Multiplex qPCR for the detection of bacteria associated with huanglongbing 'Candidatus Liberibacter asiaticus', 'Ca. L. americanus' and 16Sr IX group phytoplasma.

The occurrence of 'Candidatus Liberibacter' spp. and 'Ca. Phytoplasma' spp. associated with blotchy mottle symptoms poses challenges to huanglongbing (HLB) diagnosis using molecular techniques. The ability to detect multiple targets simultaneously and specifically is a key aspect met by qPCR. A set of primers and hydrolysis probes useful either in single or multiplex reactions for the detection and quantification of HLB-associated bacteria were developed. Sequences from conserved genes of the ribosomal proteins for Liberibacter and phytoplasma circumvent the lack of specificity and cross-reactivity problems related to 16S rDNA gene amplification, allowing precise and specific detection of HLB-associated bacteria in citrus and in the Liberibacter vector, Diaphorina citri. The triplex reaction exhibited high quality and precision as a robust tool for quantifying 'Ca. L. asiaticus' (CLas), 'Ca. L. americanus' (CLam) and 16SrIX phytoplasma. Triplex qPCR showed consistent results and comparable sensitivity to the RNR test, though Cq values were higher when compared to 16S rDNA qPCR. Detection tests using field samples indicate that the qPCR triplex can identify HLB-associated bacteria in samples with varying levels of symptoms, ranging from typical to asymptomatic. Assessment of field samples from growers indicated more than 78.6% had Cq lower than 35.0, below the cut-off established for qPCR reactions used in this work. qPCR triplex is a safe, specific, and sufficiently sensitive technique for detecting CLas, CLam and 16Sr IX phytoplasma simultaneously, in both citrus and D. citri samples. Its application is of importance in assisting growers in making decisions for HLB management.

Exploring structural and computational contrasts in Myoglobins: Implications for thermal treatment-induced Sulfmyoglobin formation.

Greening of tuna metmyoglobin (MetMb) by thermal treatment (TT) and free cysteine is associated with sulfmyoglobin (SulfMb) production. This greening reaction (GR) was once thought to occur only in tuna species. However, recent research has revealed that not all tuna species exhibit this behavior, and it can also occur in horse MetMb. Thus, the present study aimed to compare the GR-reactive (Katsuwonus pelamis and Equus caballus) and GR-unreactive (Sarda chiliensis and Euthynnus lineatus) MetMb using UV-vis spectrometry during TT (60 °C/30 min and free cysteine) to monitor the GR. We used molecular dynamics (MD) simulation to assess the stability of the heme group during TT. We discovered that using GR-unreactive MetMb resulted in an incomplete GR without producing SulfMb. Additionally, our MD simulations indicated that Met85 presence in the heme cavity from GR-unreactive is responsible for the heme group instability and displacement of distal His during TT.

Comparison of Insecticide Sprays in Alternate Rows and in All Rows on Asian Citrus Psyllid Control and Huanglongbing Progress in Sweet Orange Orchards.

One strategy to reduce huanglongbing (HLB) is controlling its insect vector, the Asian citrus psyllid (ACP) Diaphorina citri, by preventive insecticide sprays. The recommendation is to spray insecticide in all rows (conventional spray [CONV]), but some growers empirically spray in alternate rows (ALT) to increase the spray frequency without increasing the operating cost. Therefore, this work compared the effect of ALT with CONV on the ACP population and HLB incidence. The spray deposition (amount of metallic copper per leaf area), coverage (percentage of water-sensitive paper area covered by spray), and efficacy (ACP mortality) of each treatment were also evaluated on both sides of the trees. Two field trials were performed: Trial #1 compared ALT every 7 days (ALT7) with CONV every 14 days (CONV14), and trial #2 compared different spray frequencies of ALT with CONV every 7 days (CONV7). In trial #1, no differences were observed in the ACP population or HLB progress between ALT7 and CONV14 after 5 years. In trial #2, ALT7 presented the highest percentage of ACP and cumulative HLB incidence than CONV7 and ALT every 3 to 4 days, after 2 years. Hence, when the frequency of ALT was half the frequency of CONV, similar results were observed. Spray deposition, coverage, and efficacy were similar between tree sides in CONV, but they were uneven in ALT, resulting in higher values on the tree side that directly received the spray. Insecticide spray should be performed with the frequency enough to keep new shoots protected during their growth.

