Mucormicosis oral asociada a COVID-19 y diabetes mellitus: descripción de un caso / Oral mucormycosis associated with COVID-19 and diabetes mellitus: Case report and literature review

La mucormicosis es una infección fúngica oportunista e invasiva, con una elevada tasa de mortalidad. Se ha detectado principalmente en pacientes con COVID-19, especialmente en personas con enfermedades concomitantes como la diabetes mellitus. La prevalencia de las mucormicosis es de 0,005 a 1,7 casos por millón de habitantes y ha ido en aumento en países como India y Pakistán; puede afectar diferentes órganos y su forma clínica refleja el mecanismo de transmisión. Entre las formas frecuentes están la rino-orbital-cerebral y la pulmonar, por ello, debe sospecharse mucormicosis en los pacientes con lesiones necróticas en mucosas o piel. Se presenta el caso de un paciente con antecedentes de diabetes mellitus que fue diagnosticado con mucormicosis oral asociada a la COVID-19.

Mucormycosis is an invasive opportunistic fungal infection with high mortality, mainly detected in people with COVID-19, especially those with underlying diseases such as diabetes mellitus. Mucormycosis prevalence is 0.005 to 1.7 cases per million inhabitants, and it has been increasing in countries like India and Pakistan. This mycosis can affect different organs, and clinical manifestations reflect the transmission mechanism. Frequent forms are rhino-orbital-cerebral and pulmonary. This disease should be suspected in patients with necrotic injuries on mucous membranes or skin. We present a case of a patient with diabetes mellitus and diagnosed with oral mucormycosis associated with COVID-19.

In vitro combination with doxycycline plus antifungals against clinical Mucorales pathogens.

PURPOSE: Since systematic antifungals for mucormycosis showed variable MICs depending on strains, effective and safe antifungal therapy was still needed. This study is aimed to evaluate the in vitro activity of doxycycline combined with antifungal therapy against dominant Mucorales pathogens. METHODS: Multidrug susceptibility testing was performed with doxycycline and antifungals, including itraconazole, posaconazole, and amphotericin, in 21 isolates of 8 dominant Mucorales pathogens. RESULTS: The fractional inhibitory concentration index according to M38 showed one Rhizopus arrhizus isolate synergic (∑FICI = 0.375) and other isolates in addition (0.5 < ∑FICI < 4). CONCLUSIONS: Doxycycline was found to have in vitro advantages in combined antifungal treatment over antifungals alone.

Silencing of Amylomyces rouxii aspartic II protease by siRNA to increase tyrosinase activity.

Amylomyces rouxii is a zygomycete that produces extracellular protease and tyrosinase. The tyrosinase activity is negatively regulated by the proteases and, which attempts to purify the tyrosinase (tyr) enzyme that has been hampered by the presence of a protease that co-purified with it. In this work we identified genes encoding aspartic protease II (aspII) and VI of A. rouxii. Using an RNAi strategy based on the generation of a siRNA by transcription from two opposite-orientated promoters, the expression of these two proteases was silenced, showing that this molecular tool is suitable for gene silencing in Amylomyces. The transformant strains showed a significant attenuation of the transcripts (determined by RT-qPCR), with respective inhibition of the protease activity. In the case of aspII, inhibition was in the range of 43-90 % in different transformants, which correlated well with up to a five-fold increase in tyr activity with respect to the wild type and control strains. In contrast, silencing of aspVI caused a 43-65 % decrease in protease activity but had no significant effect on the tyr activity. The results show that aspII has a negative effect on tyr activity, and that the silencing of this protease is important to obtain strains with high levels of tyr activity.

COVID-19 associated rhinomaxillary mucormycosis: a case series and review of literature

During COVID-19 pandemic, fulminant deep fungal infection started emerging in India, known as Mucormycosis. This type of mucormycosis was termed as COVID-19 associated mucormycosis (CAM). These patients had previous history of COVID-19 infection. Such cases were mainly reported in immunocompromised patients such as patients with poorly controlled diabetes and chronic renal diseases etc. Rhinomaxillary mucormycosis is an aggressive, fulminant, fatal deep fungal infection of head and neck region. Early diagnosis and prompt treatment can reduce the mortality and morbidity associated with the disease; hence we present case series of rhinomaxillary mucormycosis to create awareness amongst dental surgeons

