Biological and molecular characterization of a new potexvirus isolated from Adenium obesum.

Adenium obesum plants showing virus-like symptoms were collected in several regions of Brazil. Mottling symptoms like those observed in symptomatic plants in the field were reproduced in mechanically inoculated A. obesum plants. This potexvirus was named "desert rose mottle virus" (DRMoV), and its genome sequence was first determined by high-throughput sequencing and then confirmed by Sanger sequencing. The complete genome of DRMoV is 6,781 nt in length, excluding the poly(A) tail, and five ORFs were predicted in order from 5' to 3': Rep-TGB1-TGB2-TGB3-CP. Phylogenetic analysis based on Rep amino acid sequences showed different clustering among potexviruses. These data suggest that RDMoV is a new member of the genus Potexvirus, and the binomial name "Potexvirus adenii" is proposed for its species.

Molecular and biological characterization of an isolate of the potyvirus passiflora virus Y naturally infecting soybean (Glycine max) in Brazil.

Passiflora virus Y was detected naturally infecting soybean (Glycine max) for the first time in Brazil. Here, we report the nearly complete genome sequence and molecular and biological properties of the PaVY-Br isolate. The nearly complete genome sequence is 9679 nt long and shares 84.4% nt sequence identity with a previously reported PaVY isolate from Passiflora sp. PaVY-Br induced chlorotic spots and systemic mosaic on soybean and chlorotic local lesions on yellow passion fruit (Passiflora edulis) and sesame (Sesamum indicum). The virus was successfully transmitted by Myzus persicae, indicating that this aphid vector can contribute to the spread of PaYV from passion fruit to soybean plants. Additional epidemiological research is in progress to investigate the distribution of PaVY in soybean production areas in Brazil.

Influence of Mycosphaerella and Teratosphaeria leaf diseases on chemical composition of essential oils of Eucalyptus globulus and effect of these essential oils on ascospores germination.

In a first step, essential oils were extracted from Eucalyptus globulus leaves, healthy and with symptoms and signs of Mycosphaerella leaf disease (MLD) and Teratosphaeria leaf disease (TLD), in two leaf stages. Stage 1: sessile, oval leaves covered by a waxy layer of a bluish colour, with opposite phyllotaxis, inserted along stems of quadrangular section. Stage 2: narrow and sickle leaves with a greyish green surface, mainly on the abaxial surface, inserted in alternating pairs along rounded stems. The essential oils were extracted by hydrodistillation and were analyzed by gas chromatography coupled with mass spectrometry. Chemical composition data and percentages of essential oil constituents were submitted to cluster analysis and principal component analysis. In a second step, under in vitro conditions, was evaluated the germination of Teratosphaeria nubilosa (one of the causal agents of TLD) ascospores in contact with the four types of essential oils extracted. The evaluations were performed at 24, 48 and 72 h after the experiments were assembled. The present study made it possible to distinguish and identify the chemical composition of essential oils from the eucalypt leaves used, and allowed 1,8-cineole to be identified as the major component for the essential oils investigated. The contact between essential oils and T. nubilosa spores allowed to prove the inhibition of the ascospores germination, being more efficient for the essential oils extracted from materials with the disease, which presented high amounts of 1,8-cineole.

Population Dynamics of Whiteflies and Associated Viruses in South America: Research Progress and Perspectives.

By having an extensive territory and suitable climate conditions, South America is one of the most important agricultural regions in the world, providing different kinds of vegetable products to different regions of the world. However, such favorable conditions for plant production also allow the development of several pests, increasing production costs. Among them, whiteflies (Hemiptera: Aleyrodidae) stand out for their potential for infesting several crops and for being resistant to insecticides, having high rates of reproduction and dispersal, besides their efficient activity as virus vectors. Currently, the most important species occurring in South America are Bemisia afer, Trialeurodes vaporariorum, and the cryptic species Middle East-Asia Minor 1, Mediterranean, and New World, from Bemisia tabaci complex. In this review, a series of studies performed in South America were compiled in an attempt to unify the advances that have been developed in whitefly management in this continent. At first, a background of the current whitefly distribution in South American countries as well as factors affecting them are shown, followed by a background of the whitefly transmitted viruses in South America, addressing their location and association with whiteflies in each country. Afterwards, a series of management strategies are proposed to be implemented in South American fields, including cultural practices and biological and chemical control, finalizing with a section containing future perspectives and directions for further research.

Distribution and phylogenetics of whiteflies and their endosymbiont relationships after the Mediterranean species invasion in Brazil.

The Bemisia tabaci is a polyphagous insect and a successful vector of plant viruses. B. tabaci is a species complex and in Brazil native species from the New World (NW) group, as well as the invasive species, Middle East-Asia Minor 1 (MEAM1) and Mediterranean (MED) were reported. For better understanding the distribution of the different species four years after the Mediterranean species invasion in Brazil, whiteflies were collected from 237 locations throughout the country between the years of 2013 and 2017, species were identified and the facultative endosymbionts detected. The survey revealed that MEAM1 was the prevalent species found on major crops across Brazil. It is the only species present in North, Northwestern and Central Brazil and was associated with virus-infected plants. MED was found in five States from Southeast to South regions, infesting mainly ornamental plants and was not associated with virus-infected plants. The prevalent endosymbionts identified in MEAM1 were Hamiltonella and Rickettsia; and the mtCOI analysis revealed low genetic diversity for MEAM1. In contrast, several different endosymbionts were identified in MED including Hamiltonella, Rickettsia, Wolbachia and Arsenophonus; and two distinct genetic groups were found based on the mtCOI analysis. Monitoring the distribution of the whiteflies species in Brazil is essential for proper management of this pest.

