bluuebunny/crossref_metadata_2025_split huggingface dataset:
P417 Evaluation of matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS)-Bruker Biotyper Sirius for identification of invasive molds
Authors: Dalvi Mousami , Patwardhan Sampada , Prayag Parikshit , Purandhare Bharat , Soman Rajeev 
Published: September 2022

Abstract

Poster session 3, September 23, 2022, 12:30 PM - 1:30 PM

Objectives
Susceptibility to various antifungal drugs varies between different species and subspecies within the same genus. Phenotypic identification of fungi has limitations for species-level identification. Correct identification of species and subspecies in invasive mold infections is important to initiate the appropriate antifungal therapy. Matrix Assisted Laser Desorption Ionization Time-of-Flight mass spectrometry (MALDI-TOF MS) with its proteomic analysis overcomes this limitation and helps in administering the correct anti-fungal therapy. A total of seven mold isolates from invasive fungal infections were evaluated for identification by MALDI-TOF MS and conventional morphological methods.

Methods
Total of seven isolates from invasive mold infections were identified by the conventional method of culturing specimens on Sabouraud's dextrose agar and Potato Dextrose agar with incubation at room temperature and 37°C in Biological oxygen demand (BOD) incubator. Micro-morphological identification of the fungus was done by Lacto Phenol Cotton Blue (LPCB) mount. Same isolates were processed on MALDI-TOF MS Bruker Biotyper Sirius (Bruker Daltonics, Bremen Germany) following recommended extraction protocol using ethanol absolute, acetonitrile, and 70% formic acid.

Results
As per the below Figure.

Conclusion
In four out of seven isolates phenotypic identification upto species level based on LPCB micromorphology was confirmed on MALDI-TOF MS. In the remaining three isolates we could only give a genus level identification based on LPCB mount. These three isolates were further identified upto the level of species after processing on MALDI as Aspergillus tamarii, Phaeoacremonium cinerum, and Fusarium equiseti. All mold isolates were identified with good quality mass spectra. In our experience, mold identification by MALDI-TOF MS using the Bruker Biotyper Sirius platform definitely has an edge over conventional phenotypic methods in species-level differentiation of various molds, impacting targeted antifungal management.
P419 Galactomannan enzyme immunoassay in invasive pulmonary aspergillosis: a cross sectional study in 2021 in a New Delhi
Authors: Das Sutapa 
Published: September 2022

Abstract

Poster session 3, September 23, 2022, 12:30 PM - 1:30 PM

Objective
This study aims to determine the role of GM EIA in the diagnosis of IPA as per revised EORTC/MSG 2019 criteria.

Material and Methods
A prospective cross-sectional study was performed on 87 children (from 1 to 12 years) admitted to pediatric hematology ward from Oct 2020 to February 2022. Serum of these patients was collected and GM EIA was performed using BIO-RAD Platelia™ Aspergillus Ag. Clinical, mycological workup (potassium-hydroxide mount, fungal culture) was done and furthermore, these patients were classified into proven, probable, and possible IA as per EORTC-MSG guidelines, 2019. Galactomannan indices (GMI) measured in terms of sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) were validated with revised EORTC -MSG, 2019 of IPA.

Results
A total of 53 patients out of 87 were categorized into probable IPA with routine mycological findings. On culture Aspergillus flavus was the most common pathogen, identified in 56% (25), followed by A. fumigatus 31.8% (14), A. niger 9% (4), etc. 44 (50.5%) had GMI value of &gt;0.7, 9 (10.3%) in the range of 0.5-0.7 and 34 (39%) had GMI &lt;0.5.
The maximal Youden index ‘J’ [95% confidence interval (CI) of 0.4167-0.6875] was calculated as 0.678 and sensitivity, specificity, PPV, and NPV in IPA were calculated corresponding to a GM index of &gt;1.0, 0.7, 0.5 to determine the best cutoff point. The best cutoff was found to be 0.695 where in the 84.1% and specificity was 83.7% respectively.

