A case of Neonatal Herpes Simplex virus (HSV) encephalitis with severe ocular manifestation, treated by topical, systemic and intrathecal Interferon inducer - Poly IC (Polyriboinosinic acid-Polyribocytidylic acid) - is reported. The clinical course, the mechanism of action of Interferon, and the facts known in literature on the subject are dealt with in detail.
A three- week-old female infant was born to nonrelated parents after an uneventful pregnancy and normal vaginal delivery. Birth weight was 2400 gr. After birth, she made good progress and gained weight. Four days before admission, she became pyrexic, extremely excited, and anorexic. One day prior to admission, right-sided convulsions began which slowly progressed to all of the trunk. On examination, there were no gross findings apart from generalized bilateral tonic and clonic seizures with marked motor irritability and hyperexcitability. In the right corner of the upper lip, there was a solitary yellowish vesicle. The anterior fontanelle was normotensive. Ophthalmic examination was within normal limits. Lumbar puncture yielded clear fluid under normal pressure, with up to 80 lymphocytes/mm\ with normal levels of glucose and chlorides, and total protein levels of 400 mg/dl. Blood and CSF cultures did not grow bacteria. PPD skin test was normal. Treatment with intravenous ampicillin (200 mg/kg) and garamycin (6 mg/kg) was initiated. Administration of Diazepam (5 mg IV in repeated doses), phénobarbital (30 mg/day)and chloral hydrate20 ml of 10% solution thrice daily) failed to stop seizures. Severe respiratory distress followed by respiratory arrest required intubation and mechanical ventilation. On the third hospital day multiple vesicles erupted on the face in the typical location and having the typical appearance of primary neonatal occular HSV infection (Fig. 1 ). There was a bilateral moderate injection of bulbar and palpebral conjunctivae with serosanguineous discharge but without corneal involvement.
Samples of blood, csf and fluid from the cutaneous vesicles were taken for virological, bacteriological and serological examination. At that stage we reexamined the parents for the presence of genital HSV infection, which was confirmed by them and by clinical examination.
In view of the positive history of active genital herpetic infection in both parents, and the typical cutaneous eruption on the infant's face, neonatal HSV encephalitis was diagnosed.
Intravenous and topical treatment with Poly IC was begun. An initiating IV dose of 1000 Y/ml in a bolus injection was followed by 2000 Y/day for seven successive days. A total of 600 Y Poly IC daily was applied topically on the vesicles and in the conjunctival sacs, four times a day for seven days. On the fifth day of Poly IC treatment, 100 Y was injected intrathecally followed by 200 Y for another two days. During this course of treatment, daily samples of blood, CSF and vesicular fluid were taken for virus isolation serologic work up and Interferon level measurements. Daily liver function tests and blood counts were also performed. Slight clinical improvement in vital functions followed, body temperature dropped, and mechanical ventilation was not required. The infant was still semicomatose, nonreactive to painful stimuli and continued to suffer from sporadic convulsions for short periods. An EEG tracing showed complete flattening of cortical activity and disappearance of rhythmic activity. Contrast air studies were compatible with obstructive hydrocephalus and porencephaly. On repeated ophthalmic examination, bilateral chorioretinitis was present. The retina showed large patches of massive yellowish-white exudates in the posterior pole. The infant remained in vegetative state for three months and died of terminal bronchopneumonia and sepsis.
Fig. 1. The infant showing development of herpetic skin lesions and vesicles near the lid margins.
ISOLATION OF HERPES SIMPLES VIRUS AT TIME OF POLY IC TREATMENT IN THE CASE OF NEONATAL GENERALIZED HSV INFECTION
Laboratory Data: Virus Isolation
The vesicular fluid and tears were cultured for HSV by rolling cotton swab over the surface lesions and the conjunctiva of the lower fornix of the eye. The swabs were immediately placed into vials containing 1.5 ml medium 199 (1% calf serum and antibiotics). The samples were inoculated into tubes of monolayer culture of Rabbit Kidney primary cells, which were examined for cytopathogenic effect daily.
Serum Neutralizing Antibody Titer
The neutralizing antibody titer was assayed by inhibition of the cytopathogenic effect in Vero ceils using a standard type 2 virus.' Fluorescent antibody studies were performed using the F.A. Technique of Chien Liu2 according to the indirect method of identification.
The laboratory results are summarized in Table I and Table II. No virus could be isolated from the cutaneous lesions and conjunctival sac after three days of Poly IC treatment.
Rising levels of antibodiesto Herpes Simplex using the CFT method were found in the blood. These levels. were from 1:30 to 1:60 during the illness.
Antibodies to Herpes in CSF were found on the first day and remained constant throughout the patient's illness.
