Report of Citrus tristeza virus in Diaphorina citri (Hemiotera: Liviidae) insects of different sexes, color morphs, and developmental stages.

Diaphorina citri, also known as the Asian citrus psyllid, is the main vector of 'Candidatus Liberibacter asiaticus' (CLas) associated with citrus Huanglongbing. It has been reported that D. citri could also be infected by Citrus tristeza virus (CTV), a virus that has been previously reported to be vectored by certain aphid species. In this study, the CTV and CLas profiles in different organs, color variants, developmental stages, or sexes of D. citri insects were analyzed. Although no significant differences were found between nymphs and adults in CTV titers, we found that the third instar nymph of D. citri was more efficient in CTV and CLas acquisition compared to the fourth and fifth instars and adults. With the instars of D. citri development, the relationship between the acquiring of CTV and CLas by D. citri seemed to follow an inverse trend, with the titer of CLas increased and the titer of CTV decreased. No significant differences were observed between the 2 sexes of D. citri in acquiring either CTV or CLas titers in the field. However, no differences were drawn among the 3 color morph variants for CTV titers. CTV titers in the midguts of adult D. citri were significantly higher than those in the salivary glands. Both CTV-positive incidence and CTV titers in the midguts of adult D. citri increased with increasing exposure periods. This study provides new data to deepen our understanding of the CTV-involved interaction between D. citri and CLas.

Validation of High-Throughput Sequencing (HTS) for Routine Detection of Citrus Viruses and Viroids.

The management of plant diseases relies on the accurate identification of pathogens that requires a robust and validated tool in terms of specificity, sensitivity, repeatability, and reproducibility. High-throughput sequencing (HTS) has become the method of choice for virus detection when either a complete viral status of a plant is required in a single assay or if an unknown viral agent is expected. To ensure that the most accurate diagnosis is made from an HTS data analysis, a standardized protocol per pathosystem is required. This chapter presents a detailed protocol for the detection of viruses and viroids infecting citrus using HTS. The protocol describes all the steps from sample processing, nucleic acid extraction, and bioinformatic analyses validated to be an efficient method for detection in this pathosystem. The protocol also includes a section on citrus tristeza virus (CTV) genotype differentiation using HTS data.

Stem-pitting caused by Citrus tristeza virus is associated with increased phloem occlusion.

Stem-pitting (SP) disease results from disruption of normal phloem and xylem development. In citrus, a characteristic manifestation of SP caused by Citrus tristeza virus (CTV) is phloem regeneration. We hypothesized that phloem regeneration occurs due to reduced functionality of CTV infected phloem cells. To examine phloem cell occlusions in CTV-SP, we analyzed callose and phloem-protein (PP) accumulation in Citrus macrophylla trees infected with CTV mutants exhibiting different SP phenotypes from very mild (CTVΔp13) to severe (CTVΔp33), in addition to full-length CTV and healthy plants. CTV infection was accompanied by callose and PP accumulation in the phloem. With the increase in the SP symptoms from very mild to severe, there was a constant increase in the levels of callose and PP, accompanied by an increase in PHLOEM-PROTEIN 2 and a decrease in BETA-1,3-GLUCANASE gene expression levels. These results indicate that SP symptom development is associated with increased phloem occlusion.

Bayesian phylodynamic analysis reveals the evolutionary history and the dispersal patterns of citrus tristeza virus in China based on the p25 gene.

