Mousepox, Liver, Mouse 145

logical techniques applied to the study of physiologi• Toolan HW (1968) The picodna viruses H, RV, and AA V. cal and pathological growth. Exp Mol Pathol 5: 146- Int Rev Exp Pathol6: 135-180 181 Yanoff M, Rawson AJ (1964) Peliosis hepatis. An anatom• Toolan HW (1960) Experimental production of mongoloid ical study with demonstration of two varieties. Arch Pa• hamsters. Science 131: 1446-1448 thol77: 159-165

Mousepox, Liver, Mouse

Robert O.Jacoby

Synonyms. Infectious ectromelia ration or can shrink and develop intensely eosino• philic cytoplasm. Hepatocytes with clear cyto• plasm can occur at some distance from necrotic Gross Appearance foci and have reportedly undergone glycogen de• pletion (De Burgh 1950). Nuclei of infected cells The liver is a major site of viral replication in may be transiently enlarged, but quickly become mousepox, but gross lesions, even during acute pyknotic or karyorrhectic. disease, are not readily apparent until shortly be• Two types of intracytoplasmic inclusions occur in fore death. Severely affected livers are usually mousepox: the early or type B inclusion and the swollen and friable and may occupy up to half the late or type A inclusion. Type B inclusions com• volume of the peritoneal cavity, whereas mildly monly develop in the liver, whereas type A inclu• affected livers may remain grossly normal or have sions are rare. Type B inclusions are basophilic to sparse focal necrosis. Necrotic areas appear first amphophilic and can occur singly or in groups as pinpoint yellow-white foci, but increase rapidly (Fig. 126). They may be surrounded by a thin "ha• in size and number. Confluent areas of necrosis lo" but this is often hard to see in formalin-fixed can produce a reticulated pattern of yellow• sections. Type B inclusions also are difficult to de• brown to pink discoloration on the surface and tect unless the hematoxylin staining time is at throughout the parenchyma. Areas of hemor• least doubled (Allen et al. 1981). Alternatively, be• rhage also may develop. The pale hue of severely cause they are sites of viral replication (Cairns affected livers is in part due to fatty change and 1960; Kameyama et al. 1959), they can be detect• the fat content of such livers can be as much as ed in formalin-fixed liver by immunohistochemi• four times normal. Livers from mice that survive cal methods. Immunoperoxidase staining of in• acute infection usually have a normal gross ap• fected hepatocytes will reveal intracytoplasmic pearance. A few small scars may be present, how• antigen in particulate and diffuse distribution ever, especially at the margins (Fenner 1948d, (Fig. 127). Viral antigen also can be found in cells 1949b: Allen et al. 1981). lining vascular channels, especially during early phases of infection. Type A inclusions are large and intensely eosinophilic. They can more readily Microscopic Features be detected in other tissues, particularly skin (Fig. 128), mucous membranes, and intestinal epi• The major liver lesion is coagulation necrosis, thelium, to help confirm the etiology of hepatic le• which begins in random fashion among individu• sions. al or small groups of hepatocytes approximately The inflammatory response during fulminating 5 days after infection (Fig. 125). In highly suscep• hepatic necrosis is minimal. Some mononuclear tible mice, necrotic areas enlarge rapidly and in cells and polymorphonuclear leukocytes may in• 2-4 days lead to extensive necrosis and variable filtrate portal triads or the margin of necrotic le• degrees of hemorrhage. sions. In milder or more prolonged hepatic infec• Hepatocytes in early lesions or at the margins of tion, such as occurs in genetically resistant mice, advanced lesions can undergo ballooning degene- inflammatory infiltrates are more prominent. The

T. C. Jones et al. (eds.), Digestive System © Springer-Verlag Berlin Heidelberg 1985 146 Robert O.Jacoby Mousepox, Liver, Mouse 147

mononuclear cell response is compatible with host defenses mediated by cellular immunity (Blanden 1974). The histologic sequelae of mousepox in the liver have not been described in detail, but appear to be unremarkable. This is probably due to the fact that extensive hepatic involvement commonly leads to rapid death, whereas mild hepatic lesions are quickly repaired. Large or multinucleated hepatocytes can accumulate at the margin of ne• crotic lesions during repair. Chronic hepatitis has not been reported. "Hyalinized" areas have been found and may represent local postnecrotic scar• ring, but widespread fibrosis or cirrhosis does not occur.

