The ecological role of mosquito larvae in aquatic environments

The ecological role of mosquito larvae in aquatic environments

The ecological role of mosquito larvae in aquatic environments

Assessing the community role and trophic interactions of Anopheles gambiae larvae in the Volta Region of Ghana.

IMAGE CREDIT: Michelle L. D’Souza

 The Anopheles gambiae mosquito complex, which consists of eight species, is the prevalent malaria vector in Sub-Saharan Africa1. While we have a reasonable understanding of predation rates, resource availability, and competition among aquatic invertebrate2-7, we lack a comprehensive appreciation of the larval ecology of An. gambiae, including their trophic role, diets, and population dynamics. My research aims to address this gap, crucial in integrating aquatic habitat management with vector control programs8.

With the advance of DNA technologies such as metabarcoding, we can now identify prey to a species level by examining the stomach contents of aquatic predators. When using these insights with network analyses, it is possible to quantify direct and indirect ecological interactions in the environment9. I will take advantage of these techniques for my research which aims to assess the trophic interactions of An. gambiae larvae in water bodies within two agricultural communities in Ghana.

I will collect mosquito larvae with a dipper, and a larger subset of invertebrates with an aquatic net (with a mesh size of 250 μm).  Each specimen’s gut contents will be analyzed using DNA barcoding and this data will be used to gain an understanding of the larval niche and ultimately An. gambiae’s role within the aquatic community. I will examine network interaction metrics such as connectance (the number of realized connections between species relative to what is available), degree (the number of interaction partners), betweenness (the importance of a species as a connector between different groups), and closeness (how central a focal species is in the community) to aid in niche construction. Temperature, pH, dissolved oxygen, salinity, and conductivity of the target water bodies will also be measured while sampling as these factors affect the occurrence and abundance of larvae by influencing the breeding behaviours of mosquitoes10.

In addition, I plan to study how different species of mosquito larvae compete for key resources. In the laboratory, the densities of local-caught larval populations of An. gambiae and other mosquito species will be manipulated to help determine the strongest competitor. While maintaining optimal rearing conditions by measuring the physio-chemical properties of the water daily, I will examine four indicators of overall growth and survival (the mean time to pupation, percentage of larvae that did not reach the adult stage, sex ratio, and mean female wing length) and therefore infer competitive strength among species. These data will provide important insights into predicting whether another mosquito species would dominate if the number of An. gambiae is reduced in the habitat.

An. gambiae is relatively small, constituting about half to one-third the mass of many Aedes mosquito species12. Foraging theory indicates that small, mobile insects of low profitability, do not form a preferred food source to predators unless they are massively clustered13. Though many species feed on mosquitos, these animals also feed on other small organisms that typically co-occur with An. gambiae14. For these reasons, I do not foresee that An. gambiae larvae will be a key food source for any predator in the aquatic environments in Ghana or that they will have a central role in local trophic systems.

If this is proven to be true, it will provide evidence that malaria-intervention methods that aim to suppress or reduce An. gambiae mosquitoes will not have detrimental consequences for the larger community.

Written by

Afia S. Karikari

Afia S. Karikari

African Regional Postgraduate Programme in Insect Science, University of Ghana, Accra, Ghana  

April 21, 2021

doi:10.21083/ibol.v11i1.6619 

This research is part of a larger effort by Target Malaria in Ghana to understand the role of the An. gambiae mosquito in the broader ecosystem.

For more information see:

The important interactions behind the itch

References:

  1. Service MW (1971) Studies on sampling larval populations of the Anopheles gambiae Bulletin of the World Health Organization 45:169–180.
  2. Service MW (1973) Mortalities of the larvae of the Anopheles gambiae Giles complex and detection of predators by the precipitin test. Bulletin of Entomological Research 62:359– 369.
  3. Service MW (1977) Mortalities of the immature stages of species of the Anopheles gambiae complex in Kenya: comparison between rice fields and temporary pools, identification of predators, and effects of insecticidal spraying. Journal of Medical Entomology 13:535–545.
  4. Ho BC, Ewert A, Chew LM (1989) Interspecific competition among Aedes Aegypti, albopictus and Ae. triseriatus (Diptera: Culicidae). Journal of Medical Entomology. 26:615–623.
  5. Barrera L (1996) Competition and resistance to starvation in larvae of container-inhabiting Aedes mosquitos. Ecological Entomology 21:117–127.
  6. Juliano SA, Lounibos LP and O’Meara GF (2004) A field test for competitive effects of Aedes albopictus on aegypti in south Florida: differences between sites and co-existence and exclusion? Oecologia. 139:583–593.
  7. Braks MAH, Honόrio NA, Lounibos LP, Lourenςo-de-Oliveira R, Juliano SA (2004) Interspecific competition between two invasive species of container mosquitos, Aedes aegypti and Aedes albopictus (Diptera: Culicidae), in Brazil. Annual Entomological Society of America 97:130–139.
  8. Li L, Bian L, Yakob L, Zhou U, Yan G (2009) Temporal and spatial stability of Anopheles gambiaelarval habitat distribution in western Kenya highlands. International Journal of Health Geographics. 8(70). doi:10.1186/1476-072X-8-70
  9. Fath BD, Patten BC (1998) Network synergism: emergence of positive relations in ecological systems. Ecological Modelling 107:127–143.
  10. Clements AN (1992) The Biology of Mosquitoes (Vol 1) Development, Nutrition and Reproduction. Chapman and Hall, London.
  11. Paajimans KP, Huijben S, Githeko AK, Takken W (2009) Competitive interactions between larvae of the malaria mosquitos, Anopheles arabiensis and Anopheles gambiae under semi-field conditions in western Kenya. Acta Tropica. 109:124–
  12. Koella JC, Lyimo EO (1996) Variability in the relationship between weight and wing length of Anopheles gambiae (Diptera: Culicidae). Journal of Medical Entomology 33: 261–264.
  13. Stephens DW, Brown JS, Ydenberg RC (2007) Foraging: Behavior and Ecology. University of Chicago Press, Chicago, IL.
  14. Findley JS, Black H (1983) Morphological and dietary structuring of a Zambian insectivorous bat community. Ecology 64:625–630.