BACKGROUND: Citrus tristeza virus (CTV) is one of the most serious threats to the citrus industry, and is present in both wild and cultivated citrus. The origin and dispersal patterns of CTV is still poorly understood in China. METHODS: In this study, 524 CTV suspected citrus samples from China were collected, including 354 cultivated citrus samples and 174 wild citrus samples. Finally, 126 CTV coat protein sequences were obtained with time-stamped from 10 citrus origins in China. Bayesian phylodynamic inference were performed for CTV origin and dispersal patterns study in China. RESULT: We found that CTV was mainly distributed in southern and coastal areas of China. The substitution rate of CTV was 4.70 × 10- 4 subs/site/year (95% credibility interval: 1.10 × 10- 4 subs/site/year ~ 9.10 × 10- 4 subs/site/year), with a slight increasing trend in CTV populations between 1990 and 2006. The CTV isolates in China shared a most common recent ancestor around 1875 (95% credibility interval: 1676.57 ~ 1961.02). The CTV in China was originated from wild citrus in Hunan and Jiangxi, and then spread from the wild citrus to cultivated citrus in the growing regions of Sichuan, Chongqing, Hubei, Fujian, Zhejiang, Guangxi and Guangdong provinces. CONCLUSIONS: This study has proved that CTV in China was originated from wild citrus in Hunan and Jiangxi. The spatial-temporal distribution and dispersal patterns has uncovered the population and pandemic history of CTV, providing hints toward a better understanding of the spread and origin of CTV in China.

Variation of endosymbiont and citrus tristeza virus (CTV) titers in the Huanglongbing insect vector, Diaphorina citri, on CTV-infected plants.

"Candidatus Liberibacter asiaticus" (CLas) is a notorious agent that causes Citrus Huanglongbing (HLB), which is transmitted by Diaphorina citri (D. citri). We recently found that the acquisition and transmission of CLas by D. citri was facilitated by Citrus tristeza virus (CTV), a widely distributed virus in the field. In this study, we further studied whether different CTV strains manipulate the host preference of D. citri, and whether endosymbionts variation is related to CTV strains in D. citri. The results showed that the non-viruliferous D. citri preferred to select the shoots infected with CTV, without strain differences was observed in the selection. However, the viruliferous D. citri prefered to select the mixed strain that is similar to the field's. Furthermore, D. citri effectively acquired the CTV within 2-12 h depending on the strains of the virus. The persistence period of CTV in D. citri was longer than 24 days, without reduction of the CTV titers being observed. These results provide a foundation for understanding the transmission mode of D. citri on CTV. During the process of CTV acquisition and persistence, the titers of main endosymbionts in D. citri showed similar variation trend, but their relative titers were different at different time points. The titers of the "Candidatus Profftella armatura" and CTV tended to be positively correlated, and the titers of Wolbachia and "Candidatus Carsonella ruddii" were mostly negatively related with titers of CT31. These results showed the relationship among D. citri, endosymbionts, and CTV and provided useful information for further research on the interactions between D. citri and CLas, which may benefit the development of approaches for the prevention of CLas transmission and control of citrus HLB.

Transcriptomic alterations in the sweet orange vasculature correlate with growth repression induced by a variant of citrus tristeza virus.

Citrus tristeza virus (CTV, family Closteroviridae) is an economically important pathogen of citrus. CTV resides in the phloem of the infected plants and induces a range of disease phenotypes, including stem pitting and quick decline as well as a number of other deleterious syndromes. To uncover the biological processes underlying the poorly understood damaging symptoms of CTV, we profiled the transcriptome of sweet orange (Citrus sinensis) phloem-rich bark tissues of non-infected, mock-inoculated trees and trees singly infected with two distinct variants of CTV, T36 or T68-1. The T36 and T68-1 variants accumulated in the infected plants at similar titers. With that, young trees infected with T68-1 were markedly repressed in growth, while the growth rate of the trees infected with T36 was comparable to the mock-inoculated trees. Only a small number of differentially expressed genes (DEGs) were identified in the nearly asymptomatic T36-infected trees, whereas almost fourfold the number of DEGs were identified with the growth-restricting T68-1 infection. DEGs were validated using quantitative reverse transcription-PCR. While T36 did not induce many noteworthy changes, T68-1 altered the expression of numerous host mRNAs encoding proteins within significant biological pathways, including immunity and stress response proteins, papain-like cysteine proteases (PLCPs), cell-wall modifying enzymes, vascular development proteins and others. The transcriptomic alterations in the T68-1-infected trees, in particular, the strong and persistent increase in the expression levels of PLCPs, appear to contribute to the observed stem growth repression. On the other hand, analysis of the viral small interfering RNAs revealed that the host RNA silencing-based response to the infection by T36 and that by T68-1 was comparable, and thus, the induction of this antiviral mechanism may not contribute to the difference in the observed symptoms. The DEGs identified in this study promote our understanding of the underlying mechanisms of the yet unexplained growth repression induced by severe CTV isolates in sweet orange trees.

