virus and its relationship to members of the family Betaflexiviridae
The nucleotide sequence of Dweet mottle virus (DMV) was determined and compared to sequences of members of the families Alphaflexiviridae and Betaflexiviridae. The DMV genome has 8,747 nucleotides (nt) excluding the 30 poly-(A) tail. DMV... more
The nucleotide sequence of Dweet mottle virus (DMV) was determined and compared to sequences of members of the families Alphaflexiviridae and Betaflexiviridae. The DMV genome has 8,747 nucleotides (nt) excluding the 30 poly-(A) tail. DMV genomic RNA contains three putative open reading frames (ORFs) and untranslated regions of 73 nt at the 50 and 541 nt at 30 termini. ORF1 potentially encoding a 227.48-kDa polyprotein, which has methyltransferase, oxygenase, endopeptidase, helicase, and RNA-dependent RNA polymerase (RdRP) domains. ORF2 encodes a movement protein of 40.25 kDa, while ORF3 encodes a coat protein of 40.69 kDa. Protein database searches showed 98–99% matches of DMV ORFs with citrus leaf blotch virus (CLBV) sequences. Phylogenetic analysis based on the RdRP core domain revealed that DMV is closely related to CLBV as a member of the genus Citrivirus. DMV did not satisfy the molecular criteria for demarcation of an independent species within the genus Citrivirus, family Bet...
Citrus huanglongbing (HLB) disease associated with the ‘Candidatus Liberibacter asiaticus’ (CLas) bacterium has caused significant financial damage to many citrus industries. Large-scale pathogen surveys are routinely conducted in... more
Citrus huanglongbing (HLB) disease associated with the ‘Candidatus Liberibacter asiaticus’ (CLas) bacterium has caused significant financial damage to many citrus industries. Large-scale pathogen surveys are routinely conducted in California to detect CLas early in the disease cycle by lab-based qPCR assays. We have developed an improved reference gene for the sensitive detection of CLas from plants in diagnostic duplex qPCR and analytical digital droplet PCR (ddPCR) assays. The mitochondrial cytochrome oxidase gene (COX), widely used as a reference, is not ideal because its high copy number can inhibit amplification of small quantities of target genes. In ddPCRs, oversaturation of droplets complicates data normalization and quantification. The variable copy numbers of COX gene in metabolically active young tissue, greenhouse plants, and citrus relatives suggest the need for a non-variable, nuclear, low copy, universal reference gene for analysis of HLB hosts. The single-copy nuclea...
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Asiatic citrus canker (ACC) foliar lesions were evaluated on progenies of 84 seed-source genotypes (“parent genotypes”) from the Citrus Variety Collection (CVC) of the University of California at Riverside (UCR) of Citrus trifoliata and... more
Asiatic citrus canker (ACC) foliar lesions were evaluated on progenies of 84 seed-source genotypes (“parent genotypes”) from the Citrus Variety Collection (CVC) of the University of California at Riverside (UCR) of Citrus trifoliata and hybrids between C. trifoliata and other Citrus species and hybrids. Trees were planted Aug. 2013 in a completely randomized design at the Fort Pierce U.S. Department of Agriculture (USDA) grove. Plants were assessed visually Aug. 2017, Sept. 2019, and Sept. 2020 for distinctive ACC lesion incidence and severity. Progeny were compared by parent genotypes using nonparametric analysis. Incidence of ACC [percentage of leaves displaying symptoms, verified by quantitative polymerase chain reaction (qPCR) to be associated with Xanthomonas citri pv. citri] across parent genotypes ranged from 8% to 80% (mean, 49%) of leaves affected in 2017, from 4% to 58% (mean, 29%) in 2019, and 8% to 46% (mean, 25%) in 2020. In 2017, of 49 C. trifoliata parent genotypes, o...
