Human cytomegalovirus viraemia in HIV-1-seropositive patients at various clinical stages of infection.

Eighty-two HIV-1-seropositive subjects were examined for the presence and quantification of human cytomegalovirus (HCMV) in peripheral blood polymorphonuclear leukocytes (PMNL) by polymerase chain reaction, culture and immunofluorescence in order to investigate the relationship between viraemia and immunosuppression. Patients were divided into three groups: (1) asymptomatic subjects with greater than 400 x 10(6)/l CD4 lymphocytes (n = 30); (2) asymptomatic subjects with less than 400 x 10(6)/l of CD4 lymphocytes and zidovudine (n = 20), and (3) AIDS-related complex (ARC)/AIDS patients on zidovudine (n = 32). Evidence of HCMV infection in circulating PMNL was found in 15 out of 29 ARC/AIDS patients examined (51.7%), whereas no infection was detected among the 50 asymptomatic HIV-1-seropositive subjects. HCMV-related symptoms were found only where the number of infected PMNL was greater than 50 per 2 x 10(5) cells.

Rapid detection of human rotavirus strains in stools by single-sandwich enzyme-linked immunosorbent assay systems using monoclonal antibodies.

Using murine monoclonal antibodies (MAbs) raised against the common antigen of group A rotavirus (RV), two single-sandwich ELISA systems were developed for detection of RV in stools: one using polyclonal antibody (PAb) as capture and a MAb as detector antibody (referred to as PAb-MAb assay); and the other based on the use of two different MAbs as capture and detector antibodies (referred to as MAb-MAb assay). In each single-sandwich ELISA system, samples and peroxidase-labeled MAb were incubated sequentially (two-step method) or simultaneously (one-step method). Using the two-step procedure on purified RV, 50 pg of protein was detected in the PAb-MAb as well as in the MAb-MAb assay, whereas the one-step method detected 0.4 ng and a conventional double-sandwich ELISA detected 3.2 ng of viral protein. Titration of RV samples from stools and cell cultures showed that single-sandwich ELISA titers were, on the average, 10-100-fold higher than those obtained by electron microscopy (EM), but 10-100-fold lower than those obtained by solid-phase immune EM (SPIEM). However, when 200 stool samples previously examined by EM or SPIEM were tested by the single-sandwich ELISA systems, specificity and sensitivity of these assays were 100%, and comparable to SPIEM. No false positive results were obtained when 54 samples of meconium and 91 stools from newborns in the first five days of life were tested. The two-step procedure appeared to be somewhat preferable over the one-step method, which, although faster, gave a marked prozone with a few samples in the MAb-MAb assay. The use of MAbs in rapid single-sandwich ELISA systems for RV detection in stools appears highly convenient, due to reliable results and short test performance times.

[Follow-up and long-term treatment of infections caused by human cytomegalovirus in 3 patients with AIDS]. / Monitoraggio e trattamento a lungo termine delle infezioni da cytomegalovirus umano in tre pazienti affetti da AIDS.

We report the follow-up and treatment of HCMV infections in three patients with AIDS. The patients, affected by HCMV retinitis, have been followed 24, 12 and 6 months respectively. The antiviral treatment was based on the DHPG administration which was substituted in one case of resistance to DHPG with Foscarnet. In the follow-up period, virological tests have been performed to detect the presence of the HCMV antigenemia/viremia. The results show that, to avoid the progression of the retinitis, the antiviral treatment must not be stopped or discontinued. DHPG and Foscarnet were able to limit the infection to the eye and were well tolerated. In the HCMV-infected patients, the continuous monitoring of the antigenemia/viremia is of main importance to follow the clinical and therapeutical course of the disease.

Early virus isolation, early structural antigen detection and DNA amplification by the polymerase chain reaction in polymorphonuclear leukocytes from AIDS patients with human cytomegalovirus viraemia.

