Lansing Virus Research Articles

Mapping of sequences required for mouse neurovirulence of poliovirus type 2 Lansing.

Intracerebral inoculation of mice with poliovirus type 2 Lansing induces a fatal paralysis, while most other poliovirus strains are unable to cause disease in the mouse. To determine the molecular basis for Lansing virus neurovirulence, we determined the complete nucleotide sequence of the Lansing viral genome from cloned cDNA. The deduced amino acid sequence was compared with that of two mouse-avirulent strains. There are 83 amino acid differences between the Lansing and Sabin type 2 strain and 179 differences between the Lansing and Mahoney type 1 strain scattered throughout the genome. To further localize Lansing sequences important for mouse neurovirulence, four intertypic recombinants were isolated by exchanging DNA restriction fragments between the Lansing 2 and Mahoney 1 infectious poliovirus cDNA clones. Plasmids were transfected into HeLa cells, and infectious recombinant viruses were recovered. All four recombinant viruses, which contained the Lansing capsid region and different amounts of the Mahoney genome, were neurovirulent for 18- to 21-day-old Swiss-Webster mice by the intracerebral route. The genome of neurovirulent recombinant PRV5.1 contained only nucleotides 631 to 3413 from Lansing, encoding primarily the viral capsid proteins. Therefore, the ability of Lansing virus to cause paralysis in mice is due to the viral capsid. The Lansing capsid sequence differs from that of the mouse avirulent Sabin 2 strain at 32 of 879 amino acid positions: 1 in VP4, 5 in VP2, 4 in VP3, and 22 in VP1.

A Comparative Study of Type 2 Poliomyelitis Viruses

Summary An antigenic analysis of 18 type 2 polio viruses has been made by means of cross-neutralization tests in tissue culture. Differing antigenic components entered into the constitution of the various strains making it possible to classify them into three major categories, namely, strains identical or closely related to the prototype Lansing virus, strains distinctly different from the Lansing virus, and intermediate strains. Differences in antigenic constitution of the type 2 polio viruses studied were observed in reciprocal CF tests using 1) the strains as viral antigens and studying their reaction with reference heterotypic sera, and 2) type 2 antisera and reference viral antigens of the three polio virus types and studying their reaction with antisera prepared against each of the type 2 strains under study. On the basis of these tests at least four of 15 type 2 polio viruses studied contained type 1 components in the composite antigenic structure.

A Comparative Study of Type 2 Poliomyelitis Viruses

Summary In this report, some additional evidence has been obtained for the existence of biologic variations among Type 2 strains of poliomyelitis viruses. The differences observed among such strains as Leyva and Theiler's Lansing are well defined and are clearly at variance in respect to the prototype Lansing virus. Variations encountered with those strains intermediate in position are also well defined. These strains are of interest because they have been passaged relatively infrequently and presumably possess native biologic characters. In contrast, strains passaged many times in rodents, or in monkeys, change their biologic properties and acquire additional characters, some of them peculiar to the recipient host. Hence, the designated characters assigned to the 16 strains are not viewed as stable; for it is clear that changes occur among strains of Type 2 poliomyelitis viruses studied under varying influences in the experimental laboratory. Experimental data obtained during the study provided opportunity for comparing monkey kidney epithelial cell cultures and mice and monkeys with regard to their sensitivities in delineating available units of poliomyelitis virus. In most instances, tissue cultures were the superior indicator systems.

Mixed infections with Coxsackie and Lansing poliomyelitis viruses in mice.

SummaryThe following reciprocal effects have been observed in mice infected with viruses which do not possess the same degree of pathogenicity for the host. 1. The Ohio-R strain of C virus influenced significantly both the course and outcome of experimental Lansing poliomyelitis infections in 4-week-old white Swiss mice. 2. In the presence of the Lansing infection the virulence of the Ohio-R virus seemed to be enhanced for these otherwise insusceptible animals. 3. The Nancy strain of C virus had a noticeable effect on the pathogenesis of the Lansing infection in 4-week-old mice. The frequency with which the forelimbs became paralyzed first as compared with the first appearance of hindlimb paralysis was influenced by the presence of Nancy virus in the higher levels of the CNS. The presence of Nancy virus in the lumbar region of the spinal cord, following intraspinal inoculation, seemed to limit extension of the Lansing virus thus altering the normal progress of this experimental infection. 4. There was no ...