Maxilectomia em paciente diabética tipo II, diagnosticada com Mucormicose rinomaxilar, com posterior reabilitação protética: relato de caso / Maxillectomy in a type II diabetic patient, diagnosed with rhinomaxillary mucormycosis, with subsequent prosthetic rehabilitation: case report / Maxilectomía en paciente diabética tipo II diagnosticada de mucormicosis rinomaxilar, con posterior rehabilitación protésica: reporte de caso

A mucormicose é uma infecção fúngica angioinvasiva que afeta uma ampla faixa etária, geralmente imunodeprimidos, sem predileção por gênero ou raça e com alta taxa de mortalidade. Essa infecção inicia se no nariz, devido à inalação dos esporos, podendo-se espalhar pelos seios paranasais, órbita e estruturas intracranianas. As características clínicas incluem parestesia perinasal, celulite periorbitária, rinorréia, obstrução nasal, epistaxe e diminuição de peso. O tratamento efetivo dessa comorbidade compoe uma combinação de manejo clínico e medicamentoso, conjuntamente com desbridamento cirúrgico radical do tecido infectado e/ou necrótico. O caso clínico descrito nesse trabalho refere-se a uma paciente diagnosticada com Mucormicose rino maxilar e diabetes do tipo II, a qual foi submetida a procedimento cirúrgico de hemimaxilectomia do lado esquerdo com posterior reabilitação protética para selamento de comunicação buco-naso-sino-etmoidal... (AU)

Mucormycosis is an angioinvasive fungal infection that affects a wide age group, usually immunocompromised, with no gender or race predilection, and with a high mortality rate. This infection starts in the nose, due to the inhalation of spores, and can spread through the paranasal sinuses, orbit and intracranial structures. Clinical features include perinasal paresthesia, periorbital cellulitis, rhinorrhea, nasal obstruction, epistaxis and weight loss. Effective treatment of this comorbidity comprises a combination of clinical and drug management, together with radical surgical debridement of infected and/or necrotic tissue. The clinical case described in this work refers to a patient diagnosed with Mucormycosis Rhinomaxilla and Type II diabetes, who underwent a surgical procedure of left hemimaxillectomy with subsequent prosthetic rehabilitation for sealing of the bucco nasal-sino-ethmoidal communication... (AU)

La mucormicosis es una micosis angioinvasiva que afecta a un amplio grupo de edad, habitualmente inmunodeprimidos, sin predilección de género ni raza, y con una alta tasa de mortalidad. Esta infección comienza en la nariz, debido a la inhalación de esporas, y puede extenderse a través de los senos paranasales, la órbita y las estructuras intracraneales. Las características clínicas incluyen parestesia perinasal, celulitis periorbitaria, rinorrea, obstrucción nasal, epistaxis y pérdida de peso. El tratamiento eficaz de esta comorbilidad comprende una combinación de manejo clínico y farmacológico, junto con un desbridamiento quirúrgico radical del tejido infectado y / o necrótico. El caso clínico descrito en este trabajo se refiere a una paciente diagnosticada de Mucormicosis Rinomaxilar y diabetes Tipo II, que fue sometida a un procedimiento quirúrgico de hemimaxilectomía izquierda con posterior rehabilitación protésica para sellar la comunicación buco-nasal-sino-etmoidal... (AU)

Recent developments in less known and multi-resistant fungal opportunists.

Fungal infections have increased in recent years due to host factors, such as oncohaematological and transplant-related disorders, immunosuppressive therapy, and AIDS. Additionally, molecular and proteomic facilities have become available to identify previously unrecognizable opportunists. For these reasons, reports on less-known and recalcitrant mycoses, such as those caused by black fungi, hyaline filamentous fungi, coelomycetes, Mucorales, and non-Candida yeasts have emerged. In this review, novel taxonomy in these groups, which often are multi-resistant to one or several classes of antifungals, is discussed. Clinical presentations, diagnosis and current treatment of some major groups are summarised.

Mucormycosis at a tertiary-care center in Mexico. A 35-year retrospective study of 214 cases.

