J. T. M. Van Der Logt

16S rRNA based polymerase chain reaction compared with culture and serological methods for diagnosis of Mycoplasma pneumoniae infection

The use of a 16S rRNA based polymerase chain reaction (PCR) for the detection ofMycoplasma pneumoniae infection was investigated. Sputum samples from 34 patients with respiratory illness and evidence of pneumonia as judged by chest X-ray were analyzed by PCR and microbiological culture. Throat swabs from 14 healthy individuals were used as controls. For serology, an enzyme immunoassay for the detection of immunoglobulin M antibodies and a complement fixation assay were performed. Evidence ofMycoplasma pneumoniae infection was obtained in ten patients (29 %), eight of whom were found positive by both PCR and serology. Two of the sputum samples from these eight patients were negative by culture. Of the remaining two patients positive forMycoplasma pneumoniae, one was positive by PCR and culture but negative by serology, and one was found positive by serology but negative by PCR and culture. Thirteen of the 14 controls were negative by both PCR and serology. One control, however, was negative by serology but positive by PCR, which was probably due to asymptomatic carriage ofMycoplasma pneumoniae. The results of this study indicate the suitability of the PCR for the detection ofMycoplasma pneumoniae in clinical samples as well as its potential value as an additional tool for the diagnosis of infection.

Prevalence of naturally occurring viral infections, Mycoplasma pulmonis and Clostridium piliforme in laboratory rodents in Western Europe screened from 2000 to 2003

In this report prevalence rates of rodent viruses in laboratory animals are presented based on routine serological screening of mouse and rat colonies from European institutes. The prevalences found during the period 2000–2003 are compared with those reported for 1981–1984 and 1990–1993. It is shown that some infections were eliminated from laboratory animal colonies (e.g. K-virus and polyomavirus) by taking preventative measures whereas other infections such as mouse hepatitis virus and parvoviruses remained at a high rate. Further decreases in prevalence rates in the last 10 years were found for infections such as pneumonia virus of mice, reovirus type 3, Sendai virus, sialodacryoadenitis/rat coronavirus and Mycoplasma pulmonis. The introduction of new detection methods showed that mouse parvovirus and rat parvovirus, both members of the Parvoviridae family, remain a major threat to laboratory mice and rats. Guineapig cytomegalovirus and parainfluenzavirus appeared to be the most prevalent agents among laboratory guineapigs. The importance of a standardized, up-to-date screening programme is discussed.

Enzyme-linked immunosorbent assay for measurement of antibody against cytomegalovirus and rubella virus in a single serum dilution.

Enzyme-linked immunosorbent assays (ELISA) were developed for quantifying cytomegalovirus (CMV) and rubella antibodies using a single serum dilution (1/800) in conjunction with a standard curve. A near linear relation was found between the logarithms of absorbance values of sera at a dilution of 1/800 and the titres as determined by an end point dilution ELISA. The reproducibility of the single dilution ELISA was good; the within-test coefficients of variation averaged 7.5% for CMV antibody and 12.4% for rubella antibody. A close correlation was found between ELISA and complement-fixing (CF) antibody titres to CMV and between ELISA and haemagglutination-inhibition (HI) antibody titres to rubella virus. The titres in ELISA were 200 to 1000 times higher than in CF for CMV and 50 to 100 times higher than in HI for rubella virus.

Post-transfusion non-A, non-B hepatitis in the Netherlands.

tions and hospital admissions (r=09; p<0 01). There was a tendency throughout the period, when the figures for whooping cough increased, for the number of notifications to increase proportionately more than the number of admissions. In the peak quarters of 1974 and 1978 there were 25 and 58 admissions and 159 and 666 notifications respectively. We studied Hospital Activity Analysis statistics for a range of other respiratory and infective diseases in childhood during the period but found no disease other than whooping cough for which admissions corresponded with the quarter by quarter trends in whooping-cough notifications. There were three deaths -one in 1974, one in 1977, and one in 1979-ascribed to whooping cough in the region during the period.

Antibody isotype response after human cytomegalovirus infection

The antibody isotype response to human cytomegalovirus (CMV) was studied in paired sera from patients with primary and recurrent CMV infection. The majority of sera was obtained from immunocompromised patients. In the 48 patients with primary CMV infection, CMV-specific IgM (CMV-IgM) and IgG (CMV-IgG) antibodies were found in all patients, and CMV-specific IgA (CMV-IgA) and IgE (CMV-IgE) antibodies in 46 patients. CMV-IgM, -IgA, and -IgE antibodies were found in, respectively, 21, 31 and 4 of the 53 patients with recurrent CMV infection, and in, respectively, 3, 3 and 1 of the healthy controls. The results indicate that CMV-IgE is a better marker of primary CMV infection than CMV-IgM, and confirm that detection of CMV-IgM and -IgA may also be useful for diagnosis of recurrent CMV infection.