Conclusion
GM EIA can prove as an excellent diagnostic test for IPA when done in addition to culture from nonsterile sites in high-risk populations like pediatric patients with hematological malignancy. This study reinforces the definition of the probable category of EORTC-MSG criteria, 2019 in overall diagnosis, prognosis, and management of IPA in pediatric patients with hematological malignancy.
P420 Fungal detection by means of HCR using 2D-Covalent Organic Framework Nanosheet
Authors: Gajjar Devarshi , Baghel Manoj Kumar , Thakkar Harshil , Thakore Sonal 
Published: September 2022

Abstract

Poster session 3, September 23, 2022, 12:30 PM - 1:30 PM
The timely diagnosis of fungal infections is of prime importance for prescribing appropriate anti-fungal drugs. Current methods for fungal diagnosis involve culture-based methods, antibody-based detection using lateral flow assays and RT-PCR. In the present work, we devised a non-enzymatic amplification using 2D-Covalent Organic Framework (COF) nanosheet for the detection of fungal DNA.

Objectives
(1) Validation of exfoliated 2D COF Nanosheet as an efficient DNA detection tool via Hybridization chain reaction (HCR) triggered fluorescent assay. (2) Sequence retrieval and probe generation of fungal sample and detection of extracted target DNA via fluorescent assay.

Method
A novel COF was synthesized and characterization was done using FTIR, BET, TGA, XRD, and SEM. Probes for the detection of fungi (Candida, Aspergillus, and Mucor) were designed using NUPACK software. HCR was monitored for different time and probe concentrations and standardized reaction was used for the detection of fungal RNA.

Results
FTIR, BET, TGA, XRD, and SEM confirmed the structure and formation of COF-nanosheet. H1, H2 probes at a concentration of 1μm and in presence of Target DNA (0.001μm-1μm) showed HCR reaction at 1.5 h. Fluorescence quenching was observed when probes were mixed with both, bulk COF and COF-nanosheets but increased quenching

Conclusion
Fungal detection can be done by means of HCR using the Covalent nanosheets.
P422 EQUAL PCP Score 2022—an ECMM score derived from current guidelines to measure QUAlity of clinical Pneumocystis Pneumonia management
Authors: Grothe Jan 
Published: September 2022

Abstract

Poster session 3, September 23, 2022, 12:30 PM - 1:30 PM

Background
Pneumocystis pneumonia (PCP) is a life-threatening opportunistic fungal infection requiring complex clinical management. Guidelines assist clinicians but can be challenging to comply with.

Objectives
To develop a scoring tool, that facilitates and quantifies adherence to guideline recommendations for PCP.

Methods
We reviewed current PCP guidelines and determined essential recommendations for diagnosis, treatment, and follow-up. These were weighted according to their strength of recommendation and level of evidence.

Results
The EQUAL PCP Score 2022 consists of 22 items. For diagnosis, weight was given to bronchoalveolar lavage and immunofluorescence assays as the gold standard for sampling and analysis. Beta-D-glucan was considered of similar importance due to its high negative predictive value. Trimethoprim/sulfamethoxazole and the addition of corticosteroids in respiratory failure got 3 points respectively. Alternative approaches received less points and the use of aerosolized pentamidine was discouraged with 1 minus point. HIV-specific considerations such as the start of secondary prophylaxis were factored in as well.

Conclusion
The EQUAL PCP Score 2022 weighs and aggregates factors recommended for optimal management of PCP. It provides a tool for antifungal stewardship as well as for measuring guideline adherence but remains to be correlated with patient outcomes.
P424 Comparative analysis of Galactomannan Lateral Flow Assay, Galactomannan Enzyme Immunoassay and BAL culture for diagnosis of COVID-19-associated pulmonary aspergillosis
Authors: Hedayati Mohammad , Ghazanfari Mona , Charati Jamshid Yazdani , Davoodi Lotfollah , Arastehfar Amir , Moazeni Maryam , Abastabar Mahdi , Haghani Iman , Mayahi Sabah , Hoenigl Martin , Pan Weihua 
Published: September 2022

Abstract

Poster session 3, September 23, 2022, 12:30 PM - 1:30 PM

Objectives
Invasive pulmonary aspergillosis is a well-known complication in COVID-19 patients. A reliable and early diagnosis is essential for the early initiation of antifungal therapy. On the other hand, conventional Bronchoalveolar lavage (BAL) culture may lack the specificity of differentiating between colonization and infection. Here, we aimed a comparative analysis of Galactomannan Lateral Flow Assay (GM-LFA), Galactomannan Enzyme Immunoassay (GM-EIA), and BAL culture for diagnosis of COVID-19-associated pulmonary aspergillosis (CAPA).

