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Biology Dec 2021l-asparaginase (EC 3.5.1.1) hydrolyzes l-asparagine to produce l-aspartate and ammonia and is widely found in microorganisms, plants, and some rodent sera....
l-asparaginase (EC 3.5.1.1) hydrolyzes l-asparagine to produce l-aspartate and ammonia and is widely found in microorganisms, plants, and some rodent sera. l-asparaginase used for industrial production should have good thermostability. We heterologously expressed l-asparaginase from , selected nine loci for site-directed mutagenesis by rational design, and obtained two mutants with significantly improved thermostability. The optimal temperature of mutants S302I and S302M was 50 °C. After incubating the mutant and wild-type enzymes at 45 °C for 35 h, the residual activity of the wild-type enzyme (WT) was only about 10%. In contrast, the residual activity of S302I and S302M was more than 50%. After combination mutagenesis, 168-pMA5-A344E/S302I was constructed using the food-safe host strain 168. Additionally, a 5' untranslated region (UTR) modification strategy was adopted to enhance the expression level of -derived l-asparaginase in . In a 5-L fermenter scale-up experiment, the enzyme activity of recombinant 168-pMA5-UTR-A344E/S302I reached 521.9 U·mL by fed-batch fermentation.
PubMed: 34943261
DOI: 10.3390/biology10121346 -
Frontiers in Cellular and Infection... 2024Invasive mold diseases of the central nervous (CNS IMD) system are exceedingly rare disorders, characterized by nonspecific clinical symptoms. This results in...
BACKGROUND
Invasive mold diseases of the central nervous (CNS IMD) system are exceedingly rare disorders, characterized by nonspecific clinical symptoms. This results in significant diagnostic challenges, often leading to delayed diagnosis and the risk of misdiagnosis for patients. Metagenomic Next-Generation Sequencing (mNGS) holds significant importance for the diagnosis of infectious diseases, especially in the rapid and accurate identification of rare and difficult-to-culture pathogens. Therefore, this study aims to explore the clinical characteristics of invasive mold disease of CNS IMD in children and assess the effectiveness of mNGS technology in diagnosing CNS IMD.
METHODS
Three pediatric patients diagnosed with Invasive mold disease brain abscess and treated in the Pediatric Intensive Care Unit (PICU) of the First Affiliated Hospital of Zhengzhou University from January 2020 to December 2023 were selected for this study.
RESULTS
Case 1, a 6-year-old girl, was admitted to the hospital with "acute liver failure." During her hospital stay, she developed fever, irritability, and seizures. CSF mNGS testing resulted in a negative outcome. Multiple brain abscesses were drained, and was detected in pus culture and mNGS. The condition gradually improved after treatment with voriconazole combined with caspofungin. Case 2, a 3-year-old girl, was admitted with "acute B-lymphoblastic leukemia." During induction chemotherapy, she developed fever and seizures. was detected in the intracranial abscess fluid by mNGS, and the condition gradually improved after treatment with voriconazole combined with caspofungin, followed by "right-sided brain abscess drainage surgery." Case 3, a 7-year-old girl, showed lethargy, fever, and right-sided limb weakness during the pending chemotherapy period for acute B-lymphoblastic leukemia. and was detected in the cerebrospinal fluid by mNGS. The condition gradually improved after treatment with amphotericin B combined with posaconazole. After a six-month follow-up post-discharge, the three patients improved without residual neurological sequelae, and the primary diseases were in complete remission.
CONCLUSION
The clinical manifestations of CNS IMD lack specificity. Early mNGS can assist in identifying the pathogen, providing a basis for definitive diagnosis. Combined surgical treatment when necessary can help improve prognosis.
Topics: Humans; Female; High-Throughput Nucleotide Sequencing; Child; Metagenomics; Brain Abscess; Antifungal Agents; Invasive Fungal Infections; Male; Central Nervous System Fungal Infections; Child, Preschool; Aspergillus fumigatus; Caspofungin
PubMed: 38912204
DOI: 10.3389/fcimb.2024.1393242 -
Le Infezioni in Medicina Sep 2015We describe a severe pulmonary invasive zygomycosis sustained by Rhizomucor pusillus and concomitant bacterial endocarditis in a woman diagnosed with acute promyelocytic...
