An Investigation of Culicoides (Diptera: Ceratopogonidae) as Potential Vectors of Medically and Veterinary Important Arboviruses in South Africa.

Culicoides-borne viruses such as bluetongue, African horse sickness, and Schmallenberg virus cause major economic burdens due to animal outbreaks in Africa and their emergence in Europe and Asia. However, little is known about the role of Culicoides as vectors for zoonotic arboviruses. In this study, we identify both veterinary and zoonotic arboviruses in pools of Culicoides biting midges in South Africa, during 2012-2017. Midges were collected at six surveillance sites in three provinces and screened for Alphavirs, Flavivirus, Orthobunyavirus, and Phlebovirus genera; equine encephalosis virus (EEV); and Rhaboviridae, by reverse transcription polymerase chain reaction. In total, 66/331 (minimum infection rate (MIR) = 0.4) pools tested positive for one or more arbovirus. Orthobunyaviruses, including Shuni virus (MIR = 0.1) and EEV (MIR = 0.2) were more readily detected, while only 2/66 (MIR = 0.1) Middelburg virus and 4/66 unknown Rhabdoviridae viruses (MIR = 0.0) were detected. This study suggests Culicoides as potential vectors of both veterinary and zoonotic arboviruses detected in disease outbreaks in Africa, which may contribute to the emergence of these viruses to new regions.

Towards an odour-baited trap to control Musca sorbens, the putative vector of trachoma.

Musca sorbens is a synanthropic filth fly that aggressively attacks people to feed from mucous membranes of the eyes, nose or mouth, from open sores, or from sweat. It has long been suspected that this fly contributes to the transmission of eye infections, particularly trachoma, and recent work has added to the evidence base that M. sorbens is a trachoma vector in Ethiopia. There are few options to control M. sorbens, largely due to a lack of evidence. Space spraying with insecticides is effective, but an environmentally sound and long-term sustainable solution would be better, for example, mass trapping. We tested commercially available and homemade trap types in a pilot (laboratory) study and three field studies. A homemade design, built from a bucket and two empty water bottles, baited with a commercially available lure, The Buzz, was found to be most effective. This trap caught 3848 M. sorbens over 26 trap 'events' (3- or 4-day periods); mean/median per 24 h 43.6 (standard deviation 137.10)/2.25 (IQR 0.25-12.67). The Buzz lure is cheap and effective for 4 weeks, and trap components cheap and locally available. Further studies are needed to understand the impact of this trap on local fly populations and the local transmission of trachoma.

Transcriptome response comparison between vector and non-vector aphids after feeding on virus-infected wheat plants.

BACKGROUND: Plant viruses maintain intricate interactions with their vector and non-vector insects and can impact the fitness of insects. However, the details of their molecular and cellular mechanisms have not been studied well. We compared the transcriptome-level responses in vector and non-vector aphids (Schizaphis graminum and Rhopalosiphum padi, respectively) after feeding on wheat plants with viral infections (Barley Yellow Dwarf Virus (BYDV) and Wheat dwarf virus (WDV), respectively). We conducted differentially expressed gene (DEG) annotation analyses and observed DEGs related to immune pathway, growth, development, and reproduction. And we conducted cloning and bioinformatic analyses of the key DEG involved in immune. RESULTS: For all differentially expressed gene analyses, the numbers of DEGs related to immune, growth, development, reproduction and cuticle were higher in vector aphids than in non-vector aphids. STAT5B (signal transducer and activator of transcription 5B), which is involved in the JAK-STAT pathway, was upregulated in R. padi exposed to WDV. The cloning and bioinformatic results indicated that the RpSTAT5B sequence contains a 2082 bp ORF encoding 693 amino acids. The protein molecular weight is 79.1 kD and pI is 8.13. Analysis indicated that RpSTAT5B is a non-transmembrane protein and a non-secreted protein. Homology and evolutionary analysis indicated that RpSTAT5B was closely related to R. maidis. CONCLUSIONS: Unigene expression analysis showed that the total number of differentially expressed genes (DEGs) in the vector aphids was higher than that in the non-vector aphids. Functional enrichment analysis showed that the DEGs related to immunity, growth and reproduction in vector aphids were higher than those in non-vector aphids, and the differentially expressed genes related to immune were up-regulated. This study provides a basis for the evaluation of the response mechanisms of vector/non-vector insects to plant viruses.

Innate Immune Response to Tick-Borne Pathogens: Cellular and Molecular Mechanisms Induced in the Hosts.

