Response of particle-attached and free-living bacterial communities to Microcystis blooms.

The massive proliferation of Microcystis threatens freshwater ecosystems and degrades water quality globally. Understanding the mechanisms that contribute to Microcystis growth is crucial for managing Microcystis blooms. The lifestyles of bacteria can be classified generally into two groups: particle-attached (PA; > 3 µm) and free-living (FL; 0.2-3.0 µm). However, little is known about the response of PA and FL bacteria to Microcystis blooms. Using 16S rRNA gene high-throughput sequencing, we investigated the stability, assembly process, and co-occurrence patterns of PA and FL bacterial communities during distinct bloom stages. PA bacteria were phylogenetically different from their FL counterparts. Microcystis blooms substantially influenced bacterial communities. The time decay relationship model revealed that Microcystis blooms might increase the stability of both PA and FL bacterial communities. A contrasting community assembly mechanism was observed between the PA and FL bacterial communities. Throughout Microcystis blooms, homogeneous selection was the major assembly process that impacted the PA bacterial community, whereas drift explained much of the turnover of the FL bacterial community. Both PA and FL bacterial communities could be separated into modules related to different phases of Microcystis blooms. Microcystis blooms altered the assembly process of PA and FL bacterial communities. PA bacterial community appeared to be more responsive to Microcystis blooms than FL bacteria. Decomposition of Microcystis blooms may enhance cooperation among bacteria. Our findings highlight the importance of studying bacterial lifestyles to understand their functions in regulating Microcystis blooms. KEY POINTS: • Microcystis blooms alter the assembly process of PA and FL bacterial communities • Microcystis blooms increase the stability of both PA and FL bacterial communities • PA bacteria seem to be more responsive to Microcystis blooms than FL bacteria.

Dynamic response of bacterial communities to Microcystis blooms: A three-year study.

Although nutrient availability is widely recognized as the driving force behind Microcystis blooms, identifying the microorganisms that play a pivotal role in their formation is a challenging task. Our understanding of the contribution of bacterial communities to the development of Microcystis blooms remains incomplete, despite the fact that the relationship between Microcystis and bacterial communities has been extensively investigated. Most studies have focused on their interaction for a single year rather than for multiple years. To determine key bacteria crucial for the formation of Microcystis blooms, we collected samples from three sites in the Daechung Reservoir (Chuso, Hoenam, and Janggye) over three years (2017, 2019, and 2020). Our results indicated that Microcystis bloom-associated bacterial communities were more conserved across stations than across years. Bacterial communities could be separated into modules corresponding to the different phases of Microcystis blooms. Dolichospermum and Aphanizomenon belonged to the same module, whereas the module of Microcystis was distinct. The microbial recurrent association network (MRAN) showed that amplicon sequence variants (ASVs) directly linked to Microcystis belonged to Pseudanabaena, Microscillaceae, Sutterellaceae, Flavobacterium, Candidatus Aquiluna, Bryobacter, and DSSD61. These ASVs were also identified as key indicators of the bloom stage, indicating that they were fundamental biological elements in the development of Microcystis blooms. Overall, our study highlights that, although bacterial communities change annually, they continue to share core ASVs that may be crucial for the formation and maintenance of Microcystis blooms.

Antibody-free and selective detection of okadaic acid using an affinity peptide-based indirect assay.

Okadaic acid (OA) is a type of marine biotoxin produced by some species of dinoflagellates in marine environments. Consumption of shellfish contaminated with OA can cause diarrhetic shellfish poisoning (DSP) in humans with symptoms that typically include abdominal pain, diarrhea and vomiting. In this study, we developed an affinity peptide-based direct competition enzyme-linked immunosorbent assay (dc-ELISA) for the detection of OA in real samples. The OA-specific peptide was successfully identified via M13 biopanning and a series of peptides were chemically synthesized and characterized their recognition activities. The dc-ELISA system showed good sensitivity and selectivity with a half-maximal inhibitory concentration (IC50) of 148.7 ng/mL and a limit of detection (LOD) of 5.41 ng/mL (equivalent, 21.52 ng/g). Moreover, the effectiveness of the developed dc-ELISA was validated using OA-spiked shellfish samples, and the developed dc-ELISA showed a high recovery rate. These results suggest that the affinity peptide-based dc-ELISA can be a promising tool for detecting OA in shellfish samples.

Transition metal complex-incorporated polyaniline as a platform for an enzymatic uric acid electrochemical sensor.

