De novo morphogenesis in L‐forms via geometric control of cell growth

In virtually all bacteria, the cell wall is crucial for mechanical integrity and for determining cell shape. Escherichia coli's rod‐like shape is maintained via the spatiotemporal patterning of cell‐wall synthesis by the actin homologue MreB. Here, we transiently inhibited cell‐wall synthesis in E. coli to generate cell‐wall‐deficient, spherical L‐forms, and found that they robustly reverted to a rod‐like shape within several generations after inhibition cessation. The chemical composition of the cell wall remained essentially unchanged during this process, as indicated by liquid chromatography. Throughout reversion, MreB localized to inwardly curved regions of the cell, and fluorescent cell wall labelling revealed that MreB targets synthesis to those regions. When exposed to the MreB inhibitor A22, reverting cells regrew a cell wall but failed to recover a rod‐like shape. Our results suggest that MreB provides the geometric measure that allows E. coli to actively establish and regulate its morphology.     

The liposoluble proteome of <Emphasis Type="Italic">Mycoplasma agalactiae</Emphasis>: an insight into the minimal protein complement of a bacterial membrane

Background  

Mycoplasmas are the simplest bacteria capable of autonomous replication. Their evolution proceeded from gram-positive bacteria, with the loss of many biosynthetic pathways and of the cell wall. In this work, the liposoluble protein complement of Mycoplasma agalactiae, a minimal bacterial pathogen causing mastitis, polyarthritis, keratoconjunctivitis, and abortion in small ruminants, was subjected to systematic characterization in order to gain insights into its membrane proteome composition.     

Neutrophil Response to Nontypable Haemophilus influenzae in Respiratory Infections

Sputa from patients with respiratory infections by nontypable Haemophilus influenzae (H. influenzae) were investigated by electron microscopy. The cell wall of H. influenzae appeared wavy and nonwavy. In the cell wall the peptidoglycan layer was ill-defined. These patients had adequate IgG response in the serum against H. influenzae. However neither capsule nor fimbriae were found. Different stages of phagocytosis and destruction of the bacteria by polymorphonuclear neutrophils (PMN) were observed. PMNs were also found to phagocytose the debris. Evidences were found that the debris is formed mainly by the destruction of polymorphonuclear neutrophil. Extracellular lysosomes were also observed, which may have a role in destruction of both bacteria and host tissue. It was concluded that nontypable H. influenzae are nonfimbriated and noncapsulated during infection. Debris are the end product of PMN destruction, and phagocytosis of debris by PMNs has a role in the pathogenesis of chronic respiratory diseases.     

Effect of environmental factors on the composition of lipopolysaccharides released from the <Emphasis Type="Italic">Rhodobacter capsulatus</Emphasis> cell wall

The goal of this study was to compare the environmental factors that determine viability of the gram-negative photosynthesizing bacteria Rhodobacter capsulatus (specifically, light and growth medium composition) and to assess the effect of these factors on the synthesis and composition of lipopolysaccharides released from the cell wall. Depletion of medium resulted in the release of lipopolysaccharides with a truncated polysaccharide fragment. It is concluded that, due to high viability of these bacteria, the main factor that induced lipopolysaccharide release from the cell wall, irrespective of culturing conditions, is cell division rather than cell death.Translated from Izvestiya Akademii Nauk, Seriya Biologicheskaya, No. 2, 2005, pp. 168–173.Original Russian Text Copyright © 2005 by Zubova, Melzer, Prokhorenko.     

The Mechanism of Action of the Extracellular Bacteriolytic Enzymes of Lysobacter sp. on Gram-Positive Bacteria: The Role of the Cell Wall Anionic Polymers of Target Bacteria

The study of the extracellular bacteriolytic enzymes of Lysobacter sp. showed that they can efficiently hydrolyze the peptidoglycan of gram-positive bacteria provided that there is an electrostatic interaction of these enzymes with the cell wall anionic polymers, teichoic and teichuronic acids in particular. The hydrolytic action of bacteriolytic enzymes on the cell wall largely depends on the negative charge of the teichoic and teichuronic acids rather than on their chemical composition.     