Poliomyelitis in Canadian Eskimos laboratory studies. IV. Antigenic typing of virus strains in monkeys and in tissue cultures.

Three strains of poliomyelitis virus isolated from Eskimos infected in the outbreak of poliomyelitis occurring at Chesterfield Inlet, N.W.T., in February 1949 have been typed. This typing has been carried out in monkeys and in tissue cultures. Typing was carried out by preparing in rhesus monkeys antiserum to the Eskimo strains and testing these for capacity to neutralize a fixed dose of the Brunhilde (Type 1) and Leon (Type 3) strains on thalamic inoculation of monkeys, and of the Lansing (Type 2) strain on cerebral injection of mice. Antisera prepared against Strains E3-56 and E3-227 completely neutralized Brunhilde virus, as shown on thalamic inoculation of virus-serum mixtures in groups of five monkeys. In contrast, these sera did not neutralize Leon virus. Tests carried out in mice showed that antisera to these Eskimo strains failed to neutralize Lansing virus. It was concluded therefore that Eskimo Strains E3-56 and E3-227 belong to Type 1. The same two sera were tested for their capacity to inhibit the cytopathogenic effect induced in roller-tube cultures of monkey testis by representative strains of the three types (Brunhilde, Y-SK, and Saukett). The two Eskimo sera inhibited the cytopathogenic effect induced by the Brunhilde strain but not the changes induced by the Y-SK and Saukett strains, confirming that the corresponding Eskimo viruses belonged to Type 1. Eskimo Strains E3-79 and E3-227 have also been typed in tissue cultures by a direct method; in this method antisera to the three prototype viruses were tested for capacity to inhibit the cytopathogenic effect induced by the Eskimo strains. Results of this test were in agreement, the Eskimo strains belonging to Type 1. Two strains of virus isolated from the Chesterfield Inlet outbreak have previously been typed by U.S. investigators, by techniques involving the use of monkeys. A total of five strains isolated from this epidemic have therefore been typed, and all have been found to belong to Type 1. This study illustrates that the typing of poliomyelitis strains can be carried out more rapidly and more economically in tissue culture than in monkeys.

Passive immunity in poliomyelitis. VI. Antibody decline in Rhesus monkeys inoculated with homologous Lansing antiserum.

Summary1. Serum antibody titers have been determined in a passive protection experiment in which rhesus monkeys were inoculated intramuscularly with Lansing antiserum prepared in rhesus monkeys; the monkeys were challenged thalamically with 50 PD50 of Lansing virus. 2. Four of these monkeys developed paralysis, and 9 remained clinically and histologically normal. 3. Average serum antibody titers in samples removed one week after inoculation were 10−1.87 in the paralyzed group, and 10−1.88 in the unaffected group. 4. By extrapolation, the initial antibody titer was estimated to be 10−2.1. The average titer 6 weeks after inoculation was 10−0.29. 5. The rate of decline of antiboyd in the 9 monkeys the survived the cerebral challenge was constant, the half-life being 6.8 days.

CHEMOTHERAPEUTIC EFFECT OF N1-DODECANOYL-N4-ACETYL-4-AMINONAPHTHALENE-SULFONAMIDE-(1) ON NEUROTROPIC VIRUSES

(1) PANS-No. 610 shows a remarkable effect on the Japanese encephalitis virus in vitro and in vivo. It is effective even at a longer interval after the inoculation of the virus.(2) The drug shows a weak effect in vitro and in vivo on the Lansing virus.(3) The drug possesses a comparatively low toxicity.(4) It is assumed that the effect of the drug on the Japanese encephalitis virus might be caused by its neurotropy, cerebrotropy and penetration-power into cells in host, related to its surfactant properties.(5) The drug is of promise for the treatment of the diseases caused by the Japanese encephalitis virus.(6) It may be claimed that the drug is the only compound which has been confirmed to have chemotherapeutic effects on patients suffering from the so-called small virus such as the Japanese encephalitis virus.