Many pathogens are transmitted by tick bites, including Anaplasma spp., Ehrlichia spp., Rickettsia spp., Babesia and Theileria sensu stricto species. These pathogens cause infectious diseases both in animals and humans. Different types of immune effector mechanisms could be induced in hosts by these microorganisms, triggered either directly by pathogen-derived antigens or indirectly by molecules released by host cells binding to these antigens. The components of innate immunity, such as natural killer cells, complement proteins, macrophages, dendritic cells and tumor necrosis factor alpha, cause a rapid and intense protection for the acute phase of infectious diseases. Moreover, the onset of a pro-inflammatory state occurs upon the activation of the inflammasome, a protein scaffold with a key-role in host defense mechanism, regulating the action of caspase-1 and the maturation of interleukin-1ß and IL-18 into bioactive molecules. During the infection caused by different microbial agents, very similar profiles of the human innate immune response are observed including secretion of IL-1α, IL-8, and IFN-α, and suppression of superoxide dismutase, IL-1Ra and IL-17A release. Innate immunity is activated immediately after the infection and inflammasome-mediated changes in the pro-inflammatory cytokines at systemic and intracellular levels can be detected as early as on days 2-5 after tick bite. The ongoing research field of "inflammasome biology" focuses on the interactions among molecules and cells of innate immune response that could be responsible for triggering a protective adaptive immunity. The knowledge of the innate immunity mechanisms, as well as the new targets of investigation arising by bioinformatics analysis, could lead to the development of new methods of emergency diagnosis and prevention of tick-borne infections.

A plant DNA virus replicates in the salivary glands of its insect vector via recruitment of host DNA synthesis machinery.

Whereas most of the arthropod-borne animal viruses replicate in their vectors, this is less common for plant viruses. So far, only some plant RNA viruses have been demonstrated to replicate in insect vectors and plant hosts. How plant viruses evolved to replicate in the animal kingdom remains largely unknown. Geminiviruses comprise a large family of plant-infecting, single-stranded DNA viruses that cause serious crop losses worldwide. Here, we report evidence and insight into the replication of the geminivirus tomato yellow leaf curl virus (TYLCV) in the whitefly (Bemisia tabaci) vector and that replication is mainly in the salivary glands. We found that TYLCV induces DNA synthesis machinery, proliferating cell nuclear antigen (PCNA) and DNA polymerase δ (Polδ), to establish a replication-competent environment in whiteflies. TYLCV replication-associated protein (Rep) interacts with whitefly PCNA, which recruits DNA Polδ for virus replication. In contrast, another geminivirus, papaya leaf curl China virus (PaLCuCNV), does not replicate in the whitefly vector. PaLCuCNV does not induce DNA-synthesis machinery, and the Rep does not interact with whitefly PCNA. Our findings reveal important mechanisms by which a plant DNA virus replicates across the kingdom barrier in an insect and may help to explain the global spread of this devastating pathogen.

A biological control model to manage the vector and the infection of Xylella fastidiosa on olive trees.

Xylella fastidiosa pauca ST53 is the bacterium responsible for the Olive Quick Decline Syndrome that has killed millions of olive trees in Southern Italy. A recent work demonstrates that a rational integration of vector and transmission control measures, into a strategy based on chemical and physical control means, can manage Xylella fastidiosa invasion and impact below an acceptable economic threshold. In the present study, we propose a biological alternative to the chemical control action, which involves the predetermined use of an available natural enemy of Philaenus spumarius, i.e., Zelus renardii, for adult vector population and infection biocontrol. The paper combines two different approaches: a laboratory experiment to test the predation dynamics of Zelus renardii on Philaenus spumarius and its attitude as candidate for an inundation strategy; a simulated experiment of inundation, to preliminary test the efficacy of such strategy, before eventually proceeding to an in-field experimentation. With this double-fold approach we show that an inundation strategy with Zelus renardii has the potential to furnish an efficient and "green" solution to Xylella fastidiosa invasion, with a reduction of the pathogen incidence below 10%. The biocontrol model presented here could be promising for containing the impact and spread of Xylella fastidiosa, after an in-field validation of the inundation technique. Saving the fruit orchard, the production and the industry in susceptible areas could thus become an attainable goal, within comfortable parameters for sustainability, environmental safety, and effective plant health protection in organic orchard management.

Estimating epidemiological parameters from experiments in vector access to host plants, the method of matching gradients.

