Acquisition Feeding Research Articles

Phylogenetic evaluation and haplotype mapping of phytoplasma disease in sesame: insights into leafhopper-mediated disease transmission through molecular investigation

The predominance of leafhopper species like Orosius albicinctus, Hishimonus phycitis, and Amrasca biguttula biguttula indeed poses a significant threat to sesame production in India and globally. This disease is a major challenge for sesame farming, as it can lead to yield reductions of up to 95%. The current study examined phytoplasma and leafhoppers in sesame and analyzed the phylogenetic differences in the phyllody phytoplasma. The identification of both 16Sr group I and group II phytoplasma isolates, along with the discovery of the 16SrI-B group reported for the first time in Tamil Nadu, represents a significant advancement in our understanding of phytoplasma diversity in the region. The utilization of virtual RFLP analysis in sequencing the sesame phyllody phytoplasma highlights the importance of molecular techniques in characterizing the phytoplasma. Leafhoppers in sesame were a co-occurring species complex of O. albicinctus, H. phycitis, and A. bigutulla bigutulla. The leafhoppers were identified using species-specific taxonomical traits, and a mtCOI gene-based genetic study confirmed the species. In disease transmission efficiency studies in the greenhouse, O. albicinctus transmitted phyllody at a rate ranging from 7.8% to 84%, while H. phycitis transmitted phyllody at a rate of 5.2% to 74%. Both of these species showed the highest transmission efficiency in the combination of three insects per plant with five days of acquisition feeding and one day of inoculation feeding. A comprehensive understanding of leafhopper-phytoplasma interactions through molecular studies supports effective monitoring, early detection, and the development of integrated pest management strategies.

Enhancing the Robustness of Rib-Groove Filling and Strain Homogeneity in the Isothermal Forging of Titanium Alloy Multi-Rib Components

Isothermal forging stands as an effective technology for the production of large-scale titanium alloy multi-rib components. However, challenges have persisted, including die underfilling and strain concentration due to the complex material flow and heterogeneous deformation within the forging die cavity. While approaches centered on optimized billet designs have mitigated these challenges, uncertainties in process parameters continue to introduce unacceptable variations in forming accuracy and stability. To tackle this issue, this study introduced a multi-objective robust optimization approach for billet design, accounting for the multi-rib eigenstructure and potential uncertainties. The approach includes finite element (FE) modeling for analyzing the die-filling and strain inhomogeneity within the multi-rib eigenstructure. Furthermore, it integrated image acquisition perception and feed back technologies (IAPF) for real-time monitoring of material flow and filling sequences within die rib-grooves, validating the accuracy of the FE modeling. By incorporating dimensional parameters of the billet and uncertainty factors, including friction, draft angle, forming temperature, speed, and deviations in billet and die, quantitative analyses on the rib-groove filling and strain inhomogeneity with fluctuation were conducted. Subsequently, a dual-response surface model was developed for statistical analysis of the cavity filling and strain homogeneity. Finally, the robust optimization was processed using a non-dominated sorting genetic algorithm II (NSGA-II) and validated using the IAPF technologies. The proposed approach enables robust design enhancements for rib-groove filling and strain homogeneity in titanium alloy multi-rib components.

New heterodont odontocetes from the Oligocene Pysht Formation in Washington State, U.S.A., and a reevaluation of Simocetidae (Cetacea, Odontoceti).

