Could indigenous arbuscular mycorrhizal communities be used to improve tolerance of pistachio to salinity and/or drought? Abstract Arbuscular mycorrhizal fungi (AMF) establish symbiosis with a number of plant species, including pistachio, under a broad range of soil conditions. Here we present a fully factorial pot experiment where four different indigenous AMF communities (two with high- and two with low-salinity legacies) were tested for improving pistachio seedling tolerance to salinity and/or drought in terms of maintaining its growth and/or mineral (phosphorus and zinc) nutrition under experimentally applied abiotic stress. The AMF communities were pre-cultured with a grass to reduce the load of pistachio pathogens before application to the seedlings. We detected systematic improvement of pistachio mineral nutrition by the AMF communities, whereas salinity and drought generally suppressed pistachio performance independently of each other. The AMF communities from saline soils systematically improved pistachio tolerance to salinity in terms of maintaining higher phosphorus acquisition of their host, whereas their effect on maintaining plant zinc nutrition under high salinity was less prominent. The effects of the different AMF communities on pistachio tolerance to drought were variable and without obvious link to the environmental legacy of the AMF. One of the AMF communities from saline soil (dominated by Rhizophagus) showed particular tolerance to salinity in terms of maintaining high colonization rates of pistachio roots. Interestingly, one of the AMF communities from low-salinity soils conferred a significant tolerance of pistachio to salinity in terms of maintaining its phosphorus acquisition upon the stress. This means that beneficial traits of plant symbiotic microbes may occur uncoupled from the selective pressure in their native range.

Endophytic bacteria mitigate mercury toxicity to host plants Abstract Plant communities growing in metal-contaminated areas can develop resistance mechanisms by establishing symbiotic associations with endophytic microorganisms. The functionality and diversity of endophytic communities depend on the amount and type of metal present in the soil. To characterise the response of endophytic bacterial communities to mercury-induced abiotic stress, we analysed the colonization frequency and number of bacterial isolates in the roots of Aeschynomene fluminensis (Joint Vetch) and Polygonum acuminatum (Smartweed), which represent the families Fabaceae and Polygonaceae, respectively. These two plant species are found in many mercury-contaminated areas. The isolates were characterised by morpho- and genotyping and identified by 16S rDNA gene sequencing. The bacteria belonged to the phyla Actinobacteria, Bacteriodetes, Firmicutes, and Proteobacteria. The Hill series and Venn diagram provided evidence that mercury affects the composition, diversity, and richness of the endophytic bacterial communities. Inoculation with Bacillus_sp_BacI34, Burkholderia_sp_BacI45, Enterobacter_sp_BacI14, Enterobacter_sp_BacI26, Enterobacter_sp_BacI18, Klebsiella_pneumoniae_BacI20, Lysobacter_soli_BacI39, Pantoea_sp_BacI16, and Pantoea_sp_BacI23 promoted the growth of corn (Zea mays) plants in mercury-supplemented substrata. It is noteworthy that Pantoea sp_BacI23 increased the host plant length (root and shoot) by 117.09 ± 0.28%. Endophytic bacterial strains may well provide important inoculants for plant growth promotion on metal-contaminated sites and in metal bioremediation programs.

The pioneer lichen Placopsis in maritime Antarctica: Genetic diversity of their mycobionts and green algal symbionts, and their correlation with deglaciation time Abstract Since ice-free areas in Antarctica are predicted to increase by up to 25% before the end of this century, lichens such as the genus Placopsis will be important colonizers of these newly available grounds and will still be present in later successional stages of the lichen community. The main symbionts of Placopsis species are examined for 56 specimens collected from the South Shetland Islands, Antarctica using molecular (fungal and algal nrITS, fungal RPB1, algal rbcL sequences) and morphological methods. The specimens were collected from soils with different deglaciation times. Eight uni-algal photobiont cultures were obtained and analysed from two specimens. Placopsis antarctica and P. contortuplicata proved to be monophyletic and are sister species, only the former producing vegetative diaspores (soredia). Both share the same photobiont pool and are lichenized with two closely related species, Stichococcus antarcticus and S. allas. Two haplotypes of S. antarcticus are restricted to areas deglaciated for more than 5000 years and the volcanic Deception Island indicating a shift in the photobionts of Placopsis in the course of the soil and lichen community development. These photobiont haplotypes exhibit different ecological preferences, possibly leading to adaptation of the symbiotic entity to changing environmental conditions.