Peptidoglycan turnover and recycling in Gram-positive bacteria

Bacterial cells are protected by an exoskeleton, the stabilizing and shape-maintaining cell wall, consisting of the complex macromolecule peptidoglycan. In view of its function, it could be assumed that the cell wall is a static structure. In truth, however, it is steadily broken down by peptidoglycan-cleaving enzymes during cell growth. In this process, named cell wall turnover, in one generation up to half of the preexisting peptidoglycan of a bacterial cell is released from the wall. This would result in a massive loss of cell material, if turnover products were not be taken up and recovered. Indeed, in the Gram-negative model organism Escherichia coli, peptidoglycan recovery has been recognized as a complex pathway, named cell wall recycling. It involves about a dozen dedicated recycling enzymes that convey cell wall turnover products to peptidoglycan synthesis or energy pathways. Whether Gram-positive bacteria also recover their cell wall is currently questioned. Given the much larger portion of peptidoglycan in the cell wall of Gram-positive bacteria, however, recovery of the wall material would provide an even greater benefit in these organisms compared to Gram-negatives. Consistently, in many Gram-positives, orthologs of recycling enzymes were identified, indicating that the cell wall may also be recycled in these organisms. This mini-review provides a compilation of information about cell wall turnover and recycling in Gram-positive bacteria during cell growth and division, including recent findings relating to muropeptide recovery in Bacillus subtilis and Clostridium acetobutylicum from our group. Furthermore, the impact of cell wall turnover and recycling on biotechnological processes is discussed.     

THE EFFECTS OF MECHANICAL STIMULATION AND ETIOLATION ON THE COLLENCHYMA OF DATURA STRAMONIUM

Walker , Waldo S. (Grinnell College, Grinnell, Iowa.) The effects of mechanical stimulation and etiolation on the collenchyma of Datura stramonium. Amer. Jour. Bot. 47(9) : 717–724. Illus. 1960.–In an effort to determine the effect of mechanical stimulation on collenchyma tissue, plants of Datura stramonium L. were placed on a mechanical agitator and subjected to intensive shaking for 9 hr. per day for 40 days. Measurements indicated that such stimulation greatly increased the amount of wall thickening per cell, as observed in transverse section. Measurements also indicated that such stimulation may inhibit collenchyma cell elongation. A second group of Datura stramonium plants was placed in total darkness to determine the effect of such treatment on the quantity of wall thickening in the collenchyma tissue. Measurements indicated that when plants were placed in the dark for extended periods a great reduction of wall thickening resulted. It is suggested that reduction of wall material was due to its utilization as substrate for respiratory processes which occur in the plant under such extreme conditions. The composition and structure of the collenchyma cell walls are discussed.     

Proteolytic cleavage and cell wall anchoring at the LPXTG motif of surface proteins in Gram-positive bacteria

Many surface proteins are thought to be anchored to the cell wall of Gram-positive bacteria via their C-terminus. Cell wall anchoring requires a specific sorting signal, normally located at the predicted C-terminus of surface proteins. Here we show that when placed into the middle of a polypeptide chain, the sorting signal causes the specific cleavage of the precursor as well as the cell wall anchoring of its N- terminal fragment, while the C-terminal fragment remains within the cytoplasm. N-terminal sequencing of the C-terminal cleavage fragment suggests that the cleavage site is located between threonine (T) and glycine (G) of the LPXTG motif, the signature sequence of cell wall sorting signals. All surface proteins harbouring an LPXTG sequence motif may therefore be cleaved and anchored by a universal mechanism. We also propose a novel hypothesis for the cell wall linkage of surface proteins in Gram-positive bacteria.     

