Biomedical subjects

Wun-Ju Shieh

Publications and source records attributed to Wun-Ju Shieh.

At least 19 recordsLinked to original sources

PubMed2006

Role of host cytokine responses in the pathogenesis of avian H5N1 influenza viruses in mice.

Highly pathogenic avian H5N1 influenza viruses are now widespread in poultry in Asia and have recently spread to some African and European countries. Interspecies transmission of these viruses to humans poses a major threat to public health. To better understand the basis of pathogenesis of H5N1 viruses, we have investigated the role of proinflammatory cytokines in transgenic mice deficient in interleukin-6 (IL-6), macrophage inflammatory protein 1 alpha (MIP-1alpha), IL-1 receptor (IL-1R), or tumor necrosis factor receptor 1 (TNFR1) by the use of two avian influenza A viruses isolated from humans, A/Hong Kong/483/97 (HK/483) and A/Hong Kong/486/97 (HK/486), which exhibit high and low lethality in mice, respectively. The course of disease and the extent of virus replication and spread in IL-6- and MIP-1alpha-deficient mice were not different from those observed in wild-type mice during acute infection with 1,000 50% mouse infective doses of either H5N1 virus. However, with HK/486 virus, IL-1R-deficient mice exhibited heightened morbidity and mortality due to infection, whereas no such differences were observed with the more virulent HK/483 virus. Furthermore, TNFR1-deficient mice exhibited significantly reduced morbidity following challenge with either H5N1 virus but no difference in viral replication and spread or ultimate disease outcome compared with wild-type mice. These results suggest that TNF-alpha may contribute to morbidity during H5N1 influenza virus infection, while IL-1 may be important for effective virus clearance in nonlethal H5N1 disease.

Animals↗

PubMed2006

Detection of West Nile virus in formalin-fixed, paraffin-embedded human tissues by RT-PCR: a useful adjunct to conventional tissue-based diagnostic methods.

BACKGROUND: West Nile virus (WNV), a member of genus Flavivirus, causes febrile illness, encephalitis, meningitis, myelitis, and occasional deaths in humans. Although several reverse transcription-polymerase chain reaction (RT-PCR) assays have been developed for detection of WNV in serum, cerebrospinal fluid, and fresh tissues, the usefulness of WNV RT-PCR assays for RNA extracted from formalin-fixed human tissues has not previously been demonstrated. OBJECTIVE: The objective of this study was to evaluate the application of a RT-PCR technique for the detection of WNV in routinely processed, formalin-fixed, paraffin-embedded (FFPE) human tissues, and to compare it with conventional serology and immunohistochemistry (IHC). STUDY DESIGN: We performed two WNV-specific nested RT-PCR assays targeting the viral capsid, premembrane, and envelope genes in FFPE central nervous system tissue samples from 27 patients with fatal WNV encephalitis, as confirmed by serology or IHC, and compared the results. The presence of WNV in RT-PCR-positive samples was confirmed by amplicon sequencing. RESULTS: Twenty (74%) patients were WNV RT-PCR positive while 24 (89%) were seropositive. WNV IHC staining of neurons and neuronal processes was positive in fourteen (52%) patients. The concordance between IHC and serology was 41% (11/27) and between RT-PCR and serology was 63% (17/27). All 11 seropositive/IHC-positive patients and 6 (46%) of 13 seropositive/IHC-negative patients were RT-PCR positive while all 3 seronegatives were positive by both IHC and RT-PCR. CONCLUSIONS: In this study, RT-PCR was significantly more sensitive than IHC in detecting WNV infections and provided specific sequence information about the infecting virus. RT-PCR on FFPE tissues may be a particularly useful diagnostic tool in patients who die relatively soon after disease onset and for whom serology may be negative. Combined use of serology, IHC, and RT-PCR would be expected to have the best overall sensitivity and improve detection of fatal WNV infection.

Adult↗

PubMed2006

A severe acute respiratory syndrome coronavirus that lacks the E gene is attenuated in vitro and in vivo.

