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Parasites & Vectors Jun 2018In recent years, the genus Asaia (Rhodospirillales: Acetobacteraceae) has been isolated from different Anopheles species and presented as a promising tool to combat...
Isolation and identification of Asaia sp. in Anopheles spp. mosquitoes collected from Iranian malaria settings: steps toward applying paratransgenic tools against malaria.
BACKGROUND
In recent years, the genus Asaia (Rhodospirillales: Acetobacteraceae) has been isolated from different Anopheles species and presented as a promising tool to combat malaria. This bacterium has unique features such as presence in different organs of mosquitoes (midgut, salivary glands and reproductive organs) of female and male mosquitoes and vertical and horizontal transmission. These specifications lead to the possibility of introducing Asaia as a robust candidate for malaria vector control via paratransgenesis technology. Several studies have been performed on the microbiota of Anopheles mosquitoes (Diptera: Culicidae) in Iran and the Middle East to find a suitable candidate for controlling the malaria based on paratransgenesis approaches. The present study is the first report of isolation, biochemical and molecular characterization of the genus Asaia within five different Anopheles species which originated from different zoogeographical zones in the south, east, and north of Iran.
METHODS
Mosquitoes originated from field-collected and laboratory-reared colonies of five Anopheles spp. Adult mosquitoes were anesthetized; their midguts were isolated by dissection, followed by grinding the midgut contents which were then cultured in enrichment broth media and later in CaCO agar plates separately. Morphological, biochemical and physiological characterization were carried out after the appearance of colonies. For molecular confirmation, selected colonies were cultured, their DNAs were extracted and PCR was performed on the 16S ribosomal RNA gene using specific newly designed primers.
RESULTS
Morphological, biochemical, physiological and molecular results indicated that all isolates are members of the genus Asaia.
CONCLUSIONS
Contrary to previous opinions, our findings show that Asaia bacteria are present in both insectary-reared colonies and field-collected mosquitoes and can be isolated by simple and specific methods. Furthermore, with respect to the fact that we isolated Asaia within the different Anopheles specimens from distinct climatic and zoogeographical regions, it is promising and may be concluded that species of this genus can tolerate the complicated environmental conditions of the vector-borne diseases endemic regions. Therefore, it can be considered as a promising target in paratransgenesis and vector control programs. However, we suggest that introducing the new technologies such as next generation sequencing and robust in silico approaches may pave the way to find a unique biomarker for rapid and reliable differentiation of the Asaia species.
Topics: Acetobacteraceae; Animals; Anopheles; Biological Control Agents; Digestive System; Female; Humans; Iran; Larva; Malaria; Male; Microbiota; Middle East; Mosquito Control; Mosquito Vectors; Phylogeny; Polymerase Chain Reaction; RNA, Ribosomal, 16S; Symbiosis
PubMed: 29950179
DOI: 10.1186/s13071-018-2955-9 -
Microbial Cell Factories Dec 2022The global market for sweeteners is increasing, and the food industry is constantly looking for new low-caloric sweeteners. The natural sweetener 5-keto-D-fructose is...
BACKGROUND
The global market for sweeteners is increasing, and the food industry is constantly looking for new low-caloric sweeteners. The natural sweetener 5-keto-D-fructose is one such candidate. 5-Keto-D-fructose has a similar sweet taste quality as fructose. Developing a highly efficient 5-keto-D-fructose production process is key to being competitive with established sweeteners. Hence, the 5-keto-D-fructose production process was optimised regarding titre, yield, and productivity.
RESULTS
For production of 5-keto-D-fructose with G. oxydans 621H ΔhsdR pBBR1-p264-fdhSCL-ST an extended-batch fermentation was conducted. During fructose feeding, a decreasing respiratory activity occurred, despite sufficient carbon supply. Oxygen and second substrate limitation could be excluded as reasons for the decreasing respiration. It was demonstrated that a short period of oxygen limitation has no significant influence on 5-keto-D-fructose production, showing the robustness of this process. Increasing the medium concentration increased initial biomass formation. Applying a fructose feeding solution with a concentration of approx. 1200 g/L, a titre of 545 g/L 5-keto-D-fructose was reached. The yield was with 0.98 g/g close to the theoretical maximum. A 1200 g/L fructose solution has a viscosity of 450 mPa∙s at a temperature of 55 °C. Hence, the solution itself and the whole peripheral feeding system need to be heated, to apply such a highly concentrated feeding solution. Thermal treatment of highly concentrated fructose solutions led to the formation of 5-hydroxymethylfurfural, which inhibited the 5-keto-D-fructose production. Therefore, fructose solutions were only heated to about 100 °C for approx. 10 min. An alternative feeding strategy was investigated using solid fructose cubes, reaching the highest productivities above 10 g/L/h during feeding. Moreover, the scale-up of the 5-keto-D-fructose production to a 150 L pressurised fermenter was successfully demonstrated using liquid fructose solutions (745 g/L).
