Lindy Liu

2009 Pandemic Influenza A (H1N1): Pathology and Pathogenesis of 100 Fatal Cases in the United States

In the spring of 2009, a novel influenza A (H1N1) virus emerged in North America and spread worldwide to cause the first influenza pandemic since 1968. During the first 4 months, over 500 deaths in the United States had been associated with confirmed 2009 pandemic influenza A (H1N1) [2009 H1N1] virus infection. Pathological evaluation of respiratory specimens from initial influenza-associated deaths suggested marked differences in viral tropism and tissue damage compared with seasonal influenza and prompted further investigation. Available autopsy tissue samples were obtained from 100 US deaths with laboratory-confirmed 2009 H1N1 virus infection. Demographic and clinical data of these case-patients were collected, and the tissues were evaluated by multiple laboratory methods, including histopathological evaluation, special stains, molecular and immunohistochemical assays, viral culture, and electron microscopy. The most prominent histopathological feature observed was diffuse alveolar damage in the lung in all case-patients examined. Alveolar lining cells, including type I and type II pneumocytes, were the primary infected cells. Bacterial co-infections were identified in >25% of the case-patients. Viral pneumonia and immunolocalization of viral antigen in association with diffuse alveolar damage are prominent features of infection with 2009 pandemic influenza A (H1N1) virus. Underlying medical conditions and bacterial co-infections contributed to the fatal outcome of this infection. More studies are needed to understand the multifactorial pathogenesis of this infection.

Myocardial Injury and Bacterial Pneumonia Contribute to the Pathogenesis of Fatal Influenza B Virus Infection

BACKGROUND Influenza B virus infection causes rates of hospitalization and influenza-associated pneumonia similar to seasonal influenza A virus infection and accounts for a substantial percentage of all influenza-related hospitalizations and deaths among those aged <18 years; however, the pathogenesis of fatal influenza B virus infection is poorly described. METHODS Tissue samples obtained at autopsy from 45 case patients with fatal influenza B virus infection were evaluated by light microscopy and immunohistochemical assays for influenza B virus, various bacterial pathogens, and complement components C4d and C9, to identify the cellular tropism of influenza B virus, characterize concomitant bacterial pneumonia, and describe the spectrum of cardiopulmonary injury. RESULTS Viral antigens were localized to ciliated respiratory epithelium and cells of submucosal glands and ducts. Concomitant bacterial pneumonia, caused predominantly by Staphylococcus aureus, was identified in 38% of case patients and occurred with significantly greater frequency in those aged >18 years. Pathologic evidence of myocardial injury was identified in 69% of case patients for whom cardiac tissue samples were available for examination, predominantly in case patients aged <18 years. CONCLUSIONS Our findings suggest that bacterial pneumonia and cardiac injury contribute to fatal outcomes after infection with influenza B virus and that the frequency of these manifestations may be age related.

Cytokine and Chemokine Profiles in Lung Tissues from Fatal Cases of 2009 Pandemic Influenza A (H1N1): Role of the Host Immune Response in Pathogenesis

Pathological studies on fatal cases caused by 2009 pandemic influenza H1N1 virus (2009 pH1N1) reported extensive diffuse alveolar damage and virus infection predominantly in the lung parenchyma. However, the host immune response after severe 2009 pH1N1 infection is poorly understood. Herein, we investigated viral load, the immune response, and apoptosis in lung tissues from 50 fatal cases with 2009 pH1N1 virus infection. The results suggested that 7 of the 27 cytokines/chemokines showed remarkably high expression, including IL-1 receptor antagonist protein, IL-6, tumor necrosis factor-α, IL-8, monocyte chemoattractant protein-1, macrophage inflammatory protein 1-β, and interferon-inducible protein-10 in lung tissues of 2009 pH1N1 fatal cases. Viral load, which showed the highest level on day 7 of illness onset and persisted until day 17 of illness, was positively correlated with mRNA levels of IL-1 receptor antagonist protein, monocyte chemoattractant protein-1, macrophage inflammatory protein 1-β, interferon-inducible protein-10, and regulated on activation normal T-cell expressed and secreted. Apoptosis was evident in lung tissues stained by the TUNEL assay. Decreased Fas and elevated FasL mRNA levels were present in lung tissues, and cleaved caspase-3 was frequently seen in pneumocytes, submucosal glands, and lymphoid tissues. The pathogenesis of the 2009 pH1N1 virus infection is associated with viral replication and production of proinflammatory mediators. FasL and caspase-3 are involved in the pathway of 2009 pH1N1 virus-induced apoptosis in lung tissues, and the disequilibrium between the Fas and FasL level in lung tissues could contribute to delayed clearance of the virus and subsequent pathological damages.

