-
Anais Da Academia Brasileira de Ciencias 2021Thermophilic fungi constitute an ecologically well-defined group, commonly found in environments wherever decomposition of organic matter takes place, making them...
Thermophilic fungi constitute an ecologically well-defined group, commonly found in environments wherever decomposition of organic matter takes place, making them self-heating. The importance of thermophilic fungus in ecosystems contrasts with the incompleteness of our understanding of the group's biogeography patterns, phylogenies and coevolution relationships. Actually, the lack of data about thermophilic fungi from the Brazil is a limiting factor that also contributes for this scenario. In order to reduce this gap of knowledge, we aimed to characterize thermophilic filamentous fungi in Araucaria Forest, Atlantic Forest biome. Species identification was achieved by using internal transcribed spacers (ITS) as molecular ribosomal markers. In total, 240 heat-tolerant fungal strains were isolated and identified as Thermothielavioides terrestris, Thielavia sp., Thermoascus crustaceus, Aspergillus fumigatus, Rhizomucor miehei, Rhizomucor pusillus, and Rhizopus microsporus. All thermophilic strains exhibited optimal growth at 45 °C. T. crustaceus, T. miehei e R. pusillus were the dominant species, with the frequencies of occurrence of 35.00%, 28.33% and 23.33%, respectively. Our data reveals the apparent diversity of the Neotropical realm and may serve as reference to future studies that will try to elucidate important aspects of group.
Topics: Araucaria; Brazil; Ecosystem; Eurotiales; Forests; Fungi; Rhizomucor; Rhizopus; Sordariales
PubMed: 34706012
DOI: 10.1590/0001-3765202120210714 -
Journal of Microbiology and... Feb 2017NRRL 5282 and NRRL 1526 can produce lipases with high synthetic activities in wheat bran-based solid-state culture. In this study, the purification and biochemical...
NRRL 5282 and NRRL 1526 can produce lipases with high synthetic activities in wheat bran-based solid-state culture. In this study, the purification and biochemical characterization of the lipolytic activities of these lipases are presented. SDS-PAGE indicated a molecular mass of about 55 and 35 kDa for the purified and enzymes, respectively. -Nitrophenyl palmitate (NPP) hydrolysis was maximal at 40°C and pH 7.0 for the lipase, and at 30°C and pH 5.2 for the enzyme. The enzymes showed almost equal affinity to NPP, but the of the lipase was about 1.13 times higher than that determined for using the same substrate. For both enzymes, a dramatic loss of activity was observed in the presence of 5 mM Hg, Zn, or Mn, 10 mM -bromosuccinimide or sodium dodecyl sulfate, and 5-10% (v/v) of hexanol or butanol. At the same time, they proved to be extraordinarily stable in the presence of -hexane, cyclohexane, -heptane, and isooctane. Moreover, isopentanol up to 10% (v/v) and propionic acid in 1 mM concentrations increased the NPP hydrolyzing activity of lipase. Both enzymes had 1,3-regioselectivity, and efficiently hydrolyzed -nitrophenyl (NP) esters with C8-C16 acids, exhibiting maximum activity towards NP-caprylate () and pNP-dodecanoate (). The purified lipases are promising candidates for various biotechnological applications.
Topics: Bromosuccinimide; Butanols; Caprylates; Electrophoresis, Polyacrylamide Gel; Esterification; Heptanes; Hexanes; Hexanols; Hydrogen-Ion Concentration; Hydrolysis; Kinetics; Laurates; Lipase; Manganese; Mercury; Nitrobenzenes; Palmitates; Propionates; Rhizomucor; Rhizopus; Sodium Dodecyl Sulfate; Zinc
PubMed: 27780957
DOI: 10.4014/jmb.1608.08005 -
Frontiers in Microbiology 2022To explore the potential mechanism of improving enzymatic activities in medium-temperature (MTD) by inoculation functional isolates, we inoculated a single strain of ,...
