The phyllosphere microbiome of host trees contributes more than leaf phytochemicals to variation in the Agrilus planipennis Fairmaire gut microbiome structure.

The microbiome composition of living organisms is closely linked to essential functions determining the fitness of the host for thriving and adapting to a particular ecosystem. Although multiple factors, including the developmental stage, the diet, and host-microbe coevolution have been reported to drive compositional changes in the microbiome structures, very few attempts have been made to disentangle their various contributions in a global approach. Here, we focus on the emerald ash borer (EAB), an herbivorous pest and a real threat to North American ash tree species, to explore the responses of the adult EAB gut microbiome to ash leaf properties, and to identify potential predictors of EAB microbial variations. The relative contributions of specific host plant properties, namely bacterial and fungal communities on leaves, phytochemical composition, and the geographical coordinates of the sampling sites, to the EAB gut microbial community was examined by canonical analyses. The composition of the phyllosphere microbiome appeared to be a strong predictor of the microbial community structure in EAB guts, explaining 53 and 48% of the variation in fungi and bacteria, respectively. This study suggests a potential covariation of the microorganisms associated with food sources and the insect gut microbiome.

Natural Variation in Physicochemical Profiles and Bacterial Communities Associated with Aedes aegypti Breeding Sites and Larvae on Guadeloupe and French Guiana.

Aedes aegypti develop in aquatic habitats in which mosquito larvae are exposed to physicochemical elements and microorganisms that may influence their life cycle and their ability to transmit arboviruses. Little is known about the natural bacterial communities associated with A. aegypti or their relation to the biotic and abiotic characteristics of their aquatic habitats. We characterized the physicochemical properties and bacterial microbiota of A. aegypti breeding sites and larvae on Guadeloupe and in French Guiana. In addition, we explored whether geographic location, the type of breeding site and physicochemical parameters influenced the microbiota associated with this mosquito species. We used large-scale 16S rRNA gene sequencing of 160 breeding sites and 147 pools of A. aegypti larvae and recorded 12 physicochemical parameters at the sampled breeding sites. Ordination plots and multiple linear regression were used to assess the influence of environmental factors on the bacterial microbiota of water and larvae. We found territory-specific differences in physicochemical properties (dissolved oxygen, conductivity) and the composition of bacterial communities in A. aegypti breeding sites that influenced the relative abundance of several bacteria genera (e.g., Methylobacterium, Roseoccocus) on the corresponding larvae. A significant fraction of the bacterial communities identified on larvae, dominated by Herbiconiux and Microvirga genera, were consistently enriched in mosquitoes regardless the location. In conclusion, territory-specific differences observed in the biotic and abiotic properties of A. aegypti breeding sites raise concern about the impact of these changes on pathogen transmission by different A. aegypti populations.

Microbial Control of the Emerald Ash Borer (Coleoptera: Buprestidae) Using Beauveria bassiana (Hypocreales: Cordycipitaceae) by the Means of an Autodissemination Device.

The FraxiProtec, an autodissemination device loaded with the fungus Beauveria bassiana isolate CFL-A, was tested in the field to evaluate its potential to infect emerald ash borer adults and reduce their populations. During the 2-yr experimental period, the dispersion of B. bassiana-infected adults was also documented to assess the dissemination capacity of the biocontrol agent beyond the treated areas. The mean percentage of infected emerald ash borer in 2017 and 2018 in 15 treated sites was 43.3 ± 2.9% and 39.7 ± 3.9%, respectively, and no significant variation was observed over the tested years. Furthermore, a 40% significant reduction of the mean emerald ash borer population growth per tree in treated sites was recorded when compared to the control sites. Emerald ash borer infected with B. bassiana isolate CFL-A were collected from baited sticky traps in the sentinel stations in the area surrounding the FraxiProtec-treated sites. Even at a distance of 125 m, an average of 9.4 ± 2.6% infected emerald ash borer were collected. Finally, exploratory analyzes were carried out on parameters such as the area to be treated, ash tree density, and FraxiProtec density to document potential relationships, which could be used in the determination of a prescription.

