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The emergence of multicellularity is a key event in the evolution of life and is an attractive challenge among researchers, including those investigating the artificial design of cellular behavior1. Multicellular organisms are widely distributed on Earth, and retracing the specific conditions conducive for the initial transition from unicellularity to multicellularity is difficult. However, by examining organisms that inhabit unique (e.g., isolated) environmental niches, we may be able to get a glimpse into primitive multicellularity in the context of a given environment. Here we report the discovery of a new bacterium that displayed multicellular-like characteristics and behavior. The bacterium, which was isolated adjacent to an underground stream in a limestone cave, is to be named Jeongeupia sacculi sp. nov. HS-3. On a solid surface, HS-3 self-organizes its filamentous cells to form an appearance similar to the nematic phase of a liquid crystal2. Mature colonies produce and accommodate clusters of coccobacillus progeny, and release them upon contact with water. HS-3 demonstrated novel, spatiotemporally regulated multicellularity that can resolve the so-called ‘competition-dispersal trade-off’ problem3. This study illustrates a hypothetical missing link on the emergence of multicellularity.

An aggregation-induced emission chromophore, vinylpyrrole, was prepared from a formylpyrrole derivative, Meldrum's acid, and 1,3-dimethylbarbituric acid. The optical properties of the chromophore both in the solution and solid states were investigated by UV-vis and fluorescence spectroscopy. Single crystal X-ray diffraction measurements revealed that the dimethylbarbituric acid adduct formed a J-aggregate in the solid and resulted in higher fluorescence quantum yield compared to the Meldrum's acid adduct. Emission enhancement was found to occur by the restriction of molecular rotation in the solid state.

Halomonas sp. O-1 is a halophilic bacterium with a high potential for industrial application due to its natural ability to produce polyhydroxyalkanoates (PHAs) using seawater-based media. However, a major barrier preventing industrial scale implementation of this organism is a lack of molecular methodologies capable of readily transforming members of the Halomonas genus. Currently, the only reliable method used for introducing DNA into Halomonas spp. is bacterial conjugation, a somewhat tedious and time-consuming technique compared to electroporation-based methodologies. Here we describe a rapid and reproducible method for the electroporation of Halomonas sp. O-1 with plasmid DNA. Electrocompetent cells were generated by growing Halomonas sp. O-1 in a yeast extract-tryptone medium with a final salinity of 3.5%, pH of 7.5, followed by several washes using 300mM sucrose. Results show that plasmids containing chloramphenicol (Cm(R)) and gentamicin (Gm(R)) resistance cassettes are suitable antibiotic selection markers for transformation and yields of 10(4) transformants per μg of DNA were obtained. This method is simple to perform and the materials used are readily available in most research labs. Additionally, this plasmid-based transformation procedure has the potential to be adapted for a number of applications including the creation of recombinant stains and the generation of deletion mutants of Halomonas spp.

This review highlights the recent investigations of class IV polyhydroxyalkanoate (PHA) synthases, the newest classification of PHA synthases. Class IV synthases are prevalent in organisms of the Bacillus genus and are composed of a catalytic subunit PhaC (approximately 40 kDa), which has a PhaC box sequence ([GS]-X-C-X-[GA]-G) at the active site, and a second subunit PhaR (approximately 20 kDa). The representative PHA-producing Bacillus strains are Bacillus megaterium and Bacillus cereus; the nucleotide sequence of phaC and the genetic organization of the PHA biosynthesis gene locus are somewhat different between these two strains. It is generally considered that class IV synthases favor short-chain-length monomers such as 3-hydroxybutyrate (C4) and 3-hydroxyvalerate (C5) for polymerization, but can polymerize some unusual monomers as minor components. In Escherichia coli expressing PhaRC from B. cereus YB-4, the biosynthesized PHA undergoes synthase-catalyzed alcoholytic cleavage using endogenous and exogenous alcohols. This alcoholysis is thought to be shared among class IV synthases, and this reaction is useful not only for the regulation of PHA molecular weight but also for the modification of the PHA carboxy terminus. The novel properties of class IV synthases will open up the possibility for the design of new PHA materials.