To explore the potential mechanism of improving enzymatic activities in medium-temperature (MTD) by inoculation functional isolates, we inoculated a single strain of , and the microbiota composed of and in MTD to investigate the association between the response of the functional microbiota and the enzymatic activity. The results showed that the bacterial community of MTD might be more sensitive to bioturbation than the fungal community, and the indigenous microbiota responded to the single strain more than to the microbiota. Moreover, the differential microorganisms mainly included , , and between the conventional and fortified samples. Notably, the composition of functional microbiota related to liquefying activity (LA) and saccharifying activity (SA) were significantly different, changing from and to , , and . That might be closely related to the effect of the bioturbation on LA (31.33%) and SA (43.54%) associated microorganisms was more tellingly. Furthermore, the relative abundance changes of bioturbation-sensitive modules in the co-occurrence network might also lead to the difference in enzymatic activities. Therefore, the LA and SA of MTD were improved by bioturbation significantly. These results provide diverse insights into the exogenous functional isolates to regulate the MTD microbiota and improve enzymatic activities.
PubMed: 36458186
DOI: 10.3389/fmicb.2022.1047041 -
Sheng Wu Gong Cheng Xue Bao = Chinese... Sep 2021L-asparaginase hydrolyzes L-asparagine to produce L-aspartic acid and ammonia. It is widely distributed in microorganisms, plants and serum of some rodents, and has...
L-asparaginase hydrolyzes L-asparagine to produce L-aspartic acid and ammonia. It is widely distributed in microorganisms, plants and serum of some rodents, and has important applications in the pharmaceutical and food industries. However, the poor thermal stability, low catalytic efficiency and low yield hampered the further application of L-asparaginase. In this paper, rational design and 5' untranslated region (5'UTR) design strategies were used to increase the specific enzyme activity and protein expression of L-asparaginase derived from Rhizomucor miehei (RmAsnase). The results showed that among the six mutants constructed through homology modeling combined with sequence alignment, the specific enzyme activity of the mutant A344E was 1.5 times higher than the wild type. Subsequently, a food-safe strain Bacillus subtilis 168/pMA5-A344E was constructed, and the UTR strategy was used for the construction of recombinant strain B. subtilis 168/pMA5 UTR-A344E. The enzyme activity of B. subtilis 168/pMA5 UTR-A344E was 7.2 times higher than that of B. subtilis 168/pMA5-A344E. The recombinant strain B. subtilis 168/pMA5 UTR-A344E was scaled up in 5 L fermenter, and the final yield of L-asparaginase was 489.1 U/mL, showing great potential for industrial application.
Topics: Asparaginase; Bacillus subtilis; Industrial Microbiology; Protein Engineering; Rhizomucor; Sequence Alignment
PubMed: 34622632
DOI: 10.13345/j.cjb.200759 -
Journal of Clinical Microbiology Jan 2023There has been significant increase in the use of molecular tools for the diagnosis of invasive aspergillosis (IA) and mucormycosis. However, their range of detection...
There has been significant increase in the use of molecular tools for the diagnosis of invasive aspergillosis (IA) and mucormycosis. However, their range of detection may be too limited as species diversity and coinfections are increasing. Here, we aimed to evaluate a molecular workflow based on a new multiplex PCR assay detecting the whole Aspergillus genus and the Mucorales order followed by a species-specific PCR or a DNA-sequencing approach for IA and/or mucormycosis diagnosis and species identification on serum. Performances of the MycoGENIE Aspergillus spp./Mucorales spp. duplex PCR kit were analyzed on a broad range of fungal strains and on sera from high-risk patients prospectively over a 12-month period. The kit allowed the detection of nine Aspergillus species and 10 Mucorales (eight genera) strains assessed. No cross-reactions between the two targets were observed. Sera from 744 patients were prospectively analyzed, including 35 IA, 16 mucormycosis, and four coinfections. Sensitivity varies from 85.7% (18/21) in probable/proven IA to 28.6% (4/14) in COVID-19-associated pulmonary aspergillosis. PCR-positive samples corresponded to 21 A. fumigatus, one A. flavus, and one A. nidulans infections. All the disseminated mucormycosis were positive in serum (14/14), including the four Aspergillus coinfections, but sensitivity fell to 33.3% (2/6) in localized forms. DNA sequencing allowed Mucorales identification in serum in 15 patients. Remarkably, the most frequent species identified was (eight cases), whereas it is barely found in fungal culture. This molecular workflow is a promising approach to improve IA and mucormycosis diagnosis and epidemiology.
