A large scale quasi-crystalline lamellar lattice in chloroplasts of the green alga Zygnema.

A quasi-crystalline lamellar lattice was observed in chloroplasts of the filamentous green alga Zygnema. The lattice does not appear in the cells until cultures are at the end of the log phase of growth. Pseudograna are also present and become more numerous towards the middle of the log phase. The three-dimensional lattice superficially resembles the configuration of cubic prolamellar bodies but is about 10 times larger and is entirely different in internal structure. The lattice is composed of one or two appressed thylakoids in a stroma matrix which is bounded on each side by a single thylakoid membrane. This multilayered sandwich of membranes and matrix occupies a position equivalent to the single membrane of a cubic prolamellar body.

Dietary phytochemicals as quorum sensing inhibitors.

Quorum sensing (QS) is a cell density dependent expression of species in bacteria mediated by hormone like compounds called autoinducers (AI). Several processes responsible for successful establishment of bacterial infection are mediated by QS. Inhibition of QS is therefore being considered as a new target for antimicrobial chemotherapy. Dietary phytochemicals are secondary metabolites in plants known to have several health benefits including antimicrobial activity. However, their ability to inhibit QS has never been studied. Our objective was to investigate the effect of sub-lethal concentrations (SLC) of bioactive dietary phytochemical extracts from common dietary fruit, herb and spice extracts on modulating QS mediated by AI in model bioassay test systems. QS inhibition was measured in violacein pigment producing Chromobacterium violaceum O26 (CVO26) and CV 31532 system, mediated by AI known as acylated homeserine lactone (AHL). We also investigated the effect of the sub-lethal concentrations of the extracts on swarming motility of pathogens Escherichia coli (EC)O157:H7 and Pseudomonas aeruginosa (PA-01). Our results indicate that all extracts significantly inhibited quorum sensing. The mechanism of inhibition appeared to be combination of interfering with AHL activity and modulating the synthesis of AHL's. Our results also indicated that various phytochemical extracts which inhibited QS also inhibited swarming of pathogenic bacteria, known to be modulated by QS. The observation that phytochemicals from foods can inhibit QS related processes opens up an exciting new strategy for antimicrobial chemotherapy and lead to the discovery of new category of antibiotics which can overcome the issues related to antimicrobial resistance.

The effects of gabapentin and memantine in acquired and congenital nystagmus: a retrospective study.

BACKGROUND: Pharmacological treatment has been successful in some forms of acquired neurological nystagmus. However, drugs are not known to be effective in idiopathic infantile nystagmus or nystagmus associated with ocular diseases. METHODS: The authors retrospectively analysed Snellen visual acuity (VA), subjective visual function, and eye movement recordings of 23 patients with nystagmus (13 secondary to multiple sclerosis, three associated with other neurological diseases, two idiopathic infantile, and five with associated ocular diseases) treated with gabapentin or memantine. RESULTS: With gabapentin, 10 of 13 patients with nystagmus secondary to multiple sclerosis (MS) showed some improvement. Memantine improved the VA in all three patients with MS who did not improve on gabapentin. There was no change of nystagmus in other neurological disorders. Patients with congenital nystagmus showed reduction of nystagmus and their VA changes depended on the ocular pathology. CONCLUSION: Gabapentin and memantine may be effective in acquired nystagmus secondary to MS. To the authors' knowledge this is the first series of patients showing that gabapentin is effective in improving nystagmus in congenital nystagmus/nystagmus associated with ocular pathology. Memantine may be useful as an alternative drug in treating patients with nystagmus.

Phenotype characterization of genetically defined microorganisms and growth of bacteriophage in biofilms.

Phenotypic characterization will be a pivotal aspect of future research in understanding the biofilm mode of growth. We hope that the concepts and techniques presented in this chapter will benefit other investigators in this field. Although initial studies will necessarily involve monocultures, eventually mixed culture work will have to be performed to understand biofilm growth in the natural environment. As the study of biofilm-phage interactions is new, there is considerable fundamental work that needs to be addressed. Here, we anticipate that some phage are better adapted to growth in biofilms, some are adept in growing in mixed culture biofilms, and others are better adapted to infecting planktonic organisms. Whereas biofilms are now widely accepted as a fundamental aspect of microbial growth in nature, the field of phage ecology is quite new and an exciting challenge for the future.

Evidence of autoinducer activity in naturally occurring biofilms.

