An Ascovirus Utilizes Different Types of Host Larval Regulated Cell Death Mechanisms To Produce and Release Vesicles.

Ascoviruses are insect-specific viruses that are thought to utilize the cellular apoptotic processes of host larvae to produce numerous virion-containing vesicles. In this study, we monitored the in vivo infection processes of Heliothis virescens ascovirus 3h (HvAV-3h) to illustrate the regulated cell death (RCD) of host cells. Transmission electron microscopic observations did not reveal any morphological markers of apoptosis in the fat bodies or hemocytes of HvAV-3h-infected Helicoverpa armigera or Spodoptera exigua larvae. However, several hemocytes showed the morphological criteria for necrosis and/or pyroptosis. Further in vitro biochemical tests were performed to confirm the RCD type of host cells after infection with HvAV-3h. Different morphological characteristics were found between the early (prior to 24 hours post-infection, [hpi]) and later (48 to 120 hpi) stages in both HvAV-3h infected larval fat bodies and hemocytes. In the early stages, the virions could only be found in several adipohemocytes, and the fat bodies were cleaving their contained lipid inclusions into small lipid dots. In the later stage, both fat bodies and hemocytes were filled with numerous virions. According to the morphological characteristics of HvAV-3h infected larval fat bodies or hemocytes, the pathogenic characteristics and infection patterns of HvAV-3h in the host larvae were described, and the systematic pathogenic mode of ascovirus infection was refined in this study. This study details the complete infection process of ascoviruses, which provides insights into the relationship between a pathogenesis of an insect virus and the RCD of different host tissues at different stages of infection. IMPORTANCE Viruses and other pathogens can interrupt host cellular apoptosis to gain benefits, such as sufficient resources and a stable environment that enables them to complete their replication and assembly. It is unusual for viruses to code proteins with homology to caspases, which are commonly recognized as apoptosis regulators. Ascoviruses are insect viruses with special cytopathology, and they have been hypothesized to induce apoptosis in their host larvae via coding a caspase-like protein. This enables them to utilize the process of cellular apoptosis to facilitate vesicle formation and replication. However, our previous studies revealed different trends. The fat bodies and hemocytes of Heliothis virescens ascovirus 3h (HvAV-3h)-infected larvae did not show any morphological markers of apoptosis but did display necrosis and/or pyroptosis morphological characteristics. The pathogenic characteristics and infection patterns of HvAV-3h in the host larvae were described, which can help us understand the relationship between the pathogenesis of an insect virus and host RCD.

Phylogenomic data exploration with increased sampling provides new insights into the higher-level relationships of butterflies and moths (Lepidoptera).

A robust and stable phylogenetic framework is a fundamental goal of evolutionary biology. As the third largest insect order in the world following Coleoptera and Diptera, Lepidoptera (butterflies and moths) play a central role in almost every terrestrial ecosystem as indicators of environmental change and serve as important models for biologists exploring questions related to ecology and evolutionary biology. However, for such a charismatic insect group, the higher-level phylogenetic relationships among its superfamilies are still poorly resolved. Compared to earlier phylogenomic studies, we increased taxon sampling among Lepidoptera (37 superfamilies and 68 families containing 263 taxa) and acquired a series of large amino-acid datasets from 69,680 to 400,330 for phylogenomic reconstructions. Using these datasets, we explored the effect of different taxon sampling with significant increases in the number of included genes on tree topology by considering a series of systematic errors using maximum-likelihood (ML) and Bayesian inference (BI) methods. Moreover, we also tested the effectiveness in topology robustness among the three ML-based models. The results showed that taxon sampling is an important determinant in tree robustness of accurate lepidopteran phylogenetic estimation. Long-branch attraction (LBA) caused by site-wise heterogeneity is a significant source of bias giving rise to unstable positions of ditrysian groups in phylogenomic reconstruction. Phylogenetic inference showed the most comprehensive framework to reveal the relationships among lepidopteran superfamilies, and presented some newly relationships with strong supports (Papilionoidea was sister to Gelechioidea and Immoidea was sister to Galacticoidea, respectively), but limited by taxon sampling, the relationships within the species-rich and relatively rapid radiation Ditrysia and especially Apoditrysia remain poorly resolved, which need to increase taxon sampling for further phylogenomic reconstruction. The present study demonstrates that taxon sampling is an important determinant for an accurate lepidopteran tree of life and provides some essential insights for future lepidopteran phylogenomic studies.