A deletion mutant of severe acute respiratory syndrome coronavirus (SARS-CoV) has been engineered by deleting the structural E gene in an infectious cDNA clone that was constructed as a bacterial artificial chromosome (BAC). The recombinant virus lacking the E gene (rSARS-CoV-DeltaE) was rescued in Vero E6 cells. The recovered deletion mutant grew in Vero E6, Huh-7, and CaCo-2 cells to titers 20-, 200-, and 200-fold lower than the recombinant wild-type virus, respectively, indicating that although the E protein has an effect on growth, it is not essential for virus replication. No differences in virion stability under a wide range of pH and temperature were detected between the deletion mutant and recombinant wild-type viruses. Although both viruses showed the same morphology by electron microscopy, the process of morphogenesis seemed to be less efficient with the defective virus than with the recombinant wild-type one. The rSARS-CoV-DeltaE virus replicated to titers 100- to 1,000-fold lower than the recombinant wild-type virus in the upper and lower respiratory tract of hamsters, and the lower viral load was accompanied by less inflammation in the lungs of hamsters infected with rSARS-CoV-DeltaE virus than with the recombinant wild-type virus. Therefore, the SARS-CoV that lacks the E gene is attenuated in hamsters, might be a safer research tool, and may be a good candidate for the development of a live attenuated SARS-CoV vaccine.

Animals↗

PubMed2006

Histopathologic and immunohistochemical features of fatal influenza virus infection in children during the 2003-2004 season.

BACKGROUND: The Centers for Disease Control and Prevention enhanced national surveillance for influenza-associated deaths among children because of early reports of pediatric deaths during the 2003-2004 influenza season. METHODS: We studied lung and upper airway specimens from 47 case patients who died who had at least 1 positive result for influenza virus tests using hematoxylin and eosin, special stains for bacteria and fungi, and immunohistochemical (IHC) assays for influenza A and B viruses and other potential viral and bacterial respiratory pathogens. RESULTS: Nineteen (40%) of the 47 patients were <or=2 years old, and 26 (55%) were female. Influenza IHC testing identified type A antigens in 26 patients and type B antigens in 1 patient. Influenza antigens were observed focally in bronchoepithelial cells and mucous glands of trachea, bronchi, and larger bronchioli, showing submucosal mononuclear inflammatory infiltrates. IHC assays were the only confirmatory diagnostic test for 5 patients (11%). Significant life-threatening pathological conditions that could be considered the cause of death were present in 36 patients (77%) and included diffuse alveolar damage (12 cases), extensive secondary pneumonia (11 cases), extensive intraalveolar hemorrhage (10 cases), viral pneumonitis (10 cases), myocarditis (6 cases), and meningoencephalitis (1 case). For 9 patients with bronchopneumonia, a bacterial or fungal etiology was determined with IHC assay (3 Staphylococcus infections, 3 group A streptococci infections, 1 Streptococcus pneumoniae infection, 1 Bordetella pertussis infection, and 1 Aspergillus infection). CONCLUSIONS: IHC assays are useful for the diagnosis of influenza and bacterial pneumonia. This study underscores the importance of performing autopsies to identify the causes of death in patients with influenza infection.

Adolescent↗

PubMed2006

A large outbreak of Brainerd diarrhea associated with a restaurant in the Red River Valley, Texas.

BACKGROUND: In June 1996, an outbreak of chronic diarrhea was reported to the Texas Department of Health (Austin). METHODS: We initiated active case finding, performed 2 case-control studies, and conducted an extensive laboratory and environmental investigation. RESULTS: We identified 114 persons with diarrhea that lasted > or = 4 weeks. Symptoms among 102 patients who were studied included urgency (87%), fatigue (86%), fecal incontinence (74%), and weight loss (73%); the median maximum 24-h stool frequency was 15 stools. Diarrhea persisted for > 6 months in 87% and for > 1 year in 70% of patients who were observed. Fifty-one (89%) of 57 ill persons had eaten at a particular restaurant within 4 weeks before onset, compared with 8 (14%) of 59 matched control subjects (matched odds ratio [OR], undefined; 95% confidence interval [CI], 11.2-infinity). At the restaurant, patients were more likely than their unaffected dining companions to have drunk tap water (OR, 2.8; 95% CI, 1.0-9.9) and to have eaten several specific food items, and they were less likely to have drunk iced tea made from boiled water and store-bought ice (OR, 0.3; 95% CI, 0.05-1.0). A multivariable model that included consumption of tap water and salad bar tomatoes best fit the data. The restaurant had multiple sanitary and plumbing deficiencies. Extensive laboratory and environmental testing for bacterial, parasitic, mycotic, and viral agents did not identify an etiologic agent. CONCLUSIONS: The clinical, laboratory, and epidemiologic findings are consistent with those of previous outbreaks of Brainerd diarrhea. To our knowledge, this is the largest reported outbreak of Brainerd diarrhea associated with a restaurant.

