Granulovirus Thesis Statement

Tips and Examples for Writing Thesis Statements

Summary:

This resource provides tips for creating a thesis statement and examples of different types of thesis statements.

Contributors: Elyssa Tardiff, Allen Brizee
Last Edited: 2018-01-24 02:29:37

Tips for Writing Your Thesis Statement

1. Determine what kind of paper you are writing:

  • An analytical paper breaks down an issue or an idea into its component parts, evaluates the issue or idea, and presents this breakdown and evaluation to the audience.
  • An expository (explanatory) paper explains something to the audience.
  • An argumentative paper makes a claim about a topic and justifies this claim with specific evidence. The claim could be an opinion, a policy proposal, an evaluation, a cause-and-effect statement, or an interpretation. The goal of the argumentative paper is to convince the audience that the claim is true based on the evidence provided.

If you are writing a text that does not fall under these three categories (e.g., a narrative), a thesis statement somewhere in the first paragraph could still be helpful to your reader.

2. Your thesis statement should be specific—it should cover only what you will discuss in your paper and should be supported with specific evidence.

3. The thesis statement usually appears at the end of the first paragraph of a paper.

4. Your topic may change as you write, so you may need to revise your thesis statement to reflect exactly what you have discussed in the paper.

Thesis Statement Examples

Example of an analytical thesis statement:

An analysis of the college admission process reveals one challenge facing counselors: accepting students with high test scores or students with strong extracurricular backgrounds.

The paper that follows should:

  • Explain the analysis of the college admission process
  • Explain the challenge facing admissions counselors

Example of an expository (explanatory) thesis statement:

The life of the typical college student is characterized by time spent studying, attending class, and socializing with peers.

The paper that follows should:

  • Explain how students spend their time studying, attending class, and socializing with peers

Example of an argumentative thesis statement:

High school graduates should be required to take a year off to pursue community service projects before entering college in order to increase their maturity and global awareness.

The paper that follows should:

  • Present an argument and give evidence to support the claim that students should pursue community projects before entering college

Genome Features

The assembled genome of LoobMNPV (GenBank accession number: KP763670) is 120,023-bp long with a mean coverage of 20.5 times. We have found 134 Open Reading Frames (ORFs) coding for putative proteins with at least 50 amino acid residues (Fig. 1a and Table S1). Out of these, 120 ORFs were found in other baculoviruses and among those, three ORFs (loob078, loob100 (he65) and loob113) showed higher identity with orthologs from betabaculoviruses (Table S1), confirming that horizontal transference between alphabaculovirus and betabaculovirus is indeed a common event, as previously documented11. Moreover, some unique ORFs were located within the hr3 (loob038) and hr4 (loob060 and loob061). All the 37 baculovirus core genes were present, however some genes present in almost all Alpha- and Betabaculovirus12 genomes available were not present: Ac106/107, Ac108 and gp37.

LoobMNPV homologous regions (hrs)

Seven homologous regions (hrs) were found in the LoobMNPV genome, in agreement to other baculoviruses13. These DNA palindromic repetitive elements are interspersed throughout most baculoviral genomes and are related to DNA replication14, gene transcription15, and possibly homologous recombination16. The hr1 has 684 bp, hr2 has 541 bp, hr3 has 1,037 bp, hr4 has 1,426 bp, hr5 has 679 bp, hr6 has 361 bp and hr7 has 836 bp, and their size and position were confirmed by PCR (data not shown). As shown in Fig. 2, all hrs have various copies of a common imperfect palindromic repeat of 38 bp (hr1a, hr2a, hr3a, hr3b, hr4a, hr5a, hr5b, hr6a, hr6b and hr7a).

