Clinical Case of Rhino-Orbital Mucormycosis in a Convalescent Covid-19 Patient: Diagnostic and Treatment Tactics

According to current data, SARS-CoV-2 virus has the ability to cause multi-organ pathology, leading to acute damage of various organs and systems and long-term consequences characterized by polymorphic symptoms. Recently, a high incidence of invasive mycoses, particularly mucormycosis - COVID-M, has been noted among the COVID-19 complications. The predisposing factor for the development of this pathology is diabetes mellitus, immunodeficiency states, and prolonged use of high doses of glucocorticosteroids. Mucormycosis is characterized by severe clinical manifestations and high lethality, and timely diagnostics of this pathology often represents a difficult problem. The aim of this study was to analyze a clinical case of rhino-orbital mucormycosis in convalescent COVID-19 patient. In the study, there was used mucopurulent nasal discharge from the patient previously hospitalized with a severe novel coronavirus infection. Here, we describe the methodology allowing to isolate and identify a pure mold fungus culture from the biomaterial using methods of routine bacteriology and MALDI-ToF mass spectrometry. Direct microscopy examination of nasal cavity discharge revealed branched non-septic hyphae with a characteristic branching angle, allowing to preliminarily diagnose invasive mucormycosis. Growth of mycelial fungus colony was observed by using Sabouraud's medium with potassium tellurite. Microscopy of the pure culture revealed branching mycelium without septa, broad, with irregular thickness, unsegregated hyphae, and sporangia with a typical column specific to mucormycetes. Analysis of the obtained mass spectra allowed to establish the microbial species identity as Lichtheimia corymbifera. The latter along with other members of the order Mucorales, are known to cause mucormycosis. As a result of antifungal treatment (Amphotericin B) and timely surgical intervention, the patient was discharged from the hospital with prominent clinical improvement and no complaints during further outpatient follow-up period. The analysis of this clinical case showed the lack of alertness in some clinical diagnostic laboratories to detect pathogens of invasive mycoses. To avoid errors, while making a diagnosis, attention should be paid not only to detection of fungal spores in clinical material, but also take into account the structure of mycelium underlying major difference between yeast-like fungi, higher and lower molds. The isolation and identification of a pure pathogen culture allows to confidently verify the diagnosis, timely correct the treatment tactics and monitor circulation of mycotic agents to prevent occurrence of mycoses in most vulnerable patients cohorts.

A multiplex method for detection of SARS-CoV-2 variants based on MALDI-TOF mass spectrometry.

The recent outbreak of the coronavirus disease 2019 (COVID-19) pandemic and the continuous evolution of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have highlighted the significance of new detection methods for global monitoring and prevention. Although quantitative reverse transcription PCR (RT-qPCR), the current gold standard for diagnosis, performs excellently in genetic testing, its multiplexing capability is limited because of the signal crosstalk of various fluorophores. Herein, we present a highly efficient platform which combines 17-plex assays with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS), enabling the targeting of 14 different mutation sites of the spike gene. Diagnosis using a set of 324 nasopharyngeal swabs or sputum clinical samples with SARS-CoV-2 MS method was identical to that with the RT-qPCR. The detection consistency of mutation sites was 97.9% (47/48) compared to Sanger sequencing without cross-reaction with other respiratory-related pathogens. Therefore, the MS method is highly potent to track and assess SARS-CoV-2 changes in a timely manner, thereby aiding the continuous response to viral variation and prevention of further transmission.

Structural and Immunoreactivity Properties of the SARS-CoV-2 Spike Protein upon the Development of an Inactivated Vaccine.

Inactivated vaccines are promising tools for tackling the COVID-19 pandemic. We applied several protocols for SARS-CoV-2 inactivation (by ß-propiolactone, formaldehyde, and UV radiation) and examined the morphology of viral spikes, protein composition of the preparations, and their immunoreactivity in ELISA using two panels of sera collected from convalescents and people vaccinated by Sputnik V. Transmission electron microscopy (TEM) allowed us to distinguish wider flail-like spikes (supposedly the S-protein's pre-fusion conformation) from narrower needle-like ones (the post-fusion state). While the flails were present in all preparations studied, the needles were highly abundant in the ß-propiolactone-inactivated samples only. Structural proteins S, N, and M of SARS-CoV-2 were detected via mass spectrometry. Formaldehyde and UV-inactivated samples demonstrated the highest affinity/immunoreactivity against the convalescent sera, while ß-propiolactone (1:2000, 36 h) and UV-inactivated ones were more active against the sera of people vaccinated with Sputnik V. A higher concentration of ß-propiolactone (1:1000, 2 h) led to a loss of antigenic affinity for both serum panels. Thus, although we did not analyze native SARS-CoV-2 for biosafety reasons, our comparative approach helped to exclude some destructive inactivation conditions and select suitable variants for future animal research. We believe that TEM is a valuable tool for inactivated COVID-19 vaccine quality control during the downstream manufacturing process.

CLINICAL CASE OF RHINO-ORBITAL MUCORMYCOSIS IN A CONVALESCENT COVID-19 PATIENT: DIAGNOSTIC AND TREATMENT TACTICS.