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What we know, don’t know, and think we know about the predators of mosquitoes

What we know, don’t know, and think we know about the predators of mosquitoes

What we know, don’t know, and think we know about the predators of mosquitoes

Studying ecological relationships and community perceptions of Anopheles gambiae (Diptera: Culicidae) and their predators

IMAGE CREDIT: Michelle L. D’Souza

 The malaria-transmitting mosquito An. gambiae is eaten by species of bats, birds, damselflies, dragonflies, and spiders, although the mosquito likely accounts for only a small percentage of their total diet1. While there is limited data on the predation of adult An. gambiae at blood-feeding, nectar-feeding, and breeding sites, my research will use DNA metabarcoding techniques combined with ecological network analyses to determine the diet of predators and the community structure of predator-prey interactions.

Currently, we lack an understanding of indigenous perceptions surrounding An. gambiae. While Ghana’s local communities have expressed an interest in sharing their first-hand knowledge on various problems associated with mosquitoes, researchers often fail to consult them. Local peoples’ perceptions, knowledge, and experiences of the ecological relationship between predators and mosquitoes in the area where they live and farm provide important information to consider when addressing misconceptions that could impede malaria control interventions2.

My research will group mosquito activities into feeding (e.g., blood-feeding and nectar-feeding) and breeding (e.g., oviposition and eclosion) behaviours upon sampling specimens at three site types: (1) in homes both indoors and outdoors, (2) near vegetation and flowering plants closer to homes, and (3) around clear and shallow temporary water bodies.

I will sample invertebrates using both active and passive sampling traps including Malaise traps, pitfall traps, yellow pan traps, light traps, sweep nets, and aerial nets. Insectivorous invertebrates will be sorted out from the total trap catch and their diet composition determined by DNA metabarcoding to identify mosquitoes as well as other prey. I will also ascertain whether the diet of a predator changes with the sampling site, for example, where An. gambiae is feeding or depositing eggs. I will further determine how the predators are distributed across sampling sites, and in which of the sampling sites An. gambiae is most vulnerable to predation. I will analyze the invertebrate community coexisting with adult An. gambiae and the community structure of predator-prey interactions by using ecological network analysis, thus determining the role of An. gambiae in the food web.   

To assess indigenous knowledge and local perceptions on the ecological role of An. gambiae, I plan to distribute questionnaires that will gather basic information on mosquito identification, biology, and ecology as well as observations on feeding interactions and locations, and the predators involved. Questionnaire results will be compared with those of the metabarcoding results to determine the presence and extent of misconceptions regarding predatory interactions.

My goal is to assess the extent of mosquito knowledge in the community so that I can design appropriate education measures to ensure that future vector control projects are successful. The local communities’ knowledge and understanding of mosquito ecology is vital in these efforts.

This research will fill knowledge gaps regarding the role of An. gambiae in the local ecosystem and also help predict any possible ecological consequences of suppressing the malaria-transmitting mosquito as part of a vector control strategy.

Written by

Roger Sigismund Anderson

Roger Sigismund Anderson

African Regional Postgraduate Programme in Insect Science, University of Ghana, Ghana  

April 12, 2021

doi:10.21083/ibol.v11i1.6597

This research is part of a larger effort by Target Malaria in Ghana to understand the role of the An. gambiae mosquito in the broader ecosystem.

For more information see:

The important interactions behind the itch

References:

1. Collins CM, Bonds JAS, Quinlan MM, & Mumford JD (2019) Effects of the removal or reduction in density of the malaria mosquito, Anopheles gambiae s.l., on interacting predators and competitors in local ecosystems. Medical and Veterinary Entomology, 33(1), 1-15.

2. Ahorlu CK, Dunyo SK, Afari EA, Koram KA & Nkrumah FK (1997) Malaria‐related beliefs and behaviour in southern Ghana: implications for treatment, prevention and control. Trop Med Int Health 2, 488–499.

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Pollinators or nectar thieves? The role of malaria-transmitting mosquitoes in pollination

Pollinators or nectar thieves? The role of malaria-transmitting mosquitoes in pollination

Pollinators or nectar thieves? The role of malaria-transmitting mosquitoes in pollination

Establishing the impact of the Anopheles gambiae complex on fruit sets and their role in a natural pollinator community.