Enhanced virulence of genetically engineered Autographa californica nucleopolyhedrovirus owing to accelerated viral DNA replication aided by inserted ascovirus genes.

Genetic engineering technology is an ideal method to improve insecticidal efficiency by combining the advantages of different pathogenic microorganisms. Thus, six ascovirus genes were introduced into the genomic DNA of Autographa californica nucleopolyhedrovirus (AcMNPV) to possibly transfer the intrinsically valuable insecticidal properties from ascovirus to baculovirus. The viral budded virus (BV) production and viral DNA replication ability of AcMNPV-111 and AcMNPV-165 were significantly stronger than that of AcMNPV-Egfp (used as the wild-type virus in this study), whereas AcMNPV-33 had reduced ones. AcMNPV-111 and AcMNPV-165 also exhibited excellent insecticidal efficiency in the in vivo bioassays: AcMNPV-111 showed a 24.1% decrease in the LT50 value and AcMNPV-165 exhibited a 56.3% decrease in the LD50 value compared with AcMNPV-Egfp against the 3rd instar of Spodoptera exigua larvae, respectively. Furthermore, the size of the occlusion bodies (OBs) of AcMNPV-33, AcMNPV-111, and AcMNPV-165 were significantly increased compared to that of AcMNPV-Egfp. AcMNPV-111 and AcMNPV-165 had stable virulence against the 2nd to 4th instars tested larvae and higher OB yield than AcMNPV-Egfp in the 3rd and 4th instar larvae. Correlation and regression analyses indicated that it is better to use 5 OBs/larva virus to infect the 2nd instar larvae to produce AcMNPV-111 and 50 OBs/larva virus to infect the 3rd instar larvae to produce AcMNPV-165. The results of this study obtained recombinant viruses with enhanced virulence and exhibited a diversity of ascovirus gene function based on the baculovirus platform, which provided a novel strategy for the improvement of baculovirus as a biological insecticide.

Heliothis virescens ascovirus 3h blocks the cell cycle of Spodoptera exigua fat body cells at G2 /M phase by downregulating cyclin B1 and cyclin-dependent kinase 1.

Ascoviruses are double-stranded DNA viruses that are pathogenic to noctuid larvae. In vitro infection causes the cells to fail to replicate and proliferate normally. However, the molecular mechanisms are unclear. In this study, the transmission electron microscopy data of infected-Spodoptera exigua (Hübner) fat body cells (SeFB, IOZCAS-SpexII-A cells) showed that virions were internalized in phagocytic vesicles, but not in the nucleus. FACS of cell-cycle progression was performed in SeFB cells infected with Heliothis virescens ascovirus 3h (HvAV-3h). The cell cycle phase distributions of the SeFB cells were G1 = 29.52 ± 1.10%, S = 30.33 ± 1.19%, and G2 /M = 40.06 ± 0.75%. The cell culture doubling time was approximately 24 h. The G1 , S, and G2 /M phases were each approximately 8 h. The unsynchronized or synchronized cells were arrested at G2 /M phase after infection with HvAV-3h. Our data also showed that cells with more than 4N DNA content appeared in the HvAV-3h-treated group. While the mRNA levels of cyclin B1 , cyclin H, and cyclin-dependent kinase 1 (CDK1) were downregulated after HvAV-3h infection, the mRNA expression levels of cyclin A, cyclin D, and cyclin B2 were not significantly changed. Western blotting results showed that the expression of cyclin B1 and CDK1 in infected SeFB cells within 24 h postinfection (hpi), and HvAV-3h infection inhibited the expression of cyclin B1 and CDK1 at 12-24 hpi. Overall, these data implied that HvAV-3h infection leads to an accumulation of cells in the G2 /M phases by downregulating the expression of cyclin B1 and CDK1.

Comprehensive identification of protein orthologs in the family Ascoviridae facilitates an understanding of phylogenomics, protein conservation, and phosphorylation.

Analysis of orthology is important for understanding protein conservation, function, and phylogenomics. In this study, we performed a comprehensive analysis of gene orthology in the family Ascoviridae based on identification of 366 protein homologue groups and phylogenetic analysis of 34 non-single-copy proteins. Our findings revealed 90 newly annotated proteins, five newly identified core proteins for the family Ascoviridae, and 14 core proteins for the genus Ascovirus. A phylogenomic tree of 11 Ascoviridae members was constructed based on a concatenation of 35 of the 45 ortholog groups. In combination with phosphoproteomic results and conservation estimations, 30 conserved phosphorylation sites on 17 phosphoproteins were identified from a total of 176 phosphosites on 57 phosphoproteins from Heliothis virescens ascovirus 3h (HvAV-3h), providing potential research targets for investigating the role of these protein in the regulation of viral infection. This study will facilitate genome annotation and comparison of further Ascoviridae members as well as functional genomic investigations.

