Non-mammalian Sources Research Articles

Modifying Naturally Occurring, Nonmammalian-Sourced Biopolymers for Biomedical Applications.

Natural biopolymers have a rich history, with many uses across the fields of healthcare and medicine, including formulations for wound dressings, surgical implants, tissue culture substrates, and drug delivery vehicles. Yet, synthetic-based materials have been more successful in translation due to precise control and regulation achievable during manufacturing. However, there is a renewed interest in natural biopolymers, which offer a diverse landscape of architecture, sustainable sourcing, functional groups, and properties that synthetic counterparts cannot fully replicate as processing and sourcing of these materials has improved. Proteins and polysaccharides derived from various sources (crustaceans, plants, insects, etc.) are highlighted in this review. We discuss the common types of polysaccharide and protein biopolymers used in healthcare and medicine, highlighting methods and strategies to alter structures and intra- and interchain interactions to engineer specific functions, products, or materials. We focus on biopolymers obtained from natural, nonmammalian sources, including silk fibroins, alginates, chitosans, chitins, mucins, keratins, and resilins, while discussing strategies to improve upon their innate properties and sourcing standardization to expand their clinical uses and relevance. Emphasis will be placed on methods that preserve the structural integrity and native biological functions of the biopolymers and their makers. We will conclude by discussing the untapped potential of new technologies to manipulate native biopolymers while controlling their secondary and tertiary structures, offering a perspective on advancing biopolymer utility in novel applications within biomedical engineering, advanced manufacturing, and tissue engineering.

Cell culture-derived extracellular vesicles: Considerations for reporting cell culturing parameters.

Cell culture-conditioned medium (CCM) is a valuable source of extracellular vesicles (EVs) for basic scientific, therapeutic and diagnostic applications. Cell culturing parameters affect the biochemical composition, release and possibly the function of CCM-derived EVs (CCM-EV). The CCM-EV task force of the Rigor and Standardization Subcommittee of the International Society for Extracellular Vesicles aims to identify relevant cell culturing parameters, describe their effects based on current knowledge, recommend reporting parameters and identify outstanding questions. While some recommendations are valid for all cell types, cell-specific recommendations may need to be established for non-mammalian sources, such as bacteria, yeast and plant cells. Current progress towards these goals is summarized in this perspective paper, along with a checklist to facilitate transparent reporting of cell culturing parameters to improve the reproducibility of CCM-EV research.

EXPLOITING NOVEL 3D JELLYFISH COLLAGEN SPONGES FOR BONE TISSUE REGENERATION

Jellyfish collagens exhibit auspicious perspectives for tissue engineering applications primarily due to their outstanding compatibility with a wide range of cell types, low immunogenicity and biodegradability. Furthermore, derived from a non-mammalian source, jellyfish collagens reduce the risk of disease transmission, minimising therefore the ethical and safety concerns. The current study aims to investigate the potential of 3-dimensional jellyfish collagen sponges (3D-JCS) in promoting bone tissue regeneration.Both qualitative and quantitative analyses were performed in order to assess adhesion and proliferation of MC3T3 cells on 3D-JCL, as well as cell migration and bone-like ECM production. Histological and fluorescent dyes were used to stain mineral deposits (i.e. Alizarin Red S (ARS), Von Kossa, Tetracycline hydrochloride) while images were acquired using optical and confocal microscopy.Qualitative data indicated successful adhesion and proliferation of MC3T3 cells on the 3D-JCS as well as cell migration along with ECM production both on the inner and outer surface of the scaffolds. Moreover, quantitative analyses indicated a four-fold increase of ARS uptake between 2- and 3-dimensional cultures (N=3) as well as an eighteen-fold increase of ARS uptake for the 3D-JCS (N=3) when cultured in osteogenic conditions compared to control. This suggests the augmented osteogenic potential of MC3T3 cells when cultured on 3D-JCS. Nevertheless, the cell-mediated mineral deposition appeared to alter the mechanical properties of the jellyfish collagen sponges that were previously reported to exhibit low mechanical properties (compressive modulus: 1-2 kPa before culture).The biocompatibility, high porosity and pore interconnectivity of jellyfish collagen sponges promoted adhesion and proliferation of MC3T3 cells as well as cell migration and bone-like ECM production. Their unique features recommend the jellyfish collagen sponges as superior biomaterial scaffolds for bone tissue regeneration. Further studies are required to quantify the change in mechanical properties of the cell-seeded scaffolds and confirm their suitability for bone tissue regeneration. We predict that the 3D-JCS will be useful for future studies in both bone and bone-tendon interface regeneration.AcknowledgmentsThis research has been supported by a Medical Research Scotland Studentship award (ref: -50177-2019) in collaboration with Jellagen Ltd.