Review of the Greening Reaction by Thermal Treatment: New Insights Exploring the Structural Implications of Myoglobin.

Myoglobin is the main factor responsible for muscle pigmentation in tuna; muscle color depends upon changes in the oxidative state of myoglobin. The tuna industry has reported muscle greening after thermal treatment involving metmyoglobin (MetMb), trimethylamine oxide (TMAO), and free cysteine (Cys). It has been proposed that this pigmentation change is due to a disulfide bond between a unique cysteine residue (Cys10) found in tuna MetMb and free Cys. However, no evidence has been given to confirm that this reaction occurs. In this review, new findings about the mechanism of this greening reaction are discussed, showing evidence of how free radicals produced from Cys oxidation under thermal treatment participate in the greening of tuna and horse muscle during thermal treatment. In addition, the reaction conditions are compared to other green myoglobins, such as sulfmyoglobin, verdomyoglobin, and cholemyoglobin.

Sulfmyoglobin production by free cysteine during thermal treatment: Involvement of heme iron in the production of free radicals.

The meat greening is an abnormal pigmentation related to microbiological contamination and lipid oxidation during storage. This color change results from sulfmyoglobin (SulfMb) production promoted by the reaction between metmyoglobin (MetMb), H2O2, and thiol compounds. Spectral studies on cooked meat suggested the production of SulfMb, probably due to the increment of free radicals during thermal treatment. Thus, we evaluated the involvement of free radicals and heme iron in the SulfMb production from horse MetMb and free cysteine (Cys) during thermal treatment. The results confirm that the reaction of SulfMb production at meat muscle pH (5.7-7.2) during heat treatment is a product of free radicals formed from Cys oxidation (SH) and reactive oxygen species (O2-, H2O2). This is catalyzed by the release of heme iron, thus promoting a consecutive reaction having MbFe(IV)O as a reaction intermediate.

Ineffectiveness of applying additional insecticide sprays at the border of the citrus block for the control of Huanglongbing.

BACKGROUND: Huanglongbing (HLB), mainly associated with the bacterium 'Candidatus Liberibacter asiaticus', is a devastating disease that causes negative impacts on citrus yield and fruit quality. One of the main HLB control measures is based on preventive insecticide sprays, aiming to control its vector, the Asian citrus psyllid (ACP) Diaphorina citri. In blocks on the orchard's periphery, the ACP population is more concentrated in trees closer to the block's edge and gradually decreasing towards its interior. This spread pattern of ACP has motivated growers to perform, empirically based, additional insecticide applications at the border of the block ('AB'), as an additional measure to spraying insecticides in the entire block ('EB'). Hence, this work aimed (i) to evaluate the coverage and efficiency of AB, comparing tree canopy volume, and planting direction; and (ii) to compare ACP population and HLB incidence between blocks treated under EB, and blocks that received 'EB + AB'. RESULTS: Regardless of the operational situation, as the distance increased from the orchard's edge (0 m) to the interior, spray coverage exponentially decayed from an average of 21% to less than 1% of coverage, and ACP mortality exponentially decayed from 83% to 7%. In general, the spray coverage and ACP mortality were suitable only for the tree canopy face directly sprayed at the orchard's edge (0 m). After 5.5 years, a higher ACP population and number of symptomatic trees were detected in 'EB + AB'. CONCLUSION: As the 'EB + AB' had no effect in reducing ACP population, or HLB incidence, this additional spraying had no benefit in HLB management. © 2022 Society of Chemical Industry.

Insight into resistance to 'Candidatus Liberibacter asiaticus,' associated with Huanglongbing, in Oceanian citrus genotypes.