Methods
BAL (n = 105) and serum samples (n = 101) from COVID-19 patients who were mechanically ventilated for ≥4 days in intensive care units (ICUs) were evaluated by GM-LFA and GM-EIA. All BAL samples were cultured on Sabouraud-Chloramphenicol dextrose Agar. Diagnostic performance of GM-LFA and GM-EIA in BAL (GM indexes ≥1) and serum (GM indexes &gt;0.5) were evaluated and sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) and Roc curve compared with BAL culture calculated.

Results
The Aspergillus GM-LFA for serum and BAL samples showed a sensitivity of 46.9% and 54.5%, specificity of 100% and 91.7%, PPV of 100% and 75%, and NPV of 80.2% and 81.5%, when compared with BAL culture, respectively. GM-EIA showed sensitivities of 56.3% and 60.6%, specificities of 94.2% and 88.9%, PPVs of 81.8% and 71.4%, and NPVs of 82.3% and 83.1% for serum and BAL samples, respectively.

Conclusion
According to our results BAL GM detection using both EIA and LFA may be a promising approach for early diagnosis of CAPA and differentiating between colonization and invasive infection.
P428 Evaluation of Galactomannan enzyme assay in non-hematological non-neutropenic ICU admitted patients for diagnosis of Invasive Pulmonary Aspergillosis as per EORTC criteria 2019
Authors: Kashyap Sanchi , Capoor Malini R. , Lavanya P. , Gupta Poonam , Verma P.K. , Talwar Vandana , Sachdeva Harish Chand 
Published: September 2022

Abstract

Poster session 3, September 23, 2022, 12:30 PM - 1:30 PM

Objective
The objective of this study was to evaluate galactomannan enzyme assay in non-hematological ICU patients for diagnosis of Invasive Pulmonary Aspergillosis as per EORTC criteria 2019.

Methods
Galactomannan detection in serum samples was performed by enzyme-linked immunosorbent assay in 138 patients with clinically suspected pulmonary aspergillosis between January 2012 and January 2019 in patients admitted to ICUs. Patients with hematological diseases or who underwent HSCT were excluded. Control group (n = 25) was selected from patients with no obvious immunosuppression, cough/dyspnea, or any other respiratory symptoms, etc admitted for elective surgeries from surgery wards. According to the criteria of the European Organization for Research and Treatment of Cancer/Mycoses Study Group revised in 2019, the patients were categorized as proven, probable, and possible IA. Galactomannan optical density indices ≥0.5 and ≥1 were analyzed with respect to clinical, radiological, and mycological evidence.

Results
Amongst 138 patients 2 fulfilled the criteria of proven IA, 78 were probable IA, and the rest (58) were grouped under possible IA. In the control group, the Galactomannan optical density index was ≤ 0.5. Receiver operating characteristic curve analysis showed the serum GM detection cutoff value was 0.91 (95% CI 0.885-0.973, P-value &lt;.0001) with Youden index J = 0.78, its diagnostic value for pulmonary aspergillosis was optimized, and the sensitivity and specificity reached 83.75% and 94.83% respectively. Other cutoffs had high variance between sensitivity and specificity for the diagnosis of IPA.

Conclusion
Our study highlights the usefulness of the serum GM antigen test in the early diagnosis of IA and suggests a GMI cutoff of 0.91 as it has the highest diagnostic accuracy. Also, we recommend that patients with a GM OD of ≥1.0 should be labeled as clear GM positive, whereas those with &lt;0.5 OD should be labeled as GM negative, and in those with OD ranging from 0.5 to 1.0, repeat sampling from the patient is advisable. It seems more reasonable not to overtreat all patients with fever refractory to broad-spectrum antibiotics with antifungal agents, but rather to decide taking into account the serum GM antigen positivity. A negative GM antigen can curtail the usage of unwarranted antifungal therapy, especially in resource-poor country like India.
P431 Tip to correct the variation position error in applying long-read high-throughput sequencing technology for fungal identification
Authors: Langsiri Nattapong , Worasilchai Navaporn , Irinyi Lazlo , Meyer Wieland , Chindamporn Ariya 
Published: September 2022