Fish collagen peptides and its encapsulation

In the last decade, more research has been conducted in order to find a better way for utilizing the waste product generated from food processing industries. The increasing demand of industrial by-products is one of the main reasons for the conversion of these wastes into valuable products. Among the different valuable products from the waste, the extraction of collagen could be a better way of utilizing the wastes, due to their effective applications in biomedical and pharmaceutical industries. The most abundant sources of collagen and gelatin are land-based animals, such as cows and pigs. However, the extraction of collagen and gelatin from non-mammalian sources such as fish has been highly influenced in current society due to some religious and disease transmission issues. Many studies have dealt with the extraction and physicochemical, functional properties of collagen from fish wastes. The present work is a compilation of information on encapsulation of collagen from fish processing wastes.

Critical Assessment of In Vitro Screening of α-Glucosidase Inhibitors from Plants with Acarbose as a Reference Standard.

Postprandial hyperglycemia is treated with the oral antidiabetic drug acarbose, an intestinal α-glucosidase inhibitor. Side effects of acarbose motivated a growing number of screening studies to identify novel α-glucosidase inhibitors derived from plant extracts and other natural sources. As "gold standard", acarbose is frequently included as the reference standard to assess the potency of these candidate α-glucosidase inhibitors, with many outperforming acarbose by several orders of magnitude. The results are subsequently used to identify suitable compounds/products with strong potential for in vivo efficacy. However, most α-glucosidase inhibitor screening studies use enzyme preparations obtained from nonmammalian sources (typically Saccharomyces cerevisiae), despite strong evidence that inhibition data obtained using nonmammalian α-glucosidase may hold limited value in terms of identifying α-glucosidase inhibitors with actual in vivo hypoglycemic potential. The aim was to critically discuss the screening of novel α-glucosidase inhibitors from plant sources, emphasizing inconsistencies and pitfalls, specifically where acarbose was included as the reference standard. An assessment of the available literature emphasized the cruciality of stating the biological source of α-glucosidase in such screening studies to allow for unambiguous and rational interpretation of the data. The review also highlights the lack of a universally adopted screening assay for novel α-glucosidase inhibitors and the commercial availability of a standardized preparation of mammalian α-glucosidase.

Exploring lectin–glycan interactions to combat COVID-19: Lessons acquired from other enveloped viruses

The emergence of a new human coronavirus (SARS-CoV-2) has imposed great pressure on the health system worldwide. The presence of glycoproteins on the viral envelope opens a wide range of possibilities for application of lectins to address some urgent problems involved in this pandemic. In this work, we discuss the potential contributions of lectins from non-mammalian sources in the development of several fields associated with viral infections, most notably COVID-19. We review the literature on the use of non-mammalian lectins as a therapeutic approach against members of the Coronaviridae family, including recent advances in strategies of protein engineering to improve their efficacy. The applications of lectins as adjuvants for antiviral vaccines are also discussed. Finally, we present some emerging strategies employing lectins for the development of biosensors, microarrays, immunoassays and tools for purification of viruses from whole blood. Altogether, the data compiled in this review highlights the importance of structural studies aiming to improve our knowledge about the basis of glycan recognition by lectins and its repercussions in several fields, providing potential solutions for complex aspects that are emerging from different health challenges.

