Pathology of Viral Infections PDF

Module 04: Basic Pathologies 2 Infectious Diseases Pathology of Viral Infections Maria Kariza Tolentino Molina, MD, DPSP | 10/23...

Module 04: Basic Pathologies 2 Infectious Diseases Pathology of Viral Infections Maria Kariza Tolentino Molina, MD, DPSP | 10/23/24 | Asynchronous TABLE OF CONTENTS Immunity to some viruses wanes with time → May allow the same virus to infect the host repeatedly Learning Objectives 1 C. Cytomegalovirus 7 → For example: Respiratory Syncytial Virus (RSV) I. Introduction to Viral 1 (CMV) Infections IV. Transforming Viral 8 A. MEASLES (RUBEOLA VIRUS) II. Acute (Transient 1 Infections Acute viral infection Infections A. Epstein-Barr 8 Affects multiple organs A. Measles 1 Virus (EBV) Causes a wide range of diseases B. Mumps 2 Infections → Mild, self-limited, to severe systemic manifestations C. Poliovirus Infection 3 IV. Other Acute 11 Leading cause of vaccine preventable death & illness D. West Nile Virus 4 (Transient) Infections → More than 20 million people acquire measles each year E. Viral Hemorrhagic 4 A. Zika Virus 11 2010: 139,000 deaths globally Fever B. Dengue Virus 11 → Majority occurred in children in developing countries III. Latent (Herpesvirus) 5 C. SARS-COV-2 13 → Poor nutrition, lack of access to medical care Infections (COVID-19) Severe disease in patients with defective immunity A. Herpes Simplex 5 Summary & Key 13 → HIV Virus (HSV) Points → Hematologic malignancies (such as leukemia) B. Varicella-Zoster 6 Review Questions 14 Epidemics Virus (VZV) Rationale 14 → Unvaccinated individuals Appendix 16 [Nice!] Dr. Molina’s Clerkship in San Lazaro Hospital LEARNING OBJECTIVES Outbreak of severe forms of measles in pediatric patients 1. To discuss the characteristics, pathogenesis, diagnosis, and → Increased mortality clinical manifestations of acute (transient) infections such as → Babies developing pneumonia secondary to measles measles, mumps, poliovirus, West Nile virus, and viral Mostly unvaccinated hemorrhagic fever VIRUS CHARACTERISTICS 2. To discuss the characteristics, pathogenesis, diagnosis, and clinical manifestations of latent (Herpesvirus) infections, such Single-stranded RNA as Herpes Simplex Virus, Varicella Zoster Virus, and Family Paramyxoviridae, Genus Morbillivirus (T04.34, 2027) Cytomegalovirus Only one serotype 3. To discuss the characteristics, pathogenesis, diagnosis, and Transmitted by respiratory or aerosol droplets clinical manifestations of transforming viral infections, such as 3 Cell-Surface Receptors: all bind the hemagglutinin protein Epstein-Barr Virus → CD46: complement regulatory protein 4. To discuss the characteristics, pathogenesis, diagnosis, and → SLAM: signaling lymphocytic activation molecules clinical manifestations of Zika, Dengue, and COVID-19 viruses → Necin 4: adherens junction protein Can replicate in a variety of cell types Take Note! → Epithelial cells The lecture is based on Chapter 8 of Robbins & Cotran → Leukocytes Pathologic Basis of Disease, 10th Edition PATHOGENESIS HIV and Hepatitis viruses will not be discussed in detail, as this will be covered in different chapters. 1. Initially, multiplies in the respiratory tract 2. Spreads to local lymphoid tissue I. INTRODUCTION TO VIRAL INFECTIONS 3. Replication of the virus in the lymphoid tissue Viral infections are the cause of many clinically important acute 4. Enters the bloodstream: causing viremia and chronic infections, which may affect every organ system 5. Systemic dissemination of virus and blood → Conjunctiva, skin, respiratory tract, urinary tract, small blood Important! vessels, lymphatic system, and CNS Refer to Table A under Master Tables for the Selected Human CLINICAL MANIFESTATIONS Viruses and Viral Diseases lifted from Robbins. Measles Rash → It organizes the different viruses according to the organ system it infects and what disease it causes. Most children develop T-cell mediated immunity to measles → Doc mentions that it is very helpful and well-organized. → Helps control the viral infection → Produces the measles rash II. ACUTE (TRANSIENT) INFECTIONS Hence, the rash is less frequent in people with deficiencies Characteristics of viruses that cause transient infections: in cell-mediated immunity → Structurally heterogeneous Appearance: blotchy, reddish-brown rash on the face, trunk, → All elicit effective immune responses and proximal extremities That eliminate the pathogens Microscopically: dilated skin vessels, edema, mononuclear → Limiting the durations of infections perivascular infiltrate Viruses exhibit widely differing degrees of genetic diversity → Mononuclear cells can be seen around the blood vessels Important impact on the susceptibility of host to reinfection by Koplik Spots viruses of the same type A pathognomonic physical finding in patients with measles → Mumps Virus Appearance: ulcerated mucosal lesions in oral cavity near Has only one genetic subtype infects people only once opening of Stensen’s ducts (as pointed by arrows in Figure 1) → Influenza Virus Microscopically marked by: necrosis, neutrophilic exudate, and Can repeatedly infect the same individual since new neovascularization genetic variants arise periodically in nature YL6 04.39 TG13: Trans Police [Yang] | CG4: Trans Police [Beltran] | Version 2 1 of 16  → Measles inclusion body encephalitis Occurs in immunocompromised individuals DIAGNOSIS Mainly clinical Serologic tests may be performed Detection of viral antigen in nasal exudates or urinary sediments B. MUMPS Causes acute systemic infection → Can be mild, self-limited, or severe systemic infection Initially causes pain and swelling of salivary glands Figure 1. Koplik spots Mumps vaccine reduces incidences up to 99% Germinal Cell Hyperplasia VIRUS CHARACTERISTICS In lymphoid organs, measles is characterized by germinal cell Family Paramyxoviridae, Genus Paramyxovirus hyperplasia Transmitted via respiratory droplets → Large germinal centers compared to normal Two surface glycoproteins: → Recall: germinal center of a lymphoid tissue is the → Hemagglutinin and neuraminidase activity lighter-colored zone → Cell fusion and cytolytic activity Warthin-Finkeldey Cells PATHOGENESIS Pathognomonic finding of measles 1. Enters respiratory tract through inhalation of droplets Have eosinophilic nuclei and cytoplasmic inclusion bodies 2. Spreads to draining lymph nodes or lymphoid tissue where the Commonly seen in the lung tissue and sputum virus will replicate in lymphocytes 3. Spreads to blood causing