical staining reveals the viral antigen distribution in vascular endothelial cells, surrounding smooth muscles of tunica media, neurons, glial cells, ependymal cells, and the epithelial cells of the choroid plexus. Significant pathology is also seen in lungs in natural NiV infections of cats, pigs, and humans. Borisevich et al.⁶³ in a recent in vitro study reported the susceptibility of human primary olfactory epithelial cells to infection by NiV-Malaysia and NiV-Bangladesh strains. Both the strains replicated efficiently in the olfactory epithelial cells and induced progressive syncytiation and cell death. Notably, immunohistochemical staining revealed the presence of viral antigen only in olfactory sensory neurons in the olfactory epithelium. These facts indicate that human olfactory epithelium may serve as a point of entry of NiV into the CNS. NiV can enter the CNS through disruptions in the blood-brain barrier (BBB), infected microvascular endothelial cells, or a combination of both. Breaches in the BBB resulting from extensive inflammatory changes may facilitate the direct entry of the virus into the CNS and infection of cells expressing ephrin-B2/B3. In the alternate pathway, infected brain microvascular endothelial cells can release virions through the basolateral surface, which can subsequently infect other cells, even with an intact BBB. Weise et al.⁶⁴ have reported the presence of basolateral-targeting signals in the cytoplasmic domains of G and F proteins of NiV.

Primary epithelial cell monolayers derived from bronchi and small airways also exhibit high susceptibility to NiV infection. Notably, NiV-induced inflammation occurred predominantly in the small airways but not the bronchi.⁶⁵ Bronchiolar epithelial cells are infected early in human NiV infections, and virus shedding occurs in nasopharyngeal and tracheal secretions.⁵⁶ NiV can replicate to high titers in cells of the lower respiratory tract. There is a greater chance of aerosol transmission of the infection from patients with a symptomatic respiratory infection. Histopathological changes consistent with necrotizing alveolitis, hemorrhage, pulmonary edema, and aspiration pneumonia are often evident. The other findings include multinucleated giant cells in the alveolar septum and alveolar spaces, as well as intra-alveolar inflammatory cells. Infected airway epithelia may release inflammatory cytokines including IL-6, IL-8, IL-la, MCP-1, G-CSF, and CXCL-10, and the recruitment of immune cells by these may produce acute respiratory distress syndrome (ARDS). NiV infection is also known to induce the IFN-(3 and IP-10.^66-68 With progressive infection, the viral replication spreads from respiratory epithelia to the endothelia of lungs and causes prominent pathology in small vessels and capillaries. Subsequent viremic spread can lead to multiorgan failure.

NiV-Malaysia and NiV-Bangladesh seem to be associated with subtle differences in virulence properties despite a nucleotide sequence similarity of about 91.8%. In comparison to NiV-М, the NiV-В shows a shorter incubation period as well as a narrower range for it. Respiratory symptoms predominated in most cases with NiV-В infections, while only a few of those with NiV-М infections experienced them. Myoclonus was reported in only a few of the Bangladeshi and Indian patients infected with NiV-В, while a significant number developed it during infection with NiV-М. Some of the fatal cases during the NiV outbreak caused by NiV-М in the Philippines presented with acute encephalitis syndrome.⁸

Clinical Features of Human NiV Infection

The incubation period in most human cases is about 2 weeks or less (ranging from 4 days to 2 months).⁶-^[1] After a febrile prodrome with a headache, dizziness, and vomiting, there was a rapid progression to severe encephalitis, with altered sensorium, signs of brainstem dysfunction (e.g., abnormal doll’s eye reflex and papillary responses) vasomotor disturbances, seizures, and myoclonus), cerebellar signs, etc. The spectrum of presentations included aseptic meningitis, diffuse encephalitis, and focal brainstem involvement. Respiratory disturbances developed in 14%-20% of cases in the Malaysian outbreak and two cases presented with respiratory manifestations alone in the Singapore outbreak. Respiratory involvement was prominently observed during the outbreaks of NiV infection in Bangladesh and India, with a few cases of acute respiratory distress syndrome.

Magnetic resonance imaging often reveals multiple hyperintense lesions in the cortex, pons, puta- men, and cerebral and cerebellar peduncles in NiV infections in humans. This is thought to correspond to the diffuse expression of ephrin-B2 and often correlates with the neurological symptoms seen. In contrast, the expression of ephrin-B3 is restricted to the brainstem and spinal cord. Brainstem dysfunction carries a bad prognosis in patients with Nipah encephalitis and is a frequent cause of death.^[1]

A relapse of the infection and late-onset encephalitis occurred in many patients who survived an initial infection with NiV, months to years later.⁶ Psychiatric disturbances, including depression, personality changes, and persistent neurocognitive deficits, were reported in a significant proportion of the cases from Malaysia and Singapore.^{[3] [4]}-^[5]

• [1] A clinically effective antiviral against NiV infections in humans is still lacking. Several approaches have
• [2] A clinically effective antiviral against NiV infections in humans is still lacking. Several approaches have
• [3] been employed in developing an efficacious therapy against NiV, with varying success. Inhibition of viralentry has been explored as an antiviral strategy against NiV, since the NiV G protein exhibits a high affinity
• [4] for its cognate receptor, ephrin-B2/B3. In in vitro experiments, soluble forms of NiV G protein showed potent
• [5] inhibition of virus entry.33-5 This may work as a form of passive immunotherapy, which blocks acute viralreplication and generates protective antibody responses. Fusing the G moiety to Fc region of human immu