Fifty AIDS patients were investigated for human cytomegalovirus (HCMV) viraemia when potentially HCMV-related clinical symptoms or syndromes were observed. Nine patients underwent prolonged virologic follow-up, while 41 additional patients were examined only once or sporadically. Concentrated preparations of polymorphonuclear leukocytes (PMNL) from 153 blood samples were obtained for monitoring: (1) early virus isolation in cell cultures 24 h p.i. (viraemia); (2) early structural antigen detection in cytospin preparations (antigenemia); and (3) HCMV DNA in blood (DNAemia) through DNA amplification by the polymerase chain reaction (PCR). Viraemia and antigenemia were quantitated, whereas evaluation of DNAemia was only qualitative. A good correlation between levels of viraemia and antigenemia was consistently found except during ganciclovir treatment. HCMV-related clinical symptoms were observed when the number of infected PMNL was greater than 100 per 2 x 10(5) cells examined. All 56 blood samples positive for viraemia and antigenemia were also PCR-positive, whereas 44 samples (39 of which taken from patients with ascertained HCMV infection in blood) were positive by PCR only. Viraemia and antigenemia were often unrelated to HCMV organ syndromes, such as retinitis, in which only DNAemia was often detected. Prolonged ganciclovir treatment kept viraemia, antigenemia and even DNAemia at a low or negative level, yet drug discontinuation led to rapid progression of HCMV infection in blood. In addition, prolonged antiviral treatment could induce appearance of ganciclovir-resistant HCMV strains, requiring alternative foscarnet therapy. In conclusion, determination of viraemia and antigenemia appears essential for correct clinical management and antiviral treatment of disseminated HCMV infections in AIDS patients. However, PCR is the most sensitive method for diagnosis and monitoring of HCMV infections in blood at a pre-clinical stage.

Identification of two subtypes of serotype 4 human rotavirus by using VP7-specific neutralizing monoclonal antibodies.

Two distinct subtypes of human rotavirus serotype 4 were identified by using neutralizing monoclonal antibodies directed to the major outer capsid glycoprotein, VP7, of strains ST3 (subtype 4A) and VA70 (subtype 4B). Specimens containing serotype 4 rotavirus, obtained from different countries, were examined for subtyping by using solid-phase immune electron microscopy, enzyme-linked immunosorbent assay, and, for cell culture-adapted strains, neutralization assay. All 59 human rotavirus strains identified as serotype 4 by using animal antisera were classified into either subtype by monoclonal antibodies. This suggests that the antigenic difference between the two subtypes is a consequence of critical variations within the immunodominant serotype 4-specific neutralization site of rotavirus VP7. Subtype 4A (ST3-like) strains were predominant and were detected in stools from patients with gastroenteritis, as well as from healthy infants and young children.

Electropherotypes, subgroups and serotypes of human rotavirus strains causing gastroenteritis in infants and young children in Palermo, Italy, from 1985 to 1989.

During 1985-89, an epidemiological survey was conducted in Palermo, Sicily (Southern Italy) on group A human rotavirus (HRV) strains which cause gastroenteritis in infants and young children. Two hundred and thirty eight HRV strains were characterized for subgroup and serotype using monoclonal-antibody-based ELISA systems, and for electropherotype using polyacrylamide gel electrophoresis. Subgroup II strains were largely predominant, constituting 218/238 of the positive stool samples (91.6%). Among the serotypes, 192/238 strains (80.7%) were serotype 1 and 16 strains (6.7%) were serotype 4; serotype 2 circulated intermittently and serotype 3 was nearly absent (only one subgroup I strain was detected). Two electropherotypes, bbba and cbba, accounted for the largest proportion of the 345 HRV strains examined, 74 (21.4%) and 222 (64.3%) strains, respectively. Unexpected combinations of subgroup, serotype and electropherotype were detected in 5 subgroup I strains, of which 4 possessed a "long" RNA pattern (1 serotype 3 and 3 serotype 4 strains) and one a "short" RNA pattern (a serotype 4 strain). In addition, 4 group C HRV strains (atypical HRV or pararotaviruses) were detected on the basis of electropherotype. These findings emphasize the need for continuous surveillance of HRV infections in different geographic areas of the world in order to detect the appearance of new strains early and to adopt adequate strategies for vaccine preparation and administration.

Nosocomial outbreak of infant rotavirus diarrhea due to the appearance of a new serotype 4 strain.

An outbreak of acute gastroenteritis, involving 30 infants and young children aged 2 months to 4 years, took place in a pediatric ward of the University Hospital of Pavia, Northern Italy, in the period from November 9 to December 1, 1986. Out of the 14 patients examined, ten were found to shed rotavirus with stools. All strains were characterized for serotype, using a monoclonal antibody-based enzyme-linked immunosorbent assay, and for electropherotype, by polyacrylamide gel electrophoresis of genomic RNA. It was shown that a single serotype 4 (subtype 4A) strain spread within the ward from a primary case to seven other patients. The remaining two patients were found to be infected by a serotype 1 strain that was circulating in the same area prior to the outbreak. The clinical symptoms were unusually severe, since significant dehydration was observed in four of the eight serotype 4 rotavirus-infected children. Previous epidemiological studies had shown that since 1983 serotype 4 strains had not been circulating in Pavia, and the electropherotype of the newly circulating serotype 4 strain was different from those observed in 1981-1983. Thus, the severity of the diarrheal disease appeared to be related to the circulation of both a new serotype and a new electropherotype.

