Chickenpox & Shingles: No Transmission

“Incidentally, these tests furnish interesting information as to the portal of entry of the virus of varicella. The fact that the lymph from the vesicles, when applied to the broken skin or to the mucous membranes, led to no infection must cause us to consider this investigation from quite another standpoint. How are we to explain this lack of infectivity of the contents of the vesicles, which are generally regarded as the source of infection, and are used as the criterion in prescribing the duration of quarantine?”

doi:10.1001/archpedi.1918.01910130041005

In 1925, Rufus Cole, M.D., and Ann G. Kuttner, PH.D wrote a pretty scathing review on the evidence (or lackthereof) for the successful transmission of herpes zoster to both animal and humans. They felt compelled to not only share their own negative experiments but to also analyze and critique the work of others due to the growing body of work coming out claiming successful transmission yet lacking solid proof. Contained within are numerous examples of the failed transmission attempts of varicella, herpes zoster, and herpes simplex to both animal and human. The report is 22 pages long so obviously I could not copy/paste the whole thing here, but I tried to highlight most of the pertinent sections. I also highlighted many of the inhumane and grotesque experiments the unfortunate animals were subjected to all in the name of “science.” There is much I had to leave out so I highly recommend reading the whole report when you have the time. I provided a summary at the end:

THE PROBLEM OF THE ETIOLOGY OF HERPES ZOSTER.

The nature and etiology of that group of infectious diseases of which one of the features is a vesicular eruption on the skin are at the present time much confused. Rivers (1) has constructed a table indicating a possible relationship between a series of these diseases beginning with sheep-pox and horse-pox and extending through cow-pox, smallpox, varioloid, alastrim, chicken-pox, and herpes zoster to symptomatic herpes and lethargic encephalitis. Certain of these conditions resemble each other in their clinical manifestations, others have little in common. Certainly, the symptoms of herpes simplex have little resemblance to those of smallpox. The only feature present in all of them, except lethargic encephalitis, is a vesicular eruption of the skin. In most of the conditions the skin lesions show similar histological characteristics.

At one time or another some relationship in etiology between various members of the group, or even an identical etiology in all of them has been suggested. In none of the conditions has the etiological agent been cultivated, but there is considerable evidence that the responsible agent in most of them is ultramicroscopic or filterable. It is evident that in the absence of cultivation, in order to establish the etiological relation of an ultramicroscopic virus with one of these diseases, it is necessary to reproduce, in animals or man, lesions resembling the natural infection. With certain of these diseases, notably smallpox and vaccinia, the experimental reproduction of the disease is comparatively easy. With others, such as varicella and herpes zoster, all attempts to transmit them to animals have led only to equivocal and uncertain results. In the case of herpes simplex, although a virus has been isolated which is highly infectious for rabbits, the clinical picture produced is not, as will be discussed below, identical with herpes simplex in man.

Recent investigations have suggested an especially close etiologic relationship between varicella, herpes zoster, and herpes simplex. Some of the workers, mainly on the basis of clinical observations, claim an identity of the etiological agent concerned in varicella and herpes zoster, others are of the opinion that herpes zoster is due to a modified herpes simplex virus. An etiologic relationship between these conditions is also suggested by the fact that in the epithelial cells of the skin lesions in all of them acidophilic intranuclear inclusion bodies are found. These bodies were first described by Tyzzer (2) in varicelia and later by Lipschtitz (3) in herpes simplex and herpes zoster.

Inclusion bodies, both intranuclear and extranuclear, have been studied in great detail by Lipschfitz, and are considered by him to be characteristic of diseases of the filterable virus group. Goodpasture (4) has corroborated and extended Lipschiitz’s studies of these structures in connection with herpes simplex and agrees with Lipschfitz as to the specific nature of intranuclear inclusion bodies. Rivers and Tillett (5) have demonstrated similar intranuclear inclusion bodies in the corium of skin lesions associated with a rabbit virus isolated by them. In the study of diseases of this group, the finding of these characteristic nuclear changes in experimental lesions in animals or man is therefore of importance in determining whether the reaction obtained is specific. Following the observation by Gfiiter (6) in 1920 and those of Lowenstein (7, 8) investigators in all parts of the world have demonstrated that the virus of herpes simplex, when inoculated into the scarified cornea of rabbits produces with great regularity a vesicular eruption followed by an intense keratoconjunctivitis. Inoculations into the skin less frequently give rise to lesions. As Doerr and Vochting (9) first observed, corneal inoculations are frequently followed by marked nervous symptoms and death and similar symptoms can be produced by direct inoculation into the brain. The inoculations of the virus into rabbits, therefore, gives rise to lesions which may resemble those seen in man, but in most cases the lesions and symptoms differ both in character and severity from those present in the mild and common condition in human beings known as herpes simplex. It is of importance, however, that in all the lesions produced in animals, including those in the cornea, the skin, and the brain, the most characteristic feature of the lesion of herpes simplex in man is reproduced; namely, the occurrence of ceils containing intranuclear inclusion
bodies. The intracutaneous inoculation of the vesicular fluid of herpes simplex either into an individual already infected with herpes simplex or into a normal person has not given as constant results as has the inoculation of this fluid into the rabbit’s cornea. Man’s susceptibility to herpes simplex seems to depend on certain secondary factors which are, at the prese~nt time, unknown.

