A comparative evaluation with respect to the bacterial cleanability of a range of wall and floor surface materials used in the food industry

J. H. TAYLOR AND J. T. HOLAH. 1996. A range of floor and wall materials suitable for use in the food industry were selected for comparative bacterial cleanability tests. A standard cleaning regime was carried out on samples after biofilm development of Acinetobacter calcoaceticus (CRA 296). Cleanability was measured as the log reduction in bacterial numbers. The results for the wall materials showed that there were no statistical differences, whilst those for the floor materials were more variable. The grouted joints and texture did not compromise the cleanabilities of tiled surfaces.     

Ultrastructural specialization at the host-pathogen interface in rust-infected flax

Summary Ultrastructure of the association between the rust fungus, Melampsora lini, and a compatible variety of flax, Linum usitatissimum, was studied to clarify the structural relationships and interactions at the site of host penetration and at the host-parasite interface. Results of freeze-etching as well as a special section-staining procedure consisting of periodate-chromate-phosphotungstate (PACP) are shown with a host-parasite combination for the first time. The host plasma membrane is invaginated by the fungus and forms a continuous boundary around the fungal haustoria which penetrate the host cells. No morphological continuities are observed linking the protoplasts of host and fungus. With both freeze-etching and the PACP stain, the invaginated portion of the host plasma membrane at the host-parasite interface shows distinctive features that are not characteristic of the non-invaginated portion of the membrane. This localized specialization of host plasma membrane in response to the fungus appears as a significant and consistent feature of the host-parasite interaction. The host plasma membrane is separated from the haustorial wall by an amorphous layer of sheath material which covers the body but not the neck of the haustorium. This sheath provides the environment in which the haustorium exists and functions during the course of the host-parasite association. Occasionally, a collar of wall-like material derived from the host cell forms around the haustorial neck. The collar is continuous with the host wall and is distinct and discontinuous from the haustorial sheath. In fewer than 5% of the infected cells this wall material encases entire haustoria. The fungal wall is structurally specialized around the site of host penetration, and it becomes intimately associated with the host wall where the fungus penetrates into the lumen of the host cell. During penetration, the host and fungal walls appear to be fused so that the interface between them is not clearly delineated. The haustorial wall is continuous, via the haustorial neck, with the wall of the haustorial mother cell which lies outside the host cell. Different staining properties reveal this wall continuum to consist of several well-defined regions having different structure or composition. A ring of fungal wall material midway along the haustorial neck stains densely with lead citrate, but is preferentially etched away by periodic acid. The neck ring denotes a transition in the staining reaction of the fungal wall, from that present in the region of host penetration to that of the wall surrounding the haustorium. The findings demonstrate specialization of the fungal wall in the area of host penetration as well as specialization of the host plasma membrane at the host-parasite interface to a degree not previously realized from ultrastructural information.     

How the deposition of cellulose microfibrils builds cell wall architecture

Cell walls, the extracytoplasmic matrices of plant cells, consist of an ordered array of cellulose microfibrils embedded in a matrix of polysaccharides and glycoproteins. This construction is reminiscent of steel rods in reinforced concrete. How a cell organizes these ordered textures around itself, creating its own desirable environment, is a fascinating question. We believe that nature adopted an economical solution to this design problem: it exploits the geometrical constraints imposed by the shape of the cell and the limited space in which microfibrils are deposited, enabling the wall textures essentially to 'build themselves'. This does not imply that the cell cannot control its wall texture. On the contrary, the cell has ample regulatory mechanisms to control wall texture formation by controlling the insertion of synthases and the distance between individual microfibrils within a wall lamella.     

