Virginia M. Anderson

H5N1 infection of the respiratory tract and beyond: a molecular pathology study

Summary Background Human infection with avian influenza H5N1 is an emerging infectious disease characterised by respiratory symptoms and a high fatality rate. Previous studies have shown that the human infection with avian influenza H5N1 could also target organs apart from the lungs. Methods We studied post-mortem tissues of two adults (one man and one pregnant woman) infected with H5N1 influenza virus, and a fetus carried by the woman. In-situ hybridisation (with sense and antisense probes to haemagglutinin and nucleoprotein) and immunohistochemistry (with monoclonal antibodies to haemagglutinin and nucleoprotein) were done on selected tissues. Reverse-transcriptase (RT) PCR, real-time RT-PCR, strand-specific RT-PCR, and nucleic acid sequence-based amplification (NASBA) detection assays were also undertaken to detect viral RNA in organ tissue samples. Findings We detected viral genomic sequences and antigens in type II epithelial cells of the lungs, ciliated and non-ciliated epithelial cells of the trachea, T cells of the lymph node, neurons of the brain, and Hofbauer cells and cytotrophoblasts of the placenta. Viral genomic sequences (but no viral antigens) were detected in the intestinal mucosa. In the fetus, we found viral sequences and antigens in the lungs, circulating mononuclear cells, and macrophages of the liver. The presence of viral sequences in the organs and the fetus was also confirmed by RT-PCR, strand-specific RT-PCR, real-time RT-PCR, and NASBA. Interpretation In addition to the lungs, H5N1 influenza virus infects the trachea and disseminates to other organs including the brain. The virus could also be transmitted from mother to fetus across the placenta.

Multiple organ infection and the pathogenesis of SARS

After >8,000 infections and >700 deaths worldwide, the pathogenesis of the new infectious disease, severe acute respiratory syndrome (SARS), remains poorly understood. We investigated 18 autopsies of patients who had suspected SARS; 8 cases were confirmed as SARS. We evaluated white blood cells from 22 confirmed SARS patients at various stages of the disease. T lymphocyte counts in 65 confirmed and 35 misdiagnosed SARS cases also were analyzed retrospectively. SARS viral particles and genomic sequence were detected in a large number of circulating lymphocytes, monocytes, and lymphoid tissues, as well as in the epithelial cells of the respiratory tract, the mucosa of the intestine, the epithelium of the renal distal tubules, the neurons of the brain, and macrophages in different organs. SARS virus seemed to be capable of infecting multiple cell types in several organs; immune cells and pulmonary epithelium were identified as the main sites of injury. A comprehensive theory of pathogenesis is proposed for SARS with immune and lung damage as key features.

Localization and distribution of vasoactive neuropeptides in the human placenta

Neuropeptides play an important role in the regional regulation of blood flow and hormone secretion. Few studies report the presence of peptides in the human placenta. Our experiment evaluates neuropeptides in the human placenta using immunocytochemical techniques. Representative tissue sections from full-term placentae were fixed immediately after delivery and processed into paraffin sections or frozen. They were treated with multiple immunofluorescence, streptavidin-biotin-peroxidase complex and immunogold-silver staining techniques in combination with well-established monoclonal and polyclonal antibodies, using appropriate absorption controls to ensure the validity of the staining. Vasoactive intestinal polypeptide (VIP), calcitonin gene-related peptide (CGRP), neuropeptide tyrosine (NPY), galanin, somatostatin, met-enkephaline, helodermin and substance P-like immunoreactivities were demonstrated within decidual cells. Endothelin-1 was found in both trophoblasts and endothelial cells. Peptide immunoreactivities in the human placenta especially at the decidual interface between mother and fetus supports a role for the diffuse neuroendocrine system (DNES) in the regulation of placental blood flow critical for fetal growth and development.

Tonsillar crypt epithelium is an important extra-central nervous system site for viral replication in EV71 encephalomyelitis.