Abstract

Poster session 3, September 23, 2022, 12:30 PM - 1:30 PM

Presently, long-read high-throughput sequencing technology has been started applying in the medical mycology field instead of common Sanger sequencing. With its capability to sequence nucleic acid among the large region compared to Sanger sequencing, this technology can detect and analyze both inter- and intra-species sequence variation in internal transcribed spacer (ITS), a fungal house-keeping region, resulting in high accurate fungal identification. However, to set up the long-read high-throughput sequencing technology for fungal identification is quite challenging because of several factors related to the accuracy including the variation position error. So, in this study, we summarize the tip to correct the variation position error in applying long-read high-throughput sequencing technology for fungal identification that we have learnt in the preliminary laboratory setting using the 8 clinical isolates of yeast: Candida albicans (n = 2), C. tropicalis (n = 1), C. glabrata (n = 1), Trichosporon asahii (n = 2), Pichia kudriavzevii (n = 1), Cryptococcus neoformans var. grubii (n = 1). Based on the recruited strains, we found that self-assembling reference sequence generated from raw data of reading by using an auto-program named Canu causes the size-inflated sequence, a larger size calculated as 22.83 ± 7.56% than it should be, resulting in the shift of variation position. This error can be corrected by the alignment process of the reference sequence with the known sequence, both size and position, prior to doing the raw read alignment. The advantage of this process could correct not only for position shifting caused by the analysis process but also the random error generated from nanopore system. To validate this correction protocol, more samples are needed for further study.
P434 Anti fungal susceptibility of Malassezia to azoles by broth microdilution method and their phylogenetic relationship based on multi-locus sequence analysis
Authors: Romald Packia Nancy , Kindo Anupma Jyoti , Veereraghavan Mahalakshmi , Umadevi U. 
Published: September 2022

Abstract

Poster session 3, September 23, 2022, 12:30 PM - 1:30 PM

Objectives

Method
Part 1—Epidemiological pattern, phenotypic Identification, and anti-fungal susceptibility pattern by broth microdilution method
Part 2-Molecular Identification, Sangers Sequencing, Phylogenetic Analysis and Dendogram
Molecular Identification

Results
Figure 1 Epidemiological pattern, phenotypic Identification, and AFST by broth microdilution
Figure 2 Molecular Identification, Sangers Sequencing, Phylogenetic Analysis, and Dendrogram

Conclusions
P436 A study to compare detection of Aspergillus galactomannan in patients with clinically suspected invasive Aspergillosis using the Lateral Flow Assay (LFA) in comparison with Enzyme Linked Immunosorbent Assay (ELISA) and identification of new risk factors as per BM-AspICU criteria
Authors: Padaki Priyadarshini , Savio Jayanthi , Ramachandran Priya , Krishna Bhuvana 
Published: September 2022

Abstract

Poster session 3, September 23, 2022, 12:30 PM - 1:30 PM

Objectives

Methods
A total of 50 consecutive leftover samples (30 blood and 20 BAL) received for testing by Galactomannan assay by ELISA method were included. LFA and ELISA were performed as per manufacturers’ instructions. Results were recorded using the Digital Reader provided which gives readings as Positive ≥0.5 and &lt;0.5 Negative. Additionally, results were read visually by two different people and graded as 0 (negative), 1+ (very faint band), 2+ (faint band), and 3+ (strong band). The values obtained by ELISA were reported as Galactomannan Index. Results of LFA and ELISA were compared. Patient profiles were studied to identify the possible and probable cases of IA as per BM-AspICU guidelines.

Results
Overall, 92% samples (n = 46) were positive with ELISA. Results of LFA and ELISA were comparable in 68% samples (n = 34). For all except two samples, visual readings of LFA (5 with 3+, 12 with 2 + and 19 with 1 + readings) correlated with digital readings. Sensitivity of LFA was 73.9% (95% CI). Applying BM-AspICU criteria, 28% (n = 14) were categorized as probable, 64% (n = 32) as possible cases of IA and 8% (n = 4) as NO IA. A total of 12 patients had classical host factors, 42% (n = 21) had other risk factors and we were able to identify 16% (n = 8) cases with new risk factors which were CNS involvement/brain injury/hemorrhage on mechanical ventilation. Clinical features were present in all of these 58% (n = 29) cases whereas radiological imaging was done in only 25 cases out of which 28% (n = 14) were positive. BM-AspICU criteria require ≥2 mycological criteria, but only two of the BAL samples were culture positive for Aspergillus species. These criteria could not be strictly applied to most patients in our study since invasive procedures such as BAL was not done, instead, blood was sent for galactomannan detection.
The average turn-around time was 3-7 days for ELISA since the test was run twice a week whereas it was 5-24 h for the LFA. The pre-processing of samples and time required to perform the procedure for both the assays are similar. The cost per test for ELISA was Rs.1675/- as against LFA for Rs.750/-.

