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Virus isolation or influenza virus antigen detection are the most rapid tests for diagnosis in the acute stage of influenza virus infection. As serology is easier to carry out, the synthesis of serum IgM, IgA and IgG was studied in two well-defined patient groups, infected with influenza B virus (cohort 1, n = 37) and influenza A virus (cohort 2, n = 40), diagnosed by antigen detection and/or virus isolation within 36 h after onset of symptoms. IgM was found in 13 influenza B patients (35%), IgA in 12 patients (32%), whereas a significant antibody rise was found in 33 patients (92%) by enzyme-linked immunosorbent assay (ELISA) and 74% by haemagglutination inhibition assay (HAI). For the influenza A cohort these numbers were respectively 18 (45%), 27 (68%) and 24 (62%) HAI (72%). In age-matched controls, who were bled on the first day of illness of the enrolled patient low prevalence was found for IgA and IgG, for influenza B respectively in 2 and 18%, and for influenza A in 4 and 39%. Studying the kinetics of the antibody response, we found that virus specific IgA and the bulk of IgG is synthesised within the first week of the infection. It is concluded that the finding of a specific serum IgA is highly indicative of an acute influenza infection.

Objectives: To determine which antecedent infections are specifically associated with the Guillain-Barré syndrome (GBS).Background: Infections with many agents have been reported preceding GBS. Some infections are related to specific clinical and immunologic subgroups in GBS. Most agents were reported in case reports and uncontrolled small series of GBS patients only, and their relation to GBS and its subgroups remains unclear.Methods: A serologic study for 16 infectious agents in 154 GBS patients and 154 sex- and age-matched controls with other neurologic diseases. Acute phase, pretreatment samples were used from clinically well-defined GBS patients. The seasonal distribution of serum sampling in the GBS and control group was the same.Results: Multivariate analysis showed that in GBS patients, infections with Campylobacter jejuni (32%), cytomegalovirus (13%), and Epstein-Barr virus (10%) were significantly more frequent than in controls. Mycoplasma pneumoniae infections occurred more often in GBS patients (5%) than in controls in univariate analysis. Infections with Haemophilus influenzae (1%), parainfluenza 1 virus (1%), influenza A virus (1%), influenza B virus (1%), adenovirus (1%), herpes simplex virus (1%), and varicella zoster virus (1%) were also demonstrated in GBS patients, but not more frequently than in controls. C. jejuni infections were associated with antibodies to the gangliosides GM1 and GD1b and with a severe pure motor form of GBS. Cytomegalovirus infections were associated with antibodies to the ganglioside GM2 and with severe motor sensory deficits. Other infections were not related to specific antiganglioside antibodies and neurologic patterns.Conclusions: Recent infections with C. jejuni, cytomegalovirus, Epstein-Barr virus, and M. pneumoniae are specifically related to GBS. The variety of infections may contribute to the clinical and immunologic heterogeneity of GBS.

UNLABELLED: Respiratory syncytial virus (RSV) infections are characterized by upper or lower respiratory tract symptoms including bronchiolitis and pneumonia. Apnoea may be the first sign of disease in children with RSV infection. The aims of this study were the identification of independent risk factors for RSV associated apnoea and the prediction of the risk for mechanical ventilation in children with RSV associated apnoea. Medical records of children younger than 12 months of age admitted with RSV infection between 1992 and 1995 to the Sophia Children's Hospital, were reviewed. Demographic parameters, clinical features and laboratory parameters (SaO2, pCO2 and pH) were obtained upon admission and during hospitalization. Children with and without apnoea were compared using univariate and multivariate logistic and linear regression analysis. One hundred and eighty-five patients with RSV infection were admitted of whom 38 (21%) presented with apnoea. Patients with apnoea were significantly younger, had a significantly lower temperature, higher pCO2 and lower pH and had on chest radiographs also more signs of atelectasis. The number of patients admitted to the ICU because of mechanical ventilation and oxygen administration was significantly higher in children with RSV associated apnoea. Apnoea at admission was a strong predictor for recurrent apnoea. The relative risk for mechanical ventilation increased with the number of episodes of apnoea: 2.4 (95% CI 0.8-6.6) in children with one episode of apnoea (at admission) versus 6.5 (95% CI 3.3-12.9) in children with recurrent episodes of apnoea. CONCLUSIONS: Age below 2 months is the strongest independent risk factor for RSV associated apnoea. Apnoea at admission increases the risk for recurrent apnoea. The risk for mechanical ventilation significantly increases in children who suffer from recurrent apnoea.

One hundred and fifty-three nursing home residents received 0, 5, 25 or 50 mg N-acetylglucosaminyl-N-acetylmuramyl-dipeptide (GMDP) orally, and trivalent influenza subunit vaccine intramuscularly. One day after intervention, there was a strong increase of total leucocytes, monocytes and neutrophils in the groups receiving 25 or 50 mg GMDP. A GMDP dose dependent increase in systemic, but not in local, vaccine side-effects was observed. No significant differences in post-vaccination haemagglutination inhibiting serum antibody titres were observed between the four groups, indicating that oral administration of GMDP together with influenza vaccination, does not lead to a higher vaccine efficacy.