Case-Control Studies↗

PubMed2006

Transmission of lymphocytic choriomeningitis virus by organ transplantation.

BACKGROUND: In December 2003 and April 2005, signs and symptoms suggestive of infection developed in two groups of recipients of solid-organ transplants. Each cluster was investigated because diagnostic evaluations were unrevealing, and in each a common donor was recognized. METHODS: We examined clinical specimens from the two donors and eight recipients, using viral culture, electron microscopy, serologic testing, molecular analysis, and histopathological examination with immunohistochemical staining to identify a cause. Epidemiologic investigations, including interviews, environmental assessments, and medical-record reviews, were performed to characterize clinical courses and to determine the cause of the illnesses. RESULTS: Laboratory testing revealed lymphocytic choriomeningitis virus (LCMV) in all the recipients, with a single, unique strain of LCMV identified in each cluster. In both investigations, LCMV could not be detected in the organ donor. In the 2005 cluster, the donor had had contact in her home with a pet hamster infected with an LCMV strain identical to that detected in the organ recipients; no source of LCMV infection was found in the 2003 cluster. The transplant recipients had abdominal pain, altered mental status, thrombocytopenia, elevated aminotransferase levels, coagulopathy, graft dysfunction, and either fever or leukocytosis within three weeks after transplantation. Diarrhea, peri-incisional rash, renal failure, and seizures were variably present. Seven of the eight recipients died, 9 to 76 days after transplantation. One recipient, who received ribavirin and reduced levels of immunosuppressive therapy, survived. CONCLUSIONS: We document two clusters of LCMV infection transmitted through organ transplantation.

Adult↗

PubMed2006

Molecular diagnosis of Nocardia farcinica from a cerebral abscess.

Histopathology and special stains of a brain biopsy specimen from a 42-year-old man revealed numerous gram-positive bacilli arranged in branching filaments, suggesting Nocardia infection. Antibiotic therapy with trimethoprim-sulfamethoxazole markedly decreased the abscess size, and the patient improved. DNA was analyzed from formalin-fixed sections of the cerebral abscess by a 16S ribosomal DNA polymerase chain reaction assay demonstrating the presence of either Nocardia farcinica or N otitidiscaviarum. A species-specific polymerase chain reaction assay confirmed N farcinica as the etiologic agent.

Adult↗

PubMed2006

Morphologic, immunologic, and molecular methods to detect bacillus anthracis in formalin-fixed tissues.

Due to the importance of Bacillus anthracis as a cause of naturally occurring infection among humans and as an agent of bioterrorism, there is a vital need for rapid and specific assays, including immunohistochemistry (IHC) and polymerase chain reaction (PCR) assays, to detect the bacterium in formalin-fixed tissues. Colorimetric IHC assays were developed using a multistep indirect immunoalkaline phosphatase method with anti-B. anthracis cell wall (EAII-6G6-2-3) and anti-B. anthracis capsule (FDF-1B9) mAbs to detect B. anthracis antigens in formalin-fixed, paraffin-embedded bacterial cultures and tissues. B. anthracis antigens were localized, using both antibodies, in samples from B. anthracis-infected animals and humans. The colorimetric IHC assay with both antibodies was expedient in diagnosing the presence of B. anthracis in formalin-fixed, paraffin-embedded tissue from bioterrorism-associated cases of inhalational and cutaneous anthrax and from a case of naturally occurring cutaneous anthrax. Using the same antibodies, confocal microscopy demonstrated the structure of replicating B. anthracis in tissues. B. anthracis-specific primers were successfully used with PCR to amplify and detect B. anthracis sequences derived from formalin-fixed tissues of anthrax cases. In this study, morphologic, immunologic, and molecular assays were used to study and diagnose 22 veterinary and human anthrax cases.

Animals↗

PubMed2006

A single intranasal inoculation with a paramyxovirus-vectored vaccine protects guinea pigs against a lethal-dose Ebola virus challenge.