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Citrus tristeza virus (CTV), a member of the aphid-transmitted closterovirus group, is the causal agent of the notorious tristeza disease in several citrus species worldwide. The codon usage patterns of viruses reflect the evolutionary... more
Citrus tristeza virus (CTV), a member of the aphid-transmitted closterovirus group, is the causal agent of the notorious tristeza disease in several citrus species worldwide. The codon usage patterns of viruses reflect the evolutionary changes for optimization of their survival and adaptation in their fitness to the external environment and the hosts. The codon usage adaptation of CTV to specific citrus hosts remains to be studied; thus, its role in CTV evolution is not clearly comprehended. Therefore, to better explain the host–virus interaction and evolutionary history of CTV, the codon usage patterns of the coat protein (CP) genes of 122 CTV isolates originating from three economically important citrus hosts (55 isolate from Citrus sinensis, 38 from C. reticulata, and 29 from C. aurantifolia) were studied using several codon usage indices and multivariate statistical methods. The present study shows that CTV displays low codon usage bias (CUB) and higher genomic stability. Neutra...
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Citrus tristeza virus (CTV, genus ) is one of the most serious pathogens responsible for huge loss of citrus trees worldwide. Four Indian CTV isolates, Kat1 (/Central India), D1 (/North India), B5 (/South India) and G28 (/Northeast India)... more
Citrus tristeza virus (CTV, genus ) is one of the most serious pathogens responsible for huge loss of citrus trees worldwide. Four Indian CTV isolates, Kat1 (/Central India), D1 (/North India), B5 (/South India) and G28 (/Northeast India) collected from different regions of India were characterized based on sequencing of 3' half genome (~ 8.4 kb) comprising 10 open reading frames (ORFs2-11) and 3' UTR and the sequences were submitted to NCBI database as Acc. No KJ914662, HQ912022, HQ912023 and KJ914661, respectively. The present and previously reported Indian isolates Kpg3 and B165 were analyzed and compared with other Asian and international CTV isolates. The Indian CTV isolates had 92-99% nt identities among them. The phylogenetic analysis generated overall ten genogroups/lineages. Of them, all the Asian isolates fell into seven genogroups, whereas the Indian isolates into four. Indian isolates Kat1, D1 and Kpg3 grouped together, termed "Kpg3Gr", along with Flori...
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The paucity of molecular markers limits the application of genetic and genomic research in date palm (Phoenix dactylifera L.). Availability of expressed sequence tag (EST) sequences in date palm may provide a good resource for developing... more
The paucity of molecular markers limits the application of genetic and genomic research in date palm (Phoenix dactylifera L.). Availability of expressed sequence tag (EST) sequences in date palm may provide a good resource for developing gene-based markers. This study characterizes a substantial fraction of transcriptome sequences containing simple sequence repeats (SSRs) from the EST sequences in date palm. The EST sequences studied are mainly homologous to those of Elaeis guineensis and Musa acuminata. A total of 911 gene-based SSR markers, characterized with functional annotations, have provided a useful basis not only for discovering candidate genes and understanding genetic basis of traits of interest but also for developing genetic and genomic tools for molecular research in date palm, such as diversity study, quantitative trait locus (QTL) mapping, and molecular breeding. The procedures of DNA extraction, polymerase chain reaction (PCR) amplification of these gene-based SSR m...
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Mandarin (C. reticulata), citron (C. medica) and pummelo (C. maxima) are important species of the genus Citrus and parents of the interspecific hybrids that constitute the most familiar commercial varieties of citrus: sweet orange, sour... more
Mandarin (C. reticulata), citron (C. medica) and pummelo (C. maxima) are important species of the genus Citrus and parents of the interspecific hybrids that constitute the most familiar commercial varieties of citrus: sweet orange, sour orange, clementine, lemon, lime and grapefruit. Citron produces anthocyanins in its young leaves and flowers, as do species in genera closely related to Citrus, but mandarins do not and pummelo varieties that produce anthocyanins have not been reported. We investigated the activity of the Ruby gene, which encodes a MYB transcription factor controlling anthocyanin biosynthesis, in different accessions of a range of Citrus species and in domesticated cultivars. A white mutant of lemon lacks functional alleles of Ruby, demonstrating that Ruby plays an essential role in anthocyanin production in Citrus. Almost all the natural variation in pigmentation by anthocyanins in Citrus species can be explained by differences in activity of the Ruby gene, caused b...
