Collective Motion of Spherical Bacteria

A large variety of motile bacterial species exhibit collective motions while inhabiting liquids or colonizing surfaces. These collective motions are often characterized by coherent dynamic clusters, where hundreds of cells move in correlated whirls and jets. Previously, all species that were known to form such motion had a rod-shaped structure, which enhances the order through steric and hydrodynamic interactions. Here we show that the spherical motile bacteria Serratia marcescens exhibit robust collective dynamics and correlated coherent motion while grown in suspensions. As cells migrate to the upper surface of a drop, they form a monolayer, and move collectively in whirls and jets. At all concentrations, the distribution of the bacterial speed was approximately Rayleigh with an average that depends on concentration in a non-monotonic way. Other dynamical parameters such as vorticity and correlation functions are also analyzed and compared to rod-shaped bacteria from the same strain. Our results demonstrate that self-propelled spherical objects do form complex ordered collective motion. This opens a door for a new perspective on the role of cell aspect ratio and alignment of cells with regards to collective motion in nature.     

Organized Cell Swimming Motions in Bacillus subtilis Colonies: Patterns of Short-Lived Whirls and Jets

The swimming motions of cells within Bacillus subtilis colonies, as well as the associated fluid flows, were analyzed from video films produced during colony growth and expansion on wet agar surfaces. Individual cells in very wet dense populations moved at rates between 76 and 116 μm/s. Swimming cells were organized into patterns of whirls, each approximately 1,000 μm², and jets of about 95 by 12 μm. Whirls and jets were short-lived, lasting only about 0.25 s. Patterns within given areas constantly repeated with a periodicity of approximately 1 s. Whirls of a given direction became disorganized and then re-formed, usually into whirls moving in the opposite direction. Pattern elements were also organized with respect to one another in the colony. Neighboring whirls usually turned in opposite directions. This correlation decreased as a function of distance between whirls. Fluid flows associated with whirls and jets were measured by observing the movement of marker latex spheres added to colonies. The average velocity of markers traveling in whirls was 19 μm/s, whereas those traveling in jets moved at 27 μm/s. The paths followed by markers were aligned with the direction of cell motion, suggesting that cells create flows moving with them into whirls and along jets. When colonies became dry, swimming motions ceased except in regions close to the periphery and in isolated islands where cells traveled in slow whirls at about 4 μm/s. The addition of water resulted in immediate though transient rapid swimming (> 80 μm/s) in characteristic whirl and jet patterns. The rate of swimming decreased to 13 μm/s within 2 min, however, as the water diffused into the agar. Organized swimming patterns were nevertheless preserved throughout this period. These findings show that cell swimming in colonies is highly organized.     

Dynamics of Snake-like Swarming Behavior of Vibrio alginolyticus

Swarming represents a special case of bacterial behavior where motile bacteria migrate rapidly and collectively on surfaces. Swarming and swimming motility of bacteria has been studied well for rigid, self-propelled rods. In this study we report a strain of Vibrio alginolyticus, a species that exhibits similar collective motility but a fundamentally different cell morphology with highly flexible snake-like swarming cells. Investigating swarming dynamics requires high-resolution imaging of single cells with coverage over a large area: thousands of square microns. Researchers previously have employed various methods of motion analysis but largely for rod-like bacteria. We employ temporal variance analysis of a short time-lapse microscopic image series to capture the motion dynamics of swarming Vibrio alginolyticus at cellular resolution over hundreds of microns. Temporal variance is a simple and broadly applicable method for analyzing bacterial swarming behavior in two and three dimensions with both high-resolution and wide-spatial coverage. This study provides detailed insights into the swarming architecture and dynamics of Vibrio alginolyticus isolate B522 on carrageenan agar that may lay the foundation for swarming studies of snake-like, nonrod-shaped motile cell types.     