Gut microbiome of the emerald ash borer, Agrilus planipennis Fairmaire, and its relationship with insect population density.

The gut microbial communities of beetles play crucial roles in their adaptive capacities. Environmental factors such as temperature or nutrition naturally affect the insect microbiome, but a shift in local conditions like the population density on a host tree could also lead to changes in the microbiota. The emerald ash borer (EAB), Agrilus planipennis Fairmaire, is an exotic wood borer that causes environmental and economic damage to ash trees in North America. This study aimed to describe the taxonomic structure of the EAB gut microbiome and explore its potential relationship with borer population size. The number of EAB adults collected per tree through a 75 km transect from an epicenter allowed the creation of distinct classes of population density. The Gammaproteobacteria and Ascomycota predominated in bacterial and fungal communities respectively, as determined by sequencing of the bacterial 16S rRNA gene and the fungal internal transcribed spacer ITS2. Species richness and diversity of the bacterial community showed significant dependence on population density. Moreover, α-diversity and ß-diversity analysis revealed some indicator amplicon sequence variants suggesting that the plasticity of the gut microbiome could be related to the EAB population density in host trees.

Biological H2 and CO oxidation activities are sensitive to compositional change of soil microbial communities.

Trace gas uptake by microorganisms controls the oxidative capacity of the troposphere, but little is known about how this important function is affected by changes in soil microbial diversity. This article bridges that knowledge gap by examining the response of the microbial community-level physiological profiles (CLPPs), carbon dioxide (CO2) production, and molecular hydrogen (H2) and carbon monoxide (CO) oxidation activities to manipulation of microbial diversity in soil microcosms. Microbial diversity was manipulated by mixing nonsterile and sterile soil with and without the addition of antibiotics. Nonsterile soil without antibiotics was used as a reference. Species composition changed significantly in soil microcosms as a result of dilution and antibiotic treatments, but there was no difference in species richness, according to PCR amplicon sequencing of the bacterial 16S rRNA gene. The CLPP was 15% higher in all dilution and antibiotic treatments than in reference microcosms, but the dilution treatment had no effect on CO2 production. Soil microcosms with dilution treatments had 58%-98% less H2 oxidation and 54%-99% lower CO oxidation, relative to reference microcosms, but did not differ among the antibiotic treatments. These results indicate that H2 and CO oxidation activities respond to compositional changes of microbial community in soil.

Horizontal Transmission of the Entomopathogenic Fungal Isolate INRS-242 of Beauveria bassiana (Hypocreales: Cordycipitaceae) in Emerald Ash Borer, Agrilus planipennis (Coleoptera: Buprestidae).

Emerald ash borer (EAB), Agrilus planipennis Fairmaire, is an invasive and destructive beetle that causes extensive damage to ash trees in North America. The entomopathogenic fungus Beauveria bassiana Vuillemin is considered as an effective biological control agent for EAB adult populations. Using an autodissemination device with a fungal isolate of B. bassiana, our research aims to investigate the possibility of horizontal transmission of the fungal disease from infected to uninfected EAB adults during mating. Results show that the efficiency of fungal transmission is significantly related to the sex of EAB carrying the fungal pathogen. EAB males are the promising vector to transmit mycosis to their partners during mating. Results strengthen the potential of the fungal autodissemination device as a powerful biological strategy to control EAB populations.

Phytoseiid predatory mites can disperse entomopathogenic fungi to prey patches.

