Publications

3841

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.

AbstractInfection with the bacterium ‘Candidatus Liberibacter solanacearum’ (Lso) is suspected to cause severe damage in carrot leading to high carrot weight loss. This study investigates three main aspects: (i) whether there is a reduction of carrot root weight under field conditions due to Lso infection; (ii) the correlation between Lso infection rate in carrot plants and occurrence of the psyllid Trioza apicalis as the known vector for Lso in carrot, and (iii) the comparison between symptoms described in literature and observed symptoms associated with Lso infection. Therefore, field surveys were conducted from 2018 to 2021 on organically managed carrot fields in Lower Saxony, Germany. Two Rebell orange sticky traps were placed per field replaced on a weekly basis. Captured T. apicalis were morphologically identified and counted. Carrot plant samples were collected from T. apicalis infested fields. Discolouration of foliage and further symptoms on carrots were investigated. For detection of Lso in carrot plants and psyllids, the samples were analysed using PCR. The infection rate of carrot plants varied between 2.5% and 80% per field with low abundance of T. apicalis. Of the asymptomatic plants, 39.4% were infected with Lso, while 80.9% of the symptomatic plants were infected with Lso. The weight of Lso-positive carrot samples was not significantly reduced compared to Lso-negative samples. No economically relevant losses of carrot yield were reported by farmers participating in this study. This indicates that there was no effect of Lso infection on carrot root weight during this study. Of the symptoms associated with a Lso infection in carrots, only leaf discolouration could be confirmed. Overall, the bacterium is present in organically grown carrot plants in Lower Saxony but no major carrot root weight losses were observed.

Two Gram-stain-positive, aerobic, non-spore-forming, non-motile, irregular rod-shaped actinobacteria, designated as D2-41T and D3-21, were isolated from soil samples collected in a natural cave in Jeju, Republic of Korea. Both of the isolates were shown to share 100 % 16S rRNA sequence identity. The cell wall contained meso-diaminopimelic acid, arabinose and galactose. The predominant menaquinone was MK-8(H2). The polar lipids contained phosphatidylethanolamine, phosphatidylglycerol, phosphatidylinositol, phosphatidylinositol mannoside, an unidentified aminolipid, an unidentified aminoglycolipid, an unidentified phospholipid and two unidentified lipids. The predominant fatty acids were C16 : 0 and summed feature 3 (C16 : 1  ω7c and/or iso-C15 : 0 2-OH). Mycolic acids of C30–C38 were present. The 16S rRNA gene trees showed that the organisms occupied a distinct position remotely located from recognized genera within the order Mycobacteriales, albeit with the 16S rRNA gene similarities of 97.0–97.1 % with Rhodococcus olei, Rhodococcus rhodnii and Rhodococcus triatomae. The genome sizes and DNA G+C contents of strains D2-41T and D3-21 were 4.77–4.88 Mbp and 69.8 mol%, respectively. Both of the isolates shared an average nucleotide identity of 99.4 % and digital DNA–DNA hybridization of 95.2 % to each other, revealing that strains D2-41T and D3-21 belonged to the same species. In the core genome-based phylogenomic tree, both of the isolates were found to be closely associated with members of the genus Tomitella. However, strains D2-41T and D3-21 revealed the highest amino acid identity values (mean 66.5 %, range 66.2–67.0 % with the genus Prescottella of the family Nocardiaceae, followed by the genus Tomitella (mean 64.1 %, range 63.6–64.7 %) of the family Tomitellaceae. Based on the combined data obtained here, the novel isolates belong to a new genus of the new family for which the name Speluncibacter jeojiensis gen. nov. sp. nov. is proposed, with Speluncibacteraceae fam. nov. The type strain is strain D2-41T (=KACC 17930T=DSM 101875T).