Phylogenetic relationships among Bombycinae (Lepidoptera, Bombycoidea, and Bombycidae) based on mitochondrial genomes.

The subfamily Bombycinae Latreille, [1802] is an important silk-producing group, including well-known economical insects. Although there are many studies on the development of these economic insects, the relationships between genera/species of this subfamily are still unclear. Two data sets of mitochondrial genomes, 13 protein-coding genes (13PCGs) and 13PCGs-AA, were used to estimate phylogenetic relationships based on the maximum likelihood and Bayesian inference methods. The results strongly support the subfamily Bombycinae as a monophyletic group divided into two clades.

Comparisons of pathogenic course of two Heliothis virescens ascovirus isolates (HvAV-3i and HvAV-3j) in four noctuid (Lepidoptera) pest species.

Ascoviruses are fatal double-stranded DNA viruses with a special pathogenesis in which cells are converted into vesicles with virions. Several closely related ascovirus isolates that shared more than 90% genomic DNA identity showed different pathogenic courses in previous studies. To investigate the pathogenic differences between the related ascovirus isolates, Heliothis virescens ascovirus 3i (HvAV-3i) and Heliothis virescens ascovirus 3j (HvAV-3j) were used to inoculate four noctuid pest species (Helicoverpa armigera, Mythimna separata, Spodoptera frugiperda, and Spodoptera litura), and the pathogenic indexes were recorded. The mortality of HvAV-3i infected H. armigera and S. frugiperda was approximately 60%, while the other HvAV-infected larvae had mortality rates above 90%. The maximum lethal dilution ratios of HvAV-3i in H. armigera, M. separata, S. frugiperda, and S. litura were 1.90 × 107, 1.90 × 103, 1.90 × 108, and 1.90 × 104 viral genome DNA copies/mL, respectively, while the ratios of HvAV-3j were 8.22 × 106, 8.22 × 102, 8.22 × 105, and 8.22 × 103 viral genome DNA copies/mL, respectively. Extended larval survival time was found in the HvAV-infected larvae; median survival time of the HvAV-infected larvae ranged from 13 to 19 days. An additional larval instar was found in HvAV-infected M. separata, S. frugiperda, and S. litura. Larval growth and food intake were significantly inhibited from 2 days post-infection (dpi) in the tested H. armigera, S. frugiperda, and S. litura after infection with HvAV-3i or HvAV-3j. The detoxification enzyme activity of host larvae was influenced after infection with HvAVs, and two different regulation patterns were detected, one in infected H. armigera and M. separata and the other in S. frugiperda and S. litura. The results obtained in this study provide insights into the pathogenic characteristics of ascoviruses.

Fourteen complete mitochondrial genomes of butterflies from the genus Lethe (Lepidoptera, Nymphalidae, Satyrinae) with mitogenome-based phylogenetic analysis.

The mitochondrial genome (mitogenome) can help us understand the phylogenetic relationships within the genus Lethe and the subfamily Satyrinae. In this study, we sequenced the complete mitogenomes of 14 Lethe species, which range in size from 15,225 to 15,271 bp, with both 37 genes (13 PCGs, 22 tRNAs, 2 rRNAs) and a noncoding A + T-rich region. The gene arrangement and orientation is similar to typical mitogenomes of Lepidoptera. The Ka/Ks ratio shows that cox1 has the slowest evolutionary rate. The secondary structure of trnN lacks the Pseudouracil loop (TψC loop) in most Lethe species. The inferred phylogenetic analyses show that Lethe is a well-supported monophyletic group, and reveal 2 major clades within the genus Lethe, which is consistent with previous morphological classifications.

Proteomic analysis of the Heliothis virescens ascovirus 3i (HvAV-3i) virion.