Conclusion
LFA for galactomannan detection is an easier to perform, cost-effective method for diagnosis of IA as compared with ELISA. LFA overcomes the difficulty of insufficient volume of calibrators and controls provided with the ELISA kit. Useful especially for patients admitted in ICU where reports can be issued on the same day instead of batch testing. Identification of new risk factors and inclusion of the same under guidelines is essential to suspect and send samples for testing for early initiation of treatment.
P440 General protocol applied to the identification and production of new biomarkers with potential use for the diagnosis of histoplasmosis
Authors: Arias Juan David Puerta , Isaza Juan Pablo , Moreno Ernesto , Berrio Indira , Restrepo Luz Elena Cano , Naranjo Tonny 
Published: September 2022

Abstract

Poster session 3, September 23, 2022, 12:30 PM - 1:30 PM

Objective
To identify and produce novel biomarkers with potential use for the specific diagnosis of H. capsulatum infection.

Methods
Here, we design a novel strategy to search and select new Candidate genes for biomarkers that integrates the use of a computational analysis model that includes the application of bioinformatic tools such as OrthoMCL, BLASTp, TargetP, and SignalP, applied on a local collection of proteome database obtained manually from GenBank-NCBI, and the analysis of previously published biological and experimental data sets, including a secreted proteome database obtained from pathogenic yeast-phase H. capsulatum culture filtrates, a Histoplasma yeast and mycelial transcriptomes database, and a urine-peptides database from Histoplasma-immunoassay-positive patients.
For the synthesis of the Candidates, an internal protocol for the production of recombinant proteins in prokaryotic and eukaryotic systems was applied. Obtaining polyclonal antibodies (PAb) specific for each biomarker was carried out by adapting a rapid immunization protocol for BALB/c mice.
Finally, the computational model was experimentally validated, evaluating the reactivity and specificity of PAb anti-Histoplasma with fungus culture extracts and samples from patients with histoplasmosis.

Results
Using the computational analysis model, 2 Candidate genes for diagnostic biomarkers were identified. Subsequently, the construction of expression vector for each Candidate and the production of these genes were achieved using a standardized protocol for the production of recombinant proteins.
Polyclonal antibodies (PAb) anti-histoplasma were obtained and shown to be reactive against purified H. capsulatum-antigens. Finally, we confirmed the presence of these antigens in yeast culture extracts of H. capsulatum and demonstrated the immunoreactivity of anti-Histoplasma PAb with urine samples from patients previously diagnosed with histoplasmosis.

Conclusion
The generation of novel strategies that combine data analysis, computational tools, and transcriptomic and proteomic techniques could be very useful for the identification of new biomarker genes and the development of microbiological diagnostic tests for important pathogens.
P444 Detection of causative agents of infectious keratitis in patients from western rajasthan
Authors: Shrimali Twishi , Sharma Anuradha , Ghosh Anup , Morya Arvind , Tak Vibhor , Nag Vijayalakshmi 
Published: September 2022

Abstract

Poster session 3, September 23, 2022, 12:30 PM - 1:30 PM

Objective
To determine the spectrum of causative agents, the related risk factors, and their association in patients of infectious keratitis.

Methodology
It was a prospective study conducted over a period of 18 months from August 2018 to January 2020, which included 100 patients attending the Ophthalmology OPD with features of keratitis. Ophthalmological examination was followed by corneal scrapings’ collection, which were subjected to culture, microscopy, and molecular diagnostic tools. Bacterial isolates were identified by conventional methods and MicroScan Walkaway system while the isolated fungi were identified conventionally. Pan-fungal primers were used to detect fungal elements directly from the sample.

Results
Out of 100, 41 cases were positive by culture, of which 32 (78.04%) had fungal and nine (21.95%) had bacterial keratitis. Fusarium spp. accounted for 33.33% of fungal and Pseudomonas aeruginosa accounted for 55.55% of the bacterial isolates. Fungal material was detected in 41% using pan fungal primers. Cases were maximally recorded during July-October. Traumatic history was present in 78% patients caused by vegetative matter (49%). A male preponderance (67%) was also observed. Four patients underwent evisceration in spite-of rigorous management.

Conclusion
Poor prognosis emphasizes the need for faster diagnostics, which can detect the causative agents from the clinical specimen itself, reinforcing the concept of clinical metagenomics.