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Biomedical subjects

G J Demmler

Publications and source records attributed to G J Demmler.

At least 55 records · Page 3Linked to original sources
PubMed1992

Rapid epidemiologic analysis of cytomegalovirus by using polymerase chain reaction amplification of the L-S junction region.

A technique based on polymerase chain reaction (PCR) amplification was developed to facilitate the study of the epidemiology of cytomegalovirus (CMV). Consensus oligonucleotide primers from repetitive DNA sequences were designed to amplify interspersed repetitive sequences in an area of heterogeneity within the L-S junction region of the CMV genome, and PCR products were detected by gel electrophoresis. Purified CMV DNAs from 25 CMV isolates, 13 from members of five families in which person-to-person transmission was documented, 9 random clinical isolates of CMV, and 3 laboratory reference strains of CMV (Towne, Davis, and AD169), were analyzed. The gel electrophoretic patterns of DNA bands, or PCR profiles, produced by amplification with the L-S primers were unique for epidemiologically unrelated strains and laboratory reference strains, yet similar patterns were observed for epidemiologically related strains isolated from members of the same family. This method of rapid fingerprinting of CMV DNA within the hypervariable L-S junction region by PCR to produce strain-specific, variably sized PCR products should simplify the molecular epidemiologic analysis of CMV.

Base Sequence↗
PubMed1992

Paecilomyces lilacinus catheter-related fungemia in an immunocompromised pediatric patient.

Paecilomyces lilacinus catheter-related fungemia in an immunocompromised child is reported. The presence of a central venous catheter and the patient's immunocompromised status were felt to be predisposing factors for this unusual infection. To our knowledge, this is the first description of P. lilacinus catheter-related fungemia, and our patient may be the youngest reported patient with this mycosis who was cured.

Catheterization, Central Venous↗
PubMed1992

Semiquantitative measurement of cytomegalovirus DNA in lung and heart-lung transplant patients by in vitro DNA amplification.

We report the cases of two lung transplant recipients (one heart-lung and one single lung) who eventually developed cytomegalovirus (CMV) pneumonitis after documentation of increasing CMV DNA titers in sequential bronchoalveolar lavage (BAL) specimens by polymerase chain reaction (PCR) amplification. To our knowledge, this is the first report that semiquantitation of PCR-amplified DNA can detect an increase in CMV DNA titer in BAL specimens prior to the onset of clinical symptoms or detection of infection by traditional techniques in lung transplant patients. The results obtained in these two cases suggest that DNA titer measurement on sequential BAL samples may differentiate latency from active viral replication and, thus, provide an opportunity for clinical intervention before the development of overt clinical symptoms.

Adult↗
PubMed1992

Progressive hearing loss in infants with asymptomatic congenital cytomegalovirus infection.

Congenital cytomegalovirus (CMV) infection is a major public health problem because 30,000 to 40,000 neonates with the infection are born each year in the United States. Although 90% of the congenitally infected infants are asymptomatic at birth, evidence is accumulating that these infants are at risk for audiologic, neurologic, and developmental sequelae. The current study describes the audiologic outcome of 59 infants with asymptomatic congenital CMV infection compared with 26 control infants. Eight of 59 infected infants had congenital sensorineural hearing loss (SNHL) but none of the control subjects did. Longitudinal audiologic assessments revealed that 5 of the 8 infants had further deterioration of their SNHL; a ninth infant with initially normal hearing experienced a unilateral SNHL during the first year of life, with further deterioration subsequently. The frequency of SNHL was similar for infected infants born to mothers with recurrent CMV infections during pregnancy (2 of 9) and for those born to mothers who experienced primary CMV infections (5 of 26). There was a significant difference between the occurrence of hearing loss in infected infants with normal computed tomographic scans (2 of 40) compared with those with either periventricular radiolucencies (4 of 13) or calcifications (1 of 3). Children with SNHL often have no identified cause of the loss; thus, it is likely that many of these children had asymptomatic congenital CMV infection. Given the progressive nature of SNHL associated with asymptomatic congenital CMV infection, longitudinal audiologic assessments are mandatory.

Audiometry↗
PubMed1991

Hepatocellular transplantation in acute hepatic failure and targeting genetic markers to hepatic cells.

