- Email: [email protected]
“B” Virus Disease in Monkey and Man
Brit. vet. ]. ( 1966),
122,
46
"B" VIRUS DISEASE IN MONKEY AND MAN BY
E. G.
HARTLEY
National Institute for Medical Research, Mill Hill, London
=============
SUMMARY
"B" virus causes a benign disease of monkeys characterized by a herpes-like stomatitis which heals spontaneously in seven to I4 days. However, when the virus is established in man by accidental monkey-bite or from infected tissues or fluids, it may produce a fatal encephalitis or encephalomyelitis. A clinical description of the disease is given and methods of diagnosis in monkey and man are described. Methods to reduce the risk ofinfection to those closely involved in the handling of monkeys or their tissues are suggested. HISTORY
Sabin & Wright ( I934a) first isolated a virus from the central nervous system of a laboratory worker who had died after a bite from an apparently normal rhesus monkey. The virus was called by Sabin "B" virus. No further case was reported until I 949 when a case from similar circumstances was reported by Sabin (I949). From the mid-fifties onwards, however, with the increasing use of monkeys for the production and testing of biological products, principally the poliomyelitis vaccines, and as a source of tissue culture material for virus studies, the significance of the disease increased considerably. To date more than fifteen cases have been reported in man, all fatal except two. All have occurred in persons closely associated with either monkeys or their tissues. Keeble, Christofinis & Wood (I958) first noticed herpes-like ulcers on the tongue and lips of rhesus monkeys they were examining. A virus was isolated from these lesions which they showed to be "B" virus. INCIDENCE AND AETIOLOGY
The incidence of the disease in the commoner laboratory monkeys, namely the rhesus (Macaca mulatta) and the cynamolgus (Macaca ira) is fairly high. Keeble (I 96o) gives a figure of 2 ·3 per cent clinical cases seen in I 4,400 rhesus examined by him. Hartley (I 964) gives an incidence of 3 per cent with lesions in I ,ooo cynamolgus monkeys examined. Despite the lack of evidence there is no reason to doubt that the disease can occur also in many other species of monkey. A seasonal increase in the natural disease occurs with the monsoon and post-monsoon period, i.e. from October to February, when the majority of clinical cases are encountered (Keeble, Ig6o; Hartley, I964). It is evident also from neutralizing antibodies detected in apparently normal monkeys that the incidence is in fact very widespread. Some workers have found up to I oo
"B" VIRUS DISEASE IN MONKEY AND MAN
47
per cent in batches of imported monkeys. The natural infection in the monkey seems to follow a similar pattern to the primary stomatitis seen in man. It has been suggested by Burnet, Lush & Jackson (1939) that herpes simiae ("B" virus) and herpes simplex were at one time the same virus before adaptation to their own specific hosts. There is, however, no evidence to suggest at present that, like herpes in man, "B" virus persists in monkeys in a latent form after the initial lesions have healed. The virus in monkeys appears to spread within the colony by the contamination of food and water from infective saliva, and by direct inoculation of the skin by bites or scratches. In man every reported case except one has been accompanied by a history of either bites, scratches, or other close contact with monkeys. The exception was a case reported by Davidson & Hummeler (1960) in which the fatality occurred in a laboratory worker handling tissue cultures of monkey renal cells. Their evidence also suggests that the virus may be spread by gross aerosols to personnel working in the monkey rooms. CLINICAL FEATURES
In the monkey the gross lesions are most readily recognized on the surface of the tongue (Fig. 1). These lesions take the form of small vesicles which soon rupture to form an ulcer (Fig. 2) covered by a necrotic scab. This scab is yellowish-grey in colour and is sharply demarcated from the surrounding tissues. Within seven to 14 days these ulcers heal spontaneously, leaving no scar. During the period of stomatitis the animal does not appear to suffer any discomfort and continues to eat satisfactorily. Although the majority of these lesions appear on the tongue either singly or in groups, ulcers may also be seen on the mucoepithelial border of the lips. Lesions have also been reported on various parts ofthe skin, due no doubt to bites received during fighting. Secondary infections of a bacterial or fungal nature can also occur. As with herpes simplex, occasional involvement of the central nervous system takes place, and lesions have been seen during histological examinations of monkey brain and cord. Lesions seen are mild in character, and do not appear to give rise to clinically evident signs. In man the clinical symptoms have varied considerably but the general picture is that of an encephalitis or encephalomyelitis. The local area of the bite may show pain, redness and swelling, with the formation of vesicles and exudate. Often, associated lymphangitis and lymphadenitis occur. In all cases the fatal termination was within three weeks of illness and was in one case as early as three days. In both the cases which recovered there was severe residual damage to the central nervous system. The original case described by Sabin & Wright (1934b) was of an acute ascending myelitis with paralysis ofthe lower · extremities and bladder, and finally death from respiratory failure. HISTOLOGY