Expression and purification of the human tumor suppressor protein RNASET2 in CHO–S cells

Members of the T2 extracellular ribonucleases family have long been reported as stress response proteins, often involved in host defence, in many different taxonomic groups. In particular, the human RNASET2 protein (hRNASET2) has been reported as an extracellular tumor suppressor protein, endowed with the ability to act as an “alarmin” signalling molecule following its expression and secretion in the tumor microenvironment by cancer cells and the subsequent recruitment and activation of cells belonging to the host innate immune system. Many in vitro and in vivo assays have been recently reported in support of the oncosuppressive role of hRNASET2: most of them relied on genetically engineered cell lines and the use of recombinant proteins from non-mammalian sources. In order to ensure a human-like glycosylation pattern, here we report for the first time the expression of recombinant hRNASET2 in the CHO–S cell line. We established a simple one-step chromatographic purification procedure that resulted in the production of 5 mg of endotoxin-free hRNASET2 per liter of culture, with a >95% purity degree. hRNASET2 expressed in CHO–S cells displayed a high degree of glycosylation homogeneity and a secondary structure content in agreement with that determined from the crystal structure. Indeed, recombinant hRNASET2 was active at both enzymatic and functional level, as stated by a biological activity assay. The availability of a pure, homogeneous recombinant human RNASET2 would provide a key tool to better investigate its non cell-autonomous roles in the context of cancer development and growth.

Type I Collagen from Jellyfish Catostylus mosaicus for Biomaterial Applications.

Collagen is the predominant protein in animal connective tissues and is widely used in tissue regeneration and other industrial applications. Marine organisms have gained interest as alternative, nonmammalian collagen sources for biomaterial applications because of potential medical and economic advantages. In this work, we present physicochemical and biofunctionality studies of acid solubilized collagen (ASC) from jellyfish Catostylus mosaicus (JASC), harvested from the Persian Gulf, compared with ASC from rat tail tendon (RASC), the industry-standard collagen used for biomedical research. From the protein subunit (alpha chain) pattern of JASC, we identified it as a type I collagen, and extensive molecular spectroscopic analyses showed similar triple helical molecular signatures for JASC and RASC. Atomic force microscopy of fibrillized JASC showed clear fibril reassembly upon pH neutralization though with different temperature and concentration dependence compared with RASC. Molecular (natively folded, nonfibrillized) JASC was shown to functionalize rigid substrates and promote MC3T3 preosteoblast cell attachment and proliferation better than RASC over 6 days. On blended collagen-agarose scaffolds, both RASC and JASC fibrils supported cell attachment and proliferation, and scaffolds with RASC fibrils showed more cell growth after 6 days compared with those scaffolds with JASC fibrils. These results demonstrate the potential for this new type I collagen as a possible alternative to mammalian type I collagen for biomaterial applications.

Survey of the C20 and C22 oxylipin family in marine diatoms

Diatoms are highly abundant microalgae that are prolific sources for bioactive oxylipins, which are generated by the oxidative transformation of polyunsaturated fatty acids and contribute to the chemical defense of algae. We introduce herein a targeted survey of oxylipins from four diatom species with a focus on metabolites that also occur in mammalians. Analytical data from LC–MS/MS measurements indicate that diatoms produce a surprising variety of oxylipins, including several hydroxylated and dihydroxylated C18-, C20-, and C22-polyunsaturated fatty acids. It was also found that diatoms represent the first non-mammalian source for the resolvins (Rv)E2 and RvE3, which have recently been identified as modulators of inflammation in humans and rodents. The oxylipins are produced in response to mechanical disruption of the cells, suggesting their potential involvement in the wound-activated chemical defense of these diatoms.