Detection of human cytomegalovirus immediate early antigen in leukocytes as a marker of viremia in immunocompromised patients.

Peripheral blood polymorphonuclear (PMN) cells from 35 immunocompromised patients (22 heart transplant recipients and 13 AIDS patients) and four normal subjects were tested for the presence of human cytomegalovirus (HCMV) immediate early antigen (IEA) (antigenemia) by indirect immunofluorescence (IFA) and IEA-specific monoclonal antibodies (MAb). PMN samples were tested in parallel for HCMV isolation (viremia) by using MAb to viral early antigens (EA) and the IFA technique 24-48 hr after inoculation onto human fibroblast monolayers. HCMV was isolated from 26 of 83 PMN samples examined: of these, 25 were also positive for HCMV IEA (96% sensitivity). Seven additional PMN samples negative for viral isolation resulted IEA-positive (87.7% specificity). Six of the seven discordant samples were taken from four patients during ganciclovir treatment. The transitory dissociation between positive HCMV antigenemia and negative viremia during antiviral treatment was followed, at the end of the therapy, either by virus clearance and disappearance of IEA-positive PMNs (one patient) or by reappearance of viremia (three patients). Among concordant positive samples, a significant correlation was observed between the number of IEA-positive PMN leukocytes and EA-positive nuclei of infected fibroblasts, when the same number of PMNs were used for both tests.

Quantification of human cytomegalovirus viremia by using monoclonal antibodies to different viral proteins.

Human cytomegalovirus (HCMV) viremia in peripheral blood polymorphonuclear leukocytes (PMNLs) from 187 immunosuppressed patients (79 heart transplant recipients and 108 patients with acquired immunodeficiency syndrome [AIDS]) was investigated. Five mouse monoclonal antibodies (MAbs), one specifically reactive to HCMV immediate-early antigen (IEA) in PMNLs, two specifically reactive to IEA in infected cell cultures, and two specifically reactive to late antigens, were used in immunofluorescence and/or immunoperoxidase test systems for (i) detection of HCMV IEA in human embryonic lung fibroblast (HELF) cell cultures inoculated with PMNL samples, (ii) direct detection of HCMV IEA in PMNL nuclei of cytospin preparations, and (iii) identification of HCMV isolates from PMNL samples in HELF cells. Quantification of viremia was achieved by counting the number of infected PMNLs per 2 x 10(5) cells examined directly on cytospin preparations as well as by counting the number of IEA-positive HELF cells inoculated with 2 x 10(5) PMNLs. A significant correlation was found between the number of infected PMNLs and the number of infected HELF cells. When the number of infected PMNLs per 2 x 10(5) cells was greater than 10, both methods (PMNL IEA and HELF IEA) gave concordant results; when it was greater than 80/2 x 10(5) cells, clinical symptoms were consistently associated with HCMV viremia. Ten patients with heart transplants and three patients with AIDS who had high or increasing levels of HCMV viremia underwent antiviral treatment with ganciclovir. Treatment was discontinued only after disappearance of IEA-positive PMNLs from blood (the last marker of infection to become negative). On the other hand, in the presence of low levels of viremia (less than 10 infected PMNLs per 2 x 10(5) cells), different methods often provided discordant results and overt clinical symptoms were never observed. IEA-negative results with PMNL samples or HELF cells in the presence of positive virus isolation could never be attributed to the inability of IEA MAbs to recognize individual HCMV strains, since all of the relevant viral isolates were recognized by IEA MAbs. In addition, all five MAbs used in the study were capable of identifying all 135 conventional HCMV isolates obtained from the study population. It was concluded that (i) maximal sensitivity in diagnosing HCMV viremia may be achieved by combining different techniques; (ii) direct detection of HCMV IEA in PMNLs, yielding results on the day of blood collection, is a very rapid and sensitive technique, and thus one can rely on it for clinical management of HCMV infections; and (iii) low levels of viremia are clinically irrelevant, whereas high levels are associated with clinical symptoms.

Group- and type-specific serologic response in infants and children with primary rotavirus infections and gastroenteritis caused by a strain of known serotype.