On histological examination of tongue ulcers in monkeys, the most significant finding is the presence oftype A intranuclear inclusion bodies (Figs. 3 and 4)
BRITISH VETERINARY JOURNAL,
122, 2
sited in the most recent areas of degeneration, that is, at the junction of the necrotic plaque with normal tissue. These inclusions are most easily seen in lesions of three to 'four days standing. They vary in character from an eosinophilic mass of fine granular material filling the nucleus, to a typical type A inclusion of an intensely staining mass, shrunken to leave a clear surrounding halo. These inclusion-filled nuclei tend to aggregate into clumps of several nuclei which are quite distinctive in appearance. Histological evidence of damage has also been reported in the liver and kidneys. VIRUS ISOLATION IN THE LABORATORY
In the laboratory the virus may be isolated in tissue culture or in experimental animals. In tissue culture various cell systems have been used, for example monkey kidney, rabbit kidney, HeLa and human amnion cells. The virus also grows readily on the chorioallantoic membrane of the embryonated hen's egg, producing pocks similar in appearance to those of herpes simplex. The characteristic cytopathogenic effect described by Wood & Shimada (1954) in monkey kidney cells is of scattered foci of cell necrosis which rapidly coalesce to complete cell involvement. Individual cells enlarge and become globular and giant-cell formation is soon evident. These giant cells may contain two to 20 nuclei and often each nucleus is the site of a type A inclusion. The most susceptible small laboratory animal is the rabbit, although young suckling mice and guinea pigs are also susceptible. In the rabbit, inoculation by any route usually has a fatal outcome, in the form of fatal encephalitis with local skin lesions at the initial inoculation site. This disease pattern is similar to the picture in man. Inoculation of the rabbit cornea produces keratitis and involvement of the central nervous system. Only suckling mice up to three weeks old are susceptible to inoculation with the virus. Guinea pigs are slightly more resistant and usually survive infection. They do however become temporarily paralysed and produce antibody to the virus. DIAGNOSIS
In the monkey, diagnosis is made by the clinical evidence of herpetic ulcers on the tongue, with laboratory confirmation, if this is considered necessary, by virus isolation, histological examination, or the demonstration of "B" virus antibodies in the serum of cases which recover. In man the diagnosis can only be made accurately in the laboratory. However, any encephalitis or encephalomyelitis developing in a person exposed to monkeys or monkey tissues should be suspected. The laboratory diagnosis is based on the following criteria: (i) Isolation ofthe virus from the central nervous system. (ii) The finding of typical inclusions at histopathological examination. (This is difficult except in cases where there has been a fulminating encephalitis and the patient has died within a few days.) The inclusions morphologically resemble those of herpes simplex.
"B" VIRUS DISEASE IN MONKEY AND MAN
49
(iii) Serological examination by the serum neutralization test to demonstrate neutralizing antibodies to "B" virus in serum of convalescent cases. This antibody is present after ten days but it does not reach a high titre. There is a close antigenic relationship between "B" virus and herpes simplex virus. "B" virus antiserum neutralizes herpes simplex as well as or even better than it neutralizes "B" virus, but herpes simplex antiserum will not neutralize "B" virus, or will only do so to a low titre. TREATMENT
In man treatment involves supportive measures only, although there is some evidence to suggest that steroids may be of value. Breen, Lamb & Otaki ( 1958) used 200 mg. of cortisone daily on one of the reported survivors of the disease, and a definite improvement was noted in the patient's condition coincident with the instigation of this line of treatment. PROPHYLAXIS
Basically this involves limiting the possibilityofthediseaseaccidentallyinfecting man by following a rigid safety routine when housing or using monkeys, particularly monkeys recently imported. To this end the following measures should be considered. All newly imported monkeys should be kept in strict quarantine, caged preferably singly, or at least in pairs, for six to eight weeks, so that the dissemination of the virus within the group is limited and it may safely be assumed that the healthy monkeys are free from infection. Personnel handling monkeys should wear protective clothing, to include as complete a protective covering as possible, e.g. boots, gowns, masks and gloves. No animals should be caught by hand if at all possible, and use should be made of mechanical catching devices, such as nets or boxes, or anaesthetic and tranquillizing agents should be administered. Care should be taken that all cages and equipment are of such a standard that the possibility of a worker receiving a cut or abrasion