Ultrastructure

The most complete observations have been made by Leduc and Bernhard (1962) on livers of natu• rally infected mice. The earliest signs of infection are seen among periportal hepatocytes. Cytoplas• mic changes are characterized by loss of glycogen, vesicular swelling of the endoplasmic reticulum, mitochondria, and Golgi apparatus, and sparse Fig. 129. Hepatocyte infected with mousepox. Matrix lipid droplets (Fig. 129). Irregular dense bodies zone (encircled by dotted line) contains developing virus develop and are found occasionally in nucleo• particles. Incomplete c-shaped particles presumably repre• plasm. Nucleolar fragmentation is also observed. sent immature virions (arrow). (From Leduc and Bernhard In 1962 with permission of Journal of Ultrastructural Re• moribund or necrotic cells, the cytoplasm is search). TEM, x 17 000 dense and, in addition to viral particles, contains membranous whorls, vesicles of various sizes, vacuoles with cell debris and collapsed, distorted mitochondria. Nuclear chromatin is dense and Virus formation begins in granular to reticular marginated and blisters form in the nuclear enve• cytoplasmic matrices of low electron density lope. Although cells lining hepatic sinusoids sup• (Fig. 129). Matrix zones displace the cytoplasm port viral replication they do not undergo necro• and lack sharp boundaries. They are thought to be sis. the ultrastructural correlates of type B inclusions. Three early forms of virus emerge from the ma• trix: (1) 220 m~, membrane-bound, oval particles with uniform viroplasm similar in appearance to

