Culicoides (Diptera: Ceratopogonidae) of Ontario: A Dichotomous Key and Wing Atlas
- CJAI #50
- November 30, 2023
Lauren A.A. Janke1, 2*
,
Stacey Vigil3
,
Kate G. Lindsay2
,
Tara Furukawa-Stoffer4
,
Nicole Colucci4
,
Aruna Ambagala5
,
Robert Hanner2
1John H. Daniels Faculty of Architecture, Landscape, and Design, University of Toronto, Toronto, Ontario, M5S 3E8, Canada. Email: lauren.janke@mail.utoronto.ca
2Department of Integrative Biology, University of Guelph, Guelph, Ontario, N1G 2W1, Canada. Emails: klindsay@uoguelph.ca; rhanner@uoguelph.ca
3Southeastern Cooperative Wildlife Disease Study, College of Veterinary Medicine, University of Georgia, Athens, Georgia, 30602, USA. Email: svigil@uga.edu
4National Centre for Animal Diseases, Canadian Food Inspection Agency, Lethbridge, Alberta, T1J 3Z4, Canada. Emails: tara.furukawa-stoffer@inspection.gc.ca, Nikki.Colucci@alumni.uleth.ca
5National Centre for Foreign Animal Disease, Canadian Food Inspection Agency, Winnipeg, Manitoba, R3E 3M4, Canada. Email: aruna.ambagala@inspection.gc.ca
*Corresponding author
Culicoides (Diptera: Ceratopogonidae) of Ontario: A Dichotomous Key and Wing Atlas
Culicoides (Diptera: Ceratopogonidae) of Ontario: A Dichotomous Key and Wing Atlas
- CJAI #50
- November 30, 2023
Lauren A.A. Janke1, 2*
,
Stacey Vigil3
,
Kate G. Lindsay2
,
Tara Furukawa-Stoffer4
,
Nicole Colucci4
,
Aruna Ambagala5
,
Robert Hanner2
1John H. Daniels Faculty of Architecture, Landscape, and Design, University of Toronto, Toronto, Ontario, M5S 3E8, Canada. Email: lauren.janke@mail.utoronto.ca
2Department of Integrative Biology, University of Guelph, Guelph, Ontario, N1G 2W1, Canada. Emails: klindsay@uoguelph.ca; rhanner@uoguelph.ca
3Southeastern Cooperative Wildlife Disease Study, College of Veterinary Medicine, University of Georgia, Athens, Georgia, 30602, USA. Email: svigil@uga.edu
4National Centre for Animal Diseases, Canadian Food Inspection Agency, Lethbridge, Alberta, T1J 3Z4, Canada. Emails: tara.furukawa-stoffer@inspection.gc.ca, Nikki.Colucci@alumni.uleth.ca
5National Centre for Foreign Animal Disease, Canadian Food Inspection Agency, Winnipeg, Manitoba, R3E 3M4, Canada. Email: aruna.ambagala@inspection.gc.ca
*Corresponding author
Abstract
Culicoides Latreille, 1809 (Diptera: Ceratopogonidae) is a diverse genus of tiny biting flies that are both pests and vectors of pathogens of medical, veterinary, and economic importance. Herein, we provide an identification key and wing atlas of adult female biting midges for the 40 Culicoides species that have been recorded or are likely to be present in Ontario, Canada. The key includes: C. albertensis Wirth and Jones, C. alexanderi Wirth and Hubert, C. arboricola Root and Hoffman, C. atchleyi Wirth and Blanton, C. baueri Hoffman, C. bergi Cochrane, C. bickleyi Wirth and Hubert, C. biguttatus (Coquillett), C. chiopterus (Meigen), C. cockerellii (Coquillett), C. crepuscularis Malloch, C. denticulatus Wirth and Hubert, C. downesi Wirth and Hubert, C. flukei Jones, C. footei Wirth and Jones, C. franclemonti Cochrane, C. frohnei Wirth and Blanton, C. furensoides Williams, C. guttipennis (Coquillett), C. haematopotus Malloch, C. jamnbacki Wirth and Hubert, C. nanus Root and Hoffman, C. obsoletus (Meigen), C. paraimpunctatus Borkent, C. parapiliferus Wirth and Blanton, C. piliferus Root and Hoffman, C. sanguisuga (Coquillett), C. snowi Wirth and Jones, C. sphagnumensis Williams, C. spinosus Root and Hoffman, C. stellifer (Coquillett), C. stilobezzioides Foote and Pratt, C. testudinalis Wirth and Hubert, C. travisi Vargas, C. utowana Jamnback, C. variipennis (Coquillett), C. venustus Hoffman, C. villosipennis Root and Hoffman, C. wisconsinensis Jones, and C. yukonensis Hoffman. We provide a pictorial key based on novel and existing published observations of the morphological features observed from these 40 species, and include a pictorial wing atlas. This key will allow for the identification of Culicoides from Ontario down to the lowest taxonomic level possible including: one subgenus (with three species), one species group (with 11 species), and 26 separate species.
