Validation of a new Aspergillus real-time PCR assay for direct detection of Aspergillus and azole resistance of Aspergillus fumigatus on bronchoalveolar lavage fluid.

Azole resistance in Aspergillus fumigatus is increasingly reported. Here, we describe the validation of the AsperGenius, a new multiplex real-time PCR assay consisting of two multiplex real-time PCRs, one that identifies the clinically relevant Aspergillus species, and one that detects the TR34, L98H, T289A, and Y121F mutations in CYP51A and differentiates susceptible from resistant A. fumigatus strains. The diagnostic performance of the AsperGenius assay was tested on 37 bronchoalveolar lavage (BAL) fluid samples from hematology patients and 40 BAL fluid samples from intensive care unit (ICU) patients using a BAL fluid galactomannan level of ≥1.0 or positive culture as the gold standard for detecting the presence of Aspergillus. In the hematology and ICU groups combined, there were 22 BAL fluid samples from patients with invasive aspergillosis (IA) (2 proven, 9 probable, and 11 nonclassifiable). Nineteen of the 22 BAL fluid samples were positive, according to the gold standard. The optimal cycle threshold value for the presence of Aspergillus was <36. Sixteen of the 19 BAL fluid samples had a positive PCR (2 Aspergillus species and 14 A. fumigatus samples). This resulted in a sensitivity, specificity, and positive and negative predictive values of 88.9%, 89.3%, 72.7%, and 96.2%, respectively, for the hematology group and 80.0%, 93.3%, 80.0%, and 93.3%, respectively, in the ICU group. The CYP51A real-time PCR confirmed 12 wild-type and 2 resistant strains (1 TR34-L98H and 1 TR46-Y121F-T289A mutant). Voriconazole therapy failed for both patients. The AsperGenius multiplex real-time PCR assay allows for sensitive and fast detection of Aspergillus species directly from BAL fluid samples. More importantly, this assay detects and differentiates wild-type from resistant strains, even if BAL fluid cultures remain negative.

The type I interferon response during viral infections: a "SWOT" analysis.

The type I interferon (IFN) response is a strong and crucial moderator for the control of viral infections. The strength of this system is illustrated by the fact that, despite some temporary discomfort like a common cold or diarrhea, most viral infections will not cause major harm to the healthy immunocompetent host. To achieve this, the immune system is equipped with a wide array of pattern recognition receptors and the subsequent coordinated type I IFN response orchestrated by plasmacytoid dendritic cells (pDCs) and conventional dendritic cells (cDCs). The production of type I IFN subtypes by dendritic cells (DCs), but also other cells is crucial for the execution of many antiviral processes. Despite this coordinated response, morbidity and mortality are still common in viral disease due to the ability of viruses to exploit the weaknesses of the immune system. Viruses successfully evade immunity and infection can result in aberrant immune responses. However, these weaknesses also open opportunities for improvement via clinical interventions as can be seen in current vaccination and antiviral treatment programs. The application of IFNs, Toll-like receptor ligands, DCs, and antiviral proteins is now being investigated to further limit viral infections. Unfortunately, a common threat during stimulation of immunity is the possible initiation or aggravation of autoimmunity. Also the translation from animal models to the human situation remains difficult. With a Strengths-Weaknesses-Opportunities-Threats ("SWOT") analysis, we discuss the interaction between host and virus as well as (future) therapeutic options, related to the type I IFN system.

SCAN. Schedules for Clinical Assessment in Neuropsychiatry.

After more than 12 years of development, the ninth edition of the Present State Examination (PSE-9) was published, together with associated instruments and computer algorithm, in 1974. The system has now been expanded, in the framework of the World Health Organization/Alcohol, Drug Abuse, and Mental Health Administration Joint Project on Standardization of Diagnosis and Classification, and is being tested with the aim of developing a comprehensive procedure for clinical examination that is also capable of generating many of the categories of the International Classification of Diseases, 10th edition, and the Diagnostic and Statistical Manual of Mental Disorders, revised third edition. The new system is known as SCAN (Schedules for Clinical Assessment in Neuropsychiatry). It includes the 10th edition of the PSE as one of its core schedules, preliminary tests of which have suggested that reliability is similar to that of PSE-9. SCAN is being field tested in 20 centers in 11 countries. A final version is expected to be available in January 1990.

