1310 jections and the scrapie inoculation were separated by a week (regimen A) or a day (regimen B). All mice were infected simultaneously with 0.1ml of a 10% saline homogenate of pooled brains taken from clini-
cally affected hamsters at the 4th intracerebral passage of Chandler scrapie. All homogenates were prepared in advance and stored at - 20°C; samples were thawed and rehomogenised before injection. The incubation-time of scrapie was determined by "blind" scoring of mice at weekly intervals for up to 440 days after infection. The four groups of mice that were pretreated with either brain or hamster brain had longer mean incubation periods and higher proportions of survivors than the two control groups that were pretreated with saline (table). However, mouse
pretreatment with mouse brain consistently produced effects less pronounced than with hamster brain. It is significant that these effects were produced by injecting homogenates of normal hamster brain under conditions identical to those used for infection with scrapie-i.e., pretreatment with normal brain suspension in Freund’s adjuvant were deliberately avoided. Consequently these findings suggest that part of the speciesbarrier effect observed on intraperitoneal transmission of hamster scrapie to mice may arise from the necessity of using crude scrapie inocula, the presence of a large amount of antigenically foreign material possibly stimulating a host response which increases the effective removal of infectivity. What is not clear is whether the effects observed are due to a systemic response to hamster antigens or to a local response in the peritoneum since both pretreatment and infection were carried out by the i.p. route. In a wider context these results suggest that immunosuppression of the host might be used to diminish species-barrier effects. However, such an approach may be self-defeating because some components of the lymphoid system may play an important role (other than as a defence mechanism) in scrapie7 pathogenesis, at least in mice infected by a peripheral route.6 In a collaborative study with Dr P. G. Cunnington (Beecham Pharmaceuticals Research Division, Betchworth, Surrey) we have obtained preliminary data suggesting that the i.p. injection of mice with suspensions of methanol extraction residue ofB.c.G. just before i.p. infection with low doses of mousepassaged scrapie can considerably shorten incubation time. We have yet to define the optimum conditions for this effect or to apply it to the interspecific transmission of scrapie by the i.p. route. However, these observations suggest that the use of drugs to modify agent/cell interactions in vivo may be useful in promoting experimental transmission of scrapie and related human diseases to other host species. R. H. KIMBERLIN A.R.C. Institute for Research on Animal Diseases, Compton, Newbuty, Berkshire RG16 0NN.
C. A. WALKER G. C. MILLSON
CHOLESTEROL CONTENT OF TUBE FEEDS
SiR,—We write in defence of the dietetic profession in reply Dr Craig’s letter (Nov. 8, p. 935) on this subject. Dr Craig states that her patient had previously been having a low-cholesterol diet; it therefore seems reasonable that the doctor request a "low-cholesterol" tube feed, which no doubt would have been supplied. To prevent mishaps like this, it is imperative that the dietetic department receive a written request from the doctor stating the diagnosis and type of diet required. As a profession supplementary to medicine we aim only to please
to
and to implement the doctors’ requests to the best of our ability-if only we are asked. Northwick Park Hospital, Watford Road,
Harrow, Middlesex HA1 3UJ.
P. BRERETON V. HAINSWORTH
5. Kimberlin, R. H., Collis, S. C., Walker, C. A. J. comp. Path. (in the 6. Outram, G. W., Dickinson, A. G., Fraser, H. Nature, 1974, 249, 855. 7. Outram, G. W., Dickinson, A. G., Fraser, H. Lancet, 1975, i, 198.
press).
MONONUCLEOSIS-ASSOCIATED SUBACUTE SCLEROSING PANENCEPHALITIS
SIR,-The occurrence of subacute sclerosing panencephalitis (s.s.P.E.) following infectious mononucleosis (i.M.) described by Dr Feorino and his colleagues (Sept. 20, p.530) adds to the growing list of complications associated with i.M.’ Fatal t.M. has a predilection for boys; fatalities have been described in boys in three families.2-4 Emerging information on the immunopathogenesis of !.M. suggests that an immunodeficiency to the Epstein-Barr virus (E.B.v.) occurs in patients who succumb to i.M. Likewise, abnormal immune responses to measles virus (rubeola) might lead to the development of S.S.P.E. many years after measles infection. Intranuclear "paramyxovirus-like" or "measles-like" inclusions were seen in the lymphoid cells and in the pneumocytes of two brothers who had Duncan’s disease (see figure). How-
Electron
micrograph of lung taken
at
necropsy.