CONCLUSION
We optimised the 5-keto-D-fructose production process and successfully increased titre, yield and productivity. By using solid fructose, we presented a second feeding strategy, which can be of great interest for further scale-up experiments. A first scale-up of this process was performed, showing the possibility for an industrial production of 5-keto-D-fructose.
Topics: Gluconobacter oxydans; Fructose; Fermentation; Sweetening Agents; Oxygen
PubMed: 36496372
DOI: 10.1186/s12934-022-01980-5 -
Emerging Infectious Diseases Oct 2018A clinical case study involving a man (35-49 years of age) with wounds to his lower right extremity. An isolate was sent to the Delaware Public Health Laboratory for...
A clinical case study involving a man (35-49 years of age) with wounds to his lower right extremity. An isolate was sent to the Delaware Public Health Laboratory for confirmatory testing by genetic analysis of the 16S gene. Testing identified the isolate as a novel genus and species, Haematospirillum jordaniae.
Topics: Adult; Anti-Bacterial Agents; Bacteriological Techniques; Gram-Negative Bacterial Infections; Humans; Male; Middle Aged; RNA, Ribosomal, 16S; Rhodospirillaceae; Treatment Outcome; Wound Infection
PubMed: 30226180
DOI: 10.3201/eid2410.180548 -
Journal of Bacteriology Nov 2023, an industrial vinegar-producing strain, is suffered by fermentation stress such as fermentation heat and/or high concentrations of acetic acid. By an experimental...
, an industrial vinegar-producing strain, is suffered by fermentation stress such as fermentation heat and/or high concentrations of acetic acid. By an experimental evolution approach, we have obtained a stress-tolerant strain, exhibiting significantly increased growth and acetic acid fermentation ability at higher temperatures. In this study, we report that only the three gene mutations of ones accumulated during the adaptation process, , , and , were sufficient to reproduce the increased thermotolerance of . These mutations resulted in cell envelope modification, including increased phospholipid and lipopolysaccharide synthesis, increased respiratory activity, and cell size reduction. The phenotypic changes may cooperatively work to make the adapted cell thermotolerant by enhancing cell surface integrity, nutrient or oxygen availability, and energy generation.
Topics: Acetic Acid; Thermotolerance; Acetobacter; Fermentation; Amino Acids
PubMed: 37930061
DOI: 10.1128/jb.00101-23 -
The Journal of Biophysical and... Oct 1959Cells from serial cultures of R. rubrum, grown anaerobically in the light, were harvested at intervals from (1/2) to 15 days and sectioned for electron microscopy by...
Cells from serial cultures of R. rubrum, grown anaerobically in the light, were harvested at intervals from (1/2) to 15 days and sectioned for electron microscopy by conventional methods. Cells of this species possess a multilayered outer envelope, and the external cell surface is differentiated into ridges extending parallel or obliquely to the long axis of the cell. Cells from very young cultures resemble non-photosynthetic bacteria and contain only a granular cytoplasm, scattered high-density particles, and low-density areas corresponding to the chromatin areas observed by light microscopy. They contain neither the chromatophores nor the lamellar systems assumed by previous investigators to be characteristic of this species when grown anaerobically in the light. Chromatophores appear in cells from cultures older than about 12 hours, while systems of paired lamellae appear along with the chromatophores in cells from cultures older than about 8 days. Divergent opinions concerning the occurrence of chromatophores or lamellae in this species can be resolved on the basis of the age of cultures used in previous studies. Other changes occurring in cells from cultures of increasing age include the appearance of granular and reticulate cytoplasmic bodies and vacuoles, extension of the chromatin areas, and the appearance of a single membrane enclosing several chromatophores.