Recent studies have shown that predatory mites used as biocontrol agents can be loaded with entomopathogenic fungal conidia to increase infection rates in pest populations. Under laboratory conditions, we determined the capacity of two phytoseiid mites, Amblyseius swirskii and Neoseiulus cucumeris to deliver the entomopathogenic fungus Beauveria bassiana to their prey, Frankliniella occidentalis. Predatory mites were loaded with conidia and released on plants that had been previously infested with first instar prey clustered on a bean leaf. We examined each plant section to characterize the spatial distribution of each interacting organism. Our results showed that A. swirskii delivered high numbers of conidia to thrips infested leaves, thereby increasing the proportion of thrips that came into contact with the fungus. The effect was larger when thrips infestation occurred on young leaves than on old leaves. Neoseiulus cucumeris delivered less conidia to the thrips infested leaves. These patterns result from differences in foraging activity between predatory mite species. Amblyseius swirskii stayed longer on plants, especially within thrips colonies, and had a stronger suppressing effect on thrips than N. cucumeris. Our study suggests that loading certain predatory mite species with fungal conidia can increase their capacity to suppress thrips populations by combining predation and dispersing pathogens.

Physiological traits and relative abundance of species as explanatory variables of co-occurrence pattern of cultivable bacteria associated with chia seeds.

Deciphering the rules defining microbial community assemblage is envisioned as a promising strategy to improve predictions of pathogens colonization and proliferation in food. Despite the increasing number of studies reporting microbial co-occurrence patterns, only a few attempts have been made to challenge them in experimental or theoretical frameworks. Here, we tested the hypothesis that observed variations in co-occurrence patterns can be explained by taxonomy, relative abundance, and physiological traits of microbial species. We used PCR amplicon sequencing of taxonomic markers to assess distribution and co-occurrence patterns of bacterial and fungal species found in 25 chia (Salvia hispanica L.) samples originating from eight different sources. The use of nutrient-rich and oligotrophic media enabled isolation of 71 strains encompassing 16 bacterial species, of which five corresponded to phylotypes represented in the molecular survey. Tolerance to different growth inhibitors and antibiotics was tested to assess the physiological traits of these isolates. Divergence of physiological traits and relative abundance of each pair of species explained 69% of the co-occurrence profile displayed by cultivable bacterial phylotypes in chia. Validation of this ecological network conceptualization approach to more food products is required to integrate microbial species co-occurrence patterns in predictive microbiology.

Towards the development of multifunctional molecular indicators combining soil biogeochemical and microbiological variables to predict the ecological integrity of silvicultural practices.

The impact of mechanical site preparation (MSP) on soil biogeochemical structure in young larch plantations was investigated. Soil samples were collected in replicated plots comprising simple trenching, double trenching, mounding and inverting site preparation. Unlogged natural mixed forest areas were used as a reference. Analysis of soil nutrients, abundance of bacteria and gas exchanges unveiled no significant difference among the plots. However, inverting site preparation resulted in higher variations of gas exchanges when compared with trenching, mounding and unlogged natural forest. A combination of the biological and physicochemical variables was used to define a multifunctional classification of the soil samples into four distinct groups categorized as a function of their deviation from baseline ecological conditions. According to this classification model, simple trenching was the approach that represented the lowest ecological risk potential at the microsite level. No relationship was observed between MSP method and soil bacterial community structure as assessed by high-throughput sequencing of bacterial 16S rRNA gene; however, indicator genotypes were identified for each multifunctional soil class. This is the first identification of multifunctional molecular indicators for baseline and disturbed ecological conditions in soil, demonstrating the potential of applied microbial ecology to guide silvicultural practices and ecological risk assessment.

Surveying the endomicrobiome and ectomicrobiome of bark beetles: The case of Dendroctonus simplex.

Many bark beetles belonging to the Dendroctonus genus carry bacterial and fungal microbiota, forming a symbiotic complex that helps the insect to colonize the subcortical environment of the host tree. However, the biodiversity of those bacteria at the surface of the cuticle or inside the body parts of bark beetles is not well established. The aim of this study was to characterize the bacterial microbiome associated with the eastern larch beetle, Dendroctonus simplex, using bacterial 16S rRNA gene pyrosequencing. The ecto- and endomicrobiome and the subcortical galleries were investigated. Several bacterial genera were identified, among which Pseudomonas, Serratia and Yersinia are associated with the surface of the beetle cuticle, and genera belonging to Enterobacteriaceae and Gammaproteobacteria with the interior of the insect body. The index of dissimilarity indicates that the bacterial microbiome associated with each environment constitutes exclusive groups. These results suggest the presence of distinct bacterial microbiota on the surface of the cuticle and the interior of D. simplex body. Additionally, the bacterial diversity identified in the galleries is substantially different from the ectomicrobiome, which could indicate a selection by the insect. This study reports for the first time the identification of the eastern larch beetle microbiome.