Polyhydroxyalkanoate (PHA) synthase from the bacterium Bacillus cereus YB-4 (PhaRC YB4 ), a Class IV PHA synthase, shows both PHA polymerization and alcoholysis activities. In PhaRC YB4 -expressing Escherichia coli in the presence of ethanol, biosynthesized PHA is subjected to an ethyl-esterification at the carboxy terminus as a result of alcoholytic cleavage of the PHA chain by PhaRC YB4 . In this study, we investigated the range of alcohol species available for alcoholysis by PhaRC YB4 using E. coli JM109 as an assay host by monitoring the molecular weight of PHA during the second step of a two-step cultivation. PhaRC YB4 utilized various alcohols other than ethanol for alcoholysis. Through the use of bifunctional compounds for alcoholysis, the PHA carboxy terminus could be modified with thiol, alkynyl, hydroxy, and benzyl groups, resulting in the functionalization of the PHA carboxy terminus. Furthermore, one-step production of carboxy-terminal functionalized PHA was developed using E. coli XL1-Blue as a production host.

In certain conditions, members of the Lactobacillus genus are auxotrophs that have fastidious requirements for growth. Notably, Lactobacillus cannot grow in M9 medium, a minimal synthetic medium used for Escherichia coli. However, we found that some Lactobacillus strains can be grown in M9 when co-cultured with E. coli K-12. In the co-culture, L. casei proliferates exponentially, reaching cell densities of 108 CFU (colony-forming unit) ml−1 in 6 h and dominating E. coli in the late growth phase. Spent medium from E. coli grown overnight lacked this growth-promoting effect on L. casei. Similarly, the effect was not observed when the species were separated by a 0.4-µm membrane. Microscopic observations showed that L. casei are embedded in the micro-scale clusters of E. coli in the early growth phase. This study describes for the first time the ability of a Lactobacillus species to grow in minimal medium when in close proximity with co-cultured bacteria.

Bacillus cereus and Bacillus megaterium both accumulate polyhydroxyalkanoate (PHA) but their PHA biosynthetic gene (pha) clusters that code for proteins involved in PHA biosynthesis are different. Namely, a gene encoding MaoC-like protein exists in the B. cereus-type pha cluster but not in the B. megaterium-type pha cluster. MaoC-like protein has an R-specific enoyl-CoA hydratase (R-hydratase) activity and is referred to as PhaJ when involved in PHA metabolism. In this study, the pha cluster of B. cereus YB-4 was characterized in terms of PhaJ’s function. In an in vitro assay, PhaJ from B. cereus YB-4 (PhaJYB4) exhibited hydration activity toward crotonyl-CoA. In an in vivo assay using Escherichia coli as a host for PHA accumulation, the recombinant strain expressing PhaJYB4 and PHA synthase led to increased PHA accumulation, suggesting that PhaJYB4 functioned as a monomer supplier. The monomer composition of the accumulated PHA reflected the substrate specificity of PhaJYB4, which appeared to prefer short chain-length substrates. The pha cluster from B. cereus YB-4 functioned to accumulate PHA in E. coli; however, it did not function when the phaJYB4 gene was deleted. The B. cereus-type pha cluster represents a new example of a pha cluster that contains the gene encoding PhaJ.

Developing Escherichia coli strains that are tolerant to acetate toxicity is important in light of an increased interest in the efficient utilization of lignocellulosic biomass feedstocks for the biosynthesis of value-added products. In this study, four strains known to produce polyhydroxyalkanoates (PHAs) from the typical hemicellulosic sugar xylose were tested for their tolerance to acetate. E. coli RSC10 was found to be tolerant of acetate, both in growth and fermentation studies. In the presence of acetate the strain showed a >2-fold increase in overall yields compared to using xylose alone as the feedstock. More importantly, the strain was found to be able to utilize acetate as a feedstock for biosynthesis of PHAs, with complete depletion of acetate (25 mM) at 9 h when acetate was the sole feedstock. Higher concentrations of acetate showed greater inhibition of fermentation than growth with a reduction of 90% in PHA yields at 100 mM. Additionally, the present work provides data to support the potential of acetate as a modulator for the control of composition of PHAs that incorporate lactate (LA) monomers into the copolymer from hemicellulose derived sugars.