Nucleus of pneumocyte on the bottom is filled with a dense array of filaments (x 10 540). Inset displays finer d’etail of particles (x 43 000). The filaments ranged from 228 to 297 A in length and from 22.8 to 29.7 nm in diameter (average 250 nm).
the interpretation of these intranuclear inclusions requires cautious judgement,5-’ because they differ from the viral particles observed when measles virus infects cells grown ever,
in tissue-culture8 and from the inclusions seen in s.s.r.E.9-" One of the Duncan boys had developed only a "light rash" when infected by measles 3 years before his death. His brother, who died 4 years later, had measles a week before the onset of fatal i.M. Measles virus infects T lymphocytes,l2 and thus s.s.p.E. and Duncan’s disease may be associated with a defect in T-cell function that enables measles virus to persist. In Duncan’s dis-
Penman, H. G. J. clin. Path. 1970, 23, 765. Bar, R. S., DeLor, C. J., Clausen, K. P., Hurtubise, P., Henle, W. Hewetson, J. F. New Engl. J. Med. 1974, 290, 363. 3. Purtilo, D. T., Cassel, C. K., Yang, J. P. S., Harper, R., Stephenson, S. K., Landing, B. H., Vawter, G. F. Lancet, 1975, i, 935. 4. Provisor, A. J., Iacuone, J. J., Chilcote, R. R., Neibuger, R. G., Crusse, F. G., Baehner, R. L. New Engl. J. Med. 1975, 293, 62. 5. Lampert, F., Lampert, P. Archs Neurol. 1975, 32, 425. 6. Shaw, C. M., Sumi, S. M. ibid. p.428. 7. Raine, C. S., Schaumburg, H. H., Snyder, D. H., Suzuki, K., J. neurol. Sci. 1975, 25, 29. 8. Raine, C. S., Feldman, L. A., Sheppard, R. D., Bornstein, M. B., J. Virol. 1969, 4, 169. 9 Oyangi, S., ter Meulen, V., Katz, M., Koprowski, H. ibid. 1971, 7, 176. 10. Dubois-Dalq, M., Barbosa, L. H., Hamilton, R., Sever, J. L. Lab. Invest. 1974, 30, 241. 11. Dubois-Dalaq, M., Worthington, K., Gutenson, O., Barbosa, L. H., ibid. 1975, 32, 518. 12. Valdimarsson, H., Agnarsdottir, G., Lachmann, P. J. Nature, 1975, 255, 1. 2.
554.
1311
and possibly in S.S.P.E., a progressive attrition of T-cell function may ensue. Measles infections are known to be associated with anergy13and thymic atrophy. 14 E.B.v. infects B lymphocytes and trigger their proliferation."In Duncan’s disease, T-cell function for controlling the proliferation of B cells was probably impaired by concurrent measles infection of T-cells. The association between !.M., which is caused by an E.B.v. infection of B cells, and S.S.P.E., which is probably caused by measles virus, appears more than fortuitous. Simultaneous E.B.v. and measles-virus infections could potentiate one another by impairing normal immune responses.
ease,
such complexes might then result in immune complex deposition in the vessel wall in view of the interchange between serum and cell-membrane cholesterol.
to
Central Microbiological Laboratories, Western General Hospital,
K. C. WATSON
Crewe Road, Edinburgh EH4 2XU
E.
Department of Pathology,
University of Massachusetts Medical Center, 55 Lake Avenue North,
Worcester, Massachusetts 01605, and Department of Pathology,
Joseph’s Hospital, Providence, Rhode Island, U.S.A.
St.