Effects of Pheromone Release Rate and Trap Placement on Trapping of Agrilus planipennis (Coleoptera: Buprestidae) in Canada.

The emerald ash borer, Agrilus planipennis Fairmaire (Coleoptera: Buprestidae), is a devastating insect pest in its introduced range. A trapping system that increases trap catches or detection rates in low-density populations would be beneficial for survey programs. Five trapping experiments were conducted to investigate factors influencing capture rates of male beetles on dark green traps baited with the A. planipennis pheromone, (3Z)-dodecen-12-olide ((3Z)-lactone), plus the green leaf volatile, (3Z)-hexenol. Low doses (0.001-1.0 mg) of (3Z)-lactone + (3Z)-hexenol did not consistently increase captures of male A. planipennis. In other experiments, mean captures of males were significantly higher on traps baited with a moderate dose (3.0 mg/septum) of (3Z)-lactone + (3Z)-hexenol, compared with lower doses (0.001, 0.1, and 1.0 mg) or (3Z)-hexenol alone. Next, we demonstrated that addition of (3Z)-lactone to traps baited with (3Z)-hexenol resulted in significantly greater increases in male captures when pairs of traps were placed on the same tree, than when traps were placed on adjacent trees. Moreover, significantly more A. planipennis were captured on pheromone-baited traps placed in the southern versus northern aspect of the crown. These results highlight the importance of experimental set-up for elucidating lure treatment effects and also suggests the (3Z)-lactone may be more active at close range. Our findings increase our understanding of the pheromone ecology of this species and lend support toward the use of dark green traps baited with 3.0 mg (3Z)-lactone + (3Z)-hexenol deployed in the south aspect of the canopy for detection programs for this insect.

Land-use influences the distribution and activity of high affinity CO-oxidizing bacteria associated to type I-coxL genotype in soil.

Soil carboxydovore bacteria are the biological sink of atmospheric carbon monoxide (CO). The initial oxidation of CO is catalyzed by a CO-dehydrogenase (CODH), and the gene coxL encodes the large subunit of the enzyme. Only a few carboxydovore isolates were shown to oxidize atmospheric CO and little is known about the potential impact of global change on the ecophysiology of this functional group. The main objective of this study was to assess the impact of land-use and soil properties on coxL gene diversity and identify molecular indicators for the soil uptake of atmospheric CO. Soil samples were collected in three neighboring sites encompassing different land-use types, namely deciduous forest, larch plantation and maize field. CO uptake activity was related to total carbon and nitrogen content in soil, with the highest activity observed in deciduous forest. An extensive coxL database was assembled to optimize a PCR detection assay targeting sequences belonging to functional type I-CODH and hypothetical type II-CODH. Fully replicated coxL gene libraries unveiled a unique molecular signature in deciduous forest soil, with enrichment of type I sequences. Genetic profiles of larch and maize monocultures were not statistically different and showed higher level of coxL gene richness than deciduous forest. Soil water content and CO uptake activity explained 38% of the variation of coxL gene profiles in a canonical ordination analysis, leading to the identification of sequences belonging to the δ-Proteobacteria cluster as indicator for high affinity CO uptake activity. Enrichment of type I and δ-Proteobacteria coxL sequences in deciduous forest were confirmed by qPCR in an independent soil survey. CO uptake activity in model carboxydovore bacteria suggested that a significant fraction of detected putative high affinity CO oxidizers were active in soil. Land-use was a driving force separating coxL diversity in deciduous forest from monocultures.

Occurrence of species of the genus Pityophthorus Eichhoff (Coleoptera, Curculionidae, Scolytinae) in the province of Quebec, Canada.