Psychiatric diagnosis in an outpatient population. A comparative study of PSE-Catego and DSM-III.

Two diagnostic systems are compared in a psychiatric outpatient population of 175 patients. The Present State Examination (PSE)-Catego system identified 121 patients (69%) as "cases," whereas DSM-III identified 152 patients (87%) as cases. The two systems converged in 115 patients, yielding a kappa coefficient of only .32. Approximately one third of the DSM-III cases that were not detected by the PSE-Catego system was due to the restricted coverage of the latter system; the remaining two thirds could be attributed to differences in threshold and time framing. Compared with DSM-III, the PSE-Catego system showed a strong bias toward depression, and the system was extremely insensitive to the detection of social phobias and obsessive-compulsive disorders. Only 58% of cases of depression and 46% of cases of anxiety were diagnosed by both systems. The results are compared with other studies, and some consequences are discussed.

Predicting feasibility of day treatment for unselected patients referred for inpatient psychiatric treatment: results of a randomized trial.

OBJECTIVE: Because previous studies of day treatment as an alternative to inpatient treatment had major disadvantages or methodological shortcomings, the authors conducted a randomized controlled trial to estimate and predict the extent to which day treatment is feasible for unselected patients referred for inpatient treatment. METHOD: Of 160 patients, 57 were randomly assigned to the control condition and 103 were assigned to the experimental condition. Control patients received standard clinical care. In the experimental condition, day treatment was attempted as soon as the patient's condition permitted. The average number of nights per week that experimental patients spent away from the hospital was compared to the average number of nights away for patients under standard care. RESULTS: Day treatment was satisfactory for 40% of the experimental patients but was completely infeasible for another 40%. The level of surveillance needed in the first week, physical illness, number of previous admissions, depressive symptoms, and treatment by qualified psychiatrists versus registrars were variables predictive of these differences. CONCLUSIONS: In this unselected group of patients, no absolute contraindications against day treatment were found. This suggests that the selection criteria applied in nearly all other controlled studies on the subject were unwarranted. The approach used in this study facilitated treatment in the least restrictive environment possible.

The WHO collaborative study on strategies for extending mental health care, II: The development of new research methods.

Teams in seven developing countries under sponsorship of the World Health Organization have been carrying out collaborative operational research on providing mental health care through primary health care services. New techniques of identifying mental disorders in children and adults have been developed and tested. Methods of assessing the skills and attitudes of health workers toward mental health work and of gauging community attitudes toward mental illness have also been developed. Results have been directly applied in planning better mental health care. The authors conclude that cross-cultural collaborative research is effective in improving mental health care for those in greatest need.

The WHO collaborative study on strategies for extending mental health care, III: Evaluative design and illustrative results.

Teams in seven developing countries have adopted a common research design to evaluate new community mental health care services. The nature of the intervention programs varied considerably according to the characteristics of each area. Observations made before the intervention and 18 to 24 months after showed significant changes in the attitudes, knowledge, and diagnostic accuracy of health staff and in community attitudes and reactions. A considerable number of individuals with serious mental disorders received effective care for the first time.

Childhood mental disorders in primary health care: results of observations in four developing countries. A report from the WHO collaborative Study on Strategies for Extending Mental Health Care.

