Detection of a natural point mutation in Potato virus A that overcomes resistance to vascular movement in Nicandra physaloides., and studies on seed-transmissibility of the mutant virus

Isolate M of Potato virus A (PVA‐M; genus Potyvirus) is avirulent in Nicandra physaloides L. (family Solanaceae). The inoculated leaves are infected but no systemic infection is observed. Forty plants of ‘Black Pod’ (BP) and ‘Black Pod Alba’ (BPA), two variants of N. physaloides described in this study, were inoculated with PVA‐M. Two plants of BP and one plant of BPA were systemically infected. Mosaic, blistering and dark green islands developed on the systemically infected leaves, and flowers showed colour‐break symptoms. PVAprogeny were sequence‐characterised for the 6K2 protein and viral genome‐linked protein (VPg) encoding regions known to control the long distance movement of PVA in N. physaloides. All virus progeny (designated as PVA‐Mm) in the systemically infected leaves of the plants inoculated with PVA‐M contained only a single amino acid substitution (Vail 16Met) in the central part of VPg due to a nucleotide substitution G6033A, as compared to PVA‐M. Other PVA isolates that infected N. physaloides systemically also contained Metll6 in VPg. In a previous study using chimeric viruses, Metl16 in VPg was shown to be a major determinant for vascular movement of PVA in N. physaloides, and this study reveals that the mutation for Metl16 can occur in vivo during replication of the avirulent PVA‐M in infected plants. Immunolocalisation studies on BP and BPA plants showed that the pods (berries) and seed coat contained PVA‐Mm in the developing seeds, but no virus was detected in embryons. Up to 27% of the mature seeds contained PVA‐Mm but no transmission to seedlings was observed in a total of 450 seeds tested, and no test plants were infected following mechanical inoculation with extracts prepared from the seeds.     

ULTRASTRUCTURE AND ECOLOGY OF AUREOCOCCUS ANOPHAGEFERENS GEN. ET SP. NOV. (CHRYSOPHYCEAE): THE DOMINANT PICOPLANKTER DURING A BLOOM IN NARRAGANSETT BAY,RHODE ISLAND,SUMMER 19851

Observations of a marked cessation of feeding in filter feeding animals maintained in flowing Narragansett Bay seawater in June 1985 drew our attention to a bloom of a golden alga 2 μm in diameter at unprecedented populations of 10⁹ cells. L^?1. This picoplankter lacked morphological features useful in discriminating it from other similar sized forms with either phase contrast or epifluorescence light microscopy. Natural populations of picoplankton, obtained from the height of the bloom until its decline, were examined in thin section with transmission electron microscopy. A cell with a single chloroplast, nucleus, and mitochondrion and an unusual exocellular polysaccharide-like layer was apparently the bloom alga. The ultrastructure of this alga is consistent with that of the Chrysophyceae, and a new genus and species, Aureococcus anophagefferens is described. Attempts to grow this previously unrecognized picoplanktonic alga as an obligate phototroph failed and only yielded cultures of other previously described picoalgae. Facultative and obligate phagotrophic protists with ingested cells of Aureococcus were only observed as the bloom waned and minute diatoms became common. Cells of A. anophagefferens with virus particles typical for picoalgae occurred throughout the bloom. Populations of the usually dominant photosynthetic picoplankter, the cyanobacterium Synechococcus Nägeli, were depressed during the bloom. This could be due in part to selective grazing on Synechococcus rather than Aureococcus by elevated populations of Calycomonas ovalis Wulff which accompanied the algal bloom.     

Sri Lankan passion fruit mottle virus, a potyvirus infecting golden passion fruit in Sri Lanka

A sap-transmissible virus, provisionally named Sri Lankan passion fruit mottle virus (SLPFMV), was isolated from Passiflora edulis f. flavicarpa and shown to induce leaf mottling and distortion in that host. The virus infected 23 species in five plant families with systemic infection being common in the Passifloraceae. Chenopodium amaranticolor was a good local lesion host and Passiflora foetida was a useful systemic host for purification. In P. foetida extracts, SLPFMV lost infectivity after 10 min between 70–75°C, 6–7 days at 20–23°C and at dilutions of 10^--⁵ -W^-6. The virus had flexuous, filamentous particles with a normal length of c. 841 nm. Two polypeptides of mol. wt c. 33 200 and 28 700 were detected in purified virus preparations, and a major species of double-stranded RNA (mol. wt 7.0 × 10⁶), was detected in infected plants. Pinwheels, tubular and laminated inclusions were found in ultrathin sections of infected P. edulis f. plavicurpa and cylindrical inclusions were observed in epidermal strips. SLPFMV was transmitted by the aphids Myzus persicae, Aphis spiraecola, A. gossypü and A. cruccivora after brief acquisition feeds. SLPFMV reacted with antisera to several potyviruses including passion fruit woodiness virus, passion fruit ringspot virus, potato virus Y and watermelon mosaic virus 2 and thus, apparently, is a member of the potyvirus group.     