Huanglongbing (HLB), the most destructive citrus disease, is associated with unculturable, phloem-limited Candidatus Liberibacter species, mainly Ca. L. asiaticus (Las). Las is transmitted naturally by the insect Diaphorina citri. In a previous study, we determined that the Oceanian citrus relatives Eremocitrus glauca, Microcitrus warburgiana, Microcitrus papuana, and Microcitrus australis and three hybrids among them and Citrus were full-resistant to Las. After 2 years of evaluations, leaves of those seven genotypes remained Las-free even with their susceptible rootstock being infected. However, Las was detected in their stem bark above the scion-rootstock graft union. Aiming to gain an understanding of the full-resistance phenotype, new experiments were carried out with the challenge-inoculated Oceanian citrus genotypes through which we evaluated: (1) Las acquisition by D. citri fed onto them; (2) Las infection in sweet orange plants grafted with bark or budwood from them; (3) Las infection in sweet orange plants top-grafted onto them; (4) Las infection in new shoots from rooted plants of them; and (5) Las infection in new shoots of them after drastic back-pruning. Overall, results showed that insects that fed on plants from the Oceanian citrus genotypes, their canopies, new flushes, and leaves from rooted cuttings evaluated remained quantitative real-time polymerase chain reaction (qPCR)-negative. Moreover, their budwood pieces were unable to infect sweet orange through grafting. Furthermore, sweet orange control leaves resulted infected when insects fed onto them and graft-receptor susceptible plants. Genomic and morphological analysis of the Oceanian genotypes corroborated that E. glauca and M. warburgiana are pure species while our M. australis accession is an M. australis × M. inodora hybrid and M. papuana is probably a M. papuana × M. warburgiana hybrid. E. glauca × C. sinensis hybrid was found coming from a cross between E. glauca and mandarin or tangor. Eremocitrus × Microcitrus hybrid is a complex admixture of M. australasica, M. australis, and E. glauca while the last hybrid is an M. australasica × M. australis admixture. Confirmation of consistent full resistance in these genotypes with proper validation of their genomic parentages is essential to map properly genomic regions for breeding programs aimed to generate new Citrus-like cultivars yielding immunity to HLB.

A reverse-ecology framework to uncover the potential metabolic interplay among 'Candidatus Liberibacter' species, citrus hosts and psyllid vector.

'Candidatus Liberibacter' species have developed a dependency on essential nutrients and metabolites from the host cell, as a result of substantial genome reduction. Still, it is difficult to state which nutrients they acquire and whether or not they are metabolically reliant. We used a reverse-ecology model to investigate the potential metabolic interactions of 'Ca Liberibacter' species, Citrus, and the psyllid Diaphorina citri in the huanglongbing disease pyramid. Our findings show that hosts (citrus and psyllid) tend to support the nutritional needs of 'Ca. Liberibacter' species, implying that the pathogen's metabolism has become tightly linked to hosts, which may reflect in the parasite lifestyle of this important genus.

Physical activity during recess in elementary schoolchildren in Belgium and Ecuador: The role of the physical environment at school.

To increase physical activity levels in children, knowledge on how school playgrounds can be optimally designed is important. Until now, the contribution of playground variety and greening to children's physical activity level during recess remains unclear. This study examined the associations of school playground variety and greening with energy expenditure (EE) in elementary school children in Ecuador and Belgium. Additionally, moderating effects of study site and gender were studied. Observational data were collected using SOPLAY in ten schools in East Flanders (Belgium) and Cuenca (Ecuador). Data were analysed with linear mixed-effects model analyses. Results showed that independent of study site, EE was higher in playground areas with less green elements, and a higher level of total variety. Primary surface was only associated with EE in boys, they were more active when an artificial surface was present. Implementation of loose equipment was somewhat more strongly positively associated with EE in Ecuadorian than in Belgian children. Because of the well-known restorative effects of green elements, we do not wish to suggest that less greening should be implemented at schools. However, ideally green spaces should be designed in a way that active play is stimulated by combining green elements with other improvements.

Prevalent Transmission of 'Candidatus Liberibacter asiaticus' over 'Ca. Liberibacter americanus' in a Long-Term Controlled Environment.