Edible Scaffolds Based on Non-Mammalian Biopolymers for Myoblast Growth.

In vitro meat has recently emerged as a new concept in food biotechnology. Methods to produce in vitro meat generally involve the growth of muscle cells that are cultured on scaffolds using bioreactors. Suitable scaffold design and manufacture are critical to downstream culture and meat production. Most current scaffolds are based on mammalian-derived biomaterials, the use of which is counter to the desire to obviate mammal slaughter in artificial meat production. Consequently, most of the knowledge is related to the design and control of scaffold properties based on these mammalian-sourced materials. To address this, four different scaffold materials were formulated using non-mammalian sources, namely, salmon gelatin, alginate, and additives including gelling agents and plasticizers. The scaffolds were produced using a freeze-drying process, and the physical, mechanical, and biological properties of the scaffolds were evaluated. The most promising scaffolds were produced from salmon gelatin, alginate, agarose, and glycerol, which exhibited relatively large pore sizes (~200 μm diameter) and biocompatibility, permitting myoblast cell adhesion (~40%) and growth (~24 h duplication time). The biodegradation profiles of the scaffolds were followed, and were observed to be less than 25% after 4 weeks. The scaffolds enabled suitable myogenic response, with high cell proliferation, viability, and adequate cell distribution throughout. This system composed of non-mammalian edible scaffold material and muscle-cells is promising for the production of in vitro meat.

Surface modification of PVDF using non-mammalian sources of collagen for enhancement of endothelial cell functionality.

Although polyvinylidene fluoride (PVDF) is non-toxic and stable in vivo, its hydrophobic surface has limited its bio-applications due to poor cell-material interaction and thrombus formation when used in blood contacting devices. In this study, surface modification of PVDF using naturally derived non-mammalian collagen was accomplished via direct surface-initiated atom transfer radical polymerisation (SI-ATRP) to enhance its cytocompatibility and hemocompatibility. Results showed that Type I collagen was successfully extracted from fish scales and bullfrog skin. The covalent immobilisation of fish scale-derived collagen (FSCOL) and bullfrog skin-derived collagen (BFCOL) onto the PVDF surface improves the attachment and proliferation of human umbilical vein endothelial cells (HUVECs). Furthermore, both FSCOL and BFCOL had comparable anti-thrombogenic profiles to that of commercially available bovine collagen (BVCOL). Also, cell surface expression of the leukocyte adhesion molecule was lower on HUVECs cultured on non-mammalian collagen surfaces than on BVCOL, which is an indication of lower pro-inflammatory response. Overall, results from this study demonstrated that non-mammalian sources of collagen could be used to confer bioactivity to PVDF, with comparable cell-material interactions and hemocompatibility to BVCOL. Additionally, higher expression levels of Type IV collagen in HUVECs cultured on FSCOL and BFCOL were observed as compared to BVCOL, which is an indication that the non-mammalian sources of collagen led to a better pro-angiogenic properties, thus making them suitable for blood contacting applications.Electronic supplementary materialThe online version of this article (doi:10.1007/s10856-015-5651-8) contains supplementary material, which is available to authorized users.

Potential functional applications of extracellular vesicles: a report by the NIH Common Fund Extracellular RNA Communication Consortium

The NIH Extracellular RNA Communication Program's initiative on clinical utility of extracellular RNAs and therapeutic agents and developing scalable technologies is reviewed here. Background information and details of the projects are presented. The work has focused on modulation of target cell fate by extracellular vesicles (EVs) and RNA. Work on plant-derived vesicles is of intense interest, and non-mammalian sources of vesicles may represent a very promising source for different therapeutic approaches. Retro-viral-like particles are intriguing. Clearly, EVs share pathways with the assembly machinery of several other viruses, including human endogenous retrovirals (HERVs), and this convergence may explain the observation of viral-like particles containing viral proteins and nucleic acid in EVs. Dramatic effect on regeneration of damaged bone marrow, renal, pulmonary and cardiovascular tissue is demonstrated and discussed. These studies show restoration of injured cell function and the importance of heterogeneity of different vesicle populations. The potential for neural regeneration is explored, and the capacity to promote and reverse neoplasia by EV exposure is described. The tremendous clinical potential of EVs underlies many of these projects, and the importance of regulatory issues and the necessity of general manufacturing production (GMP) studies for eventual clinical trials are emphasized. Clinical trials are already being pursued and should expand dramatically in the near future.