Introduction
The genus Culicoides Latreille (Diptera: Ceratopogonidae) is an abundant, diverse, and economically important group of hematophagous flies, known as biting midges. Culicoides can be found worldwide on most major land masses, with the exception of New Zealand and Antarctica, and together represent the most diverse genus in the family Ceratopogonidae, with more than 1,300 described species (Borkent and Dominiak 2020). These tiny flies, also called no-see-ums or punkies are typically less than 3 mm in length and well known for their irritating bites, as well as their ability to serve as vectors of a variety of pathogens that can affect humans, livestock, and wildlife (Borkent 2004).
Biology
The life cycle of Culicoides normally comprises four stages: egg, larva (with four instars), pupa and adult, although two species are parthenogenetic (Borkent 2004). Larval Culicoides require water or moisture and often inhabit streams, pools, moist soil, dung, or vegetation (Wirth and Hubert 1989). Currently, there is only taxonomic information available for the larvae and pupae for approximately 19% of the world’s Culicoides, meaning that immature stages of many species, and their associated habitats, are still undescribed (Borkent 2012b). Development takes weeks to months, and overwintering occurs in a larval stage in temperate regions (Sick et al. 2019). Most adults survive for around 10–20 days (Mellor et al. 2000) and are typically active at dusk (Blanton and Wirth 1979) or in a crepuscular pattern (Kline and Roberts 1982).
Female Culicoides are hematophagous, and most species require a blood meal for their eggs to reach maturity, although autogeny does occur in both nature and a laboratory setting (Glukhova and Dubrovskaia 1972). Blood feeding habits differ widely, but many North American species are considered either primarily ornithophilic, primarily mammalophilic, or generalist (Jamnback 1965; Blanton and Wirth 1979; Hair and Turner 1968). Within these categories, species will often exhibit further feeding specialization, such as primarily utilizing large mammal hosts, or feeding from specific areas of the host such as the lower legs or flank (Nielsen 1971). Finally, there are some Culicoides species with less typical feeding habits like C. testudinalis Wirth and Hubert, which feeds on turtles (Jamnback 1965), and others with reduced mandibular teeth, suggesting a shift from blood-feeding to flower-feeding (Wirth and Hubert 1962). Determining which species feed on which host/hosts is essential in determining whether a species is or has the potential to act as a vector of a pathogen (Kettle 1977).