Twig beetles in the genus Pityophthorus Eichhoff, 1864 include more than 300 species worldwide, with maximum diversity in tropical and subtropical regions. To date, approximately 50 species of Pityophthorus have been recorded in Canada, and these species are associated mainly with coniferous trees. Since 1981, no comprehensive study on this difficult taxonomic group has been conducted in Quebec, Canada, most likely due to their limited significance as forest pests. Based on data gathered from five years of field sampling in conifer seed orchards and compiled from various entomological collections, the distribution of Pityophthorus species in Quebec is presented. Approximately 291 new localities were recorded for the Pityophthorus species. Five species-group taxa, namely Pityophthorus puberulus (LeConte, 1868), Pityophthorus pulchellus pulchellus Eichhoff, 1869, Pityophthorus pulicarius (Zimmermann, 1868), Pityophthorus nitidus Swaine, 1917,and Pityophthorus cariniceps LeConte&Horn, 1876 were the most widespread. In contrast, Pityophthorus consimilis LeConte, 1878, Pityophthorus intextus Swaine, 1917, Pityophthorus dentifrons Blackman, 1922, Pityophthorus ramiperda Swaine, 1917, and Pityophthorus concavus Blackman, 1928 display a notably limited distribution. In addition, the first distribution records of Pityophthorus intextus and Pityophthorus biovalis Blackman, 1922 are furnished, and the subspecies Pityophthorus murrayanae murrayanae Blackman, 1922is reported from Quebec for the second time. Moreover, distribution maps are provided for all Pityophthorus species recorded in the province of Quebec.

The complex symbiotic relationships of bark beetles with microorganisms: a potential practical approach for biological control in forestry.

Bark beetles, especially Dendroctonus species, are considered to be serious pests of the coniferous forests in North America. Bark beetle forest pests undergo population eruptions, causing region wide economic losses. In order to save forests, finding new and innovative environmentally friendly approaches in wood-boring insect pest management is more important than ever. Several biological control methods have been attempted over time to limit the damage and spreading of bark beetle epidemics. The use of entomopathogenic microorganisms against bark beetle populations is an attractive alternative tool for many biological control programmes in forestry. However, the effectiveness of these biological control agents is strongly affected by environmental factors, as well as by the susceptibility of the insect host. Bark beetle susceptibility to entomopathogens varies greatly between species. According to recent literature, bark beetles are engaged in symbiotic relationships with fungi and bacteria. These types of relationship are very complex and apparently involved in bark beetle defensive mechanisms against pathogens. The latest scientific discoveries in multipartite symbiosis have unravelled unexpected opportunities in bark beetle pest management, which are discussed in this article.

Towards the development of an autocontamination trap system to manage populations of emerald ash borer (Coleoptera: Buprestidae) with the native entomopathogenic fungus, Beauveria bassiana.

Emerald ash borer, Agrilus planipennis Fairmaire (Coleoptera: Buprestidae) is an invasive species from Asia that was discovered in North America Canada, in 2002. Herein, we describe studies to develop an autocontamination trapping system to disseminate Beauveria bassiana to control beetle populations. The standard trap for emerald ash borer in Canada is a light green prism trap covered in an insect adhesive and baited with (Z)-3-hexenol. We compared of green multifunnel traps, green intercept panel traps (both with and without fluon coating) and green prism traps for capturing emerald ash borer in a green ash plantation. The coated green multifunnel traps captured significantly more males and more females than any other trap design. We examined the efficacy of two native B. bassiana isolates, INRS-CFL and L49-1AA. In a field experiment the INRS-CFL isolate attached to multifunnel traps in autocontamination chambers retained its pathogenicity to emerald ash borer adults for up to 43 d of outdoor exposure. Conidia germination of the INRS-CFL isolate was >69% after outdoor exposure in the traps for up to 57 d. The L49-1AA isolate was not pathogenic in simulated trap exposures and the germination rate was extremely low (<5.3%). Mean (+/- SEM) conidia loads on ash borer adults after being autocontaminated in the laboratory using pouches that had been exposed in traps out of doors for 29 d were 579,200 (+/- 86,181) and 2,400 (+/- 681) for the INRS-CFL and the LA9-1AA isolates, respectively. We also examined the fungal dissemination process under field conditions using the L49-1AA isolate in a green ash plantation. Beetles were lured to baited green multifunnel traps with attached autocontamination chambers. Beetles acquired fungal conidia from cultures growing on pouches in the chambers and were recaptured on Pestick-coated traps. In total, 2,532 beetles were captured of which 165 (6.5%) had fungal growth that resembled B. bassiana. Of these 25 beetles were positive for the L49-1AA isolate.