BACKGROUND: Mucormycosis is a rare, invasive disease associated with high mortality rates, produced by opportunistic pathogens related to the Mucorales order and characterised by a diverse range of clinical forms; acute rhino-orbital-cerebral and pulmonary symptoms are the most reported ones. OBJECTIVES: To report the experience of mucormycosis observed in a tertiary-care hospital in Mexico for 35 years. METHODS: This was a retrospective, descriptive and observational study on mucormycosis at a tertiary-care hospital in Mexico from January 1985 to December 2019. Demographic and clinical data and mycological and histopathological records were selected. RESULTS: Two hundred fourteen proven cases of mucormycosis for 35 years at a tertiary-care hospital in Mexico were included. Most of the cases were male patients with a median age of 45 years. The two most associated underlying diseases were diabetes mellitus (76.6%) and haematologic malignancy (15.4%). The three primary clinical forms were as follows: rhino-orbito-cerebral (75.9%), cutaneous (8.41%) and pulmonary (7.47%) mucormycosis. The most isolated agents were Rhizopus arrhizus (58.4%) and Lichtheimia corymbifera (12.3%). The overall therapeutic response was 58.5%, and the best response was observed with amphotericin B deoxycholate and surgical debridement. CONCLUSION: Mucormycosis is an emerging disease, and its incidence has increased at our hospital over the years. In this study, the rhino-cerebral clinical type was the most frequent in patients with uncontrolled diabetes; the main aetiological agent was R. arrhizus. Early diagnosis, control of the underlying disease and prompt management may increase the survival rate.

Rhino-orbital mucormycosis due to Apophysomyces ossiformis in a patient with diabetes mellitus: a case report.

BACKGROUND: The most common aetiological agents of mucormycosis are Rhizopus, Mucor, Apophysomyces and Lichtheimia. Apophysomyces is comparatively rare, as it has been reported in less than 3% of mucormycosis cases. The genus Apophysomyces includes six species, and only A. elegans, A. mexicanus, A. variabilis and A. ossiformis have been reported to cause infections in both immunocompetent and immunocompromised patients. CASE PRESENTATION: We present a case of a 46-year-old male patient with bilateral blepharoedema, corneal opacity in the left eye and poorly controlled diabetes mellitus. The patient was subjected to total maxillectomy, exenteration of the left orbit and treatment with liposomal amphotericin B. Direct mycological analysis with KOH 10% revealed hyaline, coenocytic, long and wide hyphae. Apophysomyces ossiformis was identified from maxillary biopsy using 18S-ITS1-5.8S-ITS2-28S rRNA gene amplification and sequencing. The patient requested to be transferred to another hospital to continue treatment, where he died on the ninth day after admittance. CONCLUSION: To the best of our knowledge, this is the first case of rhino-orbital mucormycosis due to A. ossiformis with a fatal outcome. This case reveals the need to identify the fungus causing mucormycosis with molecular methods to identify adequate treatment therapies for patients with this infection.

First reported case of Gilbertella persicaria in human stool: outcome of a community study from Segamat, Johor, Malaysia.

Species of fungi belonging to the order Mucorales can be found everywhere in the environment. Gilbertella persicaria, which belongs to this order, have often been isolated from fruits and in water systems. However, there has been no report of isolation of this fungus from human samples. During a gut mycobiome study, from the Segamat community, Gilbertella persicaria was isolated from a human fecal sample and was characterized through a series of morphological assessment, biochemical tests, and molecular techniques. The isolate produced a white velvety surface that turned grayish after 24 h. Although no biofilm production was observed, the results indicated that the isolate could form calcium oxalate crystals, produced urease, and was resistant to low pH. The isolate was sensitive to amphotericin but resistant to voriconazole and itraconazole. The features of this fungus that could help in its survival in the human gut are also discussed.

Biotechnological Strategies for Chitosan Production by Mucoralean Strains and Dimorphism Using Renewable Substrates.

We investigated the influence of corn steep liquor (CSL) and cassava waste water (CWW) as carbon and nitrogen sources on the morphology and production of biomass and chitosan by Mucor subtilissimus UCP 1262 and Lichtheimia hyalospora UCP 1266. The highest biomass yields of 4.832 g/L (M. subtilissimus UCP 1262) and 6.345 g/L (L. hyalospora UCP 1266) were produced in assay 2 (6% CSL and 4% CWW), factorial design 22, and also favored higher chitosan production (32.471 mg/g) for M. subtilissimus. The highest chitosan production (44.91 mg/g) by L. hyalospora (UCP 1266) was obtained at the central point (4% of CWW and 6% of CSL). The statistical analysis, the higher concentration of CSL, and lower concentration of CWW significantly contributed to the growth of the strains. The FTIR bands confirmed the deacetylation degree of 80.29% and 83.61% of the chitosan produced by M. subtilissimus (UCP 1262) and L. hyalospora (UCP 1266), respectively. M. subtilissimus (UCP 1262) showed dimorphism in assay 4-6% CSL and 8% CWW and central point. L. hyalospora (UCP 1266) was optimized using a central composite rotational design, and the highest yield of chitosan (63.18 mg/g) was obtained in medium containing 8.82% CSL and 7% CWW. The experimental data suggest that the use of CSL and CWW is a promising association to chitosan production.

