Potential areas for the establishment of citrus leprosis virus vectors, Brevipalpus spp., in Mexico.

Citrus leprosis is a viral disease vectored by the mites Brevipalpus californicus and Brevipalpus yothersi. This work aimed to determine the potential areas for establishment of both mites and viruses in Mexico, based on the geographical distribution of the hosts and the climatic suitability for the vectors. Life tables of both mites were constructed to determine their thermal requirements-base temperature and degree-days required to complete life cycle-and population growth parameters-net reproduction rate, generation time, and intrinsic growth rate. For this, the mites were confined in Citrus aurantium fruits at 20, 22.5, 25 or 30 °C, 60 ± 5% RH and L14:D10 h photoperiod. Maps were generated where the climatic suitability for establishment of the mites and the citrus leprosis viruses was estimated in citrus-producing municipalities. The climatic suitability was determined through historical temperature records to calculate the potential number of generations per year, and ecological niche modeling based on collecting localities and bioclimatic variables using the algorithm Maxent. The base temperature was 9.5 °C for B. californicus and 10.2 °C for B. yothersi; degree-days required to reach adulthood were 372.1 and 331.7 °C, respectively. Potential sites for establishment of B. yothersi are mostly lowlands, whereas for B. californicus they are both lowlands and highlands. Temperature data indicate that B. californicus has fewer sites where it can develop > 16 generations per year than B. yothersi. According to our results, the sites where citrus leprosis is most likely to present high incidence are the sweet orange cultivars bordering the Gulf of Mexico.

First detection of orchid fleck virus in orchids in Mexico.

For the first time, an isolate of the dichorhavirus orchid fleck virus (OFV, family Rhabdoviridae) was found infecting an orchid plant in Mexico. The infected sample of Epidendrum veroscriptum was collected in a nursery in Lagunillas, municipality of Zihuateutla, Edo. Puebla. Mites gathered on this plant were analyzed by light and scanning electron microscopy, which consistently indicated the presence of adults of the species Brevipalpus californicus, the common vector of OFV. Viral identification was based on symptoms, cytopathology, and reverse transcriptase-PCR/sequencing of genome fragments of the RNA1 and 2 molecules. Since isolates of OFV causing citrus leprosis have been previously detected in the Mexican states of Chiapas, Querétaro, and Jalisco, we promote a pertinent discussion and thought-provoking questions regarding the epidemiology and putative evolution of OFV.

Role Bending: Complex Relationships Between Viruses, Hosts, and Vectors Related to Citrus Leprosis, an Emerging Disease.

Citrus leprosis complex is an emerging disease in the Americas, associated with two unrelated taxa of viruses distributed in South, Central, and North America. The cytoplasmic viruses are Citrus leprosis virus C (CiLV-C), Citrus leprosis virus C2 (CiLV-C2), and Hibiscus green spot virus 2, and the nuclear viruses are Citrus leprosis virus N (CiLV-N) and Citrus necrotic spot virus. These viruses cause local lesion infections in all known hosts, with no natural systemic host identified to date. All leprosis viruses were believed to be transmitted by one species of mite, Brevipalpus phoenicis. However, mites collected from CiLV-C and CiLV-N infected citrus groves in Mexico were identified as B. yothersi and B. californicus sensu lato, respectively, and only B. yothersi was detected from CiLV-C2 and CiLV-N mixed infections in the Orinoco regions of Colombia. Phylogenetic analysis of the helicase, RNA-dependent RNA polymerase 2 domains and p24 gene amino acid sequences of cytoplasmic leprosis viruses showed a close relationship with recently deposited mosquito-borne negevirus sequences. Here, we present evidence that both cytoplasmic and nuclear viruses seem to replicate in viruliferous Brevipalpus species. The possible replication in the mite vector and the close relationship with mosquito borne negeviruses are consistent with the concept that members of the genus Cilevirus and Higrevirus originated in mites and citrus may play the role of mite virus vector.

Identification and Molecular Characterization of Nuclear Citrus leprosis virus, a Member of the Proposed Dichorhavirus Genus Infecting Multiple Citrus Species in Mexico.

Citrus leprosis is one of the most destructive diseases of Citrus spp. and is associated with two unrelated virus groups that produce particles primarily in either the cytoplasm or nucleus of infected plant cells. Symptoms of leprosis, including chlorotic spots surrounded by yellow haloes on leaves and necrotic spots on twigs and fruit, were observed on leprosis-affected mandarin and navel sweet orange trees in the state of Querétaro, Mexico. Serological and molecular assays showed that the cytoplasmic types of Citrus leprosis virus (CiLV-C) often associated with leprosis symptomatic tissues were absent. However, using transmission electron microscopy, bullet-shaped rhabdovirus-like virions were observed in the nuclei and cytoplasm of the citrus leprosis-infected leaf tissues. An analysis of small RNA populations from symptomatic tissue was carried out to determine the genome sequence of the rhabdovirus-like particles observed in the citrus leprosis samples. The complete genome sequence showed that the nuclear type of CiLV (CiLV-N) present in the samples consisted of two negative-sense RNAs: 6,268-nucleotide (nt)-long RNA1 and 5,847-nt-long RNA2, excluding the poly(A) tails. CiLV-N had a genome organization identical to that of Orchid fleck virus (OFV), with the exception of shorter 5' untranslated regions in RNA1 (53 versus 205 nt) and RNA2 (34 versus 182 nt). Phylogenetic trees constructed with the amino acid sequences of the nucleocapsid (N) and glycoproteins (G) and the RNA polymerase (L protein) showed that CiLV-N clusters with OFV. Furthermore, phylogenetic analyses of N protein established CiLV-N as a member of the proposed genus Dichorhavirus. Reverse-transcription polymerase chain reaction primers for the detection of CiLV-N were designed based on the sequence of the N gene and the assay was optimized and tested to detect the presence of CiLV-N in both diseased and symptom-free plants.

Genome assembly of citrus leprosis virus nuclear type reveals a close association with orchid fleck virus.

The complete genome of citrus leprosis virus nuclear type (CiLV-N) was identified by small RNA sequencing utilizing leprosis-affected citrus samples collected from the state of Querétaro, Mexico. The nucleotide identity and phylogenetic analysis indicate that CiLV-N is very closely related to orchid fleck virus, which typically infects Cymbidium species.