Studies on the Cultivation of Poliomyelitis Viruses in Tissue Culture

Summary In 1949, in preliminary notes, we recorded the fact that the in vitro propagation of the poliomyelitis viruses may be accomplished in suspended cell cultures of various types of human tissues. The present paper details the procedures we have employed in the preparation and maintenance of suspended cell cultures for this purpose, and summarizes observations on the behavior of poliomyelitis viruses in such cultures, as observed during the past 4 years. Each of 3 antigenic types of poliomyelitis virus has been propagated in suspended cell cultures of human embryonic skin and muscle tissue, in cultures of human embryonic brain tissue and in cultures of mature human kidney tissue. In addition it has been demonstrated that one or more of these 3 types of virus will multiply in cultures of various other human embryonic tissues, including intestine, adrenal and spleen. Likewise, mature human tissues including thyroid, foreskin, and testis have been successfully employed for the cultivation of the Lansing strain as well as materials from two tumors, an embryoma of the kidney, and a neuroblastoma. Poliomyelitis virus has also been cultivated in monkey kidney cultures. Assay of the infectivity of materials removed from the tissue cultures was perfomred by the classical method of titration in animals and also by the technique of titration in vitro employing the cytopathogenic effect of the poliomyelitis viruses as an indicator of viral activity. Materials from cultures prepared with human embryonic skin-muscle tissues, kidney tissue, or brain tissues yielded the highest infectivity titers. Serial propagation in suspended cell cultures of human embryonic skin-muscle tissue was achieved for 23 passages with the Lansing strain, for a cumulate period of 331 days in cultivation, and for 15 passages with the Brunhilde strain, during a total period of 267 days. Throughout these prolonged periods of cultivation, virus production in each instance remained essentially constant as assayed by the in vitro technique. The titers as determined in vitro were in most cases equivalent to those observed in the central nervous system of the infected animal. However, in experiments with Lansing virus prolonged cultivation in human tissues was associated with a marked decline in apparent infectivity titer of the agent for the mouse. Likewise, in one experiment with Brunhilde virus, a significant decrease in the apparent infectivity titer for the monkey was observed.

POLIOMYELITIS IN CANADIAN ESKIMOS

Studies of the basic immunology of poliomyelitis in Canadian Eskimos have been continued. The sera of 24 Eskimos living at Lake Harbour and of 99 at Pangnirtung, Baffin Island, have been tested for poliomyelitis antibody by means of the Lansing virus neutralization test. Poliomyelitis occurred in Lake Harbour in 1943 but has not been recorded in Pangnirtung. Eighteen of the 24 sera from Lake Harbour contained antibody, but only 10 from the 99 Pangnirtung residents. The positive sera from Pangnirtung were all in persons aged 18 and over. These results are compared with those previously reported by the authors with the sera of Eskimos at Chesterfield Inlet, and with those reported by Paul with the sera of North Alaskan Eskimos. The evidence suggests that poliomyelitis virus is world-wide in distribution.

Routine titrations of Lansing virus and antibody in mice: comparison of methods of estimating endpoints.

Summary1. Over a period of several months, routine titrations in mice were carried out by a single worker of a pool of Lansing virus, and of samples of plasma gamma globulin, placental gamma globulin, and immune monkey serum. Estimates of the ED50's and their standard errors were calculated for the individual titrations by the probit method. In addition ED50's were calculated by the Karber, Reed and Muench, and moving average methods. 2. The standard deviation of the titrations was found to be greater than the error of individual assays, suggesting that titrations carried out on different occasions vary more than those performed at the same time. 3. None of the methods mentioned gave widely different estimates of the ED50. In particular, the ED50's calculated by the Karber method compared very favourably with those obtained by the probit method. 4. The Karber method is proposed as a suitable alternative to the Reed and Muench procedure for the calculation of ED50 in routine titrations of virus and antiserum.

Studies on passive immunity in poliomyelitis. V. Lansing antibody levels in humans after gamma globulin administration.

Summary1. The sera of 51 children and over 300 adults were first tested for the presence of Lansing virus neutralizing antibody. 2. Of those whose serum contained no demonstrable antibody, 6 children and 4 adults were given gamma globulin intramuscularly in doses from 0.1 to 0.4 ml per pound body weight. 3. Blood was taken at frequent intervals for 6 weeks and tested for antibody. A small quantity of antibody was present in the serum, as shown by a protective effect of the undiluted sample in mice. This antibody was detected in some cases for a few weeks. 4. No antibody was found in the serum of the same 10 persons following the oral administration of gamma globulin.

