Christine Wennerås

Comparison of a new commercial test, Dermatophyte-PCR kit, with conventional methods for rapid detection and identification of Trichophyton rubrum in nail specimens.

The performance of a new commercially available duplex PCR, which combines pan-dermatophyte PCR with a Trichophyton rubrum-specific PCR, was evaluated. This Dermatophyte PCR kit, which requires one day for laboratory diagnosis, was compared with the conventional methods of microscopy and culture that necessitate up to 4 weeks for final diagnosis of dermatophytosis. We studied 177 nail samples from patients with suspected onychomycosis by fluorescence microscopy (blankophore), cultures and the Dermatophyte PCR kit. More samples were positive by PCR (78/177, 44%) than by culture (59/177, 34%). T. rubrum was present in 95% of all culture-positive nail specimens, which was confirmed by PCR in 55/56 specimens. The positive predictive value, negative predictive value, specificity and sensitivity of the duplex PCR was 93%, 87%, 94% and 85%, respectively, when confirmed by positive culture, microscopy or both. Due to its sensitivity, specificity and rapidity, we conclude that this PCR is an attractive method for routine investigation of nail dermatophytosis in a clinical setting.

Infections With the Tick-Borne Bacterium “Candidatus Neoehrlichia mikurensis” Mimic Noninfectious Conditions in Patients With B Cell Malignancies or Autoimmune Diseases

BACKGROUNDnCandidatus Neoehrlichia mikurensis is a newly discovered noncultivatable bacterium spread among ticks and rodents in Europe and Asia that can infect humans, particularly immunocompromised patients.nnnMETHODSnWe compiled clinical and laboratory data from 11 patients with hematological malignances or autoimmune diseases who were diagnosed with Candidatus N. mikurensis infection in Europe 2010-2013. Both published (6) and unpublished cases (5) were included.nnnRESULTSnThe patients had a median age of 67, were mostly male (8/11), and resided in Sweden, Switzerland, Germany, and the Czech Republic. All but one had ongoing or recent immune suppressive treatment and a majority were splenectomized (8/11). Less than half of them recalled tick exposure. The most frequent symptoms were fever (11/11), localized pain afflicting muscles and/or joints (8/11), vascular and thromboembolic events (6/11), that is, deep vein thrombosis (4), transitory ischemic attacks (2), pulmonary embolism (1), and arterial aneurysm (1). Typical laboratory findings were elevated C-reactive protein, leukocytosis with neutrophilia, and anemia. Median time from onset of symptoms to correct diagnosis was 2 months. In at least 4 cases, the condition was interpreted to be due to the underlying disease, and immunosuppressive therapy was scheduled. All patients recovered completely when doxycycline was administered.nnnCONCLUSIONSnCandidatus N. mikurensis is an emerging tick-borne pathogen that may give rise to a systemic inflammatory syndrome in persons with hematologic or autoimmune diseases that could be mistaken for recurrence of the underlying disease and/or unrelated arteriosclerotic vascular events. Awareness of this new pathogen is warranted among rheumatologists, hematologists, oncologists, and infectious disease specialists.

Comparative study of immune status to infectious agents in elderly patients with multiple myeloma, Waldenstrom's macroglobulinemia, and monoclonal gammopathy of undetermined significance.

ABSTRACT Whereas patients with multiple myeloma (MM) have a well-documented susceptibility to infections, this has been less studied in other B-cell disorders, such as Waldenstroms macroglobulinemia (WM) and monoclonal gammopathy of undetermined significance (MGUS). We investigated the humoral immunity to 24 different pathogens in elderly patients with MM (n = 25), WM (n = 16), and MGUS (n = 18) and in age-matched controls (n = 20). Antibody titers against pneumococci, staphylococcal alpha-toxin, tetanus and diphtheria toxoids, and varicella, mumps, and rubella viruses were most depressed in MM patients, next to lowest in WM and MGUS patients, and highest in the controls. In contrast, levels of antibodies specific for staphylococcal teichoic acid, Moraxella catarrhalis, candida, aspergillus, and measles virus were similarly decreased in MM and MGUS patients. Comparable titers in all study groups were seen against Haemophilus influenzae type b (Hib), borrelia, toxoplasma, and members of the herpesvirus family. Finally, a uniform lack of antibodies was noted against Streptococcus pyogenes, salmonella, yersinia, brucella, francisella, and herpes simplex virus type 2. To conclude, although MM patients displayed the most depressed humoral immunity, significantly decreased antibody levels were also evident in patients with WM and MGUS, particularly against Staphylococcus aureus, pneumococci, and varicella. Conversely, immunity was retained for Hib and certain herpesviruses in all study groups.