Antibody response to group A rotavirus (RV), investigated in paired sera from 72 infants and young children with acute gastroenteritis caused by an RV infection, was diagnosed on the basis of a fourfold or greater rise in group A common RV IgG antibody titer. Virus-specific IgM was detected in sera from 64 patients showing seroconversion; these were considered primary infection. RV was detected in stools of 56 (77.8%) patients with serologic evidence of infection and 54 were considered primary infection isolates: 39, serotype 1; 11, serotype 4; and 2, serotype 2. Two could not be typed. Neutralizing antibody studies showed that in primary infections serotype 1 induced an antibody response to serotype 4 at least fourfold lower than the homotypic response; serotype 2 elicited antibody titers to serotypes 1 and 4 at least fourfold lower than homotypic titer; and serotype 4 infections produced a response to serotype 1 as high as the homotypic response. Of 12 patients with primary infection, virus was not typed in 2 or detected in 10; however, the infecting serotype was identified on the basis of distinct patterns of homotypic and heterotypic antibody response.

Comparative evaluation of a commercial enzyme-linked immunoassay and solid-phase immune electron microscopy for rotavirus detection in stool specimens.

Using solid-phase immune electron microscopy (SPIEM) as a reference test, we examined 151 stool specimens from infants and young children with acute gastroenteritis for rotavirus detection by a one-step commercial enzyme-linked immunosorbent assay (ELISA) with labeled monoclonal antibody. Of the 83 samples determined to be positive for rotavirus by SPIEM, 82 were detected as positive by the monoclonal antibody ELISA (sensitivity, 98.7%), while 67 of the 68 specimens determined to be negative by SPIEM were correctly detected as negative by the ELISA (specificity, 98.5%). The diagnostic accuracy of the ELISA kit was 98.6%. Thus, the one-step monoclonal antibody ELISA, which can be completed in less than 90 min, appears to be highly suitable for the rapid and reliable detection of rotavirus in stools.

Nosocomial outbreak of severe Pseudomonas aeruginosa infections in haematological patients.

From June to September 1988, an outbreak of Pseudomonas aeruginosa infections in neutropenic patients admitted to the Haematological Wards of "Ospedali Riuniti" in Bergamo, Italy, was detected. Out of 11 cases of P. aeruginosa infections, 8 were bacteremic. Of these, 7 died within few days of onset (mortality rate: 87.5%). Consequently, possible sources of infection were investigated, and moist areas of the hospital environment were shown to be highly contaminated by P. aeruginosa. A clinical and microbiological follow-up of patients admitted to the Haematological Wards was performed for a 10 month period following the outbreak. Adequate measures for cleaning and disinfection were shown to reduce the frequency of P. aeruginosa hospital infections.

A Family Case Report of Schistosoma haematobium Infection in Italian Travelers.

Schistosoma haematobium infection is endemic in 53 countries and is confined to Africa and the Middle East.1,2,3 It is estimated that about 90 million persons are infected and that at least 180 million persons are exposed to the risk of infection.1 Fortunately, the morbidity caused by S. haematobium infection is probably low, even though it is still difficult to define and evaluate this morbidity with any precision. S. haematobium infection is not a significant cause of death in most endemic areas.1,2 In fact, this particular parasitic infection is mild and often without symptoms. In some cases, the only clinically relevant symptom is recurrent, painless hematuria.2 Among people living in endemic areas, almost all children are infected by the parasite and, in some cases, complications develop, such as chronic renal infection, bladder abnormalities, and carcinoma.2 The seriousness of the disease is related to the rate of infection, since the active disease is more frequently detected among children aged 5-15 years.1,3,4 In developing countries, strategies for the control and prevention of the increasing diffusion of the S. haematobium infection are mainly aimed at the following: (1) education of the population to avoid contamination of fresh water with urine, possibly containing viable eggs of the parasite; (2) education to avoid bathing in or contact with, contaminated water; (3) control of irrigation systems and snails; and (4) use of noninvasive diagnostic techniques and treatment of infected subjects.2,3 In industrialized countries, S. haematobium infection is a rare, imported disease. In most cases, it exhibits only mild symptoms related to the urinary tract. The disease is often frequently unsuspected and misdiagnosed. In fact, S. haematobium infection is usually suspected only in patients immigrating from endemic areas and suffering from urinary tract symptoms, whereas correct diagnosis is often delayed in most cases of infected patients returning from a visit to an endemic area with similar symptoms. In addition, only a few laboratories can perform the correct diagnostic procedures. One such procedure is urine filtration, which can detect the presence and determine the number of S. haematobium eggs in urine samples obtained from infected subjects. In Italy, the number of reported cases of S. haematobium infection has increased in recent years. The increase is due to the increasing presence of immigrants from endemic African countries (unpublished data). The aim of this work was to describe the case report of an Italian family, which was infected by S. haematobium while vacationing in Malawi, and to emphasize that, in addition to immigrants from endemic areas, this parasitic infection should be suspected in patients who travel to these areas and who return, suffering from urinary tract symptoms.