Effect of specific growth limitations on cell wall composition of Staphylococcus aureus H

Conditions are described for the continuous culture of a derivative of Staphylococcus aureus H in a fully defined minimal medium in which cysteine is the sole amino acid. The effects of growth under various nutrient limitations on the composition and properties of the cell wall have been studied. The proportion of ribitol teichoic acid present in the wall, and the extent to which it is substituted with N-acetylglucosamine, varies in bacteria grown under different conditions as does the composition and extent of cross-linking of the peptidoglycan. Neither the derivative nor the original strain H produced teichuronic acid when grown under phosphate limitation.Non-Standard Abbreviation SDS Sodium dodecyl sulphate     

Russian Article pp. 309–318

It is demonstrated that aliphatic hydrocarbons (alkanes) penetrate into bacteria cells by the way of passive diffusion. The mechanism of this process is different for several bacteria species. A hydrophobic cell wall is essential for that process. In saprophytic Mycobacteria hydrocarbons are solubilized in the thick hydrophobic cell wall. During the process of absorption hydrocarbons pass through the whole cell wall up to the membrane. In the case of Arthrobacteria the hydrocarbons might pass not through the whole cell wall, but through special lipophilie canals. Mobile hydrocarbon-oxidizing bacteria g. Pseudomonas form peptidoglycolipid and excrete it into the medium. The peptidoglycolipid emulsifies hydrocarbon substrate.     

Study of phototrophic purple bacterium <Emphasis Type="Italic">Rhodobacter sphaeroides</Emphasis> cell morphology of wild-type and ipt-transformant by atomic force and electron microscopy

A comparative study of phototrophic purple bacterium Rhodobacter sphaeroides cell morphology of wild-type and ipt-transformant was done by atomic force and electron microscopy. It was shown that transformation led to a decrease in the number or total disappearance of the flagella, as well as to changes in the structure of the outer membrane of the bacteria cell wall. On the wild-type cell surface phage-like structures were found, and in transformed cells at their places hollows were identified. This study significantly extends an understanding of the changes occurring in the ipt-transformants of phototrophic purple bacterium Rhodobacter sphaeroides. This investigation not only confirmed earlier obtained data about the differences in the wild-type and ipt-transformant phototrophic purple bacteria cell wall, but also showed fine changes in the structure of its outer membrane.     

α-溶血性链球菌在健康人群口咽部分布及 生物拮抗作用

目的通过采集健康人群口咽部分泌物,分析上呼吸道中α-溶血性链球菌的分布状况,并对革兰阳性化脓性球菌进行生物拮抗试验,为进一步研究上呼吸道益生菌提供理论基础。方法随机自愿原则,用无菌咽拭子采集沈阳市年龄在3～75岁的300名健康人群咽后壁分泌物,对α-溶血性链球菌进行鉴定和定量分析。对致病菌的生物拮抗试验采用小缸杯法。结果定量分析显示不同年龄人群咽后壁的α-溶血性链球菌检出率均较高。在咽后壁菌群中α-溶血性链球菌构成比最多的是幼儿组,达到60.3%。其中唾液链球菌群在幼儿组所占比重最大;老年组人群格氏链球菌占比较大;儿童、青年、成人以缓症链球菌和口腔链球菌为主。对革兰阳性化脓性球菌的生物拮抗试验显示,1株婴儿链球菌婴儿亚种能够拮抗8株致病菌;4株分离菌只能拮抗1株病原菌,提示不同的菌株拮抗病原菌的能力差异较大。结论α-溶血性链球菌在人群中分布广,数量多,不同年龄人群的菌群构成存在差异。并且某些菌株显示出对致病菌较强的生物拮抗作用,推测这些菌株在呼吸道黏膜保护中起到重要作用,可作为上呼吸道益生菌的备选菌株。     

Differentiated roles for MreB‐actin isologues and autolytic enzymes in Bacillus subtilis morphogenesis

Cell morphogenesis in most bacteria is governed by spatiotemporal growth regulation of the peptidoglycan cell wall layer. Much is known about peptidoglycan synthesis but regulation of its turnover by hydrolytic enzymes is much less well understood. Bacillus subtilis has a multitude of such enzymes. Two of the best characterized are CwlO and LytE: cells lacking both enzymes have a lethal block in cell elongation. Here we show that activity of CwlO is regulated by an ABC transporter, FtsEX, which is required for cell elongation, unlike cell division as in Escherichia coli. Actin‐like MreB proteins are thought to play a key role in orchestrating cell wall morphogenesis. B. subtilis has three MreB isologues with partially differentiated functions. We now show that the three MreB isologues have differential roles in regulation of the CwlO and LytE systems and that autolysins control different aspects of cell morphogenesis. The results add major autolytic activities to the growing list of functions controlled by MreB isologues in bacteria and provide new insights into the different specialized functions of essential cell wall autolysins.     