N-Acyl homoserine lactone (AHL) molecules have been shown to act as mediators of population density-dependent (quorum-sensing) gene expression in numerous Gram-negative bacteria. Functions associated with AHL include light production in Vibrio fischeri, expression of virulence factors in Pseudomonas aeruginosa, and conjugation in Agrobacterium tumefaciens. In nature, bacteria often grow as surface-adherent biofilm communities. As biofilms typically contain high concentrations of cells, AHL activity and quorum-sensing gene expression have been proposed as essential components of biofilm physiology. However, proof of AHL production within biofilms has heretofore been lacking. In this study we have employed a cross-feeding assay, using A, tumefaciens A136 (traI::lacZ) as an AHL-responsive reporter strain, to show the presence of naturally occurring AHL production in aquatic biofilms growing on submerged stones. AHL was detected in living biofilms and biofilm extracts, but was not present in rocks lacking a biofilm. This represents the first report of AHL activity in naturally occurring biofilms.

Bacterial biofilm formation under microgravity conditions.

Although biofilm formation is widely documented on Earth, it has not been demonstrated in the absence of gravity. To explore this possibility, Pseudomonas aeruginosa, suspended in sterile buffer, was flown in a commercial payload on space shuttle flight STS-95. During earth orbit, biofilm formation was induced by exposing the bacteria to sterile media through a 0.2-microm (pore size) polycarbonate membrane. Examination of these membranes by confocal microscopy revealed biofilms to be present and that these biofilms could persist in spite of vigorous agitation. These results represent the first report of biofilm formation under microgravity conditions.

Observations of the ultrastructure of infected kidney stones.

Struvite stones are formed as the result of urinary tract infection by urease-producing bacteria. Ultrastructural examination of calculi removed from a patient revealed bacteria incorporated throughout the stone matrix. Exopolysaccharide stained by ruthenium red was associated with most of the bacteria, but it represented only a small portion of the organic matrix in the stone. Localised deposits of calcium and phosphorus, components of carbonate-apatite, and magnesium, a struvite component, were detected in close proximity to the cells. Histochemical examinations revealed that several of the gram-negative bacteria within the stone matrix possessed high levels of urease activity. We propose that bacterial slime production, intimately involved in the initiation of stone matrix deposition, is less prominent in mature stones because of the increased incorporation of host-derived mucoproteins and mucopolysaccharides.

Pyrophosphate inhibition of Proteus mirabilis-induced struvite crystallization in vitro.

Struvite (MgNH4PO4.6H2O) crystals, the major mineral component of infectious urinary calculi, were produced in vitro by growth of a clinical isolate of Proteus mirabilis in artificial urine. P. mirabilis growth and urease-induced struvite production were monitored by phase contrast light microscopy and measurements of urease activity, pH, ammonia concentrations, turbidity, and culture viability. In the absence of pyrophosphate, struvite crystals appeared within 3-5 h due to the urease-induced elevation of pH and initially assumed a planar or 'X-shaped' crystal habit (morphology) characteristic of rapid growth. When pyrophosphate was present, initial precipitation and crystal appearance were significantly impaired and precipitates were largely amorphous. When crystals did appear (usually after 7 or 8 h) they were misshapen or octahedral in shape indicative of very slow growth. X-ray diffraction and Fourier transform infrared spectroscopy (FTIR) identified all crystals as struvite. Trace contaminates of carbonate-apatite (Ca10(PO4)6CO3) or newberyite (MgHPO4.H2O) were produced only in the absence of pyrophosphate. P. mirabilis viability and culture pH elevation were unaffected by the addition of pyrophosphate, whereas urease activity and ammonia concentrations were marginally reduced. Struvite could also be produced chemically by titration of the artificial urine with NH4OH. If pyrophosphate was present during titration, the same inhibitory effect on crystal growth occurred, so it is unlikely that urease inhibition is important. Lowering of pyrophosphate concentration from 13-0.45 mumol/l did not reduce its inhibitory activity so it is unlikely to act by chelating free Mg2+. We propose that pyrophosphate inhibits struvite growth principally through direct interference with the chemical mechanisms involved in crystal nucleation and growth, because of its effectiveness at very low concentrations.

ULTRASTRUCTURE OF SPONGIOCHLORIS TYPICA DURING SENESCENCE(1) (2).

The ultrastructural changes which occurred during senescence in the stationary phase of growth of the unicellular green alga Spongiochloris typica were observed. The cell wall consists of a membrane like primary wall and an inner secondary wall which becomes progressively thickened with age of the culture. During senescence the lamellae become more compact within the chloroplast. The major feature of aging is the appearance of lipid bodies which eventually come to occupy a major portion of the cell lumen. The ultrastructural changes observed to occur during senescence are discussed in relation to physiological data.

NEW TAXONOMIC CRITERIA IN THE CLASSIFICATION OF CHLOROCOCCUM SPECIES. I. PIGMENTATION(1) (2).