Fig. 128 (lower right). Type A inclusions of ectromelia virus (arrows) in the epidermis of a mouse. Hand E, x 450 148 Robert O.Jacoby that is not compatible with mousepox. Mouse cy• tomegalovirus and lymphocytic choriomeningitis virus can cause hepatic necrosis after experimen• tal inoculation, but infection with these agents is not prevalent among well-managed mouse colon• ies. Differentiation of the various forms of viral hepa• titis in mice can be difficult. Therefore, in the ab• sence of specific morphological changes, ancil• lary data should be collected to confirm the diag• nosis. These include one or more of the following: viral serology, immunohistochemical demonstra• tion of viral antigen in tissue, electron microscopy for viral particles, and, when feasible, viral isola• tion. Tyzzer's disease should be considered whenever Fig.130. Two types of mousepox viral particles. Oval par• hepatic necrosis is found in mice. The causative ticles with uniform viroplasm (arrow) and particles with organism, Bacillus pilijormis, is difficult to find in nucleoids of precisely aligned filaments. (From Leduc and hematoxylin and eosin stained sections, but can Bernhard 1962 with permission of Journal of Ultrastrnctu• ral Research). TEM, x 53000 be demonstrated with special stains (Giemsa, pe• riodic-acid-Schiff, Warthin-Starry) (see p. 156, this volume). Organisms are generally found in Differential Diagnosis the cytoplasm of hepatocytes bordering necrotic areas. They may be present in small numbers, Several naturally occurring murine viruses cause making it necessary to examine several sections to necrotizing hepatitis in mice. Hepatotropic strains locate them. Although the acute form of other of mouse coronaviruses (mouse hepatitis viruses) bacterial infections such as salmonellosis may may induce acute multifocal hepatic necrosis, but cause necrotizing hepatitis, these organisms are lesions, especially in adult mice, are seldom as se• rarely found in modem vivariums. Nevertheless, vere as those of lethal mousepox (see p. 134, this it is advisable to obtain bacterial cultures from volume). Because the course of coronaviral hepa• mice with hepatic necrosis. titis is often longer than that of mousepox, signs of repair (mitotic activity, large or binucleated hepatocytes) can be found more commonly at the Biologic Features margins of necrotic lesions. Inflammation also may be more prominent. Syncytia, which are The clinical expression of mousepox can vary and common in mouse coronavirus infection, are not is influenced strongly by genotype (Briody et al. typical of mousepox, whereas viral inclusions are 1956; Briody 1966; O'Neill and Blanden 1983; not found in coronaviral hepatitis. Extrahepatic Bhatt and Jacoby 1985). Broadly speaking, three manifestations, especially necrosis of spleen and clinical courses are recognized. Highly suscepti• other tissues, are also more severe in mousepox. ble mice develop acute, fatal disease wherein ap• Several strains of mouse coronavirus are neuro• parently healthy mice may die within several tropic, whereas mousepox does not cause signifi• hours from the onset of illness. This form of cant lesions of the central nervous system unless mousepox is closely associated with severe hepat• virus is inoculated intracerebrally. ic necrosis. Clinical signs are relatively nonspecif• Reovirus-3 can cause necrotizin hepatitis in infant ic and include hunched posture, rough haircoat, mice, but not in adults (see p. 151, this volume). and diarrhea. If infection has occurred through Necrosis tends to begin in centrilobular zones and skin abrasion, careful examination of the skin to spread peripherally (Walters et al. 1963). Mice may reveal a "primary lesion", which indicates that recover may develop chronic active hepatitis the initial site of viral replication (Fenner 1947). that has immunopathologic overtones expressed Moderately susceptible mice often develop a by a significant, persistent inflammatory response chronic form of mousepox with variable mortali• (Stanley 1974). The disease in infant mice is fre• ty. It is characterized by erosive or ulcerative quently accompanied by necrosis in other organs; dermatitis which is most easily observed on the most frequently among these is brain, a finding ears, tail, and feet, but which also may occur as a Mousepox, Liver, Mouse 149 general exanthema. These mice can also have ede• remia during which virus is widely distributed to ma of the face and extremities and conjunctivitis. skin and causes a typical pox rash. Recovery from Amputation of part or all of one or more limbs or mousepox and resistance to lethal infection de• the tail can occur, a lesion that gave rise to the pends on intact cellular immunity (Blanden 1974). term "infectious ectromelia." Skin lesions among Evidence that humoral responses are critical to survivors of chronic mousepox heal as hairless host survival are less compelling (Schell 1960a, scars. Resistant mice commonly