Citrus tristeza virus Promotes the Acquisition and Transmission of 'Candidatus Liberibacter Asiaticus' by Diaphorina citri.

Diaphorina citri Kuwayama (D. citri) is an insect vector of phloem-limited 'Candidatus Liberibacter asiatus' (CLas), the presumed pathogen of citrus Huanglongbing (HLB). Recently, our lab has preliminarily found it acquired and transmitted Citrus tristeza virus (CTV), which was previously suggested to be vectored by species of aphids. However, the influences of one of the pathogens on the acquisition and transmission efficiency of the other pathogen remain unknown. In this study, CLas and CTV acquisition and transmission by D. citri at different development stages under field and laboratory conditions were determined. CTV could be detected from the nymphs, adults, and honeydew of D. citri but not from the eggs and exuviates of them. CLas in plants might inhibit CTV acquisition by D. citri as lower CTV-positive rates and CTV titers were detected in D. citri collected from HLB-affected trees compared to those from CLas-free trees. D. citri were more likely to obtain CTV than CLas from host plants co-infected with the two pathogens. Intriguingly, CTV in D. citri facilitated the acquisition and transmission of CLas, but CLas carried by D. citri had no significant effect on the transmission of CTV by the same vector. Molecular detection and microscopy methods confirmed the enrichment of CTV in the midgut after a 72-h acquisition access period. Collectively, these results raise essential scientific questions for further research on the molecular mechanism of pathogen transmission by D. citri and provide new ideas for the comprehensive prevention and control of HLB and CTV.

Long Noncoding RNAs in Plant-Pathogen Interactions.

Long noncoding RNAs (lncRNAs) are commonly defined as transcripts that lack protein-coding capacity and are longer than 200 nucleotides. Since the emergence of next-generation sequencing technologies in this century, thousands of lncRNAs have been identified from nearly all living organisms. Notably, various pathogens also express their own lncRNAs in host cells during infection. In plants, many lncRNAs exhibit dynamic expression patterns in response to environmental stimuli, including pathogen attacks. In contrast to well-established methods in identifying such lncRNAs, the current understanding of lncRNAs' functional mechanisms is in its infancy. Some lncRNAs serve as precursors for generating small RNAs or serve as target mimics to sequester functional small RNAs, which have been extensively reviewed in the literature. This review focuses on the emerging evidence supporting that certain lncRNAs function as negative or positive regulators of plant immunity. A common theme is that those regulations rely on specific interactions between lncRNAs and key regulatory proteins. Viroids as single-stranded circular noncoding RNAs provide a handle to investigate how RNA local motifs render interaction specificity between lncRNAs and regulatory proteins. [Formula: see text] Copyright © 2023 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Saúde mental de puérperas durante a pandemia de Covid-19 no Brasil / Postpartum women mental health during the Covid-19 pandemic in Brazil / Salud mental de mujeres posparto durante la pandemia de Covid-19 en Brasil