viremia 4. Through the blood, it spreads to other organs such as salivary glands and other glands → Primarily infects salivary gland ductal epithelial cells Resulting in desquamation of involved cells, edema, and inflammation CLINICAL MANIFESTATIONS Classically presents as salivary gland pain and swelling Can spread to other sites and cause inflammation & swelling in: → CNS, testis, ovary, pancreas → Aseptic meningitis (15%): most common extra salivary Figure 2. Warthin-Finkeldey cells gland complication Other Clinical Manifestations Mumps Parotitis In malnourished children with poor medical care, measles virus Inflammation of the parotid gland secondary to the mumps virus may cause: Most commonly occurs bilaterally (70%) → Croup Appearance: → Pneumonia → Enlargement of the parotid gland with doughy consistency → Diarrhea → Gross cut section: moist, glistening, and reddish-brown → Protein-losing enteropathy Microscopically: In more severe cases, it may lead to: → Diffuse infiltration by macrophages, lymphocytes, and plasma → Keratitis (leading to scarring and blindness) cells → Encephalitis Infiltrates compress the acini and ducts of the gland → Hemorrhagic rashes (“black measles”) → Neutrophils and necrotic debris fill the lumen May cause focal damage to the lining epithelium Figure 3. Hemorrhagic rash (‘black measles”) Figure 4. Mumps parotitis Antibody-mediated immunity Mumps Orchitis → Protects against reinfection [!] Testicular swelling Can cause transient but profound immunosuppression Microscopically: → Secondary to bacterial and viral infections → Presence of edema, mononuclear infiltration, focal → Attributes to the measles-related morbidity and mortality hemorrhages Alterations in both innate and adaptive immune responses Parenchymal swelling can compromise blood supply because → Occur following measles infection the tunica albuginea is not expandable → Includes defects in dendritic cell and lymphocyte function → Recall: the testis is tightly contained within the tunica Rare complications include: albuginea → Subacute sclerosing panencephalitis → Can lead to areas of infarction YL6 04.39 Pathology of Viral Infections 2 of 16  May cause testicular damage resulting in: → Have nearly eradicated polio because poliovirus: ○ Scarring Infects only humans ○ Atrophy Has limited genetic variation ○ Sterility (severe cases) Maybe effectively neutralized by antibodies generated by immunization PATHOGENESIS 1. Virus binds to CD155 (epithelial adhesion molecule) 2. Virus is ingested and replicates in the mucosa of the pharynx and gut, including tonsils and Peyer’s patches of the ileum 3. Spreads to lymphatics and lymph nodes 4. Produces transient viremia and fever CLINICAL MANIFESTATIONS Mostly asymptomatic Very rare for poliovirus infection to occur after vaccination → The virus will only occur in patients with mutations of the attenuated viruses to wild-type forms CNS Complications Only 1/100 cases that occur invade the CNS → Spinal poliomyelitis Poliovirus replicated in motor neurons of the spinal cord Figure 5. Mumps orchitis → Bulbar poliomyelitis Mumps Pancreatitis Poliovirus replicated in the brainstem Infection and damage of acinar cells of the pancreas which Viral spread to the CNS may through: triggers the release of digestive enzymes → Blood (viremia) Microscopically: → Retrograde transport of the virus along axons of the motor → Parenchymal and fat necrosis neurons → Neutrophil rich inflammation Figure 6. Mumps pancreatitis Figure 7. Descending (motor) tracts in the spinal cord Mumps Encephalitis Inflammation of the brain secondary to mumps Perivenous demyelination Perivascular mononuclear cuffing DIAGNOSIS Mainly clinical → Usually enough to diagnose mumps Serology Viral culture PCR assays: definitive diagnosis C. POLIOVIRUS INFECTION Acute systemic disease Has a wide range of manifestation: → Mild and self-limited Figure 8. Axons of the motor neurons → Severe infection can cause paralysis of the limb muscle and respiratory muscles Antiviral antibodies control the disease in most cases → It is not known why these fail to contain the virus in some VIRUS CHARACTERISTICS individuals Spherical, unencapsulated (naked) RNA virus Family Picornaviridae, Genus Enterovirus Take Note! Transmitted via fecal-oral route Doc’s slides said “antiretroviral antibodies”, but Robbins & Vaccines: Salk formalin-fixed (killed) vaccine and Sabin oral, Cotran (2015) say “antiviral antibodies” Attenuated (live) vaccine Antiretrovirals are not used for poliovirus because it is not a → Contains 3 serotypes of poliovirus retrovirus YL6 04.39 Pathology of Viral Infections 3 of 16  DIAGNOSIS → Pancreatitis Main method: viral culture or PCR CNS Complications → Samples used: Throat secretions or stool CNS complications occur in 1/150 cases Alternative Method: Serology → 10% of those who experience CNS complications have the D. WEST NILE VIRUS following manifestations: Eg., Meningitis, encephalitis, and meningoencephalitis Acute systemic viral infection → Usually results in long-term cognitive and neurologic May cause: impairment → Mild, Self-limited infection Greater risk for severe form of West Nile Virus include: → Neuroinvasive disease → Immunosuppressed individuals Associated with long-term neurological sequelae → Older adults VIRUS CHARACTERISTICS DIAGNOSIS Family Flaviviridae, Genus Flavivirus Main method of diagnosis is serology Has a broad geographic distribution: Alternative methods include: → Old World → Viral culture → Africa → PCR → Middle East → Europe E. VIRAL HEMORRHAGIC FEVER (VHF) → Southeast Asia Severe, life-threatening multisystem syndrome → Australia Causes vascular dysregulation and damage Arthropod-borne virus (arbovirus) Eventually leads to shock → Transmitted by mosquitoes to birds and mammals VIRUS CHARACTERISTICS Major reservoir hosts: birds ○ Develop prolonged viremia 4 different genera of Enveloped RNA virus cause VHF: Incidental hosts: humans → Arenaviridae ○ Most infected patients acquire the infection from a → Filoviridae mosquito bite → Bunyaviridae ○ Human-to-human transmission may occur (less → Flaviviridae common) via blood transfusion, organ transplantation, PATHOGENESIS breastfeeding, or transplacental spread All these viruses pass through insects or animals in the life cycle Humans: incidental hosts → Via contact with infected hosts (rodents) or insect vectors (mosquitoes and ticks) Some viruses occur secondary to person-to-person spread: → Ebola Figure 9. Transmission of west nile virus from