We describe a severe pulmonary invasive zygomycosis sustained by Rhizomucor pusillus and concomitant bacterial endocarditis in a woman diagnosed with acute promyelocytic leukaemia.
Topics: Antifungal Agents; Drug Therapy, Combination; Endocarditis, Bacterial; Female; Humans; Immunocompromised Host; Leukemia, Promyelocytic, Acute; Lung Diseases, Fungal; Middle Aged; Pneumonectomy; Rhizomucor; Severity of Illness Index; Treatment Outcome; Zygomycosis
PubMed: 26397298
DOI: No ID Found -
Journal of Oleo Science Nov 2016A two-step enzymatic transesterification process in a solvent-free system has been developed as a novel approach to the production of biodiesel using acid oil from rice...
A two-step enzymatic transesterification process in a solvent-free system has been developed as a novel approach to the production of biodiesel using acid oil from rice bran oil soapstock. The acid oil consisted of 53.7 wt% fatty acids, 2.4 wt% monoacylglycerols, 9.1 wt% diacylglycerols, 28.8 wt% triacylglycerols, and 6.0 wt% others. Three immobilized lipases were evaluated as potential biocatalysts, including Novozym 435 from Candida antarctica, Lipozyme RM IM from Rhizomucor miehei, and Lipozyme TL IM from Thermomyces lanuginosus. The effects of molar ratio of acid oil to ethanol, temperature, and enzyme loading were investigated to determine the optimum conditions for the transesterification with the three immobilized lipases. The optimum conditions of the three immobilized lipases were a molar ratio of 1:5 (acid oil to ethanol), the temperature range of 30-40°C, and the enzyme loading range of 5-10%. The two-step transesterification was then conducted under the optimum conditions of each lipase. The stepwise use of Novozym 435 and Lipozyme TL IM or Lipozyme RM IM and Lipozyme TL IM resulted in similar or higher levels of yield to the individual lipases. The maximum yields obtained in both stepwise uses were ca. 92%.
Topics: Biocatalysis; Biofuels; Esterification; Fatty Acids, Nonesterified; Glycerides; Lipase; Oryza; Plant Oils; Rice Bran Oil; Temperature
PubMed: 27733740
DOI: 10.5650/jos.ess16092 -
The Journal of General and Applied... 2016A large number of propeptide regions from various proteins have been identified which function as intramolecular chaperones and assist the folding of the respective...
A large number of propeptide regions from various proteins have been identified which function as intramolecular chaperones and assist the folding of the respective functional domains. The same polypeptide can fold into an altered conformation because of a mutated intramolecular chaperone and can maintain the "memory" of the folding process (new physicochemical properties). Two new kinds of Rhizomucor miehei lipase (RML) were constructed by replacing its propeptide region with that from either Rhizopus chinensis lipase (RCL) or Rhizopus oryzae lipase (ROL). The enzymatic properties were also analyzed and compared between wild-type RML and the mutants. The results indicated that the same polypeptide can fold into different conformations because of changes in the propeptide region.
Topics: Amino Acid Sequence; Gene Expression; Lipase; Pichia; Protein Conformation; Protein Folding; Recombinant Fusion Proteins; Rhizomucor; Rhizopus
PubMed: 26923128
DOI: 10.2323/jgam.62.25 -
Bioengineering (Basel, Switzerland) Oct 2022Morphology plays an important role in the fermentation bioprocess of filamentous fungi. In this study, we investigated the controlling strategies of morphology that...
Morphology plays an important role in the fermentation bioprocess of filamentous fungi. In this study, we investigated the controlling strategies of morphology that improved the efficiency of lipase (RML) production using a high-yield First, the inoculated spore concentrations were optimized in seed culture, and the RML activity increased by 43.4% with the well-controlled mycelium pellets in both ideal sizes and concentrations. Then, the initial nitrogen source and agitation strategies were optimized to regulate the morphology of in a 5 L bioreactor, and the established stable fermentation system increased the RML activity to 232.0 U/mL, combined with an increase in total RML activity from 98,080 U to 487,179 U. Furthermore, the optimized fermentation strategy was verified by a high-yield and achieved an additional improvement of RML activity, up to 320.0 U/mL. Moreover, this optimized fermentation bioprocess was successfully scaled up to a 50 L bioreactor, and the RML activity reached 550.0 U/mL. This work has established a stable precision fermentation bioprocess for RML production by in bioreactors, and the controlling strategy developed in this study could potentially be extended to an industrial scale for RML production with high efficiency.