IMAGE CREDIT: Michelle L. D’Souza

Pollination is an important ecosystem service that is necessary for the reproduction of over 90% of the 250,000 species of modern vascular plants. The diversity of wild plants and food crops depend on the diversity of animal pollinators, hence a reduction and/or loss of either will affect the survival of both.

Due to their ecological and economical importance, studies on pollinator communities and conservation efforts have generally focused on bees (Hymenoptera), butterflies (Lepidoptera) and hoverflies (syrphid Diptera). However, there is a paucity of information on the potential influence of other flower-visiting insects (i.e., non-syrphid Diptera) on pollination. Non-syrphid Diptera are diverse, common, widely distributed, and among the most ubiquitous insect species in both natural and managed habitats. Hence, collectively, they may be more important pollinators than previously understood.

The malaria-transmitting mosquito complex Anopheles gambiae is not a known pollinator of any plant. Yet they visit flowers to feed on nectar, and thus have the potential to pollinate flowers. However, many malaria control strategies, including Target Malaria’s approach, target the vector, hence it is important to assess the impact of these mosquito species on pollination and seed sets, as well as to study their role in the pollination community network.

My research will provide a better understanding of the ecological role of An. gambiae using both observational and experimental studies. I will collect both male and female An. gambiae from their resting places in rooms, abandoned houses, beneath grasses, and on public toilets among others, within the study areas. Through DNA metabarcoding, I will identify pollen attached to each collected An. gambiae and use the results to determine the plants that are visited and potentially pollinated by An. gambiae.

To establish the impact of An. gambiae pollination on fruit set, I will raise several plants including those known to be visited by An. gambiae in a semi-field environment. I will designate the plants into three groups: In the first, An. gambiae will be introduced as the only pollinator; the second group will be opened to all pollinators and; the third group (i.e., control) will be kept away from all pollinators. I will use the number and quality of fruit sets and dry weight of fruit yield in each group to determine the impact of An. gambiae pollination on plant reproduction.

This research will identify potential consequences of new and existing methods to control the malaria vector, especially as current interventions typically rely on insecticide sprays and insecticide-treated bed nets that are less species-specific and therefore affect a broader range of species.

Written by

Thomas Gyimah

Thomas Gyimah

African Regional Post Graduate Programme in Insect Science, University of Ghana, Accra, Ghana

February 24, 2021

doi: 10.21083/ibol.v11i1.6540

This research is part of a larger effort by Target Malaria in Ghana to understand the role of the An. gambiae mosquito in the broader ecosystem.

For more information see:

The important interactions behind the itch

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Ecological and epidemiological insights from blood meals

Ecological and epidemiological insights from blood meals

Ecological and epidemiological insights from blood meals

Using interaction networks to explore the community structure and network ecology of biting insects and their hosts
PHOTO CREDIT: Ben Bellekom, IMAGE CREDIT: Michelle L. D’Souza

Knowledge of variations in insect biting behaviour and feeding patterns of insect disease vectors is important in understanding disease epidemiology and establishing effective control methods. Blood feeding insects are embedded within complex ecological communities and interact with other species acting as predator, prey, or competitor. These interactions are likely to impact the population dynamics of those co-occurring species1.

Biting insects exist within a variety of landscapes, where landscape change is likely to influence host-insect interactions2,3.  For example, deforestation within the Peruvian Amazon resulted in 278 times more human interactions with the malaria vector Anopheles darlingi than in undisturbed habitat, leading to a rise in malaria transmission4. Consequently, creating vector controls requires an understanding of how the landscape modifies biting insect community structure and biting interaction frequencies. Similarly, community structure, host availability, and the frequency of interactions may all be affected by time, again with consequences for disease transmission.  For example, the Leishmaniasis vector, sand flies, are most active at dawn and dusk, with a resultant increase in transmission risk to nocturnal hosts5.

Biting insect research has primarily focused on the analysis of single biting insect species or small sets of interacting species to examine host usage and to screen vectors for pathogens of interest (such as Zika virus and malaria). As many biting insects are medically important, research has also focused on the effects of specific control initiatives, such as examining Anopheles mosquito abundances following the implementation of long-lasting insecticidal nets. Whilst limited work has examined the effect of habitat modification and temporal variations on feeding patterns and species abundances, this work has broadly overlooked the community aspect6. Therefore, there is currently an incomplete understanding of the degree to which these variations and distance from human habitation7, impact interaction frequencies, and the structural properties of the wider biting insect-host community. Indeed, as insect communities themselves are rarely studied, the full ecological and epidemiological implications of their interactions is currently unknown, so that the design of control strategies is potentially compromised.

Understanding such interactions is possible by constructing interaction networks, which provide a visual and mathematical representation of a community of species, connected through their feeding interactions8,9. I will create biting insect-host interaction networks using data from insect bloodmeals, focusing on the communities of biting flies in Ghana. I will explore the impact of landscape, comparing distinct habitat categories (such as scrubland and fallow land), and temporal variations (day or night), and the effect of proximity to human habitation on biting insect-host community composition, structural properties, such as how specialised the network is, and relative species’ abundances. I will use metabarcoding to create data on species interactions as it allows for the identification of animal DNA in a mixed sample collected from biting insect bloodmeals10.

