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Under-skin browning (USB) is an unsightly physiological disorder that afflicts ‘Honey Gold’ mango fruit. Under-skin browning symptoms develop after harvest upon the interaction of physical abrasion and physiological chilling stresses. Less understood preharvest and/or harvest factors may also influence fruit susceptibility to USB. In this study, we examined the impact of harvest time during the diurnal cycle and fruit sap components on USB development. Fruits were harvested at 4- to 6-h intervals, lightly abraded with sandpaper to simulate vibration damage during refrigerated road transport, held at 12 ± 1°C for 6 days, transported to the research facilities and ripened before USB assessment. Spurt and ooze sap from the fruit were collected at each harvest time. The samples were separated and analysed by gas chromatography–mass spectrometry. Fruit harvested at 10:00, 14:00 and 18:00 h had 3- to 5-fold higher incidence of USB than did those picked at 22:00, 2:00 and 6:00 h. Sap concentrations of the key aroma volatile compounds 2-carene, 3-carene, α-terpinene, p-cymene, limonene and α-terpinolene were higher for fruit harvested at 14:00 h compared to those picked at other times. In the fruits harvested in the afternoon, abraded skin treated with spurt sap sampled at 14:00 h had 14.3- and 29.0-fold higher incidence and severity, respectively, of induced browning than did those treated with sap collected at 6:00 h. The results showed that fruit harvested in the afternoon were more susceptible to USB than those picked at night or in early morning. The diurnal variation in fruit sensitivity was evidently associated with specific compositional differences in sap phytotoxicity. Topical application to the fruit skin of pure terpinolene and limonene resulted in induced USB damage, whereas pure carene and distilled water did not. Microscopy examination showed that while skin damage caused by pure terpinolene and limonene was not identical to USB per se, similarities suggested that sap components cause USB under inductive commercial conditions. Considered collectively, these findings suggest that night and early morning harvesting will reduce USB and thus improve the postharvest quality of Honey Gold mango fruit.

Four sea snakes (two Hydrophis major, one Hydrophis platurus, one Hydrophis elegans) were found washed ashore on different beaches in the Sunshine Coast region and Fraser Island in Queensland, Australia between 2007-2013. Each snake had multiple granulomas and locally extensive regions of pallor evident in the hypaxial and intercostal musculature along the body. Lesions in two individuals were also associated with vertebral and rib fractures. Histological examination revealed granulomas scattered throughout skeletal muscle, subcutaneous adipose tissue and fractured bone. These were composed of dense aggregates of microsporidian spores surrounded by a mantle of macrophages. Sequences (ssrRNA) were obtained from lesions in three sea snakes and all revealed 99% similarity with Heterosporis anguillarum from the Japanese eel (Anguillarum japonica). However, ultrastructural characteristics of the organism were not consistent with those of previous descriptions. Electron microscopic examination of skeletal muscle revealed large cysts (not xenomas) bound by walls of fibrillar material (Heterosporis-like sporophorocyst walls were not detected). The cysts contained numerous mature microsporidian spores arranged in small clusters, sometimes apparently within sporophorous vesicles. The microspores were monomorphic, oval and measured 2.5-3.0 μm by 1.6-1.8 μm. They contained isofilar polar filaments with 11 (infrequently 9-12) coils arranged in two ranks. This is the first published report of a microsporidian infection in hydrophiid sea snakes. This discovery shows microsporidia with molecular affinities to Heterosporis anguillarum but ultrastructural characters most consistent with the genus Pleistophora (but no hitherto described species). Further studies are required to determine whether the microsporidian presented here belongs to the genus Heterosporis, or to a polymorphic species group as suggested by the recognition of a robust Pleistophora/Heterosporis clade by molecular studies. The gross and histological pathology associated with these infections are described.

Members of phylum Planctomycetes have been proposed to possess atypical cell organisation for the Bacteria, having a structure of sectioned cells consistent with internal compartments surrounded by membranes. Here via electron tomography we confirm the presence of compartments in the planctomycete Gemmata obscuriglobus cells. Resulting 3-D models for the most prominent structures, nuclear body and riboplasm, demonstrate their entirely membrane - enclosed nature. Immunogold localization of the FtsK protein also supports the internal organisation of G.obscuriglobus cells and their unique mechanism of cell division. We discuss how these new data expand our knowledge on bacterial cell biology and suggest evolutionary consequences of the findings.

Recent reports of a novel group of flaviviruses that replicate only in mosquitoes and appear to spread through insect populations via vertical transmission have emerged from around the globe. To date, there is no information on the presence or prevalence of these insect-specific flaviviruses (ISFs) in Australian mosquito species. To assess whether such viruses occur locally, we used reverse transcription-polymerase chain reaction (RT-PCR) and flavivirus universal primers that are specific to the NS5 gene to detect these viruses in mosquito pools collected from the Northern Territory. Of 94 pools of mosquitoes, 13 were RT-PCR positive, and of these, 6 flavivirus isolates were obtained by inoculation of mosquito cell culture. Sequence analysis of the NS5 gene revealed that these isolates are genetically and phylogenetically similar to ISFs reported from other parts of the world. The entire coding region of one isolate (designated 56) was sequenced and shown to have approximately 63.7% nucleotide identity and 66.6% amino acid identity with its closest known relative (Nakiwogo virus) indicating that the prototype Australian ISF represents a new species. All isolates were obtained from Coquillettidia xanthogaster mosquitoes. The new virus is tentatively named Palm Creek virus (PCV) after its place of isolation. We also demonstrated that prior infection of cultured mosquito cells with PCV suppressed subsequent replication of the medically significant West Nile and Murray Valley encephalitis viruses by 10-43 fold (1 to 1.63 log) at 48 hr post-infection, suggesting that superinfection exclusion can occur between ISFs and vertebrate-infecting flaviviruses despite their high level of genetic diversity. We also generated several monoclonal antibodies (mAbs) that are specific to the NS1 protein of PCV, and these represent the first ISF-specific mAbs reported to date.