Dodder hyphae invade the host: a structural and immunocytochemical characterization

Summary.  Dodder (Cuscuta pentagona) hyphae are unique amongst the parasitic weeds for their ability to apparently grow through the walls of the host plant. Closer examination reveals, however, that the hyphae do not grow through the host but rather induce the host to form a new cell wall (or extend the existing wall) to coat the growing hypha. This chimeric wall composed of walls from two species is even traversed by plasmodesmata that connect the two cytoplasms. Compositionally, the chimeric wall is quite different from the walls of either the host or in other cells of the dodder plant, on the basis of immunocytochemical labeling. The most striking differences were in the pectins, with much stronger labeling present in the chimeric wall than in either the host or other dodder walls. Interestingly, labeling with monoclonal antibodies specific to arabinan side chains of rhamnogalacturonan I pectin fraction was highly enriched in the chimeric wall, but antibodies to galactan side chains revealed no labeling. Arabinogalactan protein antibodies labeled the plasma membrane and vesicles at the tips of the hyphae and the complementary host wall, although the JIM8-reactive epitope, associated with very lipophilic arabinogalactan proteins, was found only in dodder cells and not the host. Callose was found in the plasmodesmata and along the forming hyphal wall but was found at low levels in the host wall. The low level of host wall labeling with anticallose indicates that a typical woundlike response was not induced by the dodder. When dodder infects leaf lamina, which have more abundant intercellular spaces than petioles or shoots, the hyphae grew both intra- and extracellularly. In the latter condition, a host wall did not ensheath the parasite and there was clear degradation of the host middle lamellae by the growing hyphae, allowing the dodder to pass between cells. These data indicate that the chimeric walls formed from the growth of the host cell wall in concert with the developing hyphae are unique in composition and structure and represent an induction of a wall type in the host that is not noted in surrounding walls. Received February 1, 2002; accepted July 8, 2002; published online November 29, 2002     

Helicoidal pattern in secondary cell walls and possible role of xylans in their construction

The helicoidal organization of secondary cell walls is overviewed from several examples. Both the plywood texture and the occurrence of characteristic defects strongly suggest that the wall ordering is relevant of a cholesteric liquid-crystal assembly that is rapidly and strongly consolidated by lignification. A preferential localization of glucuronoxylans, major matrix components, and in vitro re-association experiments emphasize their preeminent role: (1) during the construction of the composite as directing the cellulose microfibrils in a helicoidal array; (2) during the lignification of the composite as a host structure for lignin precursors.     

Fasciola hepatica: Migration of newly excysted juveniles in resistant rats

Using transmission and scanning electron microscopy, the early migration of juvenile Fasciola hepatica was examined in naive and resistant rats. In naive rats, the migration of flukes to the peritoneal cavity was uneventful. In resistant rats, flukes were rapidly coated with antibody whilst still in the gut lumen and a proportion of the flukes were unable to penetrate the intestinal wall. Those that did penetrate were unharmed as they crossed the gut wall, but on entering the peritoneal cavity they were coated with antibody and host cells including eosinophils, neutrophils, macrophages, and mast cells. Eosinophils were seen degranulating onto the fluke surface, and this appeared to result in the erosion of the tegumental syncytium.     

Local and global geometric influence on steady flow in distal anastomoses of peripheral bypass grafts

We consider the effect of geometrical configuration on the steady flow field of representative geometries from an in vivo anatomical data set of end-to-side distal anastomoses constructed as part of a peripheral bypass graft. Using a geometrical classification technique, we select the anastomoses of three representative patients according to the angle between the graft and proximal host vessels (GPA) and the planarity of the anastomotic configuration. The geometries considered include two surgically tunneled grafts with shallow GPAs which are relatively planar but have different lumen characteristics, one case exhibiting a local restriction at the perianastomotic graft and proximal host whilst the other case has a relatively uniform cross section. The third case is nonplanar and characterized by a wide GPA resulting from the graft being constructed superficially from an in situ vein. In all three models the same peripheral resistance was imposed at the computational outflows of the distal and proximal host vessels and this condition, combined with the effect of the anastomotic geometry, has been observed to reasonably reproduce the in vivo flow split. By analyzing the flow fields we demonstrate how the local and global geometric characteristics influences the distribution of wall shear stress and the steady transport of fluid particles. Specifically, in vessels that have a global geometric characteristic we observe that the wall shear stress depends on large scale geometrical factors, e.g., the curvature and planarity of blood vessels. In contrast, the wall shear stress distribution and local mixing is significantly influenced by morphology and location of restrictions, particular when there is a shallow GPA. A combination of local and global effects are also possible as demonstrated in our third study of an anastomosis with a larger GPA. These relatively simple observations highlight the need to distinguish between local and global geometric influences for a given reconstruction. We further present the geometrical evolution of the anastomoses over a series of follow-up studies and observe how the lumen progresses towards the faster bulk flow of the velocity in the original geometry. This mechanism is consistent with the luminal changes in recirculation regions that experience low wall shear stress. In the shallow GPA anastomoses the proximal part of the native host vessel occludes or stenoses earlier than in the case with wide GPA. A potential contribution to this behavior is suggested by the stronger mixing that characterizes anastomoses with large GPA.     