LoobMNPV Phylogeny

We have performed a phylogenetic analysis (Fig. 3) based on the concatenated amino acid sequence alignment of 37 baculovirus core genes of 72 baculovirus species available in GenBank (Table S2). LoobMNPV clustered with group I alphabaculovirus. Its closest relative was found to be DekiNPV and both are basally related to AcMNPV-like viruses. This result disagrees with previous analysis based on the polh gene, in which LoobMNPV clustered together with both species Amsacta albistriga nucleopolyhedrovirus (AaNPV) and Thysanoplusia orichalcea nucleopolyhedrovirus (ThorNPV)6. However, the phylogenetic signal of highly conserved genes, such as polh, is usually inaccurate17, while phylogenetic inferences based on a large set of genes is more accurate and robust18.

LoobMNPV Comparative Genomics

Based on the phylogenetic results we have selected some of the closest relatives of LoobMNPV, including AcMNPV, Antheraea pernyi nucleopolyhedrovirus (AnpeNPV), Maruca vitrata nucleopolyhedrovirus (MaviNPV), Dendrolimus kikuchii nucleopolyhedrovirus (DekiNPV) and ThorNPV for comparative genomics. Figure 1b and Table S1 reveal all the LoobMNPV ORFs compared to orthologs from these genomes by the level of similarity (in terms of percentage of identity and E-values < 10−3). Overall, some ORFs are more conserved (high similarity), such as the genes that correspond to polh (loob001), p74 (loob010), p49 (loob014), pif-1 (loob041), vlf-1 (loob067), lef-9 (loob080), lef-8 (loob093) and pif-2 (loob123); while others are more variable, presenting a lower level of similarity, but still present in almost all baculovirus genomes, such as ORF1629 (loob002), ie-2 (loob005), vp80 (loob043), desmoplakin (loob075), f protein (loob122), arif-1 (loob124) and bv/odv-e26 (loob128).

Moreover, we have investigated the genome synteny among LoobMNPV and those selected alphabaculovirus genomes (Fig. 4). A circular ideogram layout shown in Fig. 4 displays lines connecting related genes, where it is possible to observe the relative position compared to LoobMNPV genome. Interestingly, we have observed that LoobMNPV and DekiNPV show high collinearity, and both present genome inversions and rearrangement in contrast to the genomes of AcMNPV, AnpeNPV, MaviNPV, and ThorNPV. The inverted regions were adjacent or flanked by the hr1 and hr2 (Fig. S1). These findings support the phylogenetic results presented in Fig. 3 and suggests that the inversions were originated in a common ancestor of both LoobMNPV and DekiNPV. These inversions are an autapomorphy when compared to the AcMNPV-like viruses. Previous studies have shown that rearrangements in the baculovirus genomes reflect evolutionary history, with more closely related viruses presenting higher genome collinearity19.

Furthermore, when analyzing the region where all genomes overlap with LoobMNPV (Fig. 4b), we have observed that unique ORFs in LoobMNPV (gaps with no correspondence to the other genomes) are interestingly found near hrs. According to previous reports, gene rearrangements and acquisitions are of common occurrence on proximities of hrs20, confirming the possibility of gene transfers to viruses by homologous recombination21, which could be also facilitated by factors such as the prevalence of various pathogens infecting the same host22, as well as concomitant infections in field populations23.

LoobMNPV unique ORFs

LoobMNPV genome showed 12 genes that do not have any match among baculoviruses. These ORFs are loob004, loob006, loob012, loob035, loob038, loob055, loob059, loob060, loob061, loob071, loob084, and loob097. There are three possible mechanisms for gene acquisition: extensive sequence divergence, which could push homolog genes below the similarity threshold; gene recombination, which produces novel protein products; and horizontal gene transfer (HGT). This third possibility is expected to be detectable by gene similarity from phylogenetically distinct species24. For each of these LoobMNPV unique ORFs, we have searched for baculovirus promoter motifs within 200 bp upstream of the start codon. The late promoter motif TAAG, that appears to be necessary for late transcription by the viral RNA polymerase25 was found in loob004, loob012, loob030, loob038, loob055, and loob071. However, further experiments are necessary to confirm whether these ORFs encode bona fide proteins. Moreover, these novel ORFs were searched for known domains and eight of theses ORFs (loob004, loob006, loob012, loob038, loob055, loob061, loob071, and loob097) did not match any predicted domain. The remaining will be discussed below.

