Publications

4369

AbstractOf late, the presence of Candidatus Phytoplasma trifolii was reported as a serious threat to the pepper crop in Zacatecas, México; therefore, asymptomatic and symptomatic pepper plants were collected from a commercial field among three samplings after the fruit set stage was reached. Total DNA was extracted using the CTAB‐based method and tested for phytoplasma using a nested PCR assay, followed by a BLAST, and restriction fragment length polymorphism (RFLP) analysis of 16S rDNA sequences, which confirmed the presence of phytoplasma group 16SrVI, “Candidatus Phytoplasma trifolii” in the symptomatic plants. As the metabolic pathways of pathogen‐infected plants tend to change, resulting in a biochemical differentiation with the noninfected plants, the polyphenolic compound concentrations were quantified from the vegetative tissues (root, stem, leaves and developed fruit/big bud) and were analysed based on a principal component analysis (PCA). Results revealed that, in general, plants tend to a progressive increase in total phenols, flavonoids, condensed tannins and anthocyanins related to the plants exposure to “Ca. P. trifolii” infection, and PCA demonstrated that almost 90% of the observed variance was explained by the first two components. Hence, the phenolic content of the plants increases as a response of the defence mechanism, which reflects its condition and resistance.

Huanglongbing ( HLB ), caused by ‘ Candidatus Liberibacter asiaticus’ (Las), is a devastating disease of citrus trees in Florida. Previous work showed that the rootstock cultivar Cleopatra mandarin ( Citrus reticulata ) has a higher population of Las in roots than Swingle citrumelo ( C. paradisi  ×  Poncirus trifoliata ). Las reduced fibrous root biomass and sucrose content in Cleopatra mandarin more than in Swingle citrumelo. To understand the mechanisms for susceptibility to Las infection, sucrose and hormone metabolism status were evaluated in Cleopatra mandarin and Swingle citrumelo. In fibrous roots of Cleopatra mandarin, higher expression of genes related to sucrose cleavage was consistent with lower sucrose content compared to noninoculated seedlings at 5 weeks post‐root trimming (wpt). In fibrous roots of Swingle citrumelo, both sucrose content and gene expression related to sucrose cleavage were less disrupted by Las infection compared to Cleopatra mandarin at 5 wpt. Genes associated with salicylic acid ( SA ), ethylene ( ET ) and abscisic acid ( ABA ) synthesis, and ABA signalling, phospholipases D ( PLD ), and phospholipase A2 ( PLA 2) were activated by Las infection at 5 wpt in Cleopatra mandarin. Expression of downstream effectors of SA , i.e. NPR 1, WRKY 70 and PR 1, did not change in Cleopatra mandarin, suggesting inhibition of the response to SA by the elevation of ABA , ET and PLD . In contrast, the up‐regulation of PR 1, lower response of sucrose metabolism genes and down‐regulation of biosynthesis of phytohormones indicates that Swingle citrumelo activates a more effective defence against this biotrophic pathogen than Cleopatra mandarin.

AbstractA disease known as broccoli stunt, associated with “Candidatus Phytoplasma pruni”‐related strain, has been responsible by significant economic losses in crops grown in the State of São Paulo, Brazil. Previous investigations evidenced some species of leafhoppers observed in broccoli fields as potential vectors of the phytoplasma. In this study, the six species more frequently found in broccoli crops were collected to confirm that evidence. Group of five insects of each species were confined per broccoli seedling for an inoculation access period (IAP) of 48 hr. After the IAP, each group was tested for detection of phytoplasma. Evaluation of plants was performed 60 days after inoculation based on the presence of phytoplasma in their tissues. When transmission was positive, genomic fragments corresponding to 16S rDNA were sequenced both for the infected plants and its respective group of insects. The results revealed that the species Agallia albidula, Agalliana sticticollis, Atanus nitidus and Balcluta hebe were able to transmit phytoplasma to broccoli seedlings. Based on the estimates of transmission probability by single insects (P), the highest transmission rate was observed for A. nitidus (24.2%) and the lowest for B. hebe (1.9%). The sequencing of 16S rDNA revealed complete similarity between the sequences of the phytoplasma transmitted to broccoli test plants and the sequences of the phytoplasma found in the field‐collected leafhoppers. These findings support the inclusion of those species as vectors of phytoplasmas belonging to 16SrIII group in broccolis, providing additional information to improve management of this important disease of endemic occurrence.