Uric acid (UA) is the primary waste product from purine metabolism in humans. Excessive UA levels in the body will accumulate in joints and form crystals that cause a wide range of health problems. An enzymatic electrochemical biosensor for UA based on the transition metal complex-incorporated polyaniline PANI-RC functionalized with both urate oxidase (UOx) as a specific bioreceptor and horseradish peroxidase (HRP) as a signal enhancer was developed. The transition metal complex being used herein is the commonly used redox couple (RC) in electrochemical biosensors, [Fe(CN)6]3-/4-, which plays the pivotal role of electron acceptors. This PANI-RC platform then becomes a conducive environment not only for enzyme immobilization but also for signal transfer improvement. The synergistic combination of HRP near UOx and RC anchored on the backbone of PANI helps in electron transfer from the enzymatic reaction to the current collector. The resulting PANI-RC-based UA sensor demonstrates high sensitivity with a detection limit of 11.4 µM, wide linear range, good stability, and excellent selectivity even in the presence of the most problematic interference in UA assays (e.g., ascorbic acid and urea). The recovery tests using artificial biofluid-spiked UA samples also showed promising results for practical usage of the PANI-RC-based UA sensor.

Sensitive Detection of BVDV Using Gold Nanoparticle-Modified Few-Layer Black Phosphorus with Affinity Peptide-Based Electrochemical Sensor.

The lethality of the bovine viral diarrhea virus (BVDV) in cattle involves inapparent infection and various, typically subclinical, syndromes. Cattle of all ages are vulnerable to infection with the virus. It also causes considerable economic losses, primarily due to reduced reproductive performance. In the absence of treatment that can completely cure infected animals, detection of BVDV relies on highly sensitive and selective diagnosis methods. In this study, an electrochemical detection system was developed as a useful and sensitive system for the detection of BVDV to suggest the direction of diagnostic technology through the development of conductive nanoparticle synthesis. As a countermeasure, a more sensitive and rapid BVDV detection system was developed using the synthesis of electroconductive nanomaterials black phosphorus (BP) and gold nanoparticle (AuNP). To increase the conductivity effect, AuNP was synthesized on the BP surface, and the stability of BP was improved by using dopamine self-polymerization. Moreover, its characterizations, electrical conductivity, selectivity, and sensitivity toward BVDV also have been investigated. The BP@AuNP-peptide-based BVDV electrochemical sensor exhibited a low detection limit of 0.59 copies mL-1 with high selectivity and long-term stability (retaining 95% of its initial performance over 30 days).

The cyanobactericidal bacterium Paucibacter aquatile DH15 caused the decline of Microcystis and aquatic microbial community succession: A mesocosm study.

Microcystis blooms pose a major threat to the quality of drinking water. Cyanobactericidal bacteria have attracted much attention in the research community as a vehicle for controlling Microcystis blooms because of their ecological safety. Nonetheless, most studies on cyanobactericidal bacteria have been conducted on a laboratory scale but have not been scaled-up as field experiments. Thus, our understanding of the microbial response to cyanobactericidal bacteria in natural ecosystems remains elusive. Herein, we applied Paucibacter aquatile DH15 to control Microcystis blooms in a 1000 L mesocosm experiment and demonstrated its potential with the following results: (1) DH15 reduced Microcystis cell density by 90.7% within two days; (2) microcystins released by Microcystis death decreased to the control level in four days; (3) during the cyanobactericidal processes, the physicochemical parameters of water quality remained safe for other aquatic organisms; and (4) the cyanobactericidal processes promoted the growth of eukaryotic microalgae, replacing cyanobacteria. The cyanobactericidal processes accelerated turnover rates, decreased stability, and altered the functional profile of the microbial community. Network analysis demonstrated that this process resulted in more complex interactions between microbes. Overall, our findings suggest that strain DH15 could be considered a promising candidate for controlling Microcystis blooms in an eco-friendly manner.

Aquariibacter albus gen. nov., sp. nov., a new member of the order Burkholderiales, isolated from a freshwater aquarium.