Orthotopic liver transplantation (OLT) represents the only therapeutic option for many patients with end-stage liver disease as well as many inborn genetic errors of hepatic metabolism. Despite dramatic progress in methods for OLT, the utilization of this procedure is limited by its considerable morbidity and mortality, by a chronic shortage of organs for transplant, and by difficulty arranging funding for many patients. Many children with fulminant hepatic failure do not receive OLT because this technology is unavailable or unaffordable. Hepatocellular transplantation (HCT), in which isolated, heterologous hepatocytes from a donor liver would be infused into the diseased organ in order to provide essential hepatic functions, could provide a much needed therapeutic alternative to OLT in the treatment of some causes of hepatic insufficiency. Experiments in animals have demonstrated that several genetic deficiencies of hepatic metabolism as well as experimental induced hepatic failure in animals can be reversed by HCT. Despite this experience, HCT has never been attempted in human subjects. This protocol represents the first proposed clinical trial of HCT. We are proposing a clinical trial in which HCT would be attempted as a therapeutic intervention in children with acute hepatic failure who have no other medical or surgical options. This proposal is intended to establish surgical methods for HCT and to evaluate the feasibility of this procedure for treating hepatic disease in humans. It is our expectation that HCT may provide short-term support for patients awaiting organ availability, a "bridge to recovery" allowing patients with fulminant hepatic failure to recover, or a long-term repopulation of the patient's liver with healthy donor cells. One of the major limitations of many animal studies in HCT is that, since the donor hepatocytes are often indistinguishable from those of the host, it has often been difficult to demonstrate a clear correlation between engraftment and the therapeutic effect. In order to verify engraftment independent of the therapeutic response, we propose to "mark" the donor hepatocytes by transducing these cells with a recombinant retroviral vector (LNL6) carrying a marker gene (NEO-R, neomycin phosphoribosyl transferase). The presence of this marker will enhance the ability to identify transplanted cells in the host using assays for the NEO-R gene or transcribed NEO-R mRNA. The LNL6 vector has been approved for human use and has been used as a marker gene for transplanted cells in human subjects without any reported adverse effects. We would like to emphasize that this is a proposal with therapeutic intent.(ABSTRACT TRUNCATED AT 400 WORDS)

Acute Disease↗
PubMed1991

Improved amplification of cytomegalovirus DNA from urine after purification of DNA with glass beads.

Cytomegalovirus can be detected in a variety of specimens including leukocytes, urine, saliva, feces, and various tissues by polymerase chain reaction (PCR) amplification of viral DNA. Although methods for amplification are fairly standard, sample preparation is not well characterized, especially for tissue. Typically, preparation of samples for PCR amplification ranges from simple boiling to phenol/chloroform extraction and quantification before testing. Several reports have described inhibition of the PCR in some samples types. Here we show that reliable detection of cytomegalovirus DNA in urine is obtained only after some degree of DNA purification, presumably because of PCR inhibition by a yet unidentified component present in a few of the urine samples tested. Glass, in the form of fine beads, was used to adsorb DNA such that protein and other substances could be selectively eluted before the recovery of DNA for PCR amplification. Urine samples prepared by this method did not show inhibition, and results correlated well with those by tissue culture for detection of cytomegalovirus.

Acquired Immunodeficiency Syndrome↗
PubMed1990

Detection of mRNA from the immediate early gene of human cytomegalovirus in infected cells by in vitro amplification.

It would be desirable to have a facile means of detecting the presence of viral DNA or mRNA in any given biological sample, especially in view of the growing immunocompromised population in which the diagnosis of viral disease is a common problem. In vitro amplification of DNA using methods such as the polymerase chain reaction, offers a sensitive means of detecting both DNA and mRNA. We have used the polymerase chain reaction to detect DNA and mRNA from human cytomegalovirus infected human fibroblasts. Viral mRNA was differentiated from DNA using primers which flank a splice junction, resulting in a smaller product for the mRNA template. A cDNA was prepared from total RNA using a primer specific for the gene of interest, in this case the major immediate early transcript of human cytomegalovirus. The cDNA was then amplified using a modified polymerase chain reaction protocol. mRNA from the major immediate early gene was detected in cultured fibroblasts as early as 6 h after infection, and continued to be expressed for at least 96 h post infection. Sensitive and facile detection of viral mRNA should facilitate diagnostic and basic studies of viral pathogenesis.

Amino Acid Sequence↗
PubMed1989

Prevalence of cytomegalovirus antibody in subjects between the ages of 6 and 22 years.

We determined the prevalence of antibody to cytomegalovirus (CMV) in three groups between 1985 and 1987. Group I consisted of 511 subjects 6-22 y old, group II consisted of 920 subjects 18-21 y old, and group III of 113 subjects 18-22 y old. The overall prevalence of antibody in these three groups was 34%, 24%, and 28%, respectively. Prevalence of antibody in white subjects (24%, 21%, and 24%, respectively) was significantly lower than that in nonwhite subjects. In group I, there was no increase in prevalence with age in white subjects, but the percentage of individuals with antibody increased with age among nonwhite subjects. It is of obvious concern that a large proportion of white women entering childbearing years lack CMV antibody.

Adolescent↗
PubMed1988

Factors associated with cytomegalovirus excretion in hospitalized children.