I have augmented this fieldwork with a literature-based analysis of published blood meal interaction data. This provides an in-depth opinion on the value of interaction networks in understanding host-insect-disease interactions and highlights their future applications in identifying and monitoring emerging diseases and unrecognized vectors11. Further, using a global data set extracted from the literature, I aim to examine structural variations of biting insect-host communities between distinct habitat types and latitudes. The results generated by my work will provide valuable insight into biting insect community ecology, evaluate the potential impact of interventions on community structure and interactions.

Written by

Ben Bellekom

Ben Bellekom

Department of Zoology, University of Oxford, Oxford, UK

February 24, 2021

doi:10.21083/ibol.v11i1.6539

This research is part of a larger effort by Target Malaria in Ghana to understand the role of the An. gambiae mosquito in the broader ecosystem.

For more information see:

The important interactions behind the itch

References:

1. Ferguson HM, Dornhaus A, Beeche A, Borgemeister C, Gottlieb M, Mulla MS, Gimnig J, Fish D, Kileen G (2010) Ecology: A prerequisite for malaria elimination and eradication. PLOS Medicine 7. doi:10.1371/journal.pmed.1000303

2. Wolinska J, King KC (2009) Environment can alter selection in host-parasite interactions. Trends in Parasitology 25: 236–244. doi:10.1016/j.pt.2009.02.004

3. Lachish S, Knowles S, Alves R, Sepil I, Davies A, Lee S, Wood M, Sheldon B (2013) Spatial determinants of infection risk in a multi-species avian malaria system. Ecography 36:587–598. doi:10.1111/j.1600-0587.2012.07801.x

4. Vittor AY, Gilman RH, Tielsch J, Glass G, Shields T, Lozano WS, Pinedo-Cancino V, Patz J (2006) The effect of deforestation on the human-biting rate of Anopheles darlingi, the primary vector of falciparum malaria in the Peruvian Amazon. American Journal of Tropical Medicine and Hygiene 74:3–11. doi:10.4269/ajtmh.2006.74.3

5. Aklilu E, Gebresilassie A, Yared S, Kindu M, Tekie H, Balkew M, Warburg A, Hailu A, Gebre-Michael T (2017) Comparative study on the nocturnal activity of phlebotomine sand flies in a highland and lowland foci of visceral leishmaniasis in north-western Ethiopia with special reference to Phlebotomus orientalis. Parasites & Vectors 10:393. doi:0.1186/s13071-017-2339-6

6. Braack L, Almeida P, Cornel A, Swanepoel R, de Jager C (2018) Mosquito-borne arboviruses of African origin: review of key viruses and vectors. Parasites & Vectors 11:29. doi:10.1186/s13071-017-2559-9

7. Orsborne, J, Furuya-Kanamori L, Jeffries C, Kristan M, Mohammed A, Afrane Y, O’Reilly K, Massad E, Drakeley C, Walker T, Yakob L (2019) Investigating the blood-host plasticity and dispersal of Anopheles coluzzii using a novel field-based methodology. Parasites & Vectors 12:143. doi:10.1186/s13071-019-3401-3

8. Kaiser-Bunbury C.N, Blüthgen N (2015) Integrating network ecology with applied conservation: a synthesis and guide to implementation. AoB Plants 7. doi:10.1093/aobpla/plv076

9. Proulx S.R, Promislow D, Phillips P (2005) Network thinking in ecology and evolution, Trends in Ecology & Evolution 20:345–353. https://doi.org/10.1016/j.tree.2005.04.004

10. Drinkwater R, Schnell I, Bohmann K, Bernard H, Veron G, Clare E, Gilbert T, Rossiter S (2019) Using metabarcoding to compare the suitability of two blood‐feeding leech species for sampling mammalian diversity in North Borneo, Molecular Ecology Resources 19:105–117. doi:10.1111/1755-0998.12943

11. Bellekom B, Hackett TD, Lewis OT (2021) A network perspective on the vectoring of human disease. Trends in Parasitology. doi: 10.1016/j.pt.2020.12.001

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The fly specimen that awaited a growing barcode community to be dusted off the shelves and given a name

The fly specimen that awaited a growing barcode community to be dusted off the shelves and given a name

The fly specimen that awaited a growing barcode community to be dusted off the shelves and given a name

The online barcode library facilitates the description of new Diptera genus with circumpolar Holarctic distribution

Reisa National Park, Norway. The type locality of a new fungus gnat species Coelosynapha loici.

PHOTO CREDIT: J. Kjærandsen

While it can take an average of 21 years between the discovery and description of a new species1, the challenges of relating known taxa and the scarcity of high-quality specimens make the description of a new genus even more difficult and time-consuming. Our paper recently published in the Biodiversity Data Journal introduced a new fungus gnat genus–Coelosynapha–demonstrating that despite a taxon being shelved for a long time after discovery, strong collaborative networks spanning many countries combined with the power of DNA barcoding are greatly changing the pace at which we catalogue life.

This story begins some 25 years ago when the late French entomologist Loïc Matile (1938–2000) sent Geir E.E. Søli (at the Natural History Museum in Oslo, Norway) illustrations and a brief one-page description of a potential new fly genus, belonging to fungus gnats of the family Mycetophilidae, based on a single specimen collected in Finland. Matile posed a seeming simple question to Søli, “Had this for years…What do you think?” Unable to do much with the scarce material, the specimen was shelved, twice, for even after more specimens were found in Finland in 2009 and sent to a specialist in the USA, no progress was made.