DAVID T. PURTILO JAMES P. S. YANG UMBERTO DEGIROLAM SALVATORE ALLEGRA
CHOLESTEROL-BINDING CAPACITY AND ATHEROGENESIS
SIR,-We
were
J. C. KERR
VIRUS OF INFANTILE GASTROENTERITIS IN ARGENTINA
SIR,-We have followed with interest the reports on reovirus-like particles associated to acute infantile gastroenteritis in several countries and have lately started looking for them in Argentina. Children of less than five years of age with acute sporadic gastroenteritis were selected for this preliminary study. Faecal suspensions of 20 children were treated with techniques described by Bishop et aU and examined under an ’Elmiskop I’ electron microscope. In 3 cases (15%) we found particles of similar structure and size to those reported by Bishop, Flewett and their co-workers.1-3 We have not yet investigated other bacterial or viral aetiologies.
interested in the
findings of Dr Hsia and his 1000) regarding the serum reserve
colleagues (Nov. 22, p. capacity for binding added cholesterol and its possible role in atherogenesis. Using a more sensitive method for cholesterol assay-namely, the ability of free cholesterol to neutralise the haemolytic property of streptolysin 0-we found16 17 that different normal sera vary in their ability to bind added cholesterol before it becomes detectable by our method used by Hsia et al. The streptolysin method will detect less than 1 (ig/ml of cholesterol. We have also shown that antistreptolysin activity is generated in normal serum incubated with strains of Staphylococcus aureus or Pseudomonas ceruginosa. This is due to cholesterol esterase and possibly proteolytic enzymes released by the organisms, which alters the spatial orientation of cholesterol in lipoprotein fragments so that both the hydroxyl group at C3 and the side chain at C17 are available for binding. These conditions are necessary for neutralisation of streptolysin 0. When serum filtrates containing high levels of antistreptolysin activity generated in this way are added to normal serum and titrated, the activity drops in 2-6h to levels considerably lower than that expected from the dilution factor alone, suggesting that the cholesterol moiety of the lipoprotein fragments has undergone rearrangement with other lipoproteins. The findings of Dr Hsia’s group suggest that these liproproteins involve the very low density (V.L.D.L.) and high density (H.D.L.) lipoproteins subfraction components which they describe. Since low levels of antistreptolysin activity attributable to altered beta-lipoproteins appear to be found in all normal sera we believe that there is a continuing dynamic equilibrium between the catabolic production of this fraction and the V.L.D.L. and H.D.L. subfractions described. The ability of the serum antistreptolysin factor to bind to certain bacterial proteins suggests that it plays a homoeostatic role as a non-specific defence mechanism.’* If so, the V.L.D.L. and H.D.L. subfractions would act as regulators of this process. Reduction of the capacity Of H.D.L. and V.L.D.L. subfractions to bind cholesterol in arterial walls may be involved in atherogenesis as suggested by Dr Hsia and colleagues. However, we believe that the process is likely to be much more complex, possibly involving the ability of the protein moiety of altered lipoprotein exhibiting antistreptolysin activity to function as an autoantigen especially if already bound to some exogenous foreign protein molecule such as bacterial toxin. Autoantibody Sullivan, J. L., Barry, D. W., Albrecht, P., Lucas, S. J. J. Immun. 1975, 114, 1458. 14. White, R. G., Boyd, J. F. Clin. exp. Immun. 1973, 13, 343. 15. Papamichail, M., Sheldon, P. J., Holoborow, E. J. ibid. 1974, 18, 1. 16. Watson, K. C., Kerr, E. J. C. Nature, 1974, 250, 230. 17. Watson, K. C., Kerr, E. J. C. J. med. Microbiol. (in the press). 18. Watson, K. C., Kerr, E. J. C. Lancet, 1975, i, 308.
Virus particles
( x 201500).
We think it of interest that the agent that is beginning to be called "virus of infantile gastroenteritis"4 is also present in the southern tip of the American continent. Virus Department, National Institute of Microbiology, Av. Velez Sarsfield 563, Buenos Aires, Argentina.
Alejandro Posadas Polyclinic, Ramos Mejía.
GABRIEL H. LOMBARDI ALBERTO M. ROSETO DANIEL STAMBOULIAN
Virus Department, National Institute of Buenos Aires.
JULIO G. BARRERA ORO
Microbiology,
13.
1.
Bishop, R. F., Davidson, G. P., Holmes, I. H., Ruck,
B.
J. Lancet, 1974, i,
149. 2.
3.
Flewett, T. H., Bryden, A. S., Davies, H. ibid, 1973, ii, 1497. Bishop, R. F., Davidson, G. P., Holmes, I. H., Ruck, B. J. ibid. 1973, ii, 1281.