Hospital employees, especially pregnant women, have expressed concern over exposure to patients excreting cytomegalovirus (CMV). However, the vast majority of CMV excreters are asymptomatic and are not identified while hospitalized. As part of a 2-year, longitudinal study of nosocomial transmission of CMV in a children's hospital, the prevalence rates of CMV excretion by patients in different areas of the hospital were determined. The average prevalence rates were neonatal intensive care unit, 5.7%; intermediate care nurseries, 4%; premature nursery, 6.3%; normal newborn nursery, 3%; hematology-oncology, 3.7%; pediatric intensive care unit, 3.6%; general pediatric ward, 6%; and a chronic care unit, 16%. During this 2-year study, 315 patients admitted to the chronic care unit were investigated to determine which clinical factors might help predict children likely to excrete CMV. With the use of multiple logistic regression analysis of variables, prior excretion of CMV, multiple hospital admissions, female sex, and Hispanic ethnic background were correlated with excretion. Although not identified as significant factors, a history of premature, bronchopulmonary dysplasia and positive results of CMV serologic studies were more commonly associated with excretion. CMV excretion occurred in all areas within a children's hospital, and certain factors were associated with excretion, but it is unlikely that this information could adequately identify "safe" patients or safe areas within a hospital. This study reinforces the need to follow proper precautions with all patients.

Adult↗
PubMed1988

Detection of cytomegalovirus in urine from newborns by using polymerase chain reaction DNA amplification.

Polymerase chain reaction (PCR) amplification was used to detect cytomegalovirus (CMV) in tissue culture and in urine specimens from newborns. Synthetic oligonucleotide primer pairs were used to amplify DNA from the major immediate-early and the late antigen genes of CMV. Amplified products were detected by gel electrophoresis and by dot-blot hybridization with oligonucleotide probes. Using one or both of the primer pairs and associated probes, we found 46 different tissue culture isolates of CMV that were positive; no reaction products were detected when the same primers and probes were used to amplify other herpes family viruses or human genomic DNA. Urine samples from 44 congenitally infected infants were positive when tested with one or both primer pairs and probes. When compared with tissue culture, detection by gel electrophoresis provided a sensitivity of 93%, a specificity of 100%, and a predictive value of a positive result of 100%. Dot-blot analysis raised the sensitivity to 100%. We conclude that PCR amplification may be a valuable tool for diagnosing congenital CMV infection.

Blotting, Southern↗
PubMed1988

DNA hybridization assay for congenital cytomegalovirus infection.

An in vitro DNA hybridization assay was used to test 281 newborns for congenital infection with cytomegalovirus. The assay utilized an abbreviated method for DNA preparation and a dot blot assay that provided good sensitivity (100%) and specificity (98.9%) when compared with standard tissue culture, yet substantially reduced the total time of analysis. This assay would be a useful adjunct to tissue culture to diagnose newborns with congenital infection with cytomegalovirus.

Cytomegalovirus Infections↗
PubMed1987

Method of blood processing affects the prevalence of cytomegalovirus excretion in newborn nurseries.

The potential risk of exposure by hospital personnel and noninfected patients to patients excreting the cytomegalovirus (CMV) may be increased in areas of high prevalence of CMV excretion, such as the neonatal nursery. These personnel are particularly sensitized to the adverse effects of intrauterine CMV infection because they frequently care for the severely affected congenitally infected infants. Multiple transfusions with conventionally processed blood have been associated with high excretion rates in sick preterm neonates. During a 4-year period, four separate methods of blood preparation were used. Two of these methods (CMV-seronegative blood and frozen deglycerolized red blood cells) have been previously shown to be effective in reducing posttransfusion CMV infection. We were able to show that excretion rates in neonates were significantly lower when CMV-seronegative blood and frozen deglycerolized red blood cells were used than when conventionally processed blood and saline-washed red blood cells were used. In areas where posttransfusion CMV infection has been identified as a problem, the increased expense of specially processed blood can be justified, not only to reduce transfusion-associated disease but also to minimize exposure of the hospital staff and noninfected patients to patients excreting CMV.

Cross Infection↗
PubMed1987

Nosocomial cytomegalovirus infections within two hospitals caring for infants and children.

Using serology, virology, and molecular epidemiology, we investigated nosocomial transmission of cytomegalovirus (CMV) over a two-year period in two contrasting environments: a crowded, busy pediatric chronic care unit (337 patients, 43 nurses, and 76 therapists; average prevalence of CMV excretion in patients, 16%) and a small neonatal unit (293 patients and 69 nurses; average prevalence, 0.7%). In the chronic care unit no nurse or therapist acquired CMV, but two pairs of infants were infected with homologous strains of CMV, and patient-to-patient transmission was proven in one pair. In the neonatal unit no patients acquired CMV in the hospital, but two nurses seroconverted, with a nonoccupational source proven for one. Transmission from CMV-infected caretaker to patient did not occur in either environment. CMV was isolated from diapers as well as hands of patients and personnel but not from other environmental surfaces.

Adult↗