To ascertain the frequency of mental disorders in Sudan, Philippines, India, and Columbia, 925 children attending primary health care facilities were studied. Rates of between 12% and 29% were found in the four study areas. The range of mental disorders diagnosed was similar to the encountered in industrialized countries. The research procedure involved a two-stage screening in which a ten-item "reporting questionnaire" constituted the first stage. The study has shown that mental disorders are common among children attending primary health care facilities in four developing countries and that accompanying adults (usually the mothers) readily recognize and report common psychologic and behavioral symptoms when these are solicited by means of a simple set of questions. Despite this, the primary health workers themselves recognized only between 10% and 22% of the cases of mental disorder. The result have been used to design appropriate brief training courses in childhood mental disorders for primary health workers in the countries participating in the study.

Knowledge and attitudes of primary health care personnel concerning mental health problems in developing countries: a follow-up study.

Within the context of a World Health Organization coordinated collaborative study health workers in six developing countries were assessed 18 months after their training for improvement in their knowledge and attitude towards mental health problems and their management. The approaches to training varied between study areas, but the degree of improvement following the training, was of equal magnitude in all countries. The training process has formalized the recognition by the health workers that treatment of mental health problems is an integral part of their work.

General practitioners' characteristics and the assessment of psychiatric illness.

We put forward the hypothesis that general practitioners (GPs) with a family medicine orientation are more sensitive to the presence of mental health problems than GPs with a clinical orientation. To test it, GPs were divided into three subgroups on the basis of an attitude questionnaire. The General Health Questionnaire (GHQ) and the Present State Examination (PSE) were used as criteria. No differences in sensitivity to psychiatric illness were observed using either scale. Results of factor analysis with the subscales of the GP attitude questionnaire and the indices 'bias' and 'accuracy' were similar to those reported by GOLDBERG and associates. 'New' patients were defined as patients in whom the GP had not identified a mental health problem (MHP) in the past year. 'Old' patients were defined as 'not new'. GPs tended to under-identify MHPs in 'new' and over-identify them in 'old' patients. Recognition of psychiatric illness was better in 'old' than in 'new' patients. In 'new' patients, recognition depended on psychiatric diagnosis. Among 'old' patients, older people and people (especially women) with low education predominated. In their assessments GPs used information not contained in the GHQ.

Costs and benefits of day treatment with community care for schizophrenic patients.

The feasibility of day treatment with community care for schizophrenic patients was tested by means of a longitudinal randomized experiment with 34 experimentals and 16 controls: 38 percent could be treated satisfactorily in a day program that included a very active ambulatory service. The new approach did not improve prognosis with respect to psychiatric symptomatology, social role disabilities, or number of readmissions during the first year of followup. Total cost of treatment was less for day-treatment patients than for ordinary clinical patients.

Natural course of schizophrenic disorders: a 15-year followup of a Dutch incidence cohort.

Data are presented on the 15-year natural course of schizophrenia and other nonaffective functional psychoses in a cohort of 82 first-contact cases from a circumscribed area in the Netherlands. The subjects were suffering from functional psychosis with International Classification of Diseases-Ninth Revision (ICD-9) diagnoses 295, 297, or 298.3-9 (broad definition of schizophrenia) on entry. Standardized assessments of psychopathology, psychological impairments, negative symptomatology, social disability, and use of mental healthcare were used. The study reveals a pattern of chronicity and relapses with a high risk of suicide: Two-thirds of the subjects had at least one relapse and after each relapse 1 of 6 subjects did not remit from the episode; 1 of 10 committed suicide; and 1 of 7 had at least one episode with affective psychotic symptoms that started on average 6 years after the onset of the schizophrenic disorder. Diagnoses were reclassified in five patients, according to DSM-III-R criteria for a bipolar disorder. The predictive power--in terms of time in psychosis and in partial or full remission--of demographic, illness, and treatment variables at onset of the illness was very limited. Insidious onset and delays in mental health treatment are risk factors that predict a longer duration of first or subsequent episodes. The importance of mental health treatment in regard to outcome is probably subject to change because an early warning and intervention strategy could prevent further damage and deterioration. Our data support the need for an adequate relapse prevention program as a priority for our mental health services.

Coping with outpatients who cannot cope : management of persistent complainers in an African country.

