Nonseptate Filaments Research Articles

Filament formation of Salmonella Paratyphi A accompanied by FtsZ assembly impairment and low level ppGpp.

Previously, we reported that Salmonella enterica serovar Paratyphi A strain S602 grew into multinuclear, nonseptate, and nonlethal filaments on agar plates containing nitrogenous salts. Strain S602 was more sensitive to osmotic and oxidative stress than the reference strain 3P243 of nonfilamentous Salmonella Paratyphi A. Strain S602 had an amber mutation (C154T) in rpoS. The revertant of this mutant, SR603, was repressed to form filaments under conditions with abundant nitrogenous salts. However, 3PR244, an rpoS mutant of 3P243 (C154T), did not form filaments, which implies that the rpoS mutation is not the sole cause of filamentation in strain S602. Next, we examined whether the level of guanosine 5'-diphosphate 3'-diphosphate (ppGpp) in S602 strain is involved in filament formation. The intracellular ppGpp level in filamentous cells was lower than that in nonfilamentous cells. Furthermore, cells belonging to strain RE606, a derivative of S602 where the intracellular concentration of ppGpp was increased by overexpression of the relA gene, exhibited normal Z-ring formation and cell division. In the S602 strain, the decrease in the ppGpp level induced by the presence of nitrogenous salt and the rpoS mutation led to the inhibition of Z-ring formation and the subsequent filamentation of cells.

Rapid Mass Movement of Chloroplasts during Segment Formation of the Calcifying Siphonalean Green Alga, Halimeda macroloba

BackgroundThe calcifying siphonalean green alga, Halimeda macroloba is abundant on coral reefs and is important in the production of calcium carbonate sediments. The process by which new green segments are formed over-night is revealed here for the first time.Methodology/Principal FindingsGrowth of new segments was visualised by epifluorescence and confocal microscopy and by pulse amplitude modulation (PAM) fluorimetry. Apical colourless proto-segments were initiated on day 1, and formed a loose network of non-calcified, non-septate filaments, containing no chloroplasts. Rapid greening was initiated at dusk by i) the mass movement of chloroplasts into these filaments from the parent segment and ii) the growth of new filaments containing chloroplasts. Greening was usually complete in 3–5 h and certainly before dawn on day 2 when the first signs of calcification were apparent. Mass chloroplast movement took place at a rate of ∼0.65 µm/s. Photosynthetic yield and rate remained low for a period of 1 to several hours, indicating that the chloroplasts were made de novo. Use of the inhibitors colchicine and cytochalasin d indicated that the movement process is dependent on both microtubules and microfilaments.SignificanceThis unusual process involves the mass movement of chloroplasts at a high rate into new segments during the night and rapid calcification on the following day and may be an adaptation to minimise the impact of herbivorous activity.

Early stages in development of the Escherichia coli cell-division site.

Development of the Escherichia coli cell division site was studied in wild-type cells and in non-septate filaments of ftsZnull and ftsZTs mutant cells. Localized regions of plasmolysis were used as markers for the positions of annular structures that are thought to be related to the periseptal annuli that flank the ingrowing septum during cytokinesis. The results show that these structures are localized at potential division sites in non-septate filaments of FtsZ- cells, contrary to previous reports. The positions of the structures along the long axis of the cells in both wild-type cells and FtsZ- filaments were unaffected by the presence of plasmolysis bays at the cell poles. These results do not agree with a previous suggestion that the apparent association of plasmolysis bays with future division sites was artefactual. They support the view that division sites begin to differentiate before the initiation of septal ingrowth and that plasmolysis bays and the annular attachments that define them, mark the locations of these early events in the division process.

Development of the cell‐division site in FtsA‐filaments

Early changes at cell-division sites were studied in non-septate filaments induced by growth of ftsATs mutant cells under non-permissive conditions. The positions of localized regions of plasmolysis were used as markers for the locations of partial and complete annular structures that are thought to be precursors of the periseptal annuli that flank the septum during cytokinesis. The results confirmed that these structures were localized at potential division sites and suggested a model in which older division sites play a role in the generation of new sites for future use, with each older site being used only once for this purpose. The results also suggest that the details of division-site development can profitably be studied in cells in which early events in the differentiation process are uncoupled from the septation event.