Interferon Levels were measured in the blood, CSF, and tears during Poly IC treatment. Before administration of Poly IC Interferon could not be detected in the blood and CSF, but 24 hours after IV injection, there was a measureable level (1:8-1:32) in the blood, which remained constantduring the treatment. It is worth mentioning that IV treatment in the dosage scheduled in Table II did not initiate Interferon presence in CSF. Intrathecal administration of Poly IC induced a significant rise in Interferon levels in the CSF.
The pathological findings in the brain were compatible with massive destruction of normal tissue, with diffuse and focal gliosis perivascular mononuclear infiltration, which also spread to the meninges. Many calcified inclusion bodies were seen (Fig. 2).
Herpes virus hominis known also as Herpes Simplex Virus is a common cause of fatal encephalitis in adults and neonates. Its two stereotypes are completely different in their clinical and pathologic manifestations.'
Type I in adults causes lesions in the mouth, cornea, skin, and central nervous system (CNS). Type II is transmitted exclusively by genital contact, and usually causes only genital and cutaneous lesions, but may cause a diffuse disease. 4"h Many reports of ocular involvement such as conjunctivitis, chorioretinitis, uveitis, and also encephalitis in neonates with disseminated HSV infection have recently been reported.7"
The incidence of congenital infection has not been established, but some believe it to be about one case to 7500 live-births. I>u About 80 percent of the genital infections are caused by HSV type II. 14,15
The clinical course of neonatal HSVencephalitis is very similar in the majority of the reports. It usually begins with pyrexia, convulsions, deteriorating consciousness, and depression of vital centers.
The diagnosis usually is made late in the course of the disease, after failure of bacteriologie and biochemical work upto define the etiology. Only by specific virologie and serologic laboratory procedures and a high index of suspicion may the viral etiology be proved.
Brain biopsy has been the most accepted approach for viral identification and isolation. 15 Recently, more rapid and accurate methods for viral identification have been utilized. These methods include complement-fixation test (CFT) to HSV,: neutralization test,' and immunofluorescent.u'~w
According to the literature there are three therapeutic possibilities: (1) administration of IDU (5 IOD 2-Deoxyuridine); (2) administration of Cytosin-Arabinoside; and (3) use of interferon. IDU is an antimetabolite, which acts as a competitive inhibitor to Thymidine uptake into the RNA molecule.2" Since it was introduced into clinical trial good results had been achieved in the treatment of topical herpetic keratitis and mucco-cutaneous lesions."' Many side effects have been observed on systemic administration of the drug, such as stomatitis, alopecia, and severe bone marrow depression."
The established concept in the past years has been to avoid the use of IDU because of the bone marrow cytotoxicity.'3"25
Cytosine Arabinoside was not used because of the known cytotoxic effects in this very you ng age, and also because we realized that the drug would not penetrate the blood-brain barrier.26
A new approach in antiviral therapy has emerged following the discovery of interferon by Lindeman and Isaacs."7 They found that this protein with a molecular weight of 20,000 is produced in reaction to viral infection or nonviral factors, and acts by preventing the replication of DNA molecule and inhibiting the action of DNA. Interferon is synthesized in any cellular tissue, after being induced by viruses specific group of double-stranded RNA molecules, endotoxins, gram-negative bacteria and mitogens such as phytohemagglutinins and streptolysin. 2*~w The lymphocytes are considered the major site for interferon production, and this characteristic is constant beginning in fetal life and in the postnatal period.1'"12
Two mechanisms are considered responsible for Interferon induction: (1) synthesis of Interferon de novo by each cellular tissue in reaction to various stimuli as described; (2) release of Interferon stored in the reticuloendothelial system. The inactive form is activated during its release from RES cells.28
The disadvantage of endogenous Interferon is its ultrashort action and the persistance of very low serum concentration. Positive "defending" effect is achieved when Interferon is administered close to the infection.29'10'"
Fig. 2. Pathological findings in the brain: (a) Massive destruction of normal tissue: (b) Focal gliosis: and (c) Calcified inclusion bodies. Paraffin sections, H and E stains, photomicrographs.
Among the stimulating factors mentioned above, the most extensively investigated in mankind is a synthetic polymer, Poly IC.
This is practically a double stranded RNA molecule with a potent Interferon-induction ability." This preparation is very effective against various viruses as was proved in animal and human experimentation."
The conclusions drawn from experimentation with Poly IC in animals are: (1) there is a defending effect of the drug if administered before or even after the infection occurred36"37; (2) the levels of induced Interferon are linearly dependant on the dosage of the drug, route, a nd frequency of administration36'3"; and (3) there were no hepatic or hematologic complications in human trials.
