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Bacteria frequently engage in obligate metabolic mutualisms with other microorganisms. However, it remains generally unclear how the resulting metabolic dependencies affect the ecological niche space accessible to the whole consortium relative to the niche space available to its constituent individuals. Here we address this issue by systematically cultivating metabolically dependent strains of different bacterial species either individually or as pairwise cocultures in a wide range of carbon sources. Our results show that obligate cross-feeding is significantly more likely to expand the metabolic niche space of interacting bacterial populations than to contract it. Moreover, niche expansion occurred predominantly between two specialist taxa and correlated positively with the phylogenetic distance between interaction partners. Together, our results demonstrate that obligate cross-feeding can significantly expand the ecological niche space of interacting bacterial genotypes, thus explaining the widespread occurrence of this type of ecological interaction in natural microbiomes., (© 2021. The Author(s), under exclusive licence to Springer Nature Limited.)

Species of the genus Shewanella are widespread in nature in various habitats, however, little is known about phages affecting Shewanella sp. Here, we report the isolation of phages from diverse freshwater environments that infect and lyse strains of Shewanella oneidensis and other Shewanella sp. Sequence analysis and microscopic imaging strongly indicate that these phages form a so far unclassified genus, now named Shewanella phage Thanatos, which can be positioned within the subfamily of Tevenvirinae ( Duplodnaviria ; Heunggongvirae ; Uroviricota ; Caudoviricetes ; Caudovirales ; Myoviridae ; Tevenvirinae ). We characterized one member of this group in more detail using S. oneidensis MR-1 as a host. Shewanella phage Thanatos-1 possesses a prolate icosahedral capsule of about 110 nm in height and 70 nm in width and a tail of about 95 nm in length. The dsDNA genome exhibits a GC content of about 34.5%, has a size of 160.6 kbp and encodes about 206 proteins (92 with an annotated putative function) and two tRNAs. Out of those 206, MS analyses identified about 155 phage proteins in PEG-precipitated samples of infected cells. Phage attachment likely requires the outer lipopolysaccharide of S. oneidensis , narrowing the phage's host range. Under the applied conditions, about 20 novel phage particles per cell were produced after a latent period of approximately 40 min, which are stable at a pH range from 4 to 12 and resist temperatures up to 55°C for at least 24 h. Addition of Thanatos to S. oneidensis results in partial dissolution of established biofilms, however, early exposure of planktonic cells to Thanatos significantly enhances biofilm formation. Taken together, we identified a novel genus of Myophages affecting S. oneidensis communities in different ways., (Copyright © 2020 Kreienbaum, Dörrich, Brandt, Schmid, Leonhard, Hager, Brenzinger, Hahn, Glatter, Ruwe, Briegel, Kalinowski and Thormann.)

Bacterial extracellular nucleases have multiple functions in processes as diverse as nutrient acquisition, natural transformation, biofilm formation, or defense against neutrophil extracellular traps (NETs). Here we explored the properties of ExeM in Shewanella oneidensis MR-1, an extracellular nuclease, which is widely conserved among species of Shewanella, Vibrio, Aeromonas , and others. In S. oneidensis , ExeM is crucial for normal biofilm formation. In vitro activity measurements on heterologously produced ExeM revealed that this enzyme is a sugar-unspecific endonuclease, which requires Ca 2+ and Mg 2+ /Mn 2+ as co-factors for full activity. ExeM was almost exclusively localized to the cytoplasmic membrane fraction, even when a putative C-terminal membrane anchor was deleted. In contrast, ExeM was not detected in medium supernatants. Based on the results we hypothesize that ExeM predominantly interacts with DNA in close proximity to the cell, e.g., to promote biofilm formation and defense against NETs, or to control uptake of DNA.

