Diagnostic information Profiling and Evaluation of Causative Fungi of Fungal Keratitis Using High-throughput Internal Transcribed Spacer Sequencing

Abstract

Early and accurate diagnosis is essential for the targeted management of fungal keratitis (FK), which is one of the major blinding eye diseases worldwide. To elucidate the diagnostic information of high-throughput internal transcribed spacer (ITS) sequencing for identifying causative fungi of FK, 38 patients who were highly suspected of having FK were included in this research. In vivo confocal microscopy, potassium hydroxide smear, and fungal culture were performed to diagnose FK. Culture and ITS sequencing were used to identify causative fungi. We hypothesized that the dominant genus was the result of pathogen identification by ITS sequencing. Thirty-five patients were eventually diagnosed with FK, with fungal pathogens found by confocal microscopy in 27 patients (77.14%), by smear examination in 27 patients (77.14%), by culture in 25 patients (71.43%), and by ITS sequencing in 26 patients (74.29%). Eight causative fungal genera were determined by ITS sequencing, while five causative fungal genera were identified based on the morphology of the cultured pathogens. The results of ITS sequencing and culture were coincident in 10 patients with FK (28.57%). It is concluded that ITS sequencing, to some extent, challenged fungal culture and might be an optional complement in identifying fungal pathogens in corneas.

Profiling of Diagnostic Information of and Latent Susceptibility to Bacterial Keratitis From the Perspective of Ocular Bacterial Microbiota

“…High-Throughput 16S rDNA Sequencing Challenges Bacterial Culture in Identifying the Pathogenic Bacteria of BK Despite the worldwide recognition of culture in identifying microorganisms responsible for corneal infection (Austin et al, 2017;Eguchi et al, 2017), it has been facing an increasing challenge. According to our previous studies, microbiota abounding on the ocular surface of both healthy people and patients with non-infective eye disease undermines the reliability of microbial culture (Huang et al, 2016;Dong et al, 2019;Ge et al, 2019;Wang et al, 2019;Ren et al, 2020). High-throughput internal transcribed spacer (the unique barcode for fungal identification) sequencing also presents advantages over fungal culture for fungal identification in fungal keratitis (Ren et al, 2020).…”

“…According to our previous studies, microbiota abounding on the ocular surface of both healthy people and patients with non-infective eye disease undermines the reliability of microbial culture (Huang et al, 2016;Dong et al, 2019;Ge et al, 2019;Wang et al, 2019;Ren et al, 2020). High-throughput internal transcribed spacer (the unique barcode for fungal identification) sequencing also presents advantages over fungal culture for fungal identification in fungal keratitis (Ren et al, 2020). The inconformity of results between culture and highthroughput 16S rDNA sequencing seems to be within expect, but it does not mean high-throughput sequencing technology could FIGURE 2 | On the dimensionality of alpha diversities, six indices all suggest the sort and richness of ocular bacterial microbiota from the healthy eyes of BK patients had significant difference from the eyes of healthy people.…”

“…Bacterial culture is considered as the gold standard for etiological diagnosis of BK ( Sharma, 2012 ; Ren et al., 2020 ). However, this technique is becoming limited in the current diagnostic application for its frequent unsatisfying positive rates ( Ren et al., 2020 ).…”

“…Bacterial culture is considered as the gold standard for etiological diagnosis of BK ( Sharma, 2012 ; Ren et al., 2020 ). However, this technique is becoming limited in the current diagnostic application for its frequent unsatisfying positive rates ( Ren et al., 2020 ). Culture lacks specificity with its positivity for the healthy ocular surface being as high as 42% to 80% ( Willcox, 2013 ; Huang et al., 2016 ; Bograd et al., 2019 ).…”

Molecular Identification, Antioxidant Efficacy of Phenolic Compounds, and Antimicrobial Activity of Beta-Carotene Isolated from Fruiting Bodies of Suillus sp

“…In addition, ITS sequencing appears to be more cheaper with global availability than gene chipping, multi-locus sequencing or nanopore sequencing [8]. The researchers were interested in the field of natural products by extracting the active compounds from fruiting bodies of fungi, and studying the antimicrobial and antioxidant effects of these compounds against bacteria as well as fungi [9]. These natural compounds which contain many active substances were considered an important start for modern medicines and stimulated the interest of the companies specialized in manufacture of medicines, medical preparations, and cosmetics to use them as raw materials in their products.…”