Novel Harzia ixtarensis Fungus on Annona cherimola Fruit in Mexico and Its Synergistic Relationship with Colletotrichum fragariae.

Since 2005 in Íxtaro, Michoacán, symptoms of Harzia infection have been observed on immature Annona cherimola fruit with Colletotrichum fragariae-induced anthracnose lesions and mummified fruit. This study aimed to identify the Harzia sp. and evaluate its pathogenicity. Four isolates were obtained from fruit exhibiting symptoms, cultured in four types of agar under various conditions, and characterized based on concatenated internal transcribes spacer (ITS) + large subunit and ITS + small subunit sequences. Additionally, the isolates were compared with two CBS species (two-type strains and two isolates) of Harzia patula and H. tenella under the same conditions as the Harzia isolates, and all known Harzia spp. in culture were included in two phylogenetic analyses. H. ixtarensis sp. nov. was proposed. Compared with H. patula CBS isolate 121524 which was the most closely phylogenetically related species, H. ixtarensis was characterized by slower colony growth (white to salmonish-beige), different percentages of two forms of conidia (elongated and globose; unicellular and hyaline to subhyaline), and smaller conidia. The conidia mainly germinated with two hyaline tubes without an appressorium. In situ inoculations (1 × 106 ml-1 conidia suspension) of fruit showed that fruit with wounds developed larger lesions than those without wounds. Harzia inoculation on anthracnose lesions (induced by prior inoculation with C. fragariae) produced larger anthracnose lesions than C. fragariae alone. When C. fragariae or H. ixtarensis was inoculated alone, the lesion size was 51 and 99% smaller, respectively, indicating synergy between C. fragariae and H. ixtarensis. Thus, H. ixtarensis may have a parasitic-synergistic and necrotrophic lifestyle, and exhibited symptoms on anthracnose lesions.

Machine learning and analysis of genomic diversity of "Candidatus Liberibacter asiaticus" strains from 20 citrus production states in Mexico.

Background: Huanglongbing (HLB, yellow shoot disease) is a highly destructive citrus disease associated with a nonculturable bacterium, "Candidatus Liberibacter asiaticus" (CLas), which is transmitted by Asian citrus psyllid (ACP, Diaphorina citri). In Mexico, HLB was first reported in Tizimin, Yucatán, in 2009 and is now endemic in 351 municipalities of 25 states. Understanding the population diversity of CLas is critical for HLB management. Current CLas diversity research is exclusively based on analysis of the bacterial genome, which composed two regions, chromosome (> 1,000 genes) and prophage (about 40 genes). Methods and results: In this study, 40 CLas-infected ACP samples from 20 states in Mexico were collected. CLas was detected and confirmed by PCR assays. A prophage gene(terL)-based typing system (TTS) divided the Mexican CLas strains into two groups: Term-G including four strains from Yucatán and Chiapas, as well as strain psy62 from Florida, USA, and Term-A included all other 36 Mexican strains, as well as strain AHCA1 from California, USA. CLas diversity was further evaluated to include all chromosomal and prophage genes assisted by using machine learning (ML) tools to resolve multidimensional data handling issues. A Term-G strain (YTMX) and a Term-A strain (BCSMX) were sequenced and analyzed. The two Mexican genome sequences along with the CLas genome sequences available in GenBank were studied. An unsupervised ML was implemented through principal component analysis (PCA) on average nucleotide identities (ANIs) of CLas whole genome sequences; And a supervised ML was implemented through sparse partial least squares discriminant analysis (sPLS-DA) on single nucleotide polymorphisms (SNPs) of coding genes of CLas guided by the TTS. Two CLas Geno-groups, Geno-group 1 that extended Term-A and Geno-group 2 that extended Term-G, were established. Conclusions: This study concluded that: 1) there were at least two different introductions of CLas into Mexico; 2) CLas strains between Mexico and USA are closely related; and 3) The two Geno-groups provide the basis for future CLas subspecies research.

Conventional and qPCR reveals the presence of 'Candidatus Liberibacter solanacearum' haplotypes A, and B in Physalis philadelphica plant, seed, and Βactericera cockerelli psyllids, with the assignment of a new haplotype H in Convolvulaceae.

The husk tomato (Physalis philadelphica Lam.) is an important Solanaceae native to Mesoamerica that is grown for its green fruit used as an important ingredient in domestic and international cuisine. Nevertheless, husk tomato plants with symptoms resembling those caused by 'Candidatus Liberibacter solanacearum' (CLso) have been observed during the last decade in plantations located in the State of Mexico, Michoacan and Sinaloa in Mexico. These areas are located near other solanaceous crops where Bactericera cockerelli the well-known psyllid transmitter of CLso is frequently present. Thus, the goal of this study was to determine if CLso haplotypes are present in husk tomato varieties in commercial fields in Mexico. From 2015 to 2016, plants and fruit showing evident symptoms of CLso infection, as well as psyllids were collected in these states and assayed by PCR for CLso using primer sets OA2/OI2c and LpFrag 1-25F/427R. Phylogenetic reconstruction was performed with Bayesian analysis and maximum likelihood methods using amplicon sequences obtained in this work along with those deposited in the GenBank database corresponding to the CLso detected in Solanaceae, Apiaceae, and Convolvulaceae host families. In addition, all the sequences were subjected to haplotype determination through an analysis of DNA polymorphisms using the DnaSP software. Furthermore, quantitative PCR (qPCR) was performed using CLso-specific primers and probes. Phylogenetic reconstruction and qPCR confirmed the presence of CLso in plants, seeds and insect-vectors, and CLso sequences from plants and seeds completely matched haplotype B, whereas CLso haplotypes A and B were detected in B. cockerelli psyllids. Polymorphism analysis identified a novel Convolvulaceae-associated CLso haplotype, which was named haplotype H. The results of this study will enable the dissemination of infected seeds to new husk tomato production areas to be avoided.