The relationship between clinical severity of respiratory syncytial virus (RSV) infection and distribution of subtype A or B was investigated. The data of 232 children, who were admitted with RSV infection or diagnosed in the outpatient department of the Sophia Children's Hospital, Rotterdam between 1992 and 1995, were studied. The diagnosis of RSV was confirmed by a direct immunofluorescence assay. Subtyping was performed by an indirect immunofluorescence assay using specific monoclonal antibodies. Gender, age at diagnosis, gestational age and birth weight, the presence of underlying diseases, feeding difficulties, the presence of wheezing and retractions, respiratory rate, temperature, clinical diagnosis at presentation, oxygen saturation (SaO2), carbon dioxide tension (PCO2), and pH, characteristics of hospitalisation, and the need for mechanical ventilation were observed. Analysis was performed on data from all patients diagnosed with RSV infection in the period between 1992 and 1995 spanning three RSV seasons, and separately on the RSV season 1993-4. The outcome of the three year analysis (150 (64.7%) subtype A v 82 (35.3%) subtype B) was compared with the outcome of the season 1993-4, a mixed epidemic with 37 (60.7%) subtype A and 24 (39.3%) subtype B isolates. None of the variables observed in the season 1993-4 differed significantly between RSV subtype A and B. Similar results were obtained from the analysis in the period 1992 until 1995, with the exception of PCO2 (a higher PCO2 was found in subtype A, p < 0.001) and retractions (more retractions were noted in patients with subtype A, p = 0.03). After correcting for possible confounders using regression analysis, these differences were not significant anymore. The data indicate that there is no relationship between clinical severity of RSV infection and subtype.

AIM: To determine the value of the polymerase chain reaction (PCR) technique and the analysis of intraocularly produced antibodies by calculating a Goldmann-Witmer quotient (GWq) as diagnostic assays in the confirmation of a clinically diagnosed cytomegalovirus (CMV) retinitis in a group of unselected AIDS patients. METHODS: Eleven samples of undiluted ocular fluid, obtained from nine AIDS patients with a clinically diagnosed CMV retinitis were analysed for the presence of genomic DNA from CMV, HSV-1, VZV, and EBV by PCR. Nine of these samples were analysed for the presence of locally produced IgG antibodies against these herpesviruses by calculating a GWq. Ten samples obtained from patients with various entities of clinical non-herpetic uveitis and 17 samples of aqueous humour obtained at cataract surgery were used as controls. RESULTS: In 10 out of 11 samples from AIDS patients (91%) the presence of CMV DNA was demonstrated. In four out of nine (44%) patients this was accompanied by CMV DNA in the blood indicating a CMV viraemia. In one sample, VZV DNA was detected and in another sample both CMV and VZV DNA were detected. No HSV-1 or EBV DNA could be demonstrated in these 11 samples. In contrast, simultaneous analysis of locally produced IgG antibodies against herpesviruses could not confirm the initial diagnosis of CMV retinitis. Ocular fluid samples obtained from 10 control uveitis patients were negative for DNA from CMV, VZV, and EBV by PCR. In one of 10 uveitis control samples HSV-1 DNA was detected; antibody analysis did not confirm this. In the uveitis control group, a significant GWq was calculated in one sample for HSV-1 and in another sample for VZV. The cataract control samples were all herpesvirus DNA negative by PCR. CONCLUSIONS: To establish the diagnosis of CMV retinitis in AIDS patients, ophthalmoscopic examination is a sensitive method. In confirming a diagnosis in indistinctive cases, application of a PCR assay detecting CMV DNA is a more sensitive method than analysis of locally produced antibodies by calculating a GWq. In clinical non-herpetic uveitis, secondary release of HSV-1 and VZV should be considered requiring additional therapeutic anticipation.

A prospective clinical evaluation of the reverse transcriptase polymerase chain reaction (RNA PCR) for detection of influenza viruses was carried out with specimens from 342 patients of a children's hospital in The Netherlands. The RNA PCR, carried out directly on the specimens without an organic extraction, showed a sensitivity and specificity which are superior to those of direct immunofluorescence and comparable to those of cell culture combined with immunofluorescence (culture/IF). Negative results can be obtained within 2 days by the RNA PCR but may take up to 14 days by culture/IF. Because culturing is the standard technique for the detection of respiratory viruses, at this moment there are no strong arguments to replace culture/IF with RNA PCR for the detection of influenza A virus.