A novel bacterium, strain SJAQ100T, was isolated from a freshwater aquarium and was characterized taxonomically and phylogenetically. Strain SJAQ100T was a Gram-stain-negative, aerobic, rod-shaped and non-motile bacterium. The strain grew optimally with 0 % NaCl and at 25-37 °C on Reasoner's 2A agar. Phylogenetic analysis based on the 16S rRNA gene sequences revealed that the strain SJAQ100T clustered with members of Burkholderiales incertae sedis in the order Burkholderiales, but sequence similarities to known species were less than 96.5 %. The genomic DNA G+C content of strain SJAQ100T was 71.2 mol%. Genomic comparisons of strain SJAQ100T with species in the order Burkholderiales were made using the Genome-to-Genome Distance Calculator, average nucleotide identity and average amino acid identity analyses (values indicated ≤22.1, ≤78.1, and ≤68.1 % respectively). Strain SJAQ100T contained C16 : 0 and C16 : 1 ω7c/C16 : 1 ω6c as major fatty acids and Q-8 as the major quinone. The major polyamines were putrescine and cadaverine. Strain SJAQ100T contained phosphatidylethanolamine and diphosphatidylglycerol as major polar lipids. Based on the genotypic, chemotaxonomic and phenotypic results, strain SJAQ100T represents a novel genus and species, Aquariibacter albus gen. nov., sp. nov., which belongs to order Burkholderiales and the class Betaproteobacteria. The type strain is SJAQ100T (=KCTC 72203T=CGMCC 1.18869T=MCC 4385T).

In Situ Biosynthesis of a Metal Nanoparticle Encapsulated in Alginate Gel for Imageable Drug-Delivery System.

Development of drug-delivery systems that allow simultaneous in vivo imaging has gained much interest. We report a novel strategy to encapsulate metal nanoparticles (NPs) within alginate gel for in vivo imaging. The cell lysate of recombinant Escherichia coli strain, expressing Arabidopsis thaliana phytochelatin synthase and Pseudomonas putida metallothionein genes, was encapsulated within the alginate gel. Incubation of alginate gel with metal ion precursors followed by UV irradiation resulted in the synthesis of high concentrations of metal NPs, such as Au, Ag, CdSe, and EuSe NPs, within the gel. The alginate gel with metal NPs was used as a drug-delivery system by further co-encapsulating doxorubicin and rifampicin, the release of which was made to be pH-dependent. This system can be conveniently and safely used for in vitro and in vivo bioimaging, enabled by the metal NPs formed within the gel matrix without using toxic reducing reagents or surfactants.

Facile synthesis of carbon dots from Tagetes erecta as a precursor for determination of chlorpyrifos via fluorescence turn-off and quinalphos via fluorescence turn-on mechanisms.

Convenient one-pot synthetic route for the fabrication of carbon dots (CDs) from Tagetes erecta flower (TEF), named as "TEF-CDs', through solvo(hydro)-thermal carbonization of the TEF was developed. The TEF-CDs revealed high selectivity towards chlorpyrifos and quinalphos, acting as a fluorescent probe. The CDs synthesized from T. erecta flower showed a strong blue color at 495 nm when excited at 420 nm, along with the exhibition of a strong quantum yield of 63.7%. The synthesized CDs revealed their richness in the surface-active organic group that synthesized CDs from T. erecta flower are mainly comprised of C, O, and N, which were crystalline in structure that was revealed by TEM image and XRD spectra. Furthermore, when the probe was exposed to different pH conditions, no major noticeable changes were recorded. Moreover, when the probe was exposed to chlorpyrifos and quinalphos, we have noticed that fluorescence spectra was turned off when the probe was exposed to chlorpyrifos and "turned on" after the exposure quinalphos. Moreover, fluorescence spectral changes showed a good linearity with chlorpyrifos and quinalphos concentrations in the range of 0.05-100.0 µM for chlorpyrifos and 0.01-50.0 µM for quinalphos. The limit of detection are 2.1 ng mL-1 and 1.7 ng mL-1 for chlorpyrifos and quinalphos, respectively. Finally, the TEF-CDs-based fluorescent nanoprobe was successfully applied to estimate chlorpyrifos and quinalphos with an effective accuracy in rice and fruit samples with rapid detection time.

Tabrizicola algicola sp. nov. isolated from culture of microalga Ettlia sp.