It was shown that nanoparticle–polymer interactions that affect the free volume and oxygen barrier properties of poly(m-xylene adipamide)/clay nanocomposites can be tailored by the choice of organic modifier of montmorillonite clay. Three different organo-modified clay compounds based on montmorillonite (Cloisite 30B, 10A and 93A) were dispersed in the resin poly(m-xylene adipamide) at loading levels of 2 wt% clay. Samples were melt compounded and extruded using a laboratory scale twin screw micro compounder. Positron annihilation lifetime spectroscopy (PALS) was used to examine the free volume of the polymer nanocomposites. PALS results suggested that the Cloisite 10A additive should give the higher reduction in gas permeability as it results in the lowest free volume for the nylon resin when compared to all of the clay additives examined. Oxygen transmission rates (OTR) were measured on nanocomposite films and the Cloisite 10A additive was found to give the best oxygen barrier, showing a reduction of OTR of 66% compared to the neat resin. In all cases examined, PALS free volume data was found to have excellent correlation to the measured oxygen transmission rates. The addition of Cloisite 10A resulted in the highest crystallinity and an increase in glass transition temperature when compared to the neat resin. Results indicate that the improved barrier properties of the clay compounds is primarily due to an increase in the degree of crystallinity of the polymer, with the nanoparticles being more effective nucleating agents when favourable nanoparticle–polymer interactions are present. [ABSTRACT FROM AUTHOR]

Genetic tags allow rapid localization of tagged proteins in cells and tissues. APEX, an ascorbate peroxidase, has proven to be one of the most versatile and robust genetic tags for ultrastructural localization by electron microscopy (EM). Here, we describe a simple method, APEX-Gold, which converts the diffuse oxidized diaminobenzidine reaction product of APEX into a silver/gold particle akin to that used for immunogold labelling. The method increases the signal-to-noise ratio for EM detection, providing unambiguous detection of the tagged protein, and creates a readily quantifiable particulate signal. We demonstrate the wide applicability of this method for detection of membrane proteins, cytoplasmic proteins, and cytoskeletal proteins. The method can be combined with different EM techniques including fast freezing and freeze substitution, focussed ion beam scanning EM, and electron tomography. Quantitation of expressed APEX-fusion proteins is achievable using membrane vesicles generated by a cell-free expression system. These membrane vesicles possess a defined quantum of signal, which can act as an internal standard for determination of the absolute density of expressed APEX-fusion proteins. Detection of fusion proteins expressed at low levels in cells from CRISPR-edited mice demonstrates the high sensitivity of the APEX-Gold method.

Planctomycetes are distinguished from other Bacteria by compartmentalization of cells via internal membranes, interpretation of which has been subject to recent debate regarding potential relations to Gram-negative cell structure. In our interpretation of the available data, the planctomycete Gemmata obscuriglobus contains a nuclear body compartment, and thus possesses a type of cell organization with parallels to the eukaryote nucleus. Here we show that pore-like structures occur in internal membranes of G.obscuriglobus and that they have elements structurally similar to eukaryote nuclear pores, including a basket, ring-spoke structure, and eight-fold rotational symmetry. Bioinformatic analysis of proteomic data reveals that some of the G. obscuriglobus proteins associated with pore-containing membranes possess structural domains found in eukaryote nuclear pore complexes. Moreover, immunogold labelling demonstrates localization of one such protein, containing a β-propeller domain, specifically to the G. obscuriglobus pore-like structures. Finding bacterial pores within internal cell membranes and with structural similarities to eukaryote nuclear pore complexes raises the dual possibilities of either hitherto undetected homology or stunning evolutionary convergence.

Interactions between plants and microbes in soil, the final frontier of ecology, determine the availability of nutrients to plants and thereby primary production of terrestrial ecosystems. Nutrient cycling in soils is considered a battle between autotrophs and heterotrophs in which the latter usually outcompete the former, although recent studies have questioned the unconditional reign of microbes on nutrient cycles and the plants' dependence on microbes for breakdown of organic matter. Here we present evidence indicative of a more active role of plants in nutrient cycling than currently considered. Using fluorescent-labeled non-pathogenic and non-symbiotic strains of a bacterium and a fungus (Escherichia coli and Saccharomyces cerevisiae, respectively), we demonstrate that microbes enter root cells and are subsequently digested to release nitrogen that is used in shoots. Extensive modifications of root cell walls, as substantiated by cell wall outgrowth and induction of genes encoding cell wall synthesizing, loosening and degrading enzymes, may facilitate the uptake of microbes into root cells. Our study provides further evidence that the autotrophy of plants has a heterotrophic constituent which could explain the presence of root-inhabiting microbes of unknown ecological function. Our discovery has implications for soil ecology and applications including future sustainable agriculture with efficient nutrient cycles.