Genetic diversity and host relationships of endosymbiotic bacteria in the Asian cryptic species of Bemisia tabaci from Bangladesh Abstract Endosymbiotic bacteria are common in many herbivorous insects. Bemisia tabaci is a phloem-sapping pest of various crop plants and is known to harbor at least five endosymbionts. This species is a complex of at least 40 genetically distinct but morphologically indistinguishable cryptic species worldwide. Endosymbiont composition has been studied in invasive cryptic species such as MEAM1 and MED, but little information exists regarding the indigenous genetic groups in Asia. Here, we determined the endosymbiont profiles of four indigenous Asian cryptic species (Asia I, Asia II 1, Asia II 5 and Asia II 10) of B. tabaci identified in Bangladesh. Overall, the infection rates of Arsenophonus, Cardinium, Hamiltonella, Rickettsia, and Wolbachia were 93%, 86%, 0%, 31%, and 88%, respectively. Phylogenetic analysis revealed two subgroups in Arsenophonus (A1, A2) and Rickettsia (R1, R2), but only one subgroup in Cardinium (C2) and Wolbachia (W1). Each endosymbiont showed varying rates of infection in the four cryptic species and most were co-infected with various combinations. The results of this study provide important information on the relationships between the endosymbionts and cryptic species of B. tabaci indigenous to Asia.

Molecular and biochemical changes of aging-induced nodules senescence in common bean Abstract In the common bean (Phaseolus vulgaris L.), premature nodule senescence affects biological nitrogen fixation (BNF), reducing crop yield. Here, we investigated the molecular and biochemical changes in common bean nodules undergoing aging senescence. The experiments were carried out with three common bean genotypes and two nodule groups: senescent and non-senescent nodules. The expression patterns of 12 genes were assessed using RT-qPCR. In addition, global activities of ascorbate peroxidase (APX) and catalase (CAT), lipid peroxidation and total sugar content of nodules were also determined. Our results show that PvLb gene expression was down-regulated and PvGS(n-1) and PvUriII, genes involved in N metabolism, also decreased in senescent nodules. Transcripts related to the ethylene, abscisic acid and cytokinin phytohormones were up-regulated upon nodule senescence. Interestingly, PvSnakin-2 (SNAKIN-like cysteine rich protein), a gene related to plant-pathogen interaction, was induced in senescent nodules. The biochemical assays showed that in nodules undergoing senescence, APX activity was altered in only one genotype but CAT activity decreased in all common bean genotypes. Lipid peroxidation increased but total sugar content was not altered in nodules upon senescence. All these findings suggest that a delayed plant-response to Rhizobium infection is activated in common bean nodules undergoing aging senescence and this mechanism seems to be regulated by phytohormones and cell signaling as well, whereas the antioxidant defenses mediated by CAT are repressed, suggesting oxidative damage in nodules upon senescence. Our results offer insights into understanding nodule senescence metabolism and provide potential senescence markers for common bean nodules.