Research Interests: Pigmentation, Transcription Factors, Biological Sciences, Phylogeny, Plant Physiology, and 13 moreMutation, Flowers, Domestication, Citrus, Anthocyanins, Genotype, Species Specificity, Base Sequence, PLANT PROTEINS, Plant Leaves, alleles, reverse transcriptase polymerase chain reaction, and Retroelements
Citrus huanglongbing (HLB) is a destructive disease with no known cure. To identify sources of HLB resistance in the subfamily Aurantioideae to which citrus belongs, we conducted a six-year field trial under natural disease challenge... more
Citrus huanglongbing (HLB) is a destructive disease with no known cure. To identify sources of HLB resistance in the subfamily Aurantioideae to which citrus belongs, we conducted a six-year field trial under natural disease challenge conditions in an HLB endemic region. The study included 65 Citrus accessions and 33 accessions belonging to 20 other closely related genera. For each accession, eight seedling trees were evaluated. Based on quantitative polymerase chain reaction analysis of the pathogen titers and disease symptoms, eight disease-response categories were identified. We report two immune, six resistant, and 14 tolerant accessions. Resistance and tolerance observed in different accessions may be attributed to a multitude of factors, including psyllid colonization ability, absence of pathogen multiplication, transient replication of the bacterium, lack of pathogen establishment in the plant, delayed infection, or recovery from infection. Most citrus cultivars were considere...
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Citrus surveys were conducted at high (>700 m), medium (300-600 m) and low (<200 m) altitudes in Tanzania in 2014/15. Adults and nymphs of Trioza erytreae (del Guercio) were abundant in the highlands and less abundant at... more
Citrus surveys were conducted at high (>700 m), medium (300-600 m) and low (<200 m) altitudes in Tanzania in 2014/15. Adults and nymphs of Trioza erytreae (del Guercio) were abundant in the highlands and less abundant at medium altitudes. Unexpectedly, adults and nymphs of Diaphorina citri Kuwayama, the Asian citrus psyllid, were found and collected at medium altitudes, around Morogoro. No psyllids were observed at low altitudes. Severe huanglongbing symptoms and tree decline were evident at high altitudes, while mild and few symptoms were observed at intermediate and low altitudes, respectively. DNA was extracted from leaf and psyllid samples and subjected to conventional PCR (cPCR) with seven different primer sets and RT qPCR with two primer sets. cPCR bands were sequenced and subjected to phyloge-netic analysis. Candidatus Liberibacter africanus (Laf) was detected in highland leaf and T. erytreae samples from high and medium altitudes by all methods. Sequences from leaves and psyllids were similar to those from South Africa. Candidatus Liberibacter asiaticus (Las) was detected by qPCR in medium altitude leaf samples, but cross-reaction with Laf was likely because presence of Las was not confirmed by cPCR and sequencing. Neither Laf nor Las were detected in D. citri samples. This is the first reported occurrence of D. citri in Africa. Predictions were made of the potential distribution of D. citri and Las in Africa and along the Mediterranean coast using the correlative models MAXENT and Multi-Model Framework. Additional surveys at medium and low altitudes and quarantine measures are recommended.
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Citrus huanglongbing (HLB) has become a major disease and limiting factor of production in the citrus areas that have become infected. The destruction to the affected citrus industries has resulted in a tremendous increase to support... more
Citrus huanglongbing (HLB) has become a major disease and limiting factor of production in the citrus areas that have become infected. The destruction to the affected citrus industries has resulted in a tremendous increase to support research that in return has resulted in significant information on both applied and basic knowledge concerning this important disease to the global citrus industry. Recent research indicates the relationship between citrus and the causal agent of HLB is shaped by multiple elements, in which host defense responses may also play an important role. This review is intended to provide an overview of the importance of HLB to a wider audience of plant biologists. Recent advances on host pathogen interactions, population genetics, and vectoring of the causal agent are discussed.