Passive immunity in poliomyelitis. IV. Protection of rhesus monkeys against cerebral challenge.

1. Lansing virus serum antibody titers in the range of 10-2.0 to 10-3.0 can be achieved in monkeys shortly after the administration of gamma globulin intramuscularly or intravenously. 2. Gamma globulin injected intravenously failed to protect a significant number of monkeys against an intramuscularly challenge of 160 PD50 of Lansing virus. 3. In 2 further experiments, the majority of monkeys receiving high titer immune monkey serum were protected against an intracerebral challenge of 50 and 100 PD50 of Lansing virus respectively. In one experiment only 4/16 test animals became paralyzed as compared to 18/19 controls. In the other experiment 3/18 test animals developed poliomyelitis as compared to all of 20 controls.

Complement-Fixation Studies with Poliomyelitis in Humans.

SummaryThe Lansing complement-fixing (CF) antibody response to immunization in monkeys and to infection in humans appears to be transitory. One hundred thirty-one paralytic poliomyelitis cases in humans were examined for CF antibody to Lansing virus. In the serums of 85 cases, collected 2 weeks after onset of clinical disease, 11 had both neutralizing and CF antibodies. CF antibody was not found in any of 47 cases which were negative for neutralizing antibody. In a series of serum specimens collected from 46 other cases, CF antibody appeared in 10. The CF antibody rose quickly and remained elevated for from 6 weeks to 3 months. A positive CF test may indicate recent infection; a negative, either no exposure or infection at a period remote in the past.

POLIOMYELITIS IN CANADIAN ESKIMOS

This paper describes further laboratory studies undertaken to investigate the immunological aspects of the outbreak of poliomyelitis in the Canadian Eastern Arctic in the winter of 1948-1949. No evidence was found that any of three strains of virus recovered from Eskimos was pathogenic for cotton rats or mice. No strains of Coxsackie virus were recovered. Main attention is directed to the examination of 51 sera from 40 persons (34 Eskimos) involved in the outbreak for the presence of neutralizing antibody to Lansing poliomyelitis virus. Antibody was found in the acute phase serum of two of three patients, and in the con valescent phase serum of about half of those tested. The serum of six of eight healthy Eskimos and five of six white contacts contained antibody. It seemed that antibody started to appear in the serum from the age of 10 onwards, the serum of most adults proving positive. It is concluded that the presence of Lansing antibody in the serum of the Eskimos was the result of previous exposure to Lansing virus and was unrelated to the current epidemic. The world-wide prevalence of Lansing antibody and by inference of Lansing virus is evident.

The Effect of Fa and Lansing Viruses on the Cholinesterase Activity of Brain and Spinal Cord Homogenates

Summary Mice were inoculated intracerebrally with FA virus and with Lansing virus. Total brain and spinal cord homogenates and the residue and supernatant fluid from these homogenates obtained by centrifuging were compared with similar preparations from uninfected mice in the following respects: wet weight, total nitrogen content, nitrogen concentration (μg N2/mg wet weight), and the cholinesterase activity expressed both in terms of wet weight and nitrogen content. The results were: There were no significant differences from the controls in mean wet weight and mean total nitrogen content for both brain and spinal cord with both viruses.With Lansing virus, there was a significant increase over the control in nitrogen concentration for spinal cord homogenate and the residue and supernatant fluid fractions of this homogenate. With FA virus, the nitrogen concentration of spinal cord residue was significantly increased over the control, while the total homogenate and residue fraction nitrogen concentrations were not significantly different. With neither virus was there any significant difference in the nitrogen concentration of the brain preparations.With FA virus, the cholinesterase activity, expressed in terms of tissue wet weight, was significantly decreased in the brain total homogenate and brain residue fraction (8.5% and 10.6% respectively), but not in the supernatant fluid fraction.With FA virus the cholinesterase activity of the spinal cord preparations did not differ significantly from the control when the activity was expressed in terms of tissue wet weight, but when the activity was expressed in terms of nitrogen content the spinal cord total homogenate enzyme activity was significantly lower than the control (13.6% decrease).With Lansing virus the cholinesterase activity of the three spinal cord preparations, but not of the brain preparations, was significantly lower than the controls, when the activity was expressed in terms of nitrogen content. These changes can be explained by the increased nitrogen concentrations in these preparations, therefore the decrease in enzyme activity probably is not a real one.The magnitude of the changes in cholinesterase activity is not sufficient to affect the function of the central nervous system.