Bacterial species such as Shewanella oneidensis MR-1 require extracellular nucleolytic activity for the utilization of extracellular DNA (eDNA) as a source of nutrients and for the turnover of eDNA as a structural matrix component during biofilm formation. We have previously characterized two extracellular nucleases of S. oneidensis MR-1, ExeM and ExeS. Although both are involved in biofilm formation, they are not specifically required for the utilization of eDNA as a nutrient. Here we identified and characterized EndA, a third extracellular nuclease of Shewanella. The heterologously overproduced and purified protein was highly active and rapidly degraded linear and supercoiled DNAs of various origins. Divalent metal ions (Mg(2+) or Mn(2+)) were required for function. endA is cotranscribed with phoA, an extracellular phosphatase, and is not upregulated upon phosphostarvation. Deletion of endA abolished both extracellular degradation of DNA by S. oneidensis MR-1 and the ability to use eDNA as a sole source of phosphorus. PhoA is not strictly required for the exploitation of eDNA as a nutrient. The activity of EndA prevents the formation of large cell aggregates during planktonic growth. However, in contrast to the findings for ExeM, endA deletion had only minor effects on biofilm formation. The findings strongly suggest that the extracellular nucleases of S. oneidensis exert specific functions required under different conditions.

A semiautomated fluorometric method for the determination of resperpine in tablets was collaboratively studied by 7 laboratories. The method is a modification of the semiautomated method of Urbányi and Stober, which involves formation of a fluorogen with vanadium pentoxide. Collaborators were supplied with 3 composites, each from a different dosage level of commercial tablets. The results obtained agreed well with the AOAC manual fluorometric method; coefficients of variation ranged from 0.45 to 2.70%. The method has been adopted as official first action.

The stability of pilocarpine hydrochloride and pilocarpine nitrate ophthalmic solutions stored in hospital pharmacies across the United States was studied. Through a voluntary drug stability program, FDA selected 252 samples (representing 11 manufacturers) from pharmacies representing an adequate cross section of the country. The samples were analyzed for strength, identification, pH, and isopilocarpine and pilocarpic acid impurities. All samples of pilocarpine nitrate met USP requirements. Eight samples of pilocarpine hydrochloride had tablets that exceeded the USP upper limit for strength. All of these samples were in 1- and 2-mL bottles. The amount of isopilocarpine found ranged from 1 to 6.4%, and the amount of pilocarpic acid from 1.5 to 10.1%. Although pilocarpine salts in ophthalmic solution decompose into isopilocarpine and pilocarpic acid under various conditions of storage, an amount of pilocarpine is maintained that is within the compendial limits. However, there is a problem of evaporation from some of the 1- and 2-mL containers in which this product is supplied.

The stability of betamethasone sodium phosphate, hydrocortisone sodium phosphate, and prednisolone sodium phosphate, injections stored in hospital pharmacies across the United States was studied. Through a voluntary drug stability program, FDA selected 58 samples (representing two manufacturers) from pharmacies representing a cross section of the country. The samples were analyzed for strength, identification, pH, and related impurities. All of the hydrocortisone sodium phosphate and prednisolone sodium phosphate samples met USP requirements for strength and pH. Assays of the betamethasone sodium phosphate injection samples yielded results that were in compliance with the manufacturer's strength and pH limits; there are no USP requirements for betamethasone sodium phosphate injection. All samples were within USP limits for related free steroids. Betamethasone sodium phosphate, hydrocortisone sodium phosphate, and prednisolone sodium phosphate injections obtained from hospital pharmacies appear to be stable after storage under actual marketplace conditions.

A reversed-phase high-performance liquid chromatographic (HPLC) analysis of oral suspensions for nitrofurantoin (1) and 3-(5-nitrofurfurylideneamino)hydantoic acid (2), an impurity derived from 1, is presented. The concentration of the impurity ranged from 20 to 300 micrograms/mL in several lots of commercial oral suspensions. Conversion of 1 to 2 with citrate buffer, an excipient in the oral suspension, was achieved; selective hydantoin ring cleavage was accomplished in preference to the generally observed cleavage at the azomethine linkage. The hydantoic acid 2 was synthesized and identified by NMR, IR, TLC, and elemental analysis.

Contaminants in a commercial enema sample of theophylline olamine were found to be derived from parabens and ethanolamine. These contaminants, whose presence was characterized by the loss of preservatives and solubilizing agent, were isolated directly from the drug sample and identified. A high-performance liquid chromatographic (HPLC) system was developed to separate completely and to measure quantitatively theophylline, the two impurities, 4-hydroxybenzoic acid and N-(2-hydroxyethyl)-4-hydroxybenzamide, and the remaining parabens. The material balance obtained from the results of quantitative HPLC indicated the formation of these impurities at the expense of parabens. TLC, IR, and UV spectrophotometry and NMR and mass spectrometric analyses were used for identification or for comparisons of the new compound and the known N-(2-hydroxyethyl)benzamide.