Odontocetes first appeared in the fossil record by the early Oligocene, and their early evolutionary history can provide clues as to how some of their unique adaptations, such as echolocation, evolved. Here, three new specimens from the early to late Oligocene Pysht Formation are described further increasing our understanding of the richness and diversity of early odontocetes, particularly for the North Pacific. Phylogenetic analysis shows that the new specimens are part of a more inclusive, redefined Simocetidae, which now includes Simocetus rayi, Olympicetus sp. 1, Olympicetus avitus, O. thalassodon sp. nov., and a large unnamed taxon (Simocetidae gen. et sp. A), all part of a North Pacific clade that represents one of the earliest diverging groups of odontocetes. Amongst these, Olympicetus thalassodon sp. nov. represents one of the best known simocetids, offering new information on the cranial and dental morphology of early odontocetes. Furthermore, the inclusion of CCNHM 1000, here considered to represent a neonate of Olympicetus sp., as part of the Simocetidae, suggests that members of this group may not have had the capability of ultrasonic hearing, at least during their early ontogenetic stages. Based on the new specimens, the dentition of simocetids is interpreted as being plesiomorphic, with a tooth count more akin to that of basilosaurids and early toothed mysticetes, while other features of the skull and hyoid suggest various forms of prey acquisition, including raptorial or combined feeding in Olympicetus spp., and suction feeding in Simocetus. Finally, body size estimates show that small to moderately large taxa are present in Simocetidae, with the largest taxon represented by Simocetidae gen. et sp. A with an estimated body length of 3 m, which places it as the largest known simocetid, and amongst the largest Oligocene odontocetes. The new specimens described here add to a growing list of Oligocene marine tetrapods from the North Pacific, further promoting faunistic comparisons across other contemporaneous and younger assemblages, that will allow for an improved understanding of the evolution of marine faunas in the region.

Effect of endophytic fungi on the ability of Aphis craccivora Koch. in transmitting Bean common mosaic virus

Bean common mosaic virus (BCMV) and its aphid vector, Aphis craccivora, are essential limiting factors in yardlong bean production. The previous study showed that endophytic fungi (EF), Lecanicillium lecanii (LL), and H5 could suppress virus titer in BCMV-infected plants by its vector Aphis craccivora. Therefore, elucidation of its mechanisms is a necessary study. This research aimed to study the ability of aphids reared on EF-treated and untreated control plants to transmit BCMV. The adult aphids had been raised on EF-treated yard-long beans for three generations. The 3rd generation of adult aphids was used as vectors to transmit BCMV after acquisition feeding on infected plants and inoculating feeding on healthy plants. The disease assessment parameters were observed at four weeks post-transmission. Generally, the population number of aphids on EF-treated plants was significantly lower compared to the control. The transmission of BCMV using the 3rd generation of aphids reared on EF-treated plants affected by the prolonged incubation period, lower disease incidence and severity score, and significantly lower virus titer compared to control. Moreover, the detection of BCMV by ELISA in aphids after virus transmission showed an adverse reaction, indicating that the virus from acquisition feeding was transmitted to plants during inoculation feeding. These results revealed that rearing aphids on EF-treated plants interfere with the aphids’ ability to transmit BCMV by decreasing virus acquired during acquisition and inoculation feeding.

Haemaphysalis flava ticks as a competent vector of severe fever with thrombocytopenia syndrome virus

Severe fever with thrombocytopenia syndrome virus (SFTSV), a tick-borne Bunyavirus, causes an emerging hemorrhagic fever in humans with a high fatality in Asia. The tick vectors and hosts of SFTSV are not well studied. We evaluated SFTSV transmission in laboratory reared Haemaphysalis flava ticks. RT-PCR demonstrated that after acquisition feeding in SFTSV-infected rabbits, 10 % (4/40) engorged larvae, 25% (5/20) engorged nymphs, and 50% (5/10) engorged females of H. flava became SFTSV RNA positive; after engorged larvae and nymphs molted into nymphs and adults, respectively, 12.5% (3/24) newly molted nymphs and 20% (2/10) newly molted adults were SFTSV RNA positive. Among 30 engorged females that oviposited, 10% (3/30) clutches of eggs and 3.3% (1/30) colonies of larvae were RNA positive for SFTSV. RT-PCR also showed that 6 days after being infested with SFTSV-infected ticks, 100% (3/3) rabbits infested with larvae, 100% (2/2) rabbits infested with nymphs, and 100% (2/2) rabbits infested with adult ticks became SFTSV RNA positive. In conclusion, H. flava can acquire SFTSV from infected rabbits by feeding; there is transstadial and transovarial transmission of the virus and all three stages of H. flava can transmit SFTSV to rabbits by feeding. Thus, H. flava tick is an effective vector of SFTSV and may play a role in the transmission of SFTSV in wild animals and humans.

