In recent years there have been major advances in the field of rapid diagnosis of human cytomegalovirus (HCMV) infections. p72-Specific monoclonal antibodies (mAbs) have led to early identification of HCMV isolates from different clinical samples, particularly peripheral blood leukocytes (PBL). Determination and quantification of viraemia has proven to be particularly useful in the identification of immunocompromized patients with, or at risk for, HCMV disease. The antigenaemia assay represents a major breakthrough, based upon detection of the HCMV lower matrix phosphoprotein (pp65) in nuclei of PBL. Quantitative antigenaemia has been found to be an extremely useful test for diagnosing symptomatic HCMV infection as well as for monitoring the effects of specific antiviral treatment. Recently, HCMV-infected endothelial giant cells have been reported to circulate during disseminated HCMV infections in patients with overt HCMV organ syndromes. In addition, pp65-positive polymorphonuclear leukocytes have been detected in the cerebrospinal fluid of AIDS patients with polyradiculoneuropathy and encephalitis, providing an additional tool for rapid diagnosis of HCMV infections of the nervous system. Finally, DNA amplification by the polymerase chain reaction (PCR) has been extensively used for the detection of HCMV DNA and RNA in clinical samples, and particularly in blood. However, qualitative PCR does not provide clinically useful information, except when DNA is detected in aqueous humor or in cerebrospinal fluid. Quantitative PCR is of greater diagnostic relevance.
One hundred twenty-four patients underwent heart transplantation over a 3-year period. All patients were monitored for human cytomegalovirus (CMV) infection if at risk for primary CMV infection or in the presence of CMV-related symptoms. Rapid diagnosis of CMV infection relied on virus isolation and identification or viral antigen detection by using monoclonal antibodies to CMV immediate early or early antigens. In addition, "in situ" hybridization was used to detect viral DNA in tissue samples. Specimens examined included peripheral blood polymorphonuclear cells for CMV viremia and antigenemia determination, together with the most appropriate clinical samples when organ involvement was suspected. There was a 100% (6/6 patients) incidence of primary CMV infection in seronegative recipients of hearts from seropositive donors, whereas no CMV infection occurred in the three seronegative recipients receiving a transplant heart from CMV-negative donors. CMV hyperimmunoglobulin prophylaxis did not prevent primary CMV infection. Five of the six patients with primary CMV infection were symptomatic. In addition, 15 patients (13%) had symptomatic recurrent CMV infection. The most frequent symptoms associated with CMV infection (either primary or recurrent) were fever (19 patients) and pneumonia (eight patients). CMV viremia was detected in 17 patients either before or concomitantly with the appearance of fever. CMV was isolated from bronchoalveolar lavage in all cases with pneumonia; however, another pathogen was associated with CMV and appeared to be the major cause of pneumonia in 75% of these patients (6/8). Twelve patients (five with primary and seven with recurrent CMV infections) were treated with ganciclovir.(ABSTRACT TRUNCATED AT 250 WORDS)
Background. In the last few years, human cytomegalovirus (HCMV) viremia, pp65 antigenemia, and leuko- and plasma-DNAemia have been developed to quantitate virus in blood of immunocompromised patients with HCMV infection. However, thus far, no conclusive studies have been performed to define the correlation of each of the different assays with clinical symptoms in primary HCMV infections. Methods. This correlation was investigated in a population of 20 heart and heart-lung transplant recipients with primary HCMV infection using standardized virological methods. Results. Median peak HCMV viremia, antigenemia, and leukoDNAemia levels were 110(0-2,000) p72-positive fibroblasts, 450 (27-2,000) pp65-positive leukocytes, and >10,000 (1,358-10,000) genome equivalents (GE) in the 14 symptomatic patients and 18 (1-130) p72-positive fibroblasts, 86.5 (5-350) pp65-positive leukocytes, and 248 (10-863) GE in the six asymptomatic patients, respectively. The difference was statistically significant for antigenemia(P=0.009) and leukoDNAemia (P<0.0001). However, on an individual basis, unlike viremia and antigenemia, all DNA peaks of the 6 asymptomatic patients were below the DNA range of the 14 symptomatic patients(<1,000 GE), while all the 14 symptomatic patients had DNA peaks higher than those of asymptomatic patients (>1,000 GE). Follow-up confirmed these results, showing that 1,000-2,000 GE was the threshold zone for emergence of clinical symptoms. Symptoms were never observed in patients with secondary DNA peaks, except for one patient suffering from an HCMV organ localization(HCMV gastritis). Conclusions. LeukoDNAemia is the viral parameter of choice for monitoring of primary HCMV infections and antiviral treatment in heart and heart-lung transplant recipients. In this patient population, antigenemia-guided preemptive therapy could be replaced by leukoDNAemia-based antiviral therapy.