Electron Microscopic Studies on the Outer Layers of Staphylococcus aureus Using a Lytic Enzyme from Flavobacterium

The structure of the outer layers (cell wall and membrane) of Staphylococcus aureus was studied by electron microscope using a bacteriolytic enzyme from Flavobacterium sp. called the L-11 enzyme. Comparative studies on the morphology of bacteria before and after treatment with this enzyme and cell wall and membrane fractions obtained from bacteria after the enzyme treatment led to the following conclusions. (1) The cell wall of S. aureus is composed of morphologically distinct two layers which are both susceptible to the L-11 enzyme. (2) Between the cell wall and membrane, there is an electron opaque region which could not be stained using any of the methods tested. (3) Before treatment of bacteria with the enzyme the cell membrane could not be seen clearly. However, after enzyme treatment the membrane was clearly seen. (4) The infolding of the inner layer of the cell wall, forming a structure like a mesosome, was liberated by extensive enzyme treatment.     

LYSIS OF MICROCYSTIS AERUGINOSA (KÜTZ.) BY BDELLOVIBRIO-LIKE BACTERIA1

Cells of Microcystis aeruginosa (Kützing), collected from water-blooms of Lake Varese, were lysed by Bdellovibrio-like bacteria. The cells were lysed only after penetration. The cyanobacteria and lysing bacteria were characterized by a fibrous glycocalyx. Once the host cell was penetrated, the bacteria remained localized mainly between the host cell wall and cytoplasmic membrane, which appeared partially thickened. Cell lysis began by breakdown of cell structures. The cell wall appeared broken at many sites, and in completely lysed cells, was partially interrupted. The lysis of Microcystis by bacteria could be one of the causes of the death of algal blooms.     

Neutrophil Response to Pseudomonas aeruginosa in Respiratory Infection

Sputum from patients with acute exacerbation of respiratory infection by Pseudomonas aeruginosa was observed under the electron microscope. External to the cell wall of P. aeruginosa a granular, electron-dense material was observed which is suggestive of capsule. It is supposed that stabilization of capsule occurred by the host antibody, which was produced due to chronic infection by P. aeruginosa. Mucoid type of microcolonies were observed with a fibrous matrix of exopolysaccharide. Other types of microcolonies were surrounded by granular substances or fine fibers. Neutrophil was found to be partially surrounding the microcolony in an attempt to defense. Debris was formed mainly by the destruction of the neutrophil. Most neutrophils were found full of phagocytosed debris; in contrast only a few neutrophils were found to have phagocytosed P. aeruginosa. This study concludes that instead of phagocytosing bacteria, neutrophil phagocytosed debris and bacteria were not completely eradicated. Therefore, this might be one of the factors in the pathogenesis of respiratory infection and persistent colonization by P. aeruginosa.     

Extremophilic Bacillus cereus MVK04 Isolated from Thorium Ore Sample Possesses Self-Assembled Surface Layer Protein on Cell Wall to Resist Extreme Environments