THE HOST-PARASITE INTERFACE OF ALBUGO CANDIDA ON RAPHANUS SATIVUS

The fine structure of the intercellular hyphae of the obligate parasite Albugo candida infecting radish does not differ markedly from that described previously for cells of Peronospora manshurica. The stalked, capitate haustoria do not contain nuclei and are packed with mitochondria and lomasomes. The fungal plasma membrane and cell wall are continuous from the intercellular hypha throughout the haustorium except that there is no evidence of fungal cell wall around a portion of the haustorial stalk proximal to the haustorial head. Within the vacuolate host mesophyll cell, the haustorium is always surrounded by host plasma membrane and with at least a thin layer of host cytoplasm. The host cell wall invaginates at the point of haustorial penetration to form a short sheath around the region of penetration, but normally there is no host cell wall around the balance of the haustorium. About 1% of the haustoria observed were necrotic, and these were invariably walled-off completely from host cytoplasm by host cell wall. An amorphous, moderately electron-dense encapsulation lies between the haustorium proper and the host plasma membrane and extends into the penetration region between the sheath and the fungal cell wall. Invaded host cells contain more ribosomal-rich ground cytoplasm than uninfected cells. Glandular-like systems of tubules and connecting vesicles are often numerous in host cytoplasm in the vicinity of haustorial heads. These tubules open into the encapsulation, their limiting unit membranes being continuous with the host plasma membrane. We suggest that these represent a secretory mechanism of the host specifically induced by the parasite.     

In vivo and in vitro swelling of cell walls during fruit ripening

Swelling properties of the cell walls of nine temperate fruit species, selected for their different ripening and textural characteristics, were studied during ripening. Cell wall swelling was examined in intact fruit using microscopy techniques and in vitro, using cell wall material isolated from fruit tissue. In fruit which ripened to a soft melting texture (persimmon, avocado, blackberry, strawberry, plum), wall swelling was pronounced, particularly in vitro. In-vivo swelling was marked only in avocado and blackberry. Fruit which ripened to a crisp, fracturable texture [apple (two cultivars), nashi pear, watermelon] did not show either in-vivo or in-vitro swelling of the cell wall. There was a correlation between swelling and the degree of pectin solubilisation, suggesting that wall swelling occurred as a result of changes to the viscoelastic properties of the cell wall during pectin solubilisation. Chemical and enzymatic removal of pectin from kiwifruit cell wall material supported the idea that swelling is associated with movement of water into voids left in the cellulose-hemicellulose network by the solubilised pectin. However, the results also suggested that swelling in vivo was more complex than this, and that the physicochemical changes which led to swelling included other elements of cell wall modification involving the site and mechanism of pectin solubilisation and-or the cellulose-xyloglucan complex. Received: 28 January 1997 / accepted: 11 March 1997     