To determine whether intranasal inoculation with a paramyxovirus-vectored vaccine can induce protective immunity against Ebola virus (EV), recombinant human parainfluenza virus type 3 (HPIV3) was modified to express either the EV structural glycoprotein (GP) by itself (HPIV3/EboGP) or together with the EV nucleoprotein (NP) (HPIV3/EboGP-NP). Expression of EV GP by these recombinant viruses resulted in its efficient incorporation into virus particles and increased cytopathic effect in Vero cells. HPIV3/EboGP was 100-fold more efficiently neutralized by antibodies to EV than by antibodies to HPIV3. Guinea pigs infected with a single intranasal inoculation of 10(5.3) PFU of HPIV3/EboGP or HPIV3/EboGP-NP showed no apparent signs of disease yet developed a strong humoral response specific to the EV proteins. When these animals were challenged with an intraperitoneal injection of 10(3) PFU of EV, there were no outward signs of disease, no viremia or detectable EV antigen in the blood, and no evidence of infection in the spleen, liver, and lungs. In contrast, all of the control animals died or developed severe EV disease following challenge. The highly effective immunity achieved with a single vaccine dose suggests that intranasal immunization with live vectored vaccines based on recombinant respiratory viruses may be an advantageous approach to inducing protective responses against severe systemic infections, such as those caused by hemorrhagic fever agents.

Administration, Intranasal↗

PubMed2006

Diagnosis of invasive group a streptococcal infections by using immunohistochemical and molecular assays.

Invasive group A streptococcus (GAS) infections cause 1,100 to 1,300 deaths annually in the United States. Diagnosis is made when Streptococcus pyogenes is isolated from pus or body fluids; however, cultures are not always obtained, and antibiotic treatment can preclude bacterial growth. An immunohistochemical assay for GAS was applied to formalin-fixed tissue samples from 122 patients with suspect GAS infection. Immunohistochemical staining of well-defined cocci and small, granular antigen fragments was observed in 27 cases. S pyogenes was isolated in 18 cases, whereas in 8 cases, immunohistochemical staining was confirmed by amplification of the sepB gene of S pyogenes from paraffin-embedded samples in a heminested polymerase chain reaction (PCR) assay. A primary focus of infection (respiratory, mucocutaneous, or gynecologic) was present in 22 patients, whereas 5 had no identifiable primary focus of infection. Eighteen patients had systemic infection. Immunohistochemical analysis and PCR can be used for diagnosis of GAS infections in formalin-fixed, paraffin-embedded samples.

Adolescent↗

PubMed2006

Intussusception after administration of the rhesus tetravalent rotavirus vaccine (Rotashield): the search for a pathogenic mechanism.

OBJECTIVES: The rhesus tetravalent rotavirus vaccine (RRV) was withdrawn from the routine program for childhood immunization in the United States because of the rare and unexpected occurrence of intussusception in the 2-week period after administration of the first dose. METHODS: To search for the pathogenesis of this association, we compared the pathology of surgical specimens from infants who had surgical reduction of their intussusceptions within 2 weeks of receiving the vaccine (case patients; n = 8) with the pathology of specimens from children who had surgery > 2 weeks after immunization (n = 6) or who had never been immunized (n = 26). Tissue was examined for evidence of the vaccine strain of rotavirus by reverse transcriptase-polymerase chain reaction (RT-PCR), in situ hybridization, and immunohistochemical staining. RESULTS: RRV was identified by RT-PCR in tissue samples from 7 of the 8 case patients and in 2 of the 6 children who received the vaccine at a more distant time (29 and 58 days before surgery), but it was not identified in samples from any of the nonvaccinated children. No evidence of rotavirus tissue involvement was detected in any of the children by in situ hybridization or immunohistochemical staining. Pathologic evidence (for example, inclusion bodies, smudge cells) of adenovirus infection was present in 35% of the 37 specimens examined by routine staining and immunohistochemistry. CONCLUSIONS: The fact that RRV was detected by RT-PCR but not by either of the other assays could be explained by RRV being present in the lumen of the gut but not in the tissues of appendix, ileum, or Peyer's patches. The Peyer's patches were not hyperplastic, and we could not establish the pathogenic mechanism for this association.

