Publications

4466

Huanglongbing (HLB) is a major disease of citrus associated with phloem-limited bacteria in the genus Candidatus Liberibacter that affects all known citrus species and relatives, with many commercial cultivars being greatly damaged. Testing cultivar tolerance to HLB in field conditions is difficult because of the erratic spread of the bacteria, scion and rootstock interactions, and influence of many biotic and abiotic factors on the tree response to the disease. This study aimed to validate the effect of CLas infection on different citrus species and hybrids thought to have different levels of tolerance to the disease using CLas graft inoculation under controlled greenhouse conditions. Young potted seedlings from 12 different citrus germplasm selections were graft-inoculated with CLas or mock-inoculated. Plants were monitored periodically during 18 months for canopy growth, HLB and nutritional leaf symptoms, and leaf CLas titers. The leaf nutrient content was measured at the end of the experiment. Roots were also assessed at 18 months after inoculation (mai) for CLas titers and biomass distribution. There were significant differences in most analyzed variables of healthy and infected plants. Some plants of all cultivars were successfully infected; however, overall, the CLas transmission rate was low and inconsistent. Ct values of roots were generally higher than those in leaves at 18 mai. HLB symptoms were not observed on seedlings until 1 year after inoculation; at 18 mai, infected trees of all cultivars were HLB symptomatic. Significant shoot and root biomass reductions (44%–75%) in infected ‘Cleopatra’, ‘Duncan’, ‘Olinda Valencia’, ‘Sunburst’, and ‘Valencia 1-14-19’, considered susceptible to HLB, were measured. These cultivars also showed more severe HLB symptoms than the presumed tolerant cultivars such as Microcitrus inodora, Rich 16-6 trifoliate orange, and US-897. This study provides new knowledge of the efficacy and value of greenhouse screening of citrus germplasm for response to HLB to support the development of new cultivars with improved HLB tolerance or resistance.

This study delves into the evolutionary history of Anaerolineaceae, a diverse bacterial family within the Chloroflexota phylum. Employing a multi-faceted approach, including phylogenetic analyses, genomic comparisons, and exploration of adaptive features, the research unveils novel insights into the family’s taxonomy and evolutionary dynamics. The investigation employs metagenome-assembled genomes (MAGs), emphasizing their prevalence in anaerobic environments. Notably, a novel mesophilic lineage, tentatively named Mesolinea, emerges within Anaerolineaceae, showcasing a distinctive genomic profile and apparent adaptation to a mesophilic lifestyle. The comprehensive genomic analyses shed light on the family’s complex evolutionary patterns, including the conservation of key operons in thermophiles, providing a foundation for understanding the diverse ecological roles and adaptive strategies of Anaerolineaceae members.

In Germany, the phloem sucking planthopper Pentastiridius leporinus (Hemiptera: Cixiidae) currently represents the epidemiological driver for the spread of the syndrome “Basses Richesses” in sugar beet, which results in a reduced sugar content and an economic loss for the farmers. This disease is associated with the γ-proteobacterium ‘Candidatus Arsenophonus phytopathogenicus’ and the Stolbur phytoplasma ‘Candidatus Phytoplasma solani’. Recently, P. leporinus was found in potato fields in Germany associated with Stolbur-like symptoms in this crop. In this study, we confirmed that the vector completes its lifecycle on sugar beet as well as on potato when reared under controlled conditions. Transmission experiments with adults of this vector combined with molecular analyses showed for the first time that both pathogens are transmitted by this vector to potato. For an accurate assessment of the Ca. P. solani and Ca. A. phytopathogenicus titers in the vector and host plants, gBlocks derived from the hsp20- and 16S rRNA gene were employed, respectively. For Ca. P. solani the limit of detection was determined in potato and sugar beet material. The results of this study will further research on the epidemiology of the syndrome “Basses Richesses” and “Stolbur” diseases and the response of host plants and vector insects to both pathogens.