The successful inoculation of rabbits with herpes simplex material was followed by attempts to transmit varicella to animals, but so far these attempts have not been successful. (For a review of the literature, see Rivers and Tillett (1).) Many attempts have been made to reproduce varicella in man by inoculating material from active cases into normal individuals. Kling (10) reported the successful vaccination of children against chicken-pox by inoculation with vesicular fluid. Certain later observers employing the method of Kling have noted the development of a local vesicle or papule at the site of inoculation, others have described the occurrence of a generalized eruption (true chicken-pox?), others have stated that no obvious lesions result from the inoculation. So far as we are aware no histological study has been made of any of the lesions described. The difficulty in successfully inoculating animals or man with varicella virus is of interest in view of the claims which have been made regarding the identity of the viruses of chicken-pox and herpes zoster.

Inoculation of Virus of Herpes Zoster into Animals.

Prior to 1921, attempts to inoculate animals with herpes zoster had proved negative. In this year Lipschtitz (3) reported successful results. Seven cases which were apparently typical as regards clinical manifestations were studied. The vesicular fluid was obtained early in the disease and was rubbed into the scarified cornea of rabbits. In certain instances the fluid was combined with the “roofs” of vesicles. Lipschutz considers that positive results were obtained with the material from four cases. In view of the importance of his conclusions a brief review of his cases will be given.

Case I.–It is stated that the inoculation of the material into the rabbit’s eye was followed in 4 days by a slight opacity of the cornea along the lines of scarification. The eye was removed and sections were made through the cornea. Occasional giant epithelial cells were present along the lines of scarification. Under Bowman’s membrane hypertrophied and swollen connective tissue cells were seen but no leucocytes. In the nuclei of the epithelial cells and also of the swollen connective tissue cells there were to be seen occasional, typical, round, sharply circumscribed and clearly demonstrable intranuclear inclusion bodies.

Cases II and III.–These were also early cases and material from them was similarly inoculated into the eyes of four rabbits. In one of the rabbits after 4 days the cornea showed an intense circumscribed keratitis, with the appearance of a slightly elevated “eitrig getriibten,” and “daherweiszlich” appearing vesicle. The microscopic examination of this eye showed marked infiltration with pus cells. No inclusion bodies were found. In another animal a keratitis developed, but the occurrence of vesicles or the presence of inclusion bodies is not noted. The results in the other two animals were negative. In these two cases, therefore, the evidence presented in the protocols which indicates positive results is very slight.

Case IV.–This was also a typical early case of herpes zoster. Vesicular material from this case was inoculated into the eyes of two rabbits and two guinea pigs. The results in the two guinea pigs and in one of the rabbits were negative. In the second rabbit the inoculated eye showed on the 2nd day conjunctivitis and circumscribed corneal infiltration. The eye was enucleated on the 4th day and in microscopical sections very numerous intranuclear inclusion bodies were found in the epithelial cells.

The remaining three cases were studied at later periods of the disease and the results were negative.

The experiments of Lipschutz with material from these seven cases can, therefore, as judged from his brief protocols, be considered to have yielded positive results in only two animals, and in these instances the results are of importance chiefly on account of the presence of intranuclear inclusion bodies in the epithelial cells.

Lipschutz (11) considers that his positive findings have been confirmed by Marinesco and Draganesco, Truffi, Mariani, and Blanc and Caminopetros, and that the successful transmission of herpes zoster to animals has thus been accomplished. It is therefore important to review in some detail the reports of these investigators.

Marinesco (12), and Marinesco and Draganesco (13), injected material from three cases of herpes zoster.

Case I.–Herpes zoster localized on the thigh. Vesicular fluid was inoculated into the scarified corneas of four small rabbits, and into the second cervical ganglion of two small cats. The rabbits all remained unaffected. The ganglia of the cats were examined after 7 days and in one of them there was evident lymphatic infiltration and atrophy of the neurones.

Case II.–Herpes zoster of the first and second branches of the trigeminus nerve. Since the vesicles contained but little fluid, spinal fluid obtained on the 6th day of the disease was used for inoculation. The injections were made into the anterior chamber of one eye, and into the scarified cornea of the other eye in each of nine rabbits. Moreover, in five of these nine rabbits, in addition to the eye inoculations, 0.2 cc. of spinal fluid was inoculated intracerebrally. The eyes of the first three rabbits showed only injection due to injury. On the 4th day, in Rabbit 4, in the cornea of the eye in which the injection was made into the anterior chamber, there was noted a zone of infiltration. Rabbit 5 showed two points of infiltration on the scarified cornea. Rabbits 6, 8, and 9 were negative. Rabbit 7, besides a febrile reaction on the 4th day, developed an area of infiltration reaching the center of the pupil. The emulsified brain and cerebellum of this rabbit were inoculated into the cornea of four more rabbits, two of which showed on the 3rd and 4th days grayish infiltration along the lines of scarification. No statement is made concerning microscopic examination.

So far as can be judged from the protocols, therefore, inoculations made with the material from these two cases produced no characteristic lesions. A macroscopic infiltration of the cornea can hardly be regarded as specific.