A novel Gram-stain-negative, aerobic, non-spore-forming, non-motile, and rod-shaped bacterium, strain ETT8T was isolated from a chemostat culture of microalga Ettlia sp. YC001. Optimal growth was with 0-2% NaCl and at 25-37 °C on R2A medium. Phylogenetic analysis based on the 16S rRNA gene and genome sequence showed that strain ETT8T belongs to the genus Tabrizicola, with the close neighbours being T. sediminis DRYC-M-16T (98.1 %), T. alkalilacus DJCT (97.6 %), T. fusiformis SY72T (96.9 %), T. piscis K13M18T (96.8 %), and T. aquatica RCRI19T (96.5 %). The genomic comparison of strain ETT8T with type species in the genus Tabrizicola was analysed using the genome-to-genome distance calculator (GGDC), average nucleotide identity (ANI), and average amino acid identity (AAI) (values indicated ≤17.7, ≤75.4 and ≤71.9 %, respectively). The genomic DNA G+C content of strain ETT8T was 64.4 %, plus C18 : 1 ω6c and C18 : 0-iso were the major fatty acids and Q-10 the major respiratory quinone. Strain ETT8T contained diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine, phosphatidylcholine aminolipid, and four unidentified lipids as the major polar lipids. Based on the chemotaxonomic, genotypic, and phenotype results, strain ETT8T was recognized as a novel species of the genus Tabrizicola for which the name Tabrizicola algicola sp. nov. is proposed. The type strain is ETT8T (=KCTC 72206T=JCM 31893T=MCC 4339T).

Fluorescence detection of histamine based on specific binding bioreceptors and carbon quantum dots.

Histamine is primarily found in spoiled food and often used as an indicator of food safety. Compared to various existing methods for analyzing histamine, high-performance liquid chromatography (HPLC) which is accurate but time-consuming, and immunochemical methods that are difficult to produce high specificity and affinity antibodies towards small molecules have been used. In this study, we developed a newly designed, sensitive, and selective fluorescence detection platform for histamine sensing, utilizing carbon quantum dots (CQDs) and synthetic peptides. Specifically, through biopanning approaches, a series of peptides having a high affinity towards immobilized histamine hapten were selected from phage-displayed libraries. Then, CQDs were synthesized by one-pot hydrothermal treatment enabling their fluorescence to be effectively quenched by peptides via the electron transfer interactions. While, in the presence of histamine, fluorescence will be recovered because of the stronger interaction between peptide and target. In this study, from the selectivity tests towards histamine and in contrast to structurally similar compounds, peptide Hisp3 (DIDRAGKASHWP) along with its dipeptide repeat derivative (Hisp3-2-C) were chemically synthesized to be used as promising histamine receptors. Furthermore, the application of peptide along with gold-coated magnetic nanoparticles (MNP@Au NPs) was designed for purification and analysis of fish samples. These results indicate that the CQDs and peptide sensor system could detect histamine at lower concentrations with high sensitivity and selectivity.

Fabrication of a paper strip for facile and rapid detection of bovine viral diarrhea virus via signal enhancement by copper polyhedral nanoshells.

The detection of bovine viral diarrhea virus (BVDV), which is a pathogen inducing fatal gastrointestinal disease in cattle, is becoming a momentous issue in the livestock farm. In that, BVDV is related to inapparent infection and various diseases with high transmissibility; it has also led to considerable economic losses. In this study, a simple dot-blotting method was devised to construct a rapid screening system for BVDV. Based on the BVDV-specific bioreceptors, it was anchored on the gold nanoparticles (AuNPs) to generate the seeding sites for signaling; then the signals were amplified by adopting the overgrowth of copper nano-polyhedral shells on AuNPs. The developed detection system shows a low detection limit of 4.4 copies per mL, and even this could be distinguished with naked eyes. These results indicate that the designed nanobiosensor possesses not only high sensitivity and selectivity but also potential usage on a point-of-care testing platform for BVDV.

Cu-nanoflower decorated gold nanoparticles-graphene oxide nanofiber as electrochemical biosensor for glucose detection.

A novel electrospinning approach is proposed for the fabrication of copper (Cu)-nanoflower decorated gold nanoparticles (AuNPs)-graphene oxide (GO) nanofiber (NF) as an electrochemical biosensor for the glucose detection. In this study, GO was mixed with poly(vinyl alcohol) (PVA) and used as a fiber precursor, which greatly improves the electrochemical properties. The above solution was uniformly coated onto the surfaces of gold chip to form GO NFs via electrospinning. AuNPs were coated onto the surface of GO NFs and then incorporated organic-inorganic hybrid nanoflower [Cu nanoflower-glucose oxidase (GOx) and horseradish peroxidase (HRP)]. The electrochemical experiments revealed that Cu-nanoflower@AuNPs-GO NFs exhibited outstanding electrochemical catalytic nature, and selectivity for the conversion of glucose to gluconic acid in the presence of GOx-HRP-Cu nanoflower. The Cu-nanoflower@AuNPs-GO NFs coated Au chip exhibited good linear range 0.001-0.1 mM, with a detection limit of 0.018 µM. The Cu-nanoflower@AuNPs-GO NFs modified Au chip exhibited higher catalytic properties, which are attributed to the coating of unique organic-inorganic nanostructured materials on the surfaces of Au chip. These results indicate that the nano-bio hybrid materials can be applied as a promising electrochemical biosensor to monitor glucose levels in biofluids.

