Lysine provisioning by horizontally acquired genes promotes mutual dependence between whitefly and two intracellular symbionts

Horizontal gene transfer is widespread in insects bearing intracellular symbionts. Horizontally transferred genes (HTGs) are presumably involved in amino acid synthesis in sternorrhynchan insects. However, their role in insect-symbiont interactions remains largely unknown. We found symbionts Portiera, Hamiltonella and Rickettsia possess most genes involved in lysine synthesis in the whitefly Bemisia tabaci MEAM1 although their genomes are reduced. Hamiltonella maintains a nearly complete lysine synthesis pathway. In contrast, Portiera and Rickettsia require the complementation of whitefly HTGs for lysine synthesis and have lysE, encoding a lysine exporter. Furthermore, each horizontally transferred lysine gene of ten B. tabaci cryptic species shares an evolutionary origin. We demonstrated that Hamiltonella did not alter the titers of Portiera and Rickettsia or lysine gene expression of Portiera, Rickettsia and whiteflies. Hamiltonella also did not impact on lysine levels or protein localization in bacteriocytes harboring Portiera and ovaries infected with Rickettsia. Complementation with whitefly lysine synthesis HTGs rescued E. coli lysine gene knockout mutants. Silencing whitefly lysA in whiteflies harboring Hamiltonella reduced lysine levels, adult fecundity and titers of Portiera and Rickettsia without influencing the expression of Hamiltonella lysA. Furthermore, silencing whitefly lysA in whiteflies lacking Hamiltonella reduced lysine levels, adult fecundity and titers of Portiera and Rickettsia in ovarioles. Therefore, we, for the first time, demonstrated an essential amino acid lysine synthesized through HTGs is important for whitefly reproduction and fitness of both obligate and facultative symbionts, and it illustrates the mutual dependence between whitefly and its two symbionts. Collectively, this study reveals that acquisition of horizontally transferred lysine genes contributes to coadaptation and coevolution between B. tabaci and its symbionts.     

Genome‐wide identification and characterization of HSP gene superfamily in whitefly (Bemisia tabaci) and expression profiling analysis under temperature stress

Heat shock proteins (HSP)are essential molecular chaperones that play important roles in the stress stimulation of insects.Bemisia tabaci,a phloem feeder and invasive species,can cause extensive crop damage through direct feeding and transmission of plant viruses.Here we employed comprehensive genomics approaches to identity HSP superfamily members in the Middle East Asia Minor 1 whitefly genome.In total,we identified 26 Hsp genes,including three Hsp90,17 Hsp70,one Hsp60 and five sHSP (small heat shock protein)genes.The HSP gene superfamily of whitefly is expanded compared with the other five insects surveyed here.The gene structures among the same families are relatively conserved.Meanwhile,the motif compositions and secondary structures of BtHsp proteins were predicted.In addition,quantitative polymerase chain reaction analysis showed that the expression patterns of BtHsp gene superfamily were diverse across different tissues of whiteflies.Most Hsp genes were induced or repressed by thermal stress (40℃)and cold treatment (4℃)in whitefly.Silencing the expression of BtHsp70-6 significantly decreased the survival rate of whitefly under 45℃.All the results showed the Hsps conferred thermo-tolerance or cold-tolerance to whiteflies that protect them from being affected by detrimental temperature conditions.Our observations highlighted the molecular evolutionary properties and the response mechanism to temperature assaults of Hsp genes in whitefly.     

RNA interference mediated knockdown of juvenile hormone esterase gene in Bemisia tabaci (Gennadius): Effects on adults and their progeny

Juvenile hormone is responsible for regulating metamorphosis and reproduction in insects. Analysis of key elements of juvenile hormone regulation would enhance the understanding of this complex mechanism. Juvenile hormone esterase plays an important role in maintaining juvenile hormone titres in insects. In this study, effects of knockdown of juvenile hormone esterase gene (jhe) in Bemisia tabaci were studied using RNA interference (RNAi) technique. dsRNA corresponding to two conserved regions of jhe gene, substrate binding pocket site (jhe1), catalytic triad site (jhe2), green fluorescent protein gene (gfp) as control were synthesized. dsRNAs incorporated in artificial diet (20% sucrose solution) @ 2.5, 1.0, 0.5 and 0.1 μg/μl were fed to adult whiteflies for 48 h, followed by shifting whiteflies to live plants for next generation biology study. Based on qRT-PCR analyses, reduced jhe gene expression was observed in adult whiteflies after dsRNA feeding @ 2.5 and 1.0 μg/μl. jhe gene knockdown affects the survival and reproduction of whiteflies adversely in a dose-dependent manner. Moreover, oral feeding of dsRNA to adult whiteflies @ 2.5 and 1.0 μg/μl showed adverse effects on next generation of whitefly viz., lower egg hatchability and shortened egg incubation period. Minimum number of viable eggs (1.04 and 1.80 eggs/female) were observed when whiteflies were fed with highest concentration of dsjhe1 and dsjhe2 as compared to control (16.58 eggs/female). These data suggest that jhe gene acts as a major biological player in whitefly and its progeny and further indicate to be potential target for managing whitefly population.     