We analyzed the results of passive immunization against CMV in 146 heart transplant recipients. The 65 seronegative recipients were prophylactically treated with anti-CMV immunoglobulins during and after the operation. Twenty-nine of these 65 patients received a seropositive donor heart. CMV infection occurred in 21/65 seronegative and in 40/81 seropositive recipients (difference not significant). The incidence of CMV infection in seronegative recipients of a CMV-matched donor heart (3/34) was significantly lower than in seronegative recipients of a positive donor heart and lower than in seropositive recipients, but no significant difference in infection rate was found between the two latter groups (18/29 vs. 40/81). Although primary infection more frequently resulted in CMV disease than secondary infection (11/21 vs. 10/40) no difference in incidence of disease was noted between seronegative and seropositive patients (11/65 vs. 10/81), nor was there a difference in the severity of symptoms following primary or secondary infection. There was a higher incidence of CMV disease in all patients who received a heart from a seropositive donor versus a seronegative donor. However, after transplantation of a heart from a seropositive donor the incidence (27%) of CMV disease observed in our passively immunized seronegative patients was the same as in the patients with naturally acquired seropositivity. There was no difference in the prevalence of coronary artery disease between patients with and without CMV infection or disease. We conclude that using the current passive immunization scheme the occurrence of CMV infection and disease is largely dependent on the serostatus of the donor.

Thirty-two CMV seronegative heart transplant patients received prophylactic anti-CMV immunoglobulin during the first three posttransplant months. One of the 16 recipients of a heart from a seronegative donor acquired CMV infection and developed CMV disease. In eight of the 16 recipients of a heart from a seropositive donor, CMV infection was observed. Viremia was diagnosed in seven of them, but only two of these patients developed CMV disease. The incidence of CMV infection and of CMV disease in the globulin-treated CMV seronegative recipients of a heart from a seropositive donor was comparable to the incidence of CMV infection and of CMV disease in 31 nonglobulin-treated CMV seropositive recipients. This was significantly lower (percentage difference 69 percent, 95 percent CI 42–97 percent, p (Chest 1990; 97:396–99)

Respiratory syncytial virus (RSV) is the most prominent pathogen found in respiratory tract infections in children and the most important cause of bronchiolitis in the first two years of life. In the Netherlands approximately 2000 children are admitted each winter season. A serious course is mostly seen in children younger than 3 months, (ex-)prematures, children with bronchopulmonary dysplasia or congenital cardiac anomalies, children with cystic fibrosis younger then 2 years and children with impaired T cell immunity; such cases not rarely require intensive care. Treatment (fluid, nutrition, bronchodilator agents, corticosteroids, oxygen and ventilation) is usually symptomatic. Antiviral therapy is only indicated in immunodeficient patients. For prevention by passive immunization palivizumab was recently registered in the Netherlands, a monoclonal antibody against RSV that has to be administered intramuscularly from the start of the RSV season (15 mg per kg bodyweight once a month during five months). In a number of large-scale American multicenter studies both the number of hospital admissions related to RSV infection and the mean duration of hospital stay showed a statistically significant reduction in high-risk children who had been treated with palivizumab. Palivizumab appears to be indicated in children from the categories with an increased risk for serious RSV disease.

OBJECTIVE: To determine the prevalence and risk factors for hepatitis B virus (HBV) infections among individuals attending an STD clinic in a low endemic region. STUDY DESIGN: A total of 1228 women and 1648 men attending the STD clinic at the University Hospital Rotterdam, Netherlands, were examined for HBV infection by determination of hepatitis B surface antigen (HBsAg) and antibodies to hepatitis B core antigen (anti-HBc). Demographic characteristics, information on sexual behaviour, and intravenous drug use were recorded. RESULTS: The seroprevalence of HBsAg was 1.4% in women and 2.1% in men (0% in homosexual men). The seroprevalence of anti-HBc was 13% in women and 20% in men (36% in homosexual men). Native country, intravenous drug use, a history of STD, and the number of partners in the past half year (inversely) were independent risk factors for HBsAg positivity in women and heterosexual men. For anti-HBc independent associations were observed for native country, age, intravenous drug use, commercial sex, number of lifetime partners, homosexual contacts, orogenital contact (inverse), and a history of STD. CONCLUSION: The HBV prevalence in the STD clinic attendants was high, exceeding the national estimate, and indicates that the STD clinic population may be considered a high risk group. Our data confirmed an increased risk for HBV infections among established risk groups. Therefore, these risk groups should be routinely screened to identify HBV cases for counselling and contact tracing.

The Dutch Association for Paediatric Medicine has formulated guidelines regarding influenza vaccination of children with pulmonary disease. Influenza virus is the most frequent cause of airway infections in humans over two years of age. It may lead to serious morbidity in children with pulmonary disease: exacerbations, (transient) disturbances in pulmonary function, and symptoms lasting weeks, but mortality is probably very low. The effects of influenza vaccination of children with pulmonary disease are similar to those in normal healthy children. A positive long-term effect on the asthma has never been demonstrated. It is advised that children with moderate to severe asthma who require treatment to be vaccinated against influenza every year. If the first vaccination ever occurs before the age of six years, it should be followed by a booster vaccination after four weeks. In both instances, a full vaccination dose should be administered.