First report of Bemisia tabaci Mediterranean (Q biotype) species in Brazil.

BACKGROUND: The whitefly Bemisia tabaci is a major cosmopolitan pest and comprises a complex of more than 36 cryptic species that cause serious damage to agricultural crops worldwide. In this study, the Mediterranean species of B. tabaci, formerly known as Q biotype, was identified for the first time in Brazil. RESULTS: Adult B. tabaci were collected from different localities and hosts from Rio Grande do Sul, the southernmost state of the country that borders Uruguay and Argentina. Partial sequencing of the mitochondrial cytochrome oxidase I (mtCOI) gene indicated that B. tabaci MED species appears to be restricted to the province of Barra do Quaraí, infesting Capsicum annuum cultivated in greenhouses and Ipomoea batatas in open fields. The partial mtCOI sequences obtained shared 100% nucleotide identity with reference sequences for the Q biotype reported from Uruguay. The secondary endosymbionts Hamiltonella and Cardinium were detected by PCR in the new identified MED species from Brazil, similarly to the Q biotype from Uruguay. CONCLUSION: Our results indicate the presence of the MED species in Brazil. The close monitoring of this new identified species in the southern region of Brazil is essential to avoid its geographical expansion to more important agricultural areas in the country.

Diversity and localization of bacterial endosymbionts from whitefly species collected in Brazil.

Whiteflies (Hemiptera: Aleyrodidae) are sap-sucking insect pests, and some cause serious damage in agricultural crops by direct feeding and by transmitting plant viruses. Whiteflies maintain close associations with bacterial endosymbionts that can significantly influence their biology. All whitefly species harbor a primary endosymbiont, and a diverse array of secondary endosymbionts. In this study, we surveyed 34 whitefly populations collected from the states of Sao Paulo, Bahia, Minas Gerais and Parana in Brazil, for species identification and for infection with secondary endosymbionts. Sequencing the mitochondrial Cytochrome Oxidase I gene revealed the existence of five whitefly species: The sweetpotato whitefly Bemisia tabaci B biotype (recently termed Middle East-Asia Minor 1 or MEAM1), the greenhouse whitefly Trialeurodes vaporariorum, B. tabaci A biotype (recently termed New World 2 or NW2) collected only from Euphorbia, the Acacia whitefly Tetraleurodes acaciae and Bemisia tuberculata both were detected only on cassava. Sequencing rRNA genes showed that Hamiltonella and Rickettsia were highly prevalent in all MEAM1 populations, while Cardinium was close to fixation in only three populations. Surprisingly, some MEAM1 individuals and one NW2 population were infected with Fritschea. Arsenopnohus was the only endosymbiont detected in T. vaporariorum. In T. acaciae and B. tuberculata populations collected from cassava, Wolbachia was fixed in B. tuberculata and was highly prevalent in T. acaciae. Interestingly, while B. tuberculata was additionally infected with Arsenophonus, T. acaciae was infected with Cardinium and Fritschea. Fluorescence in situ hybridization analysis on representative individuals showed that Hamiltonella, Arsenopnohus and Fritschea were localized inside the bacteriome, Cardinium and Wolbachia exhibited dual localization patterns inside and outside the bacteriome, and Rickettsia showed strict localization outside the bacteriome. This study is the first survey of whitely populations collected in Brazil, and provides further insights into the complexity of infection with secondary endosymionts in whiteflies.

Fluorescence in situ hybridizations (FISH) for the localization of viruses and endosymbiotic bacteria in plant and insect tissues.

Fluorescence in situ hybridization (FISH) is a name given to a variety of techniques commonly used for visualizing gene transcripts in eukaryotic cells and can be further modified to visualize other components in the cell such as infection with viruses and bacteria. Spatial localization and visualization of viruses and bacteria during the infection process is an essential step that complements expression profiling experiments such as microarrays and RNAseq in response to different stimuli. Understanding the spatiotemporal infections with these agents complements biological experiments aimed at understanding their interaction with cellular components. Several techniques for visualizing viruses and bacteria such as reporter gene systems or immunohistochemical methods are time-consuming, and some are limited to work with model organisms and involve complex methodologies. FISH that targets RNA or DNA species in the cell is a relatively easy and fast method for studying spatiotemporal localization of genes and for diagnostic purposes. This method can be robust and relatively easy to implement when the protocols employ short hybridizing, commercially-purchased probes, which are not expensive. This is particularly robust when sample preparation, fixation, hybridization, and microscopic visualization do not involve complex steps. Here we describe a protocol for localization of bacteria and viruses in insect and plant tissues. The method is based on simple preparation, fixation, and hybridization of insect whole mounts and dissected organs or hand-made plant sections, with 20 base pairs short DNA probes conjugated to fluorescent dyes on their 5' or 3' ends. This protocol has been successfully applied to a number of insect and plant tissues, and can be used to analyze expression of mRNAs or other RNA or DNA species in the cell.