Estimation of pathogenic life-history values, for instance the duration a pathogen is retained in an insect vector (i.e., retention period) is of particular importance for understanding plant disease epidemiology. How can we extract values for these epidemiological parameters from conventional small-scale laboratory experiments in which transmission success is measured in relation to durations of vector access to host plants? We provide a solution to this problem by deriving formulae for the empirical curves that these experiments produce, called access period response curves (i.e., transmission success vs access period). We do this by writing simple equations for the fundamental life-cycle components of insect vectors in the laboratory. We then infer values of epidemiological parameters by matching the theoretical and empirical gradients of access period response curves. Using the example of Cassava brown streak virus (CBSV), which has emerged in sub-Saharan Africa and now threatens regional food security, we illustrate the method of matching gradients. We show how applying the method to published data produces a new understanding of CBSV through the inference of retention period, acquisition period and inoculation period parameters. We found that CBSV is retained for a far shorter duration in its insect vector (Bemisia tabaci whitefly) than had previously been assumed. Our results shed light on a number of critical factors that may be responsible for the transition of CBSV from sub- to super-threshold R0 in sub-Saharan Africa. The method is applicable to plant pathogens in general, to supply epidemiological parameter estimates that are crucial for practical management of epidemics and prediction of pandemic risk.

Evaluation of semiochemical based push-pull strategy for population suppression of ambrosia beetle vectors of laurel wilt disease in avocado.

Ambrosia beetles (Coleoptera: Curculionidae: Scolytinae and Platypodinae) bore into tree xylem to complete their life cycle, feeding on symbiotic fungi. Ambrosia beetles are a threat to avocado where they have been found to vector a symbiotic fungus, Raffaelea lauricola, the causal agent of the laurel wilt disease. We assessed the repellency of methyl salicylate and verbenone to two putative laurel wilt vectors in avocado, Xyleborus volvulus (Fabricius) and Xyleborus bispinatus (Eichhoff), under laboratory conditions. Then, we tested the same two chemicals released from SPLAT flowable matrix with and without low-dose ethanol dispensers for manipulation of ambrosia beetle populations occurring in commercial avocado. The potential active space of repellents was assessed by quantifying beetle catch on traps placed 'close' (~5-10 cm) and 'far' (~1-1.5 m) away from repellent dispensers. Ambrosia beetles collected on traps associated with all in-field treatments were identified to species to assess beetle diversity and community variation. Xyleborus volvulus was not repelled by methyl salicylate (MeSA) or verbenone in laboratory assays, while X. bispinatus was repelled by MeSA but not verbenone. Ambrosia beetle trap catches were reduced in the field more when plots were treated with verbenone dispensers (SPLAT) co-deployed with low-dose ethanol dispensers than when treated with verbenone alone. Beetle diversity was highest on traps deployed with low-dose ethanol lures. The repellent treatments and ethanol lures significantly altered the species composition of beetles captured in experiment plots. Our results indicate that verbenone co-deployed with ethanol lures holds potential for manipulating ambrosia beetle vectors via push-pull management in avocado. This tactic could discourage immigration and/or population establishment of ambrosia beetles in commercial avocado and function as an additional tool for management programs of laurel wilt.

Tomato chlorosis virus, an emergent plant virus still expanding its geographical and host ranges.

Tomato chlorosis virus (ToCV) causes an important disease that primarily affects tomato, although it has been found infecting other economically important vegetable crops and a wide range of wild plants. First described in Florida (USA) and associated with a 'yellow leaf disorder' in the mid-1990s, ToCV has been found in 35 countries and territories to date, constituting a paradigmatic example of an emergent plant pathogen. ToCV is transmitted semipersistently by whiteflies (Hemiptera: Aleyrodidae) belonging to the genera Bemisia and Trialeurodes. Whitefly transmission is highly efficient and cases of 100% infection are frequently observed in the field. To date, no resistant or tolerant tomato plants are commercially available and the control of the disease relies primarily on the control of the insect vector. TAXONOMY: Tomato chlorosis virus is one of the 14 accepted species in the genus Crinivirus, one of the four genera in the family Closteroviridae of plant viruses. VIRION AND GENOME PROPERTIES: The genome of ToCV is composed of two molecules of single-stranded positive-sense RNA, named RNA1 and RNA2, separately encapsidated in long, flexuous, rod-like virions. As has been shown for other closterovirids, ToCV virions are believed to have a bipolar structure. RNA1 contains four open reading frames (ORFs) encoding proteins associated with virus replication and suppression of gene silencing, whereas RNA2 contains nine ORFs encoding proteins putatively involved in encapsidation, cell-to-cell movement, gene silencing suppression and whitefly transmission. HOST RANGE: In addition to tomato, ToCV has been found to infect 84 dicot plant species belonging to 25 botanical families, including economically important crops. TRANSMISSION: Like all species within the genus Crinivirus, ToCV is semipersistently transmitted by whiteflies, being one of only two criniviruses transmitted by members of the genera Bemisia and Trialeurodes. DISEASE SYMPTOMS: Tomato 'yellow leaf disorder' syndrome includes interveinal yellowing and thickening of leaves. Symptoms first develop on lower leaves and then advance towards the upper part of the plant. Bronzing and necrosis of the older leaves are accompanied by a decline in vigour and reduction in fruit yield. In other hosts the most common symptoms include interveinal chlorosis and mild yellowing on older leaves. CONTROL: Control of the disease caused by ToCV is based on the use of healthy seedlings for transplanting, limiting accessibility of alternate host plants that can serve as virus reservoirs and the spraying of insecticides for vector control. Although several wild tomato species have been shown to contain genotypes resistant to ToCV, there are no commercially available resistant or tolerant tomato varieties to date.