Ecologically relevant radiation exposure triggers elevated metabolic rate and nectar consumption in bumblebees

Abstract Exposure to radiation is a natural part of our environment. Yet, due to nuclear accidents such as at Chernobyl, some organisms are exposed to significantly elevated dose rates. Our understanding of the effects of radiation in the environment is limited, confounded by substantial interspecific differences in radio‐sensitivity and conflicting findings. Here we study radiation impacts on bumblebees in the laboratory using principles from life‐history theory, which assume organismal investment in fitness‐related traits is constrained by resource availability and resource allocation decisions. To investigate how chronic radiation might negatively affect life‐history traits, we tested whether exposure affects bumblebee energy budgets by studying resource acquisition (feeding) and resource use (metabolic rate). We monitored metabolic rate, movement and nectar intake of bumblebees before, during and after 10 days of radiation exposure. Subsequently, we monitored feeding and body mass across a dose rate gradient to investigate the dose rate threshold for these effects. We studied dose rates up to 200 μGy/hr: a range found today in some areas of the Chernobyl Exclusion Zone. Chronic low‐dose radiation affected bumblebee energy budgets. At 200 μGy/hr nectar consumption elevated by 56% relative to controls, metabolic CO2 production increased by 18%, and time spent active rose by 30%. Once radiation exposure stopped, feeding remained elevated but CO2 production and activity returned to baseline. Our analysis indicates that elevated metabolic rate was not driven by increased activity but was instead closely associated with feeding increases. Our data suggest bumblebee nectar consumption was affected across the 50–200 μGy/hr range. We show field‐realistic radiation exposure influences fundamental metabolic processes with potential to drive changes in many downstream life‐history traits. We hypothesise that radiation may trigger energetically costly repair mechanisms, increasing metabolic rate and nectar requirements. This change could have significant ecological consequences in contaminated landscapes, including Chernobyl. We demonstrate bumblebees are more sensitive to radiation than assumed by existing international frameworks for environmental radiological protection. Read the free Plain Language Summary for this article on the Journal blog.

Studies on Transmission of Phytoplasma in Sesame

The experiment was conducted to artificially transmit phytoplasma, the causal organism of sesame phyllody by mechanical sap inoculation and with insect vectors (Orosius albicinctus and Hishimonus phycitis). The experimental findings revealed that the phytoplasma could not be transmitted by sap inoculation. The mean transmission rate was found to be 54.67 per cent, with the highest rate of phyllody transmission (93.33%) observed in both treatments T8 and T9 with 3 insects/plant + 7 Days of Acquisition Feeding (DAF) + 5 Days of Inoculation Feeding (DIF) and 3 insects/plant + 7 DAF+ 7 DIF, respectively, while the lowest was observed in the treatment T1 with 3 insects/plant + 3 DAF+ 3 DIF (13.3%) phyllody transmission from infected to healthy sesame plants. All the treatments were found to be statistically significant over control, however, the treatments T8 and T9 with three insects/plant and with acquisition feeding period of 7 days and inoculation feeding of 5 and 7 days, respectively were found to be at par with each other statistically.

Response of banana germplasms to banana bunchy top virus

Banana bunchy top virus (BBTV) is one of the important viruses causing disease in bananas and its infection has the potential to cause yield loss. This study was conducted to evaluate the response of several commercial cultivars (Cavendish, Bebek, Goroho, Tanduk, and Barangan Merah) and wild accessions (Klutuk NTT, Halabanensis, SPn 001, LNT 001, and Microcarpa) of banana to BBTV infection. Transmission of BBTV was carried out through banana aphid Pentalonia nigronervosa, using 20 adult aphids per plant with an acquisition feeding period of 24 hours on BBTV-infected plants and an inoculation feeding period on healthy test plants for 48 hours. Observation on plant growth and disease intensity was conducted for 8 weeks after inoculation. At the end of the observation period, only 5 cultivars, i.e. Cavendish, Bebek, Goroho, Barangan Merah, and Halabanensis showed typical symptoms of BBTV with disease incidence reached 80%, 60%, 20%, 20%, and 20% respectively. Significant inhibition of plant height and leaves width occurred in Cavendish, Bebek, and Goroho i.e. 44.60%, 36.31%; 12.62%, 41.08%; and 25%, 10.13%, respectively. This paper discusses the need for banana germplasm exploration to find sources of resistance to BBTV.

How many begomovirus copies are acquired and inoculated by its vector, whitefly (Bemisia tabaci) during feeding?