is minimized. Any superficial fresh cuts or scratches received outside the animal house must be adequately covered before a worker enters it. If a bite or scratch is received in the monkey building prompt treatment of the wound must be carried out. This should involve washing the site vigorously with copious supplies of soap and water before a suitable topical antiseptic is applied. The person should then report immediately to the appropriate medical authority. Gamma-globulin (human) has been given parenterally as an adjunct to this treatment, although its use must be of doubtful value. Some experimental vaccines against "B" virus have been prepared which use killed cultures of the virus, but these have not as yet reached a satisfactory stage of development. Evidence of a "herpes-like" stomatitis seen in monkeys should be viewed very seriously and it is suggested that such animals should be killed forthwith, their carcases incinerated and their cages sterilized. CONCLUSION
Despite the serious consequences of "B" virus disease in man, it is evident that
so
BRITISH VETERINARY JOURNAL, 122, 2
the susceptibility of man to a clinical infection is low in view of the vast number of monkeys kept and handled in laboratories and animal houses throughout the world and the small number of clinical cases recorded. REFERENCES
BREEN, G. E., LAMB, S. G. & OTAKI, A. T. (1958). Br. med. ]. , :~, 22. BuRNET, F . M., LuSH, D. & jAcKsoN, A. V. (1939). Aust. J. exp. Biol. med. Sci., 17, 41. DAvmsoN, W. L. & HuMMELER, K. (r96o). Ann. N.r. Acad. Sci., 85, 970. HARTLEY, E. G. (1964). Vet Rec., 76, 555· KEEBLE, S. A. (r96o). Ann. N.r. Acad. Sci., 85, 96o. KEEBLE, S. A., CHRISTOFINis, G. J. & WooD, W. (1958). ]. Path. Bact., 76, 189. SABIN, A. B. (1949). J. clin. Invest., :~8, 8o8. SABIN, A. B. & WRIGHT, A. M. (1934a). Br. J. exp. Path., 15, 248, 321. SABIN, A. B. & Wright, A.M. (1934b). J. exp. Med., 59, 115. WooD, w. & SHIMADA, F. T. (1954)· Can. J . publ. Hlth, 45· 509. (Acceptedfor publication 11 November, 1965)
122,
46
"B" VIRUS DISEASE IN MONKEY AND MAN BY
E. G.
HARTLEY
National Institute for Medical Research, Mill Hill, London
=============
SUMMARY
"B" virus causes a benign disease of monkeys characterized by a herpes-like stomatitis which heals spontaneously in seven to I4 days. However, when the virus is established in man by accidental monkey-bite or from infected tissues or fluids, it may produce a fatal encephalitis or encephalomyelitis. A clinical description of the disease is given and methods of diagnosis in monkey and man are described. Methods to reduce the risk ofinfection to those closely involved in the handling of monkeys or their tissues are suggested. HISTORY
Sabin & Wright ( I934a) first isolated a virus from the central nervous system of a laboratory worker who had died after a bite from an apparently normal rhesus monkey. The virus was called by Sabin "B" virus. No further case was reported until I 949 when a case from similar circumstances was reported by Sabin (I949). From the mid-fifties onwards, however, with the increasing use of monkeys for the production and testing of biological products, principally the poliomyelitis vaccines, and as a source of tissue culture material for virus studies, the significance of the disease increased considerably. To date more than fifteen cases have been reported in man, all fatal except two. All have occurred in persons closely associated with either monkeys or their tissues. Keeble, Christofinis & Wood (I958) first noticed herpes-like ulcers on the tongue and lips of rhesus monkeys they were examining. A virus was isolated from these lesions which they showed to be "B" virus. INCIDENCE AND AETIOLOGY
The incidence of the disease in the commoner laboratory monkeys, namely the rhesus (Macaca mulatta) and the cynamolgus (Macaca ira) is fairly high. Keeble (I 96o) gives a figure of 2 ·3 per cent clinical cases seen in I 4,400 rhesus examined by him. Hartley (I 964) gives an incidence of 3 per cent with lesions in I ,ooo cynamolgus monkeys examined. Despite the lack of evidence there is no reason to doubt that the disease can occur also in many other species of monkey. A seasonal increase in the natural disease occurs with the monsoon and post-monsoon period, i.e. from October to February, when the majority of clinical cases are encountered (Keeble, Ig6o; Hartley, I964). It is evident also from neutralizing antibodies detected in apparently normal monkeys that the incidence is in fact very widespread. Some workers have found up to I oo
"B" VIRUS DISEASE IN MONKEY AND MAN
47
per cent in batches of imported monkeys. The natural infection in the monkey seems to follow a similar pattern to the primary stomatitis seen in man. It has been suggested by Burnet, Lush & Jackson (1939) that herpes simiae ("B" virus) and herpes simplex were at one time the same virus before adaptation to their own specific hosts. There is, however, no evidence to suggest at present that, like herpes in man, "B" virus persists in monkeys in a latent form after the initial lesions have healed. The virus in monkeys appears to spread within the colony by the contamination of food and water from infective saliva, and by direct inoculation of the skin by bites or scratches. In man every reported case except one has been accompanied by a history of either bites, scratches, or other close contact with monkeys. The exception was a case reported by Davidson & Hummeler (1960) in which the fatality occurred in a laboratory worker handling tissue cultures of monkey renal cells. Their evidence also suggests that the virus may be spread by gross aerosols to personnel working in the monkey rooms. CLINICAL FEATURES