Some 15 to 20% of patients visiting general outpatient services in developing countries suffer from some emotional disorder although they present mainly with somatic complaints. Their contact with the services is generally a frustrating experience for themselves and for the health worker, resulting in undue medical attention to their problem. A series of five consecutive steps, of increasing psychiatric sophistication, to assist in the management of these cases is described, permitting the health worker to stop at a level not exceeding his competence.

Costs and benefits of hospital and day treatment with community care of affective and schizophrenic disorders.

BACKGROUND: A randomised controlled trial of day treatment with community care for patients with schizophrenic and affective disorders, referred for in-patient psychiatric treatment, was conducted to evaluate patterns of treatment and the course of illness with its psychosocial consequences over a period of two years. METHOD: Seventy patients, of whom 34 had affective and 36 had schizophrenic disorder, were assigned to the experimental condition (day treatment with ambulatory and domiciliary care), and 33 patients, of whom 16 had affective and 17 had schizophrenic disorder, were assigned to the control condition of standard clinical care. RESULTS: Day treatment with community care was feasible for 40.6% of the affective patients and 33.3% of the schizophrenic patients. The direct treatment costs of both disorders, based on numbers of in- and day-patient days and out-patient contacts over two years, appeared more or less the same. Patients benefited equally from day treatment as from in-patient treatment, although there were some gains in self-care and in functioning in the household among experimentals. Although schizophrenics were socioeconomically worse off, and also suffered from more (severe) symptoms and social disabilities than the affective patients at entry into the study, they were similar at two years. This finding is unexpected, compared with other follow-up studies. Extra cost for patients and families were not observed. Patients and their families in the experimental condition were significantly more satisfied with the treatment. Experimental patients spent much more time at home during admission, remained much less time in secluded wards, and were more compliant with treatment. CONCLUSIONS: Day treatment could be considered a cost-effective alternative to in-patient treatment.

Efficacy of IFN-λ1 to protect human airway epithelial cells against human rhinovirus 1B infection.

Impaired interferon (IFN) production has been observed in various obstructive respiratory diseases. This contributes to enhanced sensitivity towards viral infections triggering acute exacerbations. To compensate for this impaired host IFN response, there is need to explore new therapeutic strategies, like exogenous administration of IFNs as prophylactic treatment. In the present study, we examined the protective potential of IFN-λ1 and compared it with the previously established protecting effect of IFN-ß. A549 cells and human primary bronchial epithelial cells were first treated with either IFN-ß (500 IU/ml) or IFN-λ1 (500 ng/ml) for 18 h. For infection, two approaches were adopted: i) Continuous scenario: after pre-treatment, cells were infected immediately for 24 h with human rhinovirus 1B (HRV1B) in IFN-containing medium, or were cultured for another 72 h in IFN-containing medium, and then infected for 24 h with HRV1B, ii) Pre-treatment scenario: IFN-containing medium was replaced after 18 h and cells were infected for 4 h either immediately after pre-treatment or after additional culturing for 72 h in IFN-free medium. The protective effect was evaluated in terms of reduction in the number of viral copies/infectious progeny, and enhanced expression of IFN-stimulated genes (ISGs). In both cell types and in both approaches, IFN-λ1 and IFN-ß treatment resulted in pronounced and long-lasting antiviral effects exemplified by significantly reduced viral copy numbers and diminished infectious progeny. This was associated with strong up-regulation of multiple ISGs. However, in contrast to the IFN-ß induced expression of ISGs, which decreased over time, expression of ISGs induced by IFN-λ1 was sustained or even increased over time. Here we demonstrate that the protective potential of IFN-λ1 is comparable to IFN-ß. Yet, the long-lasting induction of ISGs by IFN-λ1 and most likely less incitement of side effects due to more localized expression of its receptors could make it an even more promising candidate for prophylactic treatment than IFN-ß.

The host immune response contributes to Haemophilus influenzae virulence.

