Structure‐function analysis of npr1 alleles in Arabidopsis reveals a role for its paralogs in the perception of salicylic acid

Salicylic acid (SA) is necessary for plant defence against some pathogens, whereas NPR1 is necessary for SA perception. Plant defence can be induced to an extreme by several applications of benzothiadiazole (BTH), an analogue of SA. Thus, plants that do not perceive BTH grow unaffected, whereas wild‐type plants grow stunted. This feature allows us to screen for mutants in Arabidopsis thaliana that show insensitivity to BTH in a high‐throughput fashion. Most of the mutants are npr1 alleles, with similar phenotypes in plant weight and pathogen growth. The mutations are clustered in the carboxyl‐terminal part of the protein, and no obvious null alleles were recovered. These facts have prompted a search for knockouts in the NPR1 gene. Two of these KO alleles identified are null and have an intermediate phenotype. All the evidence presented lead us to propose a redundancy in SA perception, with the paralogs of NPR1 taking part in this signalling. We show that the mutations recovered in the screening genetically interact with the paralogs preventing their function in SA signalling.     

Cl‐ homeostasis in includer and excluder citrus rootstocks: transport mechanisms and identification of candidate genes

To reveal specific Cl^‐ transport activities in the symplastic pathway, uptake, long‐distance transport and distribution of Cl^‐ have been investigated in the citrus rootstocks Carrizo citrange (CC, Cl^‐ includer) and Cleopatra mandarin (CM, Cl^‐ excluder). Using an external concentration of 4.5 mm Cl^‐, both species actively transported Cl^‐ to levels that exceeded the critical requirement concentration by one and two orders of magnitude in the excluder and the includer rootstocks, respectively. Both CC and CM modulated Cl^‐ influx according to the availability of the nutrient as uptake capacity was induced by Cl^‐ starvation, but inhibited after Cl^‐ resupply. Net Cl^‐ uptake was higher in the includer CC, an observation that correlated with a lower root‐to‐shoot transport capacity in the excluder CM. The patterns of tissue Cl^‐ accumulation indicated that chloride exclusion in the salt‐tolerant rootstock CM was caused by a reduced net Cl^‐ loading into the root xylem. Genes CcCCC1, CcSLAH1 and CcICln1 putatively involved in the regulation of chloride transport were isolated and their expression analysed in response to both changes in the nutritional status of Cl^‐ and salt stress. The previously uncharacterized ICln gene exhibited a strong repression to Cl^‐ application in the excluder rootstock, suggesting a role in regulating Cl^‐ homeostasis in plants.     

Role of aquaporins in root water transport of ectomycorrhizal jack pine (Pinus banksiana) seedlings exposed to NaCl and fluoride

Effects of ectomycorrhizal (ECM) fungus Suillus tomentosus on water transport properties were studied in jack pine (Pinus banksiana) seedlings. The hydraulic conductivity of root cortical cells (L_pc) and of the whole root system (L_pr) in ECM plants was higher by twofold to fourfold compared with the non‐ECM seedlings. HgCl₂ had a greater inhibitory effect on L_pc in ECM compared with non‐ECM seedlings, suggesting that the mercury‐sensitive, aquaporin (AQP)‐mediated water transport was largely responsible for the differences in L_pc between the two groups of plants. L_pc was rapidly and drastically reduced by the 50 mm NaCl treatment. However, in ECM plants, the initial decline in L_pc was followed by a quick recovery to the pre‐treatment level, while the reduction of L_pc in non‐ECM seedlings progressed over time. Treatments with fluoride reduced L_pc by about twofold in non‐ECM seedlings and caused smaller reductions of L_pc in ECM plants. When either 2 mm KF or 2 mm NaF were added to the 50 mm NaCl treatment solution, the inhibitory effect of NaCl on L_pc was rapidly reversed in both groups of plants. The results suggest that AQP‐mediated water transport may be linked to the enhancement of salt stress resistance reported for ECM plants.     

Do individuals in better condition survive for longer? Field survival estimates according to male alternative reproductive tactics and sex