According to current data, SARS-CoV-2 virus has the ability to cause multi-organ pathology, leading to acute damage of various organs and systems and long-term consequences characterized by polymorphic symptoms. Recently, a high incidence of invasive mycoses, particularly mucormycosis - COVID-M, has been noted among the COVID-19 complications. The predisposing factor for the development of this pathology is diabetes mellitus, immunodeficiency states, and prolonged use of high doses of glucocorticosteroids. Mucormycosis is characterized by severe clinical manifestations and high lethality, and timely diagnostics of this pathology often represents a difficult problem. The aim of this study was to analyze a clinical case of rhino-orbital mucormycosis in convalescent COVID-19 patient. In the study, there was used mucopurulent nasal discharge from the patient previously hospitalized with a severe novel coronavirus infection. Here, we describe the methodology allowing to isolate and identify a pure mold fungus culture from the biomaterial using methods of routine bacteriology and MALDIToF mass spectrometry. Direct microscopy examination of nasal cavity discharge revealed branched non-septic hyphae with a characteristic branching angle, allowing to preliminarily diagnose invasive mucormycosis. Growth of mycelial fungus colony was observed by using Sabouraud’s medium with potassium tellurite. Microscopy of the pure culture revealed branching mycelium without septa, broad, with irregular thickness, unsegregated hyphae, and sporangia with a typical column specific to mucormycetes. Analysis of the obtained mass spectra allowed to establish the microbial species identity as Lichtheimia corymbifera. The latter along with other members of the order Mucorales, are known to cause mucormycosis. As a result of antifungal treatment (Amphotericin B) and timely surgical intervention, the patient was discharged from the hospital with prominent clinical improvement and no complaints during further outpatient follow-up period. The analysis of this clinical case showed the lack of alertness in some clinical diagnostic laboratories to detect pathogens of invasive mycoses. To avoid errors, while making a diagnosis, attention should be paid not only to detection of fungal spores in clinical material, but also take into account the structure of mycelium underlying major difference between yeast-like fungi, higher and lower molds. The isolation and identification of a pure pathogen culture allows to confidently verify the diagnosis, timely correct the treatment tactics and monitor circulation of mycotic agents to prevent occurrence of mycoses in most vulnerable patients cohorts. (English) [ FROM AUTHOR] Согласно современным данным вирус SARS-CoV-2 обладает способностью вызывать полиорганную патологию, приводя к острым повреждениям различных органов и систем и долгосрочным последствиям, характеризующимся полиморфной симптоматикой. Ð’ последнее время среди осложнений COVID-19 отмечается высокая распространенность инвазивных микозов, в частности мукормикоза — COVID-M. Предрасполагающим фактором развития данной патологии являются сахарный диабет, иммунодефицитные состояния, длительное применение высоких доз глюкокортикостероидов. Мукормикоз отличается тяжестью клинических проявлений и высокой летальностью, своевременная диагностика данной патологии нередко является сложной проблемой. Целью настоящего исследования стал анализ клинического случая риноорбитального мукормикоза у больной, перенесшей COVID-19. Ð’ качестве материала для исследования послужило слизисто-гнойное отделяемое полости носа больной, находившейся ранее на стационарном лечении с диагнозом «Новая коронавирусная инфекция тяжелого течения». Ð’ статье приведена методика, позволяющая выделить и идентифицировать чистую культуру плесневого гриба из биоматериала с использованием методов классической бактериологии и MALDI-ToF массспектрометрии. При исследовании отделяемого носовой полости методом прямой микроскопии были обнаружены разветвленные несептированные гифы с характерным углом ветвления, что позволило поставить предварительный диагноз «Инвазивный мукормикоз». При использовании среды Сабуро с теллуритом калия был отмечен рост колонии мицелиального гриба. При микроскопии чистой культуры были обнаружены типичные для мукормицетов ветвистый мицелий без перегородок, широкие, неравномерные по толщине, несептированные гифы и спорангии с типичной колонкой. Анализ полученных масс-спектров позволил установить видовую принадлежность исследуемого микроорганизма: Lichtheimia corymbifera. Как известно, лихтеймии, наряду с другими представителями порядка Mucorales, являются возбудителями мукормикозов. Врезультате проводимого лечения противогрибковым препаратом (Амфотерицин Ð’) и своевременного оперативного вмешательства больная была выписана из стационара со значительным улучшением, при дальнейшем амбулаторном наблюдении жалоб не Ð¿Ñ€ÐµÐ´Ñ ÑÐ²Ð»ÑÐ»Ð°. Анализ данного клинического случая показал отсутствие ориентированности некоторых клинико-диагностических лабораторий на обнаружение возбудителей инвазивных микозов. Чтобы избежать ошибок при постановке диагноза, необходимо обращать внимание не только на обнаружение спор грибов в клиническом материале, но и принимать во внимание строение мицелия, что является основным различием между дрожжеподобными грибами, высшими и низшими плесенями. Выделение и идентификация чистой культуры возбудителя позволяет уверенно верифицировать диагноз, своевременно корректировать тактику лечения и осуществлять наблюдение за циркуляцией возбудителей мукормикозов для предотвращения возникновения микозов у особо уязвимых контингентов больных. (Russian) [ FROM AUTHOR] Copyright of Russian Journal of Infection & Immunity is the property of National Electronic-Information Consortium and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full . (Copyright applies to all s.)