The unexpected conversion to HBsAg seropositivity of three cardiac allograft recipients prompted us to conduct a multidisciplinary study to identify the source, transmission mode, and extent of the hepatitis B virus (HBV) infection among the 256 cardiac allograft recipients of our hospital.All recipients were retrospectively screened for serum markers of HBV infection. A selected genomic region defining subtypes of the viruses involved was amplified and sequenced. An epidemiologic case-control study for possible risk factors was conducted to identify the mode of transmission.Eighteen additional HBV-infected patients were identified, none of whom had shown symptoms of HBV infection. The involvement of one virus (subtype ayw 3) was shown in 20 of the 21 HBV-infected patients. This virus is found in less than 10% of HBV-infected individuals in The Netherlands. The demonstration of a common source of infection, combined with results of the epidemiologic study, identified posttransplantation endomyocardial biopsy procedures as the most likely mode of transmission. However, we also found evidence of secondary virus transmission by cardiac catheterization procedures to nonallograft recipients.The immunosuppressive therapy practiced in these patients to prevent allograft rejection may have not only facilitated virus transmission by causing high levels of viremia but also left the spreading of HBV undetected by causing a subclinical course of the infection. These findings stress the necessity of strict hygienic precautions during intravascular diagnostic procedures and indicate that vaccination against and routine monitoring for certain bloodborne infections in cardiac allograft recipients should be considered.

The decline of maternal respiratory syncytial virus (RSV) specific serum antibodies was studied in 45 children during the first 6 months of life, using a virus neutralization assay and competition ELISAs measuring fusion protein and glycoprotein specific antibodies. In all children RSV neutralizing antibodies were demonstrated at birth, with titers ranging from 33 to 1382. The calculated mean half life of these antibodies was 26 days. Furthermore, in a group of 38 children with suspected RSV infection, all younger than 6 months of age on admission, the diagnostic value of serological assays was evaluated. In 32 children RSV infection was confirmed by virus isolation, direct immune fluorescence and RT-PCR. In 7 patients of this group a significant titer rise in virus neutralization assay was demonstrated. Six additional RSV infected children could be identified by showing the presence of RSV-specific IgM or IgA serum antibodies or by showing an increase in fusion protein or glycoprotein specific antibodies. All serological tests together identified 13 (41%) of the 32 RSV infected patients. It is concluded that in children of this age group, which represent the majority of patients hospitalized with RSV infections, serological assays not only have a limited diagnostic value but are of limited value for sero-epidemiological studies.

Respiratory syncytial virus (RSV) lower respiratory tract infections are considered to be a serious disease in centres such as the Sophia Children’s Hospital (Rotterdam, the Netherlands), but as more benign infections in others such as the Geneva Children’s Hospital (Switzerland). To assess the clinical severity of RSV infections at the two sites, 151 infants primarily admitted with a virologically confirmed RSV infection were studied prospectively (1994‐5) and retrospectively (1993‐4) (55 infants in Geneva and 96 in Rotterdam). Parameters of RSV morbidity which were more severe in Rotterdam during the two winter seasons were apnoea (1.8 v23.9%),the rate of admission to the intensive care unit (3.6 v 28.1%), mechanical ventilation (0 v 7.3%), and length of stay in hospital (6.8 v 9.1 days). In Geneva higher respiratory rates (59.2 v 51.2), more wheezing (65.5 v 28.8%), and more retractions (81.8 v 63.3%) were recorded. Fewer infants younger than 4 months (54.9 v 68.7%), but more breast fed infants (94.1 v 38.5%), were admitted in Geneva, although the morbidity parameters remained diVerent after correction for these two variables in multivariate analyses. Thus unidentified local factors influence the pattern and severity of RSV infection and may aVect the results of multicentre prophylactic and therapeutic studies. (Arch Dis Child 1997;77:410‐414)

The decline of maternal respiratory syncytial virus (RSV) specific serum antibodies was studied in 45 children during the first 6 months of life, using a virus neutralization assay and competition ELISAs measuring fusion protein and glycoprotein specific antibodies. In all children RSV neutralizing antibodies were demonstrated at birth, with titers ranging from 33 to 1382. The calculated mean half life of these antibodies was 26 days. Furthermore, in a group of 38 children with suspected RSV infection, all younger than 6 months of age on admission, the diagnostic value of serological assays was evaluated. In 32 children RSV infection was confirmed by virus isolation, direct immune fluorescence and RT-PCR. In 7 patients of this group a significant titer rise in virus neutralization assay was demonstrated. Six additional RSV infected children could be identified by showing the presence of RSV-specific IgM or IgA serum antibodies or by showing an increase in fusion protein or glycoprotein specific antibodies. All serological tests together identified 13 (41%) of the 32 RSV infected patients. It is concluded that in children of this age group, which represent the majority of patients hospitalized with RSV infections, serological assays not only have a limited diagnostic value but are of limited value for sero-epidemiological studies.