Culicoides as a Vector
Various Culicoides species around the world have been implicated as vectors for at least 66 viruses, 15 protozoa, and 26 species of nematodes (Borkent 2004). Examples include veterinary arboviruses affecting livestock and wildlife such as bluetongue virus (BTV), epizootic hemorrhagic disease virus (EHDV), Schmallenberg virus, and African horse sickness virus (Borkent 2004; Harrup et al. 2015; Borkent 2017). Because of the lack of Culicoides surveillance activity in Canada, little is known of the country-level distribution of these biting midges acting as vectors. Culicoides sonorensis is considered one of the main vectors of BTV and EHDV in North America (Tabachnik 1996; Carpenter et al. 2015). It was previously suggested that the increased rates of BTV and EHDV in eastern Canada were due to a recent range expansion of the western species C. sonorensis (Jewiss-Gaines et al. 2017; Purse et al. 2015; Allen et al. 2019). However, recent data from Shults et al. (2022a, 2022b) suggested that this eastern Canadian species believed to be C. sonorensis is actually C. albertensis Wirth and Jones. Culicoides albertensis is sympatric with C. sonorensis but not a confirmed vector of BTV and EHDV (Shults et al. 2022b). An alternative explanation for higher rates of Culicoides-borne disease in eastern Canada is that other species, such as C. stellifer (Coquillett) and C. venustus Hoffman, which have been implicated as vectors of EHDV in the southern United States, could be responsible for the spread (McGregor et al. 2019; Shults et al. 2020). There has been a northward expansion of several Culicoides-borne viruses in North America, increasing the likelihood of these diseases being spread to Ontario and other disease-free regions (Ruder et al. 2015; Stallknecht et al. 2015; Allen et al. 2019). Potential reasons for this northward expansion include strong wind currents, movement of livestock hosts or range shifts associated with climate change (Zuliani et al. 2014; Elbers et al. 2015; Samy and Peterson, 2016). The paths of Culicoides-borne diseases are often ecologically complex and may include multiple vectors and host species within a single region (Purse et al. 2015). Since many species of Culicoides worldwide are vectors of economically important viruses that affect wild and domestic ruminants, such as deer, cattle, sheep, and goats, as well as equines, having sound morphological identifications is important for successful surveillance, management, and mitigation strategies (Harrup et al. 2015).
Culicoides Identification
Species level identification of Culicoides is difficult for a variety of reasons. Specimen collection is time-consuming and often requires specialized traps and/or equipment particular to each life stage. Observing such tiny organisms requires the use of high magnification, usually a microscope, to see even the most basic characteristics. Furthermore, specimens must also often be slide-mounted to observe diagnostic characters. Morphological identification of Culicoides often depends upon published species descriptions, which vary widely, and are often dated, cryptic, and based on subjective traits (Borkent and Grogan 2009). The identification process requires access to keys and descriptions from multiple sources (Swanson 2012), as well as a high degree of expertise in working with biting midges. The ability to identify Culicoides species is of increasing importance as the current climate crisis affects the distributions of several arthropod species of veterinary importance (Elbers et al. 2015; Vigil et al. 2018).
In the Nearctic region north of Mexico, there are 150 known species of Culicoides, comprising 13 subgenera, seven species groups, and nine unplaced species (Borkent and Grogan 2009). The subgenera and species groups are important for understanding and identifying this diverse genus, and are generally phenetic, or based on shared morphological characteristics. However, many researchers agree that the current taxonomic organization, which is ambiguous and fails to incorporate many species, needs to be thoroughly revised to reflect groups of evolutionarily related species based on derived traits, rather than on only their current morphology (Borkent and Grogan 2009, Harrup et al. 2015, Borkent and Dominiak 2020). In addition to these taxonomic challenges, most identification keys to Culicoides are dated, difficult to follow, and may only include species from limited geographic areas, specific subgenera, or species groups. This is especially true in Canada and the Nearctic, as there are no comprehensive keys to North American Culicoides in the literature (Borkent and Dominiak, 2020).
Key Preparation
Here we present a key for the 40 species of Culicoides in Ontario, using adult female morphology. We include C. atchleyi, which has been reported in Alaska, Nova Scotia, and New Mexico (Wirth et al. 1985) and was included due to the extremely wide distribution which extends over Ontario.
This key is based on the morphological characteristics of adult female Culicoides specimens, the specimens used for reference in this key have typically been stored in ethanol or slide mounted. For most species female wing patterns are more distinct, and male representatives are less frequently obtained via light traps (Gonzalez et al. 2013; Venter et al. 2009), which are the predominant method of collecting Culicoides. However, in some limited cases, keys using male features are available (Vargas 1960), and males can provide additional diagnostic characteristics (Swanson 2012). Additionally, there are keys for several described pupae of Culicoides species (Borkent 2012a; Shults and Borkent 2018).
