Fiber sequence heterogeneity in subgroup F adenoviruses.

The fiber gene of adenovirus type 41 was sequenced and compared to the fiber gene sequence of adenovirus type 40 (A. H. Kidd and M. J. Erasmus, 1989, Virology 172, 134-144), the other known member of subgroup F. The open reading frame, from map units 87 through 92 with transcription from the r-strand, comprised 1686 bases and was 45 bases longer than its counterpart on the Ad40 genome. The 45-base difference appears to have resulted from a block deletion on the Ad40 sequence. Apart from this one region, the Ad40 and Ad41 fiber genes showed remarkably high homology (95.6%), indicating a relatively recent evolutionary divergence. The deduced amino acid sequence of the Ad41 fiber polypeptide was analyzed according to the model of N. M. Green et al. (1983, EMBO J. 2, 1357-1365) for the structure of the adenovirus fiber. Ad41 had one more 15-residue repeat in the shaft region than Ad40, there being 22 repeat motifs. A detailed study of various Ad40 and Ad41 strains with proven genome differences indicated that the 15-amino acid difference in polypeptide length at the 14th repeat motif is a type-specific difference among the subgroup F adenoviruses. However, two uncommon Ad41 strains belonging to 2 of the 16 Ad41 genome types tested had a 15-amino acid block deletion which was different to that of the Ad40 polypeptide. The implication from this work is that the Ad40 fiber gene probably arose from its Ad41 counterpart, but the fiber gene sequences of both types of subgroup F adenovirus are so similar that genetic recombination between strains could occur with some frequency.

Sequence characterization of the adenovirus 40 fiber gene.

The fiber gene of human adenovirus type 40 has been characterized. The 6.1-kbp EcoRI fragment C of the Ad40 genome, from map units 74 through 92, was cloned and the right-most 2.8 kbp from 84 map units was sequenced. By analogy with Ad2, this region would be expected to contain the gene specifying the Ad40 fiber polypeptide. Sequencing revealed an open reading frame of 1641 bases on the r-strand, the first 53 bases of which had marked homology with the corresponding L5 (fiber) regions of Ad2 (77.2%), Ad5 (75.0%), and Ad3 (64.3%). In addition, base positions 1114 to 1146 of this open reading frame had 85% homology with base positions 1198 to 1230 of the Ad2 fiber gene. The predicted polypeptide sequence of 547 amino acids showed marked homology with the Ad2, Ad5, and Ad3 fiber polypeptides in two regions, in the first 55 amino acids from the N-terminus and from amino acids 372 through 382. Analysis of hydrophobic amino acid positions revealed a repeating pattern of approximately 15 residues between positions 42 and 374, with 21 repeats. The sequence of the Ad40 polypeptide thus fits the model of Green et al. [1983), EMBO J. 2, 1357-1365) for the structure of the adenovirus fiber, but is 35 amino acids shorter than the Ad2 fiber polypeptide, with one less 15-residue repeat in the shaft region. According to this model, the regions of highest homology between the Ad40 fiber polypeptide and those of Ad2, Ad5, and Ad3 correspond to the tail of the shaft and the base of the knob. The results of this analysis are in agreement with previously published EM data on the fiber length of subgroup F adenoviruses.

Infection by enteric adenoviruses, rotaviruses, and other agents in a rural African environment.

From February 1985 to January 1986, 432 stool samples, 310 from rural African children with diarrhea and 122 from controls, were analysed for the presence of enteric viruses known to be associated with diarrhea. Group A rotavirus ELISA indicated 12.9% positivity among patients and 2.5% positivity among controls. Only 23 of the 43 rotavirus ELISA-positive stools were also positive by electron microscopy. Nine children, three of whom were controls, were found to be shedding coronavirus-like particles, detected by electron microscopy. Stools from all but one of the nine children had been taken within 1 month of each other. Dot-blot hybridization tests for the presence of Ad40 or Ad41 DNA revealed 44 positive stools, 41 of which were from patients (13.2% positivity). Only three of the Ad40-or Ad41-positive stools by DNA hybridization were positive by electron microscopy, and only these three strains could be grown in semipermissive Chang conjunctival cells and their identity checked by restriction enzyme analysis. Further attempts to rescue the other strains using a helper virus failed, but nine of the stools proved positive by ELISA using a subgroup F-specific monoclonal antibody. On the basis of the DNA hybridization results alone, subgroup F adenoviruses were encountered as frequently as rotavirus in the study and were significantly associated with diarrhea, although the viability and intactness of virus particles by the time of laboratory analysis appeared to be very low.