A 5-day-old male infant was admitted to hospital with a history of poor feeding and fever. He was born at term to a 22-year-old woman (gravida 1, para 0). The delivery took place at home and was uncomplicated. The birth weight was 3300 g. On admission, a child with mild respiratory distress and tachypnoea was seen. The rectal temperature was 37.5 ~ C. There was an oxygen demand of 25%. Laboratory tests included haemoglobin 12.5 mmol/1, haematocrit 0.60 1/1, platelet count 306 x 109/1, WBC count 10.9 x 109/1 with 8% band forms, 33% neutrophils and 58% lymphocytes, serum CRP level 56 rag/1 and capillary blood gas analysis pH 7.28, PCO2 9 kPa, bicarbonate 32.1 mmol/l, base-excess +3.3 retool/1. A chest radiograph showed a lobar infiltrate in the right upper lobe. Bacteriological cultures of blood, sputum and CSF remained negative. The infant was treated with intravenous ampicillin and cefotaxim. When his clinical condition did not improve intravenous erythromycin was added. A new chest radiograph at day 11 showed complete consolidation of the right lung (Fig. 1). The child was subsequently transferred to our hospital. On admission a moderately ill child was seen with tachypnoea (respiratory rate: 60/min) and oxygen demand (46%). The rectal temperature was 36.7 ~ C. Auscultation of the right lung revealed decreased breathsounds and rhonchi. Laboratory results included haematocrit 0.52 1/1, platelet count 79 x 109/1, WBC count 14.6 x 109/1 and CRP level 56 rag/1. Capillary blood gas analysis revealed pH 7.39, PCO2 7.3 kPa, base-excess +7 mmol/1. CSF analysis showed 12 cells/3 mm 3, glucose and protein levels 2.5 mmol/1 and 84 mg/100 ml respectively. A bronchoscopy with broncho-alveolar lavage was performed on day 11. Purulent secretion was seen in the right upper lobe. Treatment was started with amoxicillin/clavulanic acid (25/2.5 mg/kg qid), tobramycin (3 mg/kg once daily) and erythromycin (5 mg/kg qid) intravenously. At the age of 14 days the patient developed an erythematous rash, which disappeared spontaneously after 1 day. Bacterial cultures remained negative. On day 19 results of viral cultures were obtained.

Epstein-Barr virus-specific polymerase chain reaction was used to diagnose EBV-meningo-encephalitis in a bone marrow transplant recipient. The patient made complete recovery with ganciclovir treatment. Pitfalls in diagnosis with EBV-PCR and the potential therapeutic efficacy of ganciclovir in EBV infections are discussed.

The polymerase chain reaction (RNA-PCR) was used for specific detection of respiratory syncytial virus (RSV) genomes in clinical specimens. A set of primers was selected from conserved regions of the 1B and N genes for detection of both subgroups. The primers were found to be RSV specific, all RSV strains generated a 218 bp product, and no RSV specific amplified product was obtained when nucleic acids from a variety of micro-organisms from the respiratory tract were subjected to the RNA-PCR. We took advantage of the sequence heterogeneity of the amplified products to discriminate between the A and B strains by hybridisation with subgroup specific oligonucleotide probes. This additional hybridisation assay increased the sensitivity of the RNA-PCR tenfold. The RNA-PCR was tested on clinical specimens from children with symptoms of an infection of the respiratory tract. The results were compared with isolation of RSV in cell culture and direct immunofluorescence. From 93 specimens tested, 31 were found positive by all three techniques. Six additional positive results were detected using RNA-PCR. From these 37 RSV positive specimens 33 (92%), including all 6 additional positives, were subgroup A and only 4 were subgroup B strains. Thus, the RNA-PCR is a specific and sensitive technique for the detection and subgroup classification of RSV genomes.

Detection of cytomegalovirus (CMV) DNA in peripheral blood leukocytes has been shown to be a sensitive marker of CMV infection. However, the specificity with regard to its clinical significance is less clear, since infections considered to be latent may be detected. In this report, the presence of CMV immediate early antigen (IEA) DNA and mRNA in peripheral blood leukocytes detected by PCR was investigated and related to the appearance of CMV pp65 antigen, CMV serology, and clinical status. Thirty-seven consecutive samples were submitted to the laboratory from 36 immunocompromised patients, on a routine basis for analysis because of a potential risk of CMV infection. To facilitate differentiation between DNA and mRNA, primers were chosen in exons 2 and 3 of the immediate early gene of CMV. Keratin type I mRNA and the ssu rRNA gene served as internal controls. Thirty specimens were CMV antibody positive, of which 11 were also CMV IEA DNA positive. Two of seven seronegative samples were CMV IEA DNA positive. No relation was found between serology and the presence of CMV IEA DNA as determined in 37 samples. Five of 32 samples that could be analyzed were positive for CMV IEA mRNA, of which four were also positive in the pp65 antigen detection technique. A clear relation was found between the presence of CMV IEA mRNA and CMV pp65 antigen in leukocytes and with the clinical findings as well (P < 0.01). It is concluded that detection of CMV mRNA may have a role in diagnosis of an active clinically relevant CMV infection.