Ascoviruses are enveloped, circular, double-stranded DNA viruses that can effectively control the appetite of lepidopteran larvae, thereby reducing the consequent damage and economic losses to crops. In this study, the virion of a sequenced Heliothis virescens ascovirus 3i (HvAV-3i) strain was used to perform proteomic analysis using both in-gel and in-solution digestion. A total of 81 viral proteins, of which 67 were associated with the virions, were identified in the proteome of HvAV-3i virions. Among these proteins, 23 with annotated functions were associated with DNA/RNA metabolism/transcription, virion assembly, sugar and lipid metabolism, signalling, cellular homoeostasis and cell lysis. Twenty-one viral membrane proteins were also identified. Some of the minor 'virion' proteins identified may be non-virion contaminants of viral proteins synthesized during replication, identified by more recent and highly sensitive methods. The extensive identification of the ascoviral proteome will establish a foundation for further investigation of ascoviral replication and infection.

3H-117, a structural protein of Heliothis virescens ascovirus 3h (HvAV-3h).

The open reading frame 117 (3h-117) of Heliothis virescens ascovirus 3h (HvAV-3h), which is a conserved coding region present in all completely sequenced ascovirus members, was characterized in this study. By RT-PCR detection, 3h-117 transcription began at 6-h post-infection (hpi) and remained stable until 168 hpi in HvAV-3h-infected Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae) larvae. In addition, 3h-117 putatively encodes a 21.5-kDa protein (3H-117) predicted to be a CTD-like phosphatase. Western blot analysis using a prepared rabbit polyclonal antibody specific to 3H-117 showed that the product could be detected at 24 hpi, which remained stably detectable until 168 hpi. The same analysis also demonstrated that the 3H-117 protein localized in the virions of HvAV-3h. Immunofluorescence analysis showed that at 24 hpi, 3H-117 was mainly located in the nuclei of H. armigera larval fat body cells and later spread into the cytoplasm. In summary, our results indicate that 3H-117 is a structural protein of HvAV-3h.

Genomic analysis of a novel isolate Heliothis virescens ascovirus 3i (HvAV-3i) and identification of ascoviral repeat ORFs (aros).

Ascoviruses are circular double-stranded DNA viruses that infect insects. Herein we sequenced and analyzed the genome of the previously unrecorded ascovirus isolate Heliothis virescens ascovirus 3i (HvAV-3i). The genome size is 185,650 bp with 181 hypothetical open reading frames (ORFs). Additionally, definition based on ascovirus repeated ORFs (aros) is proposed; whereby the 29 aros from all sequenced Ascoviridae genomes are divided into six distinct groups. The topological relationship among the isolates of Heliothis virescens ascovirus 3a is (HvAV-3f, {HvAV-3h, [HvAV-3e, (HvAV-3g, HvAV-3i)]}) with every clade well supported by a Bayesian posterior probability of 1.00 and a Bootstrap value of 100%.

Changes in lipid, protein and carbohydrate metabolism in Spodoptera exigua larvae associated with infection by Heliothis virescens ascovirus 3h.

Spodoptera exigua (Lepidoptera: Noctuidae) larvae were used to analyze the regulation of lipid, protein and carbohydrate metabolism in host larvae infected with the Heliothis virescens ascovirus 3h (HvAV-3h). Using histological sections, significant pathological changes were found in the fat bodies of infected larvae from 24 h to 72 h post-infection (hpi). The lipid and protein contents of the infected larvae were significantly higher than those of the uninfected larvae, while the carbohydrate content of the infected larvae was significantly lower than that of the mock-infected larvae. The selected primary metabolite metabolism-associated genes showed different expression patterns. Further co-relationship analysis of the gene expression level and content changes of primary metabolites indicated the following: the acyl-CoA dehydrogenase and fatty acid synthase genes were closely associated with lipid metabolism, and the hexokinase and the glycogen synthase gene expression levels were related to carbohydrate metabolism, while the aminopeptidase N and the protein disulfide isomerase gene expression levels were not correlated with protein metabolism. These results indicate that the HvAV-3h virus stimulates host larval lipid and protein syntheses and inhibits carbohydrate synthesis.

Pathogen-triggered ethylene signaling mediates systemic-induced susceptibility to herbivory in Arabidopsis.