Eighteen isolates of Chlorococcum were incubated under defined culture conditions for 6-7 weeks. At that time, isolates were separated into green, yellow-green, orange, or red color groups according to the color of the cultures. Total chlorophyll, carotenoid concentration, and percentage of carotenoids in relation to total pigment content were measured for each species. Individual carotenoids of each isolate were identified and quantified. Each color group had pigment characteristics which were distinctive. Other studies on Chlorococcum are discussed in relation to this work. It is proposed that pigmentation of old cultures be used as the first criterion for distinguishing species of Chlorococcum.

Bacterial colonization behaviour: a new virulence strategy in urinary infections?

The urinary bladder resists bacterial colonization and infection by a number of mechanisms, one of which involves the sloughing of colonized uroepithelial cells. Pathogens which thus become detached from bladder tissue are rapidly eliminated upon voiding of urine. During a recent study of bacterial colonization by the urinary pathogen, Proteus mirabilis, we noted that it colonized glass surfaces such that organisms became widely and evenly dispersed over the surface. In contrast, Pseudomonas fluorescens, a non-pathogen in the urinary tract, did not disperse over the surface but colonized and grew in such a manner as to form small clumps or microcolonies. Other investigators have also shown that Escherichia coli, a common urinary pathogen, initially colonizes bladders in a random, widely-dispersed fashion. We propose that successful bladder pathogens will predominantly adopt colonization behaviour that enables them to widely disperse over bladder tissue and, in so doing, avoid being cleared by the desquamation of uroepithelial cells. Colonization behaviour would therefore represent a previously uncharacterized virulence strategy.

Gram-negative outer membrane vesicles: beyond the cell surface.

Considerable interest has recently mounted regarding the biological roles of Gram-negative outer membrane vesicles (MVs). The first discovery of MVs was made over four decades ago, and it is now clear that most Gram-negative bacteria produce MVs, with Pseudomonas aeruginosa and Escherichia coli as the most extensively studied. Much of our knowledge of the biological roles of MVs and mechanism of MV formation is due to T.J. Beveridge and colleagues. Beveridge pioneered the field of MV research not only by enhancing our understanding of MV function, but also through the application of a wide variety of physical, chemical, and genetic techniques to complement his elegant electron microscopy investigations. Here we review the contributions of Beveridge's group to our understanding of MV biology.

Identification and characterization of stripe rust resistance gene Yr34 in common wheat.

An uncharacterized source of seedling resistance to Puccinia striiformis f.sp. tritici was identified in an advanced wheat breeding line WAWHT2046. Genetic analysis based on a WAWHT2046/Carnamah-derived double haploid (DH) population demonstrated monogenic inheritance of seedling stripe rust resistance in WAWHT2046. The gene controlling stripe rust resistance in line WAWHT2046 was tentatively designated YrWA. The chromosome 5AL located awn inhibitor gene B1, possessed by WAWHT2046, also showed monogenic inheritance when the DH population was scored for the presence and absence of awns. Joint segregation analysis at the B1 and YrWA loci indicated genetic linkage between the two loci. A recombination value of 12.2 cM was computed using Mapmanager. This association located YrWA in the chromosome arm 5AL. Molecular mapping using microsatellite markers placed YrWA distal to B1. All molecular markers mapped proximal to the awn inhibitor locus B1. As no other stripe rust resistance gene is reported to be located in the chromosome arm 5AL, YrWA was permanently designated as Yr34. Yr34 produced an intermediate (23C) seedling infection type and expressed very low stripe rust response (10R-MR) on adult plants in the field, similar to the resistance gene Yr17. In addition to Yr34, this mapping population segregated for three genetically independent adult plant stripe rust resistance genes. The detection of DH lines with completely susceptible response, higher than that shown by the Yr34-lacking parent Carnamah, suggested that both parents contributed adult plant resistance. The use of WAWHT2046 as a parent in breeding programs would also contribute APR in addition to Yr34.

REJUVENATION OF SENESCENT CELLS OF SPONGIOCHLORIS TYPICA(2).

Thick-walled, oil-filled akinetes of the green alga Spongiochloris typica, long in the stationary phase of growth, were transferred to fresh media. Ultra-structural changes were observed during a 51-hr period after transfer. Akinetes, when transferred to fresh media, may follow 1 of 3 pathways: 1 Remain unchanged; these cells appear to be moribund perhaps due to an over-accumulation of lipid. 2 Undergo immediate zoosporogenesis; seems to occur in cells with 4, 8, or more nuclei. 3 Return to the vegetative state; this was common in smaller cells with less than 4 nuclei; however, zoosporogenesis eventually did occur when 4, 8, or more nuclei were present.