4. Br.
med. J. 1975, iii, 555.
cally affected hamsters at the 4th intracerebral passage of Chandler scrapie. All homogenates were prepared in advance and stored at - 20°C; samples were thawed and rehomogenised before injection. The incubation-time of scrapie was determined by "blind" scoring of mice at weekly intervals for up to 440 days after infection. The four groups of mice that were pretreated with either brain or hamster brain had longer mean incubation periods and higher proportions of survivors than the two control groups that were pretreated with saline (table). However, mouse
pretreatment with mouse brain consistently produced effects less pronounced than with hamster brain. It is significant that these effects were produced by injecting homogenates of normal hamster brain under conditions identical to those used for infection with scrapie-i.e., pretreatment with normal brain suspension in Freund’s adjuvant were deliberately avoided. Consequently these findings suggest that part of the speciesbarrier effect observed on intraperitoneal transmission of hamster scrapie to mice may arise from the necessity of using crude scrapie inocula, the presence of a large amount of antigenically foreign material possibly stimulating a host response which increases the effective removal of infectivity. What is not clear is whether the effects observed are due to a systemic response to hamster antigens or to a local response in the peritoneum since both pretreatment and infection were carried out by the i.p. route. In a wider context these results suggest that immunosuppression of the host might be used to diminish species-barrier effects. However, such an approach may be self-defeating because some components of the lymphoid system may play an important role (other than as a defence mechanism) in scrapie7 pathogenesis, at least in mice infected by a peripheral route.6 In a collaborative study with Dr P. G. Cunnington (Beecham Pharmaceuticals Research Division, Betchworth, Surrey) we have obtained preliminary data suggesting that the i.p. injection of mice with suspensions of methanol extraction residue ofB.c.G. just before i.p. infection with low doses of mousepassaged scrapie can considerably shorten incubation time. We have yet to define the optimum conditions for this effect or to apply it to the interspecific transmission of scrapie by the i.p. route. However, these observations suggest that the use of drugs to modify agent/cell interactions in vivo may be useful in promoting experimental transmission of scrapie and related human diseases to other host species. R. H. KIMBERLIN A.R.C. Institute for Research on Animal Diseases, Compton, Newbuty, Berkshire RG16 0NN.
C. A. WALKER G. C. MILLSON
CHOLESTEROL CONTENT OF TUBE FEEDS
SiR,—We write in defence of the dietetic profession in reply Dr Craig’s letter (Nov. 8, p. 935) on this subject. Dr Craig states that her patient had previously been having a low-cholesterol diet; it therefore seems reasonable that the doctor request a "low-cholesterol" tube feed, which no doubt would have been supplied. To prevent mishaps like this, it is imperative that the dietetic department receive a written request from the doctor stating the diagnosis and type of diet required. As a profession supplementary to medicine we aim only to please
to
and to implement the doctors’ requests to the best of our ability-if only we are asked. Northwick Park Hospital, Watford Road,
Harrow, Middlesex HA1 3UJ.
P. BRERETON V. HAINSWORTH
5. Kimberlin, R. H., Collis, S. C., Walker, C. A. J. comp. Path. (in the 6. Outram, G. W., Dickinson, A. G., Fraser, H. Nature, 1974, 249, 855. 7. Outram, G. W., Dickinson, A. G., Fraser, H. Lancet, 1975, i, 198.
press).
MONONUCLEOSIS-ASSOCIATED SUBACUTE SCLEROSING PANENCEPHALITIS
SIR,-The occurrence of subacute sclerosing panencephalitis (s.s.P.E.) following infectious mononucleosis (i.M.) described by Dr Feorino and his colleagues (Sept. 20, p.530) adds to the growing list of complications associated with i.M.’ Fatal t.M. has a predilection for boys; fatalities have been described in boys in three families.2-4 Emerging information on the immunopathogenesis of !.M. suggests that an immunodeficiency to the Epstein-Barr virus (E.B.v.) occurs in patients who succumb to i.M. Likewise, abnormal immune responses to measles virus (rubeola) might lead to the development of S.S.P.E. many years after measles infection. Intranuclear "paramyxovirus-like" or "measles-like" inclusions were seen in the lymphoid cells and in the pneumocytes of two brothers who had Duncan’s disease (see figure). How-
Electron
micrograph of lung taken
at
necropsy.