A non-persistent aphid-transmitted Potyvirus differentially alters the vector and non-vector biology through host plant quality manipulation.

The association of plant viruses with their vectors has significant implications for virus transmission and spread. Only a few studies, with even fewer pathosystems, have explored non-persistent (NP) virus-vector interactions that are presumed to be transient. We studied how a NP virus, Papaya ringspot virus (PRSV) influenced the behavior and biology of its vector, the melon aphid (Aphis gossypii Glover) and the non-vector, silverleaf whitefly (Bemisia tabaci Gennadius). We also assessed whether the fitness effects on aphids are modulated through changes in the host plant, squash (Cucurbita pepo L.) nutrient profile. The overall performance of A. gossypii was substantially higher on PRSV-infected plants, along with increased arrestment on PRSV-infected than non-infected plants. No such PRSV-modulated fitness effects were observed with B. tabaci. PRSV-infected plants had increased concentrations of free essential amino acids: threonine, arginine and lysine; non-essential amino acids: glycine and homocysteine; and soluble carbohydrates: galactose, raffinose and cellobiose. In general, PRSV encouraged long-term feeding and enhanced fitness of A. gossypii through host plant nutrient enrichment. These findings provide evidence for a NP virus mediated positive fitness effects on its vector, with no spillover fitness benefits to the non-vector within the same feeding guild.

Rice dwarf virus infection alters green rice leafhopper host preference and feeding behavior.

Host plants, pathogens and their herbivore vectors systems have complex relationships via direct and indirect interactions. Although there are substantial gaps in understanding these systems, the dynamics of the relationships may influence the processes of virus transmission and plant disease epidemics. Rice dwarf virus (RDV) is mainly vectored by green rice leafhoppers (GRLHs), Nephotettix cincticeps (Uhler) (Hemiptera: Cicadellidae) in a persistently circulative manner. In this study, host plant selection preferences of non-viruliferous and viruliferous (carrying RDV) GRLHs between RDV-free and RDV-infected plants were tested. Non-viruliferous GRLHs preferred RDV-infected rice plants over RDV-free rice plants, and viruliferous GRLHs preferred RDV-free rice plants over RDV-infected rice plants. In odor selection preference bioassay using a four-field olfactometer, non-viruliferous GRLHs preferred odors of RDV-infected rice plants over healthy rice and viruliferous GRLHs preferred odors of RDV-free rice plants over RDV-infected ones. In 6 h plant penetration behavior bioassay using electrical penetration graphs, non-viruliferous GRLHs spent shorter time in non-penetration and much longer time in xylem feeding on RDV-infected, compared to RDV-free rice plants. Viruliferous GRLHs exhibited more salivation and stylet movement on RDV-free rice plants than on RDV-infected rice plants. We infer from these findings that RDV influences these vector behaviors by altering host plant physiology to promote viral transmission.

A systematic review of human pathogens carried by the housefly (Musca domestica L.).