Topics: Cell Membrane; Chromatophores; Light; Microscopy, Electron; Rhodospirillum; Rhodospirillum rubrum
PubMed: 14401694
DOI: 10.1083/jcb.6.2.277 -
The Journal of Biological Chemistry 2021Nitrogenase is the only enzyme capable of catalyzing nitrogen fixation, the reduction of dinitrogen gas (N) to ammonia (NH). Nitrogenase is tightly inhibited by the...
Nitrogenase is the only enzyme capable of catalyzing nitrogen fixation, the reduction of dinitrogen gas (N) to ammonia (NH). Nitrogenase is tightly inhibited by the environmental gas carbon monoxide (CO). Nitrogen-fixing bacteria rely on the protein CowN to grow in the presence of CO. However, the mechanism by which CowN operates is unknown. Here, we present the biochemical characterization of CowN and examine how CowN protects nitrogenase from CO. We determine that CowN interacts directly with nitrogenase and that CowN protection observes hyperbolic kinetics with respect to CowN concentration. At a CO concentration of 0.001 atm, CowN restores nearly full nitrogenase activity. Our results further indicate that CowN's protection mechanism involves decreasing the binding affinity of CO to nitrogenase's active site approximately tenfold without interrupting substrate turnover. Taken together, our work suggests CowN is an important auxiliary protein in nitrogen fixation that engenders CO tolerance to nitrogenase.
Topics: Bacterial Proteins; Carbon Monoxide; Catalysis; Gluconacetobacter; Kinetics; Models, Molecular; Nitrogen; Nitrogen Fixation; Nitrogenase; Oxidation-Reduction; Protein Interaction Domains and Motifs
PubMed: 33667548
DOI: 10.1016/j.jbc.2021.100501 -
World Journal of Microbiology &... Apr 2022The objective of the present study was to evaluate possible interactions between two potential plant growth-promoting bacteria (PGPB): Azospirillum oryzae strain NBT506...
The objective of the present study was to evaluate possible interactions between two potential plant growth-promoting bacteria (PGPB): Azospirillum oryzae strain NBT506 and Bacillus velezensis strain UTB96. To do this, the growth kinetic, biofilm formation, motility, surfactin production, indole-3-acetic acid (IAA) production, phosphate solubilization and enzyme activities of the strains were measured in monoculture and co-culture. The maximum biomass production for the strains in monoculture and co-culture was about 10 CFU/ml, confirming that these two strains have the potential to grow in co-culture without reduction of biomass efficiency. The co-culture system showed more stable biofilm formation until the end of day 3. Azospirillum showed the maximum IAA production (41.5 mg/l) in a monoculture compared to other treatments. Surfactin promoted both swimming and swarming motility in all treatments. The Bacillus strain in the monoculture and co-culture showed high phosphate solubilizing capability, which increased continuously in the co-culture system after 6 days. The strains showed protease, amylase and cellulase activities in both monoculture and co-culture forms. Chitinase and lipase activities were observed in both the monoculture of the Bacillus strain and the co-culture. Overall, our findings highlight the promotion of biological and beneficial effects of these bacteria when growing together in co-culture.
Topics: Azospirillum; Bacillus; Bacteria; Coculture Techniques; Phosphates
PubMed: 35486223
DOI: 10.1007/s11274-022-03283-8 -
Microbial Biotechnology Nov 2017Successful merging of chemical and biotechnological operations is essential to achieve cost-efficient industrialization of bio-based processes. The demonstration of the...
Successful merging of chemical and biotechnological operations is essential to achieve cost-efficient industrialization of bio-based processes. The demonstration of the use of syngas, derived from microwave assisted pyrolysis of municipal solid waste, for the improved growth and poly-3-hydroxybutyrate production in Rhodospirillium rubrum, stands out as an example of the synergistic contribution of chemical engineering and applied microbiology to sustainable biomaterial manufacturing, paving the way to similar applications for other syngas derived bioproducts.
Topics: Biodegradation, Environmental; Biotechnology; Hydroxybutyrates; Rhodospirillum rubrum; Solid Waste
PubMed: 27573515
DOI: 10.1111/1751-7915.12409 -
Applied Microbiology and Biotechnology Sep 2021For the acetic acid bacterium (AAB) Gluconobacter oxydans only recently the first tight system for regulatable target gene expression became available based on the...