Adult survival of Delphastus catalinae (Coleoptera: Coccinellidae), a predator of whiteflies (Hemiptera: Aleyrodidae), on diets of whiteflies, honeydew, and honey

Delphastus catalinae (Horn) is a coccinellid predator that is commercially sold for the management of whiteflies. A study was conducted to assay the effect of selected diets on the survival of adult D. catalinae. Treatments of water (as a control), 10% honey, honeydew, and whiteflies [Bemisia tabaci (Gennadius)] were provided to the beetles in laboratory assays. Newly emerged, unfed adult insects were used at the start of a survival experiment with trials lasting 50 d. Another survival experiment used mixed-aged adults from a greenhouse colony, and the trials lasted 21 d. Survival was poor on a diet of solely water; ~1% survived beyond a week at 26°C. Survival using the newly emerged insects was similar between those fed honeydew and honey diets, but those on the whitefly diet had the greatest survival (~60% on day 50). However, in the experiment with mixed-aged beetles, adults on honey, and whitefly diets performed the same over a 21-d experiment. Excluding those on the water diet, survival of beetles on the various diets ranged from ~50-80% after 21 d. In an open choice assay across 7 h, D. catalinae adults were found on the whitefly diet in a much greater incidence than on the other diets, and the number of beetles found on the whitefly diet increased over time. The data supports that when D. catalinae are employed in greenhouses or fields for whitefly management, during low prey populations, honeydew from the whitefly can help sustain the population of this predator. Moreover, a supplemental food such as a honey solution can help sustain the population of D. catalinae when the prey is decreased to low numbers. These results may help in the development of strategies to enhance the utility of predators for the management of whiteflies.     

Tissue-specific gene silencing by RNA interference in the whitefly Bemisia tabaci (Gennadius)

The hemipteran whitefly Bemisia tabaci (Gennadius) species complex and the plant viruses they transmit pose major constraints to vegetable and fiber production, worldwide. The whitefly tissue- and developmental-specific gene expression has not been exhaustively studied despite its economic importance. In 2002, a functional genomic project was initiated, which generated several thousands expressed sequence tags (ESTs) and their sequence. This project provides the basic information to design experiments aimed at understanding and manipulating whitefly gene expression. In this communication, for the first time we provide evidence that the RNA interference mechanism discovered in many organisms, including in Hemiptera, is active in B. tabaci. By injecting into the body cavity long double-stranded RNA (dsRNA) molecules, specifically directed against genes uniquely expressed in the midgut and salivary glands, we were able to significantly inhibit the expression of the targeted mRNA in the different organs. Gene expression levels in RNAi-silenced whiteflies were reduced up to 70% compared to whiteflies injected with buffer or with a green fluorescent protein (GFP)-specific dsRNA. Phenotypic effects were observed in B. tabaci ovaries following dsRNA targeting the whitefly Drosophila chickadee homologue. Disruption of whitefly gene expression opens the door to new strategies aimed at curbing down the deleterious effects of this insect pest to agriculture.     

Novel Genes Affecting the Interaction between the Cabbage Whitefly and Arabidopsis Uncovered by Genome-Wide Association Mapping

Plants have evolved a variety of ways to defend themselves against biotic attackers. This has resulted in the presence of substantial variation in defense mechanisms among plants, even within a species. Genome-wide association (GWA) mapping is a useful tool to study the genetic architecture of traits, but has so far only had limited exploitation in studies of plant defense. Here, we study the genetic architecture of defense against the phloem-feeding insect cabbage whitefly (Aleyrodes proletella) in Arabidopsis thaliana. We determined whitefly performance, i.e. the survival and reproduction of whitefly females, on 360 worldwide selected natural accessions and subsequently performed GWA mapping using 214,051 SNPs. Substantial variation for whitefly adult survival and oviposition rate (number of eggs laid per female per day) was observed between the accessions. We identified 39 candidate SNPs for either whitefly adult survival or oviposition rate, all with relatively small effects, underpinning the complex architecture of defense traits. Among the corresponding candidate genes, i.e. genes in linkage disequilibrium (LD) with candidate SNPs, none have previously been identified as a gene playing a role in the interaction between plants and phloem-feeding insects. Whitefly performance on knock-out mutants of a number of candidate genes was significantly affected, validating the potential of GWA mapping for novel gene discovery in plant-insect interactions. Our results show that GWA analysis is a very useful tool to gain insight into the genetic architecture of plant defense against herbivorous insects, i.e. we identified and validated several genes affecting whitefly performance that have not previously been related to plant defense against herbivorous insects.     