In Brazil, citrus huanglongbing (HLB) is associated with 'Candidatus Liberibacter americanus' (CLam) and 'Ca. Liberibacter asiaticus' (CLas). However, there are few studies about HLB epidemiology when both Liberibacter spp. and its insect vector, the Asian citrus psyllid (ACP, Diaphorina citri), are present. The objective of this work was to compare the transmission of HLB by ACP when both CLam and CLas are present as primary inoculum. Two experiments were performed under screenhouse conditions from April 2008 to January 2012 (experiment 1) and from February 2011 to December 2015 (experiment 2). The experiments were carried out with sweet orange plants infected with CLam or CLas as inoculum source surrounded by sweet orange healthy plants. One hundred Liberibacter-free adult psyllids were monthly confined to the source of inoculum plants for 7 days with subsequent free movement inside the screenhouse. Fortnightly, nymphs and adults of psyllids were monitored. Psyllid and leaf samples were collected periodically for Liberibacter detection by PCR or quantitative PCR. CLas was detected more frequently than CLam in both psyllid and leaf samples. No mixed infections were detected in the psyllids. A clear prevalence of CLas over CLam was observed in both experiments. The final HLB incidences were 16.7 and 14.5% of Liberibacter-positive test plants, and CLas was detected in 92.3 and 93.1% of these infected plants. Mixed infection was observed only in 3.8% of infected test plants in experiment 1. These results endorse the shift in the prevalence of CLam to CLas observed in citrus orchards of São Paulo, Brazil.

Effects of Deficit Irrigation and Huanglongbing on Sweet Orange Trees.

This study addresses the interactive effects of deficit irrigation and huanglongbing (HLB) infection on the physiological, biochemical, and oxidative stress responses of sweet orange trees. We sought to answer: (i) What are the causes for the reduction in water uptake in HLB infected plants? (ii) Is the water status of plants negatively affected by HLB infection? (iii) What are the key physiological traits impaired in HLB-infected plants? and (iv) What conditions can mitigate both disease severity and physiological/biochemical impairments in HLB-infected plants? Two water management treatments were applied for 11 weeks to 1-year-old-trees that were either healthy (HLB-) or infected with HLB (+) and grown in 12-L pots. Half of the trees were fully irrigated (FI) to saturation, whereas half were deficit-irrigated (DI) using 40% of the water required to saturate the substrate. Our results demonstrated that: reduced water uptake capacity in HLB+ plants was associated with reduced root growth, leaf area, stomatal conductance, and transpiration. Leaf water potential was not negatively affected by HLB infection. HLB increased leaf respiration rates (ca. 41%) and starch synthesis, downregulated starch breakdown, blocked electron transport, improved oxidative stress, and reduced leaf photosynthesis (ca. 57%) and photorespiration (ca.57%). Deficit irrigation reduced both leaf respiration (ca. 45%) and accumulation of starch (ca.53%) by increasing maltose (ca. 20%), sucrose, glucose, and fructose contents in the leaves, decreasing bacterial population (ca. 9%) and triggering a series of protective measures against further impairments in the physiology and biochemistry of HLB-infected plants. Such results provide a more complete physiological and biochemical overview of HLB-infected plants and can guide future studies to screen genetic tolerance to HLB and improve management strategies under field orchard conditions.

El desarrollo y beneficio de las escuelas de odontología sustentables / The development and benefit of sustainable dentistry school

Las Escuelas de Odontología como espacios educativos universitarios son un lugar idóneo para trabajar de manera urgente los procesos de ambientalización curricular que permitan realizar de forma horizontal y vertical temáticas y contenidos para crear y recrear un tipo de cultura y conciencia más amigable con el medio ambiente, y contribuir con conductas y actitudes sustentables en los estudiantes, docentes y trabajadores administrativos hacia una reducción significativa en la contaminación del ambiente local, regional y mundial, colaborando en ello con la solución a la regularización del clima y el control de los desastres ocasionados por el cambio climático. Además de esto, a ser las Escuelas de Odontología instituciones educativas que pueden funcionar bajo el paradigma de la sustentabilidad, se lograría contribuir al aseguramiento de los recursos naturales renovables y no renovables para las nuevas generaciones, además del logro de un importante ahorro económico en el gasto corriente de las escuelas (AU)

The Dental School as university educational spaces are an ideal place to urgently work on the curricular greening processes that allow working horizontally and vertically with themes and contents that allow creating and recreating a type of culture and awareness that is more friendly to the environment and contribute with sustainable behaviors and attitudes in students, teachers and administrative workers a more significant reduction in pollution of the local, regional and global environment, collaborating in this with the solution to the regularization of the climate and the control of disasters caused by change climate. In addition, being the School of Dentistry educational institutions that can function under the paradigm of sustainability, it would be possible to contribute to the assurance of renewable and non-renewable natural resource for the new generations, in addition to achieving significant economic savings in current spending from school (AU)

The Genome of "Candidatus Liberibacter asiaticus" Is Highly Transcribed When Infecting the Gut of Diaphorina citri.