Growth of research ties and a reference library

During this time, the popular and widespread use of DNA barcoding has led to an accumulation of a substantial volume of sequenced insects on the Barcode of Life Data (BOLD) system. More than 65,000 specimens belonging to the fly family Mycetophilidae have been successfully sequenced. Some 1,100 of them have public barcodes, although more than 2,400 different Barcode Index Numbers (BINs) are assigned, which indicates that the majority of the species still remain unidentified beyond the (sub)family level.

In the Nordic region, strong scientific research ties grew during the early 2000s between the Swedish, Norwegian, and Finnish biodiversity information centres, taxonomy initiatives, and the Norwegian Barcode of Life (NorBOL) and Finnish Barcode of Life (FinBOL). These alliances are ensuring that the best taxonomic expertise is building up data in the reference library for local fauna on BOLD. Hence, the vast majority of some 6,500 DNA barcoded fungus gnats from the Nordic region have been identified to species level upon submission. The reference library is then painstakingly quality-checked and curated after barcodes and BINs are assigned. This has resulted in a high-quality reference library, now covering about 90 percent of up to 1,000 known Nordic species of the family.

The reference library gives us entirely new opportunities through machine identification of insect samples. Examples range from expanded taxonomic studies on the larger Holarctic fauna as in this study, to ecological studies on how fungus gnats function in our boreal ecosystems, to more applied stakeholder science ranging from management of unprotected and protected areas to monitoring insect populations and the often claimed decrease in insect diversity caused by disturbances like commercial land use, pesticides from agriculture, and climate change.

Holotype of Coelosynapha loici sp. n. (left) and Coelosynapha heberti sp. n. (right).
PHOTO CREDIT: J. Kjærandsen (left) & CBG Photography Group (right)

A new genus is born from treasures on BOLD

Continuing the exploration of fungus gnats, we obtained more specimens of the enigmatic new species first studied by Matile way back in the mid-1980s from several, mainly old-growth, coniferous sites across the Palaearctic Taiga; ranging from Norway in the west all the way to Chukotka in the Far East of Russia. Specimens of the new taxon from Russia and Fennoscandia (a region covering the Scandinavian Peninsula, Finland, Karelia, and the Kola Peninsula) were submitted for barcoding. We were surprised to find that the BINs assigned on BOLD indicated these specimens were the nearest neighbour to another unidentified, and very similar species sampled across southern Canada between 2004–2014 during the Centre for Biodiversity Genomics’ early efforts to barcode the insects of Canada2

Both species were assigned to a new genus named Coelosynapha. The first, Eurasian Coelosynapha loici, is named in honour of Loïc Matile. The second, North American Coelosynapha heberti, is named in honour of Paul D. N. Hebert, “the father” of DNA barcoding who led efforts to barcode the insects of Canada and currently leads the International Barcode of Life (iBOL) as the organization’s scientific director. Taken together it marks a celebration of the synergy emerging from traditional morphologically based taxonomy meeting a new integrative taxonomy including DNA barcodes in its toolbox. 

The newly described genus belongs to the subfamily Gnoristinae which appears to be amongst the most difficult branches of the Mycetophilidae to classify, which certainly added to the prolonged shelf life.

It marks a celebration of the synergy emerging from traditional morphologically based taxonomy meeting a new integrative taxonomy including DNA barcodes in its toolbox.

Highly variable taxa have led to numerous small genera with few species being segregated, as well as species-rich, polyphyletic genera sometimes called “trash bin” genera because they are derived from more than one common evolutionary ancestor or ancestral group and are therefore not suitable to be placed in the same taxon.

Morphologically Coelosynapha is most similar to the genera Coelosia and Synapha, hence its name, while genetically, species of these genera appear rather distant. As the new species epithets suggest, there is a need for more integrative taxonomic studies combining classical morphology with DNA barcoding.

The BOLD archive certainly hides many similar treasures waiting to be uncovered, but for that to happen morphological expertise needs to be invoked. Through our description of Coelosynapha, we hope to inspire this kind of integrative taxonomic work on the species-rich family of fungus gnats and aspire to further phylogenetic studies of the intriguing subfamily Gnoristinae.

References:

1. Fontaine B, Perrard A, Bouchet P (2012) 21 years of shelf life between discovery and description of new species. Current Biology 22 (22). doi: 10.1016/j.cub.2012.10.029

2. Hebert PN, Ratnasingham S, Zakharov E, Telfer A, Levesque-Beaudin V, Milton M, Pedersen S, Jannetta P, deWaard J (2016) Counting animal species with DNA barcodes: Canadian insects. Philosophical Transactions of the Royal Society B: Biological Sciences 371 (1702). doi: 10.1098/rstb.2015.0333

Written by

Jostein Kjærandsen

Jostein Kjærandsen

Tromsø University Museum, UiT – The Arctic University of Norway Tromsø, Norway

Alexei Polevoi

Alexei Polevoi

Forest Research Institute of Karelian Research Centre of the Russian Academy of Sciences Petrozavodsk, Russia

