Britt-Marie Eriksson

Early detection of cytomegalovirus in cell culture by a new monoclonal antibody, CCH2

A CMV monoclonal antibody, CCH2, produced in this laboratory was evaluated for rapid detection of CMV. Two staining procedures, immunofluorescence and an immunoenzymatic technique using biotin-streptavidin peroxidase, were compared. The CCH2 monoclonal antibody was used to demonstrate early CMV antigen in cell culture 24 h after inoculation of 598 urine samples from kidney transplanted patients by indirect immunofluorescence in comparison with virus isolation. One hundred and sixty of the specimens were stained additionally by an immunoenzymatic technique and the results were compared. CMV was isolated from 170 out of 598 specimens within 6 weeks. Early CMV antigen was demonstrated in 114 of these specimens by immunofluorescence giving a sensitivity of 67% and a specificity of 95%. In the comparison with the immunoenzymatic staining procedure the results for all three tests agreed for 81% (130/160) of the specimens. After resolving discordant results into true positives and true negatives, the sensitivity was 87, 85 and 70%, respectively for virus isolation, immunoenzymatic staining and immunofluorescence and the specificity 100, 96 and 99%. The CCH2 monoclonal antibody proved to be useful for rapid detection of CMV in urine specimens and using immunoenzymatic staining with biotin-streptavidin a sensitivity comparable to that of virus isolation was found.

Liver transplantation in a boy with acute porphyria due to aminolaevulinate dehydratase deficiency.

The clinical and biochemical outcome of a liver transplantation in a seven-year-old boy with acute porphyria due to aminolaevulinate dehydratase deficiency is described. Before transplantation standard liver function tests were normal and the rationale for transplantation was that the new liver would reduce the metabolic disturbance and thus avert the porphyric symptoms. During the year after the transplantation, the functioning of the new liver has been excellent. Basal excretion of porphyrin and porphyrin precursors has remained unchanged but, with the new liver transplant the patient has been able to withstand several porphyrinogenic challenges without increasing the excretion. Episodes of neurological and respiratory crises may have been due to persistent porphyric vulnerability. Alternatively, two early attacks may have been caused by neurotoxic effects of cyclosporin in combination with the existing damage to nervous tissue.

Diagnosis of Pulmonary Infections in Immunocompromised Patients by Fiber-optic Bronchoscopy with Bronchoalveolar Lavage and Serology

Fiber-optic bronchoscopy (FOB) and bronchoalveolar lavage (BAL) were performed on 67 occasions in 57 immunocompromised patients with symptoms consistent with pulmonary infection. Diagnosis was achieved more often in renal transplant patients than in patients with hematological malignancies (85% versus 28%). Culture (bacteria, virus, fungi), staining and microscopy (bacteria, fungi, Pneumocystis carinii (PC)) and antigen detection by indirect immunofluorescence (cytomegalovirus (CMV), respiratory viruses, PC, Legionella) were used for diagnosis. On 20 occasions transbronchial biopsies with histopathologic examination were performed. In addition, serology comprising the herpes group (HHV-6) and respiratory viruses was done. A microbial diagnosis was obtained on 45% of occasions. The most common pathogens found were CMV (31%) and PC (25%). On 22 (33%) occasions a rapid diagnosis of 1 or more microbial agents was obtained within 24 h by conventional staining or indirect immunofluorescence. The clinical relevance of findings of CMV, HHV-6, and Epstein-Barr virus in BAL by polymerase chain detection on 18, 6 and 3 occasions is discussed. On 4 occasions pathogenic bacteria were found. It was not possible to relate findings of coagulase-negative staphylococci, alpha-streptococci and Candida albicans to the pulmonary infection.