The Asian citrus psyllid, Diaphorina citri, is the vector of the bacterium "Candidatus Liberibacter asiaticus" (Las), associated with the devastating, worldwide citrus disease huanglongbing. In order to explore the molecular interactions of this bacterium with D. citri during the vector acquisition process, cDNA libraries were sequenced on an Illumina platform, obtained from the gut of adult psyllids confined in healthy (H) and in Las-infected young shoots (Las) for different periods of times (I = 1/2 days, II = 3/4 days, and III = 5/6 days). In each sampling time, three biological replicates were collected, containing 100 guts each, totaling 18 libraries depleted in ribosomal RNA. Reads were quality-filtered and mapped against the Chinese JXGC Las strain and the Floridian strain UF506 for the analysis of the activity of Las genome and SC1, SC2, and type 3 (P-JXGC-3) prophages of the studied Las strain. Gene activity was considered only if reads of at least two replicates for each acquisition access period mapped against the selected genomes, which resulted in coverages of 44.4, 79.9, and 94.5% of the JXGC predicted coding sequences in Las I, Las II, and Las III, respectively. These genes indicate an active metabolism and increased expression according to the feeding time in the following functional categories: energy production, amino acid metabolism, signal translation, cell wall, and replication and repair of genetic material. Pilins were among the most highly expressed genes regardless of the acquisition time, while only a few genes from cluster I of flagella were not expressed. Furthermore, the prophage region had a greater coverage of reads for SC1 and P-JXGC-3 prophages and low coverage in SC2 and no indication of activity for the lysis cycle. This research presents the first descriptive analysis of Las transcriptome in the initial steps of the D. citri gut colonization, where 95% of Las genes were active.

Protocol for Successful Transmission of 'Candidatus Liberibacter asiaticus' from Citrus to Citrus Using Diaphorina citri.

A protocol to successfully transmit the huanglongbing (HLB) pathogen, 'Candidatus Liberibacter asiaticus', between citrus plants by using the Asian citrus psyllid (ACP) and an alternative way to help growers control ACP are proposed. Best results were obtained when pathogen acquisition by adults reared on fully symptomatic 'Ca. Liberibacter asiaticus'-positive plants, latency, and inoculation occurred at ambient air temperatures ranging from 24 to 28°C and when a single infective adult ACP was confined for 7 days on soft, newly developing vegetative shoots (stages v2 to v4). No infection resulted from confinement of infective ACP adults on mature leaves (stage v6). Under the described conditions, single ACP adults could successfully transmit 'Ca. Liberibacter asiaticus' to an average of 56.5% (35 to 83%) of plantlets with v2 to v4 shoots growing in 0.3-liter tubes and to 80.5% (76 to 86%) of plants with v2 to v4 shoots growing in 4.7-liter pots. The use of single insects and plantlets reduces labor, space, and other resources necessary to undertake transmission tests. It also reduces time needed for transmission studies and should help accelerate research on HLB. The results were used to develop an index for favorability to infection (IFI) to determine orchard vulnerabilities to 'Ca. Liberibacter asiaticus'. The IFI is based on the heterogeneous population of new shoots that occurs on tree canopies and may offer alternative or complementary alternatives to the laborious and costly insect surveys currently used in most instances to determine threshold levels for insecticide applications.

Huanglongbing and Citrus Variegated Chlorosis Integrated Management Based on Favorable Periods for Vector Population Increase and Symptom Expression.