Traditional Culicoides species descriptions often rely on characteristics that require specimens to be dissected and slide-mounted, as described in Borkent and Spinelli (2007), to observe morphology under high magnification. The goal of this publication is to provide a means for the morphological identification of Culicoides species, with a focus on ease of use and reducing the necessity for slide-mounting specimens. Wherever possible, this key uses characteristics that are visible under lower magnification, and a strong stereomicroscope will often suffice for visualizing characteristics in whole, unmounted specimens. In addition, we avoid using measurements and ratios on the wings, and we only suggest simple antennal ratios in some dichotomies. However, even when visible, some characteristics will be difficult to identify and define in an unmounted specimen, such as spermathecae. Therefore, slide-mounting may still be helpful or necessary to differentiate some species. It is also important to note that characteristics such as macrotrichia may fall off of specimens as they are collected to varying degrees, and therefore this should only be used as a diagnostic characteristic if the macrotrichia are present on a specimen.
Species not recorded in Ontario but occurring in bordering States or Provinces were not included in the current key as it was beyond the scope of this paper, but are mentioned in reference to similar species in the species information pages when relevant. Several of these species occurring at least in part in New York and/or New Brunswick (C. (Beltranmyia) hollensis Melander and Brues, C. (B.) bermudensis Williams, and C. (Oecacta) furens (Poey) (Jamnback 1965; Borkent and Grogan 2009)) are collected exclusively in salt marshes on the east coast (Jamnback 1965) and are therefore unlikely to be encountered in Ontario. Other species occurring at least in part in Quebec and/or New York and/or Michigan (C. (Amossovia) beckae Wirth and Blanton, C. (Drymodesmyia) hinmani Khalaf, C. (Avaritia) juddi Cochrane, C. (Silvaticulicoides) loisae Jamnback, C. (Avaritia) pechumani Cochrane, and C. scanloni Wirth and Hubert (Borkent and Grogan 2009) are more likely to be encountered in Ontario based on larval habitat and are mentioned in reference to similar species.
Morphological identification is imperative for building DNA barcode reference libraries for species. Future directions should focus on a larger cladistics interpretation of the genus, as this key and many other sources are purely based on morphological taxonomy (Borkent 2012b). This key helps alleviate the taxonomic impediment of Canadian Culicoides, as it gives the first comprehensive compilation of all diagnostic information and distributions of female Culicoides that are found in Ontario.
Materials and Methods
Specimens
Many whole specimens used for reference images were collected in June and July of 2019 on the University of Guelph Campus, using UV light traps (UV LED CDC Trap with 6V12AH/20HR Rechargeable Valve Regulated Lead-Acid Battery, Bioquip Products Inc., California, USA). Light traps equipped with LEDs emitting ultraviolet light were used because they have been found to catch some species of Culicoides in a higher abundance and/or diversity (Nelder et al. 2010; Viennet et al. 2011; Vigil et al. 2014). Three collection events (19 June 2019, 9 July 2019, and 19 July 2019) were conducted by setting out traps ¾ full with a concentrated salt solution made by mixing 1 L of water, 0.4 kg of sodium chloride (salt), and 2 drops of unscented soap. Traps were set up in an agricultural area: the Ontario Veterinary College Dairy Barn in the cattle enclosure (43.528696, -80.230604), and in a natural area: the University of Guelph Arboretum (43.538184, -80.218447) in the late afternoon, and collected in the early morning of the next day. From these traps, 162 individual Culicoides specimens were collected and identified representing 12 species, one subgenus, and one species group (Appendix 1). Additional whole specimens were obtained from the CFIA National Centre of Animal Diseases, Lethbridge Laboratory (Lethbridge, Alberta, Canada) from pre-existing Canadian collections that were conducted during 2014–2016. Whole specimens were stored individually in 95% ethanol. Specimens from the 2019 summer collection were not slide mounted, as they were being stored for future molecular work. Slide-mounted specimens or images of slide-mounted specimens were obtained from the CFIA National Centre of Animal Diseases Lethbridge Laboratory, the Canadian National Collection of Insects, Arachnids and Nematodes, The Smithsonian’s National Museum of Natural History, The New York State Museum, the Southeastern Cooperative Wildlife Disease Study, and the Florida State Collection of Arthropods. Distribution maps for each species were made using the online tool SimpleMappr (Shorthouse 2010).