Adenoviridae↗

PubMed2005

Influenza-associated deaths among children in the United States, 2003-2004.

BACKGROUND: Although influenza is common among children, pediatric mortality related to laboratory-confirmed influenza has not been assessed nationally. METHODS: During the 2003-2004 influenza season, we requested that state health departments report any death associated with laboratory-confirmed influenza in a U.S. resident younger than 18 years of age. Case reports, medical records, and autopsy reports were reviewed, and available influenza-virus isolates were analyzed at the Centers for Disease Control and Prevention. RESULTS: One hundred fifty-three influenza-associated deaths among children were reported by 40 state health departments. The median age of the children was three years, and 96 of them (63 percent) were younger than five years old. Forty-seven of the children (31 percent) died outside a hospital setting, and 45 (29 percent) died within three days after the onset of illness. Bacterial coinfections were identified in 24 of the 102 children tested (24 percent). Thirty-three percent of the children had an underlying condition recognized to increase the risk of influenza-related complications, and 20 percent had other chronic conditions; 47 percent had previously been healthy. Chronic neurologic or neuromuscular conditions were present in one third. The mortality rate was highest among children younger than six months of age (0.88 per 100,000 children; 95 percent confidence interval, 0.52 to 1.39 per 100,000). CONCLUSIONS: A substantial number of influenza-associated deaths occurred among U.S. children during the 2003-2004 influenza season. High priority should be given to improvements in influenza-vaccine coverage and improvements in the diagnosis and treatment of influenza to reduce childhood mortality from influenza.

Adolescent↗

PubMed2005

Fatal toxic shock syndrome associated with Clostridium sordellii after medical abortion.

Endometritis and toxic shock syndrome associated with Clostridium sordellii have previously been reported after childbirth and, in one case, after medical abortion. We describe four deaths due to endometritis and toxic shock syndrome associated with C. sordellii that occurred within one week after medically induced abortions. Clinical findings included tachycardia, hypotension, edema, hemoconcentration, profound leukocytosis, and absence of fever. These cases indicate the need for physician awareness of this syndrome and for further study of its association with medical abortion.

Abdominal Pain↗

PubMed2005

Subacute sclerosing panencephalitis: more cases of this fatal disease are prevented by measles immunization than was previously recognized.

BACKGROUND: The most severe sequela of measles virus infection is subacute sclerosing panencephalitis (SSPE), a fatal disease of the central nervous system that generally develops 7-10 years after infection. From 1989 through 1991, a resurgence of measles occurred in the United States, with 55,622 cases of measles reported. The purpose of the present study was to identify cases of SSPE that were associated with the resurgence of measles and to calculate the risk of developing SSPE. METHODS: Brain tissue samples obtained from 11 patients with a presumptive diagnosis of SSPE were tested for the presence of measles virus RNA. Measles virus genotypes were determined by reverse-transcription polymerase chain reaction (RT-PCR) and by analysis of the sequences of the PCR products. A search of the literature was conducted to identify reports of cases of SSPE in persons residing in the United States who had measles during 1989-1991. RESULTS: The measles virus sequences derived from brain tissue samples obtained from 11 patients with SSPE confirmed the diagnosis of SSPE. For 5 of the 11 patients with SSPE who had samples tested by RT-PCR and for 7 patients with SSPE who were identified in published case reports, it was determined that the development of SSPE was associated with the measles resurgence that occurred in the United States during 1989-1991. The estimated risk of developing SSPE was 10-fold higher than the previous estimate reported for the United States in 1982. CONCLUSIONS: Vaccination against measles prevents more cases of SSPE than was originally estimated.

Adolescent↗

PubMed2005

CD8-mediated protection against Ebola virus infection is perforin dependent.

CD8 T cells have been shown to play an important role in the clearance and protection against fatal Ebola virus infection. In this study, we examined the mechanisms by which CD8 T cells mediate this protection. Our data demonstrate that all normal mice infected s.c. with a mouse-adapted Ebola virus survived the infection, as did 100% of mice deficient in Fas and 90% of those deficient in IFN-gamma. In contrast, perforin-deficient mice uniformly died after s.c. challenge. Perforin-deficient mice failed to clear viral infection even though they developed normal levels of neutralizing anti-Ebola Abs and 5- to 10-fold higher levels of IFN-gamma than control mice. Using MHC class I tetramers, we have also shown that perforin-deficient mice have 2- to 4-fold higher numbers of Ebola-specific CD8s than control mice. These findings suggest that the clearance of Ebola virus is perforin-dependent and provide an additional example showing that this basic immunologic mechanism is not limited to the clearance of noncytopathic viruses.