In the monkey the gross lesions are most readily recognized on the surface of the tongue (Fig. 1). These lesions take the form of small vesicles which soon rupture to form an ulcer (Fig. 2) covered by a necrotic scab. This scab is yellowish-grey in colour and is sharply demarcated from the surrounding tissues. Within seven to 14 days these ulcers heal spontaneously, leaving no scar. During the period of stomatitis the animal does not appear to suffer any discomfort and continues to eat satisfactorily. Although the majority of these lesions appear on the tongue either singly or in groups, ulcers may also be seen on the mucoepithelial border of the lips. Lesions have also been reported on various parts ofthe skin, due no doubt to bites received during fighting. Secondary infections of a bacterial or fungal nature can also occur. As with herpes simplex, occasional involvement of the central nervous system takes place, and lesions have been seen during histological examinations of monkey brain and cord. Lesions seen are mild in character, and do not appear to give rise to clinically evident signs. In man the clinical symptoms have varied considerably but the general picture is that of an encephalitis or encephalomyelitis. The local area of the bite may show pain, redness and swelling, with the formation of vesicles and exudate. Often, associated lymphangitis and lymphadenitis occur. In all cases the fatal termination was within three weeks of illness and was in one case as early as three days. In both the cases which recovered there was severe residual damage to the central nervous system. The original case described by Sabin & Wright (1934b) was of an acute ascending myelitis with paralysis ofthe lower · extremities and bladder, and finally death from respiratory failure. HISTOLOGY
On histological examination of tongue ulcers in monkeys, the most significant finding is the presence oftype A intranuclear inclusion bodies (Figs. 3 and 4)
BRITISH VETERINARY JOURNAL,
122, 2
sited in the most recent areas of degeneration, that is, at the junction of the necrotic plaque with normal tissue. These inclusions are most easily seen in lesions of three to 'four days standing. They vary in character from an eosinophilic mass of fine granular material filling the nucleus, to a typical type A inclusion of an intensely staining mass, shrunken to leave a clear surrounding halo. These inclusion-filled nuclei tend to aggregate into clumps of several nuclei which are quite distinctive in appearance. Histological evidence of damage has also been reported in the liver and kidneys. VIRUS ISOLATION IN THE LABORATORY
In the laboratory the virus may be isolated in tissue culture or in experimental animals. In tissue culture various cell systems have been used, for example monkey kidney, rabbit kidney, HeLa and human amnion cells. The virus also grows readily on the chorioallantoic membrane of the embryonated hen's egg, producing pocks similar in appearance to those of herpes simplex. The characteristic cytopathogenic effect described by Wood & Shimada (1954) in monkey kidney cells is of scattered foci of cell necrosis which rapidly coalesce to complete cell involvement. Individual cells enlarge and become globular and giant-cell formation is soon evident. These giant cells may contain two to 20 nuclei and often each nucleus is the site of a type A inclusion. The most susceptible small laboratory animal is the rabbit, although young suckling mice and guinea pigs are also susceptible. In the rabbit, inoculation by any route usually has a fatal outcome, in the form of fatal encephalitis with local skin lesions at the initial inoculation site. This disease pattern is similar to the picture in man. Inoculation of the rabbit cornea produces keratitis and involvement of the central nervous system. Only suckling mice up to three weeks old are susceptible to inoculation with the virus. Guinea pigs are slightly more resistant and usually survive infection. They do however become temporarily paralysed and produce antibody to the virus. DIAGNOSIS
In the monkey, diagnosis is made by the clinical evidence of herpetic ulcers on the tongue, with laboratory confirmation, if this is considered necessary, by virus isolation, histological examination, or the demonstration of "B" virus antibodies in the serum of cases which recover. In man the diagnosis can only be made accurately in the laboratory. However, any encephalitis or encephalomyelitis developing in a person exposed to monkeys or monkey tissues should be suspected. The laboratory diagnosis is based on the following criteria: (i) Isolation ofthe virus from the central nervous system. (ii) The finding of typical inclusions at histopathological examination. (This is difficult except in cases where there has been a fulminating encephalitis and the patient has died within a few days.) The inclusions morphologically resemble those of herpes simplex.