Seroreactivity to Pneumocystis carinii in Patients with AIDS versus Other Immunosuppressed Patients

The aim was to study the humoral response to Pneumocystis carinii and its diagnostic use in patients with P. carinii pneumonia (PCP). The antibody response was measured by indirect immunofluorescence in AIDS patients versus other immunosuppressed patients with 122 episodes of confirmed PCP. During the early acute stage of the pneumonia, anti-P. carinii antibodies were found in 17% of AIDS and 24% of other immunosuppressed patients. In the second serum sample, antibodies were still found in 17% of the AIDS patients but in as many as 56% of the otherwise immunosuppressed patients. Antibodies were also found in 17% of HIV-positive and 15% of other immunosuppressed control patients, but only in 3% of immunocompetent controls (p < 0.001). Paired sera were available from 55 patients during 58 PCP episodes. Seroconversion or a fourfold rise in titre was detected in only 1/36 (3%) AIDS patients but in 10/22 (45%), (95% c.i.: 24-66%) other immunosuppressed patients (p < 0.001). We conclude that AIDS patients seem to have lost their ability to develop a humoral response to P. carinii during pneumonia, whereas many other immunosuppressed patients do respond. In these patients the serological test against P. carinii was of no diagnostic value in the acute phase of the infection, whereas when analysing paired sera it was a useful complement to the clinical diagnosis.

Diagnosis of Cytomegalovirus in Bronchoalveolar Lavage by Polymerase Chain Reaction, in Comparison with Virus Isolation and Detection of Viral Antigen

Bronchoalveolar lavage (BAL) products from 52 immunocompromised patients with symptoms of pulmonary infection was examined for cytomegalovirus (CMV) by virus isolation, polymerase chain reaction (PCR) and detection of CMV antigen by immunofluorescence or immunoperoxidase staining after short-term incubation in tissue culture and directly in BAL cells. We found that PCR detected all cases positive by virus isolation (15/52 samples) and the result was obtained within 5 h. PCR detected more cases of CMV than did virus isolation (22/52 samples). Positive PCR and negative virus isolation were consistent with probable CMV infection in 3/7 patients when other clinical and laboratory parameters of CMV infection were considered. The negative predictive value of PCR was high; none of 30 patients negative by PCR developed CMV pneumonia within the subsequent 2 months. Detection of CMV antigen after short-term incubation was rapid enough to be used in clinical practice, specific (100%) and with a sensitivity of 60%. Demonstration of CMV antigen in alveolar cells was highly specific (100%) but had too low a sensitivity (26.7%) to be used as the only rapid method. Our conclusion is that a combination of PCR and detection of CMV antigen after short-term incubation and directly in alveolar cells is optimal for rapid identification of CMV.

Hospital outbreak control requires joint efforts from hospital management, microbiology and infection control

An outbreak of multidrug-resistant Klebsiella pneumoniae producing the extended-spectrum beta-lactamase CTX-M15 affected 247 mainly elderly patients in more than 30 wards in a 1000-bedded swedish teaching hospital between May 2005 and August 2007. A manual search of the hospital administrative records for possible contacts between cases in wards and outpatient settings revealed a complex chain of transmission. Faecal screening identified twice as many cases as cultures from clinical samples. Transmission occurred by direct and indirect patient-to-patient contact, facilitated by patient overcrowding. Interventions included formation of a steering group with economic power, increased bed numbers, better compliance with alcohol hand disinfection and hospital dress code, better hand hygiene for patients and improved cleaning. The cost of the interventions was estimated to be euro3 million. Special infection control policies were not necessary, but resources were needed to make existing policies possible to follow, and for educational efforts to improve compliance.

Severe group A streptococcal infections in Uppsala County, Sweden: clinical and molecular characterization of a case cluster from 2006 to 2007.

This study describes a recent cluster of 30 patients (median age 52 years) with serious group A streptococcal (GAS) infections in Uppsala County, Sweden, from December 2006 to May 2007. Patients hospitalized with a severe GAS infection, i.e. cases with either invasive GAS (iGAS) disease or patients with a positive non-sterile site culture/rapid antigen test for GAS and clinically considered as having a critical disease, were included in the study. Common clinical presentations were skin and soft tissue infections (53%) and pneumonia (17%). Eight patients (27%) were diagnosed with streptococcal toxic shock syndrome. In 40% of the cases no relevant underlying disease was reported. Among the 16 patients with soft tissue infections, the upper chest, neck or upper arm area was frequently affected and the infection was associated with severe pain. Among the 20 collected isolates, the T1/emm1 type dominated (80%). The majority (86%) of 7 analysed acute sera lacked neutralizing activity against superantigens produced by the patients’ own infecting isolate. The study underscores the association between T1/emm1 and outbreaks of serious GAS infections. This highlights the importance of surveillance for prompt identification of more aggressive isolates in the community, thereby increasing awareness among healthcare professionals of these life-threatening infections.