Topics: Humans; Mucormycosis; Multiplex Polymerase Chain Reaction; Coinfection; Workflow; COVID-19; Aspergillosis; Mucorales; Invasive Fungal Infections; Aspergillus; Sequence Analysis, DNA; DNA; DNA, Fungal; COVID-19 Testing
PubMed: 36533925
DOI: 10.1128/jcm.01409-22 -
Antimicrobial Agents and Chemotherapy Aug 2019Recently, the species concept of opportunistic and its relatives has been revised, resulting in the recognition of its classical formae as independent species and the...
Recently, the species concept of opportunistic and its relatives has been revised, resulting in the recognition of its classical formae as independent species and the description of new species. In this study, we used isolates of all clinically relevant species and performed susceptibility testing using the EUCAST reference method to identify potential species-specific susceptibility patterns. susceptibility profiles of 101 mucoralean strains belonging to the genus (72), the closely related species (3), (12), (10), and (4) to six antifungals (amphotericin B, natamycin, terbinafine, isavuconazole, itraconazole, and posaconazole) were determined. The most active drug for all Mucorales was amphotericin B. Antifungal susceptibility profiles of pathogenic species were specific for isavuconazole, itraconazole, and posaconazole. The species formerly united in showed clear differences in their antifungal susceptibilities. , , ( f. ), and exhibited high MICs to all azoles tested. presented high MICs for isavuconazole and posaconazole, and and showed high MICs for isavuconazole. MIC values of spp. for posaconazole, isavuconazole, and itraconazole were high compared to those for and the Lichtheimiaceae ( and ). Molecular identification combined with susceptibility testing is recommended for species, especially if azoles are applied in treatment.
Topics: Amphotericin B; Antifungal Agents; Humans; Itraconazole; Microbial Sensitivity Tests; Mucor; Mucormycosis; Natamycin; Nitriles; Pyridines; Rhizopus; Species Specificity; Terbinafine; Triazoles
PubMed: 31182532
DOI: 10.1128/AAC.00653-19 -
Scientific Reports Apr 2022The use of enzymes immobilized on nanomagnetic supports has produced surprising results in catalysis, mainly due to the increase in surface area and the potential for...
The use of enzymes immobilized on nanomagnetic supports has produced surprising results in catalysis, mainly due to the increase in surface area and the potential for recovery and reuse. However, the meticulous control of the process and difficulties in reproducibility have made industrial-scale applications unfeasible. Furthermore, the role of conjugation strategies in the catalytic activity and recycling of catalysts is unclear. Therefore, the objective of this study was to compare the conjugation of enzymes on nanomagnetic supports through physical adsorption (naked) or covalent bonding with mercaptopropyltrimethoxysilane (MPTS) and aminopropyltriethoxysilane (APTS) ligands. The free lipase obtained from Rhizomucor miehei was used as a model enzyme. Total protein and enzyme activity were determined using spectrophotometry (UV-Vis) and the p-nitrophenyl palmitate (p-NPP) hydrolysis method. The results indicated that a more significant enzyme surface loading does not always mean better immobilization success. The physical adsorption binding strategy had higher surface loading and low catalytic activity. On the other hand, covalent coupling with free NH2 had an excellent catalytic activity with very low surface loading. Finally, we show that recyclability can be improved with conjugation mediated by disulfide bonds. The findings presented here are essential for developing nanoconjugates with high enzymatic activity, which can guarantee the success of several industrial applications.
Topics: Adsorption; Enzymes, Immobilized; Hydrolysis; Lipase; Reproducibility of Results
PubMed: 35474328
DOI: 10.1038/s41598-022-10721-y -
Infection and Drug Resistance 2022Metagenomic next-generation sequencing (mNGS) is a fast, sensitive and accurate diagnostic method for pathogens detection. However, reports on the application of mNGS in...
BACKGROUND
Metagenomic next-generation sequencing (mNGS) is a fast, sensitive and accurate diagnostic method for pathogens detection. However, reports on the application of mNGS in mucormycosis remain scarce.