Estudos internacionais têm reportado níveis elevados de sintomas de depressão e ansiedade em mulheres no período perinatal em resultado da atual pandemia. O presente estudo avaliou a sintomatologia de depressão e ansiedade em puérperas durante a pandemia de COVID-19 no Brasil. Participaram 625 mulheres, com idades entre 18 e 44 anos (M = 31.6; DP = 5.3), que tinham um bebê até 6 meses de idade. Foram administrados o Questionário de Transtorno de Ansiedade Generalizada (GAD-7) e a Escala de Depressão Pós-Natal de Edimburgo (EPDS). Os resultados mostraram níveis clinicamente significativos de depressão (EPDS ≥13) em 47.4% das participantes, níveis clinicamente significativos de ansiedade generalizada (GAD-7 ≥10) em 41.8% dos casos, e sintomas comórbidos em 33.1% da amostra. Registrou-se uma correlação positiva significativa entre os sintomas de depressão e ansiedade. Além disso, mais dias de vida do bebê, idade mais jovem da mãe e menor nível de escolaridade estavam associados a níveis potencialmente clínicos de sintomas. Assim, é prioritária a definição de programas de prevenção e intervenção na saúde mental perinatal durante o atual período pandêmico, com continuidade para o futuro.

International studies have reported high levels of depression and anxiety symptoms in perinatal women due to the ongoing pandemic. The present study examined symptoms of depression and anxiety in postpartum women during the COVID-19 pandemic in Brazil. Participants were 625 women, aged between 18 and 44 years (M = 31.6; SD = 5.3), who had an infant up to 6 months of age. The Generalized Anxiety Disorder Questionnaire (GAD-7) and the Edinburgh Postnatal Depression Scale (EPDS) were administered. Results showed clinically significant levels of depression (EPDS ≥13) in 47.4% of the participants, clinically significant levels of generalized anxiety (GAD-7 ≥10) in 41.8% of the cases, and comorbid symptoms in 33.1% of the participants. There was a significant positive correlation between symptoms of depression and anxiety. Furthermore, infant's older age, mother's younger age, and lower educational level were associated with potentially clinical levels of symptoms. Thus, prevention and intervention programs targeting perinatal mental health during the ongoing pandemic and beyond should be developed and prioritized.

Estudios internacionales han reportado altos niveles de síntomas de depresión y de ansiedad en mujeres em el período perinatal como consecuencia de la actual pandemia. El presente estudio examinó los síntomas de depresión y ansiedad en mujeres posparto durante la pandemia de COVID-19 en Brasil. Las participantes fueron 625 mujeres, con edades entre 18 y 44 años (M = 31.6; SD = 5.3), que tenían un hijo de hasta 6 meses de edad. Se administró el Cuestionario de Trastorno de Ansiedad Generalizada (GAD-7) y la Escala de Depresión Postnatal de Edimburgo (EPDS). Los resultados mostraron niveles clínicamente significativos de depresión (EPDS ≥13) en el 47.4 % de las participantes, niveles clínicamente significativos de ansiedad generalizada (GAD-7 ≥10) en el 41.8 % de los casos y síntomas comórbidos en el 33.1 % de las participantes. Hubo una correlación positiva significativa entre los síntomas de depresión y ansiedad. Además, más días de vida del bebé, menor edad de la madre y menor nivel educativo se asociaron con niveles potencialmente clínicos de síntomas. Por lo tanto, se debe priorizar la definición de programas de prevención e intervención dirigidos a la salud mental perinatal durante la pandemia en curso, con continuidad para el futuro.

Simple template-based reverse transcription-recombinase polymerase amplification assay for routine diagnosis of citrus tristeza virus.

This study reports a simple template-based reverse transcription-polymerase amplification assay (ST-RT-RPA) for detection of citrus tristeza virus (CTV) from crude plant extract lysed in NaOH:EDTA (1:1) without the need of tedious RNA isolation. The developed assay showed versatility in its usage as amplification can be performed at wide temperature range (14°C to 42°C) and incubation time (4 to 32 min), although the best conditions were 38°C for 30 min. The developed ST-RT-RPA assay could detect the CTV up to 10-8 dilution of crude plant extract of NaOH:EDTA and up to 0.01 fg µl-1 of RNA of CTV-infected plant tissues and 0.001 ag µl-1 of plasmid DNA containing viral insert, thus exhibiting sufficient sensitivity. ST-RT-RPA assay showed high specificity without any cross-reaction with other citrus pathogens (Indian citrus ringspot virus, citrus yellow mosaic virus, citrus yellow vein clearing virus, and Candidatus Liberibacter asiaticus) and was more sensitive in detection of CTV infection in field samples as compared to standard reverse transcription-polymerase chain reaction (RT-PCR) with later showing false negative in 7.92% of samples tested after 1 week of sampling. The developed ST-RT-RPA assay used minimally processed crude plant extract as template, tolerant to sample degradation in transit and storage, while it can be easily performed at wide temperatures and could be adopted in resource-poor setup.