have asympto• b). matic infection (Fenner 1982). Ectromelia virus is an orthopoxvirus that is mor• From an epizootiologic aspect, acute, lethal phologically identical and antigenically similar to mousepox is a major hazard to a mouse colony, vaccinia virus, a relationship which enables vacci• but is self-limiting provided that additional sus• nia virus to elicit protective immunity to mouse• ceptible animals are not introduced during an pox (Briody 1959). The virus grows in a number of outbreak. Chronic or asymptomatically infected continuous cell lines including L cells and Vero mice are a relatively greater hazard in the long run cells and classic pox lesions are produced if virus because they can perpetuate enzootic infection is grown on the chorioallantoic membrane of em• (Fenner 1948 a, b). Introduction of highly suscep• bryonated hen eggs (Fenner 1982). A line of kid• tible mice to an enzootically infected colony may ney cells (BS-C-I), derived from African green initiate a clinically explosive and devastating out• monkeys, are highly susceptible to ectromelia vi• break. rus (Bhatt and Jacoby 1985). The prevalence and biologic significance of la• Several strains of ectromelia virus have been stud• tent, persistent mousepox infection (carrier state) ied intensively. The Moscow strain is highly viru• is unsettled. Evidence for a carrier state is sparse lent, whereas the Hampstead strain is less so (Fen• (Gledhill 1962; Horzinek and Hapken 1965), but ner 1949b). Avirulent strains of Hampstead virus has not been thoroughly or systematically dis• replicate poorly in hepatic Kupffer's cells, but counted. If a carrier state occurs, its expression is readily invade hepatocytes. It has been proposed likely to be influenced by mouse genotype. Briody from this that virulence may depend on the ability and coworkers (Briody et aL 1956) and Briody of virus to infect Kupffer's cells and that survival (1959), for example, studied responses of mice to of the host may depend on the ability of Kupffer's natural epizootics of mousepox and found that cells to protect hepatocytes from viral invasion some inbred strains such as DBAlI, A and C3H (Roberts 1964). A strain of ectromelia virus (NIH-- were highly susceptible to lethal infection, where• 79) was isolated from a recent mousepox epizoo• as other strains such as C57BL/6, AKR, and tic in the United States (Allen et aL 1981). Its be• BALBIc were highly resistant. Although these havior in mice is under scrutiny, but it seems to be findings underscore the significance of genotype moderately to highly virulent (Bhatt and Jacoby for the outcome of infection, they should be ex• 1985). trapolated cautiously, because recent evidence All strains of ectromelia virus studied thus far ex• suggests that susceptibility to lethal mousepox press an immunogenic hemagglutinin. Infected can vary even among sublines of a given inbred animals develop hemagglutinin-inhibiting anti• strain (Bhatt and Jacoby 1985). body which can be detected by routine serologic Fenner (1947, 1948 a, b, c, d; 1949a, b) is largely methods (Briody 1966). The sensitivity and speci• responsible for deciphering the pathogenesis of ficity of hemagglutination inhibition has been the mousepox. Although ectromelia virus can enter center of some controversy and current evidence the body by several routes, including the respira• suggests that the hemagglutination inhibition test, tory tract, the most common mode of entry is although still widely used, may elicit occasional thought to be through abraded skin. Virus repli• false-positive results (Collins et al. 1981; Wallace cates at the site of entry and infects the draining et aL 1981). Other serologic tests used to detect in• lymph node. It reenters the lymphatics and finally fection include complement fixation (relatively reaches the blood to produce a primary viremia. insensitive), immunofluorescence (Christensen et During primary viremia, virus invades parenchy• aL 1966) virus neutralization, and enzyme-linked mal organs, including the liver. Immunofluores• immunoadsorbent assay (ELISA). The ELISA is cent studies by Mims (1964) disclosed that blood• highly sensitive, but it must be refined before it borne virus first infects littoral cells lining vascu• can be used to discriminate ectromelia-infected lar channels and that infected cells seed virus to from vaccinated mice (Buller et al. 1983; Collins hepatocytes. Mice that survive initial infection of et al. 1981). Hemagglutinin inhibition (HAl) is parenchymatous tissues develop a secondary vi- still the test of choice for this purpose. The strain 150 Robert O.Jacoby of vaccinia virus used to immunize mice (IHD-T) Cairns J (1960) The initiation of vaccinia infection. Virolo• is hemagglutinin deficient (Briody 1959). Thus gy 11: 603-623 vaccinia-immune mice should remain free of HAl Christensen LR, Weisbroth S, Matanic B (1966) Detection of ectromelia virus and ectromelia antibodies by immu• antibody to either