PubMed: 36354521
DOI: 10.3390/bioengineering9110610 -
Animal : An International Journal of... Dec 2016Molecular techniques are important tools for microbiological studies in different habitats, and the internal transcribed spacer (ITS) has been proved to be useful for...
Molecular techniques are important tools for microbiological studies in different habitats, and the internal transcribed spacer (ITS) has been proved to be useful for analyzing fungal diversity. The aim of this study was to use the ITS region to generate ruminal yeast profile and to identify ruminal yeast. DNA from ruminal digesta was extracted to amplify the ribosomal ITS region. The profile from the PCR products was visualized and the excised bands from the profile were identified as the genera Millerozyma, Pichia, Rhizomucor and Hyphopichia. Overall, the ITS resulted to be a simple, fast and sensitive approach that allowed profiling and identification of ruminal yeast that have not been previously described (Millerozyma and Hyphopichia) in the rumen microbial community.
Topics: Animals; Cattle; DNA, Ribosomal Spacer; Female; Genetic Markers; Hydrogen-Ion Concentration; Rumen; Saccharomyces cerevisiae; Sensitivity and Specificity; Yeast, Dried
PubMed: 27133003
DOI: 10.1017/S1751731116000768 -
Persoonia Jun 2013The order Mucorales comprises predominantly fast-growing saprotrophic fungi, some of which are used for the fermentation of foodstuffs but it also includes species known...
The order Mucorales comprises predominantly fast-growing saprotrophic fungi, some of which are used for the fermentation of foodstuffs but it also includes species known to cause infections in patients with severe immune or metabolic impairments. To inventory biodiversity in Mucorales ITS barcodes of 668 strains in 203 taxa were generated covering more than two thirds of the recognised species. Using the ITS sequences, Molecular Operational Taxonomic Units were defined by a similarity threshold of 99 %. An LSU sequence was generated for each unit as well. Analysis of the LSU sequences revealed that conventional phenotypic classifications of the Mucoraceae are highly artificial. The LSU- and ITS-based trees suggest that characters, such as rhizoids and sporangiola, traditionally used in mucoralean taxonomy are plesiomorphic traits. The ITS region turned out to be an appropriate barcoding marker in Mucorales. It could be sequenced directly in 82 % of the strains and its variability was sufficient to resolve most of the morphospecies. Molecular identification turned out to be problematic only for the species complexes of Mucor circinelloides, M. flavus, M. piriformis and Zygorhynchus moelleri. As many as 12 possibly undescribed species were detected. Intraspecific variability differed widely among mucorealean species ranging from 0 % in Backusella circina to 13.3 % in Cunninghamella echinulata. A high proportion of clinical strains was included for molecular identification. Clinical isolates of Cunninghamella elegans were identified molecularly for the first time. As a result of the phylogenetic analyses several taxonomic and nomenclatural changes became necessary. The genus Backusella was emended to include all species with transitorily recurved sporangiophores. Since this matched molecular data all Mucor species possessing this character were transferred to Backusella. The genus Zygorhynchus was shown to be polyphyletic based on ITS and LSU data. Consequently, Zygorhynchus was abandoned and all species were reclassified in Mucor. Our phylogenetic analyses showed, furthermore, that all non-thermophilic Rhizomucor species belong to Mucor. Accordingly, Rhizomucor endophyticus was transferred to Mucor and Rhizomucor chlamydosporus was synonymised with Mucor indicus. Lecto-, epi- or neotypes were designated for several taxa.
PubMed: 24027345
DOI: 10.3767/003158513X665070 -
Journal of Fungi (Basel, Switzerland) Apr 2018Mucormycosis is an emerging opportunistic fungal infection. Increasing immunocompromization, widespread use of antibacterial and antifungal agents (such as voriconazole...