Initiation of Swarming Motility by Proteus mirabilis Occurs in Response to Specific Cues Present in Urine and Requires Excess l-Glutamine

Proteus mirabilis, a leading cause of catheter-associated urinary tract infection (CaUTI), differentiates into swarm cells that migrate across catheter surfaces and medium solidified with 1.5% agar. While many genes and nutrient requirements involved in the swarming process have been identified, few studies have addressed the signals that promote initiation of swarming following initial contact with a surface. In this study, we show that P. mirabilis CaUTI isolates initiate swarming in response to specific nutrients and environmental cues. Thirty-three compounds, including amino acids, polyamines, fatty acids, and tricarboxylic acid (TCA) cycle intermediates, were tested for the ability to promote swarming when added to normally nonpermissive media. l-Arginine, l-glutamine, dl-histidine, malate, and dl-ornithine promoted swarming on several types of media without enhancing swimming motility or growth rate. Testing of isogenic mutants revealed that swarming in response to the cues required putrescine biosynthesis and pathways involved in amino acid metabolism. Furthermore, excess glutamine was found to be a strict requirement for swarming on normal swarm agar in addition to being a swarming cue under normally nonpermissive conditions. We thus conclude that initiation of swarming occurs in response to specific cues and that manipulating concentrations of key nutrient cues can signal whether or not a particular environment is permissive for swarming.     

Hybridization Hotspots at Bat Swarming Sites

During late summer and early autumn in temperate zones of the Northern Hemisphere, thousands of bats gather at caves, mainly for the purpose of mating. We demonstrated that this swarming behavior most probably leads not only to breeding among bats of the same species but also interbreeding between different species. Using 14 nuclear microsatellites and three different methods (the Bayesian assignment approaches of STRUCTURE and NEWHYBRIDS and a principal coordinate analysis of pairwise genetic distances), we analyzed 375 individuals belonging to three species of whiskered bats (genus Myotis) at swarming sites across their sympatric range in southern Poland. The overall hybridization rate varied from 3.2 to 7.2%. At the species level, depending on the method used, these values ranged from 2.1–4.6% in M. mystacinus and 3.0–3.7% in M. brandtii to 6.5–30.4% in M. alcathoe. Hybrids occurred in about half of the caves we studied. In all three species, the sex ratio of hybrids was biased towards males but the observed differences did not differ statistically from those noted at the population level. In our opinion, factors leading to the formation of these admixed individuals and their relatively high frequency are: i) swarming behaviour at swarming sites, where high numbers of bats belonging to several species meet; ii) male-biased sex ratio during the swarming period; iii) the fact that all these bats are generally polygynous. The highly different population sizes of different species at swarming sites may also play some role. Swarming sites may represent unique hybrid hotspots, which, as there are at least 2,000 caves in the Polish Carpathians alone, may occur on a massive scale not previously observed for any group of mammal species in the wild. Evidently, these sites should be treated as focal points for the conservation of biodiversity and evolutionary processes.     

Genetic control of morphogenesis in the multicellular alga Ulva mutabilis

Distinct ultrastructural differences between the cell walls of the mutant lumpy (lu) and wild-type are demonstrated. The polyglucan fibrils in the wild-type wall appear straight and densely packed compared to the more uneven and loosely woven fibrils in the mutant wall. The mutant grows as an aggregate of loosely connected cells. The three cell types seen in the wild type do not differentiate, and consequently no blade and holdfast forms. It is suggested that the defect in the cell wall production takes away a necessary condition for normal morphogenesis. The lumpy mutant shows epistasis over two other chromosomal mutants, bu and Sl, described earlier to be morphogenic.     

Difference in antifungal activity of morphotypes of clavicipitaceous endophytes within and between species

In a previous study, a total of 484 endophytic fungi were isolated and purified from seven populations of Achnatherum sibiricum (L.) Keng collected at six geographical locations in Inner Mongolia, China. Based on growth rates as well as morphological characteristics, the isolates were classified into five morphotypes. Among them, morphotypes A, B and C were ascribed to the same species, Neotyphodium chisosum, based on ITS sequences. Morphotype E was identified as Epichloë amarillans. In the present study, four morphotypes, A, B, C and E, belonging to two species, were chosen for an in vitro pathogen trial. The results showed that both endophyte colonies and endophyte filtrate of all morphotypes could inhibit the mycelia growth and spore germination of the pathogen fungi tested. The magnitude of inhibition varied not only between species, but also among morphotypes of the same species. Overall, the antifungal ability of E. amarillans (morphotype E) was higher than that of N. chisosum. Within N. chisosum, the antifungal ability was highest in morphotype C, followed by morphotype A, and lowest in morphotype B. This variability suggests that different morphotypes might represent different genotypes of endophyte. The effect of endophyte infection on the host grass should be examined not only on the species level but also on the morphotype level to determine the possible interactions.     