mosquito to birds and humans → Marburg → Lassa viruses PATHOGENESIS Pathogenesis varies among the viruses 1. Individual is bitten by a mosquito Common Features in Pathogenesis 2. Virus replicates in skin dendritic cells 3. From the dendritic cells, it migrates to the lymph nodes for viral Prominent damage to blood vessels caused by: replication → Direct damage to endothelial cells 4. After replication in the lymphatic tissue, the virus migrates to the → Infection of macrophages and dendritic cells leading to blood → viremia production of inflammatory cytokines 5. From the blood, the virus will be disseminated to the different CLINICAL MANIFESTATIONS organs (e.g., brain) Viral infection may cause a spectrum of illnesses, ranging from → West Nile virus can cross the blood brain barrier mild acute to life-threatening 6. In the CNS, the neurons get infected Mild acute disease → Chemokines are activated → E.g., Fever, headache, myalgia, rash, neutropenia, Critical in recruiting leukocytes for viral clearance thrombocytopenia → CCR5 Life-threatening disease Chemokine receptor → E.g., Sudden hemodynamic deterioration and shock Contributes to resistance to neuroinvasive infection Mutations in both copies of CCR5 gene that lead to loss of Bleeding Manifestations function are associated with increased rate of symptomatic Bleeding, including petechiae, occurs secondary to: infection → Thrombocytopenia/platelet dysfunction → Endothelial injury CLINICAL MANIFESTATIONS → Cytokine-induced disseminated intravascular coagulation Usually asymptomatic (DIC) Around 20% of those affected by the virus may present with: → Deficiency of clotting factors due to hepatic injury → Fever Hemorrhages are prominent in some types of VHF → Headache → Congo-Crimean fever → Myalgia Rarely life-threatening → Fatigue → Necrosis of tissues → Anorexia May also be appreciated in this group of viral infections → Nausea Secondary to vascular lesions → Around half of symptomatic cases may develop Hemorrhages vary from mild, focal to massive maculopapular rash Rare complications include: → Hepatitis → Myocarditis YL6 04.39 Pathology of Viral Infections 4 of 16  PATHOGENESIS ACTIVE RECALL In immunocompetent hosts: 1. T/F: Hepatitis B, D, & C cause acute viral hepatitis. → Primary HSV infection resolves in a few weeks and the virus 2. Identification: Name a genera that can cause Viral remains latent in nerve cells Hemorrhagic Fever. → During latency: 3. Identification: Germinal cell hyperplasia is present Viral DNA remains within the nucleus of the neuron particularly in which organ/s of a patient with measles Only latency-associated viral RNA transcripts (LATs) are synthesized infection? No viral proteins appear to be produced during latency Answer Key: 1F, 2 Arenaviridae, Filoviridae, Bunyaviridae, or Flaviviridae Reactivation of HSV-1 and HSV-2 3 Lymphoid organs → May occur repeatedly with or without symptoms III. LATENT (HERPESVIRUS) INFECTIONS Results in the spread of virus from neurons to the skin or Latency mucous membranes → Persistence of viral genomes in cells that do not produce Can occur in the presence of host immunity infectious virus/disease HSVs have developed ways to avoid immune recognition Latent infection Events in Pathogenesis → Characteristics: Viruses persist in non-infectious form With periodic reactivation Eventual shedding of infectious virus → Dissemination of the infection and tissue injury stem from and may only occur upon reactivation of the latent virus Herpesvirus → Large, encapsulated viruses → Double-stranded DNA genomes → Cause acute infection followed by latent infection → 8 human types Figure 10. Pathogenesis of HSV → 3 subgroups Defined by: 1. Transmission occurs at primary sites of replication ○ Cell type most frequently infected → HSV-1: through the mouth ○ Site of latency → HSV-2: through the genital area via sexual intercourse Table 1. Herpesvirus subgroups 2. Virus spreads to the sensory neurons that innervate the primary Subgroup Description sites of replication 3. Viral nucleocapsids are transported along axons to the neuronal Infect epithelial cells Latent infection in neurons cell bodies where the viruses establish latent infection Viruses: CLINICAL MANIFESTATIONS α-group ○ Herpes simplex virus-1 (HSV-1) ○ Herpes simplex virus-2 (HSV-2) Corneal Blindness ○ Varicella zoster virus (VZV) HSV-1: major infectious cause of corneal blindness Infect and produce latent infection in a variety of → Direct viral damage affecting the corneal epithelium cell types → Immune-mediated damage leading to corneal stromal Viruses: β-group disease ○ Cytomegalovirus (CMV) ○ Human herpesvirus 6 (HHV-6) Fatal Sporadic Encephalitis ○ Human herpesvirus 7 (HHV-7) HSV-1: major cause of fatal sporadic encephalitis Cause Kaposi sarcoma Occurs in the temporal lobe or orbital gyri of frontal lobes Latent infection in lymphoid cells Mutations in Toll-Like receptor 3 (TLR3) or components of the Viruses: γ-group ○ Epstein-Barr virus (EBV) signaling pathway increase the risk of HSV encephalitis ○ Kaposi sarcoma’s herpesvirus (KSHV)/ Patients with compromised cellular immunity (i.e., undergoing HHV-8 chemotherapy or diagnosed with HIV) → At risk for disseminated herpesvirus infection Herpesvirus simiae (Monkey B virus) → Old world monkey virus that resembles HSV-1 Cowdry Type A Bodies → Can cause fatal neurologic disease in animal handlers HSV-infected cells → Usually from animal bites A. HERPES SIMPLEX VIRUS (HSV) HSV has two serotypes: → HSV-1: occurs in the oropharyngeal area → HSV-2: occurs in the genitals Both serotypes: → Differ serologically, but are closely related genetically → Similar set of primary and recurrent infections → Same viral sites of entry and replication: Skin and mucous membranes → Produce infectious virions Figure 11. HSV-infected cell exhibiting Cowdry type A bodies → Cause vesicular lesions of the epidermis HSV-2 Figure 11. HSV-infected cell exhibiting Cowdry type A bodies → Increases risk of HIV transmission by 4-fold Microscopically: → Increases risk of HIV acquisition by 2-3 fold → Pink to purple intranuclear inclusions [Robbins & Cotran, 2015] → Consist of viral replication proteins and virions YL6 04.39 Pathology of Viral Infections 5 of 16  Take Note! Doc used the same picture for Cowdry Type A bodies and Reed-Sternberg-like cells in her lecture Figure 6 is lifted from the book (Robbins & Cotran, 2015) to illustrate Cowdry Type A bodies Figure 15. Herpes epithelial keratitis Herpes Stromal Keratitis Another form