Huanglongbing (HLB, associated with Candidatus Liberibacter asiaticus and transmitted by the Asian citrus psyllid Diaphorina citri) and citrus variegated chlorosis (CVC, caused by Xylella fastidiosa subsp. pauca and transmitted by sharpshooter species) have been managed by vector control and removal of symptomatic trees. Although vectors and new symptomatic trees can be detected year round, peaks of vector populations are higher in spring and summer, and the most symptomatic trees are found in autumn and winter. This work aimed to compare the management of both diseases during these favorable periods. The experiment was conducted during 5 years in a commercial orchard and had a three-by-two factorial design. The factor "vector control" had three levels: monthly vector control year round (VCYR), monthly vector control in spring and summer (VCSS), and vector control when a threshold level of 10% occupancy was detected (VCOT). The factor "inoculum removal" had two levels: monthly eradication year round (TEYR) and monthly eradication in autumn and winter (TEAW). Host flush, both HLB and CVC vector populations, and the number of symptomatic citrus plants were visually assessed. The level of vectors over the seasons, as measured using the average area under the curve (AUC), was similar for all treatments with the exception of psyllid abundance, which was approximately 4.5 times higher for VCSS than for other treatments. For both diseases, no difference in the average AUC of disease progress and disease final incidence was observed. VCOT or adjusted VCSS associated with TEAW could be integrated for sustainable citrus production.

Temporal Progress of Huanglongbing Epidemics and the Effect of Noncommercial Inoculum Sources on Citrus Orchards in São Paulo State, Brazil.

Huanglongbing (HLB) incidence is increasing and threatening citrus production in São Paulo State, Brazil, despite multiple efforts to control the disease and its vector, the Asian citrus psyllid (ACP) (Diaphorina citri). The objective of this research was to study the temporal dynamics of HLB epidemics, under intensive disease management, in 177 individual commercial citrus blocks on a single property in São Paulo State. The effect of internal and external sources of HLB-associated bacteria and its vector were explored based on the disease epidemics and vector dynamics in the studied area. To manage HLB, the property owner used healthy nursery plants, eradicated symptomatic trees, and used insecticides to control ACPs. Logistic and Gompertz models were fitted to the data to describe dynamics of HLB incidence for all blocks. The average number of ACPs per yellow sticky trap was determined for the same blocks for a period of four consecutive years. Both logistic and Gompertz models described the HLB epidemics well, although the Gompertz model provided a slightly better fit. Disease progress rates, HLB incidences, and average ACP count per trap in the 177 blocks were low compared with reports in the literature. HLB incidence and number of ACPs per trap were higher (P ≤ 0.05) in some citrus blocks located on the periphery of the property. A large number of noncommercial trees were found near the property and were a potential primary inoculum source of HLB-associated bacteria, accounting for the higher incidence of HLB and ACPs per trap in blocks located on the periphery of the property. These results support the recommended preventive measures to HLB management and the necessity of external actions, to include trees in commercial orchards, and noncommercial trees located near commercial citrus properties, in an attempt to maximize the effectiveness of these preventive measures.

Genome Sequence Resource of 'Candidatus Liberibacter asiaticus' strain 9PA From Brazil.

'Candidatus Liberibacter asiaticus', an unculturable α-proteobacterium, is associated with citrus huanglongbing (HLB), a devastating disease threatening citrus production in Brazil and worldwide. In this study, a draft whole-genome sequence of 'Ca. L. asiaticus' strain 9PA from a sweet orange (cultivar Pera) tree collected in São Paulo State, Brazil, is reported. The 9PA genome is 1,231,881 bp, including two prophages, with G+C content of 36.7%. This is the first report of a whole-genome sequence of 'Ca. L. asiaticus' from Brazil or South America. The 9PA genome sequence will enrich 'Ca. L. asiaticus' genome resources and facilitate HLB research and control in Brazil and the world.

Water Vapor Pressure Deficit in Portugal and Implications for the Development of the Invasive African Citrus Psyllid Trioza erytreae.