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ABSTRACT Citron (Citrus medica L.) constitutes a commercially important group of Citrus in many parts of the world. It is one of the three original species from which most cultivated citrus accessions are derived. Morphologically, citron... more
ABSTRACT Citron (Citrus medica L.) constitutes a commercially important group of Citrus in many parts of the world. It is one of the three original species from which most cultivated citrus accessions are derived. Morphologically, citron fruits are diverse ranging from small ellipsoid to gigantic ellipsoid to elaborate fingered types. We have studied the genetic variability of 48 citrons collected from various parts of the world. Thirty accessions were sampled from Yunnan province of China, part of the region where Citrus medica originated and maximum diversity of citrons exist in the wild. Eighteen citrons were sampled from the Citrus Variety Collection in Riverside, CA, and were originally obtained from different parts of the world. Citrons are known to be mono-embryonic, and previous studies have generally found little heterozygosity. For our study, we obtained data from 23 microsatellite markers distributed throughout the genome. We recorded 97 alleles and an average of 4.21 alleles per locus. Only 7 samples were homozygous in our study. 24 samples showed 5-20% heterozygosity, 15 samples had 22-52% heterozygosity. A distance matrix was created using a Microsatellite analyzer software (Nei’s standard genetic distance algorithm) and Neighbor joining trees were constructed. The 48 citrons clustered into 3 distinct groups with an average genetic distance ranging from 0.3-0.4 between the groups. Cluster A had citrons from Mediterranean and South American countries; cluster B had fingered and some non-fingered citrons indigenous to the Yunnan province; Cluster C had some wild citrons from China. Similar results were obtained using population STRUCTURE analysis.
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DNA barcoding facilitates proper and easy identification of species. Two chloroplast genes, rbcL and matK have been recognized as potentially ideal candidates for barcoding of plants. The sub-family Aurantioideae, including citrus, is... more
DNA barcoding facilitates proper and easy identification of species. Two chloroplast genes, rbcL and matK have been recognized as potentially ideal candidates for barcoding of plants. The sub-family Aurantioideae, including citrus, is known to have a high level of hybridization and incomplete lineage sorting resulting in maintenance of ancestral polymorphisms. Use of uniparentally inherited markers for barcoding will not be adequate to identify this biologically complex taxonomic group. We have used a low copy nuclear gene, malate dehydrogenase (MDH), as a barcoding gene to identify members of Aurantioideae. Sequences of a 1600 bp fragment of MDH were generated using vouchered specimens from 29 genera and 120 cultivars of Aurantioideae. The region selected has several introns providing the variability needed to distinguish closely related cultivars. We have identified regions that are short enough for easy sequencing and variable enough to provide discrimination. A two tiered approa...
Citron (Citrus medica L.) is believed to have originated in China. The wild citrons in China exhibit considerable phenotypic diversity. Our dataset for this study had 50 accessions including exotic fingered citrons, over-sized citrons and... more
Citron (Citrus medica L.) is believed to have originated in China. The wild citrons in China exhibit considerable phenotypic diversity. Our dataset for this study had 50 accessions including exotic fingered citrons, over-sized citrons and other local varieties collected mainly from the Yunnan province of China in addition to representative citrons from various parts of the world, available at the Citrus Variety Collection, Riverside, CA. We conducted molecular analysis using simple sequence repeat (SSR) markers and single nucleotide polymorphisms (SNP), two types of co-dominant markers. SSRs distributed throughout the genome were studied by PCR amplification and gel electrophoresis to detect polymorphic alleles. We tested 37 SSR markers and identified 23 that could discriminate among the different taxa. Seventy five alleles were identified and the band sizes obtained for the various accessions in the study were recorded. Citrons are generally homozygous but we found considerable het...