Identification
Culicoides that were collected in the summer of 2019 were identified to genus using the key to Ceratopogonidae found in Downes and Wirth (1981), and then were identified further down to subgenus, species group, or species (Appendix 1). Additional taxonomic keys reviewed for general information on Culicoides included Fox 1955, Hair et al. 1966, Wirth et al. 1985, Wirth and Jones 1956, Foote and Pratt 1954, Cochrane 1974, Hoffman 1925, Jamnback 1965, Williams 1955, Root and Hoffman 1937, and Wirth and Blanton 1969a. To identify the Culicoides haematopotus group (C. footei, C. haematopotus), we used Atchley and Wirth (1979). To identify the subgenus Avaritia (C. obsoletus, C. sanguisuga, C. chiopterus), we used Jamnback and Wirth (1963). To identify the guttipennis species group (C. guttipennis, C. villosipennis, C. flukei, C. arboricola), we used Wirth and Blanton (1967). To identify the pulicaris species group (C. yukonensis, C. paraimpunctatus) and cockerellii (C. cockerellii, C. frohnei), we used Wirth and Blanton (1969a). To identify the piliferus species group (C. alexanderi, C. bickleyi, C. denticulatus, C. downesi, C. franclemonti, C. jamnbacki, C. parapiliferus, C. piliferus, C. snowi, C. testudinalis, C. utowana), we used Wirth and Hubert (1962), Wirth and Blanton (1974), Cochrane (1974), and Jamnback (1965).
Imaging
All images in this publication were taken by authors L. Janke, S. Vigil, and T. Furukawa-Stoffer. For many of the images used in this publication, the following is the protocol used by the first author. Photographs for whole specimens were taken while they were submerged in 95% ethanol. The imaging program Leica LAS X (version 3.6.0.20104; Leica Microsystems, Wetzlar, Germany), the camera Leica MC170 HD (Leica Microsystems, Wetzlar, Germany), and the scope M205 A (Leica Microsystems, Wetzlar, Germany) were used for most pictures. Specimens with the least amount of wing damage and desiccation were selected for imaging. Pictures of six orientations were taken and used for each whole specimen: right lateral, left lateral, ventral, close-up of the head, close-up of external genitalia, and wings. Slide-mounted specimens were also imaged using the same microscope, and usually included a lateral, head, internal genitalia, and wing image.
Checklist
Table 1. Checklist of the Culicoides Latreille of Ontario, organized by Subgenus and then species group or section, if applicable.
Table 2. Checklist of the nine Culicoides Latreille species that are found in bordering Canadian provinces and/or United States, in alphabetical order.
| C. (Amossovia) beckae Wirth and Blanton, 1967 |
| C. (Beltranmyia) hollensis (Melander and Brues, 1903) |
| C. (Beltranmyia) bermudensis Williams, 1956 |
| C. (Oecacta) furens (Poey, 1853) |
| C. (Drymodesmyia) hinmani Khalaf, 1952 |
| C. (Avaritia) juddi Cochrane, 1973 |
| C. (Silvaticulicoides) loisae Jamnback, 1965 |
| C. (Avaritia) pechumani Cochrane, 1974 |
| C. scanloni Wirth and Hubert, 1962 (piliferus species group Root and Hoffman) |
Glossary/Characteristics
For the identification of the genus Culicoides using the key to Ceratopogonidae in Downes and Wirth (1981), these features must be visible on the entire specimen: the empodia, flagellomeres, eyes, thorax shape, and palpus (Fig. 1). Diagnostic characters for the wing are: r-m crossvein, radial cells (r1 & r2+3), medial veins (M1, M2), costal vein (C) (Fig. 2), as well as the macrotrichia and wing pattern of pale and dark spots (Fig. 3).