A method for the extraction of high quality fungal RNA suitable for RNA-seq.

Transcriptomic analysis is an OMICs technology that is becoming indispensable to understand and get a complete picture of cell functioning and adaptation to the environmental cues the cell is continuously receiving. Among the techniques available to perform transcriptomics, RNA-seq is becoming the method of choice. The quality of the RNA used for the generation of cDNA libraries and subsequent sequencing is crucial for the success of the process. Good RNA-seq performance is often limited by problems such as low RNA yield and/or integrity, RNA stability, and contamination with DNA, salts or chemicals. RNA isolation from fungi usually faces these problems and is particularly sensitive to degradation due to the high RNase activity content present in many species. Here we describe the development of a robust, highly reproducible and simple RNA purification method for filamentous fungi, which combines various strategies to get fully DNA-free RNA samples of high purity and integrity without the need to use a DNase I digestion step. The obtained RNA samples complied with all required standards to be used for RNA-seq and showed an excellent performance when subjected to Illumina-HiSeq 2500.

A Collagenolytic Aspartic Protease from Thermomucor indicae-seudaticae Expressed in Escherichia coli and Pichia pastoris.

Proteases are produced by the most diverse microorganisms and have a wide spectrum of applications. However, the use of wild microorganisms, mainly fungi, for enzyme production has some drawbacks. They are subject to physiological instability due to metabolic adaptations, causing complications and impairments in the production process. Thus, the objective of this work was to promote the heterologous expression of a collagenolytic aspartic protease (ProTiN31) from Thermomucor indicae seudaticae in Escherichia coli and Pichia pastoris. The pET_28a (+) and pPICZαA vectors were synthesized containing the gene of the enzyme and transformed into E. coli and P. pastoris, respectively. The recombinant enzymes produced by E. coli and P. pastoris showed maximum activity at pH 5.0 and 50 °C, and pH 5.0 and 60 °C, respectively. The enzyme produced by P. pastoris showed better thermostability when compared to that produced by E. coli. Both enzymes were stable at pH 6.0 and 6.5 for 24 h at 4 °C, and sensitive to pepstatin A, ß-mercaptoethanol, and Hg2+. Comparing the commercial collagen hydrolysate (Artrogen duo/Brazil) and gelatin degradation using protease from P. pastoris, they showed similar peptide profiles. There are its potential applications in a wide array of industrial sectors that use collagenolytic enzymes.

Production of cellulases by Thermomucor indicae-seudaticae: characterization of a thermophilic ß-glucosidase.

The current study evaluated the production and characterization of ß-glucosidase by the thermophilic fungus Thermomucor indicae-seudaticae in solid-state fermentation of wheat bran. Isolated fungi have significant amounts of ß-glucosidase, an enzyme that may be applied to different industrial processes, such as the production of fuels, food, and other chemical compounds. Maximal enzyme activity occurred in pH 3.5-4.5 and at 70 °C. The enzyme exhibited high thermostability, for 1 h, up to 60 °C, and good tolerance to glucose (10 mM) and ethanol (10%). The optimization of fermentative parameters on the production of ß-glucosidase was carried out by evaluating the best supplementary nutrient source, pH of nutrient solution, initial substrate moisture and fermentation temperature. The optimization of the above fermentation parameters increased enzyme activity by 120.0%. The highest enzymatic activity (164.0 U/g) occurred with wheat bran containing 70% initial moisture, supplemented with 1.0% (NH4)2SO4 solution at pH 5.5-6.0 and fungus incubated at 40 °C. A more detailed study of ß-glucosidase suggested that Sulfur is an important component of the main amino acid present in this enzyme. The enhancer of the enzyme activity occurred when the fungus was grown on wheat bran supplemented with a sulfur-containing solution. In fact, increasing the concentration of sulfur in the solution increased its activity.

In Vitro Activity of Combinations of Zinc Chelators with Amphotericin B and Posaconazole against Six Mucorales Species.

Mucormycosis is an emerging disease with high mortality rates. Few antifungal drugs are active against Mucorales. Considering the low efficacy of monotherapy, combination-therapy strategies have been described. It is known that fungi are susceptible to zinc deprivation, so we tested the in vitro effect of the zinc chelators clioquinol, phenanthroline, and N,N,N',N'-tetrakis(2-pyridylmethyl)ethane-1,2-diamine combined with amphotericin B or posaconazole against 25 strains of Mucorales. Clioquinol-posaconazole was the most active combination, although results were strain dependent.