Case III.–Herpes zoster lesions on the thigh. Vesicular fluid was inoculated into the scarified cornea of three rabbits, two of which on the 4th day showed a linear infiltration. The writer states (12) that the sections of the cornea of one of these rabbits showed swollen, edematous cells, and that here and there could be seen the “specific nuclear lesion, consisting in atrophy of the chromatin which is pushed toward the membrane, while the acidophilic mass, which has developed, offers a striking resemblance to the inclusions described by Lipschiitz in animals injected with herpes.” It is undoubtedly on this last statement that Lipschutz bases the view that his own observations have been confirmed by Marinesco and Draganesco. Certainly nothing else in the protocols indicates the occurrence of a specific lesion.

Truffi (14) studied three cases of herpes zoster. The results in the first two were negative. Vesicular fluid from the lesion of Case 3, cervicobrachial in distribution, was obtained on the 3rd day of the disease and inoculated into the scarified cornea of one rabbit. After 48 hours a slight opacity along some of the lines of scarification, and an intense conjunctivitis were noted. The corneal opacity disappeared rapidly and the eye regained its normal appearance by the 7th day. 22 days after inoculation the rabbit showed symptoms of encephalitis and was killed 10 days later. The microscopical examination of the brain was negative. The presence of intranuclear inclusion bodies in the brain cells is not noted. The inoculation of the brain emulsion into the scarified corneas of two rabbits and two guinea pigs failed to produce lesions.

Most of the attempts made by Mariani (15) to inoculate the cornea of rabbits with herpes zoster resulted negatively. In one instance he obtained a very acute keratoconjunctivitis with hypopyon and purulent ophthalmia. In only one case did there result a keratitis which he was able to transmit in series. The lesion produced was clinically and symptomatically very similar to the keratitis produced by herpes simplex virus. No description of the case of herpes zoster from which the material for inoculation was obtained, is given. No statement concerning microscopic examination of the corneas is made. Mariani himself considers this single experiment inconclusive.

Material from nine cases of herpes zoster was inoculated by Blanc and Caminopetros (16) into the eyes, cornea, conjunctiva, skin, brain, and spinal cord of a series of animals, including rabbits, mice, sheep, pigeons, monkeys, and a dog. Three monkeys (Macacus rhesus) were inoculated as follows: one into the eye, one into the spinal canal, and the third into the skin of the thoracic region which had previously been shaved and excoriated. The inoculations in the first two monkeys resulted negatively. The third monkey showed a slight inflammatory reaction at the site of inoculation but recovered without the appearance of vesicles. All the other animal experiments gave negative results with the following exceptions; two rabbits developed a late paralysis which, however, the authors considered was probably not specific, and one rabbit and one sheep, both inoculated with material from the same case, developed a definite keratitis, which spread from the point of inoculation. The writers think that this lesion might easily be interpreted as a reaction resulting from the injection. They conclude that the problem of the transmission of herpes zoster to animals remains open and they apparently consider their own experiments negative or inconclusive.

Meineri (17) claims to have produced encephalitis in a guinea pig by the intracerebral inoculation of vesicle fluid from a case of herpes zoster. A careful analysis of his experiments, however, in our opinion, indicates that his findings can best be interpreted as the result of trauma. The writer also injected vesicle fluid obtained on the 3rd day of the disease from one of his cases of herpes zoster into the skin of the arm of the patient and into the skin of a normal man. These injections in both instances were without visible result.

The review of the publications of those writers whom Lipschutz quotes as having confirmed his work shows that two of the writers regard their own results as inconclusive. Only Marinesco and Draganesco found microscopical lesions which might be interpreted as specific.

On the other hand, many other authors report entirely negative results following the inoculation of herpes zoster material into the sacrified corneas of rabbits: Kraupa (18); Baum (19); Lowenstein (8), Teissier, Gastinel, and Reilly (20); Kooy (21); Netter and Urbain (22); Bloch and Terris (23); Simon and Scott (24); and Doerr (25).

It is evident, therefore, that the results of attempts to inoculate animals with material from cases of herpes zoster must be considered at present to be inconclusive.

Herpes Simplex and Herpes Zoster.

Although it has not been possible to demonstrate conclusively any specific virus associated with herpes zoster, certain writers have presented evidence which suggests that, in certain cases at least, the symptoms and lesions of herpes zoster may result from the presence of the virus of herpes simplex.

Luger and Lauda (26) have published several papers on the problem of the etiology of herpes zoster. In their first paper they give the results obtained by inoculation with material from seven cases of typical herpes zoster, employing the technique used by Lipschfitz. In none of the eyes inoculated did any macroscopic reaction occur. On microscopical examination there was found fairly regularly edematous swelling of the epithelial cells, giant cell formation, and “ballonierende” degeneration, but in no instances were cell inclusions or characteristic changes of the nuclei seen. They themselves considered the results in this series of experiments negative.”

“Gruter (27) inoculated material from three cases of herpes zoster into the scarified corneas of rabbits. A mild keratitis resulted. No detailed description of the lesion or results of microscopic examination are given. Griiter, however, believes the lesion obtained was specific and attributes it to herpes simplex virus of a low grade of virulence. He states that there is no evidence for assuming a specific virus for herpes zoster. The data presented, however, are not sufficient to establish the isolation of a true herpes simplex virus from these cases.