Begomoviruses are transmitted by whitefly (Bemisia tabaci Gennadius, Hemiptera: Aleyrodidae) in a persistent-circulative way. Once B. tabaci becomes viruliferous, it remains so throughout its life span. Not much is known about the copies of begomoviruses ingested and/or released by B. tabaci during the process of feeding. The present study reports the absolute quantification of two different begomoviruses viz. tomato leaf curl New Delhi virus (ToLCNDV, bipartite) and chilli leaf curl virus (ChiLCV, monopartite) at different exposure of active acquisition and inoculation feeding using a detached leaf assay. A million copies of both the begomoviruses were acquired by a single B. tabaci with only 5 min of active feeding and virus copy number increased in a logarithmic model with feeding exposure. Whereas, a single B. tabaci could inoculate 8.21E+09 and 4.19E+11 copies of ToLCNDV and ChiLCV, respectively in detached leaves by 5 min of active feeding. Virus copies in inoculated leaves increased with an increase in feeding duration. Comparative dynamics of these two begomoviruses indicated that B. tabaci adult acquired around 14-fold higher copies of ChiLCV than ToLCNDV 24 hrs post feeding. Whereas, the rate of inoculation of ToLCNDV by individual B. tabaci was significantly higher than ChiLCV. The study provides a better understanding of begomovirus acquisition and inoculation dynamics by individual B. tabaci and would facilitate research on virus-vector epidemiology and screening host resistance.

Plasticity of the lettuce infectious yellows virus minor coat protein (CPm) in mediating the foregut retention and transmission of a chimeric CPm mutant by whitefly vectors.

Transmission of the crinivirus, lettuce infectious yellows virus (LIYV), is determined by a minor coat protein (CPm)-mediated virion retention mechanism located in the foregut of its whitefly vector. To better understand the functions of LIYV CPm, chimeric CPm mutants engineered with different lengths of the LIYV CPm amino acid sequence and that of the crinivirus, lettuce chlorosis virus (LCV), were constructed based on bioinformatics and sequence alignment data. The 485 amino acid-long chimeric CPm of LIYV mutant, CPmP-1, contains 60 % (from position 3 to 294) of LCV CPm amino acids. The chimeric CPm of mutants CPmP-2, CPmP-3 and CPmP-4 contains 46 (position 3 to 208), 51 (position 3 to 238) and 41 % (position 261 to 442) of LCV CPm amino acids, respectively. All four mutants moved systemically, expressed the chimeric CPm and formed virus particles. However, following acquisition feeding of the virus preparations, only CPmP-1 was retained in the foreguts of a significant number of vectors and transmitted. In immuno-gold labelling transmission electron microscopy (IGL-TEM) analysis, CPmP-1 particles were distinctly labelled by antibodies directed against the LCV but not LIYV CPm. In contrast, CPmP-4 particles were not labelled by antibodies directed against the LCV or LIYV CPm, while CPmP-2 and -3 particles were weakly labelled by anti-LIYV CPm but not anti-LCV CPm antibodies. The unique antibody recognition and binding pattern of CPmP-1 was also displayed in the foreguts of whitefly vectors that fed on CPmP-1 virions. These results are consistent with the hypothesis that the chimeric CPm of CPmP-1 is incorporated into functional virions, with the LCV CPm region being potentially exposed on the surface and accessible to anti-LCV CPm antibodies.

Nuances of Whitefly Vector-Crinivirus Interactions Revealed in the Foregut Retention and Transmission of Lettuce Chlorosis Virus by Two Bemisia tabaci Cryptic Species.

Lettuce infectious yellows virus is the first crinivirus for which the retention of purified virions ingested into the whitefly (Bemisia tabaci New World (NW)) vector’s foregut, has been demonstrated to be a requisite for successful virus transmission. This key finding supports the hypothesis that the determinant of foregut retention and transmission is present on the virion itself. However, whether this is also true for other criniviruses has not been established. Here, we provide evidence that lettuce chlorosis virus (LCV) acquired from plants is retained in the foreguts of both the B. tabaci NW and Middle East–Asia Minor 1 (MEAM1) vector species and transmitted upon inoculation feeding. An association between foregut retention and transmission by NW vectors is also observed following the acquisition and inoculation feeding of LCV virions purified using a standard procedure involving 2% or 4% (v/v) Triton™ X-100 (TX-100). However, while virions purified with 2% or 4% TX-100 are also retained in the foreguts of MEAM1 vectors, transmission is observed with the 4% TX-100-purified virions or when more vectors are used for acquisition and inoculation feeding. These results suggest that an intrinsic difference exists between NW and MEAM1 vectors in their interactions with, and transmission of, LCV virions.