Many microorganisms harbor genes necessary to synthesize biodegradable plastics known as polyhydroxyalkanoates (PHAs). We surveyed a genomic database and discovered a new cluster of class IV PHA synthase genes (phaRC). These genes are different in sequence and operon structure from any previously reported PHA synthase. The newly discovered PhaRC synthase was demonstrated to produce PHAs in recombinant Escherichia coli.

Polyhydroxyalkanoates (PHAs) are eco-friendly plastics that are synthesized by bacteria as an intracellular storage material. Ethanol is a key compound in molecular weight regulation of PHA during biosynthesis. In this study, three strains of Escherichia coli expressing PHA synthase (PhaCRe, PhaCAc or PhaCHe) were cultured to compare the effect of ethanol level on PHA molecular weight. As a result, it was revealed that PhaCAc tends to produce low molecular weight PHA due to its high ethanol sensitivity whereas PhaCHe produces high molecular weight PHA even in the presence of ethanol.

The choice of an appropriate microbial host cell and suitable production conditions is crucial for the downstream processing of pharmaceutical- and food-grade products. Although Escherichia coli serves as a highly valuable leading platform for the production of value-added products, like most Gram-negative bacteria, this bacterium contains a potent immunostimulatory lipopolysaccharide (LPS), referred to as an endotoxin. In contrast, Gram-positive bacteria, notably Bacillus, lactic acid bacteria (LAB), Corynebacterium, and yeasts have been extensively used as generally recognized as safe (GRAS) endotoxin-free platforms for the production of a variety of products. This review summarizes the currently available knowledge on the utilization of these representative Gram-positive bacteria for the production of eco- and bio-friendly products, particularly natural polyesters, polyhydroxyalkanoates, bacteriocins, and membrane proteins. The successful case studies presented here serve to inspire the use of these microorganisms as a main-player or by-player depending on their individual properties for the industrial production of these desirable targets.

The biofilm degradation of Aggregatibacter actinomycetemcomitans is essential as a complete periodontal disease therapy, and here we show the effects of potential probiotic bacteria such as Lactobacillus spp. for the biofilm of several serotypes of A. actinomycetemcomitans strains. Eight of the 13 species showed the competent biofilm degradation of ≥ 90% reduction in biofilm values in A. actinomycetemcomitans Y4 (serotype b) as well as four of the seven species for the biofilm of A. actinomycetemcomitans OMZ 534 (serotype e). In contrast, the probiotic bacteria did not have a big impact for the degradation of A. actinomycetemcomitans SUNY 75 (serotype a) biofilm. The dispersed A. actinomycetemcomitans Y4 cells through the biofilm detachment were still viable and plausible factors for the biofilm degradation were not due to the lactic acid and low pH conditions. The three enzymes, protease, lipase, and amylase may be responsible for the biofilm degradation; in particular, lipase was the most effective enzyme for the biofilm degradation of A. actinomycetemcomitans Y4 along with the protease activity which should be also important for the other serotypes. Remarkable lipase enzyme activities were detected from some of the potential probiotics and a supporting result using a lipase inhibitor presented corroborating evidence that lipase activity is one of the contributing factors for biofilm degradation outside of the protease which is also another possible factor for the biofilm of the other serotype of A. actinomycetemcomitans strains. On the other hand, the biofilm of A. actinomycetemcomitans SUNY 75 (serotype a) was not powerfully degraded by the lipase enzyme because the lipase inhibitor was slightly functional for only two of potential probiotics.