Summary A systematic review and meta-analysis was done on the use of PCR tests for the diagnosis of invasive aspergillosis. Data from more than 10 000 blood, serum, or plasma samples obtained from ...1618 patients at risk for invasive aspergillosis were retrieved from 16 studies. Overall, the mean diagnostic odds ratios (DORs) of PCR for proven and probable cases were similar whether two consecutive positive samples were required to define positivity (DOR 15·97 95% CI 6·83–37·34) or a single positive PCR test was required (DOR 16·41 95% CI 6·43–41·88). Sensitivity and specificity of PCR for two consecutive positive samples were 0·75 (95% CI 0·54–0·88) and 0·87 (95% CI 0·78–0·93), respectively, and if only a single positive sample was required, these values were 0·88 (95% CI 0·75–0·94) and 0·75 (95% CI 0·63–0·84), respectively. Whereas specificity based on a single positive test was significantly lower (p=0·027) than two positive tests, the sensitivity and DOR did not differ significantly. A single PCR-negative result is thus sufficient to exclude a diagnosis of proven or probable invasive aspergillosis. However, two positive tests are required to confirm the diagnosis because the specificity is higher than that attained from a single positive test. Populations at risk varied and there was a lack of homogeneity of the PCR methods used. Efforts are underway to devise a standard for Aspergillus sp PCR for screening, which will help enable formal validation of PCR and estimate its use in patients most likely to benefit.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UL, UM, UPUK
Aspergillus (A.) fumigatus is an opportunistic fungal mold inducing invasive aspergillosis (IA) in immunocompromised patients. Although antifungal activity of human natural killer (NK) cells was ...shown in previous studies, the underlying cellular mechanisms and pathogen recognition receptors (PRRs) are still unknown. Using flow cytometry we were able to show that the fluorescence positivity of the surface receptor CD56 significantly decreased upon fungal contact. To visualize the interaction site of NK cells and A. fumigatus we used SEM, CLSM and dSTORM techniques, which clearly demonstrated that NK cells directly interact with A. fumigatus via CD56 and that CD56 is re-organized and accumulated at this interaction site time-dependently. The inhibition of the cytoskeleton showed that the receptor re-organization was an active process dependent on actin re-arrangements. Furthermore, we could show that CD56 plays a role in the fungus mediated NK cell activation, since blocking of CD56 surface receptor reduced fungal mediated NK cell activation and reduced cytokine secretion. These results confirmed the direct interaction of NK cells and A. fumigatus, leading to the conclusion that CD56 is a pathogen recognition receptor. These findings give new insights into the functional role of CD56 in the pathogen recognition during the innate immune response.
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
The COVID-19 pandemic has placed a huge strain on global healthcare and been a significant cause of increased morbidity and mortality, particularly in at-risk populations. This disease attacks the ...respiratory systems and causes significant immune dysregulation in affected patients creating a perfect opportunity for the development of invasive fungal disease (IFD). COVID-19 infection can instill a significant, poorly regulated pro-inflammatory response. Clinically induced immunosuppression or pro-inflammatory damage to mucosa facilitate the development of IFD and
, Mucorales, and
infections have been regularly reported throughout the COVID-19 pandemic. Corticosteroids and immune modulators are used in the treatment of COVID-19. Corticosteroid use is also a risk factor for IFD, but not the only reason for IFD in COVID -19 patients. Specific dysregulation of the immune system through functional exhaustion of Natural killer (NK) cells and T cells has been observed in COVID-19 through the expression of the exhaustion markers NK-G2A and PD-1. Reduced fungicidal activity of neutrophils from COVID-19 patients indicates that immune dysfunction/imbalance are important risk factors for IFD. The COVID-19 pandemic has significantly increased the at-risk population for IFD. Even if the incidence of IFD is relatively low, the size of this new at-risk population will result in a substantial increase in the overall, annual number of IFD cases. It is important to understand how and why certain patients with COVID-19 developed increased susceptibility to IFD, as this will improve our understanding of risk of IFD in the face of future pandemics but also in a clinical era of increased clinical immuno-suppression/modulation.
Antifungal Drug Resistance Loeffler, Juergen; Stevens, David A.
Clinical infectious diseases,
01/2003, Volume:
36, Issue:
Supplement-1
Journal Article
Peer reviewed
The increasing incidence of invasive fungal infections is the result of many factors, including an increasing number of patients with severe immunosuppression. Although new drugs have been introduced ...to combat this problem, the development of resistance to antifungal drugs has become increasingly apparent, especially in patients who require long-term treatment or who are receiving antifungal prophylaxis, and there is growing awareness of shifts of flora to more-resistant species. The frequency, interpretation, and, in particular, mechanism of resistance to current classes of antifungal agents, particularly the azoles (where resistance has climbed most prominently) are discussed in this review.