PHYSIOLOGICAL CHANGES OF SPONGIOCHLORIS TYPICA IN CULTURE(1) (2).

The physiological changes in the green alga Spongiochloris typica were followed in liquid and agar for 8 weeks. Agar cultures changed drastically in lipid content, pigment composition, and heat and desiccation resistance. Changes in liquid cultures were gradual and not as extreme as those of agar cultures. However, rate and extent of dry-weight increments were similar in liquid and on agar. Liquid cultures are seemingly more stable in the physiological responses than are agar cultures where the environmental changes are conspicuous.

NEW TAXONOMIC CRITERIA IN CLASSIFICATION OF CHLOROCOCCUM SPECIES. II. PYRENOID FINE STRUCTURE(2).

The pyrenoids of 18 Chlorococcum species (including 3 now transferred to Tetracystis) have been investigated at the ultrastructural level. Three categories of pyrenoids were observed, including pyrenoids with unfragmented perforate starch plates in 11 species; those with many separate fragmented starch plates in 6 species; and those with hemispherical starch plates in 1 species. Pyrenoids were further distinguishable on the basis of the number of thylakoid disks or tubules which penetrate the ground substance. Stability and diversity of pyrenoid fine structure as a taxonomic criterion at the specific level have been demonstrated.

Impact of rpoS deletion on Escherichia coli biofilms.

Slow growth has been hypothesized to be an essential aspect of bacterial physiology within biofilms. In order to test this hypothesis, we employed two strains of Escherichia coli, ZK126 (DeltalacZ rpoS(+)) and its isogenic DeltarpoS derivative, ZK1000. These strains were grown at two rates (0.033 and 0.0083 h(-1)) in a glucose-limited chemostat which was coupled either to a modified Robbins device containing plugs of silicone rubber urinary catheter material or to a glass flow cell. The presence or absence of rpoS did not significantly affect planktonic growth of E. coli. In contrast, biofilm cell density in the rpoS mutant strain (ZK1000), as measured by determining the number of CFU per square centimeter, was reduced by 50% (P < 0.05). Deletion of rpoS caused differences in biofilm cell arrangement, as seen by scanning confocal laser microscopy. In reporter gene experiments, similar levels of rpoS expression were seen in chemostat-grown planktonic and biofilm populations at a growth rate of 0.033 h(-1). Overall, these studies suggest that rpoS is important for biofilm physiology.

Influence of metal ions and temperature on the conformation of Escherichia coli K1 capsular polysaccharide.

Escherichia coli K1 secretes a homopolymer capsular polysaccharide (CPS) consisting of alpha 2, 8 linked N-acetylneuraminic acid (poly alpha 2,8NeuNAc). Typically poly alpha 2,8NeuNAc is arranged in low and high order alpha helices with carboxyl and hydroxyl groups extending from the helices. Several properties of CPS such as antigenicity and metal binding can be influenced by its structural conformation. We examined the influences of metal ions and temperature on the secondary structure of poly alpha 2,8NeuNAc. Conformation alteration was detected by ultraviolet (UV) spectroscopy and circular dichroism (CD). The majority of metal ions tested had no detectable influence on poly alpha 2,8NeuNAc structure. In contrast, Yb3+, Hg2+, and Cu2+ ions greatly altered the UV and CD spectra, which suggests that these ions had disrupted the alpha helical structure of poly alpha 2,8NeuNAc. These changes were influenced by the metal ion concentration. When poly alpha 2,8NeuNAc was incubated at temperatures ranging from 20-60 degrees C, alterations in its UV absorption spectra were also seen. The most significant change occurred between 35 and 40 degrees C. In summary, this study suggests that the higher order structure and function of bacterial CPS may be influenced by environmental factors.

Rhizoplast and rootlet system of the flagellar apparatus of Chlamydomonas moewusii.

The ultrastructure of the flagellar apparatus of Chlamydomonas moewusii was examined in detail. Two rhizoplasts were found associated with the basal bodies of this biflagellate and were observed to extend to the central area of the cell. A segment of smooth endoplasmic reticulum ran parallel to each rhizoplast. These 2 segments anastomose beneath the basal bodies and the tubule proceeds over the top of the distal connecting fibre. A functional role for the smooth endoplasmic reticulum in these locations is discussed. Four sets of rootlet microtubules emanate from a region between the 2 basal bodies and the distal connecting fibre. Two sets have a three-over-one arrangement and the other 2 sets are doublets. Cytoplasmic microtubules were seen associated with possible nucleating sites on the rootlet microtubules. The association of the observed structures are discussed and compared to the flagellar apparatus of C. reinhardtii.