To evaluate the prevalence and the natural course of hepatitis B infection in children.Retrospective longitudinal.Four university pediatric centres.To explore the possibility of starting a trial with interferon alpha, data of viral and biochemical tests and biopsies of children younger than 16 years were studied.In a period of 10 years (1980-1990) 145 patients, of whom 74% were not of original Dutch descent, were found positive for HBsAg. The data of 142 patients could be analysed. Seroconversion was seen in 27 patients and 42 were already anti-HBe positive at the time of presentation. Chronic hepatitis, representing the category which could benefit from interferon alpha treatment, was found in 24 patients. Severe complications of the hepatitis were found in 4% of the children, including hepatocellular carcinoma and cirrhosis. Follow-up was insufficient so the seroconversion rate could only be estimated at 12% for the first year following the diagnosis.As a result of this study the authors present a proposal for a therapeutic trial with interferon alpha. This is a national protocol under the auspices of the section for pediatric gastroenterology of the Nederlandse Vereniging voor Kindergeneeskunde (Netherlands Pediatric Association).

Allograft heart valves obtained from donor hearts have been cryopreserved in the Heart Valve Bank in Rotterdam for transplantation purposes. In contrast to hepatitis B screening of organ donors, which consists of only a rapid HBV surface antigen (HBsAg) assay, tissue donors can be screened more completely for hepatitis B virus (HBV) by HBsAg and antibodies against HBV core antigen (anti-HBc) tests, and when necessary, anti-HBs and HBV-DNA tests. The value of this complete HBV screening was investigated by evaluation of the HBV screening results of 676 donor sera. HBsAg was positive in 1 serum. Anti-HBc was positive in 63 sera, of which 52 also had positive antibodies against HBV surface antigen (anti-HBs) tests (no risk of transmission) and 10 had negative anti-HBs tests. In 3 cases with a negative anti-HBs test the HBV-DNA test was positive (risk of transmission). In 3 cases not enough serum was available to perform all tests, resulting in a total of 7 rejected donors. Single HBsAg testing would have resulted in the rejection of only 1 donor. In the presented group of selected donors, approximately 0.5% of the HBsAg-negative donors were lower-level chronic carriers of hepatitis B. Complete HBV screening decreases the risk of transmission of hepatitis B in allograft heart valve transplantation.

In the period 1985-1990, 111 patients with AIDS were treated in the University Hospital Rotterdam-Dijkzigt. In 8 out of 111 patients malignant non-Hodgkin lymphoma developed. The unusual and bizarre representation is highlighted by several clinical histories and a review of literature. HIV-related non-Hodgkin lymphoma is characterized by widespread extranodal disease, often at unusual sites, and high grade B-cell malignancy. The therapeutic outcome and survival in these cases has been disappointing. Prognosis is better for patients without a prior AIDS diagnosis, higher total CD4 cell counts, good performance score and absence of an extranodal site of disease. Treatment should be tailored to individual patients based upon a variety of prognostic features.

Until recently, prevention and treatment of congenital cytomegalovirus infection was not possible. However, several studies on the epidemiology of congenital CMV infection and the development of vaccines, diagnostic tests and antiviral drugs such as ganciclovir may improve the perspectives for patients with congenital CMV disease. In this article we will discuss several of those developments that may offer new approaches for prevention and treatment of congenital CMV disease.

After the presentation of 3 case histories of children who became infected with HIV through different modes of transmission the disease is further discussed. Special attention is paid on epidemiology, transmission pathways, pathophysiology, making the diagnosis, clinical symptoms, therapy and prognosis of HIV infected children.

Central to immunization programmes is that hepatitis B virus infection in infancy is much more likely to result in the chronic carrier state than if infection occurs in adulthood. In areas of high endemicity like south-eastern Asia most infections occur when chronically infected women transmit hepatitis B virus to their newborns or when chronically infected children transmit the virus to other children. In the Netherlands, the endemicity is considered low; seroprevalence studies showed that 0.8% of the pregnant women are HBsAg-positive. To prevent mother to child transmission of hepatitis B a national immunization program has recently been implemented to prevent perinatal infection in approximately 450 neonates each year. Horizontal transmission of hepatitis B virus also seems possible in children since persistent carriers are found in (medical) centres and institutions for mentally handicapped. Since hepatitis B infections acquired during early childhood are likely to progress to chronicity and liver disease in adult life some risk factors for children in the Netherlands to contract hepatitis B infection are described and the immunization strategy is discussed.

A case of a fullterm infant with severe neonatal respiratory distress due to mumps infection is reported. Pregnancy was complicated by a self-limited febrile illness of the mother about two weeks before birth. Foetal heart rate patterns and delivery were normal. Immediately after birth the infant needed mechanical ventilation. Bacterial infections, as well as congenital cardiac or pulmonary malformations were excluded. The infant showed serologic evidence of mumps infection: IgM antibodies to mumps virus were highly positive. He expired on the 9th day of life due to bilateral pneumothoraces and pneumopericardium. Post-mortem examination showed interstitial pneumonia with intra-alveolar multinucleated giant cells, suggesting viral disease. This case demonstrates, that mumps pneumonia should be included in the differential diagnosis of severe neonatal respiratory distress in fullterm neonates.