of corneal lesion caused by HSV Microscopically: → Characterized by infiltrates of mononuclear cells around keratinocytes and endothelial cells via immunologic reaction → Leads to neovascularization, scarring, opacification of cornea → Can cause eventual blindness Figure 12. Cowdry type A bodies producing multinucleated syncytia (black arrows) Figure 12. Cowdry type A bodies producing multinucleated syncytia (black arrows) Microscopically: → Chromatin is pushed out to the edge of the nucleus → Cell fusion produces inclusion-bearing multinucleated syncytia Take Note! Figure 16. Herpes stromal keratitis The succeeding manifestations were mentioned only briefly. Thus, information regarding these was lifted from Robbins & Fever Blisters or Cold Sores Cotran (2015). Appearance: Doc recommends reading the “brown boxes” in Robbins for → Lesions that favor formation on the skin around mucosal oral other manifestations not mentioned in the lecture. orifices (i.e., lips, nose) Herpetic Whitlow Often bilateral patterns Independent of skin dermatomes Swollen, painful, erythematous lesions caused by HSV that Intraepithelial vesicles (blisters) are formed from intracellular appear on the fingers or palm edema and ballooning degeneration of epidermal cell Occur in infants → These often burst and crust over, sometimes resulting in Occasionally seen in healthcare workers (occupational exposure) superficial ulcerations Figure 13. Herpetic whitlow Gingivostomatitis Figure 17. Cold sores Appearance: Genital Herpes → Ulcers in the oral cavity → Vesicular eruption that extends from the tongue to the More often caused by HSV-2 retropharynx Appearance: Causes cervical lymphadenopathy → Vesicles on the genital mucous membranes and external Usually seen in children and is primarily caused by HSV-1 genitalia rapidly converted into superficial ulcerations → Rimmed by an inflammatory infiltrate HSV-2 infected mothers can transmit the virus to neonates during passage through the birth canal B. VARICELLA-ZOSTER VIRUS (VZV) Acute infection with VZV: chickenpox → Milder infections in children → More severe primary infection in adults and immunocompromised individuals Figure 14. Gingivostomatitis → Self-limiting disease → Infects mucous membranes, skin, and neurons Herpes Epithelial Keratitis Reactivation of latent VSV: shingles (herpes zoster) One of two forms of corneal lesions caused by HSV → Occurs in older and immunosuppressed individuals → Exhibits typical virus-induced cytolysis of the superficial → Seen in neurons and/or satellite cells around neurons in epithelium dorsal root ganglia → May recur many years after primary infection YL6 04.39 Pathology of Viral Infections 6 of 16  DIAGNOSIS → Involvement of the geniculate nucleus (rare) will induce facial paralysis Main methods of diagnosis include: → Viral culture → Detection of viral antigen in cells scraped from superficial lesions Vaccinations are recommended for individuals greater than 60 years old or immunosuppressed individuals → Due to them being more prone to developing severe forms of the disease or reactivation of the VZV latent phase CLINICAL MANIFESTATIONS Chickenpox Figure 21. Ramsay-Hunt Syndrome C. CYTOMEGALOVIRUS (CMV) Beta-group herpesvirus Disease manifestations depend on host’s age and immune status Infects monocytes and bone marrow progenitors → Healthy individuals: asymptomatic or mononucleosis-like symptoms → Neonates/immunocompromised individuals: devastating systemic infections TRANSMISSION Figure 18. Chickenpox lesions Occurs via several mechanisms that depend on the age group Primary infection of chickenpox (primary infection of VZV) affected → Lesions usually occur 2 weeks after respiratory infection Transplacental Transmission (Congenital CMV) → Appears as erythematous macule to vesicular lesions (“dew drops on a rose petal” appearance) Mother → Fetus → Centrifugal distribution of maculo-vesicular rash (spreads Occurs from a primary infection in a mother who does not have from the torso to the head to the extremities) protective antibodies Microscopic features Neonatal Transmission (Perinatal CMV) → Shows intraepithelial vesicles Mother → Fetus Characterized by intranuclear inclusions in epithelial cells Occurs through cervical/vaginal secretion at birth at the base of the vesicle Can also occur through ingestion of breast milk of mother with active infection Transmission via Saliva Children → Parents Occurs in children during preschool years (especially in daycare centers) Toddlers may readily transmit virus to parents via saliva Transmission via Genital Route Dominant route beginning 15 years old Other routes: respiratory secretions and fecal-oral route Iatrogenic Transmission Occurs at any age through organ transplantation or blood transfusion PATHOGENESIS Figure 19. Skin lesion of chickenpox (varicella-zoster virus) with intraepithelial vesicle Acute CMV Shingles → Induces transient but severe infections → Can infect dendritic cells Virus infects the keratinocytes and causes vesicular lesions Impairs antigen processing and ability of dendritic cells to → Unlike in chickenpox, the vesicular lesions are associated stimulate T lymphocytes with intense itching, burning, and sharp pain (due to concomitant radiculoneuritis) Radiculoneuritis: inflammation of one or more roots of the spinal nerves → Pain is especially severe when trigeminal nerve is involved Figure 20. Vesicular lesions in shingles Figure 22. “Owl’s eye” or classical Reed-Sternberg-like cells Ramsay Hunt Syndrome YL6 04.39 Pathology of Viral Infections 7 of 16  Infected cells are strikingly enlarged (up to 40 μm) Diagnosis: → Shows cellular and nuclear pleomorphism: → Serology Prominent intranuclear basophilic inclusions can also be Virus may continue to be secreted through the urine and seen saliva for months to years ○ Span half the nuclear diameter Infected individual remains seropositive for life ○ Set off from nuclear membranes by a halo/perinuclear Virus is never cleared clearing ○ Persisting in latently infected individuals Smaller basophilic inclusions can also be seen within the CMV in Immunosuppressed Individuals cytoplasm Commonly observed in transplant recipients and HIV patients Infected cells are also noted to have an “Owl’s eye” or Susceptible to severe CMV manifestations: Reed-Sternberg-like appearance → Pneumonitis (lungs) CMV evades immune defenses by: Can progress to respiratory distress syndrome (RDS) → Downmodulating MHC class I and II molecules → Colitis (GI tract) → Producing