The chemoattractant Trp-Lys-Tyr-Met-Val-D-Met activates eosinophils through the formyl peptide receptor and one of its homologues, formyl peptide receptor-like 1

Whereas prokaryotes use L‐ and D‐isomers of amino acids in their protein synthesis, eukaryotic proteins as a rule incorporate only L‐isomers. Hence, D‐isomers may constitute danger signals to the innate immune system. A D‐methionine‐containing peptide, Trp‐Lys‐Tyr‐Met‐Val‐D‐Met‐NH2 (WKYMVm), has been shown to be a stronger activator of neutrophils than f‐Met‐Leu‐Phe. The aim of this study was to compare the responsiveness of eosinophils to WKYMVm with that of neutrophils. The peptide was found to induce chemotaxis and respiratory burst in eosinophils. However, it did not mobilize granule constituents, as evidenced by a lack of eosinophil cationic protein, eosinophil peroxidase, and interleukin‐5 in the supernatants of stimulated eosinophils. In contrast, WKYMVm caused the release of complement receptor 3 from secretory vesicles in neutrophils. Different members of the formyl peptide receptor family were preferentially engaged by the peptide in the two classes of granulocytes: the formyl peptide receptor itself in eosinophils and formyl peptide receptor‐like 1 in neutrophils.

How to interpret serum levels of beta-glucan for the diagnosis of invasive fungal infections in adult high-risk hematology patients: optimal cut-off levels and confounding factors

Detection of the fungal cell wall component beta-glucan (BG) in serum is increasingly used to diagnose invasive fungal infections (IFI), but its optimal use in hematology patients with high risk of IFI is not well defined. We retrospectively analyzed the diagnostic accuracy, optimal cut-off level, and potential confounding factors of BG reactivity. The inclusion criteria were: adult patients with hematologic disease who were admitted to the hematology ward during the 2-year study period and who had two or more consecutive BG assays performed. In total, 127 patients were enrolled. Thirteen patients with proven or probable IFI, as defined by the 2008 European Organization for Research and Treatment of Cancer/Mycoses Study Group (EORTC/MSG) criteria, were identified. Receiver operating characteristic (ROC) curve analysis showed a high overall diagnostic performance (area under the ROC curveu2009=u20090.98) and suggested an optimal cut-off level of 158xa0pg/ml, with a sensitivity and a specificity of 92xa0% and 96xa0%, respectively. Multiway analysis of variance indicated that treatment with pegylated asparaginase (pu2009<u20090.001), admission to the intensive care unit (ICU; pu2009=u20090.0007), and treatment with albumin, plasma, or coagulation factors (pu2009=u20090.01) are potential confounding factors of BG reactivity. We propose that a higher cut-off level than that recommended by the manufacturer should be used to monitor adult hematology patients at high risk for IFI. Our results also suggest that elevated BG levels in patients treated with pegylated asparaginase, albumin, plasma, or coagulation factors, or those admitted to the ICU should be interpreted with caution.