Choristoneura fumiferana Granulovirus pk-1: a baculoviral protein kinase.

Open reading frame (ORF) 3 on the Choristoneura fumiferana granulovirus (ChfuGV), located in the 11 kb fragment of the BamHI genomic bank encodes a predicted 32-kDa putative kinase protein. Bioinformatics analysis on the predicted amino acid sequence of ChfuGV PK-1 revealed the existence of 11 catalytic subdomains. Sequence analysis within the 5'-untranslated region (5'-UTR) of ChfuGV pk- 1 indicates the presence of both putative early and late promoter motifs, indicating that pk-1 may be expressed throughout the infection cycle. Promoter sequence analysis reveals that pk-1 is deprived of a TATA box and appears instead to be regulated by other cis-acting transcriptional regulatory elements. Temporal transcription analysis by RT-PCR confirms the appearance of transcripts detected from 2 h p.i. until 72 h p.i. Northern blot hybridization characterizes pk-1 transcription as a 1.2 kb transcript. Homology comparisons reveal that ChfuGV PK-1 protein is most closely related to Phthorimaea operculalla granulovirus (PoGV) with 80 % amino acid identity.

Transcription, translation, and immunolocalization of ODVP-6E/ODV-E56 and p74 proteins: two highly conserved ODV-associated envelope proteins of Choristoneura fumiferana Granulovirus.

Choristoneura fumiferana granulovirus (ChfuGV) infection results two types of enveloped virions: Occlusion-derived virus (ODV) and budded virus (BV). Structural proteins ODVP-6E/ODV-E56 and p74 are two major conserved ODV-associated proteins that may be involved in the initiation of viral infection cycle in susceptible host insect larvae. This study presents the characterization of ChfuGV odvp-6e/odv-e56 and p74 transcription and translation as well as immunolocalization of these proteins in the occluded ChfuGV virion. Our results revealed that the transcription of odvp-6e/odv-e56 and p74 genes, both, start at 24 hours post infection (h p.i.). Using monospecific polyclonal antibodies made against ODVP-6E/ODV-E56 and p74 we demonstrated that these proteins are both expressed late in infection (24 h p.i.). Immunogold labeling using antisera against ODVP-6E/ODV-E56 and p74 proteins demonstrated that ODVP-6E/ODV-E56 and p74 proteins are both associated with the ODV envelop of ChfuGV.

Identification, characterization and phylogenic analysis of conserved genes within the p74 gene region of Choristoneura fumiferana granulovirus genome.

The genes located within the p74 gene region of the Choristoneura fumiferana granulovirus (ChfuGV) were identified by sequencing an 8.9 kb BamHI restriction fragment on the ChfuGV genome. The global guanine-cytosine (GC) content of this region of the genome was 33.02%. This paper presents the ORFs within the p74 gene region along with their transcriptional orientations. This region contains a total of 15 open reading frames (ORFs). Among those, 8 ORFs were found to be homologues to the baculoviral ORFs: Cf-i-p , Cf-vi, Cf-vii, Cf-viii (ubiquitin), Cf-xi (pp31), Cf-xii (lef-11), Cf-xiii (sod) and Cf-xv-p (p74). To date, no specific function has been assigned to the ORFs: Cf-i, Cf-ii, Cf-iii, Cf-iv, Cf-v, Cf-vi, Cf-vii, Cf-ix and Cf-x. The most noticeable ORFs located in this region of the ChfuGV genome were ubiquitin, lef-11, sod, fibrillin and p74. The phylogenetic trees (constructed using conceptual products of major conserved ORFs) and gene arrangement in this region were used to further examine the classification of the members of the granulovirus genus. Comparative studies demonstrated that ChfuGV along with the Cydia pomonella granulovirus (CpGV), Phthorimaea operculella granulovirus (PhopGV), Adoxophyes orana granulovirus (AoGV) and Cryptophlebia leucotreta granulovirus (ClGV) share a high degree of amino acids sequence and gene arrangement preservation within the studied region. These results support a previous report, which classified a granuloviruses into 2 distinct groups: Group I: ChfuGV, CpGV, PhopGV and AoGV and Group II: Xestia c-nigrum granulovirus (XcGV) and Plutella xylostella granulovirus (PxGV). The phylogenetic and gene arrangement studies also placed ClGV as a novel member of the Group I granuloviruses.