homologues of: Can progress to pseudomembranous colitis TNF receptor ○ Presents as debilitating diarrhea IL10 Diagnosis: MHC class I molecules → Antigen detection → Can evade NK cells by producing: → PCR-based assays Ligands that block activating receptors → Diagnostic methods have revolutionized the approach to Class I like proteins that engage inhibitory receptors CMV monitoring Congenital Infections 95% are asymptomatic ACTIVE RECALL If transplacental transmission occurs, classic cytomegalic 1. Cytomegalovirus can be acquired through the following inclusion disease develops mechanisms, EXCEPT: → This resembles erythroblastosis fetalis A. Transmission through saliva → Infant may develop: B. Blood transfusions Intrauterine growth restriction (IUGR) Jaundice C. Cervical/vaginal secretions Hepatosplenomegaly D. Respiratory droplets Anemia 2. T/F: After entry, HSV spreads to the motor neurons that Bleeding secondary to thrombocytopenia innervate the primary sites of replication. Encephalitis 3. T/F: Around 10% of adults have CMV antibodies. Fatal cases: microcephaly with calcifications Answer Key: 1D, 2F, 3F Diagnosis: → Viral culture or IV. TRANSFORMING VIRAL INFECTIONS → PCR of viral DNA in urine or saliva A. EPSTEIN-BARR VIRUS Infants with CMV or infected with transplacental transmission Causes infectious mononucleosis who survived: → Usually benign → May develop permanent deficits: → Self-limiting lymphoproliferative disorder Intellectual disability → Associated with several human tumors: Hearing loss Lymphomas Neurologic impairments Nasopharyngeal Carcinoma (NPCA) → Congenital Infections: Characterized by: Interstitial pneumonitis → Fever Hepatitis → Sore throat Neurologic disorders → Generalized lymphadenopathy Perinatal Infections → Splenomegaly Transmitted through birth canal or breast milk → Appearance of mononucleosis cells in the blood Usually asymptomatic due to protective maternal anti-CMV In the peripheral blood, atypical activated T lymphocytes antibodies may be appreciated → these are mononucleosis cells Many continue to secrete CMV in their urine and saliva for → Some develop hepatitis, meningoencephalitis, pneumonitis months to years All symptoms appear upon initiation of the host immune Less common complications: response → Interstitial pneumonitis Transmitted by close human contact → Failure to thrive → Frequently through the saliva during kissing → Rash EBV is also called “Kissing Disease” → Hepatitis Principally occurs in late adolescents or young adults → More common among upper socioeconomic classes in CMV Mononucleosis developed nations Commonly infects healthy young children and adults → May present with: Nearly always asymptomatic Fever 50-100% of adults have CMV antibodies around the world, Malaise indicating previous exposures Fatigue Most common clinical manifestation: infectious Lymphadenitis (most common) mononucleosis-like illness In the rest of the world → Fever → Primary infection occurs in childhood → Atypical lymphocytosis → Usually asymptomatic → Lymphadenopathy → Classically may present with: → Hepatitis Fever → Marked hepatomegaly Sore throat → Abnormal liver function test Lymphadenitis YL6 04.39 Pathology of Viral Infections 8 of 16  Resolves within 4 to 6 weeks of infection → 5-80% of these are large lymphocytes → Sometimes fatigue lasts longer Atypical lymphocytes (mononucleosis cells) Most common complication: Marked hepatic dysfunction → Sufficiently distinctive to strongly suggest the diagnosis → May present with jaundice, elevated hepatic enzymes, and disturbed appetite PATHOGENESIS 1. Transmitted by close human contact → Frequently through the saliva (during kissing) 2. EBV infects B cells and epithelial cells of the oropharynx → It is not known whether the source of the virus is from B cells, oropharyngeal epithelial cells or both 3. Spreads to the underlying lymphatic tissue → Hypothesized that EBV initially infects the oropharyngeal epithelial cells before spreading to underlying lymphoid tissue (tonsils and adenoids), where mature B cells are infected 4. Spread towards the tonsils and adenoids where the mature B Figure 24. Atypical lymphocytes cells are infected 5. Infection of B cells may take 2 forms: Minority and Majority → An EBV enveloped glycoprotein binds CD21 (CR2), the receptor for the C3D component of the complement, which is present on B cells → Minority of B cells Infection is lytic Leads to viral replication and eventual cell lysis accompanied by release of virions May infect other B cells in the process → Majority of B cells Figure 25. EBV specific CD8+ cytotoxic T cells and CD16+ NK cells EBV establishes latent infection Figure 25. EBV specific CD8+ cytotoxic T cells and CD16+ NK Virus persists as an extrachromosomal episome cells Large atypical lymphocyte → Has abundant cytoplasm with large nucleus Normal lymphocyte → More condensed chromatin/nucleus → More scant cytoplasm Lymph Nodes Typically discrete and enlarged throughout the body Reactive proliferation of T cells is largely centered in the lymphoid tissues → Accounts for lymphadenopathy and splenomegaly Lymphadenopathies may be observed in the posterior cervical, axillary, and inguinal regions Figure 23. Pathogenesis of EBV Take Note! Individuals with X-linked Agammaglobulinemia lack B cells and do not become latently infected with EBV nor shed virus This supports the theory that B-cells are the main reservoir of the infection Figure 26. Enlarged lymph nodes in patient with EBV infection CLINICAL MANIFESTATIONS Major alterations involve: → Blood → Lymph nodes → Spleen → Liver → Central nervous system → Occasionally other organs HISTOLOGICAL ANALYSIS Peripheral Blood Absolute lymphocytosis → >60% of WBCs are lymphocytes Figure 27. Comparison of lymphoid tissues YL6 04.39 Pathology of Viral Infections 9 of 16  → Showing expansion of white pulp follicles (paracortical area, Figure 27. Lymphoid tissues B-cell area, cortex, and germinal center) and red pulp Normal Lymphoid (left) sinusoids → See the paracortical area (dark purple area surrounding Vulnerable to rupture the germinal center) → Expansion of the paracortical area is due to the activation of T cells (immunoblasts) Hyperplastic lymphoid (right) → T cell proliferation may be exuberant Makes it difficult to distinguish the nodal morphology → Follicles (B cell area) may show mild hyperplasia Figure 30. Enlarged Spleen Liver Usually involved up to some degree Moderate hepatomegaly Histology → Atypical lymphocytes in portal areas and