In using this key for identification to subgenus, species group, and species for female Culicoides, these diagnostic characteristics must be seen on the specimen: the tibiae, which may be uniformly brown or have pale bands (Fig. 1), a clear view of the wing veins and cells (Fig. 2), macrotrichia and wing patterns (Fig. 3), the 3rd palpal segment shape, size, and sensory pit (Figs. 4, 5), and in some cases, the shape and relative size of the spermatheca(e) (Fig. 6).
All images used in this publication are from females. However, Culicoides species exhibit sexual dimorphism in various morphological characteristics, including but not limited to: flagellomere shape and size, wing shape and pattern, and third palpal segment shape and size (Swanson 2012) (Fig. 10).
Terminology
Antennal sensoria: Hair-like sensory detectors found on the antenna.
Empodium/Empodia (pl): A small appendage between the claws on the tarsi.
Flagellomeres: Terminal segments of the antennae. Strictly speaking the antennomeres refer to the three basic parts of the antennae (scape, pedicel, flagellum), and the flagellomeres represent the more terminal antennomeres (Cumming and Wood 2017).
Macrotrichia: Small hairs on the wings.
Mesonotum: the dorsal surface of the thorax.
Palpus/Palpi (pl): A sensory structure for touching or tasting in the mouth area of insects.
Spermatheca/Spermathecae (pl): The internal genitalia of a female fly for sperm storage.
Stigma: Dark spot at apical area of the costal vein (C).
Wing cells: The areas enclosed by veins on the wings.
Diagnostic Characters
Figure 1. Lateral view of Culicoides biguttatus (female) showing important external diagnostic characteristics. The empodia cannot be seen clearly in this image.
Comparison of Features
A. Swollen and globular segment of Culicoides nanus. B. Moderately swollen, triangular segment of Culicoides sphagnumensis. C. Narrow, elongate segment of Culicoides variipennis. Black arrows indicate the sensory pit.
Wing Atlas
Click on the wing image to open the corresponding species information sheet within the key.
Amossovia Glukhova 1989
Avaritia Fox 1955
Beltranmyia Vargas, 1953
Culicoides Latreille, 1809
Diphaomyia Vargas 1960
Hoffmania Fox, 1948
Monoculicoides Khalaf, 1954
Oecacta Poey 1853
Silvaticulicoides Glukhova, 1977
Wirthomyia Vargas 1973
Unplaced subgenus
piliferus species group Root and Hoffman
saundersi species group
Unplaced species group/Misc. species
Identification Key
Acknowledgements
We would like to thank Y. Milián-García, M. Madden, R. Young, as well as many other personnel in the Hanner Lab at the University of Guelph for aiding with collections and support of this project. We would also like to thank J. Hutcheson for his guidance, organization, edits, and supervision throughout this whole project. We acknowledge the Ontario Veterinary College Dairy Barn and R. Leighton for allowing us to collect on their premises. We also acknowledge the University of Guelph Arboretum, C. Earley, and A. Fazekas for allowing us to collect on their premises. We thank the Canadian National Collection of Insects, Arachnids, and Nematodes, The Smithsonian’s National Museum of Natural History, The New York State Museum, and the Florida State Collection of Arthropods for loaning slide mounts of Culicoides species. We thank the personnel at the Ontario Agricultural College Insect Collection for their support and knowledge, especially S. Paiero. We thank L. Cohnstaedt at the USDA Arthropod-borne Animal Diseases Research, Manhattan, KS, USA for providing specimens to the CFIA. We thank the many researchers and volunteers who were involved in the Culicoides surveillance program with the CFIA that collected specimens from 2014–2016. We also thank A. Borkent for the distribution information of his collections of Culicoides, presented and paid for by Environment Canada. Finally, we thank D. Swanson, P. Shults, and an anonymous referee for their comments and feedback on a previous version of our manuscript. UoG-BIO acknowledges the financial support of the Canadian Food Inspection Agency.
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