Population Genetic Structure Within and among Seasonal Site Types in the Little Brown Bat (Myotis lucifugus) and the Northern Long-Eared Bat (M. septentrionalis)

During late summer and early autumn, temperate bats migrate from their summering sites to swarming sites, where mating likely occurs. However, the extent to which individuals of a single summering site migrate to the same swarming site, and vice versa, is not known. We examined the migratory connectivity between summering and swarming sites in two temperate, North American, bat species, the little brown bat (Myotis lucifugus) and the northern long-eared bat (Myotis septentrionalis). Using mitochondrial and microsatellite DNA markers, we examined population structuring within and among summering and swarming sites. Both species exhibited moderate degrees of mitochondrial DNA differentiation (little brown bat: F_ST(SWARMING)= 0.093, F_ST(SWARMING)= 0.052; northern long-eared bat: F_ST(SWARMING)= 0.117, F_ST(SWARMING)= 0.043) and little microsatellite DNA differentiation among summering and among swarming sites. Haplotype diversity was significantly higher at swarming sites than summering sites, supporting the idea that swarming sites are comprised of individuals from various summering sites. Further, pairwise analyses suggest that swarming sites are not necessarily comprised of only individuals from the most proximal summering colonies.     

Apophyses,verticiles, and denticles in skeletons of Late Paleozoic radiolarians of the subfamily Entactiniinae

Morphostructural analysis of secondary skeletal elements, such as apophyses on the main spines and verticiles on the secondary spines of certain Late Paleozoic radiolarians of the subfamily Entactiniinae allow their taxonomic rank to be raised to genus. The presence of denticles on the secondary spines is only evidence of particular species. Three skeleton morphotypes are established: primary morphotype A, verticillate morphotype B, and apophyseal morphotype C. A new species, Entactinia mariannae sp. nov., and two new genera, Apophysiacus gen. nov. with five species, A. ichikawai (Caridroit et De Wever, 1984), A. minuta (Feng, 2007), A. praepycnoclada (Nazarov et Ormiston, 1983), A. pycnoclada (Nazarov et Ormiston, 1985), and A. sakmaraensis (Kozur et Mostler, 1989), and Verticillides gen. nov. with three species, V. nazarovi nom. nov., V. nestellae sp. nov., and V. verticillatus sp. nov., are described.     

Changes in the organization of the outer membrane of Proteus mirabilis during swarming: freeze-fracture structure and membrane fluidity analysis.

Freeze-fracture studies of short, nonswarming Proteus mirabilis revealed the characteristic gram-negative profile of fractured inner membrane with densely packed particles and sectioned outer membrane with little or no fracture plane. Long swarming cells, however, fractured easily along both the inner membrane and a second membrane, probably the outer membrane. The inner membrane had a typical profile, whereas the outer membrane had fewer but more prominent particles. Isolation and purification of the inner and outer membranes of the short and long bacteria and examination of them with electron paramagnetic resonance measurements after spinlabeling supported the above observations. The outer membrane of swarmer cells allowed higher mobility of the spin label than did the outer membrane of the nonswarming short cells, which showed a typical rigid profile. These results suggest that regions of lipid bilayer appear in the outer membrane during swarmer formation. Previous observation of the behavior and biochemistry of P. mirabilis during swarming are discussed in light of these results.     