A U.S. isolate of Theileria orientalis, Ikeda genotype, is transmitted to cattle by the invasive Asian longhorned tick, Haemaphysalis longicornis

BackgroundTheileria orientalis is a tick-borne hemoparasite that causes anemia, ill thrift, and death in cattle globally. The Ikeda strain of T.orientalis is more virulent than other strains, leading to severe clinical signs and death of up to 5% of affected animals. Within the Asia–Pacific region, where it affects 25% of Australian cattle, T.orientalis Ikeda has a significant economic impact on the cattle industry. In 2017, T.orientalis Ikeda was detected in a cattle herd in Albermarle County, Virginia, United States. Months earlier, the U.S. was alerted to the invasion of the Asian longhorned tick, Haemaphysalis longicornis, throughout the eastern U.S. Abundant H.longicornis ticks were identified on cattle in the T.orientalis-affected herd in VA, and a subset of ticks from the environment were PCR-positive for T.orientalis Ikeda. A strain of T.orientalis from a previous U.S. outbreak was not transmissible by H.longicornis; however, H.longicornis is the primary tick vector of T.orientalis Ikeda in other regions of the world. Thus, the objective of this study was to determine whether invasive H.longicornis ticks in the U.S. are competent vectors of T.orientalis Ikeda.MethodsNymphal H.longicornis ticks were fed on a splenectomized calf infected with the VA-U.S.-T.orientalis Ikeda strain. After molting, a subset of adult ticks from this cohort were dissected, and salivary glands assayed for T.orientalis Ikeda via qPCR. The remaining adult ticks from the group were allowed to feed on three calves. Calves were subsequently monitored for T.orientalis Ikeda infection via blood smear cytology and PCR.ResultsAfter acquisition feeding on a VA-U.S.-T.orientalis Ikeda-infected calf as nymphs, a subset of molted adult tick salivary glands tested positive by qPCR for T.orientalis Ikeda. Adult ticks from the same cohort successfully transmitted T.orientalis Ikeda to 3/3 naïve calves, each of which developed parasitemia reaching 0.4–0.9%.ConclusionsOur findings demonstrate that U.S. H.longicornis ticks are competent vectors of the VA-U.S.-T.orientalis Ikeda strain. This data provides important information for the U.S. cattle industry regarding the potential spread of this parasite and the necessity of enhanced surveillance and control measures.Graphical

Virus Vector Relationship of Yellow Mosaic Virus and Whitefly, Bemisia tabaci (Gennadius) in Soybean

Background: Whitefly is one of the most destructive sucking pest in the tropical and subtropical regions of the world and causing significant crop losses directly by sucking sap from the plants and indirectly through the transmission of viral diseases specifically caused by the genus Begomovirus. The Begomovirus species viz., Mungbean yellow mosaic India virus (MYMIV) and Mungbean yellow mosaic virus (MYMV) are causing yellow mosaic virus disease in soybean, which is transmitted by whiteflies. The disease accounts to 30-70 per cent yield loss and increases up to 80 - 100 per cent during severe incidence. Hence, there is a need for development of integrated pest management strategies against disease and whiteflies, for this the knowledge on virus-vector relationship is required. But, the studies on biological relationship of yellow mosaic virus disease and whitefly in soybean are scarce. At this juncture, considering the importance of disease in soybean, the present investigation was carried out to know the virus -vector relationship of the YMV and whitefly in soybean.Methods: The experiment on virus-vector relationship of yellow mosaic virus and whitefly in soybean was conducted at Department of Entomology, College of Agriculture, G. B. Pant University of Agriculture and Technology, Pantnagar, Uttarakhand during 2016-17. The data on number of whiteflies per plant, acquisition and inoculation access feeding period and pre and post starvation period required for effective transmission of virus was recorded.Result: A single viruliferous whitefly was able to transmit virus and ten viruliferous whiteflies per plant were required for cent per cent transmission of virus. The minimum acquisition access and inoculation access feeding periods required for virus transmission was 0.25h (15 min) each; while the 100 per cent virus transmission was recorded with acquisition and inoculation period of 12h, each. The per cent transmission was increased with the increase of acquisition and inoculation periods. The rate of transmission was positively correlated with pre-acquisition starvation period and negatively correlated with post- acquisition starvation period.