Development and clinical significance of a diagnostic assay based on the polymerase chain reaction for detection of human cytomegalovirus DNA in blood samples from immunocompromised patients.

The presence of human cytomegalovirus (HCMV) DNA in blood was investigated by the polymerase chain reaction (PCR) in 293 blood samples from 86 immunocompromised patients. Of the 86 patients, 23 underwent clinical and virologic follow-up for HCMV infection. In parallel, blood samples were examined for viremia and antigenemia. Concordant results between PCR and assays for viremia and antigenemia were obtained on 124 positive and 110 negative samples, with an overall concordance of 79.8%, while 59 samples (most from patients with HCMV infection) were positive by PCR alone. PCR is a new powerful tool for detection of HCMV infections in blood samples from immunocompromised patients. However, its clinical significance appears to be restricted to the indication of a risk of reactivation of HCMV infection.

Epidemiological characteristics of Pseudomonas aeruginosa strains causing infection in an Italian general hospital. A one-year surveillance.

During the 1989 calendar year, P. aeruginosa caused clinical infections in 0.46% of patients admitted to Ospedali Riuniti (a general hospital), Bergamo, Italy. Strains (n = 267) of P. aeruginosa were collected during this period, and epidemiological characteristics were studied. The mean prevalence of P. aeruginosa infection in inpatients was 1.1% (range 0.06-7.3), whereas outpatients showed a significantly lower prevalence of infection (0.05%). Strains were recovered from inpatients of surgical wards (n = 126; 47.2%), and outpatients (n = 15; 5.6%). Males were more often affected than females (2.7:1). Infection of the urinary tract was the most common (34.1%). Pseudomonas aeruginosa was also involved in lower respiratory tract infections (18.7%) and septicaemia (17.6%). Four typing methods were performed, i.e. serotyping, antibiotyping, pyocin typing, and restriction endonuclease analysis (REA). Serotypes O:11 and O:6 were endemic in the hospital. Some serotypes correlated with specific clinical wards. Pyocin typing was an unreliable epidemiological tool. However, antibiotyping showed the presence of some epidemic clusters, probably related to the antibiotic consumption of the patients. REA suggested the circulation of edemic P. aeruginosa strains in both the obstetrics and neurosurgery wards.

Isolation and characterization of two distinct human rotavirus strains with G6 specificity.

Two new human rotavirus (HRV) strains, PA151 and PA169, with subgroup I specificity and a long RNA pattern, yet with a serotype G (VP7) specificity different from those of any of the six well-established HRV serotypes (G1 to G4, G8, and G9), were isolated 3 months apart from two children with acute gastroenteritis in Sicily, southern Italy, in the winter season of 1987 and 1988. The HRV isolates were adapted to growth in cell cultures and were then characterized by neutralization and RNA-RNA (Northern blot) hybridization. Cross-neutralization studies with type-specific immune sera to RV serotypes 1 to 10 showed the antigenic relatedness of the two strains with serotype 6 bovine strains UK and NCDV. Monoclonal antibodies to VP7 of UK were able to recognize UK and NCDV strains as well as both HRV isolates. Cross-hybridization studies showed a genetic relatedness of PA151 and PA169 to bovine strains for all genes except gene 4. Gene 4 of PA151 appeared to be genetically related to that of AU228 (a human strain of subgroup I and with serotype G3 specificity that belongs to a feline genogroup), whereas gene 4 of PA169 appeared to be unique, yet it was related to gene 4 of two recently reported subgroup I HRV strains, one (PA710) with serotype G3 specificity and the other (HAL1271) with serotype G8 specificity. The new HRV strains must be taken into consideration when deciding strategies for the development of an effective RV vaccine.

Human cytomegalovirus infection of the major leukocyte subpopulations and evidence for initial viral replication in polymorphonuclear leukocytes from viremic patients.