Biomedical and Pharmaceutical Application of Fish Collagen and Gelatin: A Review

In last decade, more research has been conducted in order to find the better way for utilizing the wastes product generated from food processing industries. The increasing demand of industrial by-products is one of the main reasons for the conversion of these wastes into valuable products. Among the different valuable products from the waste, the extraction of collagen and gelatin could be a better way of utilizing the wastes, due to their effective applications in biomedical and pharmaceutical industries. The most abundant source of collagen and gelatin are land-based animals, such as cow and pig. However, the extraction of collagen and gelatin from non-mammalian sources such as fish has been high influences in current society due to some religious and disease transmission issues. Many studies have dealt with the extraction and functional properties of collagen and gelatin from fish wastes. The present work is a compilation of information on biomedical and pharmaceutical application of collagen and gelatin from fish processing wastes.

Expression of recombinant antibodies.

Recombinant antibodies are highly specific detection probes in research, diagnostics, and have emerged over the last two decades as the fastest growing class of therapeutic proteins. Antibody generation has been dramatically accelerated by in vitro selection systems, particularly phage display. An increasing variety of recombinant production systems have been developed, ranging from Gram-negative and positive bacteria, yeasts and filamentous fungi, insect cell lines, mammalian cells to transgenic plants and animals. Currently, almost all therapeutic antibodies are still produced in mammalian cell lines in order to reduce the risk of immunogenicity due to altered, non-human glycosylation patterns. However, recent developments of glycosylation-engineered yeast, insect cell lines, and transgenic plants are promising to obtain antibodies with “human-like” post-translational modifications. Furthermore, smaller antibody fragments including bispecific antibodies without any glycosylation are successfully produced in bacteria and have advanced to clinical testing. The first therapeutic antibody products from a non-mammalian source can be expected in coming next years. In this review, we focus on current antibody production systems including their usability for different applications.

Evaluation of a bioresorbable collagen membrane of fish origin in the treatment of periodontal intrabony defects: A prospective clinical study.

Recently, there has been interest in non-mammalian collagen sources such as fish collagen in the development of biomatrices and scaffolds for periodontal regeneration. In the present study, a novel collagen barrier membrane of fish origin was assessed in the treatment of periodontal intra-bony defects. Ten systemically healthy chronic periodontitis patients having an osseous defect in the mandibular posterior teeth were selected and following the open flap debridement, a collagen membrane was placed over the defect and the flap was sutured with interrupted sutures. Clinical parameters such as Plaque Index, Gingival Bleeding Index, probing pocket depth (PPD), relative attachment level (RAL), and recession (R) were recorded at baseline, 6 and 9 months, whereas radiographic evaluation was done to assess alveolar crestal bone level and defect depth fill at 6 and 9 months using Auto-computer aided design (ACAD) 2007 software. Statistical significance was set at 5% level of significance. There was statistical significant differences with respect to periodontal clinical parameters such as Plaque Index, Gingival Bleeding Index, PPD, RAL, and gingival recession assessed at baseline, at 6 and 9 months respectively (P < 0.05), and radiographic evaluation showed a defect fill of 58.62 median % at 9 months. This preliminary study has shown predictable results in using fish collagen membrane, for treating periodontal intra-bony defects. Further, long-term clinical trials are needed to validate the effectiveness of this membrane.

Clinical and radiographical evaluation of a bioresorbable collagen membrane of fish origin in the treatment of periodontal intrabony defects: A preliminary study.