Capybaras (Hydrochoerus hydrochaeris) as amplifying hosts of Rickettsia rickettsii to Amblyomma sculptum ticks: Evaluation during primary and subsequent exposures to R. rickettsii infection

Rickettsia rickettsii is the agent of Brazilian spotted fever (BSF), the most lethal tick-borne disease in the western hemisphere. Amblyomma sculptum is the main vector of R. rickettsii in southeastern Brazil. Capybaras act as primary hosts for this tick species, and as amplifying hosts for R. rickettsii, creating new infected lineages of A. sculptum in BSF-endemic areas. In a recent study, we demonstrated that capybaras successively exposed to R. rickettsii-infected A. sculptum ticks developed a mean rickettsemic period of 9.2 days during primary infection, but no rickettsemia during subsequent expositions, when capybaras were immune to R. rickettsii. During the primary and subsequent infections, capybaras were also infested with uninfected A. sculptum ticks. These infestations compose the present study, which aimed to evaluate: (i) if either rickettsemic or non-rickettsemic capybaras could serve as infection sources of R. rickettsii for A. sculptum larvae and nymphs; (ii) the vector competence of the subsequent nymphs and adults (molted from the engorged ticks collected from capybaras); and (iii) if there were R. rickettsii-acquisition by uninfected ticks co-feeding with infected ticks on immune capybaras (without rickettsemia). Through experimental infection of capybaras with R. rickettsii via tick feeding, simulating a natural condition, we demonstrated that primarily infected capybaras developed rickettsemia that resulted in successful acquisition feeding of larvae and nymphs of A. sculptum, since part of these ticks maintained R. rickettsii transstadially, and the resultant molted ticks (either nymphs or adults) successfully transmitted the bacterium by feeding on susceptible rabbits. Contrastingly, all rabbit infestations with ticks derived from acquisition feeding on R. rickettsii-immune capybaras (including when acquisition ticks fed in direct contact with donor ticks) evidenced absence of R. rickettsii transmission due to lack of clinical signs and antibody response in those rabbits. Our results indicate that capybaras could serve as R. rickettsii-amplifying hosts for A. sculptum ticks only during the capybara’s primary infection, but not during subsequent infections on immune capybaras. Finally, the probable co-feeding nonsystemic transmission of R. rickettsii seems to be irrelevant in the context of BSF epidemiology, in areas where capybaras are incriminated as main amplifying hosts of R. rickettsii for A. sculptum ticks.

Quantitative analysis of Anaplasma marginale acquisition and transmission by Dermacentor andersoni fed in vitro

In this study, we describe a new in vitro tick feeding system that facilitates the study of ticks and tick-borne pathogens. To optimize the system, we used Dermacentor andersoni and Anaplasma marginale as a tick-pathogen interaction model. Ticks were fed on bovine blood containing 10-fold dilutions of the pathogen to determine the effect of dose on tick infection rate. After feeding on infected blood, ticks were transferred to uninfected blood to stimulate bacterial replication within the tick vector. During stimulation feeding, blood samples were collected daily to determine if infected ticks secreted viable A. marginale. The results demonstrated similar attachment rates between the first and second tick feeding. Tick midgut and salivary glands were infected with A. marginale. However, salivary gland infection rates decreased as the percentage of parasitized erythrocytes decreased during tick acquisition feeding. Bacteria recovered from the in vitro system were able to infect a naïve bovine host. Using the highly transmissible A. marginale St. Maries strain, we demonstrated that the artificial tick feeding system is a suitable tool to study tick-pathogen interactions and that A. marginale tick salivary gland infection is dose dependent. This work demonstrates the utility of an artificial tick feeding system to directly study the association between the number of acquired pathogens and transmissibility by ticks.