Tomato yellow leaf curl geminivirus (TYLCV-Is) is transmitted among whiteflies (Bemisia tabaci) in a sex-related manner

Tomato yellow leaf curl virus (TYLCV) is the name given to a complex of geminiviruses infecting tomato cultures worldwide. TYLCV is transmitted by a single insect species, the whitefly Bemisia tabaci. Herein we show that a TYLCV isolate from Israel (TYLCV-Is) can be transmitted among whiteflies in a sex-dependent manner, in the absence of any other source of virus. TYLCV was transmitted from viruliferous males to females and from viruliferous females to males but not among insects of the same sex. Transmission took place when insects were caged in groups or in couples, in a feeding chamber or on cotton plants, a TYLCV nonhost. The recipient insects were able to efficiently inoculate tomato test plants. Insect-to-insect virus transmission was instrumental in increasing the number of whiteflies capable of infecting tomato test plants in a whitefly population. TYLCV was present in the hemolymph of whiteflies caged with viruliferous insects of the other sex; therefore, the virus follows, at least in part, the circulative pathway associated with acquisition from infected plants. Taken as a whole, these results imply that a plant virus can be sexually transmitted from insect to insect.     

Pantothenate mediates the coordination of whitefly and symbiont fitness

Intracellular symbionts in insects often have reduced genomes. Host acquisition of genes from bacteria is an important adaptation that supports symbionts. However, the function of horizontally transferred genes in insect symbiosis remains largely unclear. The primary symbiont Portiera housed in bacteriocytes lacks pantothenate synthesis genes: panB and panC, which is presumably complemented by a fused gene panB-panC (hereafter panBC) horizontally transferred from bacteria in Bemisia tabaci MEAM1. We found panBC in many laboratory cultures, and species of B. tabaci shares a common evolutionary origin. We demonstrated that complementation with whitefly panBC rescued E. coli pantothenate gene knockout mutants. Portiera elimination decreased the pantothenate level and PanBC abundance in bacteriocytes, and reduced whitefly survival and fecundity. Silencing PanBC decreased the Portiera titer, reduced the pantothenate level, and decreased whitefly survival and fecundity. Supplementation with pantothenate restored the symbiont titer, PanBC level, and fitness of RNAi whiteflies. These data suggest that pantothenate synthesis requires cooperation and coordination of whitefly PanBC expression and Portiera. This host–symbiont co-regulation was mediated by the pantothenate level. Our findings demonstrated that pantothenate production, by the cooperation of a horizontally acquired, fused bacteria gene and Portiera, facilitates the coordination of whitefly and symbiont fitness. Thus, this study extends our understanding on the basis of complex host–symbiont interactions.Subject terms: Applied microbiology, Functional genomics     

Cardinium infection alters cotton defense and detoxification metabolism of its whitefly host

Field monitoring revealed that the infection ratio of the bacterial symbiont Cardinium in the whitefly (Bemisia tabaci MED) was relatively low in northern China. However, the role of this symbiont and the symbiont–whitefly–host plant interaction mechanism are poorly understood. We investigated the influence of Cardinium on the competitiveness of the host whitefly and the physiological interaction between the host plants and host whiteflies. Cardinium-infected whiteflies were displaced by uninfected whiteflies after 5 generations, which showed that Cardinium infection reduced whitefly competitiveness. The defense response genes of cotton significantly decreased under infestation by infected whiteflies compared to uninfected whiteflies. The expression of detoxification metabolism genes, especially the uridine 5ʹ-diphospho-glucuronyltransferase and P450 genes, in infected whiteflies significantly decreased. These results demonstrated that Cardinium could inhibit the defense response of the host plant and decrease the detoxification metabolism ability of the host whitefly. The reduced competitiveness of infected whiteflies may be associated with the inhibition of the whitefly detoxification metabolism by Cardinium, resulting in the reduced performance of infected whiteflies. However, Cardinium infection can suppress plant defenses, which may benefit both infected and uninfected whiteflies when they coexist. This research illustrates the symbiont–whitefly–host plant interaction mechanism and the population dynamics of the whitefly.     

A newly recorded Rickettsia of the Torix group is a recent intruder and an endosymbiont in the whitefly Bemisia tabaci

The bacterium Rickettsia is found widely in phytophagous insects and often exerts profound effects on the phenotype and fitness of its hosts. Here, we decrypt a new, independent, phylogenetically ancient Torix Rickettsia endosymbiont found constantly in a laboratory line of an economically important insect Asia II 7, a putative species of the Bemisia tabaci whitefly complex (Hemiptera: Aleyrodidae), and occasionally in field whitefly populations. This new Rickettsia distributes throughout the body of its whitefly host. Genetically, compared to Rickettsia_bellii_MEAM1 found earlier in whiteflies, the new Rickettsia species has more gene families and pathways, which may be important factors in shaping specific symbiotic relationships. We propose the name ‘Candidatus Rickettsia_Torix_Bemisia_tabaci (RiTBt)’ for this new endosymbiont associated with whiteflies. Comparative genomic analyses indicate that RiTBi may be a relatively recent intruder in whiteflies given its low abundance in the field and relatively larger genome compared to Rickettsia_bellii_MEAM1.