Jukka Salmela

Jukka Salmela

Regional Museum of Lapland & Arctic Centre, University of Lapland Rovaniemi, Finland
December 4, 2020

doi: 10.21083/ibol.v10i1.6401

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The butterfly effect: geographic patterns of DNA barcode variation in subtropical Lepidoptera

The butterfly effect: geographic patterns of DNA barcode variation in subtropical Lepidoptera

The butterfly effect: geographic patterns of DNA barcode variation in subtropical Lepidoptera

Forest dynamics, spatial distribution patterns, and sampling scale are associated with mitochondrial DNA variation in Argentinian butterflies

Andean forests in northwestern Argentina.
PHOTO CREDIT: Ezequiel Núñez Bustos

Argentina harbours more than 1,200 species of butterflies, most of them found in two biodiversity hotspots and priority areas for conservation: the Atlantic Forest and the Andean forests1.

Figure 1: Sampling localities in northwestern Argentina (NWA, black squares) and northeastern Argentina (NEA, white triangles)2. The Atlantic Forest (dark blue) extends along the Brazilian coast and reaches its southernmost portion in NEA, while the Central Andean forests (red) descend from southern Peru and reach NWA. The distribution of eight other ecoregions indicated.

Despite their current isolation (Figure 1), these two areas have been cyclically and transiently connected in the past, promoting the interchange of flora and fauna, and resulting in a pattern of disjunctly co-distributed taxa. While the historical relationship between these allopatric forests and its evolutionary effects on shared fauna has been the subject of recent (and ongoing) research, studies have been concentrated mostly on vertebrates.

This study explores the butterflies of the Atlantic Forest and the Andean forests providing new insights into both the diversification patterns in southern South America and the impact of increasing the geographic and taxonomic scale of sampling on DNA barcoding performance in the region.

Atlantic Forest in northeastern Argentina. PHOTO CREDIT: Ezequiel Núñez Bustos

In 2017, we assembled and analyzed a DNA barcode reference library for 417 species from northeastern Argentina (NEA)2, focusing on the Atlantic Forest and covering around one-third of the butterfly fauna of the country. To expand the geographic and taxonomic distribution of this library, we generated DNA barcodes for 213 butterfly species from northwestern Argentina (NWA) with a focus on the Andean forests.

We then used these libraries to examine three themes, outlined below.

1.The effectiveness of DNA barcodes for species discrimination and identification

 

The mean intraspecific distance for the butterflies of NWA was 0.29%, while mean interspecific distance among congeneric species was 7.24% (Figure 2). More importantly, mean distance to the nearest neighbour (7.56%) was nearly 13 times larger than the mean distance to the furthest conspecific (0.60%), resulting in a distinct barcode gap for all but two species represented by two or more individuals (Figure 2).

Genetic distance or sequence variation in the COI sequences within and between species was estimated using the Kimura-2-parameter (K2P) model of nucleotide substitution

Substitution models describe the process of genetic variation through fixed mutations, constituting the foundation of evolutionary analysis at the molecular level.

Arenas M (2015) Trends in substitution models of molecular evolution. Frontiers in Genetics 6(319). 

Figure 2: Frequency histogram of COI sequence distances within species (orange) and among congeneric species (blue) of butterflies in NWA. The inset graph shows the barcode gap analysis for species represented by two or more COI sequences, where each dot represents a specimen. Red dots correspond to individuals with a maximum intraspecific distance higher than the distance to the nearest heterospecific. The vertical dashed line shows the 95th percentile of all intraspecific distances (2.02%), while the horizontal line corresponds to the lower 5% of all congeneric distances (3.36%).

Consistently, sequence-based specimen identification simulations showed that this library is extremely effective in the identification of the butterflies of NWA, exceeding a 98% success rate regardless of the identification criteria implemented.

We then used different clustering algorithms to assess the presence of cryptic species. Overall, these methods generated between 1.4–9.9% more Molecular Operational Taxonomic Units (MOTUs) than the number of reference species, suggesting that the butterfly diversity of NWA might be higher than currently recognized.

Figure 3: Taxonomic coverage of the complete DNA barcode reference library for the butterflies of Argentina. Dark shading indicates the proportion of species covered within each family based on the total known for the country.

Merging the NWA and NEA databases resulted in a DNA barcode reference library for nearly 500 butterfly species, covering ~40% of the butterfly fauna of Argentina (Figure 3) and representing 549 barcode clusters (BINs) on BOLD (170 of which are new to the platform).

2.The impact of increasing the spatial and taxonomic coverage on DNA barcoding performance

When we compared the two reference libraries, we found that the barcode gap was significantly narrower in the NEA than in the NWA library (Figure 4). This is most likely associated with the higher geographic and taxonomic coverage of the former, since expanding the spatial scale of sampling is expected to not only increase intraspecific variation as a result of isolation by distance but also reduce interspecific divergences as more closely related species appear.

Figure 4: Maximum intraspecific distance (blue) and minimum interspecific distance (red) for the three datasets. Note the different scales.

When we tried to identify specimens from NWA by using the reference library of NEA, a considerably high proportion of individuals representing shared species between these regions could not be identified or resulted in an ambiguous identification, even when we allowed a maximum intraspecific distance of as high as 2% in the identification procedure. This was due to the existence of deep intraspecific divergences between conspecifics from northeastern and northwestern Argentina, two regions separated on average by more than 1,000 km.