FtsL, an Essential Cytoplasmic Membrane Protein Involved in Cell Division in Escherichia coli

We have identified a gene involved in bacterial cell division, located immediately upstream of the ftsI gene in the min 2 region of the Escherichia coli chromosome. This gene, which we named ftsL , was detected through characterization of Tn phoA insertions in a plasmid containing this chromosomal region. Tn phoA topological analysis and fractionation of alkaline phosphatase fusion proteins indicated that the ftsL gene product is a 13.6-kDa cytoplasmic membrane protein with a cytoplasmic amino terminus, a single membrane-spanning segment, and a periplasmic carboxy terminus. The ftsL gene is essential for cell growth and division. A null mutation in ftsL resulted in inhibition of cell division, formation of long, nonseptate filaments, ultimate cessation of growth, and lysis. Under certain growth conditions, depletion of FtsL or expression of the largest ftsL-phoA fusion produced a variety of cell morphologies, including Y-shaped bacteria, indicating a possible general weakening of the cell wall. The FtsL protein is estimated to be present at about 20 to 40 copies per cell. The periplasmic domain of the protein displays a sequence with features characteristic of leucine zippers, which are involved in protein dimerization.

Chemoautotrophic, sulfur-oxidizing symbiotic bacteria on marine nematodes: Morphological and biochemical characterization

The marine, free-living Stilbonematinae (Nematoda: Desmodorida) inhabit the oxygen sulfide chemocline in marine sands. They are characterized by an association with ectosymbiotic bacteria. According to their ultrastructure the bacteria are Gram-negative and form morphologically uniform coats that cover the entire body surface of the worms. They are arranged in host-genus or host-species specific patterns: cocci form multilayered sheaths, rods, and crescent- or filament-shaped bacteria form monolayers. The detection of enzymes associated with sulfur metabolism and of ribulose-1,5 bisphosphate carboxylase oxygenase, as well as elemental sulfur in the bacteria indicate a chemolithoautotrophic nature of the symbionts. Their reproductive patterns appear to optimize space utilization on the host surface: vertically standing rods divide by longitudinal fission, whereas other bacteria form non-septate filaments of up to 100 μm length.

FtsL, an Essential Cytoplasmic Membrane Protein Involved in Cell Division in Escherichia coli

We have identified a gene involved in bacterial cell division, located immediately upstream of the ftsI gene in the min 2 region of the Escherichia coli chromosome. This gene, which we named ftsL, was detected through characterization of TnphoA insertions in a plasmid containing this chromosomal region. TnphoA topological analysis and fractionation of alkaline phosphatase fusion proteins indicated that the ftsL gene product is a 13.6-kDa cytoplasmic membrane protein with a cytoplasmic amino terminus, a single membrane-spanning segment, and a periplasmic carboxy terminus. The ftsL gene is essential for cell growth and division. A null mutation in ftsL resulted in inhibition of cell division, formation of long, nonseptate filaments, ultimate cessation of growth, and lysis. Under certain growth conditions, depletion of FtsL or expression of the largest ftsL-phoA fusion produced a variety of cell morphologies, including Y-shaped bacteria, indicating a possible general weakening of the cell wall. The FtsL protein is estimated to be present at about 20 to 40 copies per cell. The periplasmic domain of the protein displays a sequence with features characteristic of leucine zippers, which are involved in protein dimerization.

High-level expression of the FtsA protein inhibits cell septation in Escherichia coli K-12

DNA fragments encoding the ftsA gene were subcloned into plasmids downstream of a lac promoter or a tac promoter. These plasmid constructs, when transformed into wild-type and mutant strains, inhibited normal cell septation, causing the formation of long nonseptate filaments. This phenotype is due to overproduction of the FtsA protein.

Latest proterozoic microfossils from the nama group, namibia (south west Africa)

Gray to black shales from the Kuibis and Schwarzrand subgroups of the Nama Group, South West Africa/Namibia, contain a distinctive, if depauperate, assemblage of organic-walled microfossils. The assemblage is dominated by leiosphaerid acritarchs and fragments of the ribbon-like macrofossil Vendotaenia. Small unicells and non-septate filaments interpreted as the sheaths of oscillatorian cyanobacteria occur with variable frequency throughout the subgroups, while Chuaria circularis, Bavlinella faveolata, and a Comasphaeridium-like form (known from a single specimen) are found as rarer components of the biota. Assemblage composition shows no strong stratigraphic trends within the subgroups. The Nama assemblage compares closely with biotas previously described from uppermost Proterozoic (Valdaian) sequences of the Baltic region, Scandinavia, and Australia, and confirms a latest Proterozoic age for the Kuibis and Schwarzrand subgroups and their contained metazoan fossils. Although its significance is unclear, Nama microfossils tend to be strongly corroded. This diagenetic feature is widespread among Valdaian assemblages, but appears to be much less characteristic of older or younger biotas.