vaccinia virus or ectromelia vi• nofluorescence. Lab Anim Care 16: 129-141 rus. Collins MJ Jr, Peters RL, Parker JC (1981) Serological de• Naturally occurring mousepox is limited to mice. tection of ectromelia virus antibody. Lab Anim Sci 31: Outbreaks in laboratory mice have occurred in 595-598 Europe, Asia, Australia, and the United States. De Burgh PM (1950) Cytochemical changes in early ec• The risk of introducing infection to a susceptible tromelia infection of mice. Aust J Exp BioI 28: 214-218 Fenner F (1947) Studies in infectious ectromelia of mice. population is increased by extensive exchange of II. Natural transmission: the portral of entry of the vi• mice and mouse tissues among laboratories. The rus. Aust J Exp BioI 25: 275-282 spread of mousepox in a newly infected colony Fenner F (1948a) The epizootic behavior of mouse-pox depends on a combination of factors such as (infectious ectromelia). Br J Exp Pathol29: 69-91 mouse genotype, husbandry, viral virulence, and Fenner F (1948 b) The epizootic behavior of mousepox (in• experimental manipulation. Although mousepox fectious ectromelia of mice). II. The course of events in is an infectious disease, it can spread slowly even long-continued epidemics. J Hyg 46: 383-393 Fenner F (1948c) The pathogenesis of the acute exan• among mice housed in the same room (Wallace et thems. An interpretation based upon experimental in• al. 1981). In utero infection has been produced vestigations with mousepox (infectious ectromelia of experimentally (Schwanzer et al. 1975; Wylek• mice). Lancet 2: 915-920 shanin 1935), but its prevalence and significance Fenner F (1948 d) The clinical features and pathogenesis of in naturally occurring disease have not been es• mousepox (infectious ectromelia of mice). J Pathol Bac• tablished. teriol60: 429-552 Fenner F (1949a) Studies in mousepox (infectious ectro• melia of mice). VII. The effect of the age of the host up• on the response to infection. Aust J Exp BioI 27: 45-53 Comparison with Other Species Fenner F (1949b) Mouse-pox (infectious ectromelia of mice): a review. J Immunol63: 341-373 The skin form of mousepox has been studied ex• Fenner F (1982) Mousepox. In: Foster HL, Small JD, Fox tensively as a model of a viral exanthem because it JG (eds) The mouse in biomedical research, vol II, Dis• resembles human smallpox (Fenner 1948c). He• eases. Academic, New York, chap 11 patic lesions, however, are not characteristic of Gledhill AW (1962) Latent ectromelia. Nature 196: 298 smallpox or of poxvirus diseases of domestic Greene HSN (1934) Rabbitpox. I. Pathology of the ep• idemic disease. J Exp Med 60: 441-457 mammals. A notable exception is rabbit pox, Horzinek M, Hopken W (1965) Untersuchungen fiber die which tends to be severe and generalized and is inapparente Infektion mit dem Ektromelievirus. Arch frequently accompanied by hepatic necrosis Ges Virusforsch 17: 125-138 (Greene 1934). Kameyama S, Takahashi M, Toyoshima ~ Kato S, Kama• hora J (1959) Studies on the inclusion bodies of ectro• melia virus using the fluorescent antibody technique. Bi• Reforences ken J 2: 341-344 Leduc EH, Bernhard W (1962) Electron microscope study of mouse liver infected by ectromelia virus. J Ultrastruct Allen AM, Clarke GL, Ganaway JR, Lock A, Werner RM Res 6: 466-488 (1981) Pathology and diagnosis of mousepox. Lab Anim Mims CA (1964) Aspects of the pathogenesis of virus dis• Sci 31: 599-608 eases. Bact Rev 28: 30-71 Bhatt PN, Jacoby RO (1985) The pathogenesis of mouse• O'Neill HC, B1anden RV (1983) Mechanisms determining pox in genetically resistant and genetically susceptible innate resistance to ectromelia virus infection in C57BL inbred mice. (in preparation) mice. Infect Immun 41 : 1391-1394 B1anden RV (1974) T cell response to viral and bacterial in• Roberts JA (1964) Growth of ectromelia virus in the liver fection. Transplant Rev 19: 56-88 parenchymal cells of different strains of mouse. Nature Briody BA (1959) Response of mice to ectromelia and vac• 202: 1140-1141 cinia viruses. Bacteriol Rev 23: 61-95 Schell K (1960a) Studies on the innate resistance of mice Briody BA (1966) The natural history of mousepox. Nat! to infection with mousepox. I. Resistance and antibody Cancer Inst Monogr 20: 105-115 production. Aust J Exp BioI Med Sci 38: 271-288 Briody BA, Hauschka TS, Mirand EA (1956) The role of Schell K (1960b) Studies on the innate resistance of mice genotype in resistance to an epizootic of mouse pox (ec• to infection with mousepox. II. Route of inoculation tromelia). Am J Hyg 63: 59-68 and resistance; and some observations on the inheri• Buller RM, Bhatt PN, Wallace GD (1983) Evaluation of an tance of resistance. Aust J Exp BioI Med Sci 38: 289-300 enzyme-linked imrilUnosorbent assay for the detection Schwanzer V, Deerberg F, Frost J, Liess B, Schwanzerova of ectromelia (mousepox) antibody. J Clin Microbiol18: I, Pitterman W (1975) Zur intrauterinen Infektion der 1220-1225 Reovirus Type 3 Infection, Liver, Mouse 151