Bastai and Busacca (28), in a general article on herpes, state that they inoculated material from three cases of herpes zoster into the cornea of rabbits and into the cornea of one monkey (Macacus). Rabbits were also inoculated intracerebrally. None of the animals showed any reaction, with the exception of one rabbit which developed a slight kerafitis. No attempts were made to transmit this lesion, and no microscopical examinations are reported. These authors also are of the opinion that herpes zoster is probably a manifestation of infection with herpes simplex virus. The experimental data presented, however, are hardly sufficient to justify this point of view.

Teague and Goodpasture (30) were able to produce zoster-like lesions in the skin of rabbits and guinea pigs by the inoculation of herpes simplex virus into areas of the skin previously treated with coal tar. The study of the corresponding posterior root ganglia showed lesions comparable to those found in man in the ganglia innervating the area of zonal eruption. The writers do not maintain that they have reproduced the human disease herpes zoster in animals, but they believe there is a close analogy between the experimental condition produced by them and true herpes zoster. In their opinion the herpes simplex virus first multiplies at the site of inoculation in the skin and passes up the corresponding spinal nerve to its spinal ganglion; the virus then seems to pass centrifugally along the nerve and its branches to the skin, where it multiplies rapidly and gives rise to characteristic herpetic vesicles. They draw attention to the difficulty of sharply separating cases of herpes simplex and herpes zoster and discuss the occurrence of intermediate cases. From a case of the intermediate type inoculations were made into the tarred skin of a rabbit. A zonal eruption, as described above, resulted. They feel that the evidence presented strongly suggests that the virus of herpes zoster is closely allied to the virus which causes herpes simplex, probably differing only in virulence.

The interesting hypothesis presented by Teague and Goodpasture (30) and by Luger and Lauda (26) concerning the relation of herpes zoster to herpes simplex does not find acceptance, however, by Lipschutz (31). He emphasizes the point of view that in the production of an experimental herpes zoster it is of prime importance that the starting point be a typical clinical case and not a border line case.

At the present time the evidence that herpes zoster may result from infection with herpes simplex virus rests upon the isolation of a virus apparently identical with that of herpes simplex from a small number of cases. No description of the type of case from which the material employed for inoculation was obtained is given by Gruter or by Bastai and Busacca. The case described by Teague and Goodpasture and the first case described by Luger and Lauda belong to the intermediate type of cases. The second case of Luger and Lauda, and the so called symptomatic cases of Cipolla, seem to have been clinically typical cases of herpes zoster. It is possible, therefore, that in certain instances the virus of herpes simplex may be isolated from cases clinically characteristic of herpes zoster, but the evidence for this is not complete and the conclusion that herpes zoster may be the result of infection with herpes simplex virus needs further verification.

Varicella and Herpes Zoster.

Several observers (Lipschiitz, Meineri, and others) have made isolated attempts to inoculate human volunteers with herpes zoster, but always with negative results. Recent studies of Kundratitz (32) seem to show that herpes zoster can be successfully transmitted to very young children. This author wished to test out Von Bokay’s (33) hypothesis, based on clinical observation, that the virus of varicella, under certain unknown conditions, may produce a typical picture of herpes zoster and that the virus from this lesion may in turn cause varicella. He therefore attempted to immunize children against varicella by the inoculation of material from herpes zoster cases. His first results were negative, but his later attempts proved successful. He now reports that he has inoculated material from ten typical cases of thoracic herpes zoster and has had positive results with the material from five of these cases. Positive reactions were obtained only in children under 5 years of age. Children who reacted positively were subsequently shown to be immune to varicella.

Kundratitz’s work seems to indicate that the virus of varicella and that of herpes zoster are identical or, at least, closely related. It is unfortunate that Kundratitz does not give a description of the cases of herpes zoster used by him for inoculation. It would be interesting to know whether there were any clinical differences between the five cases of herpes zoster with which he was able to make successful transfers and the five cases in which transfers resulted negatively for, as Von Bokay and others have shown, the vesicles of varicella may be quite localized, resulting in lesions resembling herpes zoster. The relation between herpes zoster and varicella will, in all probability, not be entirely cleared up until we are able to transmit either one or both of these diseases to animals.

EXPERIMENTAL.

Nine cases of herpes zoster have been studied by the writers and inoculations have been made into a series of animals. The following are abstracts of the histories and protocols of the experimental studies.

Case I.–A. M. Age 13. Patient admitted to the hospital Oct. 4, 1924, suffering from subacute rheumatic fever and chronic cardiac disease. She gave no history of a previous attack of herpes zoster or chicken-pox. The arthritis had almost entirely disappeared and the cardiac lesion was well compensated, when on Dec. 6, 1924, the patient complained of pain and itching over the upper scapular area, in the axilla, and posterior part of the upper arm. On examination of this area there was discovered a rash consisting of small, discrete papules and vesicles distributed in patches over a zone on the upper chest from the midline behind to the midsternal line in front, and over the inner and posterior surface of the arm. The area of distribution corresponded to Head’s second and third dorsal areas. During the following days the vesicles became larger. The temperature was not higher than 99.8 ° until Dec. 11, when some of the vesicles had become pustular, and now the temperature rose to 101.4 °. The pain was severe and characteristic of herpes zoster, and the appearance and distribution of the lesions were typical. A small piece of skin was removed and microscopical sections showed characteristic vesicles with numerous intranuclear inclusion bodies in the epithelial cells.