Multicellular eukaryotic organisms are attacked by numerous parasites from diverse phyla, often simultaneously or sequentially. An outstanding question in these interactions is how hosts integrate signals induced by the attack of different parasites. We used a model system comprised of the plant host Arabidopsis thaliana, the hemibiotrophic bacterial phytopathogen Pseudomonas syringae, and herbivorous larvae of the moth Trichoplusia ni (cabbage looper) to characterize mechanisms involved in systemic-induced susceptibility (SIS) to T. ni herbivory caused by prior infection by virulent P. syringae. We uncovered a complex multilayered induction mechanism for SIS to herbivory. In this mechanism, antiherbivore defenses that depend on signaling via (1) the jasmonic acid-isoleucine conjugate (JA-Ile) and (2) other octadecanoids are suppressed by microbe-associated molecular pattern-triggered salicylic acid (SA) signaling and infection-triggered ethylene signaling, respectively. SIS to herbivory is, in turn, counteracted by a combination of the bacterial JA-Ile mimic coronatine and type III virulence-associated effectors. Our results show that SIS to herbivory involves more than antagonistic signaling between SA and JA-Ile and provide insight into the unexpectedly complex mechanisms behind a seemingly simple trade-off in plant defense against multiple enemies.

Analysis on the Complete Mitochondrial Genome of Andraca theae (Lepidoptera: Bombycoidea).

The bombycid moth, Andraca theae (Matsumura) (Lepidoptera: Bombycoidea) is an important pest of tea in southeastern China. In the present study, the mitochondrial genome (mitogenome) of A. theae was amplified by polymerase chain reaction and sequenced. The complete mitogenome of A. theae, encoding 37 genes, was 15,737 bp in length (Genbank no. KX365419), and consisted of 13 protein-coding genes (PCGs), 22 tRNA genes, 2 ribosomal RNA genes and an adenine (A) + thymine (T)-rich region (AT-rich region). The gene order of A. theae mitogenome was typical for Lepidoptera mitogenomes. Except for cox1, which started with CGA, all other 12 PCGs started with ATN. Eleven of the 13 PCGs ended with TAA, expect for cox1 and cox2, which ended with a single T. The maximum likelihood method and the Bayesian method were used to analyze the phylogenetic relationship among 22 representative bombycoid species with a matrix consisting of the 13 PCGs of the mitogenomes of the 22 species. The topological structures of the two phylogenetic trees we constructed were almost identical, with the results indicating that the bombycid species, including A. theae, clustered into a single clade with a bootstrap value of 58% and a posterior probability of 0.98. The phylogenetic relationship among the Bombycoidea species analyzed was Lasiocampidae + (Bombycidae + (Saturniidae + Sphingidae)) which was supported by a high bootstrap value of 100% and a posterior probability of 1.00.

Anti-tumor activities of active ingredients in Compound Kushen Injection.

Kushen (Radix Sophorae Flavescentis) has a long history of use for the treatment of tumors, inflammation and other diseases in traditional Chinese medicine. Compound Kushen Injection (CKI) is a mixture of natural compounds extracted from Kushen and Baituling (Rhizoma Smilacis Glabrae). The main principles of CKI are matrine (MT) and oxymatrine (OMT) that exhibit a variety of pharmacological activities, including anti-inflammatory, anti-allergic, anti-viral, anti-fibrotic and cardiovascular protective effects. Recent evidence shows that these compounds also produce anti-cancer actions, such as inhibiting cancer cell proliferation, inducing cell cycle arrest, accelerating apoptosis, restraining angiogenesis, inducing cell differentiation, inhibiting cancer metastasis and invasion, reversing multidrug resistance, and preventing or reducing chemotherapy- and/or radiotherapy-induced toxicity when combined with chemotherapeutic drugs. In this review, we summarize recent progress in studying the anti-cancer activities of MT, OMT and CKI and their potential molecular targets, which provide clues and references for further study.

Genome sequence and organization analysis of Heliothis virescens ascovirus 3f isolated from a Helicoverpa zea larva.

The complete genome sequence of Heliothis virescens ascovirus 3f (HvAV-3f) was obtained. The HvAV-3f genome has a circular genome of 198,157bp with a G+C content of 46.0%, and encodes 190 open reading frames (ORFs) longer than 69 amino acids. Two major homologous regions (hrs) and 29 'baculovirus repeat ORFs' (bro) were found in the genome. BLAST analyses revealed that three HvAV-3f genes were homologous to that of lepidopteran insects. Nine ORFs were unique to HvAV-3f, in which two ORFs showed significant levels of similarity to genes that have not been previously described for ascoviruses in the Genbank database.

Genomic sequence of Heliothis virescens ascovirus 3g isolated from Spodoptera exigua.