African citrus psyllid (Trioza erytreae (Del Guercio)) is a vector insect of the bacterium Candidatus Liberibacter africanus, the putative causal agent of Huanglongbing, the most devastating citrus disease in the world. The insect was found on the island of Madeira in 1994 and in mainland Portugal in 2015. Present in the north and center of the country, it is a threat to Algarve, the main citrus-producing region. Trioza erytreae eggs and first instar nymphs are sensitive to the combination of high temperatures and low relative humidity. Daily maximum air temperature and minimum relative humidity data from 18 weather stations were used to calculate the water vapor pressure deficit (vpd) from 2004 to 2018 at various locations. Based on the mean vpd and the number of unfavorable days (vpd < 34.5 and vpd < 56 mbar) of two time periods (February to May and June to September), less favorable zones for T. erytreae were identified. The zones with thermal and water conditions like those observed in the Castelo Branco and Portalegre (Center), Beja (Alentejo), Alte, and Norinha (Algarve) stations showed climatic restrictions to the development of eggs and first instar nymphs of African citrus psyllid. Effective control measures, such as the introduction and mass release of Tamarixia dryi (Waterson), a specific parasitoid, and chemical control are necessary in favorable periods for T. erytreae development, such as in spring and in areas with limited or no climate restrictions.

The Enemy is Outside: Releasing the Parasitoid Tamarixia radiata (Hymenoptera: Eulophidae) in External Sources of HLB Inocula to Control the Asian Citrus Psyllid Diaphorina citri (Hemiptera: Liviidae).

Huanglongbing (HLB), the most destructive citrus disease worldwide, was first recorded in Brazil in 2004, and since then, more than 50 million trees identified with this disease have been eliminated. The disease is managed mainly by controlling the psyllid vector Diaphorina citri Kuwayama, 1908 (Hemiptera: Liviidae). Although the presence of the insect in commercial citrus groves is low, HLB infection rates increase in areas bordering the groves. The disease is transmitted by psyllids from host citrus plants in areas outside the managed groves, such as abandoned or organic groves and residential trees, and from orange jasmine plants in urban settings. In order to provide information to support HLB control, this study evaluated the biotic and abiotic variables that affect the dynamics of D. citri populations after releases of the parasitoid wasp Tamarixia radiata (Waterston, 1922) (Hymenoptera: Eulophidae) in external sources of HLB inocula. The study was divided into two parts. After releasing the parasitoids in non-commercial areas, we determined the following: (a) the variables that significantly affected the number of nymphs collected on shoots in the same non-commercial area; (b) the variables that significantly affected the number of adult psyllids collected in a neighboring commercial citrus area. Our results indicated that the number of nymphs in external areas was affected only by the host plant and rainfall. However, periodic parasitoid releases significantly reduced the number of adult psyllids collected in the commercial area. The results indicate that the release of parasitoids in external sources of inocula has the potential to maximize actions for D. citri control, contributing to the reduction of psyllid populations in commercial areas. Consequently, this strategy may help to manage the disease infection without an increase in insecticide use.

Resistance to 'Candidatus Liberibacter asiaticus,' the Huanglongbing Associated Bacterium, in Sexually and/or Graft-Compatible Citrus Relatives.

Huanglongbing (HLB) is the most destructive, yet incurable disease of citrus. Finding sources of genetic resistance to HLB-associated 'Candidatus Liberibacter asiaticus' (Las) becomes strategic to warrant crop sustainability, but no resistant Citrus genotypes exist. Some Citrus relatives of the family Rutaceae, subfamily Aurantioideae, were described as full-resistant to Las, but they are phylogenetically far, thus incompatible with Citrus. Partial resistance was indicated for certain cross-compatible types. Moreover, other genotypes from subtribe Citrinae, sexually incompatible but graft-compatible with Citrus, may provide new rootstocks able to restrict bacterial titer in the canopy. Use of seedlings from monoembryonic species and inconsistencies in previous reports likely due to Las recalcitrance encouraged us to evaluate more accurately these Citrus relatives. We tested for Las resistance a diverse collection of graft-compatible Citrinae species using an aggressive and consistent challenge-inoculation and evaluation procedure. Most Citrinae species examined were either susceptible or partially resistant to Las. However, Eremocitrus glauca and Papua/New Guinea Microcitrus species as well as their hybrids and those with Citrus arose here for the first time as full-resistant, opening the way for using these underutilized genotypes as Las resistance sources in breeding programs or attempting using them directly as possible new Las-resistant Citrus rootstocks or interstocks.