sinusoids → Scattered isolated cells or foci of parenchymal necrosis Figure 28. Photomicrograph of Centrocytes vs Other Cells Figure 28. Photomicrograph of Centrocytes vs Other Cells Centrocytes → Cleft nucleus compared tingible-body macrophages and mantle cells → Small, cleaved cells → Nucleus is at the center Centroblasts → Nucleus at the periphery → Larger cells → Open chromatin → Several nucleoli → Modest amount of cytoplasm Figure 31. Liver Biopsy Figure 31. Liver Biopsy Hepatitis caused by EBV Image IA (upper left): a portal and lobular lymphocytic infiltrate is spilling into the lobules with minimal hepatic cellular injury Image IC (lower left): sinusoidal lymphocytosis forms a string-of-beads pattern Antibodies IgM antibodies → Found early in the course → Formed against viral capsid antigen Figure 29. Centrocytes, centroblasts, and immunoblasts IgG antibodies → Found later on Figure 29. Centrocytes, centroblasts, and immunoblasts → Persists for life Centrocytes EBV CONTRIBUTES TO BURKITT LYMPHOMA → Cleaved cells Centroblasts Critical oncogenic event: chromosomal translocation involving → Larger cells MYC oncogenes → Open chromatin → Most commonly an 8:14 translocation → Multiple nucleoli Immunoblasts → Prominent single nucleoli → Heterogeneous population Spleen Enlarged in most cases (300-500 grams) Soft and fleshy Hyperemic cut surface Histology is analogous to lymph nodes Figure 32. Burkitt Lymphoma YL6 04.39 Pathology of Viral Infections 10 of 16  EBV AND HIV / IMMUNOSUPPRESSIVE THERAPY CLINICAL MANIFESTATIONS Those with HIV or are receiving immunosuppressive therapy: In adults: usually mild and nonspecific → EBV elicits lack of T-cell immunity → Fever → Unimpeded B-cell proliferation → Myalgia → Can be initiated by acute infection or reactivation of latent → Arthralgia B-cell proliferation → transforms into B-cell lymphoma → Conjunctivitis → Manifests in Duncan disease → Maculopapular rash (lasting a few days to weeks) An X-linked lymphoproliferative syndrome Neurologic complications (in small number of cases) Caused by mutations in the SH2D1A gene → Primarily Guillain-Barré syndrome ○ Encodes a signaling protein that participates in T-cell and NK cell activation and antibody production Take Note! EBV DIAGNOSIS Zika virus is sometimes misdiagnosed as dengue since its mild and nonspecific presentations are similar. Take Note! Perinatal transmission can result in: The procedures for EBV diagnosis are listed in increasing → Fetal death sensitivity, with rising titer of specific antibodies for EBV → Moderate to severe brain defects in the fetus/newborn antigens being the most sensitive. → Microcephaly Diagnosis: Occurs in a small number of infants (3.4% are those born → Lymphocytosis with characteristic atypical lymphocytes in to infected women with Zika virus) peripheral blood Half of the cases are described as proportionate → Positive heterophile antibody reaction (Monospot test) microcephaly Patient’s blood with IgM and IgG ○ Head size is small relative to gestational age ○ (+) EBV patients have IgM against viral proteins Other half have disproportionate microcephaly Mixed with sheep or horse RBCs and tested for reactivity ○ Head size is small relative to size of child ○ (+) reactivity = agglutination Sensitivity: high (90-95%) after 1st week but only 75% in the 1st week Specificity: not that specific (94%) ○ False positives: tests positive in cytomegalovirus (CMV), toxoplasmosis, acute HIV, malignancy, autoimmune diseases (e.g., SLE) Not recommended by the CDC given false (+) and (-). Figure 34. Microcephaly Other common manifestations in children: → Ocular abnormalities: Pigment mottling Chorioretinal atrophy Optic nerve abnormalities Figure 33. Monospot test for heterophile antibodies → Rising titer of specific antibodies for EBV antigens Viral capsid antigens, early antigens, or Epstein-Barr nuclear antigen V. OTHER ACUTE INFECTIONS A. ZIKA VIRUS Discovered in 1947 → Outbreaks in 2007, 2014, 2015) Figure 35. Morphology of a 2 month-old child with congenital Zika virus; (A) LPO Widespread in Africa, Asia, Middle East view (right), (B) HPO view (right) [Source, Year] Usually brought by travelers returning from affected areas Figure 35. → There are documented cases of community transmission of the virus Figure A (left) → Subcortical band with degenerating cells (red circle) Mosquito-borne (primarily by Aedes aegypti) Dark blue cells: degenerating neurons Perinatal transmission is documented Darker blue figures: calcifications → In Brazil, the rate of adverse effects on newborns following Figure B (right) maternal Zika virus infections was up to 46% → Degenerating neurons (green arrow heads) Highest rates were in the 1st and 2nd trimesters Hyperchromatic → Increased cases of microcephaly in newborns in Brazil Clumped up chromatin Individuals may also be infected through blood transfusion and Some nucleoplasm is spilling out of the cell → Viable neurons sexual contact Intact nucleus and nucleoli → Virus detectable in semen, vaginal secretions, breast milk, and urine B. DENGUE VIRUS Can be isolated from these bodily fluids up to several Transmitted by Aedes aegypti months CLINICAL MANIFESTATIONS The viability or degree of infectiousness of the virus is not Clinical manifestations vary from: yet documented → Fever with headache → Macular rash → Severe myalgias (breakbone fever) → Severe dengue (Dengue hemorrhagic fever) YL6 04.39 Pathology of Viral Infections 11 of 16  Bleeding Liver failure Reduced consciousness Organ failure Plasma leakage Severe dengue leads to shock and respiratory distress → There is widespread hemorrhages throughout the body with: Hepatic necrosis Mononuclear infiltrates Septal thickening Hyaline membrane formation in the lung IMMUNE RESPONSE Immune response against dengue virus is determined by the severity of infection Dengue virus has 4 serotypes Figure 38. Photomicrograph of a lung infected with dengue virus (higher magnification) [Molina, 2024] → Infection with each serotype stimulates protective immunity against that serotype Figure 38. → A cross-reactive antibody response is also stimulated Weak and non-protective of the other serotypes AM: mononuclear cells Being exposed to one serotype does not protect one from → Alveolar macrophages → Lymphocytes the other serotypes Mononuclear cells are increased in severe dengue Antibody-dependent enhancement → Manifested as difficulty breathing → Severe dengue usually occurs to those who have been previously infected with a different