"B" VIRUS DISEASE IN MONKEY AND MAN
49
(iii) Serological examination by the serum neutralization test to demonstrate neutralizing antibodies to "B" virus in serum of convalescent cases. This antibody is present after ten days but it does not reach a high titre. There is a close antigenic relationship between "B" virus and herpes simplex virus. "B" virus antiserum neutralizes herpes simplex as well as or even better than it neutralizes "B" virus, but herpes simplex antiserum will not neutralize "B" virus, or will only do so to a low titre. TREATMENT
In man treatment involves supportive measures only, although there is some evidence to suggest that steroids may be of value. Breen, Lamb & Otaki ( 1958) used 200 mg. of cortisone daily on one of the reported survivors of the disease, and a definite improvement was noted in the patient's condition coincident with the instigation of this line of treatment. PROPHYLAXIS
Basically this involves limiting the possibilityofthediseaseaccidentallyinfecting man by following a rigid safety routine when housing or using monkeys, particularly monkeys recently imported. To this end the following measures should be considered. All newly imported monkeys should be kept in strict quarantine, caged preferably singly, or at least in pairs, for six to eight weeks, so that the dissemination of the virus within the group is limited and it may safely be assumed that the healthy monkeys are free from infection. Personnel handling monkeys should wear protective clothing, to include as complete a protective covering as possible, e.g. boots, gowns, masks and gloves. No animals should be caught by hand if at all possible, and use should be made of mechanical catching devices, such as nets or boxes, or anaesthetic and tranquillizing agents should be administered. Care should be taken that all cages and equipment are of such a standard that the possibility of a worker receiving a cut or abrasion is minimized. Any superficial fresh cuts or scratches received outside the animal house must be adequately covered before a worker enters it. If a bite or scratch is received in the monkey building prompt treatment of the wound must be carried out. This should involve washing the site vigorously with copious supplies of soap and water before a suitable topical antiseptic is applied. The person should then report immediately to the appropriate medical authority. Gamma-globulin (human) has been given parenterally as an adjunct to this treatment, although its use must be of doubtful value. Some experimental vaccines against "B" virus have been prepared which use killed cultures of the virus, but these have not as yet reached a satisfactory stage of development. Evidence of a "herpes-like" stomatitis seen in monkeys should be viewed very seriously and it is suggested that such animals should be killed forthwith, their carcases incinerated and their cages sterilized. CONCLUSION
Despite the serious consequences of "B" virus disease in man, it is evident that
so
BRITISH VETERINARY JOURNAL, 122, 2
the susceptibility of man to a clinical infection is low in view of the vast number of monkeys kept and handled in laboratories and animal houses throughout the world and the small number of clinical cases recorded. REFERENCES
BREEN, G. E., LAMB, S. G. & OTAKI, A. T. (1958). Br. med. ]. , :~, 22. BuRNET, F . M., LuSH, D. & jAcKsoN, A. V. (1939). Aust. J. exp. Biol. med. Sci., 17, 41. DAvmsoN, W. L. & HuMMELER, K. (r96o). Ann. N.r. Acad. Sci., 85, 970. HARTLEY, E. G. (1964). Vet Rec., 76, 555· KEEBLE, S. A. (r96o). Ann. N.r. Acad. Sci., 85, 96o. KEEBLE, S. A., CHRISTOFINis, G. J. & WooD, W. (1958). ]. Path. Bact., 76, 189. SABIN, A. B. (1949). J. clin. Invest., :~8, 8o8. SABIN, A. B. & WRIGHT, A. M. (1934a). Br. J. exp. Path., 15, 248, 321. SABIN, A. B. & Wright, A.M. (1934b). J. exp. Med., 59, 115. WooD, w. & SHIMADA, F. T. (1954)· Can. J . publ. Hlth, 45· 509. (Acceptedfor publication 11 November, 1965)