Background:Recently, there has been interest in non-mammalian collagen sources such as fish collagen in periodontal regeneration. In the present study, collagen barrier membrane of fish origin was assessed in the treatment of periodontal intrabony defects.Materials and Methods:Ten systemically healthy chronic periodontitis patients having a paired osseous defect in the mandibular posterior teeth were selected and randomly assigned to receive a collagen membrane (test) or open flap debridement (control) in a split mouth design. Clinical parameters such as Plaque index, Gingival bleeding index, Probing pocket depth, Relative attachment level, and Recession were recorded at baseline, 3, 6, and at 9 months, while radiographic evaluation was done to assess alveolar crestal bone level and percentage of defect fill at 6 and 9 months using autoCAD 2007 software. Student's t test (two-tailed, dependent) was used to find the significance of study parameters on continuous scale. Significance was set at 5% level of significance. Wilcoxon signed rank test was used to find the significance of percentage change of defect fill.Results:The comparison between the two groups did not show any statistically significant differences in the parameters assessed (P > 0.05) but, within each group, clinical parameters showed statistically significant differences from baseline to 9 months (P < 0.05).Conclusion:Within the limits of the study, it can be inferred that no significant differences were found either by using collagen membrane of fish origin or open flap debridement in the treatment of periodontal intrabony defects.

Hyaluronic acid viscosupplements from avian and non‐mammalian sources exhibit biocompatibility profiles with unique, source‐specific, antigenic profiles

The objective of this study was to evaluate and compare the biocompatibility profiles of hyaluronic acid (HA) viscosupplements from avian and non-mammalian sources. Inflammatory and immune reactions were assessed in models of both clinically relevant and stringent immunological exposure conditions. Experiments were conducted to evaluate tissue reactions and immunological responses and assess antibody formation with the capacity to bind directly to and cross-react with the different viscosupplements. Mice were exposed to viscosupplements using the air pouch inflammation model and specific immunization using Freund's complete adjuvant (FCA). Murine pouch membrane tissue reactions revealed generally mild to moderate responses, with cellular infiltration and cytokine profiles of pouch tissue characteristic of predominantly fibroblastic responses rather than marked inflammatory reactions. In vitro testing indicated that pouch injections did not elicit detectable T-cell proliferative responses, while antibody assays revealed that mice immunized with viscosupplements in FCA and subsequently boosted were capable of mounting an antibody response with a range of specificities. High reactivity to avian serum albumin was seen in sera from mice injected with HA from an avian source, while low positive reactivity to the bacterial antigen Staphylococcal Lipotechoic Acid was observed in sera from mice injected with HA from bacterial sources. These specificities did not indicate any propensity to cross-react, suggesting that patients with adverse immune responses to HA from an avian source should be unresponsive to subsequent injection with HA from a non-avian source. Overall, the findings demonstrate that viscosupplements exhibit good biocompatibility profiles in the murine air pouch, but when provoked to elicit immunological reactions exhibit unique antigenic spectra. These findings suggest that an immunologically mediated immune reaction directed against avian proteins should not necessarily be a contraindication for the administration of non-avian viscosupplements.

Terminal disialylated multiantennary complex-type N-glycans carried on acutobin define the glycosylation characteristics of the Deinagkistrodon acutus venom