Survey and detection for citrus tristeza virus in Florida groves with an unconventional tool: The Asian citrus psyllid.

The citrus industry of Florida faces insurmountable challenges against the destructive diseases citrus tristeza and Huanglongbing (HLB, or citrus greening). Though the tristeza causal agent, citrus tristeza virus (CTV), has been in Florida decades longer than HLB, growers have concentrated most of their efforts on combating the more detrimental HLB. The Asian citrus psyllid (Diaphorina citri; ACP) is the insect vector of the bacterial pathogen Candidatus Liberibacter asiaticus and transmits the incurable HLB to all commercial citrus. During our searches for biological and viral controls against the ACP, we consistently detected sequences of CTV in Florida field populations of ACP. This unexpected finding led us to investigate whether ACPs collected from young shoots could be used as a tool to survey CTV in Florida citrus groves. We first surveyed for the most common CTV strains in Florida (T30, T36, and VT/T68) in citrus trees on mostly sour orange (Citrus aurantium) rootstock, the rootstock susceptible to CTV decline. Out of 968 trees sampled across five years (2018-2022), approximately 8.2% were positive for CTV, with more than half of the CTV-positive trees infected with strain T30. Simultaneously, we looked at CTV strains in ACPs during this time and found that approximately 88% of pooled adult and nymph ACPs also had CTV, with over half the positive samples having the T36 strain. As a result of the much higher CTV incidences in the ACPs, we conducted a second investigation into whether we could more easily detect the same CTV strains in ACP nymphs as in CTV-infected citrus tissue. After individually sampling 43 trees and pooling the nymphs from each tree, we detected CTV at about the same incidence in the citrus tissue and the nymphs, but with much less ACP tissue, time, and resources required for detection compared to citrus tissue. Results from this study illustrate the sustained threat of CTV to Florida citrus and demonstrate the ACP as a potential bioindicator for CTV.

Population dynamics of Citrus tristeza virus (CTV) in single aphid-transmitted sub-isolates of the South African GFMS12 isolate.

Grapefruit trees in South Africa have been cross protected against severe stem pitting genotypes of Citrus tristeza virus (CTV) since the 1920s using a mild strain initially called 'Nartia' but later referred to as grapefruit mild strain 12 (GFMS12). In the current study, the GFMS12 isolate was used as the source for single aphid transmissions (SAT) using Toxoptera citricida, commonly called the brown citrus aphid (BrCA). The BrCA-transmitted CTV sub-isolates were analyzed by the heteroduplex mobility assay (HMA), serological assays, genetic marker analysis (GMA), and selected sub-isolates were biologically indexed. Reverse transcription PCR of genomic regions was conducted using universal primers followed by cloning the PCR products, HMA and sequence analysis; nine genotypes of CTV were identified in the complex of GFMS12, including both severe and mild genotypes. A single BrCA transmitted up to six CTV genotypes simultaneously in one sub-isolate. The HMA was found to be a rapid, reliable tool for the identification of genotypes and can be useful in the development of CTV management strategies and budwood certification programs.

Comparative transcriptome analysis of Citrus macrophylla tree infected with Citrus tristeza virus stem pitting mutants provides new insight into the role of phloem regeneration in stem pitting disease.