The present study investigates the extremophilic nature of bacteria present in thorium rich mine ore samples collected from Manavalakuruchi, Tamilnadu, India. Six different bacteria strains were isolated from these ore samples and screened for its resistance against varying pH, uranium and gamma radiation. Deinococcus radiodurans ATCC 13939 and Escherichia coli MTCC 1687 was used as positive and negative control respectively. Among the six different bacterial strains, MVK04 strain was found to resist higher pH, uranium concentration and gamma radiation. The organism was identified as Bacillus cereus based on biochemical, 16S rRNA and MALDI TOF fingerprinting studies. The interaction of uranium ions with MVK04 bacterial cell wall was investigated using high resolution transmission electron microscopy (HRTEM) and energy dispersive spectroscopy (EDS). The bacterial MVK04 was further investigated for the presence of surface layer protein (S-layer protein) on the bacterial cell wall. The S-layer protein was isolated, partially purified and self-assembled onto TEM copper grids and the self-assembled protein nanostructures were characterized using HRTEM. The presence of surface layer protein on bacterial cell wall may be the possible reason for its extremophilic characteristics and its escape from lethal effect of higher concentration of uranium, lower pH and increased gamma radiation.     

Expression of Fimbriae and Host Response in Branhamella catarrhalis Respiratory Infections

Sputum during the acute exacerbation of chronic respiratory diseases were observed under the electron microscope, to determine the in vivo expression of surface structures of Branhamella catarrhalis (B. catarrhalis), the polymorphonuclear neutrophil (PMN) response to B. catarrhalis infections, and the composition of sputum. It was found that during infection fimbriae are expressed in B. catarrhalis. However, there were sparsely to densely fimbriated bacteria in each sputum sample. The length of the fimbriae were from 50 to 76 nm. In the sparsely fimbriated B. catarrhalis, external to the cell wall, a thin, granular, electron-dense layer was observed. Due to the presence of fimbriae, this layer was not seen in densely fimbriated B. catarrhalis. Blebs were also found in B. catarrhalis. PMNs were found to phagocytose both B. catarrhalis and debris. Evidence was found that debris were formed mainly by the destruction of PMNs. Bacteria as well as debris were phagocytosed by PMNs.     

CELL WALL CONSTITUENTS OF APHANOTHECE HALOPHYTICA (CYANOPHYTA)1

Some aspects of the cell wall and extracellular polysaccharide (ECPS) of the obligate halophile Aphanothece halophytica Frémy (Chroococcales) have been investigated. Extracellular polysaccharide concentration was found to remain constant on a per cell basis in medium containing from 1–3 M NaCl. The rate of ECPS production remained constant during mid-log growth phase and increased substantially as the culture reached stationary phase. The lipopolysaccharide of this organism was found to possess a low and unusual fatty acid content when compared to other chroococcalean forms. The cell wall appears to contain a typical gram-negative peptidoglycan. The covalently attached protein resembles the envelope protein of extremely halophilic bacteria in its possession of a similar molar percentage of amino acids with lipophilic R-groups and a high acidic amino acid fraction. The ECPS and cell wall fractions of A. halophytica were found to chemically more closely resemble those from other non-halophilic, chroococcalean bluegreen algae than those from the obligately halophilic bacteria.     

Dynamics of intracellular mannan and cell wall folding in the drought responses of succulent Aloe species

Plants have evolved a multitude of adaptations to survive extreme conditions. Succulent plants have the capacity to tolerate periodically dry environments, due to their ability to retain water in a specialized tissue, termed hydrenchyma. Cell wall polysaccharides are important components of water storage in hydrenchyma cells. However, the role of the cell wall and its polysaccharide composition in relation to drought resistance of succulent plants are unknown. We investigate the drought response of leaf‐succulent Aloe (Asphodelaceae) species using a combination of histological microscopy, quantification of water content, and comprehensive microarray polymer profiling. We observed a previously unreported mode of polysaccharide and cell wall structural dynamics triggered by water shortage. Microscopical analysis of the hydrenchyma cell walls revealed highly regular folding patterns indicative of predetermined cell wall mechanics in the remobilization of stored water and the possible role of homogalacturonan in this process. The in situ distribution of mannans in distinct intracellular compartments during drought, for storage, and apparent upregulation of pectins, imparting flexibility to the cell wall, facilitate elaborate cell wall folding during drought stress. We conclude that cell wall polysaccharide composition plays an important role in water storage and drought response in Aloe.