Insect immune system-associated domain

The loob060 has an immunoglobulin-like domain, which has been found in some insect proteins, such as the hemolin, an hemolymph component that plays a role in bacterial surface binding, forming a protein complex that initiates the immune response26. Hemolin has also been found in the transcripts of L. obliqua bristles2. Several immunomodulators encoded by viruses have been described27 and may be involved in regulating the immune system and protecting virus-infected cells from the attack of other cells from the immune system28,29. For viruses, the expression of these proteins may indicate beneficial susceptibility in multiple pathogen infections, by protecting the host against opportunistic pathogens, reducing competition and benefiting viral propagation29. For instance, other saturniid-related alphabaculovirus species HespNPV expresses a functional insect-related serine protease inhibitor (serpin) in its genome that is likely related to host immunity modulation and virulence8,28.

Transcription factor-related domain (loob035)

One unique ORF demonstrated high correspondence to the eukaryotic transcription terminator factor type 2 (TTF2) from the butterfly Danaus plexippus (GenBank: EHJ68439), with 44% pairwise identity and E-value equals to 3e10−11. However, when filtering this result, in order to focus only on the family Baculoviridae, the referred gene presented higher similarity to the Global Transactivator (GTA) gene from the AnpeNPV (YP_611073), with 66% of identity and an E-value of 1e10−6.

Transcription Factors (TFs) in general are fundamental in a broad array of any cellular processes due to its ability of causing changes in downstream gene expression patterns30. GTA genes are observed in members from the group I alphabaculovirus. According to a previous study31, baculovirus GTAs play an important role in transcriptional activation of viral genes and were probably originated by HGT from the host to the common ancestor of the clade that includes AcMNPV, Bombyx mori nucleopolyhedrovirus (BmNPV), Orgyia pseudotsugata multicapsid nucleopolyhedrovirus (OpMNPV), and Epiphyas postvittana nucleopolyhedrovirus (EppoNPV). Katsuma, et al.32 found that a GTA homolog from BmNPV acts as a viral virulence factor in insect larvae, and may be required for activation of host and/or viral genes, increasing the speed of host killing. Based on domain analysis, we have found that the loob035, TTF2, and GTA genes are members of the SNF2 family. This family of genes encodes proteins with sequence motifs similar to those found in many DNA and RNA helicase protein families, and also proteins from a variety of species with roles in cellular processes such as transcriptional regulation, DNA recombination, chromatin unwinding and various other types of DNA repair33. In this regard, the possible acquisition of loob035 might be involved in the inhibition of the host transcriptional machinery in order to benefit viral expression.

To analyze whether loob035 has been independently acquired from the host insect through HGT, or is a divergent baculovirus GTA gene, a phylogenetic analysis has been performeddone, based on an amino acid alignment containing both TTF2 and GTA sequences. As shown in Fig. 5, loob035 presented a long branch size, which indicates great divergence from the other sequences, possibly due to positive selection34. Actually, TFs are overrepresented among genes predicted to be positively selected in previous genome-wide selection studies35. To confirm whether loob035 is in fact a new acquisition from insect host or a divergent baculovirus GTA gene, we compared the likelihood of a tree constraining loob035 to the GTA alphabaculovirus group (lnL = −14940.61) with the likelihood of a tree constraining loob035 into TTF2 group (lnL = −1455.06). We argue that the likelihood differences in combination with high non-parametric bootstrap values and high posterior probabilities constitute considerable evidence that loob035 clusters with the group of TTF2 genes, corroborating to the possibility of a novel HGT.

