Cystic fibrosis (CF) is the most common inherited genetic disease in Caucasian populations. Besides bacteria, many species of fungi may colonize the respiratory tract of these patients, sometimes leading to true respiratory infections. In this study, an oligonucleotide array capable of identifying 20 fungal species was developed to directly detect fungi in the sputum samples of CF patients. Species-specific oligonucleotide probes were designed from the internal transcribed spacer (ITS) regions of the rRNA operon and immobilized on a nylon membrane. The fungal ITS regions were amplified by PCR and hybridized to the array for species identification. The array was validated by testing 182 target strains (strains which we aimed to identify) and 141 nontarget strains (135 species), and a sensitivity of 100% and a specificity of 99.2% were obtained. The validated array was then used for direct detection of fungi in 57 sputum samples from 39 CF patients, and the results were compared to those obtained by culture. For 16 sputum samples, the results obtained by the array corresponded with those obtained by culture. For 33 samples, the array detected more fungal species than culture did, while the reverse was found for eight samples. The accuracy of the array for fungal detection in sputum samples was confirmed (or partially confirmed) in some samples by cloning and resequencing the amplified ITS fragments. The present array is a useful tool for both the simultaneous detection of multiple fungal species present in the sputa of CF patients and the identification of fungi isolated from these patients.

Objective. Anti-double-stranded DNA (antidsDNA) antibodies may contribute to the pathogenesis of glomerulonephritis (GN) by cross-reacting with ␣-actinin in murine models and in some patients with systemic lupus erythematosus (SLE). We therefore sought to determine possible disease associations with serologic and clinical features and to characterize this new autoantibody specificity.Methods. One hundred patients with SLE were recruited into this multicenter study, as well as 100 rheumatic disease controls and 2,100 healthy blood donors. Clinical disease was evaluated by the SLE Disease Activity Index (SLEDAI; excluding the anti-DNA component). Anti-dsDNA antibodies were detected by conventional enzyme-linked immunosorbent assay (ELISA) and by a commercial enzyme immunoassay (EIA). Anti-␣-actinin antibodies were detected by ELISA, and their specificity was confirmed by Western blotting and by indirect immunofluorescence using rat kidney sections and mesangial cells as substrates. Highly positive sera were selected for absorption experiments and were affinity-purified for cross-reactivity studies and measurement of antibody avidity.Results. Sera from 62 of the SLE patients had anti-dsDNA antibodies; 21 of these sera also had anti-␣-actinin antibodies, as compared with 1 of the 38 sera without anti-dsDNA antibodies. Of the 22 patients with anti-␣-actinin antibodies, 10 had GN, as compared with 14 of the 78 without anti-␣-actinin antibodies (P < 0.01). In patients with GN, anti-␣-actinin, but not anti-dsDNA, antibodies correlated with the SLEDAI score (minus the anti-DNA component) and with treatment. The fraction of serum anti-dsDNA antibodies that cross-reacted with ␣-actinin exhibited high avidity for dsDNA, as determined using a commercial EIA for high-avidity anti-dsDNA antibodies and an in-house conventional ELISA.Conclusion. The ␣-actinin-binding antibodies are significantly associated with GN in SLE. Whether such autoantibodies may anticipate the development of this complication of SLE remains to be verified.

Using SYSMEX analyzers, Pv, Po, and Pm infections are easy to ascertain as DIFF scattergrams are almost specific (specificity = 99.9%). Pf infection diagnosis by CBC may be a more promising tool.

In murine systemic lupus erythematosus (SLE) models, nephritogenic anti-dsDNA IgG has been shown to cross-react with a kidney antigen, alpha-actinin, and to be critical in renal pathogenesis. In humans, studies of anti-alpha-actinin antibodies (Abs) are scarce, and these antibodies remain to be evaluated. We have thus far tested sera from patients with SLE (n = 103), rheumatoid arthritis (RA, n = 93), and primary Sjögren syndrome (pSS, n = 34), and from healthy subjects (n = 160), for the presence of anti-alpha-actinin and anti-DNA Abs. The latter were tested using several methods [IIF on Crithidia luciliae (Crit) and ELISA using dsDNA]. Anti-alpha-actinin Abs were confirmed by Western blot. Sera from 23 of 103 SLE patients, 3 of 93 RA patients, 1 of 33 pSS patients, and 1 of 160 controls scored positive for anti-alpha-actinin Abs. In SLE, the positivity was significantly associated with anti-dsDNA reactivity (22 of 23): 19 of 23 sera were alpha-actinin-positive/dsDNA-positive and 13 were alpha-actinin-positive/Crit-positive. Few cases were alpha-actinin-positive/dsDNA-negative: 1 SLE, 3 RA, and 1 control. Furthermore, anti-alpha-actinin Abs have been detected at high level before or at the early stage of lupus nephritis when compared with active and inactive SLE without kidney manifestations.

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