Inhibition of Quorum Sensing in Serratia marcescens AS-1 by Synthetic Analogs of N-Acylhomoserine Lactone

Quorum sensing is a regulatory system for controlling gene expression in response to increasing cell density. N-Acylhomoserine lactone (AHL) is produced by gram-negative bacteria, which use it as a quorum-sensing signal molecule. Serratia marcescens is a gram-negative opportunistic pathogen which is responsible for an increasing number of serious nosocomial infections. S. marcescens AS-1 produces N-hexanoyl homoserine lactone (C₆-HSL) and N-(3-oxohexanoyl) homoserine lactone and regulates prodigiosin production, swarming motility, and biofilm formation by AHL-mediated quorum sensing. We synthesized a series of N-acyl cyclopentylamides with acyl chain lengths ranging from 4 to 12 and estimated their inhibitory effects on prodigiosin production in AS-1. One of these molecules, N-nonanoyl-cyclopentylamide (C₉-CPA), had a strong inhibitory effect on prodigiosin production. C₉-CPA also inhibited the swarming motility and biofilm formation of AS-1. A competition assay revealed that C₉-CPA was able to inhibit quorum sensing at four times the concentration of exogenous C₆-HSL and was more effective than the previously reported halogenated furanone. Our results demonstrated that C₉-CPA was an effective quorum-sensing inhibitor for S. marcescens AS-1.     

Spirotrichonymphea (Parabasalia) symbionts of the termite Paraneotermes simplicicornis

The desert dampwood termite Paraneotermes simplicicornis harbors several species of obligately symbiotic protists that support its nutrition by fermenting lignocellulose. Among them are three morphotypes with the dexiotropic spiraling flagellar bands characteristic of Spirotrichonymphea (Parabasalia). The largest morphotype, characterized by an elongated cell apex with axial columella and internally positioned spiraling flagellar bands, was previously described as Spirotrichonympha polygyra. A smaller morphotype, with similarly internalized flagellar bands but a more rounded posterior without a protruding axostyle, was previously reported but not named. The smallest morphotype has surface flagellar bands and can attach to other protist cells by its apex. In this study, we combine light microscopy of live specimens and 18S rRNA gene sequencing of individually isolated cells to better understand the diversity of symbionts in P. simplicicornis. We found that S. polygyra branches distantly from true Spirotrichonympha, which are associated with Reticulitermes termites. Thus, we propose the new genus Cuppa to accommodate C. polygyra n. comb. (type species) and the similar but smaller morphotype Cuppa taenia n. sp. The undescribed smallest morphotype can be excluded from all previously described Spirotrichonymphea genera by molecular and behavioral evidence, so we propose Fraterculus simplicicornis n. gen., n. sp., to accommodate this organism.     

Effects of Metal Phytoextraction Practices on the Indigenous Community of Arbuscular Mycorrhizal Fungi at a Metal-Contaminated Landfill

Phytoextraction involves use of plants to remove toxic metals from soil. We examined the effects of phytoextraction practices with three plant species (Silene vulgaris, Thlaspi caerulescens, and Zea mays) and a factorial variation of soil amendments (either an ammonium or nitrate source of nitrogen and the presence or absence of an elemental sulfur supplement) on arbuscular mycorrhizal (AM) fungi (Glomales, Zygomycetes) at a moderately metal-contaminated landfill located in St. Paul, Minn. Specifically, we tested whether the applied treatments affected the density of glomalean spores and AM root colonization in maize. Glomalean fungi from the landfill were grouped into two morphotypes characterized by either light-colored spores (LCS) or dark-colored spores (DCS). Dominant species of the LCS morphotype were Glomus mosseae and an unidentified Glomus sp., whereas the DCS morphotype was dominated by Glomus constrictum. The density of spores of the LCS morphotype from the phytoremediated area was lower than the density of these spores in the untreated landfill soil. Within the experimental area, spore density of the LCS morphotype in the rhizosphere of mycorrhizal maize was significantly higher than in rhizospheres of nonmycorrhizal S. vulgaris or T. caerulescens. Sulfur supplement increased vesicular root colonization in maize and exerted a negative effect on spore density in maize rhizosphere. We conclude that phytoextraction practices, e.g., the choice of plant species and soil amendments, may have a great impact on the quantity and species composition of glomalean propagules as well as on mycorrhiza functioning during long-term metal-remediation treatments.     