In order to find suitable markers for selection and monitoring of antiviral therapy in asymptomatic HIV-infected patients, we evaluated 18 anti-HIV positive individuals at three monthly intervals by HIV culture, HIV antigen, and core (p24) antibody testing as well as by measurement of lymphocyte subsets. Consistent results were obtained with HIV antigen, p24 antibody testing and T4 cell enumeration, whereas results of virus detection were variable. Therefore cumbersome and expensive virus culture is not of use in selecting patients for antiviral therapy. On the basis of our results and recent literature we currently propose using absence of p24 antibodies, presence of HIV antigen and low or falling T4 cells as eligibility criteria for antiviral therapy in asymptomatic infected individuals.

The case is reported of a 39-year-old pregnant woman who presented with fever, abdominal complaints, and diarrhea. Laboratory investigation revealed mononucleosis in the peripheral blood. All microbiological studies were negative, with the exception of finding cytomegalovirus (CMV). Seroconversion was documented; the virus was cultured from urine and subsequently was demonstrated to be present in the inflamed mucosa of the rectum and distal sigmoid, which was found at sigmoidoscopy. This woman was delivered of a neonate with congenital CMV infection but without apparent malformations. The patient experienced recurrences of the bowel disease, in the first of which CMV could still be cultured from a biopsy specimen. In the follow-up period, an otherwise aspecific chronic inflammatory bowel disease remained present. No immunological abnormalities were found, and antibodies to human immunodeficiency virus were negative. This case demonstrates that inflammatory bowel disease can develop as a result of primary infection with CMV.

Thirty-two CMV seronegative heart transplant patients received prophylactic anti-CMV immunoglobulin during the first three posttransplant months. One of the 16 recipients of a heart from a seronegative donor acquired CMV infection and developed CMV disease. In eight of the 16 recipients of a heart from a seropositive donor, CMV infection was observed. Viremia was diagnosed in seven of them, but only two of these patients developed CMV disease. The incidence of CMV infection and of CMV disease in the globulin-treated CMV seronegative recipients of a heart from a seropositive donor was comparable to the incidence of CMV infection and of CMV disease in 31 nonglobulin-treated CMV seropositive recipients. This was significantly lower (percentage difference 69 percent, 95 percent CI 42-97 percent, p less than 0.001) than expected on the basis of the data from the literature and indicates that passive immunization with anti-CMV immunoglobulins induces the same protection against CMV disease as natural acquired anti-CMV resistance. This protective effect was temporary, as one patient developed symptomatic CMV infection four months after transplantation at a time when the anti-CMV immunoglobulin levels had decreased to pretransplantation values.

We prospectively studied the role of MP in childhood LTRI. Children aged 3 months to 12 years with a chest X-ray infitrate were included. Excluded were immunodeficiency disorders, aspiration, congenital malformations, nasogastric tube feeding, cystic fibrosis and severe retardation. A complement fixation test and an IF assay measuring IgM and IgG antibodies on day 1 and 14 were performed to confirm infection with MP. A four-fold rise in IgG titers or a positive IgM titer were considered diagnostic. On day 1 a nasopharyngeal aspirate was taken for culture of MP and PCR analysis. The primers used in PCR analysis were: Myc 16S and Myc PL Over a period of 11 months 34 children (15F, 19M) with a mean age of 5.5 years were included. In 10 patients (29%) MP infection was detected. The results of serologic testing(SER.), culture(CULT.) and PCR are shown below

The case is reported of a 39-year-old pregnant woman who presented with fever, abdominal complaints, and diarrhea. Laboratory investigation revealed mononucleosis in the peripheral blood. All microbiological studies were negative, with the exception of finding cytomegalovirus (CMV). Seroconversion was documented; the virus was cultured from urine and subsequently was demonstrated to be present in the inflamed mucosa of the rectum and distal sigmoid, which was found at sigmoidoscopy. This woman was delivered of a neonate with congenital CMV infection but without apparent malformations. The patient experienced recurrences of the bowel disease, in the first of which CMV could still be cultured from a biopsy specimen. In the follow-up period, an otherwise aspecific chronic inflammatory bowel disease remained present. No immunological abnormalities were found, and antibodies to human immunodeficiency virus were negative. This case demonstrates that inflammatory bowel disease can develop as a result of primary infection with CMV. ( Arch Intern Med. 1990;150:1749-1751)