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Citrus psorosis virus (CPV) is a multicomponent ssRNA virus with a coat protein of approximately 48 kDa. The viral genome is encapsidated in short and long particles that are readily separated by sucrose density-gradient centrifugation.... more
Citrus psorosis virus (CPV) is a multicomponent ssRNA virus with a coat protein of approximately 48 kDa. The viral genome is encapsidated in short and long particles that are readily separated by sucrose density-gradient centrifugation. CPV particles are spiral filaments that are referred to as spiroviruses (SV). A cDNA library of purified short particles from isolate CPV-4 was prepared in a Lambda vector and screened for expression of the coat protein gene (CPG) with a monoclonal antibody to the coat protein. Sequencing of immunopositive clones indicated a single ORF encoding a 49 kDa protein. This ORF, when expressed in E. coli, gave a protein identical in size and immunoreactivity to the CPV coat protein. A full-length clone of the CPG was transcribed and used in Northern hybridization assays to establish that short particle RNA of CPV is negative sense and contains the CPG. Moreover, the CPG was not found on RNA extracted from long particles or on the sedimentable dsRNA from CPV...
Research Interests: Biological Sciences, General, Nucleic acid hybridization, Polymerase Chain Reaction, Capsid, and 11 moreCitrus, Density gradient, Molecular cloning, Monoclonal Antibody, Amino Acid Sequence, Base Sequence, Full Length Movies, cDNA library, Molecular Sequence Data, Nucleotide sequence, and Medical and Health Sciences
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Tristeza, caused by Citrus tristeza virus (CTV), has long been present in Florida but outbreaks of decline on sour orange rootstock were occasional events until the late 1970s. Sour orange rootstock was valued for the high quality of... more
Tristeza, caused by Citrus tristeza virus (CTV), has long been present in Florida but outbreaks of decline on sour orange rootstock were occasional events until the late 1970s. Sour orange rootstock was valued for the high quality of fruit produced and was widely used because of its tolerance of citrus blight, a disease of unknown etiology. Research was directed towards the selection and screening of mild strains of CTV which could protect against sour orange decline strains. Following the introduction of Toxoptera citricida (also known as the brown citrus aphid) in 1995 there was a greater concern for maintaining production of existing blocks of citrus on sour orange rootstock. Availability of the CTV genome sequence around the same time as well as molecular characterization of in planta CTV populations led to the selection of mild CTV isolates which when inoculated into existing field trees, extended the productive life of the groves and enabled a more graduate replanting of trees...
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The potato psyllid, Bactericera cockerelli (Sulc) (Hemiptera: Triozidae), is a serious pest of potatoes that can cause yield loss by direct feeding on crop plants and by transmitting a bacterial pathogen. Current pest management practices... more
The potato psyllid, Bactericera cockerelli (Sulc) (Hemiptera: Triozidae), is a serious pest of potatoes that can cause yield loss by direct feeding on crop plants and by transmitting a bacterial pathogen. Current pest management practices in potatoes rely on the use of insecticides for control of the potato psyllid to lower disease incidences and increase yields. While many studies have focused on psyllid mortality caused by insecticides, little is known regarding the behavioral responses of the potato psyllid to insecticides or if insecticides can decrease pathogen transmission. Thus, the objectives of this study were 1) to determine and characterize the effects of imidacloprid on adult potato psyllid feeding behavior through direct observations and with electrical penetration graphs (EPGs); 2) determine the residual effects of imidacloprid on potato psyllid feeding behavior over time; and 3) determine the effects of imidacloprid on transmission of Candidatus Liberibacter psyllauro...
The draft whole-genome sequence of “ Candidatus Liberibacter solanacearum” strain R1, isolated from and maintained in tomato plants in California, is reported. The R1 strain has the genome size of 1,204,257 bp, G+C content of 35.3%, 1,101... more
The draft whole-genome sequence of “ Candidatus Liberibacter solanacearum” strain R1, isolated from and maintained in tomato plants in California, is reported. The R1 strain has the genome size of 1,204,257 bp, G+C content of 35.3%, 1,101 predicted open reading frames, and 57 RNA genes.