Swarmer cell differentiation of Proteus mirabilis in fluid media.

After 3-4 h in a rich fluid medium such as brain--heart infusion broth, motile nonseptate filaments developed from normal short rods and formed about 80% of the cell mass of Proteus mirabilis PM23. This developmental pattern was not observed in any of the other nine representatives of the species. These filaments were considered to be equivalent to swarmer cells formed on agar media because these cells ceased tumbling (i.e., chemotaxis was repressed), they developed large numbers of flagella (i.e., flagella synthesis and insertion was derepressed), and the distribution of nuclei in the filaments indicated that there was normal segregation. The population of cells grown in a minimal medium supplemented with amino acids and nicotinic acid consisted only of short cells with tumbling motility, despite the production of long cells and swarming on the same medium solidified with ordinary agar (refined agar was not effective). These short cells differentiated in 1-1.5 h in brain--heart infusion broth at 37 degrees C after an initial division. The requirements for initiation of differentiation were good basal nutrition, suitable cations (probably Ca2+ and Na+, or K+), and unknown heat-stable organic factors (molecular weight less than 10 000) present in crude agar and yeast extract. Other components of media promoted swarmer differentiation if it was initiated and these included organic acids (lactate), amino acids (proline or serine), phosphate, and an appropriate ionic environment. Comparison of the observed sequence of length classes in brain--heart infusion broth culture with computer generated growth models suggested that, at the outset of growth, 50% of the products of each short cell division ceased septation but grew in length for about five doubling periods and then divided cells from each end at a faster rate (3-5 times per hour) for return to the short cell pool.

Study on the cell cycle of Bacillus subtilis using temperature-sensitive mutants

Among temperature-sensitive mutants which were defective in septum formation and formed nonseptate filaments at nonpermissive temperatures three (ts31, ts341, ts526) were identified among 434 temperature-sensitive mutants isolated at random from a mutagenized population of Bacillus subtilis 168. The results of morphological observations and characterization of these mutants showed that ts31 and ts341 were septum-initiation mutants and that ts526 was a DNA elongation mutant. The above mutations, and other mutations affecting septum initiation (div355 and tms12) were mapped by PBS1-mediated transduction on the chromosome in three separate regions as follows: pur A16-ts526-div355-cysA14; metC3-(ts31, tms12)-pyrD1-recA1; ebr-2-ts341-uvrA1-hisA1-cysB3. Our results suggest that the initiation process of septum formation requires at least four kinds of gene product. In addition, the sesult obtained with ts526 suggests an intimate connection between septum initiation and DNA replication.

Suppression of Bacterial Adherence by Subminimal Inhibitory Concentrations of β-Lactam and Aminoglycoside Antibiotics

Nonseptate filaments of Escherichia coli obtained by growth of a temperature-sensitive mutant of the organism at its restrictive temperature in the absence of antibiotics or at its permissive temperature in the presence of approximately one-half the MIC of penicillin (27 micrograms/ml; MIC of penicillin, 50 micrograms/ml) lacked the ability both to bind to mannose and to adhere to host tissues. Addition of low concentrations (0.5-10 micrograms/ml) of streptomycin to cultures of E. coli resulted in marked suppression of the mannose-binding and adhering ability of streptomycin-sensitive E. coli (MIC, 30 micrograms/ml). In contrast, up to 5,000 microgram of streptomycin/ml had no effect on an isogenic streptomycin-resistant mutant of E. coli (MIC, 20,000 micrograms/ml). No concentration of penicillin or streptomycin that was tested was able to suppress either the mannose-binding or the adhering ability of E. coli once those activities had been acquired by the organism. These results suggest that subminimal inhibitory concentrations of antibiotics suppress the ability of bacteria to adhere to cells.