Reticulocytes in fixed human blood samples were stained for RNA with the fluorescent dye pyronin Y and measured by flow cytometry. The resulting relative frequency distributions of the RNA fluorescence intensities concurred with the different stages in maturation from early reticulocytes to mature red cells. A computer program was written to calculate from these frequency distributions the relative number of reticulocytes, their relative RNA content, and the median of the reticulocyte population (RNA index). This method was applied to 30 healthy blood bank donors (control group), as well as to patients with various hematologic disorders showing abnormal erythropoietic activity. The measured percentage of reticulocytes, RNA content, and RNA index were found to correlate well with the various hematologic disorders. Changes in erythropoiesis could be clearly followed, as was demonstrated by analyzing blood samples from children with aplastic anemia or acute myeloid leukemia, who were treated with allogeneic bone marrow transplantation. Measurements on blood samples from healthy blood bank donors showed that with this method, small changes in the reticulocyte population, such as the appearance of polychromatic erythrocytes in the peripheral blood 5–8 hr after donation, can be detected. The statistical reliability and the information provided on the maturation stage of the entire reticulocyte population make flow cytometry of peripheral blood reticulocytes a more informative method for the study of hematologic abnormalities than conventional methods for reticulocyte counting and classification.

A recently described immunoperoxidase method for the detection of nuclear human cytomegalovirus (HCMV) immediate early antigen (IEA) directly on periphiral blood leucocytes suffers from the drawback that the antigen is vulnerable to endogenous peroxidase inactivation procedures. To solve this problem a procedure is developed in which endogenous peroxidase is inactivated after binding and immobilization of the primary antibody with 4% formaldehyde. In combination with this procedure, three types of inactivation were investigated: glucose/glucose oxidase, hydrochloric acid and methanol/H2O2. Of these three, the first gives optimal results, especially in combination with methanol/acetic acid (20/1 v/v) as the primary fixative. This produre results in preparations which allow for a more objective evaluation and enable automated examination using bright field microscopy.As a second improvement we developed a simple adherence method in order to diminish the risk of infection for the laboratory staff during processing of unknown blood samples. The protocol described shows great clinical potentual for the diagnosis of HCMV infections.

During the winter of 1986-1987, 64 children with respiratory syncytial virus (RSV) infection were admitted to our hospital. The diagnosis was made by direct immunofluorescent antibody technique. Twenty-three children (36%) needed intensive care treatment. Nearly 11 (52%) had a preexisting disease state, identified as a risk factor i.e., prematurity (n = 8), bronchopulmonary dysplasia (n = 2), congenital heart disease (n = 1). Twelve patients (50%) were intubated and ventilated. Conditions for intubation and ventilation were repetitive apnea with or without bradycardia (n = 4), clinical deterioration (n = 3) or hypercarbia (n = 5). Seventy-five percent of the patients who needed intensive care management were under three months of age compared to 34% of the children who were admitted to the clinical ward. The mean age for ventilated patients was 7.9 weeks. The mean duration of ventilation was 5.5 days. Volume controlled ventilation was initially applied to all patients. Pulmonary complications (atelectasis, pneumonia, pneumothorax or adult respiratory distress syndrome) were present in 15 (65%) IC patients. Nine (39%) of them also had symptoms of inappropriate antidiuretic hormone secretion (IADHS). Only two patients had symptoms of IADHS and two others had convulsions. Three children (5%) died as a result of respiratory insufficiency. Two of these infants belonged to the risk group.

An assay is described for handling clinical specimens for the detection of cytomegalovirus. It consists of low-speed centrifugation of the specimen on human embryonic lung cells, using a technique adapted from chlamydial culture, followed by detection using a monoclonal antibody against the cytomegalovirus early antigen in an immunofluorescence technique. This assay was compared with the conventional cell culture system. 161 specimens obtained from 52 patients were studied; from 14 patients CMV was isolated in at least one specimen (in total 28 specimens). The centrifugation technique led to positive results generally within 24 to 48 h, whereas the cell culture took an average of 16.5 days to develop the typical cytopathic changes. No cross-reactions between the cytomegalovirus monoclonal antibody and other viruses present (herpes simplex virus and adenovirus) were observed. The centrifugation technique is a reproducible and rapid method in the diagnosis of cytomegalovirus infection.

A four-month-old girl was hospitalized with pneumococcal sepsis from which she recovered. Subsequently she developed various other infectious diseases, including chronic diarrhoea caused by Cryptosporidium. After a period with neurological symptoms, later sagittal sinus thrombosis and cerebral atrophy, she died at age 13 months. It was found that the child suffered from AIDS. The mother was seropositive and the virus had probably been transmitted via the placenta. Rapid recognition of infants and young children with AIDS is necessary, for in the near future more cases of this disease will occur. Antiviral treatment may bring about improvement in children as well.

Two hundred and thirty seven semen samples from 10 institutes for artificial insemination by donor (AID) in Belgium and the Netherlands were tested for the presence of Neisseria gonorrhoeae, Chlamydia trachomatis, Mycoplasma hominis, Ureaplasma urealyticum, herpes simplex virus, and cytomegalovirus. The incidence of these micro-organisms in the semen samples was 0%, 6.3%, 4.6%, 35.9%, 0%, and 0.4% respectively, and 47% of all samples were infected with one or more of the micro-organisms. As the ejaculates from which the samples had been taken had already been, or would be, used for AID, the exclusion of microbiological contamination with sexually communicable micro-organisms before insemination is indicated.