Pretreatment of sinus aspirates with dithiothreitol improves yield of fungal cultures in patients with chronic sinusitis

Chisholm, Karen M.; Getsinger, D. L.; Vaughan, Winston C.; Hwang, Peter H.; Banaei, Niaz

doi:10.1002/alr.21230

Cited by 5 publications

(2 citation statements)

References 22 publications

(43 reference statements)

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“…Sodium pyrophosphate weakens the hydrophilic links in mucus allowing for viruses and bacteria to be separated [ 30 ]. Dithiothreitol (DTT) has been used on sinus samples to improve the yield of fungal cultures and in studies involving quantification of inflammatory cells in nasal secretions [ 31 – 33 ]. Sputasol contains DTT and has been used to liquefy mucus in nasal lavage [ 34 ].…”

Section: Discussionmentioning

confidence: 99%

Enumerating Virus-Like Particles and Bacterial Populations in the Sinuses of Chronic Rhinosinusitis Patients Using Flow Cytometry

et al. 2016

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There is increasing evidence to suggest that the sinus microbiome plays a role in the pathogenesis of chronic rhinosinusitis (CRS). However, the concentration of these microorganisms within the sinuses is still unknown. We show that flow cytometry can be used to enumerate bacteria and virus-like particles (VLPs) in sinus flush samples of CRS patients. This was achieved through trialling 5 sample preparation techniques for flow cytometry. We found high concentrations of bacteria and VLPs in these samples. Untreated samples produced the highest average bacterial and VLP counts with 3.3 ± 0.74 x 107 bacteria ml-1 and 2.4 ± 1.23 x 109 VLP ml-1 of sinus flush (n = 9). These counts were significantly higher than most of the treated samples (p < 0.05). Results showed 103 and 104 times inter-patient variation for bacteria and VLP concentrations. This wide variation suggests that diagnosis and treatment need to be personalised and that utilising flow cytometry is useful and efficient for this. This study is the first to enumerate bacterial and VLP populations in the maxillary sinus of CRS patients. The relevance of enumeration is that with increasing antimicrobial resistance, antibiotics are becoming less effective at treating bacterial infections of the sinuses, so alternative therapies are needed. Phage therapy has been proposed as one such alternative, but for dosing, the abundance of bacteria is required. Knowledge of whether phages are normally present in the sinuses will assist in gauging the safety of applying phage therapy to sinuses. Our finding, that large numbers of VLP are frequently present in sinuses, indicates that phage therapy may represent a minimally disruptive intervention towards the nasal microbiome. We propose that flow cytometry can be used as a tool to assess microbial biomass dynamics in sinuses and other anatomical locations where infection can cause disease.

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Section: Discussionmentioning

confidence: 99%

Enumerating Virus-Like Particles and Bacterial Populations in the Sinuses of Chronic Rhinosinusitis Patients Using Flow Cytometry

et al. 2016

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“…However, allergic fungal sinusitis (AFS) remains a significant entity and the role of fungi in the pathogenesis may indeed be broader, acting synergistically with bacterial biofilms. Chisolm et al re‐evaluate whether pretreatment of sinus aspirates with the mucolytic dithiothreitol (DTT) does indeed increase the fungal recovery rate on culture as had been initially published by Ponikau . Their study confirms that DTT does indeed increase the rate of fungal culture, more than doubling the recovery rate to 17.3%, but they did not reproduce the previously published very high recovery rates.…”

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confidence: 96%

Editorial

2013

Int Forum Allergy Rhinol

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Characterization of epithelial cells, connective tissue cells and immune cells in human upper airway mucosa by immunofluorescence multichannel image cytometry: a pilot study

Giotakis

Dudás

Glueckert

et al. 2020

Histochem Cell Biol

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Epithelial, connective tissue and immune cells contribute in various ways to the pathophysiology of chronic rhinosinusitis (CRS). However, data of their distribution in upper airway mucosa are sparse. We aimed to provide quantitative, purely informative data on the distribution of these cell lineages and their coexpression patterns, which might help identifying, e.g., cells in the epithelium undergoing through epithelial–mesenchymal transition (EMT). For this purpose, we used immunofluorescence multichannel image cytometry (IMIC). We examined fixed paraffin-embedded tissue samples (FFPE) of six patients with chronic rhinosinusitis (CRS) and of three patients without CRS (controls). The direct-conjugated antibodies pancytokeratin, vimentin and CD45/CD18 were used for coexpression analysis in epithelial layer and lamina propria. Image acquisition and analysis were performed with TissueFAXS and StrataQuest, respectively. To distinguish positive from negative expression, a ratio between cell-specific immunostaining intensity and background was developed. Isotype controls were used as negative controls. Per patient, a 4.5-mm2 tissue area was scanned and a median of 14,875 cells was recognized. The most common cell types were cytokeratin-single-positive (26%), vimentin-single-positive (13%) and CD45/CD18-single-positive with CD45/CD18–vimentin-double-positive cells (29%). In the patients with CRS, CD45/CD18-single-positive cells were 3–6 times higher compared to the control patients. In the epithelial layer, cytokeratin–vimentin-double-positive EMT cells were observed 3–5 times higher in the patients with CRS than in the control patients. This study provided quantitative data for the distribution of crucial cell types in CRS. Future studies may focus on the distribution and coexpression patterns of different immune cells in CRS or even cancer tissue.

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Pretreatment of sinus aspirates with dithiothreitol improves yield of fungal cultures in patients with chronic sinusitis

Abstract: This study confirms improved recovery of mold from sinus cultures after pretreatment of samples with dithiothreitol. Further studies are needed to correlate these findings with clinical outcome.

Cited by 5 publications

References 22 publications

Enumerating Virus-Like Particles and Bacterial Populations in the Sinuses of Chronic Rhinosinusitis Patients Using Flow Cytometry

Enumerating Virus-Like Particles and Bacterial Populations in the Sinuses of Chronic Rhinosinusitis Patients Using Flow Cytometry

Editorial

Characterization of epithelial cells, connective tissue cells and immune cells in human upper airway mucosa by immunofluorescence multichannel image cytometry: a pilot study

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