Probing the herbivore’s responses to plant defenses using plant mediated RNAi

Nicotiana attenuata (Solanaceae) produces 17-hydroxygeranyllinalool diterpene glycosides (HGL-DTGs) in starch-equivalent concentrations that thwart the growth of its specialist lepidopteran herbivore Manduca sexta (Lepidoptera, Sphingidae). Lyciumoside IV and its malonylated forms, nicotianoside I and II, constitute ~80% of N. attenuata’s HGL-DTG pool. Upon ingestion, the malonylated forms are rapidly and non-enzymatically demalonylated to lyciumoside IV by the alkalinity of larval oral secretion. Ingested lyciumoside IV (44%) is excreted as a novel compound, 3-O-α-L-rhamnopyranosyl-(1-4)-β-D-glucopyranosyl-17-hydroxygeranyllinalool (RGHGL). It differs from lyciumoside IV only in its lack of the C-17-glucose. Of M. sexta's midgut-expressed β-glucosidases, only β-glucosidase1 is upregulated upon lyciumoside IV ingestion, and when silenced by plant-mediated RNAi (PMRi), larvae are impaired in lyciumoside IV deglucosylation and showed molting impairments and higher mortality than control larvae. To examine the consequences of this detoxification process on natural tritrophic interactions, β-glucosidase1-silencing PMRi plants were planted into native habitats; control and β-glucosidase1-silenced larvae’s survival was similar; however, while Camptocosa parallela spiders captured and killed the control and lyciumoside IV-replete β-glucosidase1-silenced larvae similarly, the spiders ingested only 25% of β-glucosidase1-silenced larvae and ingestion resulted in locomotor distress in the spiders. While spiders attacked and ingested RGHGL-coated or -ingested larvae equally, they were deterred by the lyciumoside IV-coated larvae. Although lyciumoside IV deters spiders, it is not defensively co-opted by M. sexta larvae, perhaps to avoid its deleterious effects such as molting impairment and mortality. We further show that these PMRi plants also silence the homologous genes in native M. quinquemaculata larvae, feeding on these transgenic plants in nature

Detoxification of hostplant’s chemical defense rather than its anti-predator co-option drives beta-glucosidase-mediated lepidopteran counter-adaptation

The evolutionary plant–herbivore arms race sometimes gives rise to remarkably unique adaptation strategies. Here we report one such strategy in the lepidopteran herbivore Manduca sexta against its hostplant Nicotiana attenuata's major phytotoxins, 17-hydroxygeranyllinalool diterpene glycoside, lyciumoside IV and its malonylated forms. We show that alkalinity of larval regurgitant non-enzymatically demalonylates the malonylated forms to lyciumoside IV. Lyciumoside IV is then detoxified in the midgut by β-glucosidase 1-catalysed deglycosylation, which is unusual, as typically the deglycosylation of glycosylated phytochemicals by insects results in the opposite: toxin activation. Suppression of deglucosylation by silencing larval β-glucosidase 1 by plant-mediated RNAi causes moulting impairments and mortality. In the native habitat of N. attenuata, β-glucosidase 1 silencing also increases larval unpalatability to native predatory spiders, suggesting that the defensive co-option of lyciumoside IV may be ecologically advantageous. We infer that M. sexta detoxifies this allelochemical to avoid its deleterious effects, rather than co-opting it against predators

Feasibility of parental collected nasal swabs for virus detection in young children with cystic fibrosis

BACKGROUND: The detrimental role of viruses has been well described in CF, although the pattern of virus infections has not been investigated in a longitudinal study. The primary aim was to determine the feasibility of fortnightly parent collected swabs in young children with CF. METHODS: Children under three years with CF were recruited. Nasal swabs were collected by parents every fortnight and during periods of symptoms over 12 months. Nasal swabs were posted and virus detected using real-time PCR. RESULTS: Only 27% of the patients completed the study to 10 months, although 98% of the swabs returned were adequate for analysis. Mould was observed growing on 23% of the returned swabs. There was no evidence to demonstrate relationships with symptoms and viruses, prolonged symptoms, prolonged shedding or patterns of virus infections. CONCLUSIONS: This study highlights the need to further investigate the role of viruses in children with CF using a robust method of frequent collection in children for a longitudinal study, with appropriate storage and shipping techniques to avoid mould growth or other potential contaminants

Type 1 Phototherapeutic Agents. 2. Cancer Cell Viability and ESR Studies of Tricyclic Diarylamines

Type 1 phototherapeutic agents based on diarylamines were assessed for free radical generation and evaluated in vitro for cell death efficacy in the U937 leukemia cancer cell line. All of the compounds were found to produce copious free radicals upon photoexcitation with UV-A and/or UV–B light, as determined by electron spin resonance (ESR) spectroscopy. Among the diarylamines, the most potent compounds were acridan (<b>4</b>) and 9-phenylacridan (<b>5</b>), with IC₅₀ values of 0.68 μM and 0.17 μM, respectively

Novel intramolecular reductive Carbon-carbon coupling during borohydride reduction of dispiro[naphthalene-1,2′2^{\prime}(1′1^{\prime}H)-naphtho[2,1-b]pyran-3′3^{\prime},1^{\prime}^{\prime}-naphthalene]-2(1H),2^{\prime}^{\prime}(1^{\prime}^{\prime}H)-dione and similar dispironaphthalenones

$NaBH_4$ reduction of the title compounds I $(R=H, Br, Me_3C, 1-methyl-1-cyclohexyl)$ gave the coupling products II, III and IV $(R1=Me, Et, Me_2CH)$

Roles of Free Radicals in Type 1 Phototherapeutic Agents: Aromatic Amines, Sulfenamides, and Sulfenates

Detailed analyses of the electron spin resonance (ESR) spectra, cell viability, and DNA degradation studies are presented for the photolyzed Type I phototherapeutic agents: aromatic amines, sulfenamides, and sulfenates. The ESR studies provided evidence that copious free radicals can be generated from these N–H, N–S, and S–O containing compounds upon photoirradiation with UV/visible light. The analyses of spectral data allowed us to identify the free radical species. The cell viability studies showed that these agents after exposure to light exert cytotoxicity to kill cancer cells (U937 leukemia cell lines HTC11, KB, and HT29 cell lines) in a dosage- and time-dependent manner. We examined a possible pathway of cell death via DNA degradation by a plasmid cleavage assay for several compounds. The effects of photosensitization with benzophenone in the presence of oxygen were examined. The studies indicate that planar tricyclic amines and sulfenamides tend to form π-electron delocalized aminyl radicals, whereas nonplanar ones tend to yield nitroxide radicals resulting from the recombination of aminyl radicals with oxygen. The ESR studies coupled with the results of cell viability measurements and DNA degradation reveal that planar N-centered radicals can provide higher potency in cell death and allow us to provide some insights on the reaction mechanisms. We also found the formation of azatropylium cations possessing high aromaticity derived from azepines can facilitate secondary electron transfer to form toxic O₂^•– radicals, which can further exert oxidative stress and cause cell death

Early detection of lung function abnormalities in young children with cystic fibrosis

The onset of lung disease in cystic fibrosis (CF) begins early in life with respiratory infection, inflammation, and structural lung damage all reported in infants with CF in the first months of life. As new treatments become available, it is essential that we have outcome measures that can be used to track disease progression and treatment efficacy. In this review, we have examined the role of lung function testing in infants and preschool children with CF. In particular, we have focused on the ability of the various lung function tests to detect the presence of respiratory pathogens and structural lung disease, increased inflammation, and the onset of acute exacerbations