There is a gap in terms of the supposed survival differences recorded in the field according to individual condition. This is partly due to our inability to assess survival in the wild. Here we applied modern statistical techniques to field‐gathered data in two damselfly species whose males practice alternative reproductive tactics (ARTs) and whose indicators of condition in both sexes are known. In Paraphlebia zoe, there are two ART: a larger black‐winged (BW) male which defends mating territories and a smaller hyaline‐winged (HW) male that usually acts as a satellite. In this species, condition in both morphs is correlated with body size. In Calopteryx haemorrhoidalis, males follow tactics according to their condition with males in better condition practicing a territorial ART. In addition, in this species, condition correlates positively with wing pigmentation in both sexes. Our prediction for both species was that males practicing the territorial tactic will survive less longer than males using a nonterritorial tactic, and larger or more pigmented animals will survive for longer. In P. zoe, BW males survived less than females but did not differ from HW males, and not necessarily larger individuals survived for longer. In fact, size affected survival but only when group identity was analysed, showing a positive relationship in females and a slightly negative relationship in both male morphs. For C. haemorrhoidalis, survival was larger for more pigmented males and females, but size was not a good survival predictor. Our results partially confirm assumptions based on the maintenance of ARTs. Our results also indicate that female pigmentation, correlates with a fitness component – survival – as proposed by recent sexual selection ideas applied to females.     

Use of a crude extract or purified antigen from first‐instar cattle grubs,Hypoderma lineatum,for the detection of anti‐Hypoderma antibodies in free‐ranging cervids from southern Spain

During the 2003–2005 hunting seasons, a total of 120 Cervidae, including 39 red deer (Cervus elaphus hispanicus) and 81 fallow deer (Dama dama), were examined for subcutaneous myiasis. Animals were shot from January to June in southern Spain. Specific antibodies against Hypodermatinae (Diptera: Oestridae) were detected by indirect enzyme‐linked immunosorbent assay (iELISA) using a crude larval extract (CLE) and a purified antigen [hypodermin C (HC)] obtained from first instars of Hypoderma lineatum (De Villers) (Diptera: Oestridae). Hypoderma actaeon Brauer was the only species detected in this study, which represents the first confirmation of this species in fallow deer from Spain. The overall prevalence of animals presenting subcutaneous larvae (14.2%) was considerably lower than the prevalences determined by iELISA with CLE (43.3%) and HC (40.0%). Red deer showed a higher prevalence of Hypoderma than fallow deer. The concordance between larval examination during the hunting season and iELISA using both antigens was low, whereas the concordance between the CLE and HC ELISAs was good. Larval antigens obtained from H. lineatum constitute a good tool for the diagnosis of H. actaeon in Cervidae, especially when the hunting season does not coincide with the maximum presence of larvae on the back.     

Enzyme inhibition by iminosugars: Analysis and insight into the glycosidase–iminosugar dependency of pH

Imino- and azasugar glycosidase inhibitors display pH dependant inhibition reflecting that both the inhibitor and the enzyme active site have groups that change protonation state with pH. With the enzyme having two acidic groups and the inhibitor one basic group, enzyme–inhibitor complexes with three (EH₃I), two (EH₂I), one (EHI), or no protons (EI), are possible. In the present work an analysis method is presented that from pH-inhibition data allows one to distinguish between the different complexes and determine which protonation state is preferred. It is also possible to determine the pH-independent binding constants of the inhibitor. Analysis of pH data for imino- and azasugar inhibition of β-glucosidases revealed that basic glycosidase inhibitors bind as the monoprotonated (EHI) complex. Three neutral inhibitors were also studied and two of these were also bound exclusively as the EHI complex while a third bound both as a EHI and a EH₂I complex.     

Non-Polar Chemical Constituents of Atemoya and Evaluation of the Cytotoxic and Antimicrobial Activity

This study aimed to identify the chemical composition of essential oil from fruits (EOAF) and the hexanic crude extract from aerial parts (At-Hex) of atemoya (Annona cherimola x Annona squamosa), a hybrid belonging to the Annonaceae family. Cytotoxic and antimicrobial activity was also evaluated. OEAF was obtained by hydrodistillation using a Clevenger apparatus, and their composition was analyzed by gas chromatography-mass spectrometry (GC-MS) analyses. Cytotoxicity was tested against human tumor cell lines HCT-116 (colon carcinoma), SF-295 (glioblastoma), OVCAR-8 (ovarian carcinoma) and HL60 (leukemia) using 3-(4,5-dimethyl-2-thiazolyl)-2,5- diphenyl-2H-tetrazolium bromide (MTT) assay, while antimicrobial activity was conducted by bioauthography method against eleven microorganisms strains. Twenty-four compounds were identified in the EOAF and twenty-nine in At-Hex. The monoterpenes linalool (25.70%), α-pinene (10.38%), β-pinene (9.12%), transocimene (7.43%), and the sesquiterpene bicyclogermacrene (12.58%) were the major constituents of EOAF, whereas the sesquiterpene spathulenol (13.91%) was the main compound of At-Hex. At-Hex showed a high cytotoxicity against SF-295 (glioblastoma). These findings show an important chemotaxonomic contribution for the Annonaceae family, mainly for the Annona genus. Atemoya proved to be a promising source of substances with potential cytotoxic activity.