Nucleus of pneumocyte on the bottom is filled with a dense array of filaments (x 10 540). Inset displays finer d’etail of particles (x 43 000). The filaments ranged from 228 to 297 A in length and from 22.8 to 29.7 nm in diameter (average 250 nm).
the interpretation of these intranuclear inclusions requires cautious judgement,5-’ because they differ from the viral particles observed when measles virus infects cells grown ever,
in tissue-culture8 and from the inclusions seen in s.s.r.E.9-" One of the Duncan boys had developed only a "light rash" when infected by measles 3 years before his death. His brother, who died 4 years later, had measles a week before the onset of fatal i.M. Measles virus infects T lymphocytes,l2 and thus s.s.p.E. and Duncan’s disease may be associated with a defect in T-cell function that enables measles virus to persist. In Duncan’s dis-
Penman, H. G. J. clin. Path. 1970, 23, 765. Bar, R. S., DeLor, C. J., Clausen, K. P., Hurtubise, P., Henle, W. Hewetson, J. F. New Engl. J. Med. 1974, 290, 363. 3. Purtilo, D. T., Cassel, C. K., Yang, J. P. S., Harper, R., Stephenson, S. K., Landing, B. H., Vawter, G. F. Lancet, 1975, i, 935. 4. Provisor, A. J., Iacuone, J. J., Chilcote, R. R., Neibuger, R. G., Crusse, F. G., Baehner, R. L. New Engl. J. Med. 1975, 293, 62. 5. Lampert, F., Lampert, P. Archs Neurol. 1975, 32, 425. 6. Shaw, C. M., Sumi, S. M. ibid. p.428. 7. Raine, C. S., Schaumburg, H. H., Snyder, D. H., Suzuki, K., J. neurol. Sci. 1975, 25, 29. 8. Raine, C. S., Feldman, L. A., Sheppard, R. D., Bornstein, M. B., J. Virol. 1969, 4, 169. 9 Oyangi, S., ter Meulen, V., Katz, M., Koprowski, H. ibid. 1971, 7, 176. 10. Dubois-Dalq, M., Barbosa, L. H., Hamilton, R., Sever, J. L. Lab. Invest. 1974, 30, 241. 11. Dubois-Dalaq, M., Worthington, K., Gutenson, O., Barbosa, L. H., ibid. 1975, 32, 518. 12. Valdimarsson, H., Agnarsdottir, G., Lachmann, P. J. Nature, 1975, 255, 1. 2.
554.
1311
and possibly in S.S.P.E., a progressive attrition of T-cell function may ensue. Measles infections are known to be associated with anergy13and thymic atrophy. 14 E.B.v. infects B lymphocytes and trigger their proliferation."In Duncan’s disease, T-cell function for controlling the proliferation of B cells was probably impaired by concurrent measles infection of T-cells. The association between !.M., which is caused by an E.B.v. infection of B cells, and S.S.P.E., which is probably caused by measles virus, appears more than fortuitous. Simultaneous E.B.v. and measles-virus infections could potentiate one another by impairing normal immune responses.
ease,
such complexes might then result in immune complex deposition in the vessel wall in view of the interchange between serum and cell-membrane cholesterol.
to
Central Microbiological Laboratories, Western General Hospital,
K. C. WATSON
Crewe Road, Edinburgh EH4 2XU
E.
Department of Pathology,
University of Massachusetts Medical Center, 55 Lake Avenue North,
Worcester, Massachusetts 01605, and Department of Pathology,
Joseph’s Hospital, Providence, Rhode Island, U.S.A.
St.