De Clerque et al39 were the first to report intrathecal injection of Interferon to an infant suffering from herpetic meningo-encephalitis. They found high titers of Interferon in the CSF, but failed to show any clinical improvement. It is worth mentioning that they began their treatment on the 20th day of the infant's illness. In view of the conclusions above, it seems that the results could have been improved if Interferon had been injected much earlier and close to the onset of the disease.
Our considerations favoring administration of local and systemic Poly IC to the infant presented were based on the following criteria:
1 . Purified human Interferon is not commercially available, is expensive and must be repetitively injected because of it's ultrashort action.
2. We have accumulated a clinical experience with Poly IC administered topically and systemically to 61 patients with severe Herpetic Dentritic Keratitis and Iritis, who completely recovered, without evident side effects of the drug.21
3. We were aware of the side effects of the other mentioned antiviral drugs.
4. The commercially available Poly IC used by us has been proved to be purified, nonpyrogenic, nontoxic in animal experimentations, and highly effective as an Interferon inducer.
Analysis of our clinical results show that:
1 . Virus Clearance from the conjunctiva and cutaneous lesions was remarkably rapid under topical and systemic Poly IC treatment, as previously reported.21
2. Intravenous Poly IC injection did not cause a rising titer of Interferon in CSF as expected in experimental models.""35 A significant rise of Interferon levels in cerebrospinal fluid was evident, on the other hand, after intrathecal injection of Poly IC.
3. There was a mild clinical improvement in the infant's general condition and vital functions, and possibly some alterations in the EEG tracings.
4. There was no defencing effect on the deteriorating cortical degeneration, and progressive hydrocephalus. This supports the basic concept that "defending" effect is achieved when Poly IC or Interferon are administered very shortly after infection has occurred.
For these reasons we believe that Poly IC should be given to any suspected or proved case of viral encephalitis, as soon as suspicion is raised in order to prevent irreversible neuronal damage. In order to initiate rapid rise of Interferon in the CSF, we believe that Poly IC should be injected intrathecally.
We believe that more therapeutic experience must be accumulated in order to define the optimal dosage and adequate period of therapy.
A case of Neonatal Herpes Simplex Virus (RSV) Encephalitis with severe ocular manifestation treated by topical systemic and intrathecal Interferon inducer. Poly IC (Polyriboinosinic acid-Polyribocytidylic acid) is reported. Prior to the administration of Poly IC, no Interferon could be detected neither in blood nor in CSF and vesicular fluid from conjunctiva. Intravenous administration of the drug initiated measurable levels in blood and tears, but not in CSF. However, intrathecal injection induced persistent high levels of Interferon in CSF. Virus clearance from conjunctiva tears and vesicular fluid occurred after three days of topical application of Poly IC.
The mechanism of action of Poly IC, the laboratory results and the therapeutic conclusions are discussed in detail. This is the first report of intrathecal administration of Poly IC in a case of HSV Encephalitis.
1. Rawls WE, Iwamoto K, Adam E, et al: Measurements of antibodies to HSV type I and II in human sera. J Immunol 3:104, 1970.
2. Chien Liu: Fluorescent antibody techniques. In Lennette EH, Schmidt NJ: Diagnostic Procedures for Viral and Rickettsial Infections, 4th ed. 1969, pp 179-204.
3. Nahmias AJ: Infections caused by herpes virus hominis. In Hoeprich PD (ed): Infectious Diseases. New York, Harper & Row, 1972, p 841.
4. Rawls WE, Gradner LH, Flanders WR, et al: Genital herpes in two social groups. Am J Obstet Gynecol 110:682, 1971.
5. Nahmias AJ, Josey EW, Nalb MZ, et al: Perinatal risk associated with maternal genital HSV infection. Am J Obstet Gynecol 1 10:825, 1971.
6. Allen JC, Waiters G, Ogorman M, etal: J Pediatr 89:947, 1976.
7. Belantini JA, Guin GJ, Grassi RM, etal: Herpes simplex encephalitis: brain biopsy a nd treatment with 5' iodeo deoxyuridine. J Pediatr 72:266, 1968.
8. Florman AL, Mindlin RL: Generalized Herpes simplex in an 1 1 -day-old premature infant. Am J Dis Child 83:481, 1952.
9. Golden B, Bell WE, McKee AP: Disseminated herpes simplex with encephalities in a neonate. J Am Med Assoc 209:1219, 1952.
10. Hagler WS, Walters PV, Nahmias AJ: Occular involvement in neonatal herpes simplex virus infection. Arch Ophthalmol 82:169, 1976.
11. Nahmias AJ, Alfords SB, Korones SB: Infection of the newborn with herpes virus hominis. Adv Pediatr 17:185, 1970.
12. Torphy DE, Ray GC, McAHster R, et al: Herpes simplex infection in infants - a spectrum of disease. J Pediatr 76:405, 1970.