Sub-Decadal Resolution in Sediments of Late Miocene Lake Pannon Reveals Speciation of Cyprideis (Crustacea,Ostracoda)

Late Miocene "Lake Pannon" (~11.3 Ma) was a remnant of the Central Paratethyan Sea. Successive freshening and constantly changing environmental conditions, like oxygenation, nutrition and substrate led to a well-documented radiation in molluscs and ostracods. Among ostracods (small crustaceans), Cyprideis is one of the most common genera in "Lake Pannon", as well as in several other ancient lakes, showing numerous adaptations and speciations. Here, we present high-resolution data from an early transgression of "Lake Pannon" in the Eastern Styrian Basin (SE Austria). Mataschen clay pit is in the focus of geologic and paleontologic research since 20 years and its geologic and paleoecologic evolution is well-documented. We drilled five cores covering a ~2.3 m long section and completely sampled it in 5-mm thick intervals to reconstruct minute changes in the ostracod fauna over a transgression of a brackish water body. The dominant genus, Cyprideis, is represented by three species C. mataschensis, C. kapfensteinensis and C. ex gr. pannonica. Through morphometric analyses we highlight the variance of each taxon and suggest that there is no direct ecologic control on size or shape. Furthermore, we found a second, co-occurring morphotype of C. kapfensteinensis which is directly related to an elevation of salinities above 13 psu. The presence of two intermediate specimens between the two morphotypes in the sample directly below the first appearance of C. kapfensteinensis B leads us to the conclusion that we are facing a speciation event leading to four sympatric species of Cyprideis.     

Caenorhabditis briggsae and C. elegans: partial characterization of cuticle surface carbohydrates.

The presence of galactose, glucose, mannose, and N-acetylglucosamine on the exposed surface of the nematodes Caenorhabditis briggsae and C. elegans was indicated by specific binding of three iodinated plant lectins. Proteolysis experiments suggested the absence of digestible glycoproteins on the exposed surfaces of the two nematode species. High resolution micrographs of cuticle surface preparations labeled with cationized ferritin indicated that the negative charge-bearing molecules are more densely packed on the nematode surface than on animal plasma membranes.     

Fine‐scale spatial distribution of murundus structures in the semi‐arid region of Brazil

Murundus are earth mounds widespread in most landscapes in the semi‐arid region of Brazil. Evidence obtained from predictive modelling has suggested a termite origin for these structures, opening up new opportunities for further research. Distribution of densely packed murundus at larger spatial scales is most related to climatic regime and soil nutrient availability. However, factors and processes underlying their distribution and density at smaller spatial scales are not yet fully understood. In this study, we adopted an approach based on mapping point data using high‐resolution satellite imagery, multi‐scale second‐order analysis and general linear models to examine the fine‐scale spatial distribution and density of murundus. Our results suggest that the distribution of those structures within densely packed areas is regulated by more than one process acting or interacting across multiple spatial scales. All densely packed murundus showed a significant regular distribution at the distance scale of up to 50 m radially and a completely random distribution across all other upper distance scales. We interpret the regular pattern as a result of competition for foraging territories between different termite colonies during the process formation of densely packed murundus. The random pattern at larger distance scales (above 50 m radially) can be attributed to habitat selection preferences by termite species builders of murundus mediated by local environmental resources and conditions (i.e. availability of food resources and nesting and open habitat), which would be randomly distributed in space. Thus, at finer spatial scales murundus distributions are associated with biotic interactions acting on an abiotic template. On the basis of significant linear correlations, we suggest that the density of murundus is strongly related to local temperature regime with soil‐type influencing its effect on the murundus densities. Our findings provide novel evidences that mound‐building termites are involved in the formation of murundus in the semi‐arid region of Brazil.     