DAVID T. PURTILO JAMES P. S. YANG UMBERTO DEGIROLAM SALVATORE ALLEGRA
CHOLESTEROL-BINDING CAPACITY AND ATHEROGENESIS
SIR,-We
were
J. C. KERR
VIRUS OF INFANTILE GASTROENTERITIS IN ARGENTINA
SIR,-We have followed with interest the reports on reovirus-like particles associated to acute infantile gastroenteritis in several countries and have lately started looking for them in Argentina. Children of less than five years of age with acute sporadic gastroenteritis were selected for this preliminary study. Faecal suspensions of 20 children were treated with techniques described by Bishop et aU and examined under an ’Elmiskop I’ electron microscope. In 3 cases (15%) we found particles of similar structure and size to those reported by Bishop, Flewett and their co-workers.1-3 We have not yet investigated other bacterial or viral aetiologies.
interested in the
findings of Dr Hsia and his 1000) regarding the serum reserve
colleagues (Nov. 22, p. capacity for binding added cholesterol and its possible role in atherogenesis. Using a more sensitive method for cholesterol assay-namely, the ability of free cholesterol to neutralise the haemolytic property of streptolysin 0-we found16 17 that different normal sera vary in their ability to bind added cholesterol before it becomes detectable by our method used by Hsia et al. The streptolysin method will detect less than 1 (ig/ml of cholesterol. We have also shown that antistreptolysin activity is generated in normal serum incubated with strains of Staphylococcus aureus or Pseudomonas ceruginosa. This is due to cholesterol esterase and possibly proteolytic enzymes released by the organisms, which alters the spatial orientation of cholesterol in lipoprotein fragments so that both the hydroxyl group at C3 and the side chain at C17 are available for binding. These conditions are necessary for neutralisation of streptolysin 0. When serum filtrates containing high levels of antistreptolysin activity generated in this way are added to normal serum and titrated, the activity drops in 2-6h to levels considerably lower than that expected from the dilution factor alone, suggesting that the cholesterol moiety of the lipoprotein fragments has undergone rearrangement with other lipoproteins. The findings of Dr Hsia’s group suggest that these liproproteins involve the very low density (V.L.D.L.) and high density (H.D.L.) lipoproteins subfraction components which they describe. Since low levels of antistreptolysin activity attributable to altered beta-lipoproteins appear to be found in all normal sera we believe that there is a continuing dynamic equilibrium between the catabolic production of this fraction and the V.L.D.L. and H.D.L. subfractions described. The ability of the serum antistreptolysin factor to bind to certain bacterial proteins suggests that it plays a homoeostatic role as a non-specific defence mechanism.’* If so, the V.L.D.L. and H.D.L. subfractions would act as regulators of this process. Reduction of the capacity Of H.D.L. and V.L.D.L. subfractions to bind cholesterol in arterial walls may be involved in atherogenesis as suggested by Dr Hsia and colleagues. However, we believe that the process is likely to be much more complex, possibly involving the ability of the protein moiety of altered lipoprotein exhibiting antistreptolysin activity to function as an autoantigen especially if already bound to some exogenous foreign protein molecule such as bacterial toxin. Autoantibody Sullivan, J. L., Barry, D. W., Albrecht, P., Lucas, S. J. J. Immun. 1975, 114, 1458. 14. White, R. G., Boyd, J. F. Clin. exp. Immun. 1973, 13, 343. 15. Papamichail, M., Sheldon, P. J., Holoborow, E. J. ibid. 1974, 18, 1. 16. Watson, K. C., Kerr, E. J. C. Nature, 1974, 250, 230. 17. Watson, K. C., Kerr, E. J. C. J. med. Microbiol. (in the press). 18. Watson, K. C., Kerr, E. J. C. Lancet, 1975, i, 308.
Virus particles
( x 201500).
We think it of interest that the agent that is beginning to be called "virus of infantile gastroenteritis"4 is also present in the southern tip of the American continent. Virus Department, National Institute of Microbiology, Av. Velez Sarsfield 563, Buenos Aires, Argentina.
Alejandro Posadas Polyclinic, Ramos Mejía.
GABRIEL H. LOMBARDI ALBERTO M. ROSETO DANIEL STAMBOULIAN
Virus Department, National Institute of Buenos Aires.
JULIO G. BARRERA ORO
Microbiology,
13.
1.
Bishop, R. F., Davidson, G. P., Holmes, I. H., Ruck,
B.
J. Lancet, 1974, i,
149. 2.
3.
Flewett, T. H., Bryden, A. S., Davies, H. ibid, 1973, ii, 1497. Bishop, R. F., Davidson, G. P., Holmes, I. H., Ruck, B. J. ibid. 1973, ii, 1281.
4. Br.
med. J. 1975, iii, 555.