We further confirmed this independent acquisition hypothesis by analyzing the genomic context of baculovirus GTA genes. As shown in Fig. 6a, GTA genes are encountered in all group I alphabaculoviruses:AcMNPV, Anticarsia gemmatalis nucleopolyhedrovirus (AgMNPV), AnpeNPV, BmNPV, Bombyx mandarina nucleopolyhedrovirus (BomaNPV), Choristoneura fumiferana multicapsid nucleopolyhedrovirus (CfMNPV), Choristoneura occidentalis nucleopolyhedrovirus (ChocNPV), Choristoneura murinana nucleopolyhedrovirus (ChmuNPV), Choristoneura nucleopolyhedrovirus roaceana (ChroNPV), EppoNPV, Hyphantria cunea nucleopolyhedrovirus (HycuMNPV), Orgyia pseudotsugata multicapsid nucleopolyhedrovirus (OpMNPV), Philosamia cynthia nucleopolyhedrovirus (PhcyNPV), Plutella xylostella multiple nucleopolyhedrovirus (PlxyMNPV), Rachiplusia ou multicapsid nucleopolyhedrovirus (RoMNPV) and ThorNPV; except in MaviMNPV, LoobMNPV and DekiNPV, within a conserved position between lef-12 and odv-e66. However, in LoobMNPV, the gene located in this position is loob102, that corresponds to an AcMNPV-like gene (ac044). On the other hand, loob035 is inserted in a completely different genome context, located between both the ac110- and the ac111-like genes (Fig. 6b), confirming that loob035 has probably a different origin not related to the GTA gene. Besides, loob035 homologs found in DekiNPV (Orf 138) and in ThorNPV (Orf 117), according to Table S1, are also inserted in a different context (Fig. 6b).

Notably, loob035 diverges greatly from all the other compared sequences (Fig. S2), showing that these sequences present similarity only because they all contain the SNF2 conserved domain. Hughes & Friedman31 found that SNF2 baculovirus gene family has homologs in cellular organisms, and clustered closer to homologs in insects (Drosophila), according to the reconstruction of the evolutionary relationship among genes that were potentially acquired through HGT in comparison to baculovirus phylogeny.

The absence of cathepsin and chitinase genes in LoobMNPV

Interestingly, LoobMNPV does not encode two common baculovirus genes that are responsible for the post mortem host melanization and liquefaction benefiting virus dissemination: the enzymes cathepsin (v-cath) and chitinase (chiA) genes36. Besides the involvement of these genes in the horizontal spread of the virus in the field37, it has been reported that the chiA gene and the pro-form of v-cath interact directly and are dependent on each other for the promotion of host liquefaction, and therefore, they are usually acquired or lost together38, since they are adjacent genes on the genome. Among alphabaculovirus from group I, only LoobMNPV, AgMNPV and PhcyNPV lack v-cath and chiA genes. The recombinant introduction of v-cath and chiA genes from Choristoneura fumiferana DEF multiple nucleopolyhedrovirus (CfDEFNPV) into AgMNPV genome improves production of occlusion bodies and insecticidal activity during A. gemmatalis larvae infection39.

Several hosts from the family Saturniidae40, as well as from Noctuidae, present gregarious behavior41, which may facilitate virus dispersion to new susceptible individuals and reduce selective pressure for the maintenance of v-cath and chiA genes, as observed for LoobMNPV, AgMNPV, and PhcyNPV. However, HespNPV and AnpeNPV also infect gregarious hosts from Saturniidae family, but have v-cath and chiA genes in their genomes. Therefore, it remains to be determined whether the host behavior it is indeed a selective factor for cath and chiA genes maintenance.

Conclusions

In this work, we described the first complete genome sequence of a baculovirus isolated from a species of medical interest. LoobMNPV is located on a basal position of group I alphabaculovirus and presents inversions in large proportions when compared to the other related genomes. During evolution, fluxes in the genomes content, such as genes acquisitions and losses, pressured by positive selection, could possibly implicate in shifts on the evolutionary dynamics, by the occurrence of events of adaptation. Therefore, the elucidation of novel genomes will help the studies on baculovirus evolution, proportioning deeper knowledge and understanding of baculovirus as a whole group.

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