Antibody-capture enzyme-linked immunosorbent assays that use enzyme-labelled antigen for detection of virus-specific immunoglobulin M, A and G in patients with varicella or herpes zoster

Antibody-capture enzyme-linked immunosorbent assays (AC-ELISA) which use enzyme-labelled antigen were developed for detection of varicella-zoster virus-(VZV) specific IgM, IgA and IgG antibody in patients with varicella or herpes zoster and in sera from healthy individuals. All 18 patients with varicella developed a VZV-IgM and a VZV-IgG response, 17 also a VZV-IgA response. In contrast, all 19 patients with herpes zoster were shown to be positive for VZV-IgA whereas only 13 of these reacted positively for VZV-IgM. A VZV-IgM response was detected in only two sera from 100 healthy individuals and an IgA response in only one. The presence of virus-specific IgA and IgG in the cerebrospinal fluid as determined by AC-ELISA was a useful indicator of VZV infection of the central nervous system. By AC-ELISA, VZV-IgG was detected predominantly in sera from patients with acute or recent VZV infection. Only 14 sera from 100 healthy individuals were positive for VZV-IgG by AC-ELISA, whereas all were positive by an indirect ELISA. These results indicate that AC-ELISAs may be useful assays for determination for acute or recurrent VZV infection, but are not suitable for determination of past infection with this virus.

Synthesis of lens protein in vitro. The lens cell-free system.

Abstract 1. Conditions for optimal peptide chain initiation and elongation have been determined in a cell-free system prepared from calf eye lens cells by using [3 5S] Met-tRNAfM et and [3 5S] Met-tRNAM et. 2. This cell-free system is capable of translating heterologous messenger RNA. 3. The lens proteins (crystallins), which comprise the structural matrix for the lens body, have been synthesized in the cell-free system. The amounts of newly synthesized polypeptides in vitro have been examined and compared with the proteins synthesized in organ culture. 4. The cell-free system has been employed to investigate some aspects of crystallin synthesis. We have found that synthesis of one class of protein ( β H - crystallin ) is negligible. It further appeared that differences in rate of synthesis between the polypeptide chains of α-crystallin do exist. The possible implications with respect to lens aging are discussed.

Use of the enzyme-linked immunosorbent assay for the early diagnosis ofMycoplasma pneumoniae infection

IgM and IgG antibodies toMycoplasma pneumoniae were measured in 147 sera from four groups of patients by means of an indirect enzyme-linked immunosorbent assay (ELISA) and the results compared with those obtained by other methods. A good correlation was demonstrated between the complement fixation test and ELISA-IgM and to a lesser extent ELISA-IgG; for the metabolic inhibition test the reverse was the case. The indirect haemagglutination test appeared to detect mainly IgM antibodies. Low levels of IgM antibodies were dectected by ELISA in 60 sera of children not suffering from pneumonia. However, if only high titres (> 800) were regarded as indicative ofMycoplasma pneumoniae infection, a presumptive diagnosis could have been made in 42 of 73 single acute phase sera from patients. Comparable results were obtained with IHA. The diagnostic level of IgM antibodies was reached during the second week of the disease. It is concluded that examination of a single serum sample by either ELISA-IgM or IHA may assist in early diagnosis ofMycoplasma pneumoniae infection.

Mutual viral and bacterial infections after housing rats of various breeders within an experimental unit

Fifteen athymic rat strains from 11 breeding colonies were housed within an experimental facility for an immunological study. Health status records supplied with 14 of the strains listed infections by Kilhams rat virus (KRV), Clostridium piliforme (Bacillus piliformis) and Pasteurella pneumotropica for 2, 2 and 1 colonies respectively. In sera taken previous to the study from euthymic rats of 10 strains, antibodies to KRV were detected in 3 strains, to Pneumonia virus of mice (PVM), Rat corona virus (RCV) and Sendai virus in one strain each and to P. pneumotropica in 2 strains. Only 2 of the KRV infections had been reported by the supplier. During the study rats of all 10 strains developed antibodies to 2-4 of viral antigens. Eight out of 10 rat strains seroconverted to 1-5 of the antigens C. piliforme (B. piliformisl, Bordetella bronchiseptica, Haemophilus spp., P. pneumotropica and Streptobacillus moniliformis. Two rat strains housed in filtertop cages did not develop antibodies to bacterial antigens. The potential detrimental effects of intercurrent infections on the outcome of the comparative immunological study are discussed.

Microbiological effects and quality control in laboratory rodents

Numerous viruses, mycoplasmas, bacteria and parasites have been associated with infectious diseases in laboratory animals. It is clear that pathogenic agents causing overt disease represent a serious hazard to research results in both short- as well as long-term studies. However, these organisms may contaminate colonies without causing any clinical or pathological symptom. This makes research less reliable because of the more subtle effects of the silent infections, especially in long-term studies as in aging research. The establishment of animal colonies that were free from these (micro-) organisms has increased substantially the value of animals used in biomedical research. Characterization of the health status and microbiological monitoring of the animals in experiments are particularly important. This paper reviews many of the major considerations in the efforts to maintain animals free of unwanted organisms, including quality and sources of animals, transportation and quarantine, maintenance during experimentation, microbiological characterization and monitoring of animals and environment. (Aging Clin. Exp. Res. 5: 317–323, 1993)Numerous viruses, mycoplasmas, bacteria and parasites have been associated with infectious diseases in laboratory animals. It is clear that pathogenic agents causing overt disease represent a serious hazard to research results in both short- as well as long-term studies. However, these organisms may contaminate colonies without causing any clinical or pathological symptom. This makes research less reliable because of the more subtle effects of the silent infections, especially in long-term studies as in aging research. The establishment of animal colonies that were free from these (micro-) organisms has increased substantially the value of animals used in biomedical research. Characterization of the health status and microbiological monitoring of the animals in experiments are particularly important. This paper reviews many of the major considerations in the efforts to maintain animals free of unwanted organisms, including quality and sources of animals, transportation and quarantine, maintenance during experimentation, microbiological characterization and monitoring of animals and environment. (Aging Clin. Exp. Res. 5: 317–323, 1993)