Antibody response to Candida albicans cell wall antigens

The cell wall of Candida albicans is not only the structure where many essential biological functions reside but is also a significant source of candidal antigens. The major cell wall components that elicit a response from the host immune system are proteins and glycoproteins, the latter being predominantly mannoproteins. Both carbohydrate and protein moieties are able to trigger immune responses. Proteins and glycoproteins exposed at the most external layers of the wall structure are involved in several types of interactions of fungal cells with the exocellular environment. Thus, coating of fungal cells with host antibodies has the potential to profoundly influence the host-parasite interaction by affecting antibody-mediated functions such as opsonin-enhanced phagocytosis and blocking the binding activity of fungal adhesins to host ligands. In this review we examine various members of the protein and glycoprotein fraction of the C. albicans cell wall that elicit an antibody response in vivo. Some of the studies demonstrate that certain cell wall antigens and anti-cell wall antibodies may be the basis for developing specific and sensitive serologic tests for the diagnosis of candidiasis, particularly the disseminated form. In addition, recent studies have focused on the potential of antibodies against the cell wall protein determinants in protecting the host against infection. Hence, a better understanding of the humoral response triggered by the cell wall antigens of C. albicans may provide the basis for the development of (i) effective procedures for the serodiagnosis of disseminated candidiasis, and (ii) novel prophylactic (vaccination) and therapeutic strategies to control this type of infections.     

Role of the host-arbuscule interface in the VA mycorrhizal symbiosis: ultracytological studies of processes involved in phosphate and carbohydrate exchange

Summary The host-arbuscule interface does not vary greatly between different VAM associations of herbaceous and woody plants. The presence in the interfacial matrix of living fine arbuscule branches, where the host wall material is reduced or nonexistent and the endophyte wall is thin, of host membranous vesicles, polysaccharides. ATPase and neutral phosphatase activities, clearly indicates that this structure is not an inert space, but a physiologically complex zone. Possible roles for these interfacial matrix components in host-fungus nutrient exchange is discussed.     

The effects of parasite age and intensity on variability in acanthocephalan-induced behavioural manipulation

Numerous parasites with complex life cycles are able to manipulate the behaviour of their intermediate host in a way that increases their trophic transmission to the definitive host. Pomphorhynchus laevis, an acanthocephalan parasite, is known to reverse the phototactic behaviour of its amphipod intermediate host, Gammarus pulex, leading to an increased predation by fish hosts. However, levels of behavioural manipulation exhibited by naturally-infected gammarids are extremely variable, with some individuals being strongly manipulated whilst others are almost not affected by infection. To investigate parasite age and parasite intensity as potential sources of this variation, we carried out controlled experimental infections on gammarids using parasites from two different populations. We first determined that parasite intensity increased with exposure dose, but found no relationship between infection and host mortality. Repeated measures confirmed that the parasite alters host behaviour only when it reaches the cystacanth stage which is infective for the definitive host. They also revealed, we believe for the first time, that the older the cystacanth, the more it manipulates its host. The age of the parasite is therefore a major source of variation in parasite manipulation. The number of parasites within a host was also a source of variation. Manipulation was higher in hosts infected by two parasites than in singly infected ones, but above this intensity, manipulation did not increase. Since the development time of the parasite was also different according to parasite intensity (it was longer in doubly infected hosts than in singly infected ones, but did not increase more in multi-infected hosts), individual parasite fitness could depend on the compromise between development time and manipulation efficiency. Finally, the two parasite populations tested induced slightly different degrees of behavioural manipulation.     

Erosion of root epidermal cell walls by Rhizobium polysaccharide-degrading enzymes as related to primary host infection in the Rhizobium-legume symbiosis

