R. Wigand

New human adenovirus (candidate adenovirus 36), a novel member of subgroup D.

SummaryA new adenovirus (candidate Ad36) was isolated from the feces of a girl with enteritis. It has the biological and biophysical properties of an adenovirus, with a particle diameter of 750±10 Å in thin sections and with a fiber length of 190 Å. The virus belongs to subgroup D. It is distinct both in neutralization and hemagglutination-inhibition from all other human adenoviruses; it also shows a unique DNA restriction pattern.

Neutralization of the Adenoviruses Types 1 to 28: Specificity and Antigenic Relationships.

Cross-neutralization tests of rabbit immune sera against adenoviruses types 1 through 28 revealed a high degree of specificity in many instances. Reciprocal cross-reactions were observed between types 4 and 16, types 15 and 25, types 3 and 7, types 7, 11, and 14, types 11 and 21 and, with some exceptions, between types 1, 2, 5, 6, and 12. The majority of the cross-neutralizations were of low titer. No antigenic difference was found between type 7 and 7a. The antigenically related subgroups correspond well with subgroups of the human adenoviruses, as defined by other criteria.

Analysis of antigenically intermediate strains of subgenus B and D adenoviruses from AIDS patients.

SummaryWe carried out detailed antigenic and restriction enzyme (RE) analyses on the subgenus B and D adenovirus isolates from 48 AIDS patients. These isolates were unusual both in the diversity of serotypes and in the number of intermediate strains identified. All unusual isolates were strain-purified and tested by serum neutralization (SN) and hemagglutination-inhibition (HI) tests with reference horse and rabbit antisera to all the prototype human adenoviruses; conversely, rabbit antisera were prepared to 16 selected strains from this study and tested by SN and HI with all prototype viruses. Among subgenus B strains, 6 DNA variants of Ad 11 isolates were distinguished by endonucleases BamHI, BglII, BstEII, HindIII, and SmaI. The D2 variant of Ad 11 was prominant with 5 isolates, and 7 other isolates differed from D 2 in only 1 or 2 enzymes. HindIII was the most discriminative endonuclease for Ad 11 and related strains. Within subgenus D, there were 16 isolates of intermediate strains, including 4 intermediate types related to new Ad types 43–47. The RE patterns of subgenus D strains showed fragment distributions typical of subgenus D, with various unique patterns. Particular care was taken to analyze multiple strains from the same patient, recovered as much as 8 months apart, for evidence of genetic changes. The possible long-term infections with these viruses may provide the opportunity for mutations and recombination to occur, with the resultant generation of a variety of new strains of adenoviruses.

Isolation and Identification of Enteric Adenoviruses

Thirty‐four out of 64 faecal samples with adenovirus particles, as seen by electron microscopy, were found to contain adenovirus 40 or 41 by direct isolation and neutralization in Changs conjunctival cells, mostly within one week. (Ad40 and 41 candidate viruses are serologically related.) 6 other adenovirus specics were isolated; 6 samples gave equivocal results, and 18 were negative. A genus‐specific ELISA with an antihexon coat yielded positive results in 40 out of 55 samples; the test failed to identify adenovirus antigen in 10 out of 17 specimens, which were found negative by culture. All of them were negative by immunfluorescence of inoculated Chang cell cultures. Hence the failures are probably due to insufficient amount of virus in the samples. The predominance of only two adenovirus species associated with gastroenteritis in children and the ease of cultivating and identifying them should help to elucidate their etiological significance.

Immunological relationship among human adenoviruses of subgenus D

SummaryAntigenic relationships between adenoviruses of subgenus D were determined by neutralization tests in HeLa cell cultures by CPE inhibition. For cross-testing, several antisera of the same species were tested against the prototype viruses 39 wild strains belonging to 12 different virus species were also studied. Marked variation in the degree of cross-neutralization between individual sera of the same species was often observed. However, virus strains within a species mostly showed identical serological reactions. Hence, antigenic specificity appears to be a fairly constant property of any one species.Strong cross-neutralizations between species are presumably due to a relationship of the ε (hexon) antigen, whereas weak cross-neutralizations found between viruses related by hemagglutination-inhibition are due to the γ (fiber) antigen.Viruses related to adenovirus 15(Mastadenovirus h 15) showed a variety of cross-reactions in neutralization tests. In view of the new species definitions of adenoviruses and to facilitate identification, changes in the classification of Ad 15, 25, 29, and 15/H9 are proposed. The prototypes of Ad 13, 15, 25, 29, and 30 have been cloned by terminal dilution.