Allergen extracts directly mobilize and activate human eosinophils

Allergic diseases are characterized by the presence of eosinophils, which are recruited to the affected tissues by chemoattractants produced by T cells, mast cells and epithelium. Our objective was to evaluate if allergens can directly activate human eosinophils. The capacity of purified allergen extracts to elicit eosinophil chemotaxis, respiratory burst, degranulation and up‐regulationof the adhesion molecule complement receptor 3 (CR3) was determined in eosinophils isolated from healthy blood donors. Eosinophils stimulated with an extract from house dust mite (HDM) released thegranule protein major basic protein (MBP) and up‐regulated the surface expression of CR3. Cat allergen extracts also induced the up‐regulation of CR3, but not the release of MBP; instead cat, as well as birch and grass allergens, elicited the release of eosinophil peroxidase (EPO). In addition, grass pollen extract caused the secretion of MBP. None of the allergens stimulated eosinophilic cationic protein release, nor production of free oxygen radicals. Both HDM and birch extracts were chemotactic for eosinophils. These findings establish that common aeroallergens can directly activate eosinophils in vitro. We propose that eosinophil activation in vivo is not exclusively mediated by cytokines and chemokines of the allergic inflammatory reaction, but could partly be theresult of direct interaction between allergens and eosinophils.

A chair of one's own.

The upper reaches of academe remain stubbornly inaccessible to women.

Multi-locus sequence analysis (MLSA) of clinical “CandidatusNeoehrlichia mikurensis” strains from Europe

ABSTRACT “Candidatus Neoehrlichia mikurensis” is the tick-borne agent of neoehrlichiosis, an infectious disease that primarily affects immunocompromised patients. So far, the genetic variability of “Ca. Neoehrlichia” has been studied only by comparing 16S rRNA genes and groEL operon sequences. We describe the development and use of a multilocus sequence analysis (MLSA) protocol to characterize the genetic diversity of clinical “Ca. Neoehrlichia” strains in Europe and their relatedness to other species within the Anaplasmataceae family. Six genes were selected: ftsZ, clpB, gatB, lipA, groEL, and 16S rRNA. Each MLSA locus was amplified by real-time PCR, and the PCR products were sequenced. Phylogenetic trees of MLSA locus relatedness were constructed from aligned sequences. Blood samples from 12 patients with confirmed “Ca. Neoehrlichia” infection from Sweden (n = 9), the Czech Republic (n = 2), and Germany (n = 1) were analyzed with the MLSA protocol. Three of the Swedish strains exhibited identical lipA sequences, while the lipA sequences of the strains from the other nine patients were identical to each other. One of the Czech strains had one differing nucleotide in the clpB sequence from the sequences of the other 11 strains. All 12 strains had identical sequences for the genes 16S rRNA, ftsZ, gatB, and groEL. According to the MLSA, among the Anaplasmataceae, “Ca. Neoehrlichia” is most closely related to Ehrlichia ruminantium, less so to Anaplasma phagocytophilum, and least to Wolbachia endosymbionts. To conclude, three sequence types of infectious “Ca. Neoehrlichia” were identified: one in the west of Sweden, one in the Czech Republic, and one spread throughout Europe.

Reply to Raoult

TO THE EDITOR—We share Dr Raoult’s opinion that most likely it will be possible to cultivate Candidatus Neoehrlichia mikurensis in the future, as has been the case for other species of the Anaplasmataceae family. The statement in the abstract of our manuscript [1] that the bacterium is not cultivable simply referred to the fact that nobody has reported its culture since the first description of its 16S ribosomal RNA sequence 15 years ago [2]. In the introductory section of our manuscript, we specified the current status of Candidatus Neoehrlichia mikurensis [3] as an uncultured bacterium and cited the initial studies that reported on the same bacterium under different names (references 10–15 of our article [1]). It should be emphasized that unlike these earlier publications, the study of Kawahara et al [3] not only detected the bacterium by polymerase chain reaction in ticks, but also found it in mammalian hosts (wild rats on Mikura Island, Japan) and even showed that Candidatus Neoehrlichia mikurensis was transferable to laboratory rats. The culture of Candidatus Neoehrlichia mikurensis would allow for the removal of “Candidatus” from its name, the development of serological assays (eg, indirect fluorescent antibody tests) similar to those used for the diagnosis of anaplasmosis and ehrlichiosis, and analysis of its pathogenicity and the mechanisms of control by the immune system.However, already now it is possible