To test the hypothesis that human cytomegalovirus (CMV) gB genotype may differ with geographic origin or patient demographics, CMV DNA was amplified for gB typing from immunocompromised patients in Italy and Africa and compared with previously reported frequencies in California. Increased gB2 frequency occurred in Italian homosexual AIDS patients, as compared with both Italian heterosexual injection drug users with AIDS and heterosexual Zimbabwe AIDS patients. Occurrence of gB3 in Italy was higher in injection drug users than in homosexual AIDS patients. The incidence of gB4 was higher overall in the Italian as compared with the California patients. Therefore geographic and demographic differences in patients affect gB distribution and should be considered before associations of gB genotypes and virulence are made.
To define diagnostic and prognostic markers of parvovirus B19 (B19V) fetal infection, two groups were investigated: 1) pregnant women with specific symptoms or contacts with symptomatic households (n=37); 2) mothers with pathological ultrasound findings and the relevant fetus at the time of prenatal diagnosis (n=16). In the first group, diagnosis of B19V infection was achieved using IgM detection in 29/37 (78.3%) of patients, while B19V DNA was detected in 36/37 (97.3%) of infected women. In the second group, intrauterine infection was investigated by amniocentesis (n=5), cordocentesis (n=3) or both (n=5). Median B19V DNA load in amniotic fluid was 8.2x107 copies/ml and in fetal blood was 2x109 copies/ml. Maternal blood was positive for B19V DNA (median 3.8x104 copies/ml) in 14/16 (87.5%) women examined. At time of fetal US investigation, all mothers were B19V IgG positive and B19V IgM were detected in 10/16 (62.5%), while fetal B19V IgG and IgM were detected in 1/8 (12.5%) and 5/8 (62.5%), respectively. Phylogenetic analysis revealed that all B19V maternal and fetal strains belonged to genotype 1A. Diagnosis of maternal, fetal and neonatal B19V infection should be based on both IgM and DNA detection. Prognostic markers of congenital B19V infection need to be defined.
To improve our understanding of the natural history of Zika virus (ZIKV) infection in humans, we described the dynamics of ZIKV RNA shedding in different body fluids and antibody responses in patients with acute infection. Twenty-nine adults with travel-associated infection and 1 case of sexual transmission were enrolled and followed up with weekly ZIKV RNA testing in blood, urine, saliva, and semen samples and antibody testing. ZIKV RNA was detected in plasma, urine, and saliva of 57%, 93.1%, and 69.2% of participants, with estimated median times to clearance of 11.5 days (interquartile range [IQR] 6–24 days), 24 days (IQR, 17–34), and 14 days (IQR, 8–31), respectively. In 2 pregnant women, ZIKV RNA persisted in blood until delivery of apparently healthy infants. ZIKV RNA was detected in semen of 5 of 10 tested men; median time to clearance was 25 days (IQR 14–29), and the longest time of shedding in semen was 370 days. In flavivirus-naive patients, the median times to detection of ZIKV nonstructural protein 1 (NS1)–specific immunoglobulin M (IgM) and immunoglobulin G (IgG) antibodies were estimated as 8 days (IQR, 5–15 days) and 17 days (IQR, 12–26 days), respectively. ZIKV NS1 IgM antibodies were undetectable in patients with previous dengue. Prolonged viremia and ZIKV RNA shedding in urine, saliva, and semen occur frequently in patients with acute ZIKV infection. At the time of diagnosis, about half of patients are ZIKV IgM negative. ZIKV NS1 IgM antibodies remain undetectable in patients with previous dengue. Estimates of the times to viral clearance and seroconversion are useful to optimize diagnostic algorithms.