Heliothis virescens ascovirus 3a (HvAV-3a), a member of the family Ascoviridae, has the highest diversity among ascovirus species that have been reported in Australia, Indonesia, China, and the United States. To understand the diversity and origin of this important ascovirus, the complete genome of the HvAV Indonesia strain (HvAV-3g), isolated from Spodoptera exigua, was determined to be 199,721 bp, with a G+C content of 45.9%. Therefore, HvAV-3g has the largest genome among the reported ascovirus genomes to date. There are 194 predicted open reading frames (ORFs) encoding proteins of 50 or more amino acid residues. In comparison to HvAV-3e reported from Australia, HvAV-3g has all the ORFs in HvAV-3e with 6 additional ORFs unique to HvAV-3g, including 1 peptidase C26 gene with the highest identity to Drosophila spp. and 2 gas vesicle protein U (GvpU) genes with identities to Bacillus megaterium. The five unique homologous regions (hrs) and 25 baculovirus repeat ORFs (bro) of HvAV-3g are highly variable.

Genome of Thysanoplusia orichalcea multiple nucleopolyhedrovirus lacks the superoxide dismutase gene.

Thysanoplusia orichalcea multiple nucleopolyhedrovirus (ThorMNPV) has high virulence to Trichoplusia ni and Pseudoplusia includens larvae, with a potential for biological control of insect pests. The genome of ThorMNPV was sequenced and found to be 132,978 bp, with a G+C content of 37.9%. There are 145 predicted open reading frames (ORFs), encoding proteins of 50 or more amino acid residues with minimal overlap. Of the 145 ORFs, 141 appeared to be homologous to those of Autographa californica MNPV (AcMNPV). In comparison to AcMNPV, 9 ORFs of AcMNPV were absent in ThorMNPV, including the superoxide dismutase (sod) gene.

Four new species of Gracillariidae (Lepidoptera) from China and Japan, and description of the pupal morphology of the genera Corythoxestis, Eumetriochroa, Guttigera, and Metriochroa.

Four new leaf mining Oecophyllembiinae (Gracillariidae) species are described from China and Japan: Metriochroa symplocosella sp. nov. (host plants: Symplocos anomala, S. sumuntia, Symplocaceae) from China, Guttigera schefflerella sp. nov. (host plant: Schefflera octophylla, Araliaceae), Eumetriochroa araliella sp. nov. (host plants: Dendropanax trifidus, Evodiopanax innovans, Eleutherococcus sciadophylloides and Fatsia japonica, Araliaceae) and Corythoxestis tricalysiella sp. nov. (host plant: Tricalysia dubia, Rubiaceae) from Japan. Corythoxestis sunosei (Kumata, 1998) is recorded from new host plants: Adinapilulifera and Mussaenda parviflora, Rubiaceae, from Japan. The female adult and pupal morphologies, life history and host plant of the genus Guttigera are described for the first time. Pupae of seven species of four genera: Corythoxestis, Eumetriochroa, Guttigera, and Metriochroa, are described for the first time. We provide morphological diagnostic differences between species and genera of Oecophyllembiinae and Phyllocnistis. Our preliminary data suggest that Oecophyllembiinae species have three valuable pupal diagnostic characters: 1) cocoon cutter with unique lateral processes or setae on the clypeus, 2) tergal spines with only a pair of dorsal setae, and 3) cremaster with more than two pairs of caudal processes, while Phyllocnistis species possess 1) cocoon cutter without lateral processes or setae on clypeus, 2) tergal spines with a pair of dorsal setae and dorsal hooks, and 3) cremaster with only a pair of caudal processes.

An Efficient Feature Selection Algorithm for Gene Families Using NMF and ReliefF.

Gene families, which are parts of a genome's information storage hierarchy, play a significant role in the development and diversity of multicellular organisms. Several studies have focused on the characteristics of gene families, such as function, homology, or phenotype. However, statistical and correlation analyses on the distribution of gene family members in the genome have yet to be conducted. Here, a novel framework incorporating gene family analysis and genome selection based on NMF-ReliefF is reported. Specifically, the proposed method starts by obtaining gene families from the TreeFam database and determining the number of gene families within the feature matrix. Then, NMF-ReliefF is used to select features from the gene feature matrix, which is a new feature selection algorithm that overcomes the inefficiencies of traditional methods. Finally, a support vector machine is utilized to classify the acquired features. The results show that the framework achieved an accuracy of 89.1% and an AUC of 0.919 on the insect genome test set. We also employed four microarray gene data sets to evaluate the performance of the NMF-ReliefF algorithm. The outcomes show that the proposed method may strike a delicate balance between robustness and discrimination. Additionally, the proposed method's categorization is superior to state-of-the-art feature selection approaches.