Stem pitting is a complex and economically important virus-associated disease of perennial woody plants. Molecular mechanisms and pathways occurring during virus-plant interaction that result in this phenomenon are still obscure. Previous studies indicated that different Citrus tristeza virus (CTV) mutants induce defined stem pitting phenotypes ranging from mild (CTVΔp13) to severe (CTVΔp33) in Citrus macrophylla trees. In this study, we conducted comparative transcriptome analyses of C. macrophylla trees infected with CTV mutants (CTVΔp13 and CTVΔp33) and a full-length virus in comparison to healthy plants as control. The mild CTV stem pitting mutant had very few differentially expressed genes (DEGs) related to plant defense mechanism and plant growth and development. In contrast, substantial gene expression changes were observed in plants infected with the severe mutant and the full-length virus, indicating that both the p13 and p33 proteins of CTV acted as a regulator of symptom production by activating and modulating plant responses, respectively. The analysis of transcriptome data for CTVΔp33 and the full-length virus suggested that xylem specification has been blocked by detecting several genes encoding xylem, cell wall and lignin degradation, and cell wall loosening enzymes. Furthermore, stem pitting was accompanied by downregulation of transcription factors involved in regulation of xylem differentiation and downregulation of some genes involved in lignin biosynthesis, showing that the xylem differentiation and specification program has been shut off. Upregulation of genes encoding transcription factors associated with phloem and cambium development indicated the activation of this program in stem pitting disease. Furthermore, we detected the induction of several DEGs encoding proteins associated with cell cycle re-entry such as chromatin remodeling factors and cyclin, and histone modification. This kind of expression pattern of genes related to xylem differentiation and specification, phloem and cambium development, and cell cycle re-entry is demonstrated during secondary vascular tissue (SVT) regeneration. The microscopy analysis confirmed that the regeneration of new phloem is associated with stem pitting phenotypes. The findings of this study, thus, provide evidence for the association between stem pitting phenotypes and SVT regeneration, suggesting that the expression of these genes might play important roles in development of stem pitting symptoms. Overall, our findings suggest that phloem regeneration contributes to development of stem pitting symptoms.

Identification of Endogenous Genes for Normalizing Titer Variation of Citrus Tristeza Virus in Aphids at Different Post-acquisition Feeding Times.

Citrus tristeza virus (CTV) is efficiently transmitted in a semi-persistent manner by the brown citrus aphid (Toxoptera citricida (Kirkaldy)). Currently, the most sensitive method for detecting plant viruses in insect vectors is reverse-transcription quantitative polymerase chain reaction (RT-qPCR). In this study, the elongation factor-1 alpha (EF-1α) gene and acidic p0 ribosomal protein (RPAP0) gene were confirmed to be suitable reference genes for RT-qPCR normalization in viruliferous T. citricida aphids using the geNorm, NormFinder, and BestKeeper tools. Then the relative CTV titer in aphids (T. citricida) at different post-acquisition feeding times on healthy plants was quantified by RT-qPCR using EF-1α and RPAP0 as reference genes. The relative CTV titer retained in the aphids gradually decreased with increasing feeding time. During the first 0.5 h of feeding time on healthy plants, the remaining CTV titer in aphids showed about 80% rapid loss for the highly transmissible isolate CT11A and 40% loss for the poorly transmissible isolate CTLJ. The relative CTV titer in aphids during more than 12 h post-acquisition times for CT11A was significantly lower than at the other feeding times, which is similar to the trend found for CTLJ. To our knowledge, this is the first report about the relative titer variation of CTV remaining in T. citricida at different post-acquisition feeding times on healthy plants.

Better together: the use of virus-induced gene silencing technique to repress the expression of two endogenous citrus genes simultaneously.