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Twenty-one Citrus tristeza virus (CTV) isolates from India were characterized, using genotype-specific multiple molecular markers (MMM) from the... more
Twenty-one Citrus tristeza virus (CTV) isolates from India were characterized, using genotype-specific multiple molecular markers (MMM) from the 54'-terminal region and two other overlapping primer pairs (CN487/489 and CN488/491) from ORF1a (697-1484 nucleotides (nt)). The 5'-terminal genotype-specific primer pairs amplified about 500 bases from the 5'-end of the CTV genomic RNA (gRNA). With the three different MMM, the VT genotype-specific primers amplified 19 Indian CTV isolates. The T30-specific primers amplified five isolates, and the T36 primer amplified only one isolate T36. All isolates were amplified with CN488/491 primers; however, only 20 isolates were amplified with CN487/489 pair. A phylogenetic tree, derived from the sequences of the different MMM primer-amplified products, placed all the isolates into four distinct genogroups. Three of these four groups were typified by the reference isolates T30, T36, and VT. The fourth group, represented by the isolate BAN-2, was considered as a new genogroup. A phylogenetic tree based on sequences of the CN487/491 amplified products and other published sequences placed all of the isolates in eight genogroups. Phylogenetic correlation over the three different regions sequences of these CTV isolates showed more sequence variability between 1082 and 1484nt than between 1 and 500 or 697-1105 nt of the CTV gRNA. Based on three different 5' regions sequences and phylogenetic analysis, it is hypothesized that isolates BAN-1, BAN-2, and B165 are three naturally occurring variants that add to the complexity of the CTV populations in India.
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Research Interests: Portugal, Biological Sciences, Plant diseases, Animals, Aphids, and 15 moreFlorida, Trees, Citrus, Immunoassay, Citrus Tristeza Virus, Amino Acid Sequence, Sensitivity and Specificity, Disease Management, Mediterranean Basin, Epitopes, Closterovirus, Aphid, Insect Vectors, Molecular Sequence Data, and reverse transcriptase polymerase chain reaction
The viral replicase complex of positive-stranded RNA viruses interacts with cis-acting elements that are usually located at the termini of the viral RNAs. On comparison of the replication requirement of a tobacco mosaic virus (TMV)-based... more
The viral replicase complex of positive-stranded RNA viruses interacts with cis-acting elements that are usually located at the termini of the viral RNAs. On comparison of the replication requirement of a tobacco mosaic virus (TMV)-based defective RNA (dRNA) and its helper virus, we found different requirements for replication of TMV RNAs in cis and in trans. The level of replication of full-length TMV RNA decreased substantially in the absence of pseudoknot (pk) 1 and/or 2, whereas identical deletions in dRNAs did not affect their replication. However, pk3 was required for replication of both full-length TMV RNAs and dRNAs. The requirements for homologous sequences were greater for dRNA replication than for replication of full-length TMV RNAs. Defective RNAs with heterologous 3' nontranslated regions (NTRs) failed to be replicated or replicated minimally, whereas replication of similarly mutated full-length RNAs was much less affected. Increasing amounts of contiguous heterologous sequences in the dRNAs compensated for the impaired interactions between the replicase and 3' NTR. The precision requirement appeared to involve the terminal 28 nucleotides, specifically the pseudoknot in the aminoacyl acceptor arm of the tRNA like structure, which was important in replication of both dRNAs and full-length TMV RNAs.