BACKGROUND: The synanthropic house fly, Musca domestica (Diptera: Muscidae), is a mechanical vector of pathogens (bacteria, fungi, viruses, and parasites), some of which cause serious diseases in humans and domestic animals. In the present study, a systematic review was done on the types and prevalence of human pathogens carried by the house fly. METHODS: Major health-related electronic databases including PubMed, PubMed Central, Google Scholar, and Science Direct were searched (Last update 31/11/2017) for relevant literature on pathogens that have been isolated from the house fly. RESULTS: Of the 1718 titles produced by bibliographic search, 99 were included in the review. Among the titles included, 69, 15, 3, 4, 1 and 7 described bacterial, fungi, bacteria+fungi, parasites, parasite+bacteria, and viral pathogens, respectively. Most of the house flies were captured in/around human habitation and animal farms. Pathogens were frequently isolated from body surfaces of the flies. Over 130 pathogens, predominantly bacteria (including some serious and life-threatening species) were identified from the house flies. Numerous publications also reported antimicrobial resistant bacteria and fungi isolated from house flies. CONCLUSIONS: This review showed that house flies carry a large number of pathogens which can cause serious infections in humans and animals. More studies are needed to identify new pathogens carried by the house fly.

Description of a new species of Nesotriatoma Usinger, 1944 from Cuba and revalidation of synonymy between Nesotriatoma bruneri (Usinger, 1944) and N. flavida (Neiva, 1911) (Hemiptera, Reduviidae, Triatominae).

Nesotriatoma confusa sp. nov. (Hemiptera, Reduviidae, Triatominae) is described based on specimens from Cuba. From one male, one female, and eleven nymphs of a then-undescribed species of Nesotriatoma collected in Cuba, a colony was formed and its specimens were used to describe N. confusa sp. nov. Characters were observed on the head, thorax, abdomen, female external genitalia, and male genitalia with optical microscopy and scanning electron microscopy. We concluded that N. bruneri (Usinger, ) was indeed a synonym of N. flavida (Neiva, ) as previously proposed.

Entomological surveys of Lutzomyia flaviscutellata and other vectors of cutaneous leishmaniasis in municipalities with records of Leishmania amazonensis within the Bragança region of Pará State, Brazil.

In southeast Amazon, Lutzomyia (Nyssomyia) flaviscutellata is the incriminated vector of Leishmania (Leishmania) amazonensis, a causative agent of zoonotic cutaneous leishmaniasis (CL). The optimal methods for surveying Lu. flaviscutellata were investigated in the Bragança region, northeast Pará State, Brazil, selected for the presence of Le. amazonensis. The performances of modified Disney traps and CDC light traps were compared in four ecotopes within and around four village transects during the wet and dry seasons. The physiological age of female sand flies was estimated and natural infection by flagellates was evaluated by dissection. Disney traps were better for detecting the presence of Lu. flaviscutellata, while CDC traps performed well for detecting Lutzomyia (Nyssomyia) antunesi, suspected vector of Leishmania lindenbergi. The former was more abundant during the wet season, when female flies were naturally infected with Le. amazonensis. These findings identified the environments of local transmission. In order to improve surveys of Lu. flaviscutellata as part of integrated epidemiological surveillance of CL, our recommendations include focusing vector surveys with Disney traps on forest fragments where people work, during the seasonal peak of the vector. Further field studies are required to make model-based predictions of seasonal variations in the vectorial capacity of vector populations.

Nuevos retos en gestión de vectores en salud pública: la mosca negra en Murcia (España) / New public health challenges in vector management: black flies in Murcia (Spain)

Hasta ahora no se habían detectado molestias causadas por simúlidos (Diptera: Simuliidae) en la Región de Murcia. En septiembre de 2016, el ayuntamiento de Ojós contactó con la Dirección General de Salud Pública en relación con una plaga de insectos en la localidad, probablemente mosquitos. Tras muestrear con una trampa BG-sentinel 2, capturar insectos adultos con aspirador entomológico, y larvas y pupas en muestras de cañas sumergidas, la especie implicada se identificó como Simulium sergenti. Esta especie no parece ser especialmente antropófila y, por tanto, el riesgo para la salud de los vecinos se consideró bajo, pero las molestias causadas por la gran cantidad de moscas afectaron a la calidad de vida de las personas en la localidad. Se recomendó un plan de gestión de simúlidos para la zona afectada, tratando el río con Bacillus thuringiensis var. Israelensis (AU)

Historically, no black fly (Diptera: Simuliidae) nuisance has been reported in the Murcia Region. Back in September 2016 the Ojós City Council has contacted the Regional Public Health General Directorate for help regarding a local insect nuisance, most probably based on mosquitoes. After sampling with a BG-sentinel 2 trap, collecting adult specimens with an entomological aspirator, and collect larvae and pupae on submerged giant cane stalks at the river, it turned out that Simulium sergenti was the insect species causing the nuisance. This species is not considered particularly anthropophilic; therefore, a low vector risk for human health was considered. However, the high fly density impaired the life quality of the people at the village. A management plan was recommended, treating the river with Bacillus thuringiensis var israelensis (AU)

An advection-deposition-survival model to assess the risk of introduction of vector-borne diseases through the wind: Application to bluetongue outbreaks in Spain.