METHODS
From January 2019 to December 2021, we recruited 13 patients with hematological malignancies who were suspected of mucormycosis and completed mNGS in D20. Then we retrospectively analyze the clinical data, diagnosis, therapeutic process, and outcomes. In order to evaluate the diagnostic value of mNGS in hematological malignancies patients with suspected mucormycosis.
RESULTS
All patients had high risk factors of Invasive Fungal Disease, including hematopoietic stem cell transplantation, immunosuppression, glucocorticoids, etc. The clinical presentations were pulmonary (n=9), rhino-orbito-cerebral (n=4). But the manifestations were nonspecific. All enrolled patients completed mNGS. And most (8/13, 61.54%) of the samples were from blood. Fungi can be detected in all specimens, including Rhizopus (n=7), Rhizomucor (n=4) and Mucor (n=2). In addition, 7/13 (53.85%) specimens were detected bacteria at the same time and virus were detected in 5/13 (38.46%). Histopathological examination was completed in 5 patients, 3 of which were completely consistent with the results of mNGS. After treatment, 6 patients were cured, while the other 7 patients died.
CONCLUSION
mNGS may be a complementary method for early diagnosis, especially for patients who are not suitable for histopathology examination or unable to obtain culture specimen. mNGS can also help detect bacteria and viruses simultaneously, allowing for appropriate and timely antibiotic administration and thus improving patient outcomes.
PubMed: 36570711
DOI: 10.2147/IDR.S393201 -
Annals of the Royal College of Surgeons... Apr 2021Fungal infections are generally observed in immunosuppressed patients only, with a diagnostic challenge due to non-specific symptoms. For this reason, appropriate...
Fungal infections are generally observed in immunosuppressed patients only, with a diagnostic challenge due to non-specific symptoms. For this reason, appropriate management may be delayed. This case report concerns a 36-year-old man with history of pancreas and kidney transplantation. He had chemotherapy for post-transplant B-cell lymphoma and presented with left upper abdominal pain and fever. Multiple investigations led to a final diagnosis of disseminated abdominal mucormycosis with multiple abscesses in the liver, spleen and kidney transplant. Treatment was antifungal therapy and laparotomy with splenectomy, wedge resection of two fungal abscesses in segments II and IVb, and segmental left colic resection.
Topics: Abdominal Abscess; Adult; Hepatectomy; Humans; Kidney Transplantation; Liver Diseases; Male; Mucormycosis; Pancreas Transplantation; Postoperative Complications; Rhizomucor; Splenectomy; Splenic Diseases
PubMed: 33682478
DOI: 10.1308/rcsann.2020.7125 -
Current Medical Mycology Mar 2016Soil is the main habitat of saprophytic and pathogenic fungi. Mucoromycotina constitutes a large group of soil fungi, with certain opportunistic members causing systemic...
BACKGROUND AND PURPOSE
Soil is the main habitat of saprophytic and pathogenic fungi. Mucoromycotina constitutes a large group of soil fungi, with certain opportunistic members causing systemic infections in immunocompromised hosts. The majority of human and animal infections are caused by the members of the genera , , , (, and . Accordingly, in the present study, we aimed to isolate and identify the main genera of the order Mucorales, using molecular assays and morphological features
MATERIALS AND METHODS
In total, 340 soil samples were collected from seven public parks throughout the city and sidewalk gardens in 14 municipal districts in Isfahan, Iran. All the samples were cultured on the appropriate media, incubated at 27°C for 2- 4 days, and examined daily for visible fungal growth. Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method was applied and macroscopic, microscopic, and physiological characteristics were assessed to identify fungal colonies.
RESULTS
400 pure colonies, belonging to the orders Mucorales and Mortierellales, including the genera , and were identified The genus (35.5%) was the most frequent isolate, followed by (32.25%) and (27.5%).
CONCLUSION
The results emphasize the importance of opportunistic fungi in public areas and indicate the risk of exposure for immunocompromised individuals.
PubMed: 28681007
DOI: 10.18869/acadpub.cmm.2.1.13