serotype Pathophysiology of Plasma Leakage in Dengue Video → Cross reactive antibodies enhance the uptake of virus into (CDC, 2018) macrophages via Fc receptors Scan the QR code to access the video → Infants who have maternal antibodies also have increased risk of developing severe dengue Plasma leakage → Process in which the protein-rich fluid component of the blood leaks from blood vessels into the surrounding tissue → Most serious complication that distinguishes dengue from severe dengue For some dengue patients, as fever begins to disappear, severe dengue will develop Figure 36. Photomicrograph of a normal lung [Molina, 2024r] Immune response process → Infected mosquito inserts its proboscis into the epidermal Figure 36. layer of human skin → Viral particles released then are taken up by antigen- A: alveolar spaces presenting cells (APCs) AS: alveolar septa → Virus replicates within the APC, causing it to mature → Mature APC enters an afferent lymphatic vessel → APC travels from the site of infection to the lymph node → In the lymph node, the mature APC produces chemical messengers that attract and activate T-cells In a patient with dengue, these activated T-cells react and secrete proinflammatory molecules (i.e., TNF-alpha, IFN-gamma, IL-6, and IL-8) In a patient with severe dengue, the production of pro-inflammatory molecules is significantly higher than in a patient with dengue → When these molecules enter the bloodstream, they help eliminate the virus and might also play a role in plasma leakage Glycocalyx → Endothelial surface glycocalyx along with the cell-to-cell junctions maintain the integrity of blood vessel → Network of proteoglycans and glycoproteins that projects from the surface of endothelial cells Figure 37. Photomicrograph of a lung infected with dengue virus [Molina, 2024] → Acts as the primary barrier against leakage of proteins and fluid across the vascular wall Figure 37. Mechanism of increasing vascular permeability in severe Mononuclear infiltrates dengue → Increased blue-stained cells along the septa → Not clearly understood Alveolar septa is thickened (blue circle; upper middle) → However, there is evidence suggesting that reactive Hyaline membrane formation oxygen species, enzymes, and proinflammatory molecules → Pink bands (black circle; lower right) start to breakdown the glycocalyx layer Septal necrosis → This allows plasma to reach the underlying intercellular → Crumbly-looking (blue circle; lower right) junctions and leak out into the tissues → The integrity of the blood vessel wall is altered YL6 04.39 Pathology of Viral Infections 12 of 16  Although plasma leaks from the blood vessels, the red HISTOLOGIC ANALYSIS blood cells are too large to pass into the tissue Diffuse Alveolar Damage (DAD) This causes a progressive increase in hematocrit (hemoconcentration) level beginning three to four Observed in the histological analysis of lung tissue from acute days after fever onset patients → Patients with severe plasma leakage will have a 20% or Previously known as acute respiratory distress (ARDS) more increase in hematocrit levels compared to baselines Characterized by alveolar collapse Effects of severe plasma leakage → Hypovolemic shock → Some are collapsed while others are distended → Pleural effusions and ascites → Lined by hyaline membrane → Respiratory problems Dense, thick, pink bands of proteinaceous material Treatment Unlike in necrosis where hyaline is crumbly, granular, or → Isotonic intravenous fluids and colloids are administered to fibrinous replace plasma Accompanied by inflammation Amount must be carefully monitored → Majority of inflammatory infiltrates are mononuclear cells → Vital signs, fluid intake and output, and hematocrit levels must be recorded similar to those seen in dengue (e.g., macrophages and Excessive IV fluid therapy can lead to pulmonary edema lymphocytes) and respiratory failure → Patients can improve rapidly over 1-2 hours if resuscitated properly However, care must be taken as leakage continues for 24-48 hours, during which time the patient may have several episodes of shock and look relatively well in-between C. SARS-COV2 (COVID-19) A novel coronavirus SARS-CoV-2-mediated disease (COVID-19) 1st detected in Wuhan, China (WHO, December 2019) Became a worldwide pandemic by March 2020 → 800,000 cases → 40,000 deaths Figure 40. Left: Diffuse alveolar damage (acute respiratory distress syndrome); Caused a major social and economic disruption, in addition to Right: Normal lung tissue [Molina, 2024] severe health consequences Figure 40. Diffuse alveolar damage Origin: seafood and animal market in Wuhan, China → Consistent with initial animal-to-human transmission Left figure: → Followed by person-to-person transmission → Collapsed and distended alveoli Sequencing of the full viral genome led to the development of → Hyaline membranes (arrows) → Blue cells are inflammatory infiltrates rapid molecular diagnostic assays (e.g., RT-PCR) Right figure is a normal lung tissue CLINICAL MANIFESTATIONS → A: Alveolar spaces → AS: Alveolar septae Range from mild to severe respiratory illness The vast majority of individuals who contract the virus recover ACTIVE RECALL after a flu-like disease In severe form and sometimes fatal illness: 1. Diffuse alveolar damage is characterized as the following → Respiratory compromise except: → Often associated with bilateral ground-glass opacities on A. Caseous pulmonary necrosis chest imaging B. Lined by hyaline membrane → Occurs mainly in older individuals and those with C. Accompanied by inflammation comorbidities D. Alveolar collapse Diabetes mellitus 2. T/F: Patients with X-linked Agammaglobulinemia lack T COPD Heart failure cells and do not become latently infected with EBV Obese 3. What do you call the phenomenon when severe dengue ○ Studies suggest that even younger obese individuals cases occur to those who have been previously infected also develop severe forms of COVID-19 with a different serotype? Genomic sequence shows COVID-19 is related to bat 4. Which cell does EBV infect? coronaviruses and the SARS coronavirus A. Macrophages B. T Cells C. B Cells D. Plasma Cells Answer Key: 1A, 2F, 3Antibody-dependent enhancement, 4C SUMMARY & KEY POINTS The different types of viral infections are the acute (transient) infections, latent (herpesvirus infections), and transforming viral infections. Acute infections are structurally heterogeneous and can elicit effective immune responses that eliminate pathogens. While latent