At the same time, however, we observed that the effect of increasing the geographic (and taxonomic) scale was more profound on the minimum interspecific distances than on the maximum intraspecific distances. Therefore, it is possible that butterfly species in NEA are also naturally more variable than in NWA based on our current sampling. While specimens from NWA came almost exclusively from the montane forest on the east slope of the Andes, the sampling in NEA covered not only larger geographic distances but also a more heterogeneous landscape, characterized by the existence of different ecoregions (Figure 1) and physical barriers such as river, specifically the Paraná-Paraguay River axis. Regardless, our results show that both large geographic distances and increased taxonomic coverage can affect DNA barcoding identification performance, especially when using a local library to identify the fauna from another distant region.

As expected, the maximum intraspecific distance was significantly higher and minimum interspecific distance was significantly lower in the complete database (NEA + NWA) than within the NWA and NEA libraries alone (Figure 4). However, the logical and anticipated reduction in the barcode gap did not have, in this case, a significant impact on the identification performance of DNA barcodes, which were able to correctly identify ~99% of the individuals. This reflects the importance of increasing the spatial and taxonomic coverage of DNA barcode libraries to improve identification success, and of considering the use of a local database to identify regional fauna when a more comprehensive COI database is not available.

Doxocopa cyane burmeisteri
Doxocopa cyane burmeisteri
Parides erithalion erlaces

Parides erithalion erlaces

Pteronymia ozia tanampaya

Pteronymia ozia tanampaya

Butterfly species from the Andean forests. PHOTO CREDIT: Ezequiel Núñez Bustos

3.Geographic patterns of intraspecific variation across Argentina

A total of 135 butterfly species are shared between the databases of NEA and NWA. Mean intraspecific distance for these species was significantly higher between regions (1.02%) than within them (NEA, mean 0.35%; NWA, mean 0.33%), especially for a subset of 43 species that showed particularly deeper distance (mean 2.43%) between NEA and NWA.

We then focused only on the 85 species that are present in both the Atlantic Forest and the Andean forests (Figure 5), 27 of which have a disjunct distribution between forests, being absent from intermediate ecoregions, while the remaining 57 have a continuous range across northern Argentina.

Figure 5: Proportion of shared species between NEA and NWA that occur in both forests. The spatial distribution pattern (disjunct vs continuous) and the percentage of species with a deep intraspecific divergence between forest populations indicated.

We found that mean intraspecific distance between forest populations was significantly higher for the disjunctly distributed species (1.65%) than for species with continuous ranges (0.78%), showing that spatial distribution patterns have an influence on the level of intraspecific variation. Moreover, the proportion of species showing the deep divergence between populations from the Atlantic Forest and the Andean forests was notably higher among species with fragmented distributions (nearly 50%) than for species with continuous ranges (less than 30%) (Figure 5).

Lastly, based on standard molecular rates and COI sequence divergence, all diversification events between forest populations were dated to the last two million years, a time period when the currently isolated Atlantic Forest and Andean forests experienced multiple transient connections across the open vegetation corridor, a diagonal of more open and drier savanna-like environments (Caatinga, Cerrado and Chaco) that isolates the Atlantic Forest from the Andean forests (and the adjacent Amazonia) (Figure 1). These past connections were promoted mainly by Pleistocene climatic changes and habitat shifts.

Catonephele numilia neogermanica

Catonephele numilia neogermanica

Callicore hydaspes

Callicore hydaspes

Doxocopa agathina vacuna

Doxocopa agathina vacuna

Butterfly species from the Atlantic Forest.
PHOTO CREDIT: Ezequiel Núñez Bustos

Conclusions

Our study has not only expanded the DNA barcode reference library for the butterflies of Argentina, but it also constitutes, to our knowledge, the first multi-species assessment of the historical relationship between the currently isolated Atlantic Forest and Andean forests using butterfly species as model organisms.

Importantly, our research supports the fact that, even in the era of genomic data, large-scale analyses of mitochondrial DNA variation are still extremely useful for evolutionary studies, as they unveil spatial diversification patterns and highlight cases that deserve further investigation.

ACKNOWLEDGEMENTS:

We thank our colleagues from the Museo Argentino de Ciencias Naturales and the staff at the Centre for Biodiversity Genomics (CBG) for their help during different stages of this ongoing investigation. We also thank Michelle D’Souza for her helpful comments and suggestions that improved this contribution. This project is supported by Richard Lounsbery Foundation, the CBG, the Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET), the Agencia Nacional de Promoción de la Investigación, el Desarrollo Tecnológico y la Innovación, Fundación Williams, Fundación Bosques Nativos Argentinos and Fundación Temaiken. For granting the permits and transit guides, we thank the Offices of Fauna of the Argentinian provinces in which fieldwork was conducted, the Administración de Parques Nacionales, and the Ministerio de Ambiente y Desarrollo Sostenible from Argentina.

References:

1. Klimaitis J, Núñez Bustos E, Klimaitis C, Güller R (2018) Mariposas-Butterflies-Argentina. Guía de Identificación-Identification Guide. Vazquez Mazzini Editores. Buenos Aires. pp. 327.