Peptidoglycan biosynthesis in a thermosensitive division mutant of Escherichia coli.

Peptidoglycan biosynthesis during a bacterial division cycle was investigated in the thermosensitive division mutant Escherichia coli PAT 84 Synchronous cell division of this organism was initiated by a shift down from restrictive to permissive growth temperature. Cells harvested at different times after a shift down of temperature served as representatives of the various stages during cell division. These cells were made permeable to peptidoglycan nucleotide-sugar precursors by pretreatment with ether and were found capable of catalyzing the polymerization of externally added precursors as well as the covalent attachment of the newly synthesized peptidoglycan strands to those preexisting in the cell wall. Differences were observed in the rats of peptidoglycan synthesis and in the extent of peptide side-chain corss-linkage at the various stages of division. Nonseptate filaments, formed at the restrictive temperature, incorporated significantly more peptidoglycan which was more cross-linked than in normally dividing cells grown at the permissive temperature. Quantitative analyses of the carboxypeptidase and transpeptidase reactions in cells at different stages of division were performed and the inhibitory effect of a number of beta-lactam antibiotics was investigated. Of special significance was the finding that low doses of penicillin or growth at restrictive temperature, which did not affect transpeptidation, partially inhibited the carboxypeptidase activity. This inhibition was paralleled by an increase in incorporation of newly synthesized peptidoglycan into the preexisting cell wall. We therefore propose that carboxypeptidase activity regulates the availability of peptidoglycan precursor(s) for attachment to the preexisting peptidoglycan by transpepidation.

Strain-specific difference that affects the inhibition of division of Escherichia coli filaments by chloramphenicol.

The Escherichia coli mutations ts1882 and ts2158 cause temperature-sensitive septum formation and result in growth of cells as long, multinucleate, nonseptate filaments at 42 C. When filaments are transferred to 28 C, they divide into short cells. Chloramphenicol, when added to cultures of filaments at the time of temperature reduction. Inhibited division of filaments when these temperature-sensitive mutations were present in the K-12 strain AB1157. However, when the ts1882 and ts2158 mutations were present in another k-12 strain, UTH4113, filaments of these strains divided in the presence of chloramphenicol.

The morphology and reproduction of Derbesia clavaeformis (J. Agardh) De Toni (Chlorophyta)

Abstract Derbesia clavaeformis grows in heavily-shaded pools on rough-water coasts, from Kangaroo Island in South Australia to Cape Patterson in Victoria. Field and culture studies show that it reproduces by multiflagellate zoospores which produce a prostrate pad, developing into a disc-like attachment stage characterized by an internal skeletal structure of calcium carbonate, deposited as aragonite. From this attachment stage, or directly from floating zoospores, fine, much-branched, non-septate filaments develop and later produce the adult, clavate, branches which bear sporangia. The life-history is thus direct and sporophytic, and has been followed in culture through successive generations. In the field, the adult thalli with clavate branches occur in summer, and the discoid attachment stage in winter. D. clavaeformis is retained within the Derbesiales, but recognized as a distinct genus, Pedobesia gen. nov., characterized by its direct life history and distinctive, calcified, attachment stage.

Isolation and Characterization of an Escherichia coli ruv Mutant Which Forms Nonseptate Filaments After Low Doses of Ultraviolet Light Irradiation

Two ultraviolet light (UV)-sensitive mutants have been isolated from Escherichia coli K-12. These mutants, designated RuvA(-) and RuvB(-), were controlled by a gene located close to the his gene on the chromosome map. They were sensitive to UV (10- to 20-fold increase) and slightly sensitive to gamma rays (3-fold increase). Host cell reactivation, UV reactivation and genetic recombination were normal in these mutants. Irradiation of the mutants with UV resulted in the production of single-strand breaks in deoxyribonucleic acid, which was repaired upon incubation in a growth medium. After UV irradiation, these mutants resumed deoxyribonucleic acid synthesis at a normal rate, as did the parent wild-type bacteria, and formed nonseptate, multinucleate filaments. From these results we concluded that the mutants have some defect in cell division after low doses of UV irradiation, similar to the lon(-) or fil(+) mutant of E. coli. The ruv locus was divided further into ruvA and ruvB with respect to nalidixic acid sensitivity and the effect of minimal agar or pantoyl lactone on survival of the UV-irradiated cell. The ruvB(-)mutant was more sensitive to nalidixic acid than were ruvA(-) and the parent strain. There was a great increase in the surviving fraction of the UV-irradiated ruvB(-) mutant when it was plated on minimal agar or L agar containing pantoyl lactone. No such increase in survival was observed in the ruvA(-) mutant.