Development of a rapid and sensitive electrochemical biosensor for detection of human norovirus via novel specific binding peptides.

Human noroviruses cause acute foodborne gastroenteritis outbreaks worldwide. In this study, a highly sensitive and selective electrochemical biosensor was fabricated for the detection of human norovirus using novel peptides as recognition elements. The electrochemical biosensor was fabricated by assembling of eight novel peptides separately on the gold electrode and investigated their efficiencies for sensing human noroviruses. Among eight peptides, NoroBP peptide coated onto the gold electrode exhibited a high binding affinity towards human noroviruses, resulting a progressive decrease in current signals with increasing concentration of human norovirus (0-105 copies/mL). As a result, NoroBP-nonFoul(FlexL)2-coated gold electrode acts an efficient electrochemical biosensor for highly selective detection of human norovirus with a detection limit of 1.7 copies/mL, which is 3-fold lower than the reported methods. The developed electrochemical biosensor was successfully applied to detect human norovirus prepared by standard procedure from oyster, which suggests that the developed biosensor can be used as a very sensitive and selective point-of-care bioanalytical platform for the detection of human norovirus in various food samples.

Morphological evolution of upconversion nanoparticles and their biomedical signal generation.

Advancements in the fabrication of upconversion nanoparticles (UCNPs) for synthetic control can enable a broad range of applications in biomedical systems. Herein, we experimentally verified the role of the hydrothermal reaction (HR) time in the synthesis of NaYF4:20%Yb3+/3%Er3+ UCNPs on their morphological evolution and phase transformation at different temperatures. Characterizations of the as-prepared UCNPs were conducted using X-ray diffraction (XRD), electron microscopy and spectroscopy, and thermogravimetric and upconversion (UC) luminescence analysis. We demonstrated that determining the optimal HR time, also referred to here as the threshold time, can produce particles with good homogeneity, hexagonal phase, and UC luminescence efficiency. Subsequently, the polymer coated UCNPs maintained their original particle size distribution and luminescence properties, and showed improved dispersibility in a variety of solvents, cellular nontoxicity, in vitro bioimaging, and biocompatibility as compared to the bare UCNP. Besides this, polyacrylic acid conjugated UCNPs (UCNP@PAA) also revealed the strong anticancer effect by conjugating with doxorubicin (DOX) as compared to the free DOX. Based on these findings, we suggest that these particles will be useful in drug-delivery systems and as in vivo bioimaging agents synchronously.

Hexagonal tungsten oxide nanoflowers as enzymatic mimetics and electrocatalysts.

Tungsten oxide (WOx) has been widely studied for versatile applications based on its photocatalytic, intrinsic catalytic, and electrocatalytic properties. Among the several nanostructures, we focused on the flower-like structures to increase the catalytic efficiency on the interface with both increased substrate interaction capacities due to their large surface area and efficient electron transportation. Therefore, improved WOx nanoflowers (WONFs) with large surface areas were developed through a simple hydrothermal method using sodium tungstate and hydrogen chloride solution at low temperature, without any additional surfactant, capping agent, or reducing agent. Structural determination and electrochemical analyses revealed that the WONFs have hexagonal Na0.17WO3.085·0.17H2O structure and exhibit peroxidase-like activity, turning from colorless to blue by catalyzing the oxidation of a peroxidase substrate, such as 3,3',5,5'-tetramethylbenzidine, in the presence of H2O2. Additionally, a WONF-modified glassy carbon electrode was adopted to monitor the electrocatalytic reduction of H2O2. To verify the catalytic efficiency enhancement by the unique shape and structure of the WONFs, they were compared with calcinated WONFs, cesium WOx nanoparticles, and other peroxidase-like nanomaterials. The results indicated that the WONFs showed a low Michaelis-Menten constant (km), high maximal reaction velocity (vmax), and large surface area.