13. Zavoral MJ, Ray LW, Kinnard GP, et al: Neonatal herpetic infection. A fatal consequence of penile herpes in servicemen. J Am Med Assoc 213:1492. 1970.
14. Nahmias AJ, Dowdle WR, Josey WE, et al: Newborn infection with herpesvirus type I and II. J Pediatr 75:1 194, 1969.
15. McCallum FO, Potter JM, Edwards DH: Early diagnosis of herpes simplex encephalitis by brain biopsy. Lancet 2:332, 1964.
16. Nachmias AJ, Delbuno I, Schneweis KE, et al: Type specific surface antigens of cells infected with HSV (1 & 2). Proç Soc Expl Biol Med Vol. 136, 1971.
17. Liu C: Fluorescent antibody techniques. In Lenette EJ, Schmidt NJ (ed): Diagnostic Procedures for Viral and Ricketsial Infection, 4th ed. 1969, p 197.
18. Taber LH, et al: Diagnosis of herpes simplex virus by immunofluorescence. J Clin Microbiol 3(3):309, 1976.
19. Lerner AM, Wilson FM, Lauter CB, et al: An estimate of the course of herpes simplex encephalitis. Scand J Inf Dis 8:37, 1976.
20. Kaufman HE, Nesburn AB, Maloney ED: IDU therapy of herpes simplex. Arch Ophthalmol 67:583, 1962.
21. Eylan E, Romano A, Ben-Tovim T, et al: Infectious inflammations of the outer eye. Harefuah 85:552, 1973.
22. Breeden CJ, Hall TC, Tyler RH: Herpes simplex encephalities treated with systemic IVDR. Ann Intern Med 65:1050, 1966.
23. Liversedge LA: Current ideas and treatment, plans for future. Proc Roy Soc Med 69:193, March 1976.
24. Rappel M: Postgraduate Med J, 49:419, 1973.
25. Boston Interhospital Virus Study Group and the NIAID Sponsored Cooperative Antiviral Clinical Study. ? Eng J Med 292:599, 1975.
26. Chien LT, Whitley RJ, Nahmias A, et al: Antiviral chemotherapy and neonatal HSV infection: a pilot study - experience with adenine-arabinose. Pediatr 55:678, 1 975.
27. Isaacs A, Lindermann C: Virus interference. I. The interferon. Proc Roy Soc Series ? 147:191, 1957.
28. Levine S, Nichel FR: Interferon inducers. Bioscience 20:696, 1970.
29. Hilleman MR, Tytell AA: The unduction of interferon. Sci Am 225:26, 1971.
30. Grossberg SE: The interferon and their inducers: molecular and therapeutic consideration. ? Engl J Med 287:13-19, 79-85, 122-128, 1972.
31 . Canthel K, Strander H, Saxen I, et al: Interferon response of human lymphocytes during intrauterine and post natal life. Jlmmunol 100:1304, 1969.
32. Green JA, Cooperband SE, Kibrick S: Immune specific induction of interferon production in cultures of human blood lymphocytes. Science 164:1415, 1969.
33. Park JH, Baron S: Herpetic karato conjunctivitis: therapy with synthetic double stranded RNA. Science 162:811, 1968.
34. DeClerque E, Nuwer MR, Merigan TC: The role of interferon in the protective effect of a synthetic double-stranded polyribonucleotide against intranasal vesicular stomatitis virus challenge in mice. J Clin Investig 49:1565, 1970.
35. Hilleman MR: Double stranded RNA's Poly I.C.) in prevention of viral infection. Arch Inter Med 126:109, 1970.
36. Hilleman MR: Prospects for use of double stranded ribonucleic acid (Poly I.C.) inducers in man. J Infect Dis 121:196, 1970.
37. Tazulakhova EB, Novokhatsky AS, Yershov Fl: Interferon induction by, and antiviral effect of Poly I.C. in experimental viral infections. Acta Virologia 17:487, 1973.
38. HO M, Nash C, Morgan CW, et al: Interferon administration in the cerebro spinal space and its effects on rabies in rabbits. Infect Immun 9:286, 1974.
39. DeClerque E, Edy VG, De Uliger H, et al: Intrathecal administration of interferon in neonatal herpes simplex encephalitis. J Pediatr 86:736, 1975.
40. Johnson RT, Olson LC, Buescher EL: Herpes simplex virus infection of the nervous system. Problems in laboratory diagnosis. Arch Neurol 18:260, 1968.
ISOLATION OF HERPES SIMPLES VIRUS AT TIME OF POLY IC TREATMENT IN THE CASE OF NEONATAL GENERALIZED HSV INFECTION