Manduca sexta, known as the tobacco hornworm or Carolina sphinx moth, is a lepidopteran insect that is used extensively as a model system for research in insect biochemistry, physiology, neurobiology, development, and immunity. One important benefit of this species as an experimental model is its extremely large size, reaching more than 10 g in the larval stage. M. sexta larvae feed on solanaceous plants and thus must tolerate a substantial challenge from plant allelochemicals, including nicotine. We report the sequence and annotation of the M. sexta genome, and a survey of gene expression in various tissues and developmental stages. The Msex_1.0 genome assembly resulted in a total genome size of 419.4 Mbp. Repetitive sequences accounted for 25.8% of the assembled genome. The official gene set is comprised of 15,451 protein-coding genes, of which 2498 were manually curated. Extensive RNA-seq data from many tissues and developmental stages were used to improve gene models and for insights into gene expression patterns. Genome wide synteny analysis indicated a high level of macrosynteny in the Lepidoptera. Annotation and analyses were carried out for gene families involved in a wide spectrum of biological processes, including apoptosis, vacuole sorting, growth and development, structures of exoskeleton, egg shells, and muscle, vision, chemosensation, ion channels, signal transduction, neuropeptide signaling, neurotransmitter synthesis and transport, nicotine tolerance, lipid metabolism, and immunity. This genome sequence, annotation, and analysis provide an important new resource from a well-studied model insect species and will facilitate further biochemical and mechanistic experimental studies of many biological systems in insects.

Aggressive forms of breast cancer, such as Her2+ and triple negative breast cancer (TNBC), are enriched in breast cancer stem cells (BCSC) and have limited therapeutic options. BCSC represent a key cellular reservoir for relapse, metastatic progression and therapeutic resistance. Their ability to resist common cytotoxic therapies relies on different mechanisms, including improved detoxification. The cystine-glutamate antiporter protein xCT (SLC7A11) regulates cystine intake, conversion to cysteine and subsequent glutathione synthesis, protecting cells against oxidative and chemical insults. Our previous work showed that xCT is highly expressed in tumorspheres derived from breast cancer cell lines and downregulation of xCT altered BCSC function in vitro and inhibited pulmonary metastases in vivo. We further strengthened these observations by developing a virus-like-particle (VLP; AX09-0M6) immunotherapy targeting the xCT protein. AX09-0M6 elicited a strong antibody response against xCT including high levels of IgG2a antibody. IgG isolated from AX09-0M6 treated mice bound to tumorspheres, inhibited xCT function as assessed by reactive oxygen species generation and decreased BCSC growth and self-renewal. To assess if AX09-0M6 impacts BCSC in vivo seeding, Her2+ TUBO-derived tumorspheres were injected into the tail vein of AX09-0M6 or control treated female BALB/c mice. AX09-0M6 significantly inhibited formation of pulmonary nodules. To evaluate its ability to impact metastases, AX09-0M6 was administered to mice with established subcutaneous 4T1 tumors. AX09-0M6 administration significantly hampered tumor growth and development of pulmonary metastases. These data show that a VLP-based immunization approach inhibits xCT activity, impacts BCSC biology and significantly reduces metastatic progression in preclinical models.

Extracellular serine proteinase cascades stimulate prophenoloxidase (proPO) activation and antimicrobial peptide production in insect innate immune responses. Serpins in plasma regulate such cascades by selective inhibition of proteinases, in reactions which result in the formation of covalent serpin-proteinase complexes. We carried out experiments to identify plasma proteinases that are inhibited by Manduca sexta serpin-3, an immune-inducible serpin known to regulate proPO activation. Immunoaffinity chromatography, using antiserum to serpin-3, yielded serpin-3 complexes with proteinases identified by immunoblot analysis as prophenoloxidase-activating proteinase (PAP)-1, PAP-2, PAP-3, and hemolymph proteinase 8 (HP8). HP8 can cleave and activate the Toll ligand, Spätzle, leading to synthesis of antimicrobial peptides. Analysis by mass spectrometry of tryptic peptides derived from the serpin-3 complexes confirmed the presence of PAP-1, PAP-3, and HP8. Purified recombinant serpin-3 and active HP8 formed an SDS-stable complex in vitro. Identification of serpin-3-proteinase complexes in plasma provides insight into proteinase targets of serpin-3 and extends the understanding of serpin/proteinase function in the immune response of M. sexta.

SeroNet institutions have developed or implemented 61 diverse COVID-19 serological assays and are collaboratively working to harmonize these assays using reference materials to establish standardized reporting units. This will facilitate clinical interpretation of serology results and cross-comparison of research data.

Entomopathogenic nematode infective juveniles are likely to encounter both uninfected and infected insects and host quality depends on the stage of the infection. We hypothesized that nematode response to infected hosts will change over the course of an infection. Here, we tested this hypothesis by focusing on the influence of host infection status on long-range attraction to host volatile cues. The attraction response of 3 nematode species (Steinernema carpocapsae, S. glaseri and S. riobrave) with different foraging strategies to infected and uninfected insects (Galleria mellonella and Tenebrio molitor) was tested at 24 h intervals from start of infection to emergence of infective juveniles from depleted host. As expected, based on their foraging strategies, S. carpocapsae was not very responsive to hosts, S. glaseri was highly responsive and S. riobrave was intermediate. Generally, the level of attraction did not change with time after infection and was similar between infected and uninfected hosts. An exception was S. glaseri infected T. molitor, which tended to be less attractive to S. glaseri than uninfected hosts. These results suggest that any influence of host infection status on infection behaviour is occurring at subsequent steps in the host-infection process than host attraction, or involves non-volatile cues.