Identification, characterization and phylogenic analysis of conserved genes within the odvp-6e/odv-e56 gene region of Choristoneura fumiferana granulovirus.

The genes that are located within the odvp-6e/odv-e56 region of the Choristoneura fumiferana granulovirus (ChfuGV) were identified by sequencing the 11 kb BamHI restriction fragment on the ChfuGV genome. The global GC content that was calculated from the data obtained from this genomic region was 34.96%. The open-reading frames (ORFs), located within the odvp-6e/odv-e56 region, are presented and compared to the equivalent ORFs that are located at the same region in other GVs. This region is composed of 14 ORFs, including three ORFs that are unique to ChfuGV with no obvious homologues in other baculoviruses as well as eleven ORFs with homologues to granuloviral ORFs, such as granulin, CfORF2, pk-1, ie-1, odv-e18, p49, and odvp-6e/odv-e56. In this study, the conceptual products of seven major conserved ORFs (granulin, CfORF2, IE-1, ODV-E18, p49 and ODVP-6E/ODV-E56) were used in order to construct phylogenetic trees. Our results show that granuloviruses can be grouped in 2 distinct groups as follows: Group I; Choristoneura fumiferana granulovirus (ChfuGV), Cydia pomonella granulovirus (CpGV), Phthorimaea operculella granulovirus (PhopGV), and Adoxophyes orana granulovirus (AoGV). Group II; Xestia c-nigrum granulovirus (XcGV), Plutella xylostella granulovirus (PxGV), and Trichoplusia ni granulovirus (TnGV). The ChfuGV conserved proteins are most closely related to those of CpGV, PhopGV, and AoGV. Comparative studies, performed on gene arrangements within this region of genomes, demonstrated that three GVs from group I maintain similar gene arrangements.

Choristoneura fumiferana Granulovirus p74 protein, a highly conserved baculoviral envelope protein.

A gene that encodes a homologue to baculoviral p74, an envelope-associated viral structural protein, has been identified and sequenced on the genome of Choristoneura fumiferana granulovirus (ChfuGV). A part of the ChfuGV p74 gene was located on an 8.9 kb BamHI subgenomic fragment using different sets of degenerated primers. These were designed using the results of the protein sequencing of a major 74 kDa structural protein that is associated with the occlusion-derived virus (ODV). The gene has a 1992 nucleotide (nt) open-reading frame (ORF) that encodes a protein with 663 amino acids with a predicted molecular mass of 74,812 Da. Comparative studies revealed the presence of two major conserved regions in the ChfuGV p74 protein. This study also shows that all of the p74 proteins contain two putative transmembrane domains at their C-terminal segments. At the nucleotide sequence level, two late promoter motifs (TAAG and GTAAG) were located upstream of the first ATG of the p74 gene. The gene contained a canonical poly(A) signal, AATAAA, at its 3 non-translated region. A phylogenetic tree for baculoviral p74 was constructed using a maximum parsimony analysis. The phylogenetic estimation demonstrated that ChfuGV p74 is related the closest to those of Cydia pomonella granulovirus (CpGV) and Phthorimaea operculella granulovirus (PhopGV).