Do silver nanoparticles stimulate the formation of ectomycorrhizae in seedlings of pedunculate oak ( Quercus robur L.)? Abstract Metal nanoparticles are gaining ever-wider application in agriculture and forestry, as alternatives to chemical agents used as fertilisers, growth stimulators and pesticides, establishing a need for eco-toxicological risk assessment of these agents. We tested the effects of foliar-applied silver nanoparticles (AgNPs) on chlorophyll a fluorescence and on abundance and species composition of ectomycorrhizal (ECM) colonisation. The application of AgNPs at concentrations of 5, 25 and 50 ppm was found to stimulate the formation of mycorrhizae in seedlings of pedunculate oak, with the highest effect at intermediate concentrations (25 ppm). There were non-linear effects on the relative abundance of ECM fungal species. The proportion of dominant T. terrestris was highest in the control group, whereas the shares of ECM formed by the two other species, S. brunnea and P. involutus, were higher in the treatments with intermediate and maximal concentrations of AgNPs, respectively. Maximum quantum yield of photosystem II (Fv/fm) assessed by chlorophyll a fluorescence measurements revealed slight debilitation of oak seedlings irrespective of the application of AgNPs and their concentrations. This result offered an indirect indication that photosynthesis capacity had no influence on the level of mycorrhization. We hypothesise that foliar AgNPs treatments at concentrations below thresholds of acute toxicity and in the absence of significant effects on chlorophyll a fluorescence may still exert significant influence on biotic interactions including mycorrhizal symbioses by impacting plant hormonal balance, particularly ethylene, and regulatory pathways involved in host control of ECM colonisation.

Comparative study of secondary metabolites and bioactive properties of the lichen Cladonia foliacea with and without the lichenicolous fungus Heterocephalacria bachmannii Abstract The phenolic, flavonoid, tannin and proanthocyanidin content of the lichen Cladonia foliacea with and without its lichenicolous fungus Heterocephalacria (Syzygospora) bachmannii was investigated. The phenolic compounds were quantified in organic extracts using ultrasonic extraction (acetone and methanol) and in milled material (the ground material diluted with microcrystalline cellulose). The total phenolic content depended on the solvent polarity, the extraction technique and the species. The results demonstrated that the highest total phenolic content was recorded in untreated milled material (935.75 μg GAE/g DW) of H. bachmannii plus C. foliacea, followed by C. foliacea (668.29 μg GAE/g DW). The antioxidant activities were evaluated by the in vitro scavenging capacity, iron reducing power, and iron chelating power. The results showed that the highest scavenging capacity were obtained in methanol extracts of C. foliacea with IC50 = 0.015 mg/mL, followed by methanolic extract of H. bachmannii plus C. foliacea that had a scavenging capacity and iron reducing power of (IC50 = 0.030 mg/mL and IC50 = 0.054 mg/mL, respectively). The milled material showed the highest iron chelating power (IC50 = 0.279 mg/mL). We conclude that Cladonia foliacea when parasitized by H. bachmannii possesses a high antioxidant potential in the methanolic extract. Acetone and methanol extracts, showed that extracts from lichen plus lichenicolous fungus contained different and possibly more effective bioactive molecules than the lichen alone. These included phenolic acids, alkanes and aromatic compounds. This is the first study to investigate the phenolic content and antioxidant capacity of a lichenicolous fungus, albeit based on differences between the lichen with and without the mycoparasite H. bachmannii .

