Rapid progress in various advanced analytical methods, such as single-cell technologies, enable unprecedented and deeper understanding of microbial ecology beyond the resolution of conventional approaches. A major application challenge exists in the determination of sufficient sample size without sufficient prior knowledge of the community complexity and, the need to balance between statistical power and limited time or resources. This hinders the desired standardization and wider application of these technologies. Here, we proposed, tested and validated a computational sampling size assessment protocol taking advantage of a metric, named kernel divergence. This metric has two advantages: First, it directly compares data set-wise distributional differences with no requirements on human intervention or prior knowledge-based preclassification. Second, minimal assumptions in distribution and sample space are made in data processing to enhance its application domain. This enables test-verified appropriate handling of data sets with both linear and nonlinear relationships. The model was then validated in a case study with Single-cell Raman Spectroscopy (SCRS) phenotyping data sets from eight different enhanced biological phosphorus removal (EBPR) activated sludge communities located across North America. The model allows the determination of sufficient sampling size for any targeted or customized information capture capacity or resolution level. Promised by its flexibility and minimal restriction of input data types, the proposed method is expected to be a standardized approach for sampling size optimization, enabling more comparable and reproducible experiments and analysis on complex environmental samples. Finally, these advantages enable the extension of the capability to other single-cell technologies or environmental applications with data sets exhibiting continuous features.

Bisphosphonic acids, which are structural analogs of pyrophosphate, constitute a class of compounds with very high potential for the construction of effective inhibitors of enzymes operating on oligo- and polyphosphates. The bisphosphonate-based methodology was applied for the discovery of inhibitors of two families of polyphosphate kinases (PPK1 and PPK2). Screening of thirty-two structurally diverse bisphosphonic acids and related compounds revealed several micromolar inhibitors of both enzymes. Importantly, selectivity of bisphosphonates could be achieved by application of the appropriate side chain.

Hemolytic-uremic syndrome (HUS) is a thrombotic microangiopathy that is characterized by microangiopathic hemolytic anemia, thrombocytopenia, and renal failure. Excess complement activation underlies atypical HUS and is evident in Shiga toxin-induced HUS (STEC-HUS). This Spotlight focuses on new knowledge of the role of Escherichia coli-derived toxins and polyphosphate in modulating complement and coagulation, and how they affect disease progression and response to treatment. Such new insights may impact on current and future choices of therapies for STEC-HUS.

The impact of polyphosphate-accumulating organism (PAO) and glycogen-accumulating organism (GAO) populations as well as of the chemical profile on the performance of Unit-3 (open elutriation tanks) and Unit-5 (covered elutriation tank) of the City of Johannesburg Northern Wastewater Treatment Works was determined. Physicochemical parameters of wastewater samples were measured using standard methods. Bacterial diversity was determined using 16S rRNA gene amplicon pyrosequencing of the variable region V1-3. Results showed soluble COD concentrations from settled sewage for Unit-3 at 192.8 mg COD/L and for Unit-5 at 214.6 mg COD/L, which increased to 301.8 mg COD/L and 411.6 mg COD/L in the overflow from elutriation tanks and decreased to 170.9 mg COD/L and 256.3 mg COD/L at the division boxes, respectively. Both long-chain volatile fatty acids (heptanoic acid, isobutyric acid, 3-methylbutanoic acid, pentanoic acid, 4-methylpentanoic acid, methylheptanoic acid) and short-chain volatile fatty acids (acetic acid, propionic acid, isobutyric acid) were present within concentration ranges of 17.19 mg/L to 54.98 mg/L and 13.64 mg/L to 87.6 mg/L for Unit 3 and 38.61 mg/L to58.85 mg/L and 21.63 mg/L to 92.39 mg/L for Unit 5, respectively. In the secondary settling tanks, the phosphate-removal efficiency in Unit-5 appeared to be slightly higher (0.08 mg P/L) compared to that of Unit-3 (0.11 mg P/L). The average DO concentrations (2.1 mg/L and 2.2 mg/L) as well as the pH values (pH 7 to pH 7.5) were found to be slightly higher in Unit-5 in the aerobic zones. The high presence of PAOs in the bioreactors (Unit-5: Dechloromonas (14.96%), Acinetobacter (6.3%), Zoogloea (4.72%) in the anaerobic zone and Dechloromonas (22.37 %) in the aerobic zone; Unit-3: Dechloromonas (37.25%) in the anaerobic zone and Dechloromonas (23.97%) in the aerobic zone) confirmed the phosphate-removal efficiencies of both units. Negligible GAOs were found in the aerobic zones (Defluviicoccus spp.: 0.33% for Unit-5 and 0.68% for Unit-3) and in the anaerobic zones (Defluviicoccus: 9.8% for Unit-3). The high microbial diversity and a negligible percentage of GAOs in Unit-5 could contribute to its high phosphate-removal efficiency, although results did not indicate statistically significant differences between the unit with a covered elutriation tank (Unit-5) and that with open elutriation tanks (Unit-3).

The kinetics of in vitro drug release from nanoparticulate systems is extensive, though uncritically, being studied by dialysis. Evaluating the actual relevance of dialysis data to drug release was the purpose of this study. Diclofenac- or ofloxacin-loaded chitosan nanoparticles crosslinked with tripolyphosphate were prepared and characterized. With each drug, dynamic dialysis was applied to nanoparticle dispersion, solution containing dissolved chitosan·HCl, and solution of plain drug. Drug kinetics in receiving phase (KRP), nanoparticle matrix (KNM) and nanoparticle dispersion medium (KDM) were determined. Release of each drug from nanoparticles was also assessed by ultracentrifugation. Although KRP data may be interpreted in terms of sustained release from nanoparticles, KNM and KDM data show that, with both drugs, the process was in fact controlled by permeation across dialysis membrane. Analysis of KRP data reveals a reversible interaction of diclofenac with dispersed nanoparticle surface, similar to the interaction of this drug with dissolved chitosan·HCl. No such interactions are noticed with ofloxacin. The results from the ultracentrifugation method agree with the above interpretation of dialysis data. This case study shows that dialysis data from a nanoparticle dispersion is not necessarily descriptive of sustained-release from nanoparticles, hence, if interpreted uncritically, it may be misleading.

The principal polyphosphate-accumulating organism (PAO) in a biological-phosphate-removal activated-sludge process was assessed microscopically. The organism was recognized by its distinct morphotype most easily after polyphosphate staining. The PAO occurred in large, homogeneous clusters. The cells of the PAO were the biggest cells abounding in the sludge--clearly bigger than average sludge bacteria. Typical of the principal PAO was a variation of cell size, even in fresh sludge. In acetate minimal medium containing ampicillin, the original principal PAO clusters were converted to clusters of clearly larger, polyphosphate-containing, vegetative yeast-like cells. Cycloheximide addition inhibited this and caused flock disintegration, disappearance of the principal PAO clusters and growth of free bacteria. The cell wall of the principal PAO was not of the usual bacterial character. It showed anomalous Gram staining, stained for chitin (not found in bacteria) and bound concanavalin A, like cell walls of many yeasts. In addition, the PAO cell wall was resistant to lysozyme, but sensitive to an enzyme mixture that lyses yeast cell walls. It was concluded that the principal PAO cells in the studied sludge were clustered spores of a yeast.

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