For the acetic acid bacterium (AAB) Gluconobacter oxydans only recently the first tight system for regulatable target gene expression became available based on the heterologous repressor-activator protein AraC from Escherichia coli and the target promoter P. In this study, we tested pure repressor-based TetR- and LacI-dependent target gene expression in G. oxydans by applying the same plasmid backbone and construction principles that we have used successfully for the araC-P system. When using a pBBR1MCS-5-based plasmid, the non-induced basal expression of the Tn10-based TetR-dependent expression system was extremely low. This allowed calculated induction ratios of up to more than 3500-fold with the fluorescence reporter protein mNeonGreen (mNG). The induction was highly homogeneous and tunable by varying the anhydrotetracycline concentration from 10 to 200 ng/mL. The already strong reporter gene expression could be doubled by inserting the ribosome binding site AGGAGA into the 3' region of the P sequence upstream from mNG. Alternative plasmid constructs used as controls revealed a strong influence of transcription terminators and antibiotics resistance gene of the plasmid backbone on the resulting expression performance. In contrast to the TetR-P-system, pBBR1MCS-5-based LacI-dependent expression from P always exhibited some non-induced basal reporter expression and was therefore tunable only up to 40-fold induction by IPTG. The leakiness of P when not induced was independent of potential read-through from the lacI promoter. Protein-DNA binding simulations for pH 7, 6, 5, and 4 by computational modeling of LacI, TetR, and AraC with DNA suggested a decreased DNA binding of LacI when pH is below 6, the latter possibly causing the leakiness of LacI-dependent systems hitherto tested in AAB. In summary, the expression performance of the pBBR1MCS-5-based TetR-P system makes this system highly suitable for applications in G. oxydans and possibly in other AAB. KEY POINTS: • A pBBR1MCS-5-based TetR-P system was tunable up to more than 3500-fold induction. • A pBBR1MCS-5-based LacI-P system was leaky and tunable only up to 40-fold. • Modeling of protein-DNA binding suggested decreased DNA binding of LacI at pH < 6.
Topics: Acetic Acid; Gene Expression; Gluconobacter; Gluconobacter oxydans; Plasmids
PubMed: 34448898
DOI: 10.1007/s00253-021-11473-x -
Journal of Applied Microbiology Sep 2016A high-quality inoculum of Gluconacetobacter xylinus is important to produce bacterial cellulose (BC), a versatile biomaterial. This work aims to develop a method of...
AIMS
A high-quality inoculum of Gluconacetobacter xylinus is important to produce bacterial cellulose (BC), a versatile biomaterial. This work aims to develop a method of preparing an inoculum of this bacterium with high cell density and without mutants.
METHODS AND RESULTS
Inocula of G. xylinus ACCC 10220 without and with cellulase or carboxymethyl cellulose (CMC) were prepared in shaken culture. BC pellets and BC-negative mutants were present in the inoculum without additives but absent in the inoculum with additives. Based on BC weights statically produced in fresh BC-producing media initiated by different seed culture, the 24-h-shaken inoculum with 1·50% (w/v) CMC was the best because of high biomass and absence of mutants. The BC weights in fresh media inoculated by the 96-h-static inoculum and 24-h-shaken CMC inoculum at 7% (v/v) were 0·70 and 1·05 g l(-1) , respectively, implying significant difference (P < 0·01) in BC weights. However, structure properties of the two BC samples, including the crystallinity index, mass fraction of cellulose Iα , degree of polymerization (DP) and micromorphology were slightly different.
CONCLUSIONS
The 24-h-shaken CMC inoculum was the most suitable for a starter culture of BC.
SIGNIFICANCE AND IMPACT OF THE STUDY
A novel method of preparing G. xylinus inoculum in shaken culture was developed, featuring high biomass, absence of mutants and no BC entanglements. Cellulase or CMC added into the medium completely suppressed mutation of G. xylinus, and CMC facilitated to form colloidal BC with the low DP in shaken culture, indicating less BC stress to cells. These findings suggested the mutation could be induced by BC stress, and not by shear stress commonly accepted.
Topics: Bacteriological Techniques; Carboxymethylcellulose Sodium; Cellulase; Cellulose; Gluconacetobacter xylinus; Industrial Microbiology; Microscopy, Electrochemical, Scanning; Spectroscopy, Fourier Transform Infrared
PubMed: 27249070
DOI: 10.1111/jam.13193