On Dec. 9, the 3rd day of the disease, fluid was pipetted from a number of vesicles and a small piece of the involved skin was obtained. The skin was ground between two glass slides, the ground material was washed off in a small amount of normal saline solution and was added to the vesicular fluid. Small amounts of this emulsion were rubbed into the scarified 2 corneas of two rabbits, Nos. 1 and 2, and also injected intracutaneously into the shaved skin of Rabbit 2 and into the skin of Guinea Pig 1. The area of skin in the guinea pig where the injection was made had been painted several days previously with coal tar solution. The emulsion was also rubbed into the scarified skin of Rabbit 3, which had received one painting of tar 5 days before, and into the scarified skin of a similarly tarred guinea pig, No. 2. (The rabbit and guinea pigs were painted with a refined coal tar solution obtained through the courtesyof Dr.Jas. B. Murphy. This refined coal tar was much less toxic than ordinary tar and could be applied in a fairly thick coat, so that one painting resulted in a marked reaction.) On the following day, Dec. 10, vesicular fluid was again obtained from fresh vesicles and also another piece of skin. This material was treated in the same way as that obtained on the preceding day, and inoculated in the following ways. The cornea of Rabbit 1 and the scarified tarred skin of Rabbit 3 and Guinea Pig 2 were reinoculated. Some of the material was also inoculated intracerebrally into Rabbit 4 and into the scarified cornea of Rabbit 5. Thus, with fresh material obtained on the 3rd and 4th days of the disease, five rabbits and two guinea pigs were inoculated in various ways.

The animals were carefully observed each day following the inoculations, but in none of the animals were any macroscopic changes seen which could be ascribed to the inoculations. One of the eyes of Rabbit 2 was removed on the 3rd day and the other on the 7th day following the inoculations and sections were made through the corneas. The sections show in places what are apparently the results of mechanical injuries and in the section of the eye removed on the 7th day, loci of slight infiltration of the substantia propria with small round cells. Some swelling of certain of the epithelial ceils is also seen. But nowhere are there any signs of vesicle formation or marked inflammatory reaction and no inclusion bodies were found.

Although no definite reactions were obtained in this first series of animals it was thought that by inoculating from one cornea to another and from one brain to another, the virus might possibly become adapted to the rabbit and produce definite lesions in subsequent transfers. Therefore, starting with Rabbit 1 inoculations were made from one rabbit to another by scraping the cornea and washing out the conjunctival sac with a small amount of saline and inoculating the material thus obtained into the scarified cornea of another rabbit. Fourteen corneal passages were thus made, at 2 and 3 day intervals. In many of the rabbits, the scarified eye on the day following the inoculation showed a slight degree of opacity along the lines of scarification and a slight exudation. However, it was found during the course of the study that slight changes of this character frequently occur following the inoculation of an emulsion of normal rabbit cornea, and even after scarification alone without the injection of any foreign matter whatever. Except for these slight non-specific reactions no changes were observed in any of the eyes of the series. In certain instances, although no gross changes were present, the cornea was sectioned but no lesions which could be considered specific and no intranuclear inclusion bodies were found.

Starting with Rabbit 4 inoculated intracerebrally with the material from this case, ten brain to brain transfers were made at 5 day intervals. Each animal was killed with ether, and the brain removed with sterile precautions. An emulsion of the brain was made with Locke’s solution in a sterile mortar, the suspension centrifuged at low speed, and 0.2 cc. of the supernatant fluid injected intracerebrally into a normal animal. At the same time, some of the brain emulsion was inoculated intracorneally and intradermally into each of two other rabbits. It was found that the inoculation of brain emulsion into the scarified cornea usually was followed by conjunctivitis of considerable severity which, however, proved to be wholly non-specific. The temperature of the intracerebrally inoculated rabbits was taken daily, and sections of the brain of each of the inoculated animals made. None of the rabbits showed a significant rise in temperature and careful study of the brain failed to reveal any characteristic lesions. No intranuclear inclusion bodies were found.”

SUMMARY AND DISCUSSION.

“In making the animal experiments we employed various methods which were suggested largely by the technique used by previous observers, especially by those who have reported results which were considered positive. In making inoculations into the corneas the technique recommended by Lipschutz was employed as far as possible. Young rabbits were used and the material was obtained from fresh vesicles early in the disease and inoculated with as little delay as possible. The material injected into rabbits’ eyes was obtained from seven cases and twenty-four rabbits were used. In judging of the results obtained in this kind of experimentation great caution must be observed. Our experience convinces us that slight opacities occurring along the lines of scarification and mild conjunctivitis cannot be held to indicate the effect of a specific virus. As regards the interpretation of the microscopic changes found, we were quite familiar with the appearance of intranuclear inclusion bodies as seen in the lesions of experimental herpes simplex and the filterable virus (Virus III) indigenous to rabbits described by Rivers and Tillett (5). We also had no difficulty in finding intranuclear inclusions in the sections of skin removed from patients. It is not likely, therefore, that these structures were overlooked in our study of the sections. Briefly stated, although the material studied was satisfactory and in spite of the fact that a considerable number of animals were used for each case, we have been unable to confirm the observations of Lipschtitz regarding the experimental production of specific lesions in the corneas of rabbits. We realize that this is only negative evidence and therefore not of conclusive importance in view of Lipschiitz’s observations. It indicates, however, that the production of specific lesions in rabbits’ eyes with material from herpes zoster vesicles is extremely difficult and that successful results may be a matter of chance, depending, possibly, on peculiar susceptibility on the part of the rabbits. In view of the fact, however, that a careful analysis of the positive results reported by other observers shows that the conclusions were based on insufficient evidence, we believe that further work is necessary before the successful inoculation of the rabbits’ corneas with herpes zoster virus can be accepted as fully demonstrated. To make the evidence convincing specific lesions should be obtained with a fair degree of regularity and the virus should be successfully transmitted through at least two generations. Apparently the latter was not attempted by Lipschutz.