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Citrus exocortis viroid (CEVd) exists as populations of heterogeneous variants in infected hosts. In vivo generated CEVd progeny variants (CEVd-PVs) populations from citrus protoplasts, seedlings and mature plants, following inoculation... more
Citrus exocortis viroid (CEVd) exists as populations of heterogeneous variants in infected hosts. In vivo generated CEVd progeny variants (CEVd-PVs) populations from citrus protoplasts, seedlings and mature plants, following inoculation with transcripts of a single CEVd cDNA-clone (wild-type, WT), were studied. The CEVd-PVs population in protoplasts was heterogeneous and became progressively more homogeneous in seedlings and mature plants. The infectivity and pathogenicity of selected CEVd-PVs was evaluated in citrus and herbaceous experimental hosts. The CEVd-PVs U30C, G128A and U182C were not infectious; G50A and 108U+ were infectious but reverted back to WT and 62A+, U129A and U278A were infectious, genetically stable and more severe than WT. The 62A+ and U278A and U129A accumulated at higher levels than WT in protoplasts and seedlings respectively. The effect of specific mutations on the predicted secondary structure of the CEVd-PVs' RNA coupled with the infectivity and replication studies suggested complex structure-to-function relationships for CEVd.
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Research Interests: Multidisciplinary, Western blotting, Escherichia coli, Monoclonal Antibodies, Capsid, and 12 morePlant viruses, Molecular cloning, Molecular Characterization, Western blot, Citrus Tristeza Virus, Monoclonal Antibody, Amino Acid Sequence, Recombinant Proteins, Epitopes, Site-directed Mutagenesis, Nucleotides, and Molecular Sequence Data
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The citrus industries of North and South America are endangered by Huanglongbing (HLB), also known as citrus greening, a devastating disease associated with ‘Candidatus Liberibacter asiaticus’ and ‘Ca. L. americanus’, two species of... more
The citrus industries of North and South America are endangered by Huanglongbing (HLB), also known as citrus greening, a devastating disease associated with ‘Candidatus Liberibacter asiaticus’ and ‘Ca. L. americanus’, two species of fastidious phloem-limited bacteria spread by the Asian citrus psyllid (ACP), Diaphorina citri, Kuwayama. The first reports of HLB from the Americas were from Brazil in 2004 followed by Florida in 2005 (3). The ACP was found in Belize in 2005 (S. Williams, personal communication) and is now present throughout Central America. On the basis of the report that the HLB-associated bacteria can be easily detected in the ACP vector (4), an initial sampling of ACP from 67 locations was collected in February 2009 from trees in the Belize, Corozal, Orange Walk, Stann Creek, and Toledo Districts of Belize, and shipped in 95% ethanol to Riverside, CA for analysis. DNA was extracted from lots containing three to five psyllids from each of the 67 samples with Fast DNA ...
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Antibodies specific for the recombinant coat protein (rCP) of the p25 gene of Citrus tristeza virus (CTV) were developed in goats and rabbits and further evaluated as a complete kit for the detection of the virus using healthy and... more
Antibodies specific for the recombinant coat protein (rCP) of the p25 gene of Citrus tristeza virus (CTV) were developed in goats and rabbits and further evaluated as a complete kit for the detection of the virus using healthy and CTV-infected tissue. The combination of goat T1 used as primary (coating) and rabbit C3 as intermediate (detecting) rCP antibodies reacted efficiently, with optical density at 405 nm (OD405) values between 0.250 and 2.000 with samples from an international collection of diverse CTV isolates. The CTV isolates tested cause a broad spectrum of disease syndromes in different citrus hosts. The OD405 values for healthy tissue were less than 0.100. Likewise, the combination of goat T1 and rabbit C3 rCP antibodies gave consistent results for CTV-positive and -negative sample discrimination when directly compared with the Central California Tristeza Eradication Agency (CCTEA) antibodies used for large-scale CTV detection and a commercially available CTV serological...