Virus-induced gene silencing is a promising technique for functional genomics studies. Citrus tristeza virus was employed successfully to create an infectious clone that was used to silence endogenous citrus genes. Phytoene desaturase (PDS) and delta (δ)-aminolevulinic acid dehydratase (ALAD) were targeted successfully in citrus. Silencing PDS usually results in a photo-bleached leaf phenotype while silencing ALAD causes discrete yellow spots in leaves. Silencing two or more genes simultaneously using the same infectious clone could be difficult due to the capacity of the plasmid and subsequent cloning. On the other hand, inoculating a new construct into a citrus plant pre-infected with another construct fails due to the superinfection exclusion phenomenon. Herein, I report our successful trials whereby we simultaneously graft-inoculate constructs targeting PDS and ALAD. The budwoods were graft-inoculated into the same tree but on two different branches. Interestingly, a new phenotype was produced because of the silencing of the two genes, which we called "color-breaking". The phenotype was observed in both branches. Gene expression analysis showed a significant reduction of PDS and ALAD transcripts. This finding suggests the possibility of targeting more than one gene using different constructs, however, the graft-inoculation must be at the same time.

Citrus Tristeza Virus: From Pathogen to Panacea.

Citrus tristeza virus (CTV) is the most destructive viral pathogen of citrus. During the past century, CTV induced grave epidemics in citrus-growing areas worldwide that have resulted in a loss of more than 100 million trees. At present, the virus continues to threaten citrus production in many different countries. Research on CTV is accompanied by distinctive challenges stemming from the large size of its RNA genome, the narrow host range limited to slow-growing Citrus species and relatives, and the complexity of CTV populations. Despite these hurdles, remarkable progress has been made in understanding the CTV-host interactions and in converting the virus into a tool for crop protection and improvement. This review focuses on recent advances that have shed light on the mechanisms underlying CTV infection. Understanding these mechanisms is pivotal for the development of means to control CTV diseases and, ultimately, turn this virus into an ally.

Knowledge of teachers about suicidal behavior in adolescents / Conocimiento de los profesores sobre el comportamiento suicida en los adolescentes / Conhecimento de professores sobre comportamento suicida em adolescentes

Objective. To identify the knowledge of teachers about suicidal behavior in adolescents. Methods. Qualitative exploratory-descriptive study conducted in a state school in the municipality of Porto Alegre, RS, Brazil. Twelve teachers participated in the study. Data were collected through semi-structured interviews analyzed using Bardin's Content Analysis. Results. Three categories were built: "Warning signs of suicide", related to the signs identified by the professionals; "Risk factors for suicide", which indicate the reasons that may lead adolescents to present this type of behavior; and "Difficulties in dealing with the behaviors", referring to the behaviors adopted by adolescents and the difficulties of teachers before the theme. Conclusion. It was possible to identify that teachers recognize some signs of suicidal behavior, as well as some risk factors. Nonetheless, it is necessary to qualify them to approach the subject, since they feel insecure to act in more critical moments, thus generating mainly feelings of sadness, guilt and powerlessness.

Objetivo. Identificar el conocimiento de los profesores sobre el comportamiento suicida en adolescentes. Métodos. Estudio exploratorio-descriptivo y cualitativo realizado en una escuela pública del municipio de Porto Alegre, RS, Brasil. Doce profesores participaron en el estudio. Los datos se recogieron mediante entrevistas semiestructuradas, que fueron analizadas utilizando el Análisis de Contenido de Bardin. Resultados. Se construyeron tres categorías: "Señales de alerta de suicidio", relacionada con las señales identificadas por los profesionales; "Factores de riesgo de suicidio", que indican las razones que pueden llevar a los adolescentes a presentar este tipo de comportamiento; y "Dificultades para lidiar con los comportamientos", relativa a las conductas adoptadas por los adolescentes y a las dificultades de los profesionales frente al tema. Conclusión. Se pudo identificar que los profesores reconocen algunas señales de comportamiento suicida, así como algunos factores de riesgo. Sin embargo, es necesario cualificarlas para el abordaje del problema, pues se sienten inseguros para actuar en los momentos más críticos, generando principalmente sentimientos de tristeza, culpa e impotencia.