Intracerebral inoculations into three rabbits with material from two cases (Nos. I and IV) were made. Two rabbits were also inoculated intraspinally with material from one case (No. IV). None of these animals showed any reaction. In the case of one of the animals inoculated into the brain (Case I) although this rabbit showed no symptoms, we thought it conceivable that the susceptibility of the species for the virus might be so slight that no obvious lesion had been produced. Nevertheless it was thought that the virus might possibly remain alive at the seat of inoculation and by repeated transfers become adapted to the rabbit. This phenomenon has been observed by Noguchi with vaccine virus, and by Rivers and Tillett with the rabbit virus isolated by these workers. This possibility was tested by us by making serial corneal and brain inoculations. Corneal transfers were carried through fourteen animals in series, and brain transfers through ten. No specific lesions developed in any of the animals.

The work of Teague and Goodpasture suggested that the skin might be rendered more susceptible to infection by previous treatment with tar. Material from two cases (Nos. I and VIII) was inoculated into the tarred skin of guinea pigs and rabbits. The material was injected intracutaneously and also rubbed into the scarified skin. No reaction was obtained in any of the animals.

Finally, the transmission of herpes zoster to monkeys was attempted. Blanc and Caminopetros, and Bastai and Busacca, as discussed in the review of the literature, inoculated monkeys (Macacus) in various ways, without success.

It was thought possible that although monkeys of the genus Macacus might be refractory, monkeys of another genus might prove susceptible. Consequently, besides the inoculation of two Macacus monkeys, attempts were made to infect five vervets. Moreover, in view of the fact that the virus of vaccinia and the rabbit virus of Rivers and Tillett could be successfully cultivated in the testicle, intratesticular inoculations were employed. The testicles were removed at varying periods following inoculation. Numerous sections of these testicles were made and examined, but in no instance were any lesions found which could be interpreted as specific. No cells containing intranuclear inclusion bodies were found. These experiments, therefore, have also led to purely negative results.

This report of our work is made at the present time because a considerable amount of literature has been published which gives the impression that herpes zoster has been successfully transmitted to animals. Although the observations of Lipschutz are suggestive, it is important that they be confirmed by further investigations.

Until herpes zoster can be regularly transmitted to animals and cross-immunity tests be carried out, the relation of the virus of herpes zoster to that of herpes simplex remains a matter of speculation. In view of the fact that herpes simplex can be easily and regularly transmitted to rabbits, whereas in the hands of a large number of investigators similar experiments with herpes zoster are completely negative, it does not seem likely that the etiological agent concerned in these two diseases can be absolutely identical.

The question of the identity or non-identity of herpes zoster and varicella is even more difficult to answer, because at present neither of these infections is readily transmissible to animals. The work of Kundratitz is extremely interesting. His observations, aside from indicating a close immunological relationship between herpes zoster and varicella, are important in that they seem to show the presence of a transmissible virus in the vesicles of herpes zoster. The only question that arises is whether the cases of herpes zoster from which Kundratitz was able to make successful transfers were true cases of idiopathic herpes zoster.

CONCLUSION.

Attempts to inoculate rabbits, guinea pigs, and monkeys with material obtained from nine cases of herpes zoster have proved uunsuccessful.

doi: 10.1084/jem.42.6.799.

In Summary:

• The nature and etiology of the infectious diseases where one of the features is a vesicular eruption on the skin are confused
• Rivers stated there was a possible relationship between a series of these diseases beginning with sheep-pox and horse-pox and extending through cow-pox, smallpox, varioloid, alastrim, chicken-pox, and herpes zoster to symptomatic herpes and lethargic encephalitis
• Certain of these conditions resemble each other in their clinical manifestations while others have little in common but they all share eruptions of the skin
• In most of the conditions the skin lesions show similar histological characteristics
• At one time or another some rrelationship in etiology between various members of the group, or even an identical etiology in all of them has been suggested
• The etiological agent had not been cultivated in any of the above conditions
• Since no “viruses” had been cultivated, the only “proof” was reproducing the same lesions in animals or man
• Attempts to transmit varicella and herpes zoster to animals were largely uunsuccessful
• In the case of herpes simplex, they could not reproduce an identical disease in animals to that experienced by humans
• Investigations suggested an especially close etiologic relationship between varicella, herpes zoster, and herpes simplex
• Some of the workers, mainly on the basis of clinical observations, claimed an identity of the etiological agent concerned in varicella and herpes zoster, others were of the opinion that herpes zoster is due to a modified herpes simplex “virus”
• Inclusion bodies, both intranuclear and extranuclear, have been studied in great detail by Lipschutz, and are considered by him to be characteristic of diseases of the filterable “virus” group
• Quick sidenote: Intranuclear inclusion bodies (INB) are aggregates of protein that are said to be frequently encountered in “viral” infections, where they are thought to be accumulations of “viral” particles, yet for RNA “viruses” replicating in the cytoplasm, this compartmentalization represents a paradox not consistent with the “viral” replication cycle https://pubmed.ncbi.nlm.nih.gov/1332364/
• In the study of diseases of this group, the finding of characteristic nuclear changes in experimental lesions in animals or man was of importance in determining whether the reaction obtained is specific
• Injecting herpes simplex into the scarified eyes of rabbits consistently produced vesicular eruptions and keratoconjunctivitis while injecting it into the skin did not regularly produce lesions
• In most cases the lesions and symptoms in rabbits differed both in character and severity from those present in the mild and common condition in human beings known as herpes simplex
• Intracutaneous inoculation of the vesicular fluid of herpes simplex either into an individual already infected with herpes simplex or into a normal person did not give constant results
• Man’s susceptibility to herpes simplex seems to depend on certain secondary factors which are unknown
• Attempts to transmit varicella to animals have not been successful
• There has been much difficulty in successfully inoculating animals or man with varicella “virus”
• Attempts using similar inoculation methods produced a papule at the site of injection or a generalized eruption (which they were unsure if it were “true” chickenpox) while others produced no lesions whatsoever
• Atempts to inoculate animals with herpes zoster were unsuccessful
• Various experiments on rabbits and Guinea pigs were performed by inoculation of vesicular fluid into their eyes
• The emulsified brain and cerebellum of one rabbit were inoculated into the cornea of four more rabbits
• In another experiment, the inoculation of the brain emulsion into the scarified corneas of two rabbits and two guinea pigs failed to produce lesions
• The experiments of Lipschutz with material from these seven cases can be considered to have yielded positive results in only two animals, and in these instances the results are of importance chiefly on account of the presence of intranuclear inclusion bodies in the epithelial cells
• Vesicular fluid was inoculated into the scarified corneas of four small rabbits and they all remained unaffected
• In another case, injections were made into the anterior chamber of one eye, and into the scarified cornea of the other eye in each of nine rabbits while in five of these nine rabbits, in addition to the eye inoculations, 0.2 cc. of spinal fluid was inoculated intracerebrally
• The eyes of the first three rabbits showed only injection due to injury
• Rabbits 6, 8, and 9 were negative
• Inoculations made with the material from these two cases produced no characteristic lesions
• A macroscopic infiltration of the cornea can hardly be regarded as specific
• Truffi studied three cases of herpes zoster and the results in the first two were negative
• The microscopical examination of the brain of the third case was negative and the presence of intranuclear inclusion bodies in the brain cells was not noted
• The inoculation of the brain emulsion into the scarified corneas of two rabbits and two guinea pigs failed to produce lesions
• Most of the attempts made by Mariani to inoculate the cornea of rabbits with herpes zoster resulted negatively
• In only one case did there result a keratitis which he was able to transmit in series, however:
  1. No description of the case of herpes zoster from which the material for inoculation was obtained, was given
  2. No statement concerning microscopic examination of the corneas is made
  3. Mariani himself considered this single experiment inconclusive
• Material from nine cases of herpes zoster was inoculated by Blanc and Caminopetros into the eyes, cornea, conjunctiva, skin, brain, and spinal cord of a series of animals, including rabbits, mice, sheep, pigeons, monkeys, and a dog
• Three monkeys (Macacus rhesus) were inoculated as follows: one into the eye, one into the spinal canal, and the third into the skin of the thoracic region which had previously been shaved and excoriated
• Inoculations in the first two monkeys resulted negatively
• The third monkey showed a slight inflammatory reaction at the site of inoculation but recovered without the appearance of vesicles
• All the other animal experiments gave negative results
• The writers of one study thought that the lesion brought about by their experiment might easily be interpreted as a reaction resulting from the injection
• These authors concluded that the problem of the transmission of herpes zoster to animals remained open and they considered their own experiments negative or inconclusive
• Analysis of the experiments by Meineri indicated that his findings can best be interpreted as the result of trauma
• His attempts to inoculate two humans with herpes zoster both failed
• 2 out of the 3 researchers Lipschutz cited as backing up his results stated their results were inconclusive
• Many other authors reported entirely negative ressults following the inoculation of herpes zoster material into the scarificed corneas of rabbits: Kraupa; Baum; Lowenstein, Teissier, Gastinel, and Reilly; Kooy; Netter and Urbain; Bloch and Terris; Simon and Scott; and Doerr
• According to Cole, it was evident that the results of attempts to inoculate animals with material from cases of herpes zoster must be considered to be inconclusive