Über den cytopathogenen Effekt der Adenoviren des Menschen

Bei Hellfeldmikroskopie lebender ungefärbter HeLa-Zellkulturen nach Infektion mit Adenoviren ließen sich drei Arten des cytopathogenen Effektes (CPE) unterscheiden, die als Rundzelltyp, Retraktionstyp und Sondertyp bezeichnet werden. Die Untersuchung von 28 Prototypviren und 32 Wildstämmen verschiedener Typen ergab ein konstantes Verhalten der Virusstämme und, mit wenigen Ausnahmen, der Stämme innerhalb der Serotypen hinsichtlich ihres CPE. Die daraus resultierende Einteilung der Adenoviren des Menschen nach ihrem CPE (Tabelle 1) stimmt mit keiner der anderen bekannten Einteilungen dieser Virusgruppe überein. Die praktische Bedeutung dieser Einteilung zur Typisierung von Adenoviren wird dadurch eingeschränkt, daß die Art der Ausprägung des Retraktionstyps von dem jeweils benutzten Zellstamm abhängt. So waren die Unterschiede in HeLa-Zellen am deutlichsten. Rundzelltyp und Sondertyp waren dagegen in allen untersuchten permanenten Kulturen gleich. Eine Beziehung zwischen Retraktionen und Virustoxin ist wahrscheinlich; jedoch führt auch toxinfreies Virus (nach Zentrifugieren, Trypsinbehandlung oder in hoher Verdünnung) zu einem CPE mit Retraktionen. Living unstained HeLa cell cultures infected with various adenoviruses exhibited three kinds of cytopathic effect (CPE), which were designated as round cell type, retraction type, and special type. The investigation of 28 proto-type viruses and 32 wild strains of various types revealed a constant association of virus strain and CPE. This was valid also, with few exceptions, for different virus strains of the same serotype. The human adenoviruses can be classified according to the kind of CPE they produce (Table 1). These groups do not correspond to any of the other known classifications of this virus family. The practical significance of these groups for typing adenoviruses is limited by the fact that the aspect of the retraction CPE is different in different cell strains. Thus the differences were particularly striking in HeLa T cells. On the other hand, round cell and special CPE were similar in all permanent cell cultures investigated. A relation between retractions and virus toxin is likely; on the other hand, toxin-free virus, provided by centrifugation, trypsin treatment or by passage in minimal amounts, also exhibited a CPE with retractions.

Inhibition of adenovirus multiplication by metabolic inhibitors.

Under conditions of one-step growth the multiplication of adenovirus 19 in HeLa cells was inhibited by 7 inhibitors of DNA synthesis (see Table 1), by cycloheximide, streptovitacin A, actinomycin D and mitomycin D. The inhibition involved the formation of infectious virus and of capsid proteins. The viral CPE was not inhibited by inhibitors of DNA synthesis; cycloheximide, streptovitacin and actinomycin inhibited the CPE with viral multiplicities up to 2. With medium doses of IUdR and BUdR the formation of infectious virus was inhibited with almost normal formation of capsid proteins. A selective formation of group-specific CF antigen was also observed under a number of conditions. The inhibitory effect of several drugs was reversible on removal. A reversion of the inhibition mediated by amethopterin, fluoruracil and FUdR was achieved only by addition of thymidine. No reversion was seen after removal of actinomycin. Deoxycytidine had an antagonistic effect against the inhibition mediated by cytosine arabinoside, when added 8 to 22 hours after the drug. The time during the replication cycle until which the inhibitors are fully effective was found to be 8 to 10 hours for inhibitors of DNA synthesis, 12 hours for actinomycin and 14 hours for cycloheximide and streptovitacin. This inhibition time was largely independent of inhibitor doses and viral multiplicity. When inhibitors were added during the exponential rise of viral activity, cycloheximide and streptovitacin stopped the further rise. Cycloheximide and actinomycin also stopped the progression of viral CPE. Apparently continuous protein synthesis is required for both viral maturation and viral CPE. Eor cycloheximide two inhibitor-sensitive periods (early proteins from 6 hpi, structural proteins from 14 hpi) were distinguished, while the period of synthesis of viral DNA lasted from 8 to 18 hours.

Eigenschaften der Adenovirus-Hämagglutinine

SummaryAdenovirus hemagglutinins were classified into five groups designated as A to E according to differences in heat stability, sensitivity to trypsin treatment or to trypsin plus subsequent heating, adsorption of hemagglutinin and infectious virus to red cells and separation of hemagglutinin from infectious virus by ultracentrifugation. These properties are summarized in Table 12. Hemagglutination by A hemagglutinin is demonstrable only in the presence of heterologous adenovirus immune serum, probably of all serotypes. Hemagglutinin A may modify red cell receptors in absence of heterologous serum; these receptors appear to be different from those involved in hemagglutination by myxoviruses. The mechanism of hemagglutination by the various hemagglutinins is discussed.

Genome type analysis of adenoviruses: isolates from one year from the Hannover area

SummaryAdenoviruses (AV), isolated from 138 children during the year 1981 in the Hannover area, were studied by DNA restriction analysis with the enzymes BamHI, Bg1II, BstEII, EcoRI, HindIII, KpnI, and SmaI and compared with the respective prototypes. Varying fragment patterns were depicted and genome types analyzed. Prototype-like strains of AV1 and 5 were not found. Types 2, 5, 1, 3, and 7 showed decreasing genetic variation in that order. Altered restriction sites were physically mapped on the genome; they appeared to be randomly distributed. The high genetic variability of AV2 and 5 is remarkable for this study population, limited in time and space.Clinical and epidemiological data were also presented in relation to serotypes.

Serologische Beziehungen der Adenoviren der Gruppe II

Thirty-six wild strains from 11 serotypes of group II adenoviruses were compared with the respective prototype viruses by hemagglutination-inhibition tests with prototype rabbit antisera. With a few exceptions, the cross-reactions observed were identical for all strains of a given type. This was true also for strains, which were not neutralized by the homologous antiserum, but corresponded to the prototype in HA-inhibition. Strong cross-reactions were found between types 8, 9 and 9–15, types 10 and 19, types 15 and 22, types 27 and 28, types 29 and 25.