AbstractBackground: The symbiosis between the olive fruit fly,Bactrocera oleae, andCandidatusErwinia dacicola has been demonstrated as essential for the fly’s larval development and adult physiology. The mass rearing of the olive fruit fly has been hindered by several issues, including problems which could be related to the lack of the symbiont, presumably due to preservatives and antibiotics currently used in the laboratory. To better understand the mechanisms underlying symbiont removal or loss during the rearing of lab colonies of the olive fruit fly, we performed experiments that focused on bacterial transfer from wild female flies to their eggs. In this research, eggs laid by wild females were treated with propionic acid solution, which is often used as an antifungal agent, a mixture of sodium hypochlorite and Triton X, or water (as a control). The presence of the bacterial symbiont on eggs was evaluated by real-time PCR and scanning electron microscopy.Results: DGGE analysis showed a clear band with the same migration behavior present in all DGGE profiles but with a decreasing intensity. Molecular analyses performed by real-time PCR showed a significant reduction inCa. E. dacicola abundance in eggs treated with propionic acid solution or a mixture of sodium hypochlorite and Triton X compared to those treated with water. In addition, the removal of bacteria from the surfaces of treated eggs was highlighted by scanning electron microscopy.Conclusions: The results clearly indicate how the first phases of the colony-establishment process are important in maintaining the symbiont load in laboratory populations and suggest that the use of products with antimicrobial activity should be avoided. The results also suggest that alternative rearing procedures for the olive fruit fly should be investigated.

ABSTRACT Members of the genus Rickettsiella are bacterial pathogens of insects and other arthropods. Recently, a novel facultative endosymbiont, “ Candidatus Rickettsiella viridis,” was described in the pea aphid Acyrthosiphon pisum , whose infection causes a striking host phenotype: red and green genetic color morphs exist in aphid populations, and upon infection with the symbiont, red aphids become green due to increased production of green polycyclic quinone pigments. Here we determined the complete genome sequence of the symbiont. The 1.6-Mb circular genome, harboring some 1,400 protein-coding genes, was similar to the genome of entomopathogenic Rickettsiella grylli (1.6 Mb) but was smaller than the genomes of phylogenetically allied human pathogens Coxiella burnetii (2.0 Mb) and Legionella pneumophila (3.4 Mb). The symbiont’s metabolic pathways exhibited little complementarity to those of the coexisting primary symbiont Buchnera aphidicola , reflecting the facultative nature of the symbiont. The symbiont genome harbored neither polyketide synthase genes nor the evolutionarily allied fatty acid synthase genes that are suspected to catalyze the polycyclic quinone synthesis, indicating that the green pigments are produced not by the symbiont but by the host aphid. The symbiont genome retained many type IV secretion system genes and presumable effector protein genes, whose homologues in L. pneumophila were reported to modulate a variety of the host's cellular processes for facilitating infection and virulence. These results suggest the possibility that the symbiont is involved in the green pigment production by affecting the host’s metabolism using the secretion machineries for delivering the effector molecules into the host cells. IMPORTANCE Insect body color is relevant to a variety of biological aspects such as species recognition, sexual selection, mimicry, aposematism, and crypsis. Hence, the bacterial endosymbiont “ Candidatus Rickettsiella viridis,” which alters aphid body color from red to green, is of ecological interest, given that different predators preferentially exploit either red- or green-colored aphids. Here we determined the complete 1.6-Mb genome of the symbiont and uncovered that, although the red-green color transition was ascribed to upregulated production of green polycyclic quinone pigments, the symbiont genome harbored few genes involved in the polycyclic quinone biosynthesis. Meanwhile, the symbiont genome contained type IV secretion system genes and presumable effector protein genes, whose homologues modulate eukaryotic cellular processes for facilitating infection and virulence in the pathogen Legionella pneumophila . We propose the hypothesis that the symbiont may upregulate the host’s production of polycyclic quinone pigments via cooption of secretion machineries and effector molecules for pathogenicity.