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The incidence of Citrus tristeza virus (CTV) was found to increase significantly in southern Florida within 2 years after the establishment of its most efficient vector, Toxoptera citricida (Kirkaldy). Increased incidence of both mild and... more
The incidence of Citrus tristeza virus (CTV) was found to increase significantly in southern Florida within 2 years after the establishment of its most efficient vector, Toxoptera citricida (Kirkaldy). Increased incidence of both mild and severe strains was documented, with the incidence of severe strains increasing more than mild strains. Molecular probes capable of differentiating mild, quick decline and various types of stem-pitting strains demonstrated that trees often were infected with more than one strain of CTV, with trees containing up to five different strains. Some CTV strains detected in the southeast urban corridor of Florida and in commercial groves in southwest Florida were found to react with probes specific for stem-pitting strains known from elsewhere in the world. The implications of the presence of these CTV strains in Florida and their possible presence in citrus budwood scion trees are discussed.
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'Candidatus Liberibacter asiaticus' (CLas) is a Gram-negative α-proteobacterium, and the prominent species of Liberibacter associated with a devastating worldwide citrus disease known as huanglongbing (HLB). This fastidious bacterium resides in phloem sieve cells of host plants and is vectored by the Asian citrus psyllid (Diaphorina citri). Due to its uneven distribution in planta and highly variable bacterial titers, detection of HLB bacteria can be challenging. Here we demonstrated a new utility of nearly identical tandem-repeats of two CLas prophage genes for real-time PCR by SYBR Green 1 (LJ900fr) and TaqMan(®) (LJ900fpr). When compared with conventional 16S rDNA-based real-time PCR, targeting the repeat sequence reduced the relative detectable threshold by approximately 9 and 3 real-time PCR cycles for LJ900fr and LJ900fpr, respectively. Additionally, both LJ900 methods detected CLas from otherwise non-detectable samples by other methods. CLas was also detected from globally derived samples including psyllids, various citrus varieties, periwinkle, dodder, and orange jasmine, suggesting the new detection method can be applicable worldwide. Additionally, we demonstrated the presence of the hyv(I)/hyv(II) repeat sequence within the 'Ca. Liberibacter americanus' strain. The method thereby provides sensitive HLB detection with broad application for scientific, regulatory, and citrus grower communities.
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... Rocha-Peña, unpublished. MX14, Tamaulipas, México, Severe isolate, similar to B247 stem pitting in sweet orange from Venezuela, +, Iracheta-Cárdenas et al., 2002; Silva-Vara et al., 2001; Rocha-Peña, unpublished. B227, India, ...
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Research Interests: Zoology and Regulations
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Research Interests: Pythium, Biological Sciences, Sequence alignment, Polymerase Chain Reaction, Species identification, and 13 morePlant Roots, Phytophthora, Capsicum, Plant Pathogen, Species Specificity, Base Sequence, Sensitivity and Specificity, Ribosomal DNA, Enzyme Linked Immunosorbent Assay, Lycopersicon esculentum, Molecular Sequence Data, Internal transcribed spacer, and Medical and Health Sciences
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The potato psyllid, Bactericera cockerelli (Sulc) (Hemiptera: Triozidae), is a major pest of potatoes that can cause yield loss by direct feeding on crop plants and by transmitting a bacterial pathogen, Candidatus Liberibacter psyllaurous... more
The potato psyllid, Bactericera cockerelli (Sulc) (Hemiptera: Triozidae), is a major pest of potatoes that can cause yield loss by direct feeding on crop plants and by transmitting a bacterial pathogen, Candidatus Liberibacter psyllaurous (a.k.a. Candidatus Liberibacter solanacearum) associated with zebra chip disease of the crop. In recent years, there have been no studies regarding resistance of potato to the
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Immunocapture reverse transcription coupled with PCR (RT-PCR). The CTV particles adsorbed from infected tissue to the ELISA plates were used for RT-PCR, using a modification of the procedure by Noiasco et al (18). After reading the color... more
Immunocapture reverse transcription coupled with PCR (RT-PCR). The CTV particles adsorbed from infected tissue to the ELISA plates were used for RT-PCR, using a modification of the procedure by Noiasco et al (18). After reading the color reac-tions, the plates were washed once ...