CMV disease in CMV-mismatched renal transplant recipients with prophylactic low dose valaciclovir

BACKGROUND Valaciclovir (VACV) 2 g q.i.d. for 3 months after kidney transplantation has been shown, (Lowance et al., NEJM 1999; 340: 1462-70), to reduce CMV disease from 45 to 16% and rejection from 52 to 26% in CMV-negative (D+R-) recipients. Neurotoxic side effects, however, were frequent, and 5% of the patients experienced hallucinations. We hypothesised that a lower dosage of VACV would prevent CMV disease with fewer CNS side effects. METHODS Since September 1998 all CMV-mismatched renal transplant recipients received VACV 1 g t.i.d. for 3 months posttransplantation (PT). The incidence of CMV disease, rejection and neurotoxic side effects during 6 months PT was studied retrospectively in, up to now, 25 patients. RESULTS 24% (6/25) of the patients developed CMV disease. The mean time for onset of symptoms was 145 days (92-191). Five of the patients had mild-moderate symptoms and recovered after ganciclovir therapy for 3 weeks. One patient was diagnosed with a CMV-associated retinitis on day 191 PT. The rate of biopsy-confirmed acute graft rejection was 32% (8/25). 20% (5/25) of the patients had a serum creatinine of >200 micromol/l after 6 months, including one patient on hemodialysis. CNS adverse effects were not observed. None out of nine patients with basiliximab induction and VACV developed CMV disease. One patient with basiliximab that did not receive VACV, developed a symptomatic CMV-infection. CONCLUSIONS The incidence of CMV disease was lower than in historical controls at our centre, and the time to onset of symptoms was prolonged. Compared to the 8 g VACV/day study, CMV disease and graft rejection was more frequent, but no neurotoxic side effects were observed. A combination with basiliximab induction and VACV 3 g/day shows promising results, but randomised studies are needed for confirmation.

Polymerase chain reaction with double primer pairs for detection of human parvovirus B19 induced aplastic crises in family outbreaks

Parvovirus B19 DNA can be detected by polymerase chain reaction with double primer pairs (nested PCR). Recent infection was documented by a retrospective serological study using Parvoscan-B19 enzyme linked immunosorbent assay (EIA) for detection of B19 human parvovirus IgM and IgG antibodies in serum or plasma specimens. In 3 families B19 outbreaks caused aplastic crises necessitating blood transfusion in 5 children and 1 adult with hereditary sphaerocytosis. Four members from 2 of the families had clinically overt haemolytic anaemia prior to the event. Two members in another family presented with an aplastic crisis disclosing the underlying chronic haemolytic disease. All 7 patients were identified as PCR positive in serum samples taken 3-14 days after the onset of symptoms. Comparison with dot blot hybridization revealed detectable DNA in only 2/3 PCR positive patients. Thus, nested PCR is more sensitive than the dot blot hybridization method and is therefore a suitable complement to the antibody assay for identifying recent B19 infection.

Human Parvovirus B19 Infection with Severe Anemia Affecting Mother and Son

Two patients, a 43-year-old mother and her 12-year-old son developed severe anemia after a disease with fever, myalgia, rash and gastrointestinal symptoms. The infectious proved to be caused by human parvovirus B19 (IgM antibody titer rises). Investigations revealed that both patients suffered from spherocytosis, apparently hereditary and not known before. This emphasizes the importance of thorough hematological investigations in patients showing symptoms resembling a viral disease when seen in connection with severe anemia in order to reveal underlying chronic hemolytic disease. In patients with these diseases transient aplastic crisis will give a much more pronounced anemia than in otherwise healthy individuals.