Mucormycosis is an emerging opportunistic fungal infection. Increasing immunocompromization, widespread use of antibacterial and antifungal agents (such as voriconazole prophylaxis), carcinomas, transplantation and lifestyle diseases such as diabetes are the main contributors to this situation. The predominant clinical manifestations of mucormycosis vary from host to host, with rhino-orbital-cerebral, pulmonary, cutaneous, and gastrointestinal infections being the most common. In India, the prevalence of mucormycosis is approximately 0.14 cases/1000 population, which is about 70 times the worldwide-estimated rate for mucormycosis. The present study was undertaken over a period of five years (January 2009-December 2014) to determine the prevalence of mucormycosis. The samples suspected of mucormycosis were examined by direct KOH wet mount and cultured on Sabouraud's dextrose agar without actidione and on blood agar as per standard mycological techniques. Histopathological correlation was done for most of the cases. Antifungal susceptibility testing was performed by the EUCAST reference method. We identified a total of 82 cases of mucormycosis out of a total of 6365 samples received for mycological culture and examination during the said time period. Out of these, 56 were male patients and 27 were females. Most common presentation was rhino-orbito-cerebral (37), followed by cutaneous (25), pulmonary (14), oral cavity involvement (4) and gastrointestinal (2). The most common risk factors were diabetes and intramuscular injections. The fungi isolated were (17), (12), (9), (8), (5), (4), (2), (1), (1) and (1). The mainstay of the treatment was amphotericin B, along with extensive surgical debridement whenever feasible. Most of the patients (50) recovered, but 25 died. The rest of the patients left against medical advice. "Nip in the Bud" should be the mantra for clinicians/surgeons for a favorable prognosis. Early diagnosis, prompt institution of appropriate antifungal therapy, surgical debridement whenever necessary, knowledge of risk factors and their timely reversal is the key for management.
PubMed: 29642408
DOI: 10.3390/jof4020046 -
Cureus Jul 2023Background Mucormycosis is a consequence of the angioinvasive disease caused by filamentous fungi that belong to the order Mucorales, particularly , , and . is the most...
Sinonasal and Orbital Imaging Findings in COVID-Associated Rhino-Orbito-Cerebral Mucormycosis During the Second Wave of COVID-19: A Retrospective Cohort Study in a Tertiary Hospital in Central India.
Background Mucormycosis is a consequence of the angioinvasive disease caused by filamentous fungi that belong to the order Mucorales, particularly , , and . is the most prevalent form. The invading hyphae lead to damage of blood vessels leading to thrombosis and consequent tissue necrosis. The incidence of this disease entity witnessed a significant rise during the second wave of the coronavirus disease 2019 (COVID-19) pandemic. Timely diagnosis and prompt treatment are crucial to diminish both the mortality and morbidity associated with this disease. Imaging plays a pivotal role in diagnosing the ailment, evaluating its extent, identifying complications such as thrombosis, and facilitating surgical planning. It demonstrates exceptional sensitivity in detecting the disease at its early stages, often before symptoms manifest. Due to the angioinvasive nature of , early detection assumes utmost importance as it necessitates intensive antifungal therapy and the removal of devitalized tissue through debridement. Methodology We conducted a retrospective cohort study to analyze computed tomography (CT) imaging findings in patients with COVID-associated rhino-orbito-cerebral mucormycosis (ROCM) confirmed by histopathological examination. We compared these findings with CT findings of the nose and paranasal sinuses in patients without mucor following COVID-19 sinusitis (non-ROCM). Results All 16 cases in the non-ROCM group were in stage 1 disease. In contrast, in the ROCM group, three patients had stage 1 disease, five patients had stage 2 disease, and 10 patients had stage 3 disease (p = 0.0001). The pterygopalatine fossa was significantly affected in 10 of 18 ROCM patients and in none of the non-ROCM patients. Conclusions Imaging plays a crucial role in the early detection of mucormycosis. It assists treating physicians in initiating prompt and aggressive treatment, thereby improving the prognosis of this frequently fatal disease.
PubMed: 37649953
DOI: 10.7759/cureus.42674