Bilayer stabilizing peptides and the inhibition of viral infection: antimeasles activity of carbobenzoxy-Ser-Leu-amide

A number of carbobenzoxy-dipeptide-amides raise the bilayer to hexagonal phase transition temperature of dielaidoylphosphatidylethanolamine (stabilizes the bilayer). The potency of the peptides in stabilizing the bilayer phase is Z-Tyr-Leu-NH₂= Z-Gly-Phe-NH₂>Z-Ser-Leu-NH₂>Z-Gly-Leu-NH₂>Z-Gly-Gly-NH₂. A linear correlation was found between the respective HPLC retention time parameterk for the peptide and the slope of the bilayer stabilization curve determined with model membranes by differential scanning calorimetry. One dipeptide, Z-Ser-Leu-NH₂, reduces measles virus cytopathic effect (CPE) in Vero cells. The mechanism by which this peptide reduces the CPE is not known, although some peptides which raise the bilayer to hexagonal phase transition temperature of phospholipids inhibit membrane fusion.Abbreviations Z carbobenzoxy - DEPE dielaidoylphosphatidylethanolamine - DSC differential scanning calorimetry - HPLC high pressure liquid chromatography - CPE cytopathic effect To whom correspondence should be addressed.     

Detection of CaMV gene I and gene VI protein products in vivo using antisera raised to COOH-terminal β-galactosidase fusion proteins

Specific antisera were prepared to the inclusion body protein (gene VI product) and the gene I product of cauliflower mosaic virus (CaMV). Translational fusions between the lacZ gene and gene VI or gene I were constructed by cloning the relevant DNA fragments into the expression vectors pUR290, pUR291 or pUR292. Large amounts of fusion protein were synthesized when the inserted DNA fragment was in frame with the lacZ gene of the expression vector. These fusion proteins were used to raise specific antisera to gene VI and gene I proteins of CaMV. Antiserum to the gene VI product detected a range of proteins in crude extracts and in a subcellular fraction enriched for virus inclusion bodies. This range of proteins was further shown to be related to gene VI by Staphylococcus aureus V8 partial proteolysis. Antiserum to the gene I product detected viral specific proteins of 46, 42 and 38 K in preparations of CaMV replication complexes from infected plants but not in any other subcellular fraction.     

Gene expression in Brassica campestris showing a hypersensitive response to the incompatible pathogen Xanthomonas campestris pv. vitians

Xanthomonas campestris pv. vitians, a pathogen of lettuce, elicits a hypersensitive response within 12 hours of inoculation into Brassica leaves, characterized by tissue collapse, loss of membrane integrity, vein blockage and melanin production. In contrast, the compatible pathogen, X. c. pv. campestris, has no visible effects on leaves for 48 hours, after which inoculated areas show chlorosis which eventually spreads, followed by rotting.mRNA was prepared from leaves inoculated with suspensions of both pathovars or with sterile medium up to 24 hours following inoculation. In vitro translation of total and poly A⁺ RNA in rabbit reticulocyte lysate in the presence of ³⁵S methionine followed by separation of the polypeptide products by 2D-PAGE, allowed comparison of the effects of these treatments on plant gene expression. Major changes in gene expression were observed as a consequence of the inoculation technique. In addition, after inoculation with X. c. vitians, up to fifteen additional major polypeptides appeared or greatly increased by four hours. Some of these had disappeared by nine hours and several more had appeared. No major polypeptides disappeared or decreased greatly in intensity following inoculation with X. c. vitians.     

DNase I hypersensitivity and expression of the Shrunken-1 gene of maize

The local chromatin structure of the Shrunken-1 (Sh) gene of maize was probed by analyzing DNase I hypersensitivity. Sh encodes the gene for sucrose synthetase, a major starch biosynthetic enzyme, which is maximally expressed in the endosperm during seed maturation. In addition to general DNase I sensitivity, specific DNase I hypersensitive sites were identified in endosperm chromatin that mapped near the 5 end of the Sh gene. The pattern of hypersensitive sites and their relative sensitivity were altered in other non-dormant tissues that produce little or no enzyme. However, some changes in chromatin structure appear to be independent of Sh gene expression and may reflect general alterations associated with plant development. The chromatin structure of several sh mutations, induced by Ds controlling element insertions, was also analyzed. Although the insertions perturbed expression of the gene, there were no notable effects on local chromatin structure.