This work develops a methodology for estimating risk of wind-borne introduction of flying insects into a country, identifying areas and periods of high risk of vector-borne diseases incursion. This risk can be characterized by the role of suitable temperatures and wind currents in small insects' survival and movements, respectively. The model predicts the number density of introduced insects over space and time based on three processes: the advection due to wind currents, the deposition on the ground and the survival due to climatic conditions. Spanish livestock has suffered many bluetongue outbreaks since 2004 and numerous experts point to Culicoides transported by wind from affected areas in North Africa as a possible cause. This work implements numerical experiments simulating the introduction of Culicoides in 2004. The model identified southern and eastern Spain, particularly between June and November, as being at greatest risk of wind-borne Culicoides introduction, which matches field data on bluetongue outbreaks in Spain this year. This validation suggests that this model may be useful for predicting introduction of airborne pathogens of significance to animal productivity.

What's eating you? sand flies.

As thousands of Americans descended upon Brazil for the Olympic games in the summer of 2016, the mosquito-borne Zika virus became a source of great concern among the countless athletes and travelers in Rio. As is often the case, the media frenzy that ensued drew travelers' attention away from a lesser known flying vector that often carries with it grave consequences. The Phlebotominae, commonly known as sand flies, are biting insects known for their ability to transmit the protozoa Leishmania as well as a number of other viruses and bacteria. As the impact of sand flies continues to grow in the United States and worldwide, knowledge of the vector is important for proper treatment and prevention of the diseases they carry.

Entomofagia: ¿una alternativa a nuestra dieta tradicional? / Entomophagy: an alternative to our traditional diet?

El autor comenta el interés creciente hacia dietas exóticas, entre las cuales destaca la entomofagia. Tras describir las propiedades de los insectos, como alimento, el autor advierte del riesgo que puede suponer la ingesta de artrópodos cuando no se observan las debidas garantías sanitarias. La entomofagia puede ser, no obstante, una posibilidad ante una carestía ocasional, sequías, hambrunas y conflictos bélicos o un hipotético desastre nuclear. Las granjas experimentales de insectos, como fuente de proteínas, podrían aportar conocimientos de interés para las fuerzas armadas

The author comments on the growing interest in exotic diets, including entomophagy. After describing the properties of insects as food, the author warns of the risk that arthropod intake may entail when due sanitary guarantees are not observed. However, entomophagy may be a possibility in the face of occasional starvation, drought, starvation and war or a hypothetical nuclear disaster. Experimental insect farms, as a source of protein, could bring knowledge of interest to the armed forces

Metabolic alterations in the nymphal instars of Diaphorina citri induced by Candidatus Liberibacter asiaticus, the putative pathogen of huanglongbing.

Currently, huanglongbing is the most damaging disease of citrus causing huge economic losses. The disease is caused by the Gram-negative bacterium Candidatus Liberibacter asiaticus (CLas). The pathogen is transmitted in a persistent propagative circulative manner within its vector, the Asian citrus psyllid, Diaphorina citri. Exploring the metabolic alteration in the vector may lead to a better understanding of the nutritional needs of CLas and to designing an artificial medium for culturing the pathogen. It has been shown that the nymphal stages have a greater role in transmission mainly because they feed on plants more actively than adults. In this study, we carried out an untargeted comparative metabolomic analysis for healthy and CLas-infected 4th / 5th instar nymphs. The metabolic analysis was performed using trimethylsilylation and methyl chloroformate derivatization followed by Gas Chromatography-Mass Spectrometry (GC-MS). Overall, the changes in the nymph metabolism due to the infection with CLas were more pronounced than in adults, as we previously published. Nymphs reared on CLas-infected Valencia sweet orange were higher in many metabolites, mainly those of the TCA cycle, C16 and C18 fatty acids, glucose, sucrose, L-proline, L-serine, pyroglutamic acid, saccharic acid, threonic acid and myo-inositol than those reared on healthy plants. In contrast, CLas-infected nymphs were lower in putrescine, glycine, L -phenylalanine, L -tyrosine, L -valine, and chiro-inositol. The information provided from this study may contribute in acceleration of the availability of CLas in culture and consequent screening of antibacterial compounds to discover a definitive solution for huanglongbing.