Detección de geminivirus asociados a la alstroemeria (alstroemeria l.) En villa guerrero, estado de México / Detection of geminivirus associated to alstroemeria (alstroemeria l.) In villa guerrero, state of Mexico / Detecção de geminivírus associados a alstroemeria (alstroemeria l.) Em villa guerrero, estado do México

En plantaciones de alstroemeria (Alstroemeria L.) de Villa Guerrero, Estado de México, se han detectado plantas con síntomas similares a los inducidos por geminivirus en otros cultivos hortícolas. En dichas plantaciones también se ha observado la presencia de la mosquita blanca, considerada como el vector más eficiente de estos virus. El objetivo del presente trabajo fue detectar la presencia de geminivirus en plantas de alstroemeria. Mediante PCR, usando los iniciadores MotCP2118/MotCP2123, se obtuvo un segmento de ~600pb, similar al del control positivo correspondiente a chile infectado con el begomovirus pepper huasteco yellow vein virus (PHYVV, antes pepper huasteco virus) en plantas sintomáticas, mientras que en las asintomáticas la detección fue negativa. Plantas de Nicotiana glutinosa, N. benthamiana, N. rustica, N. tabacum var. xanthi y Datura stramonium inoculadas por biobalística con ADN total obtenido de alstroemerias con síntomas y positivas a PHYVV mediante PCR, mostraron mosaicos leves y deformación de hojas, mientras que en plantas de Capsicum annuum se observaron mosaicos, necrosis en nervaduras y abultamientos en hojas. Con el ADN de estas plantas también se obtuvieron bandas correspondientes al PHYVV, pero en las monocotiledóneas bombardeadas, incluyendo alstroemeria, no fue detectado el fragmento. La secuencia de oligonucleótidos de los productos de PCR mostró 98% de homología con el begomovirus PHYVV. Aunque no fue posible reproducir en alstroemeria los síntomas observados en campo, sí se evidenció mediante PCR la presencia de un geminivirus similar al PHYVV en tejido de plantas sintomáticas.

In alstroemeria (Alstroemeria L.) plantations located in Villa Guerrero, Mexico State, plants with symptoms similar to those induced by geminivirus in other horticultural crops have been detected. In addition, the presence of whiteflies, which are considered the most efficient vectors of these viruses, has been observed in these plantations. The goal of this work was to detect the presence of this geminivirus species in alstroemeria plants. By means of PCR analysis using primers MotCP2118/MotCP2123, a fragment of ~600pb similar to the amplicon obtained from PHYVV-infected positive control was amplified only from symptomatic plants. Nicotiana glutinosa, N. benthamiana, N. rustica, N. tabacum var. xanthi and Datura stramonium plants were inoculated by bombardment with total DNA obtained from symptomatic alstroemerias and positive to PHYVV by means of PCR. Inoculated plants showed mild mosaics and deformation of leaves, whereas in the leaves of Capsicum annum plants, mosaics, vein necrosis and blisters were observed. Using DNA from these plants as template in PCR, amplicons corresponded to PHYVV were also obtained; however, in bombarded monocotyledons, including alstroemeria, this fragment was not detected. The sequence of oligonucleotides from the PCR products showed 98% homology to PHYVV geminivirus. Even though symptoms presented by alstroemeria plants in the field were not reproduced, the presence of a geminivirus similar to PHYVV in tissue of symptomatic plants was evidenced through PCR.

Em plantações de alstroemeria (Alstroemeria L.) de Villa Guerrero, Estado do México, se tem detectado plantas com sintomas similares aos induzidos por geminivírus em outros cultivos hortícolas. Em ditas plantações também tem sido observada a presença da mosquinha branca, considerada como o vetor mais eficiente destes vírus. O objetivo do presente trabalho foi detectar a presença de geminivírus em plantas de alstroemeria. Mediante PCR, usando os iniciadores MotCP2118/MotCP2123, obteve-se um segmento de ~600pb, similar ao do controle positivo correspondente a chile infectado com o begomovírus pepper huasteco yellow vein virus (PHYVV, antes pepper huasteco virus) em plantas sintomáticas, enquanto que nas assintomáticas a detecção foi negativa. Plantas de Nicotiana glutinosa, N. benthamiana, N. rustica, N. tabacum var. xanthi e Datura stramonium inoculadas por biobalística com DNA total obtido de alstroemerias com sintomas e positivas a PHYVV mediante PCR, mostraram mosaicos leves e deformação de folhas, enquanto que em plantas de Capsicum annuum se observaram mosaicos, necrose em nervaduras e protuberâncias em folhas. Com o DNA destas plantas também se obtiveram bandas correspondentes ao PHYVV, mas nas monocotiledóneas bombardeadas, incluindo alstroemeria, não foi detectado o fragmento. A sequência de oligonucleotídeos dos produtos de PCR mostrou 98% de homologia com o begomovírus PHYVV. Embora não foi possível reproduzir em alstroemeria os sintomas observados em campo, sím foi evidenciada, mediante PCR, a presença de um geminivírus similar ao PHYVV em tecido de plantas sintomáticas.