• It had not been possible to demonstrate conclusively any specific “virus” associated with herpes zoster
• Certain writers presented evidence which suggested that, in certain cases at least, the symptoms and lesions of herpes zoster (shingles) may result from the presence of the “virus” of herpes simplex (herpes)
• Luger and Lauda published several papers on the problem of the etiology of herpes zoster
• In their first paper they give the results obtained by inoculation with material from seven cases of typical herpes zoster, employing the technique used by Lipschutz and in none of the eyes inoculated did any macroscopic reaction occur
• They considered the results in this series of experiments negative
• Gruter stated that there is no evidence for assuming a specific “virus” for herpes zoster
• The data he presented was not sufficient to establish the isolation of a true herpes simplex “virus” from these cases
• Bastai and Busacca were of the opinion that herpes zoster was probably a manifestation of infection with herpes simplex “virus” but experimental data presented was not sufficient to justify this point of view
• None of the animals they inoculated presented any lesions
• Teague and Goodpasture were able to produce zoster-like lesions in the skin of rabbits and guinea pigs by the inoculation of herpes simplex “virus” into areas of the skin previously treated with coal tar
• However, they did not maintain that they reproduced the human disease herpes zoster in animals, only that there is a close analogy between the experimental condition produced and “true herpes zoster”
• They drew attention to the difficulty of sharply separating cases of herpes simplex and herpes zoster and discuss the occurrence of intermediate cases
• Teague and Goodpasture felt that the evidence presented strongly suggested that the “virus” of herpes zoster is closely allied to the “virus” which causes herpes simplex, probably differing only in virulence
• Lipschutz did not accept the hypothesis by several researchers about the relation of herpes zoster to herpes simplex
• He emphasized the point of view that in the production of an experimental herpes zoster it is of prime importance that the starting point be a typical clinical case and not a border line case
• The evidence that herpes zoster may result from infection with herpes simplex “virus” rests upon the “isolation” of a “virus” apparently identical with that of herpes simplex from a small number of cases
• However, while Cole felt it may be possible these “viruses” are related, it is inconclusive and in need of verification
• Several researchers attempted to infect humans with herpes zoster with negative results
• Kundratitz tried to infect children yet his first attempts were negative
• His second attempts were “successful” in infecting 5 of 10 children all under the age of five
• Kundratitz did not provide any details on the herpes zoster cases used for infection
• Cole states the relation between herpes zoster and varicella will not be entirely cleared up until they are able to transmit either one or both of these diseases to animals
• A case is presented of a girl who gave no history of a previous attack of herpes zoster or chicken-pox
• She ended up with a shingles-like rash around her torso with severe pain and and the appearance and distribution of the lesions typical and characteristic of herpes zoster
• The vesicular fluid from the girl with suspected herpes zoster was inoculated in various ways (eyes, skin, brain) into 5 rabbits and 2 Guinea pigs
• There were no signs of vesicle formation or marked inflammatory reaction and no inclusion bodies were found
• Inoculations were made from one rabbit to another by scraping the cornea and washing out the conjunctival sac with a small amount of saline and inoculating the material into the scarified cornea of another rabbit – fourteen corneal passages were made in this way
• They noticed slight changes in the cornea but found that these changes frequently occur following the inoculation of an emulsion of normal rabbit cornea, and even after scarification alone without the injection of any foreign matter whatsoever
• No lesions could be considered specific and no intranuclear inclusion bodies were found
• 10 brain to brain transfers were made at 5 day intervals in some rabbits
• An emulsion of the brain was made with Locke’s solution in a sterile mortar, the suspension centrifuged at low speed, and 0.2 cc. of the supernatant fluid injected intracerebrally into a normal animal
• Some of the brain emulsion was inoculated intracorneally and intradermally into each of two other rabbits
• None of the rabbits showed a significant rise in temperature and careful study of the brain failed to reveal any characteristic lesions
• No intranuclear inclusion bodies were found
• 8 other cases of the disgusting torture of animals in order to transmit herpes zoster are presented in detail in the paper and not one of them produced any intranuclear inclusion bodies
• Cole states that their experience convinces them that slight opacities occurring along the lines of scarification and mild conjunctivitis cannot be held to indicate the effect of a specific “virus”
• Careful analysis of the positive results reported by other observers showed that the conclusions were based on insufficient evidence
• Their experiments indicated that the production of specific lesions in rabbits’ eyes with material from herpes zoster vesicles is extremely difficult and that successful results nay be a matter of chance
• They state further work is necessary before the successful inoculation of the rabbits’ corneas with herpes zoster “virus” can be accepted as fully demonstrated
• Cole determined that to make the evidence convincing, specific lesions should be obtained with a fair degree of regularity and the “virus” should be successfully transmitted through at least two generations
• Material from two cases was inoculated into the tarred skin of guinea pigs and rabbits intracutaneously and also rubbed into the scarified skin yet no reaction was obtained in any of the animals
• Attempts to transfer herpes zoster to monkeys by various reseachers were all uunsuccessful
• Numerous sections of these testicles were made and examined, but in no instance were any lesions found which could be interpreted as specific
• No cells containing intranuclear inclusion bodies were found and these experiments led to purely negative results
• Cole detailed their negative experiments with attempting herpes zoster transmission to animals due to the growing amount of literature suggestive of successful transmission of herpes zoster to animals when this had not been the case
• He stated that until herpes zoster can be regularly transmitted to animals and cross-immunity tests are carried out, the relation of the “virus” of herpes zoster to that of herpes simplex remains a matter of speculation
• Both varicella and herpes zoster had not been shown to be transmissible to animals
• Cole questioned whether the cases outlined by Kundratitz were “true” herpes zoster cases
• He concluded that attempts to inoculate rabbits, guinea pigs, and monkeys with material obtained from nine cases of herpes zoster proved unsuccessful

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