Differences in gene expression in whitefly associated with CYSDV-infected and virus-free melon, and comparison with expression in whiteflies fed on ToCV- and TYLCV-infected tomato

BackgroundCucurbit yellow stunting disorder virus (CYSDV; genus Crinivirus, Closteroviridae) is transmitted in a semipersistent manner by the whitefly, Bemisia tabaci, and is efficiently transmitted by the widely prevalent B. tabaci cryptic species, MEAM1. In this study, we compared transcriptome profiles of B. tabaci MEAM1, after 24 h, 72 h and 7 days of acquisition feeding on melon plants infected with CYSDV (CYSDV-whiteflies) with those fed on virus-free melon, using RNA-Seq technology. We also compared transcriptome profiles with whiteflies fed on tomato plants separately infected with Tomato chlorosis virus (ToCV), a crinivirus closely related to CYSDV, and Tomato yellow leaf curl virus (TYLCV), a member of the genus Begomovirus, which has a distinctly different mode of transmission and their respective virus-free controls, to find common gene expression changes among viruliferous whiteflies feeding on different host plants infected with distinct (TYLCV) and related (CYSDV and ToCV) viruses.ResultsA total of 275 differentially expressed genes (DEGs) were identified in CYSDV-whiteflies, with 3 DEGs at 24 h, 221 DEGs at 72 h, and 51 DEGs at 7 days of virus acquisition. Changes in genes encoding orphan genes (54 genes), phosphatidylethanolamine-binding proteins (PEBP) (20 genes), and AAA-ATPase domain containing proteins (10 genes) were associated with the 72 h time point. Several more orphan genes (20 genes) were differentially expressed at 7 days. A total of 59 common DEGs were found between CYSDV-whiteflies and ToCV-whiteflies, which included 20 orphan genes and 6 lysosomal genes. A comparison of DEGs across the three different virus-host systems revealed 14 common DEGs, among which, eight showed similar and significant up-regulation in CYSDV-whiteflies at 72 h and TYLCV-whiteflies at 24 h, while down-regulation of the same genes was observed in ToCV-whiteflies at 72 h.ConclusionsDynamic gene expression changes occurred in CYSDV-whiteflies after 72 h feeding, with decreased gene expression changes associated with 7 days of CYSDV acquisition. Similarities in gene expression changes among CYSDV-whiteflies, ToCV-whiteflies and TYLCV-whiteflies suggest the possible involvement of common genes or pathways for virus acquisition and transmission by whiteflies, even for viruses with distinctly different modes of transmission.

Squash Print Reverse Transcription Loop-Mediated Isothermal Amplification Assay for Detection of Potato leafroll virus in Single Aphid and in Potato

A squash print reverse transcription loop-mediated isothermal amplification amplification (SP-RT-LAMP) assay was developed for detection of Potato leafroll virus (PLRV) in a single aphid. Healthy aphids were given an acquisition feeding on potato plants infected with PLRV, and the acquired aphids were used for RNA isolation. The RT-LAMP assay was carried out using LAMP primers targeting the coat protein gene of PLRV. The amplified product was run on agarose gel; a typical ladder-like pattern in the acquired aphids indicated that the assay was able to detect PLRV from an individual aphid. The results of the RT-LAMP assay for the detection of PLRV in a single aphid were also visualized directly by adding fluorescent dye in LAMP amplified product. These results were confirmed by RT-PCR detection where an amplicon of expected size of 492 bp was observed in virus-acquired single aphids. The assay was optimized with respect to its isothermal amplification temperature and its incubation period. The technique was examined for its relative selectivity with other potato viruses transmitted by aphids where it did not show any cross-reactivity with the tested viruses indicating its specificity. With respect to its relative sensitivity, it was found to be equally sensitive as the existing RT-PCR assay. It was also found to be robust and highly reproducible as it was able to detect PLRV from known and unknown single aphid samples. The assay was successful in detection of PLRV in potato plants and tubers as well. Hence, we conclude that the assay is simple, sensitive and economical for detection of PLRV in single aphid and in potato plants and tubers.