Congenital complete heart block in an infant with free plasma carnitine deficiency.

A newborn male presenting with congenital complete heart block was found to have low levels of free plasma carnitine. Among the many complications of carnitine deficiency is a structural disruption of myocardial tissue. A deficiency of carnitine may be causally related to the presence of cardiac dysrhythmias in this patient on the basis of mechanical disturbance to the cardiac conducting system.

Epidemiologic and clinical features of Crimean-Congo hemorrhagic fever in southern Africa.

Following the diagnosis in 1981 of the first case of Crimean-Congo hemorrhagic fever (CCHF) in South Africa, an antibody survey was undertaken on cattle sera to determine the distribution of the virus and specific diagnostic tests were routinely applied to specimens from suspected cases of hemorrhagic fever to establish the medical significance of its presence. Antibody to CCHF virus was demonstrated by reversed passive hemagglutination-inhibition technique in 2,460/8,667 (28%) cattle sera and in 140/180 herds tested in South Africa, as well as in 347/763 (45%) cattle sera and in 32/34 (94%) herds tested in Zimbabwe. The antibody was found in all major cattle farming areas, but was of low prevalence along the southern coast where 2 of the 3 species of Hyalomma tick which occur in South Africa are absent. From February 1981 to January 1986, inclusive, 29 indigenous cases of CCHF were diagnosed in 16 outbreaks which arose in various locations throughout South Africa. A further 2 imported cases of CCHF arose in Zaire and Tanzania. The clinical features of infection conformed to the classical descriptions of CCHF in the Soviet Union. The fatal outcome in 11/31 cases indicates that the African disease is no less severe than that which occurs in Eurasia. It is inferred that the virus is widespread in all countries in Africa and Eurasia which lie within the limits of world distribution of ticks of the genus Hyalomma.

Comparison of techniques for demonstrating antibodies to Rift Valley fever virus.

Nine serological techniques were compared by monitoring the response to infection with Rift Valley fever (RVF) virus in three sheep. Antibodies were monitored daily for the first 14 days after infection, then weekly and later fortnightly up to week 24. The earliest antibody response was detected in one sheep on day 3 by a plaque reduction neutralization test, and by day 6 antibodies were demonstrable in all three sheep by haemagglutination-inhibition, reversed passive haemagglutination-inhibition, immunodiffusion, indirect immunofluorescence (IF), enzyme-linked immunosorbent assay and neutralization of cytopathic effect in cell cultures. Antibodies were demonstrable by complement fixation on day 8 at the earliest. IF and the two neutralization techniques produced the highest titres, but all tests could be used satisfactorily for the serological diagnosis of RVF. Inactivated antigen could be used for all except the neutralization tests. A radioimmunoassay technique using 125I-labelled staphylococcal protein A detected antibodies on day 8 at the earliest and produced lower mean titres than some of the other techniques. This was probably because sheep immunoglobulins bind protein A poorly.

Comparative pathogenicity and antigenic cross-reactivity of Rift Valley fever and other African phleboviruses in sheep.