Enterovirus 71 (EV71; family Picornaviridae, species human Enterovirus A) usually causes hand, foot, and mouth disease, which may rarely be complicated by fatal encephalomyelitis. We investigated extra-central nervous system (extra-CNS) tissues capable of supporting EV71 infection and replication, and have correlated tissue infection with expression of putative viral entry receptors, scavenger receptor B2 (SCARB2), and P-selectin glycoprotein ligand-1 (PSGL-1). Formalin-fixed, paraffin-embedded CNS and extra-CNS tissues from seven autopsy cases were examined by IHC and in situ hybridization to evaluate viral antigens and RNA. Viral receptors were identified with IHC. In all seven cases, the CNS showed stereotypical distribution of inflammation and neuronal localization of viral antigens and RNA, confirming the clinical diagnosis of EV71 encephalomyelitis. In six cases in which tonsillar tissues were available, viral antigens and/or RNA were localized to squamous epithelium lining the tonsillar crypts. Tissues from the gastrointestinal tract, pancreas, mesenteric nodes, spleen, and skin were all negative for viral antigens/RNA. Our novel findings strongly suggest that tonsillar crypt squamous epithelium supports active viral replication and represents an important source of viral shedding that facilitates person-to-person transmission by both the fecal-oral or oral-oral routes. It may also be a portal for viral entry. A correlation between viral infection and SCARB2 expression appears to be more significant than for PSGL-1 expression.

The spleen as a target in severe acute respiratory syndrome

It has been proposed that immune injury is the central mechanism of pathogenesis of the infectious disease, severe acute respiratory syndrome (SARS). To gain a better understanding of immune injury in the spleen, we investigated the number and distribution of various immune cell types in the spleens of SARS patients. We performed autopsies on six confirmed SARS cases, with six normal subjects as controls; spleen samples from these autopsies were examined with hematoxylin and eosin (H&E) sections, in situ hybridization for SARS virus genomic sequences, and immunohistochemistry with seven monoclonal antibodies to five cell types. The number and distribution of these cells were measured and analyzed using an image analysis system. SARS genomic sequences were detected in all SARS spleens. The SARS spleens all had severe damage to the white pulp and showed an alteration of the normal distribution of various cell types. Immunocytes in the red pulp were decreased by 68.0–90.7% except for CD68 macrophages and human leukocyte antigen (HLA)‐DR positive antigen‐presenting cells (APC), which were decreased to a lesser degree. On average, CD68 macrophages were increased in size by 2.21‐fold. We hypothesize that the collapse of the splenic immune system plays a key role in the clinical outcome of these patients.—Zhan, J., Deng, R., Tang, J., Zhang, B., Tang, Y., Wang, J. K., Li, F., Anderson, V. M., McNutt, M. A., Gu, J. The spleen as a target in severe acute respiratory syndrome (SARS). FASEB J. 20, 2321–2328 (2006)

The HIV-exposed placenta morphologic observations and interpretation

Summary Pediatric AIDS was first recognized in 1983 by Oleske and Rubinstein. HIV transmission from an infected mother to her infant is responsible for nearly 90% of all cases. HIV may cross the placental barrier or be transmitted intrapartum in a manner similar to hepatitis B infection. The placenta, bathed in the blood of HIV infected mothers during gestation, provides a protective barrier against HIV for some fetuses. Histopathologic study of HIV exposed placentae seems to suggest the presence of protective mechanisms against intrauterine HIV transmission. The purpose of this preliminary report is to study the histopathology of HIV exposed placentae and to postulate placental defense mechanisms against HIV infection. The best test of placental function is the growth and development of the fetus. Women with HIV have small for gestational age term infants with large HIV exposed placentae. This may indicate insufficient placental function throughout pregnancy

A genetic male infant with female phenotype in compatomelic syndrome: A possible relationship to exposure to oral contraceptives during pregnancy

Camptomelic syndrome is a severe malformation disorder affecting infant cartilage and bone formation. This syndrome is also characterized by sex reversal in a significant proportion of phenotypic females. In this case report, the authors describe a typical case of camptomelic syndrome in a black infant who had been exposed in utero to an oral contraceptive (OC). The infant was born after a full-term pregnancy. The mother had taken an OC containing 0.5-1.0 mg norethindrone and 0.035 mg ethinyl estradiol. Exposure had occurred for 6 months after conception. Parents were healthy and unrelated. The infant exhibited significant bone malformation in her legs, arms, feet, spine, and rib cage. Chromosome analysis yielded a normal 46,XY G-banded karyotype. The infant died at the age of 3 years, 6 months. Autopsy findings evidenced a female reproductive system. Microscopic examination of ovarian tissues revealed only immature sex cords; no oocytes were found. The authors briefly comment on camptomelic syndrome cases previously reported and implications of X-Y chromosome-gene effects associated with this syndrome. This may be the second reported case involving exposure to OCs early in pregnancy and sex reversal.