Animals↗

PubMed2005

Transmission of rabies virus from an organ donor to four transplant recipients.

BACKGROUND: In 2004, four recipients of kidneys, a liver, and an arterial segment from a common organ donor died of encephalitis of an unknown cause. METHODS: We reviewed the medical records of the organ donor and the recipients. Blood, cerebrospinal fluid, and tissues from the recipients were tested with a variety of assays and pathological stains for numerous causes of encephalitis. Samples from the recipients were also inoculated into mice. RESULTS: The organ donor had been healthy before having a subarachnoid hemorrhage that led to his death. Encephalitis developed in all four recipients within 30 days after transplantation and was accompanied by rapid neurologic deterioration characterized by agitated delirium, seizures, respiratory failure, and coma. They died an average of 13 days after the onset of neurologic symptoms. Mice inoculated with samples from the affected patients became ill seven to eight days later, and electron microscopy of central nervous system (CNS) tissue demonstrated rhabdovirus particles. Rabies-specific immunohistochemical and direct fluorescence antibody staining demonstrated rabies virus in multiple tissues from all recipients. Cytoplasmic inclusions consistent with Negri bodies were seen in CNS tissue from all recipients. Antibodies against rabies virus were present in three of the four recipients and the donor. The donor had told others of being bitten by a bat. CONCLUSIONS: This report documenting the transmission of rabies virus from an organ donor to multiple recipients underscores the challenges of preventing and detecting transmission of unusual pathogens through transplantation.

Antibodies, Viral↗

PubMed2005

Immunohistochemical, in situ hybridization, and ultrastructural localization of SARS-associated coronavirus in lung of a fatal case of severe acute respiratory syndrome in Taiwan.

This article describes the pathological studies of fatal severe acute respiratory syndrome (SARS) in a 73-year-old man during an outbreak of SARS in Taiwan, 2003. Eight days before onset of symptoms, he visited a municipal hospital that was later identified as the epicenter of a large outbreak of SARS. On admission to National Taiwan University Hospital in Taipei, the patient experienced chest tightness, progressive dyspnea, and low-grade fever. His condition rapidly deteriorated with increasing respiratory difficulty, and he died 7 days after admission. The most prominent histopathologic finding was diffuse alveolar damage of the lung. Immunohistochemical and in situ hybridization assays demonstrated evidence of SARS-associated coronavirus (SARS-CoV) infection in various respiratory epithelial cells, predominantly type II pneumocytes, and in alveolar macrophages in the lung. Electron microscopic examination also revealed coronavirus particles in the pneumocytes, and their identity was confirmed as SARS-CoV by immunogold labeling electron microscopy. This report is the first to describe the cellular localization of SARS-CoV in human lung tissue by using a combination of immunohistochemistry, double-stain immunohistochemistry, in situ hybridization, electron microscopy, and immunogold labeling electron microscopy. These techniques represent valuable laboratory diagnostic modalities and provide insights into the pathogenesis of this emerging infection.

Aged↗

PubMed2005

Persistent Yersinia pestis antigens in ischemic tissues of a patient with septicemic plague.

In November 2002, a couple from New Mexico traveled to New York where both had fever and unilateral inguinal adenopathy. The husband was in septic shock when he sought medical care and was admitted to an intensive care unit, where he developed ischemic necrosis of his feet which later required bilateral amputation. Yersinia pestis was grown from his blood. Immunohistochemical assays using anti-Y pestis antibodies demonstrated multiple bacteria and granular antigens in and around vessels of the ischemic amputation tissues obtained 20 days after initiation of antibiotics; however, no evidence of Y pestis was present in viable tissues. Immunohistochemical evidence of Y pestis inside vessels of gangrenous feet in this patient underscores the importance of adequate excision of necrotic or partially necrotic tissues because antibiotics cannot be effectively delivered to necrotic and poorly perfused tissues.

Amputation, Surgical↗