Role of putrescine (Put) in imparting salt tolerance through modulation of put metabolism, mycorrhizal and rhizobial symbioses in Cajanus cajan (L.) Millsp. Abstract Salt stress is a major environmental constraint that limits growth and nitrogen-fixation in legumes. Role of putrescine (Put) and arbuscular mycorrhiza (AM) in improving functional efficiency of legumes has gained importance in recent years. Present investigations assessed the impact of Put (1 mM) seed priming and/or Rhizophagus irregularis inoculation on growth, mycorrhizal and rhizobial symbioses with an emphasis to correlate the same with nodular Put metabolism in pigeonpea (Cajanus cajan L.) genotypes (Tolerant-Pusa 2001 and Sensitive-Pusa 991) under salt stress. Salinity declined plant biomass, with greater negative effects on roots than shoots which reduced the mycorrhizal colonization as well as nodulation ability of Sinorhizobium fredii AR-4. The decline in nitrogen-fixing efficiency could be correlated with increase in Na+ concentrations in roots as well as nodules. Salinity reduced endogenous Put by increasing diamine oxidase (DAO) as well as decreasing arginine decarboxylase (ADC) as well as ornithine decarboxylase (ODC) activities in nodules. Put priming enhanced per cent mycorrhizal colonization which further increased the rhizobial symbiotic efficiency, more in Pusa 2001 than Pusa 991. Both Put priming and AM inoculations reduced Na+ uptake and improved nutrient status especially P in both underground organs, with AM more effective than Put. The further decline in Na+ uptake was recorded when both amendments were given together which enhanced nitrogen-fixing ability of nodules by modulating anabolic and catabolic enzyme activities responsible for Put biosynthesis. Hence, +Put+AM can be used as an effective strategy to improve symbiotic potential and arrest nodule senescence in pigeonpea under salt stress.

Interactive effects of Epichloë fungal and host origins on the seed germination of Achnatherum inebrians Abstract Cool-season grasses have developed a symbiotic relationship with Epichloë endophytes. In many environments, Epichloë endophytes have been shown to be mutualistic symbionts of plants by increasing the fitness of their host against abiotic or biotic stresses. The effects of Epichloë endophytes on other fitness-correlated plant characteristics are less intensively studied, and the results are usually variable and contradictory. In this study, we evaluated the effects of endophyte infection on seed germination in Achnatherum inebrians from four origins. Our results indicate that the germination rate of the seeds collected from alpine regions was higher at low temperatures than that of seeds with desert and arid grassland origins. By contrast, a higher germination percentage was detected in seeds with desert and arid grassland origins than in those with alpine origins in higher temperatures. Epichloë endophyte infection affects the cardinal temperatures of seeds from different origins. Endophyte-infected seeds have a lower base temperature and a higher ceiling temperature than their endophyte-free counterparts. The value of the base temperature was higher in seeds with alpine grassland origins than in those with desert and arid grassland origins. However, the ceiling temperature was higher in seeds with desert and arid grassland origins than in those with alpine grassland origins. Consequently, future experiments should consider the effects of endophytes on seed germination and seedling recruitment in suboptimal climatic conditions.

Diversity and structure of the bacterial microbiome of the American dog tick, Dermacentor variabilis , is dominated by the endosymbiont Francisella Abstract The American dog tick, Dermacentor variabilis Say, is the principal vector of spotted fever group rickettsiae that cause illness in people. Because the composition and structure of microbial communities in D. variabilis are poorly understood, we conducted high-throughput amplicon sequencing of the 16S rRNA gene to characterize the bacterial communities of adult D. variabilis ticks collected from field populations in North Carolina. Sequence analyses were performed using QIIME 2 with the DADA2 plugin and taxonomic assignments using the Greengenes database. After quality filtering and rarefaction, the bacterial DNA sequences were assigned to 432 operational taxonomic units (OTUs) and were dominated by a single OTU classified as Francisella spp. Subsequent cloning, sequencing and phylogenic analysis of nearly full length 16S rRNA gene fragments revealed that most Francisella were similar to Francisella-like endosymbionts (FLEs) of D. variabilis reported previously. One cloned Francisella, however, was genetically distinct and more related to the FLE of D. occidentalis. The core microbiome of D. variabilis was comprised of Francisella spp., Sphingomonas spp., Delftia spp., and Hymenobacter spp. The taxonomic resolution and detection of less abundant bacterial taxa, including Rickettsia, are discussed. Alpha diversity metrics revealed lower bacterial community diversity in females. Beta-diversity also distinguished the composition of female bacterial communities from those of males. These findings advance current knowledge of the microbial ecology and structure and composition of D. variabilis microbiome and sets the foundation for further studies to determine the influence of microbiota on vector susceptibility to pathogens.