Fourteen immunocompromised patients were examined for viremia, pp65 and p72 antigenemia, and presence of viral DNA in leukocyte fractions of polymorphonuclear leukocytes (PMNL), monocytes/macrophages (M/M), and B and T lymphocytes after purification by fluorescence-activated cell sorting. Nearly all PMNL and M/M fractions were positive for DNA and pp65 antigenemia, while p72 antigenemia was detected in 73% and 62%, respectively. The virus isolation rate was 45% from PMNL and 17% from M/M. T lymphocytes were positive for DNA in 50% of cases and for pp65 and p72 antigenemia in only 11%, while B lymphocytes were DNA-positive in 43% of samples and consistently negative for antigenemia; neither T nor B lymphocytes had virus isolated. Immediate-early (IE)1 RNA was present in 23 (85.2%) of 27 dextran-enriched DNA-positive p72-positive PMNL samples and, in sequential PMNL samples from two heart-transplanted patients, was detected during peak infection in association with p72. Thus, PMNL and M/M are the subpopulations primarily involved in HCMV infection; PMNL may undergo IE replicative events and are not merely passive carriers of phagocytized viral material.

Comparative sensitivities of solid-phase immune electron microscopy and enzyme-linked immunosorbent assay for serotyping of human rotavirus strains with neutralizing monoclonal antibodies.

Suspensions of 24 rotavirus strains, 6 for each known human rotavirus serotype, were serially diluted and titrated by (i) enzyme-linked immunosorbent assay (ELISA) for rotavirus detection, using monoclonal antibodies (MAbs) specific for group-specific sites of the VP6 inner capsid protein; (ii) ELISA for subgrouping, using MAbs reactive with subgroup-specific determinants of rotavirus VP6; (iii) ELISA for serotyping, using MAbs directed to serotype-specific sites of the VP7 outer capsid glycoprotein; and (iv) solid-phase immune electron microscopy (SPIEM) for serotyping, using VP7-specific MAbs. In addition, in each preparation the proportion of double-shelled rotavirus particles were determined by direct electron microscopy. Results showed that SPIEM was 2- to 16-fold more sensitive than ELISA for serotyping of rotavirus. The titers in VP7-specific tests correlated well with the proportion of double-shelled virus particles in each of the samples. Titers obtained by ELISA for serotyping of suspensions containing 20% or fewer complete particles were up to 4,096-fold lower than those obtained by ELISA for detection. ELISA serotyping titers of samples containing 20 to 80% double-shelled rotavirus particles were up to 128-fold lower than ELISA detection titers, whereas preparations with nearly 100% complete particles had ELISA titers that were less different from each other. ELISA subgrouping titers were four- to eightfold lower than corresponding rotavirus detection titers. It was concluded that, although SPIEM appears to be more sensitive than ELISA, the amount of complete virus particles in the specimens is of critical importance for successful serotyping of human rotavirus strains. Samples rich in single-shelled particles but containing low amounts of VP7 outer capsid glycoprotein might even be strongly reactive in assays for rotavirus detection and subgrouping but virtually unreactive in tests for serotyping.

Identification of human cytomegalovirus isolates by the polymerase chain reaction.

Fifty human cytomegalovirus (HCMV) isolates were recovered from different clinical specimens (buffy coat, throat washing and urine) obtained from fifty patients (23 AIDS patients, 20 heart transplant recipients, 1 bone marrow transplant recipient, 2 newborns with congenital HCMV infection and 4 immunocompetent individuals with acute HCMV infection). The isolates were previously identified by immunological methods and then examined for identification by the polymerase chain reaction. In parallel, reference HCMV strains as well as other human members of the Herpesviridae family (reference and wild strains) were examined as controls. Two pairs of primers relevant to the immediate-early and late regions of HCMV DNA, respectively, were used. The DNA amplification product was highly specific; in addition, all fifty HCMV isolates were amplified by both pairs of primers and thus identified as HCMV. These preliminary results show that selected pairs of primers are able to amplify DNA regions conserved enough to allow virus identification among a large number of HCMV strains. In addition, they are highly promising in view of the use of PCR for direct detection of HCMV DNA in clinical samples.

Characterization of rotavirus subgroup-specific monoclonal antibodies and use in single-sandwich ELISA systems for rapid subgrouping of human strains.

Two subgroup-specific monoclonal antibodies (MAb) raised in mice against group A human rotavirus were shown to react by immunoblotting with the trimeric form of VP6 of the homologous subgroup and successfully applied to development of new single-sandwich ELISA systems for rapid subgrouping of human strains. All of the 344 strains tested could be subgrouped, but for two of them prior propagation in cell cultures was required.