Maus mit Ektromelie-virus. Z Versuchstierkd 17: Walters MN, Joske RA, Leak PJ, Stanley NF (1963) Mu• 110-120 rine infection with reovirus. I. Pathology of the acute Stanley NF (1974) The reovirus murine models. Prog Med phase. Brit J Exp Pathol44: 427-436 Viro118: 257-272 Wylekshanin AJ (1935) Transplancental transmission of Wallace GO, Werner RM, Golway PL, Hernandez OM, the filterable virus of infectious ectromelia. Z Ihikrobiol Alling OW, George OA (1981) Epizootiology of an out• (Moscow) 15: 433 break of mousepox at the National Institutes of Health. Lab Anim Sci 31: 609-615

Reovirus Type 3 Infection, Liver, Mouse

Stephen W. Barthold

Synonyms. Reo-3 virus infection; hepatoence• lations of mixed leukocytes. Initially, bile ducts phalomyelitis virus (HEV) infection; ECHO 10 are dilated and contain esosinophilic and necrotic virus infection. cellular debris, followed by infiltration of portal areas with leukocytes (Fig. 132). Parenchymal mi• totic activity is increased. Vascular congestion, Gross Appearance hemorrhage, foci of neuronal degeneration and Natural infection of adult mice with reovirus type necrosis, perivascular leukocytic infiltrates, and 3 is asymptomatic. Infection of neonatal mice can nonsuppurative meningitis are present in the result in runting, emaciation, jaundice, bilirubinu• ria, conjunctivitis, incoordination, tremor, paraly• sis, and oily hair effect. Survivors are runted with transient alopecia, which is most marked along the dorsum of the head, neck, and rear legs. Yel• low or blood-tinged peritoneal exudate can be present. Livers are enlarged and dark with multi• ple sharply demarcated yellow foci, up to 3 mm in diameter on all surfaces. Gallbladders are dis• tended with dark bile. The intestine is reddened and distended with yellow digesta. Fibrinous ex• udate and small gray foci have been observed on the epicardium. Other lesions include general pal• lor, focal pulmonary hemorrhages, pale kidneys, and atrophy of thymus, lymph nodes, and spleen. Chronically infected mice can have recrudescence of jaundice and liver lesions (Bennette et al. 1967 a; Cook 1963; Stanley et al. 1953, 1954; Wal• ters et al. 1963).

Microscopic Features Lesions are generalized in mice exposed as neo• nates. Sharply demarcated foci of acute hepato• cellular coagulation necrosis surrounded by vari• able leukocytic infiltrates are evident, particularly in the subcapsular parenchyma and near centri• Fig.131. Focal necrotizing hepatitis in an infant mouse lobular veins (Fig. 131). This is followed by prolif• experimentally infected with reovirus type 3. Hand E, eration of Kupffer's cells and sinusoidal accumu- x 100