Exserohilum Infections Associated with Contaminated Steroid Injections: A Clinicopathologic Review of 40 Cases

September 2012 marked the beginning of the largest reported outbreak of infections associated with epidural and intra-articular injections. Contamination of methylprednisolone acetate with the black mold, Exserohilum rostratum, was the primary cause of the outbreak, with >13,000 persons exposed to the potentially contaminated drug, 741 confirmed drug-related infections, and 55 deaths. Fatal meningitis and localized epidural, paraspinal, and peripheral joint infections occurred. Tissues from 40 laboratory-confirmed cases representing these various clinical entities were evaluated by histopathological analysis, special stains, and IHC to characterize the pathological features and investigate the pathogenesis of infection, and to evaluate methods for detection of Exserohilum in formalin-fixed, paraffin-embedded (FFPE) tissues. Fatal cases had necrosuppurative to granulomatous meningitis and vasculitis, with thrombi and abundant angioinvasive fungi, with extensive involvement of the basilar arterial circulation of the brain. IHC was a highly sensitive method for detection of fungus in FFPE tissues, demonstrating both hyphal forms and granular fungal antigens, and PCR identified Exserohilum in FFPE and fresh tissues. Our findings suggest a pathogenesis for meningitis involving fungal penetration into the cerebrospinal fluid at the injection site, with transport through cerebrospinal fluid to the basal cisterns and subsequent invasion of the basilar arteries. Further studies are needed to characterize Exserohilum and investigate the potential effects of underlying host factors and steroid administration on the pathogenesis of infection.

Molecular Detection and Typing of Dengue Viruses from Archived Tissues of Fatal Cases by RT-PCR and Sequencing: Diagnostic and Epidemiologic Implications

Diagnosis of dengue virus (DENV) infection in fatal cases is challenging because of the frequent unavailability of blood or fresh tissues. For formalin-fixed, paraffin-embedded (FFPE) tissues immunohistochemistry (IHC) can be used; however, it may not be as sensitive and serotyping is not possible. The application of reverse transcription-polymerase chain reaction (RT-PCR) for the detection of DENV in FFPE tissues has been very limited. We evaluated FFPE autopsy tissues of 122 patients with suspected DENV infection by flavivirus and DENV RT-PCR, sequencing, and DENV IHC. The DENV was detected in 61 (50%) cases by RT-PCR or IHC. The RT-PCR and sequencing detected DENV in 60 (49%) cases (DENV-1 in 16, DENV-2 in 27, DENV-3 in 8, and DENV-4 in 6 cases). No serotype could be identified in three cases. The IHC detected DENV antigens in 50 (40%) cases. The RT-PCR using FFPE tissue improves detection of DENV in fatal cases and provides sequence information useful for typing and epidemiologic studies.

Phylogeography of Rickettsia rickettsii Genotypes Associated with Fatal Rocky Mountain Spotted Fever

Rocky Mountain spotted fever (RMSF), a tick-borne zoonosis caused by Rickettsia rickettsii, is among the deadliest of all infectious diseases. To identify the distribution of various genotypes of R. rickettsii associated with fatal RMSF, we applied molecular typing methods to samples of DNA extracted from formalin-fixed, paraffin-embedded tissue specimens obtained at autopsy from 103 case-patients from seven countries who died of RMSF. Complete sequences of one or more intergenic regions were amplified from tissues of 30 (29%) case-patients and revealed a distribution of genotypes consisting of four distinct clades, including the Hlp clade, regarded previously as a non-pathogenic strain of R. rickettsii. Distinct phylogeographic patterns were identified when composite case-patient and reference strain data were mapped to the state and country of origin. The phylogeography of R. rickettsii is likely determined by ecological and environmental factors that exist independently of the distribution of a particular tick vector.

A pilot study of host genetic variants associated with influenza-associated deaths among children and young adults.

Low-producing MBL2 genotypes may have increased risk for MRSA co-infection.

Diagnosis of Influenza from Respiratory Autopsy Tissues Detection of Virus by Real-Time Reverse Transcription-PCR in 222 Cases

The recent influenza pandemic, caused by a novel H1N1 influenza A virus, as well as the seasonal influenza outbreaks caused by varieties of influenza A and B viruses, are responsible for hundreds of thousands of deaths worldwide. Few studies have evaluated the utility of real-time reverse transcription-PCR to detect influenza virus RNA from formalin-fixed, paraffin-embedded tissues obtained at autopsy. In this work, respiratory autopsy tissues from 442 suspect influenza cases were tested by real-time reverse transcription-PCR for seasonal influenza A and B and 2009 pandemic influenza A (H1N1) viruses and the results were compared to those obtained by immunohistochemistry. In total, 222 cases were positive by real-time reverse transcription-PCR, and of 218 real-time, reverse transcription-PCR-positive cases also tested by immunohistochemistry, only 107 were positive. Although formalin-fixed, paraffin-embedded tissues can be used for diagnosis, frozen tissues offer the best chance to make a postmortem diagnosis of influenza because these tissues possess nucleic acids that are less degraded and, as a consequence, provide longer sequence information than that obtained from fixed tissues. We also determined that testing of all available respiratory tissues is critical for optimal detection of influenza virus in postmortem tissues.

Determinants for Autopsy after Unexplained Deaths Possibly Resulting from Infectious Causes, United States

Autopsy findings, clinical history, and diagnostic tools can aid surveillance and investigation of infectious diseases.

Estimation of Undiagnosed Naegleria fowleri Primary Amebic Meningoencephalitis, United States1

Primary amebic meningoencephalitis is an acute, rare, typically fatal disease. We used epidemiologic risk factors and multiple cause-of-death mortality data to estimate the number of deaths that fit the typical pattern for primary amebic meningoencephalitis; we estimated an annual average of 16 deaths (8 male, 8 female) in the United States.