O Huanglongbing (HLB) destaca-se como a doença mais significativa que afeta os cultivos de citros, sendo atribuível a três espécies distintas de α-proteobactérias do gênero Candidatus Liberibacter: Ca. Liberibacter asiaticus (CLas), Ca. Liberibacter americanus (CLam) e Ca. Liberibacter africanus (CLaf). No Brasil, o principal vetor para a transmissão de CLas é o psilídeo asiático dos citros, Diaphorina citri (Kuwayama) (Hemiptera: Liviidae). Este estudo tem como objetivo verificar a presença de D. citri na jurisdição da ADAPAR - PR, a Agência de Defesa Agropecuária do Estado do Paraná, e correlacionar sua incidência com o HLB. A pesquisa abrangeu todo o estado do Paraná, incluindo municípios-chave envolvidos em diversas atividades agrícolas relacionadas ao citros, desde pomares comerciais até instalação de pesquisa. Em locais georreferenciados, agentes da ADAPAR instalaram armadilhas adesivas amarelas para monitorar D. citri nos anos de 2018 e 2019. As armadilhas, identificadas com data, local e coordenadas específicas, foram coletadas, acondicionadas em sacos plásticos transparentes e armazenadas sob refrigeração até a identificação. A classificação dos insetos capturados foi realizada utilizando uma lupa binocular. Espécimes adultas de D. citri foram submetidas à extração total de DNA, seguida pela PCR direcionada ao gene 16S rRNA de CLas. Os produtos resultantes da PCR foram analisados em gel de agarose a 2% corado com brometo de etídio. A análise da triagem e classificação de insetos confirmou a presença de D. citri em vários municípios, incluindo Altônia, Ampére, Ângulo, Antonina, Cruzeiro do Oeste, Dois Vizinhos, Faxinal Guaratuba, Iporã, Ivaiporã, Nova América da Colina, Ponta Grossa, Presidente Castelo Branco e Santa Cruz de Monte Castelo. Uma análise adicional dos dados sobre temperatura média anual e precipitação, juntamente com o IDH-Paraná, indicou que Presidente Castelo Branco teve o maior número de psilídeos coletados, totalizando 96 insetos, todos em 2018. Concentrações significativas também foram observadas em Altônia, Ampére, Cruzeiro do Oeste, Iporã, Santa Cruz de Monte Castelo e Vila Rural em Jandaia do Sul em 2018. Em 2019, foram registradas contagens elevadas de psilídeos em Ampére, Foz do Iguaçu e Ponta Grossa. Após a coleta, as amostras foram submetidas à extração de DNA e amplificação por PCR de Candidatus Liberibacter asiaticus. Embora todos os psilídeos coletados tenham testado positivo para a infecção por CLas, aqueles obtidos em armadilhas em Ponta Grossa, Paranavaí, Dois Vizinhos, Ivaiporã e Moreira Sales foram negativos. Isso destaca que fatores como temperatura, umidade relativa, precipitação e disponibilidade de planta hospedeira são determinantes críticos da ocorrência de D. citri em uma determinada região. Além disso, o estudo enfatiza o papel fundamental de uma agência eficaz de defesa agropecuária no monitoramento de doenças e pragas, especialmente em um patossistema que envolve patógeno, hospedeiro e vetor.

Entomopathogenic nematodes from the genus Steinernema (Nematoda: Steinernematidae) are capable of causing the rapid killing of insect hosts, facilitated by their association with symbiotic Gram-negative bacteria in the genus Xenorhabdus (Enterobacterales: Morganellaceae), positioning them as interesting candidate tools for the control of insect pests. In spite of this, only a limited number of species from this bacterial genus have been identified from their nematode hosts and their insecticidal properties documented. This study aimed to perform the genome sequence analysis of fourteen Xenorhabdus strains that were isolated from Steinernema nematodes in Argentina. All of the strains were found to be able of killing 7th instar larvae of Galleria mellonella (L.) (Lepidoptera: Pyralidae). Their sequenced genomes harbour 110 putative insecticidal proteins including Tc, Txp, Mcf, Pra/Prb and App homologs, plus other virulence factors such as putative nematocidal proteins, chitinases and secondary metabolite gene clusters for the synthesis of different bioactive compounds. Maximum-likelihood phylogenetic analysis plus average nucleotide identity calculations strongly suggested that three strains should be considered novel species. The species name for strains PSL and Reich (same species according to % ANI) is proposed as Xenorhabdus littoralis sp. nov., whereas strain 12 is proposed as Xenorhabdus santafensis sp. nov. In this work, we present a dual insight into the biocidal potential and diversity of the Xenorhabdus genus, demonstrated by different numbers of putative insecticidal genes and biosynthetic gene clusters, along with a fresh exploration of the species within this genus.

‘Candidatus Phytoplasma prunorum’ is one of the most destructive pathogens of Prunus species, where susceptible species render unproductive several years after infection. In epidemiology, the molecular characterization of phytoplasmas is based on sequence analysis of variable nonribosomal genes. In this study aceF, pnp, imp and secY genes were used for characterization of the ‘Ca. P. prunorum’ genotypes present in the Czech Republic. In total, 56 plant and 33 vector (Cacopsylla pruni) samples positive to ‘Ca. P. prunorum’ collected in seven localities were used in the study. Based on sequence analysis, four aceF, two pnp, six imp, and three secY genotypes were identified in analyzed samples. The most abundant in both plant and insect samples were the A6, P2, I4, and S2 genotypes. Most of the Czech ‘Ca. P. prunorum’ haplotypes clustered together in the haplotype network analysis. Next, two isolates representing the two most abundant Czech haplotypes (A6-P2-I4-S2 and A5-P2-I4-S2) were used in the susceptibility test of three apricot rootstock types (St. Julien A, M-VA-1, GF-305). Susceptibility was analyzed by phytoplasma quantification using quantitative real-time PCR and evaluation of symptom manifestation. Based on the results, the influence of the rootstock type on the phytoplasma titer and symptom manifestation was greater than of the phytoplasma isolate, while the year of analysis had no influence on the results. The results also showed that the phytoplasma titer is increasing in plant tissues during the vegetation period.