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BFBNIB, NUK, PNG, UL, UM, UPUK
PCR has been used as an aid in the diagnosis of invasive aspergillosis for almost 2 decades. A lack of standardization has limited both its acceptance as a diagnostic tool and multicenter clinical ...evaluations, preventing its inclusion in disease-defining criteria. In 2006, the European Aspergillus PCR Initiative was formed. The aim of the initiative was to provide optimal standardized protocols for the widespread clinical evaluation of the Aspergillus PCR to determine its diagnostic role and allow inclusion in disease diagnosis criteria. Quality control panels were developed and circulated to centers for evaluation of the existing methodology before recommendations based on the initial results were proposed for further panels. The centers were anonymously classified as "compliant" or "noncompliant," according to whether they had followed the proposed recommendations before the performance parameters were determined and meta-regression analysis was performed. Most PCR amplification systems provided similar detection thresholds, although positivity was a function of the fungal burden. When PCR amplification was combined with DNA extraction, 50% of the centers failed to achieve the same level of detection. Meta-regression analysis showed positive correlations between sensitivity and extraction protocols incorporating the proposed recommendations and the use of bead beating, white cell lysis buffer, and an internal control PCR. The use of elution volumes above 100 μl showed a negative correlation with sensitivity. The efficiency of the Aspergillus PCR is limited by the extraction procedure and not by PCR amplification. For PCR testing of whole blood, it is essential that large blood volumes (≥3 ml) be efficiently lysed before bead beating to disrupt the fungal cell and performance of an internal control PCR to exclude false negativity. DNA should be eluted in volumes of <100 μl.
High-throughput RNA sequencing (RNA-seq) is routinely applied to study diverse biological processes; however, when performed separately on interacting organisms, systemic noise intrinsic to RNA ...extraction, library preparation, and sequencing hampers the identification of cross-species interaction nodes. Here, we develop triple RNA-seq to simultaneously detect transcriptomes of monocyte-derived dendritic cells (moDCs) infected with the frequently co-occurring pulmonary pathogens Aspergillus fumigatus and human cytomegalovirus (CMV). Comparing expression patterns after co-infection with those after single infections, our data reveal synergistic effects and mutual interferences between host responses to the two pathogens. For example, CMV attenuates the fungus-mediated activation of pro-inflammatory cytokines through NF-κB (nuclear factor κB) and NFAT (nuclear factor of activated T cells) cascades, while A. fumigatus impairs viral clearance by counteracting viral nucleic acid-induced activation of type I interferon signaling. Together, the analytical power of triple RNA-seq proposes molecular hubs in the differential moDC response to fungal/viral single infection or co-infection that contribute to our understanding of the etiology and, potentially, clearance of post-transplant infections.
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•Triple RNA-seq measures gene expression of co-infected immune cells•Gene correlation networks reveal different hub gene sets under co-infection•Co-infection expression includes synergies and interferences between host and pathogens•Molecular basis of viral/fungal pulmonary infection has potential for the clinic
Seelbinder et al. demonstrate simultaneous sequencing of RNA isolated from human immune cells infected with Aspergillus fumigatus and cytomegalovirus, two pulmonary pathogens regularly affecting the lungs of immunosuppressed patients. They detect characteristic gene expression patterns for single and co-infections and reveal synergistic virulence strategies between the two pathogens.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPUK, ZAGLJ, ZRSKP
The saprophytic fungus Aspergillus fumigatus reproduces by generation of conidia, which are spread by airflow throughout nature. Since humans are inhaling certain amounts of spores every day, the ...(innate) immune system is constantly challenged. Even though macrophages and neutrophils carry the main burden, also NK cells are regarded to contribute to the antifungal immune response. While NK cells reveal a low frequency, expression and release of immunomodulatory molecules seem to be a natural way of their involvement.
In this study we show, that NK cells secrete chemokines such as CCL3/MIP-1α, CCL4/MIP-1β and CCL5/RANTES early on after stimulation with Aspergillus fumigatus and, in addition, adjust the concentration of chemokines released to the multiplicity of infection of Aspergillus fumigatus.
These results further corroborate the relevance of NK cells within the antifungal immune response, which is regarded to be more and more important in the development and outcome of invasive aspergillosis in immunocompromised patients after hematopoietic stem cell transplantation. Additionally, the correlation between the multiplicity of infection and the expression and release of chemokines shown here may be useful in further studies for the quantification and/or surveillance of the NK cell involvement in antifungal immune responses.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
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