Studies of septation and separation in a mutant ofSalmonella typhimurium

Salmonella typhimurium strain 4a is a temperature-sensitive division mutant. At 39° or 42° in several media a septation lesion occurs since non-septate filaments form at these temperatures. At lower temperatures in some media separation is aberrant since long chains of cells are formed. Since these division abnormalities are corrected either by revertion to ts+ or phenotypically by growth with sucrose, it is likely that both are the result of one lesion and that the component (S) which is temperature-sensitive in strain 4a is involved in both septation and separation. At 42° S is irreversibly inactivated but at 39° inactivation appears to be reversible since division restarts rapidly when 39° filaments are returned to 25° and protein synthesis is not required for such division. Comparison of recovery times for 39° and 42° filaments suggests that the synthesis of enough S for division takes about 20 min at 25° while the septation and separation stages of division take only about 10 min at this temperature. Since these latter processes occur so quickly it seems likely that events other than septation and separation take up a substantial fraction of the D period.

Bacterial cell division regulation: lysogenization of conditional cell division lon - mutants of Escherichia coli by bacteriophage.

The lon(-) mutants of Escherichia coli grow apparently normally except that, after temporary periods of inhibition of deoxyribonucleic acid synthesis, septum formation is specifically inhibited. Under these conditions, long, multinucleate, nonseptate filaments result. The lon(-) mutation also creates a defect such that wild-type bacteriophage lambda fails to lysogenize lon(-) mutants efficiently and consequently forms clear plaques on a lon(-) host. Two lines of evidence suggest that this failure probably results from interference with expression of the lambdacI gene, which codes for repressor, or with repressor action:-(i) when a lon(-) mutant was infected with a lambdacII, cIII, or c Y mutant, there was an additive effect between the lon(-) mutation and the lambdac mutations upon reduction of lysogenization frequency; and (ii) lon(-) mutants permitted the growth of the lambdacro(-) mutant under conditions in which the repressor was active. The isolation of lambda mutants (lambdatp) which gained the ability to form turbid plaques on lon(-) cells is also reported.

ANALYSIS OF A GENE CONTROLLING CELL DIVISION AND SENSITIVITY TO RADIATION IN ESCHERICHIA COLI.

Adler, Howard I. (Oak Ridge National Laboratory, Oak Ridge, Tenn.), and Alice A. Hardigree. Analysis of a genetic locus controlling cell division and sensitivity to radiation in Escherichia coli. J. Bacteriol. 87:720-726. 1964.-Radiation sensitivity in Escherichia coli is under control of genes that are transferred during conjugation between donor and recipient strains. Conjugation experiments establish that one of these genes occupies a locus on the E. coli K-12 linkage map between the genes controlling ability to utilize lactose and galactose. It affects sensitivity to both ionizing and ultraviolet (2,537 A) radiation. A strain possessing a mutation at this locus fails to show increases in resistance to ionizing radiation during late lag and early log phases, and increases in resistance when grown to stationary phase in a glucose-containing complete medium. The primary effect of the mutation at this locus may be an interference with the mechanism by which cells form cross plates. Cells of the mutant form long, nonseptate filaments when grown after exposure to ionizing radiation. The filaments do not give rise to macrocolonies. Pantoyl lactone, an agent that initiates cross plate formation, allows the filaments to divide normally and produce macrocolonies. When plated after irradiation on complete medium containing pantoyl lactone, the survival of the mutant is greatly increased.

Krassilnikoviae: a new class of micro-organisms found in sea and ocean depths.

Summary: A new class of micro-organisms, the Krassilnikoviae, found in the Black Sea, the Pacific and the Arctic Oceans, is described. These organisms developed on slides immersed in the waters; they have not been cultivated in the laboratory. They consist of non-branching, non-septate filaments with a head at one end consisting of a cluster of round bodies. The filaments often become interlaced and form strings which can just be seen by the naked eye. They may or may not be contained in a sheath.