A central event of the infection process in the Rhizobium-legume symbiosis is the modification of the host cell wall barrier to form a portal of entry large enough for bacterial penetration. Transmission electron microscopy (TEM) indicates that rhizobia enter the legume root hair through a completely eroded hole that is slightly larger than the bacterial cell and is presumably created by localized enzymatic hydrolysis of the host cell wall. In this study, we have used microscopy and enzymology to further clarify how rhizobia modify root epidermal cell walls to shed new light on the mechanism of primary host infection in the Rhizobium-legume symbiosis. Quantitative scanning electron microscopy indicated that the incidence of highly localized, partially eroded pits on legume root epidermal walls that follow the contour of the rhizobial cell was higher in host than in nonhost legume combinations, was inhibited by high nitrate supply, and was not induced by immobilized wild-type chitolipooligosaccharide Nod factors reversibly adsorbed to latex beads. TEM examination of these partially eroded, epidermal pits indicated that the amorphous, noncrystalline portions of the wall were disrupted, whereas the crystalline portions remained ultrastructurally intact. Further studies using phase-contrast and polarized light microscopy indicated that (i) the structural integrity of clover root hair walls is dependent on wall polymers that are valid substrates for cell-bound polysaccharide-degrading enzymes from rhizobia, (ii) the major site where these rhizobial enzymes can completely erode the root hair wall is highly localized at the isotropic, noncrystalline apex of the root hair tip, and (iii) the degradability of clover root hair walls by rhizobial polysaccharide-degrading enzymes is enhanced by modifications induced during growth in the presence of chitolipooligosaccharide Nod factors from wild-type clover rhizobia. The results suggest a complementary role of rhizobial cell-bound glycanases and chitolipooligosaccharides in creating the localized portals of entry for successful primary host infection.     

EnP1, a microsporidian spore wall protein that enables spores to adhere to and infect host cells in vitro

Microsporidia are spore-forming fungal pathogens that require the intracellular environment of host cells for propagation. We have shown that spores of the genus Encephalitozoon adhere to host cell surface glycosaminoglycans (GAGs) in vitro and that this adherence serves to modulate the infection process. In this study, a spore wall protein (EnP1; Encephalitozoon cuniculi ECU01_0820) from E. cuniculi and Encephalitozoon intestinalis is found to interact with the host cell surface. Analysis of the amino acid sequence reveals multiple heparin-binding motifs, which are known to interact with extracellular matrices. Both recombinant EnP1 protein and purified EnP1 antibody inhibit spore adherence, resulting in decreased host cell infection. Furthermore, when the N-terminal heparin-binding motif is deleted by site-directed mutagenesis, inhibition of adherence is ablated. Our transmission immunoelectron microscopy reveals that EnP1 is embedded in the microsporidial endospore and exospore and is found in high abundance in the polar sac/anchoring disk region, an area from which the everting polar tube is released. Finally, by using a host cell binding assay, EnP1 is shown to bind host cell surfaces but not to those that lack surface GAGs. Collectively, these data show that given its expression in both the endospore and the exospore, EnP1 is a microsporidian cell wall protein that may function both in a structural capacity and in modulating in vitro host cell adherence and infection.     

Helicoidal cell-wall texture in root hairs

It is shown that root hairs of most aquatic plants have a helicoidal cell-wall texture. Cell walls of root hairs of the aquatic/marshland plant Ranunculus lingua, however, have an axial microfibril alignment. The occurrence of a helicoidal wall texture is not limited to root hairs of aquatic plants: the terrestrial plant Zebrina purpusii has a helicoidal root-hair wall texture, too. With the exception of the grasses, the occurrence of root hairs with helicoidal cell walls pertains to species with predetermined root-hair-forming cells, trichoblasts. The rotation mode of the helicoid is species-specific. The average angle between fibrils of adjacent lamellae varies from 23° to 40°. In Hydrocharis morsus-ranae, cortical microtubules have a net-axial orientation and thus do not parallel nascent microfibrils. The deposition of the helicoidal cell wall is discussed.In honour of Prof. Dr. H.F Linskens (Nijmegen) on the occasion of his 65th birthday     

Observations with cytochemistry and ultracryotomy on the fine structure of the expanding walls in actively elongating plant cells.