infections persist in a non-infectious form that is periodically reactivated. Measles as an acute viral infection can vary from mild, self-limited to severe or systemic. It can present as Measles Figure 39. Bilateral ground-glass opacities seen in COVID-19 patients Rash, Koplik Spots, Germinal Cell Hyperplasia, and/or Warthin-Finkeldey Cells. YL6 04.39 Pathology of Viral Infections 13 of 16  Mumps can lead to an acute systemic infection with varying 5. What is the correct sequence of events in the pathogenesis severity. Initially, the manifestation is pain and swelling of salivary of HSV? glands. Main clinical manifestations include mumps parotitis I. Virus spreads to the sensory neurons that innervate the (inflammation of the parotid gland), mumps orchitis (testicular primary sites of replication swelling), and mumps pancreatitis. II. HSV-1 enters through the mouth while HSV-2 enters Poliovirus has a wide range of manifestations. It can be mild and through the genital area through sexual intercourse self-limited severe infection can cause paralysis of the limb III. Viral nucleocapsids are transported along axons to the neuronal cell bodies where the viruses establish latent muscle and respiratory muscles. infection In the West Nile virus, humans are accidental hosts. A. III – I – II Viral Hemorrhagic Fever, caused by Arenaviridae, Filoviridae, B. II – I – III Bunyaviridae, and Flaviviridae, may result in vascular C. I – II – III dysregulation & damage, and eventual shock. D. III – II – I Herpesviruses that can cause latent infections can be divided into the following groups: α-group (HSV-1, HSV-2, VZV), β-group 6. What is a pathognomonic physical finding of measles infection? (CMV, HHV-6, HHV-7), and γ-group (EBV, HHV-8). Varicella-Zoster Virus (VSV) manifests as chickenpox in an acute A. Measles rash infection and manifests as shingles as a reactivation of latent B. Koplik spots VSV. Chickenpox appears as erythematous macule to vesicular C. Warthin-Finkeldey cells lesions (“dew drops on a rose petal” appearance) while shingles D. Keratitis appears as vesicular lesions are associated with intense itching, 7. Which of the following is NOT TRUE about Herpesvirus? burning, and sharp pain. A. α-group include HSV-1, HSV-2, and CMV Epstein-Barr Virus (EBV), transmitted through close contact B. It causes acute infections followed by latent infections (often via saliva – “Kissing Disease”), causes infectious C. An acute Herpesvirus infection may be followed up by a latent mononucleosis, typically presenting with fever, sore throat, infection lymphadenopathy, and splenomegaly. It is usually benign and D. It is a large, encapsulated virus with a double-stranded DNA self-limiting, though it may be associated with certain tumors and 8. [True or False] Lungs infected with the dengue virus present complications like hepatitis, with symptoms resolving in 4-6 with thickened alveolar septa and polymorphonuclear weeks. infiltrates in their morphology. Dengue virus is transmitted by the Aedes aegypti species of 9. Modified T/F Question Format mosquitoes presents with symptoms such as fever with headache and macular rash The SARS-CoV2 was initially spread via person-to-person Severe dengue (dengue hemorrhagic fever) can cause bleeding, transmission in a seafood & animal market in Wuhan, China Because of this, the virus quickly became a global pandemic by reduced consciousness, plasma leakage and organ failure. It can March of 2020. ultimately lead to shock and respiratory distress COVID-19 patients may show bilateral ground-glass opacities on A. The first statement is true chest imaging and may have diffuse alveolar damage B. The second statement is true characterized by alveolar collapse. C. Both statements are true D. Both statements are false REVIEW QUESTIONS 10. Which is an incorrect statement regarding COVID-19? 1. Which species is INCORRECTLY matched with the disease it causes? A. Diffuse alveolar damage in acute patients B. Bilateral ground-glass opacities on chest imaging A. Rotavirus: Gastroenteritis C. Hyaline membrane is dense, thick, and pink bands of B. Hepatitis A: Acute viral hepatitis proteinaceous material C. Herpes simplex virus 2: Oral herpes (“cold sore”) D. Hyaline membrane is crumbly, granular, or fibrinous D. JC virus: Progressive multifocal leukoencephalopathy (opportunistic) Answer Key 1C, 2C, 3T, 4A, 5B, 6B, 7A, 8F, 9B, 10D 2. The following connections are true, EXCEPT: RATIONALE TO REVIEW QUESTIONS A. Reed-Sternberg-like cells = Cytomegalovirus B. Vascular lesions with intense itching, burning, and sharp pain = 1. [C] — Herpes simplex virus 1 (HIV-1) causes oral herpes, also Shingles known as the cold sore. Herpes simplex virus 2 (HIV-2) causes C. Ramsay-Hunt Syndrome = Epstein-Barr Virus genital herpes. D. “Dew drops on a rose petal” appearance of lesions = 2. [C] — Ramsay-Hunt Syndrome can occur when the geniculate Chickenpox nucleus is involved in a Shingles infection. The rest of the corrections are correct. 3. [True or False] Perinatal Infections are transmitted through 3. [T] — Perinatal infections are transmitted through the birth the birth canal or breast milk. canal or breast milk. 4. Modified T/F Question Format 4. [A] — In an EBV infection, it is in the majority of B cells that EBV is transmitted by close human contact. the virus persists as an extrachromosomal episome, and thus, In such an infection, the virus establishes a latent infection infection is latent. Meanwhile, infection is lytic for a minority of B where it persists as an extrachromosomal episome in a minority cells. of B cells. 5. [B] — This is the correct sequence of events. A. The first statement is true 6. [B] — Koplik spots are the pathognomonic physical finding in B. The second statement is true patients with measles infection C. Both statements are true 7. [A] — Cytomegalovirus (CMV) belongs to the β-group. The rest D. Both statements are false are all characteristics of Herpesvirus 8. [F] — Lungs infected with dengue virus present with thickened alveolar septa and mononuclear infiltrates 9. [B] — Statement 1 is false because SARS-CoV2 initially spread via animal-to-person transmission 10. [D] — Crumbly, granular, and fibrinous are descriptions of hyaline membrane in tissue necrosis, not COVID-19 patients. YL6 04.39 Pathology of Viral Infections 14 of 16  REFERENCES REQUIRED REFERENCES Kumar, V., Abbas, A. K., & Aster, J. C. (2021). Robbins & Cotran [Textbook] Pathologic Basis Of Disease (10th ed., pp. 349–358). Elsev