Identification of sesame phyllody transmitting insect vectors in Assam, India

Different leafhopper species transmitting sesame phyllody phytoplasma were reported. Leafhoppers were collected from sesame phyllody infected field at different time interval and their population was recorded. Four leafhopper species were predominantly observed in the field conditions: Hishimonus phycitis, Exitianus indicus, Cofana unimaculata and Nephotettix nigropictus. Transmission studies were carried out by giving an acquisition and inoculation feeding period of 7 days each. Among all the leafhoppers, H. phycitis was found to transmit sesame phyllody phytoplasma (83.33%) to healthy sesame plants as confirmed by PCR and nested PCR assays followed by sequencing. The BLAST analysis of 16S rDNA sequences from H. phycitis confirmed the phytoplasma presence indicating that is clusters with the aster yellows phytoplasmas.

Evaluation of the vector competence of six ixodid tick species for Rangelia vitalii (Apicomplexa, Piroplasmorida), the agent of canine rangeliosis

Rangelia vitalii is the etiologic agent of canine rangeliosis, a severe piroplasmosis that affects domestic dogs in Brazil, Uruguay and Argentina. While R. vitalii is one of the most pathogenic tick-borne pathogens for dogs in the world, its tick vector has remained unknown. The present study evaluated the vector competence of Rhipicephalus sanguineus sensu lato (both tropical and temperate species), Amblyomma aureolatum, Amblyomma ovale, Amblyomma tigrinum, and Amblyomma sculptum for R. vitalii. These six tick species were selected for the study because they comprise the main tick species infesting dogs within the distribution area of canine rangeliosis in South America. Acquisition feeding of the above six tick species was performed on domestic dogs showing clinical signs of canine rangeliosis, after being experimentally infected through intravenous inoculation or infestation with R. vitalii-infected ticks. Thereafter, engorged ticks were evaluated for transstadial and transovarial passages of R. vitalii through molecular analysis after molting or oviposition and egg hatching. The resultant ticks were evaluated for their competence to transmit R. vitalii to susceptible dogs. Among the six tick species, only A. aureolatum was able to acquire and perpetuate R. vitalii by transstadial and transovarial passages, as demonstrated by >5% infection rates of ticks after hatching or molting. When exposed to transmission feeding, only A. aureolatum ticks were competent to transmit R. vitalii to dogs, which became severely ill, and the results confirmed by molecular methods and blood smear examination to have acquired rangeliosis. Results of the present study, coupled with epidemiological data, indicate that A. aureolatum is a natural vector of R. vitalii. Our results also indicate that R. vitalii is the first Piroplasmorida agent to be transovarially transmitted in Amblyomma ticks.

Preferensi Wereng Batang Cokelat terhadap Varietas Padi dan Ketahanan Varietas Padi terhadap Virus Kerdil Hampa

<p>Rice ragged stunt virus (RRSV) is transmitted by brown planthopper in the persistent manner. This disease in recent years has become a serious problem in Indonesia and some other countries, such as China, Vietnam, Philippines, and Thailand. Disease control is usually conducted by the vector control using insecticides, so often causes an environmental pollution. An alternative control method is using the environmentally friendly of resistant varieties. The purpose of this study was to determine the level of preference of brown planthopper (BPH) to some varieties and rice germplasms, the resistance of several varieties against rice ragged stunt virus (RRSV) disease and its effectiveness as an inoculum source of virus (RRSV). A total of 15 varieties of rice were used in preference test of BPH and resistance test to RRSV. The test for resistance varieties to RRSV was conducted by transmission of 2nd instar of BPH for 3 days of acquisition feeding period, 10 days incubation period and 24 hours inoculation period with population density of 3 BPH/plant. Disease index was calculated and used to determine the level of plant resistance. Varieties that showed resistant, moderately resistant and susceptible responses were selected and were used as a source of inoculum to be transmitted on to TN1 variety susceptible check variety to know the effectiveness of varieties as source of virus inoculum. The results showed that of the 15 varieties tested, Situ Bagendit, Utri Merah, Mentik Wangi, Mahsuri, and Inpari 1 each was less favored by BPH to settle and to multiply. Mentik Wangi, Tetep, Utri Merah, and Swarnalata each showed resistant response to RRSV. Transmission test to susceptible variety (TN1) showed that the variety of Situ Bagendit, Inpari 13, Mentik Wangi, and Tetep each has a fairly low effectiveness as a source of inoculum as indicated by the lower percentage of infection and disease index on the transmited test plants.</p>