Passive immunity in poliomyelitis: II Lansing antibody content of human gamma globulins.

ConclusionsBoth placental and plasma gamma globulins contain Lansing antibody. The 50% neutralizing titer of these products is approximately 10-3.5, as determined in mice. It would seem that a more potent gamma globulin could be obtained if prospective donors were first tested for serum Lansing antibody level. Then only those with high levels would be used to furnish plasma for the pools to be fractionated. Placental and plasma gamma globulins have a significant sparing effect when inoculated in mice 24 hours before a cerebral challenge of Lansing virus.

Failure to demonstrate antibody in feces of monkeys vaccinated with poliomyelitis virus, Lansing strain.

Summary and ConclusionFour monkeys were vaccinated with active preparations of the Lansing strain of poliomyelitis virus by parenteral routes. One animal received a total inoculum of 1.4 g of infected tissue, at the time of intracerebral challenge had an antibody titer in the serum of 800 against 10 LD50 of virus, but succumbed to an acute poliomyelitic infection. The remaining 3 received 2.4 g of inoculum, had antibody titers in their sera of approximately 2000 against 10 LD50 of virus and did not develop the disease. Each of 2 additional inoculations resulted in an increase in antibody of the serum to levels of 5100 and 8300 against 10 LD50 of virus. Despite the fact that Lansing virus antibody in the serum of three monkeys reached high titers, no evidence was obtained for the presence of antibody in the feces. Antibody was not detected in single specimens in the feces obtained from 6 humans whose sera had moderate titers of antibody to the Lansing strain of poliomyelitis virus. It is concluded on the b...

Monkey pathogenicity of various strains of murine poliomyelitis virus; experiments with Lansing, MEF, and Yale-SK group.

ConclusionThe data presented above demonstrate that strains of mouse-adapted human poliomyelitis virus may exhibit wide variations in the degree of their pathogenicity for rhesus monkeys. The Lansing virus, for instance, harvested from remote mouse passages, gave no evidence of being pathogenic for rhesus monkeys in repeated tests. The MEF virus, originally rhesus-pathogenic, apparently suffered a progressive loss of its pathogenicity for rhesus monkeys during serially maintained rapid mouse passages. The Y-SK virus, finally, seemed to have undergone but little change in its pathogenic power for rhesus monkeys as the strain was passed rapidly through mice. It is realized that the described phenomena may not always be duplicated in different laboratories since the Lansing virus has frequently been reported by others as being fully rhesus-pathogenic. Yet the authenticity of our strains is attested by the fact that all three murine strains had fully preserved their biological and immunological characteristic...

Some observations on interference between neurotropic viruses.

Studies have been carried out on the interference phenomenon with certain neurotropic viruses in experimental animals. The primary object was to investigate claims that material containing human poliomyelitis virus interferes with the progress of the infection by the MM virus in hamsters, or the Lansing virus in mice. We have failed to find evidence that the inoculation of specimens known to contain human poliomyelitis virus interferes with the progress of MM or Lansing virus infections, and we do not believe that the interference phenomenon can be used in this way as a diagnostic test. In the course of this investigation, it was found that interference occurs when hamsters are inoculated cerebrally with lymphocytic choriomeningitis virus, and, four to seven days later, with the antigenically unrelated MM virus by the peritoneal route. A significant number of such treated animals are spared from developing paralysis due to the MM virus. Evidence was also obtained that under certain circumstances the inoculation of MM virus may lessen the severity of the illness due to lymphocytic choriomeningitis virus injected several days earlier.

Monkey pathogenicity of various strains of murine poliomyelitis virus; experiments with Col. SK-MM group.

Conclusion1) Col-SK, MM, High Lansing and EMC virus, harvested from remote rodent passages, produce typical poliomyelitis in m-tracerebrally infected cynomolgus monkeys. 2) Two strains of highly virulent Theiler virus, GDVII and FA, are non-pathogenic for cynomolgus monkeys, 3) The Col-SK-MM group of virus can be carried in cynomolgus monkeys only for a limited number of passages. 4) Judging from serological and biological characteristics, monkey passage fails to bring about a permanent conversion of Col-SK virus into Y-SK virus.