Alishewanella solinquinati sp. nov., Isolated from Soil Contaminated with Textile Dyes

A novel Gram-negative, motile, rod-shaped, facultative anaerobic bacterial strain, KMK6^T, was isolated from soil contaminated with textile dyes from an industrial estate located at Ichalkaranji, Maharashtra, India, and its taxonomical position was established by using a polyphasic approach. The major cellular fatty acids included C_17:1ω8c, summed feature 3 (C_16:1ω7c and/or iso-C_15:0 2-OH), C_17:0, C_16:0, and C_18:1ω7c. The DNA G+C content of strain KMK6^T was 48.8 mol %. 16S rRNA gene sequence analysis confirmed its placement in the genus Alishewanella, and exhibited sequence similarity levels of below 97 % to the type strains of validly published Alishewanella species. On the basis of genotypic and phenotypic evidence, strains KMK6^T is considered to be a novel species of the genus Alishewanella, for which we propose that strain KMK6^T (=NCIM 5295^T =BCRC 17848^T) is assigned to a novel species, Alishewanella solinquinati sp. nov.     

Microstructure of Colonies of Rod-Shaped Bacteria

Whole colonies of Bacillus cereus, B. megaterium, B. mycoides CN2495, Corynebacterium hofmanni NCTC1938, Escherichia coli, Lactobacillus acidophilus NCIB1899, Nocardia graminis NCTC4728, Pseudomonas viscosa, and Serratia marcescens were prepared for scanning electron microscopic examination by freeze-drying and metal-coating. The arrangement of individual cells within colonies could be seen. Cells of Bacillus colonies tended to be longer than in liquid culture and irregular in shape and to give the appearance of branching. B. megaterium colonies frequently had a dense covering film. Colonies of gram-negative bacteria consisted of fairly short rods covered by much adherent extracellular material. L. acidophilus had colonies comprised of densely packed, well-oriented rods. C. hofmanni colonies contained coccobacilli, packed together. Correlations were observed between plano-convex colony form and densely packed cells, rough colony form and random arrangement of well-separated microorganisms, and irregular colony edge and tendency of cells to grow out from the colony in filaments.     

Hydrolases on silica surfaces: Coverage-activity–molecular property relationships revealed

A set of recommendations to maintain high activity of immobilized enzymes is developed based on direct observation via AFM. This helps to close knowledge gaps that often lead to poor performance of nanobiocatalysts for chemical synthesis. Molecule-level height and volume distribution analyses from high-resolution AFM images were applied to Candida antarctica Lipase B (CALB), subtilisin Carlsberg, and the Lipase from Thermomyces lanuginosus (TLL) deposited on model silica surfaces. Ensembles of flexible or “soft” enzymes appear separated when interactions with the surface are considerable at low surface coverage but form highly entangled structures of increased conformational stability at high surface coverage. By contrast, ensembles of rigid or “hard” enzymes appear to maintain stable aggregates even under strong interaction with the surface. The more rigid the enzyme the higher its tendency to remain in a densely packed state that is able to withstand surface-induced conformational transitions detrimental to catalysis. Weakening of surface-protein interactions for “soft” enzymes will prevent single-molecule immobilization, which reduces catalytic competency through structural changes. Multi-layer coverage in enzyme immobilization should generally be avoided due to mass transfer limitations.     

Interaction between bacteriophage DMS3 and host CRISPR region inhibits group behaviors of Pseudomonas aeruginosa

Bacteriophage infection has profound effects on bacterial biology. Clustered regular interspaced short palindromic repeats (CRISPRs) and cas (CRISPR-associated) genes are found in most archaea and many bacteria and have been reported to play a role in resistance to bacteriophage infection. We observed that lysogenic infection of Pseudomonas aeruginosa PA14 with bacteriophage DMS3 inhibits biofilm formation and swarming motility, both important bacterial group behaviors. This inhibition requires the CRISPR region in the host. Mutation or deletion of five of the six cas genes and one of the two CRISPRs in this region restored biofilm formation and swarming to DMS3 lysogenized strains. Our observations suggest a role for CRISPR regions in modifying the effects of lysogeny on P. aeruginosa.