Homologous and heterologous haemagglutination-inhibition (HAI), complement-fixation (CF), immunodiffusion (ID) and mouse neutralization tests were performed with the Lunyo (LUN) and a Zimbabwean strain of Rift Valley fever (RVF) virus, the prototype and a South African strain of Arumowot (AMT) virus and prototype strains of Gordil (GOR), Saint-Floris (SAF) and Gabek Forest (GF) viruses, using immune mouse ascitic fluids prepared against these viruses. Reactions of identity occurred in all tests between LUN and the Zimbabwean strains of RVF and between the two strains of AMT virus. Otherwise, cross-reactions occurred between all the phleboviruses in HAI tests, while reactions in CF, ID and neutralization tests were monospecific for virus serotypes, except that weak cross-reaction occurred between GOR and SAF viruses in CF and ID tests. Four sheep infected subcutaneously with the Zimbabwean strain of RVF virus developed transient fever, viraemia, leucopaenia, relative thrombocytopaenia, haemoconcentration and raised serum enzyme levels, which indicated that the sheep had developed necrotic hepatitis. Disseminated focal necrotic hepatitis was confirmed in a sheep killed for examination on day 4 post-infection. The other three sheep recovered uneventfully after only mild depression and anorexia. Groups of three sheep infected with SAF, GOR, AMT and GF viruses had no demonstrable viraemia or other sign of infection or illness, except that the sheep infected with AMT developed mild fever lasting less than 24 h. Antibody responses were monitored at intervals over a period of 24 weeks in all sheep by homologous and heterologous HAI, CF and cell culture neutralization (CPENT) tests. Homologous antibody responses were marked in the RVF-infected sheep and their sera cross-reacted strongly in HAI tests with antigens of the other viruses. The sera of the RVF-infected sheep cross-reacted less markedly in CF and CPENT tests. Homologous antibody responses were poor in all the sheep infected with phleboviruses other than RVF, and the cross-reactivity of their sera for RVF antigen or virus was negligible. All sheep were challenged with RVF virus 48 weeks after their initial infection. The sheep which had originally been infected with RVF virus were immune and developed neither fever nor viraemia. All other sheep developed fever, viraemia and antibodies to RVF virus. It was concluded that the African phleboviruses, other than RVF, are unlikely to cause disease in livestock or to induce antibodies which could cause confusion in the diagnosis of RVF.

Characterization of rotaviruses and subgroup F adenoviruses from acute summer gastroenteritis in South Africa.

Six hundred and sixteen specimens were collected from black children hospitalised with acute gastroenteritis during the summer and autumn of 1982-1983 (October to May). Eighty-five children (13.8%) shed rotavirus and at least 40 (6.5%) shed adenovirus (Ad) type 40 or 41 belonging to subgroup F. The highest monthly prevalence of shedding subgroup F adenoviruses (10.1%) coincided with a peak in admissions in midsummer, whereas the highest monthly prevalence of shedding rotaviruses (41.9%) coincided with a peak in admissions in autumn. There were at least five genome types of rotavirus, at least three genome types of Ad40, and at least five genome types of Ad41 circulating in the Johannesburg-Soweto area during the study period. The high rate of rotavirus shedding in autumn could not be attributed to an upsurge in infections by any particular rotavirus strain.

Specific detection and typing of adenovirus types 40 and 41 in stool specimens by dot-blot hybridization.

A dot-blot hybridization test was developed which allowed the direct detection of fastidious enteric adenovirus DNA in stool specimens from children with diarrhea and simultaneous typing of the viruses as adenovirus type 40 (Ad40) or Ad41. Cloned PstI fragments of Ad40 and Ad41 were used as 32P-labeled probes in the test, which allowed detection of picogram quantities of viral DNA in 20 to 40 microliter of stool suspension. Results were obtained within 48 h. The type specificity of the test was evaluated with 76 specimens known to contain either Ad40 or Ad41 by restriction enzyme analysis. Sixty-one specimens had sufficient DNA to be detected without any removal of protein. Thirty-one adenoviruses were typed as Ad40, and 30 were typed as Ad41, giving 100% correlation with the results of restriction enzyme analysis. The other 15 specimens were detected and typed as Ad40 or Ad41 only after removal of protein by a phenol extraction method. The dot-blot hybridization method is particularly useful for identifying those Ad40 and Ad41 strains which defy all attempts at culture and will be a useful tool in the epidemiology of fastidious enteric adenovirus infections.