Conversion of renewable substrates for biosurfactant production by Rhizopus arrhizus UCP 1607 and enhancing the removal of diesel oil from marine soil

BACKGROUND: The use of agro-industrial wastes to produce high value-added biomolecules such as biosurfactants is a promising approach for lowering the total costs of production. This study aimed to produce biosurfactants using Rhizopus arrhizus UCP 1607, with crude glycerol (CG) and corn steep liquor (CSL) as substrates. In addition, the biomolecule was characterized, and its efficiency in removing petroderivatives from marine soil was investigated. RESULTS: A 22 factorial design was applied, and the best condition for producing the biosurfactant was determined in assay 4 (3% CG and 5% CSL). The biosurfactant reduced the surface tension of water from 72 to 28.8 mN/m and produced a yield of 1.74 g/L. The preliminary biochemical characterization showed that the biosurfactant consisted of proteins (38.0%), carbohydrates (35.4%), and lipids (5.5%). The compounds presented an anionic character, nontoxicity, and great stability for all conditions tested. The biomolecule displayed great ability in dispersing hydrophobic substrates in water, thereby resulting in 53.4 cm2 ODA. The best efficiency of the biosurfactant in removing the pollutant diesel oil from marine soil was 79.4%. CONCLUSIONS: This study demonstrated the ability of R. arrhizus UCP1607 to produce a low-cost biosurfactant characterized as a glycoprotein and its potential use in the bioremediation of the hydrophobic diesel oil pollutant in marine soil

Micosis rino-orbito-cerebral: coinfección por Aspergillus sclerotiorum y Rhizopus microsporum en un huésped inmunosuprimido. Reporte de un caso / Rhino-orbital-cerebral mycoses: Aspergillus sclerotiorum infection and Rhyzopus microsporum in an immunosuppres sed host. A case report

Las micosis rino-cerebral, rino-orbito-cerebral y sinusopulmonar ocasionadas por especies de Mucorales y de Aspergillus se constituyen como una causa importan te de infección fúngica invasora asociada a una mortalidad mayor al 30%. La coinfección por dos o más especies en la forma rino-orbito-cerebral es infrecuente. Se describe un paciente con linfoma no Hodgkin, expuesto a múltiples esquemas quimioterápicos, en remisión completa, que presentó micosis con compromiso rino-orbito-cerebral por Aspergillus sclerotiorum y Rhizopus microsporum de rápida progresión con necesidad de tratamiento quirúrgico agresivo y terapia anti fúngica sistémica y local.

Rhino-orbital-cerebral and pulmonary mycosis caused by Mucorales and Aspergillus species have become an important cause of invasive fungal infection, with a 30% overall mortality rate. Rhino-orbital-cerebral disease caused by two or more species is uncommon. We present a patient with non-Hodgkin lymphoma, ex posed to aggressive chemotherapy, under complete remission, with acute onset of rhino-orbital-cerebral disease caused by Aspergillus sclerotiorum and Rhizopus microsporum, treated with aggressive surgery and both local and systemic antifungal therapy.

Catalytic and thermodynamic properties of ß-glucosidases produced by Lichtheimia corymbifera and Byssochlamys spectabilis.

The objective of the present study was to optimize parameters for the cultivation of Lichtheimia corymbifera (mesophilic) and Byssochlamys spectabilis (thermophilic) for the production of ß-glucosidases and to compare the catalytic and thermodynamic properties of the partially purified enzymes. The maximum amount of ß-glucosidase produced by L. corymbifera was 39 U/g dry substrate (or 3.9 U/mL), and that by B. spectabilis was 77 U/g (or 7.7 U/mL). The optimum pH and temperature were 4.5 and 55 °C and 4.0 and 50 °C for the enzyme from L. corymbifera and B. spectabilis, respectively. ß-Glucosidase produced by L. corymbifera was stable at pH 4.0-7.5, whereas the enzyme from B. spectabilis was stable at pH 4.0-6.0. Regarding the thermostability, ß-glucosidase produced by B. spectabilis remained stable for 1 h at 50 °C, and that from L. corymbifera was active for 1 h at 45 °C. Determination of thermodynamic parameters confirmed the greater thermostability of the enzyme produced by the thermophilic fungus B. spectabilis, which showed higher values of ΔH, activation energy for denaturation (Ea), and half-life t(1/2). The enzymes were stable in the presence of ethanol and were competitively inhibited by glucose. These characteristics contribute to their use in the simultaneous saccharification and fermentation of vegetable biomass.