Glycosylation analysis of nonmammalian sources often springs surprises and conjures up intriguing views of evolutionary adaptation. Many of the constituents of snake venoms are known to be glycosylated and yet very few were fully characterized and accorded specific functions. In the process of glycomic screening through the venoms from Asian pit vipers, a partially O-acetylated NeuAcα2-8NeuAcα2-3Galβ1-4GlcNAcβ1-terminal epitope was found to be the predominant glycosylation characteristic of the snake venom produced by the monotypic Deinagkistrodon acutus, with acutobin, a highly specific fibrinogenase, being identified as a primary protein carrier. Full structural definition and glycosylation site mapping were completed through advanced mass spectrometry analyses at both the glycan and glycopeptide levels in conjunction with chemical and enzymatic cleavages. Although similar occurrence of such terminal disialyl cap on the N-glycans of several mammalian glycoproteins has been implicated, most of these correspond to only minor constituents of the full glycomic heterogeneity and remain poorly characterized. In contrast, each antennae of the hybrid- and complex-type N-glycans derived from acutobin was found to be rather homogeneously disialylated. With up to eight sialic acids evenly distributed on nonextended tetraantennary core structure, these unusual N-glycans are among those of highest sialic acid density ever identified without actually carrying polysialic acid chains. It remains to be tested whether they may serve as multivalent disialyl ligands for several of the human Siglecs and thus meddle with the natural immuno-recognition systems of snakebite victims, apart from affecting the general efficacy of acutobin as anticoagulant in biomedical applications.

Molecular Identification of Host Feeding Patterns of Snow-Melt Mosquitoes (Diptera: Culicidae): Potential Implications for the Transmission Ecology of Jamestown Canyon Virus

We collected blood-fed, snow-melt mosquitoes (Culicidae: Culiseta and Aedes) to describe the feeding patterns of potential mosquito vectors of Jamestown Canyon virus (JCV, Bunyaviridae: Orthobunyavirus). JCV is an arthropod-borne, zoonotic virus with deer as the primary amplifying host in western alpine ecosystems. We collected mosquitoes from natural resting areas, fiber pots, and carbon-dioxide baited miniature light traps in the Colorado Rocky Mountains in 2007. We conducted two polymerase chain reactions to amplify and sequence vertebrate DNA extracted from blood-fed mosquitoes, which yielded comparable, but not identical, results. Mammal-specific primers found mule deer (Odocoileus hemionus) and elk (Cervus elaphus canadensis) as the source of all bloodmeals. To determine if unamplified bloodmeals were from nonmammalian sources, we screened all samples with conserved vertebrate primers, which confirmed the initial polymerase chain reaction results, but also found porcupine (Erethizon dorsatum) and human (Homo sapiens) as additional bloodmeal sources. We consistently found that mule deer were the primary hosts for mosquitoes in this system. These results suggest that snow-melt mosquitoes, in particular A. cataphylla, may be important vectors in western JCV alpine systems and may also act as a bridge vector for JCV from cervid virus reservoirs to humans.

Molecular Identification of Host Feeding Patterns of Snow-Melt Mosquitoes (Diptera: Culicidae): Potential Implications for the Transmission Ecology of Jamestown Canyon Virus

We collected blood-fed, snow-melt mosquitoes (Culicidae: Culiseta and Aedes) to describe the feeding patterns of potential mosquito vectors of Jamestown Canyon virus (JCV, Bunyaviridae: Orthobunyavirus). JCV is an arthropod-borne, zoonotic virus with deer as the primary amplifying host in western alpine ecosystems. We collected mosquitoes from natural resting areas, fiber pots, and carbon-dioxide baited miniature light traps in the Colorado Rocky Mountains in 2007. We conducted two polymerase chain reactions to amplify and sequence vertebrate DNA extracted from blood-fed mosquitoes, which yielded comparable, but not identical, results. Mammal-specific primers found mule deer (Odocoileus hemionus) and elk (Cervus elaphus canadensis) as the source of all bloodmeals. To determine if unamplified bloodmeals were from nonmammalian sources, we screened all samples with conserved vertebrate primers, which confirmed the initial polymerase chain reaction results, but also found porcupine (Erethizon dorsatum) and human (Homo sapiens) as additional bloodmeal sources. We consistently found that mule deer were the primary hosts for mosquitoes in this system. These results suggest that snow-melt mosquitoes, in particular A. cataphylla, may be important vectors in western JCV alpine systems and may also act as a bridge vector for JCV from cervid virus reservoirs to humans.