Objetivo. Identificar o conhecimento de professores sobre comportamento suicida em adolescentes. Métodos. Estudo exploratório-descritivo, qualitativo, realizado em uma escola estadual no município de Porto Alegre, RS, Brasil. Participaram no estudo 12 professores. Os dados foram coletados mediante entrevistas semiestruturadas analisadas por meio da Análise de Conteúdo de Bardin. Resultados. Foram construídas três categorias: "Sinais de alerta para o suicídio", relacionada com os sinais identificados pelos profissionais; "Fatores de risco para o suicídio", que apontam os motivos que podem levar os adolescentes a apresentarem este tipo de comportamento; e, "Dificuldades em lidar com os comportamentos" referentes às condutas adotadas pelos adolescentes e as dificuldades dos professores frente ao tema. Conclusão. Foi possível identificar que os professores reconhecem alguns sinais de comportamento suicida assim como alguns fatores de risco. Porém, é preciso qualificá-los para a abordagem ao assunto, visto que se sentem inseguros para agir em momentos mais críticos, gerando principalmente sentimentos de tristeza, culpa e impotência.

A Comprehensive Analysis of Citrus Tristeza Variants of Bhutan and Across the World.

Mandarin orange is economically one of the most important fruit crops in Bhutan. However, in recent years, orange productivity has dropped due to severe infection of citrus tristeza virus (CTV) associated with the gradual decline of citrus orchards. Although the disease incidence has been reported, very limited information is available on genetic variability among the Bhutanese CTV variants. This study used reverse transcription PCR (RT-PCR) to detect CTV in collected field samples and recorded disease incidence up to 71.11% in Bhutan's prominent citrus-growing regions. To elucidate the extent of genetic variabilities among the Bhutanese CTV variants, we targeted four independent genomic regions (5'ORF1a, p25, p23, and p18) and analyzed a total of 64 collected isolates. These genomic regions were amplified and sequenced for further comparative bioinformatics analysis. Comprehensive phylogenetic reconstructions of the GenBank deposited sequences, including the corresponding genomic locations from 53 whole-genome sequences, revealed unexpected and rich diversity among Bhutanese CTV variants. A resistant-breaking (RB) variant was also identified for the first time from the Asian subcontinent. Our analyses unambiguously identified five (T36, T3, T68, VT, and HA16-5) major, well-recognized CTV strains. Bhutanese CTV variants form two additional newly identified distinct clades with higher confidence, B1 and B2, named after Bhutan. The origin of each of these nine clades can be traced back to their root in the north-eastern region of India and Bhutan. Together, our study established a definitive framework for categorizing global CTV variants into their distinctive clades and provided novel insights into multiple genomic region-based genetic diversity assessments, including their pathogenicity status.

Analysis of Citrus Tristeza Virus Incidences within Asian Citrus Psyllid (Diaphorina citri) Populations in Florida via High-Throughput Sequencing.

The destructive citrus disease, Huanglongbing (HLB) or citrus greening, continues to devastate Florida's citrus industry. A hemipteran insect, the Asian citrus psyllid (ACP), disperses Candidatus Liberibacter asiaticus, one of the putative bacterial pathogens of HLB. This study builds upon ongoing research utilizing high-throughput sequencing to analyze the virome of ACP populations collected from citrus groves throughout Florida. Following the widespread detection of sequences aligning to the genome of citrus tristeza virus (CTV) across consecutive years in the Florida ACP virome, we continued to detect a pervasive amount of CTV in Florida ACPs during subsequent years. Simultaneously, we also detected mixed infections of CTV strains in pooled ACPs from different Florida regions. Predating the HLB epidemic, CTV has been present in Florida for many years and our results confirm its widespread and diverse persistence in Florida citrus groves through a unique lens, the ACP. CTV presence in the ACP likely results from feeding on CTV-infected citrus trees in Florida citrus groves, which may help to understand an overlapping presence of CTV and HLB, both endemic citrus pathosystems in the state, and their role in future integrated pest management strategies.