Ultracryotomy with negative staining and cytochemistry (periodic acid - thiocarbohydrazide - silver proteinate test for polysaccharides, in conjunction with mild extractions) were used to study the architecture of the cell wall and its modifications during expansion. Those techniques were applied to the study in situ of the walls of actively elongating parenchyma of mung bean (Phaseolus aureus), and pea (Pisum sativum) root and of collenchyma of celery (Apium graveolens) petioles. These complementary techniques provide information on the 3-dimensional disposition and fine structure of the subunits of the wall. In all the examples examined, the bulk of growing primary wall appears well-ordered and no progressive evolution from a transverse texture near the plasmalemma to a scattered texture near the middle lamella was observed. It seems unlikely that the development of the wall structure in relation to growth could be explained mechanically by a passive shift of the fibrillar elements in response to cellular stress. There is no evidence for an inert change in fibrillar orientation in the major part of the wall. If such occurs the process is limited to the outermost and senescent part of the wall. Thus, the texture observed does not agree with the classical multinet growth hypothesis but rather with the idea of an ordered structure of the primary wall. With the latter, the components should be able to respond in different ways to specific growth regulators and other environmental signals and thus exert a more positive control over the processes of oriented cell growth.     

The Mode of Transmission of the Acephaline Gregarine Parasites of Earthworms

SYNOPSIS. Speculations regarding the mode of transmission of monocystid parasites of earthworms over a period of more than 100 years have never been tested experimentally under controlled conditions. In order to do so a stock of infectionfree Eisenia foetida (Sav.) was raised from cocoons and experimental infections were induced in this host using sporocysts of the gregarine parasites Apolocystis elongata Phillips & Mackinnon 1946, and Nematocystis elmassiani (Hesse, 1909). Experimental infections were obtained by feeding to uninfected worms sporocysts obtained directly from infected host worms. This proved that the intervention of a vector is not a necessary condition of infection. Infections could not be induced by injecting sporocysts through the body wall into the body cavity. Infections are thus probably acquired in nature by the ingestion of sporocysts. Sporocysts do not leave the body of the host by being passed from coelom to lumen of the gut, nor do they pass directly to the exterior through apertures of the body wall. There was no evidence of parasitic autotomy. It is therefore concluded that death and decay of the host is the normal method of dissemination of sporocysts. Sporocysts were not infective after drying in air for three weeks. Other sporocysts lost potency after storage in moist conditions for several months. Infections involving the organisms specified were sporadic and unpredictable; modifying factors, such as variations in host susceptibility and latency in infection, appeared to be operating.     

Chalara elegal-Infected Tobacco Roots: Ultrastructural Observations on in vitro Host-Fungus Relationships

Evidence, based on ultrastructural observations of stages involved in root infection oi Nicotiana tabacum cv. Xanthi n.c. in vitro by the black root rot fungus Chalara elegans, indicates that host cells from various layers react differently when challenged by the pathogenic fungus. All the host responses observed were associated with host cell wall modifications. Host reaction to fungal invasion occurring in the epidermal cells was limited to a disorganization of the cytoplasm. In the hypodermal cell layer, fibrillar cell wall outgrowths and wall thickenings were the earliest and the most obvious host reactions. In parenchymal cells, the host reacted by depositing papilla-like wall appositions directly adjacent to the infecting hyphae; with secondary infection of these cells, a densely staining material was laid down, mainly around the distal region of the infecting hyphae. In all these tissues, infection also led to disorganization of the host cytoplasm. Colonization of the endodermis did not lead to any rapid lethal modifications in either the host or the fungus, and a biotrophic-like state seemed to occur at this stage of the infection. No hyphal infection occurred in the central cylinder.     

Characterization of a Mycobacterium smegmatis gene that confers resistance to phages L5 and D29 when overexpressed

Bacteriophage infection requires a specific interaction with the outer surface of a bacterial host followed by interaction with the cell membrane and phage DNA injection. Phages of the mycobacteria encounter a cell wall that is rich in unusual lipid- and sugar-containing components which form a formidable barrier that must be passed to gain access to the membrane. We describe here a gene of Mycobacterium smegmatis that confers resistance to mycobacteriophages L5 and D29. The phage-resistance phenotype results not from mutation but from elevated expression of a wild-type gene. It appears that the product of this multicopy phage-resistance ( mpr ) gene may alter the structure of the host cell wall or membrane, thereby inhibiting productive phage DNA injection.