BACKGROUND: There is compelling evidence that infections with non-typeable Haemophilus influenzae (NTHi) are associated with exacerbations in COPD patients. However, NTHi has also been isolated frequently during clinically stable disease. In this study we tested the hypothesis that genetically distinct NTHi isolates obtained from COPD patients differ in virulence which could account for dissimilarities in the final outcome of an infection (stable vs. exacerbation). RESULTS: NTHi isolates (n = 32) were obtained from stable COPD patients, or during exacerbations. Genetically divergent NTHi isolates were selected and induction of inflammation was assessed as an indicator of virulence using different in vitro models. Despite marked genomic differences among NTHi isolates, in vitro studies could not distinguish between NTHi isolates based on their inflammatory capacities. Alternatively, when using a whole blood assay results demonstrated marked inter-, but not intra-individual differences in cytokine release between healthy volunteers irrespective of the origin of the NTHi isolate used. CONCLUSION: Results suggest that the individual immune reactivity might be an important predictor for the clinical outcome (exacerbation vs. no exacerbation) following NTHi infection.

Interferon-ß induces a long-lasting antiviral state in human respiratory epithelial cells.

OBJECTIVES: Interferon-ß (IFNß) induces strong antiviral effects and is therefore an attractive agent to prevent or reduce the incidence of virus-mediated exacerbations in asthmatic or chronic obstructive pulmonary disease (COPD) patients. We therefore investigated the effects of prophylactic IFNß on respiratory epithelial cells infected with rhinovirus (RV). METHODS: A549 cells and primary bronchial epithelial cells (PBECs) were exposed for 18 h to IFNß. Then, IFNß was either removed or maintained in the supernatant for the rest of the experiment and cells were infected with RV-1B at t = 0 or 72 h after the initial exposure to IFNß. RESULTS: Viral RNA levels were decreased in both cell types. Furthermore, both viral RNA and infectious virus levels in the supernatant of infected A549 cells were still significantly reduced at 72 h after removal of IFNß. This pronounced antiviral pre-treatment effect was associated with increased expression of the antiviral genes IFN-stimulated protein of MR15000 (ISG15) and Myxovirus resistance 1 (Mx1) and the effect was maintained even when IFNß levels in the supernatant of A549 cells were undetectable. CONCLUSIONS: These data show that IFNß has not only a strong, but also a long-lasting protective effect against RV infection of respiratory epithelium.

CpG and poly(I:C) stimulation of dendritic cells and fibroblasts limits herpes simplex virus type 1 infection in an IFNß-dependent and -independent way.

Viral activation of toll-like receptors (TLRs) on dendritic cells (DCs) leads to production of various cytokines, including antiviral type I interferons (IFNs). Synthetic ligands specific for TLRs are also able to induce the production of type I IFNs (IFNα/ß) by DCs, suggesting that these ligands have potential as antiviral drugs. In this in vitro study we extensively investigated the antiviral activity of various TLR ligands. Mouse bone marrow (BM) cells were differentiated into plasmacytoid and conventional DCs (pDCs and cDCs), stimulated with various TLR ligands and tested the antiviral abilities of collected supernatants in an in vitro herpes simplex virus type 1 (HSV-1) infection model. We observed a significant IFNß-, (but not IFNα-) dependent reduction in HSV-1 infection when a mixed pDC/cDC population was stimulated with the TLR9 ligand CpG. In the absence of pDCs, TLR stimulation resulted in less pronounced antiviral effects. The most pronounced antiviral effect was observed when both DC subsets were stimulated with poly(I:C). A similar noticeable antiviral effect was observed when fibroblasts (L929 cells) were stimulated directly with poly(I:C). These poly(I:C)-mediated antiviral effects were only partially IFNß-mediated and probably TLR independent. These data demonstrate that TLR ligands are not only able to produce type I IFN but can indeed act as antiviral drugs. In particular poly(I:C), which exerts its antiviral effects even in the absence of DCs, may become a promising drug e.g. to prevent respiratory infections by topical intranasal application.