2. Lavinia P, Núñez Bustos E, Kopuchian C, Lijtmaer D, García N, Hebert P, Tubaro P (2017) Barcoding the butterflies of southern South America: Species delimitation efficacy, cryptic diversity and geographic patterns of divergence. PLOS ONE 12(10), e0186845. https://dx.doi.org/10.1371/journal.pone.0186845

Written by

Natalí Attiná

Natalí Attiná

Ezequiel Núñez Bustos

Ezequiel Núñez Bustos

Darío A. Lijtmaer

Darío A. Lijtmaer

Pablo L. Tubaro

Pablo L. Tubaro

Pablo D. Lavinia

Pablo D. Lavinia

Museo Argentino de Ciencias Naturales “Bernardino Rivadavia” (MACN–CONICET)

July 31, 2020

doi: 10.21083/ibol.v10i1.6256  

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Hunting for a water mite neotype in southern Norway

Hunting for a water mite neotype in southern Norway

Hunting for a water mite neotype in southern Norway

Scientists rediscover lost specimens of water mite in Norway 120 years after they were first described

A stream near the church of Vanse at Lista in southern Norwaythe type locality of Lebertia porosa Thor, 1900.

PHOTO CREDIT: Torbjørn Ekrem

Did you know that scientists can assess natural water quality by monitoring the diversity of aquatic invertebrates? Freshwater insect and arachnid populations are often important indicators of environmental change. This is evident in particularly species-rich groups, such as water mites and biting or non-biting midges, which have great potential for monitoring water quality. The problem is only that they are too difficult and time consuming to identify in routine water quality assessments. This hurdle can be overcome with DNA metabarcoding, but only if a good reference barcode library is available.

Elisabeth Stur of the Norwegian University of Science and Technology (NTNU) University Museum, along with her team, have been doing summer fieldwork for the Water Mites and Midges in southern Norway (Water M&M) project. One of the many goals for this year’s fieldwork was not only to contribute to the reference barcode library, but also to sample the type locality of the water mite Lebertia porosa, described 120 years ago by Sig Thor, a Norwegian priest and acarologist.

The Great Lakes
Phaenopsectra flavipes (Diptera: Chironomidae) with water mite larvae attached. PHOTO CREDIT: Aina Mærk Aspaas, NTNU University Museum

Barcode data indicate that there are at least six cryptic genetic lineages within this species, but it is unknown which of these applies to the nominal species. Since the original type material is lost, re-sampling L. porosa from its type locality is important in designating a neotype that most likely belongs to the species described by Thor in 1900. This way, researchers can stabilize the definition of the L. porosa species name, such that potential new species could be described. This species delineation is part of a MSc. project by Valentina Tyukosova at NTNU: Integrative taxonomy and species delimitation in the Lebertia porosa species complex (Acari, Parasitengona: Hydrachnidia).

The type locality of L. porosa was vaguely described in Thor’s original publication as a “stream near the church of Vanse”. After studying maps of the surrounding area, researchers learned that this church still stands, and were able to locate two nearby streams.

Now they wondered, would these streams still be in good condition 120 years later? As the team of researchers approached what they thought might be the stream in June 2020, they were pleased to see running, clear water under the bridge. Next mystery: could the streams hold a population of L. porosa 120 years after first collection? They found that yes, the waters could, and the water mite populations were bountiful!

The Great Lakes DNA Barcoding Project team

Water mites from the type locality of Lebertia porosa Thor, 1900.
PHOTO CREDIT: Torbjørn Ekrem

Stur and her team are now looking forward to getting these critters under the compound microscope. Using DNA analysis, they hope to identify which barcode clusters they match with, potentially revealing the nominal species of L. porosa. We’re sure that Sig Thor would be thrilled to learn that his identified species is still thriving, 120 years later.

Written by

Katherine Perry

Katherine Perry

Centre for Biodiversity Genomics, Guelph, ON, Canada

July 24, 2020

doi: 10.21083/ibol.v10i1.6243

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GBOL III: Dark Taxa

GBOL III: Dark Taxa

GBOL III: Dark Taxa

Researchers launch new BIOSCAN project that aims to illuminate thousands of new insect species on Germany's doorstep.

A “pixelated” Diptera.

Currently, around 1.4 million species of animals are known. For tropical regions, many species are still unknown, with estimates of global biodiversity ranging from five to 30 or even 100 million species. More recent studies suggest that there are about 10 million species on our planet. In contrast to the tropics, the Central European fauna is considered to be very well studied. However, specialists have mostly concentrated on less diverse and easy-to-study organisms, neglecting the species-rich, often taxonomically difficult groups, like many Diptera and Hymenoptera. This led to a mismatch between high species numbers and a small number of researchers, often referred to as the ‘taxonomic impediment’. This is most prominent for the megadiverse faunas of tropical regions. Less known is that this also applies, to some extent, for countries with a long history of taxonomic research like Germany, covering 200 or more years. For example, for the compilation of the German checklist of Hymenoptera, 32 specialists were available for 247 species of digger wasps (Crabronidae), while for parasitoid wasps of the family Ichneumonidae one specialist had to deal with 3,332 species.

In Germany, about 48,000 species of animals have been documented, including about 33,300 species of insects. In little-studied groups such as insects and arachnids, preliminary results of earlier DNA barcoding initiatives indicate the presence of thousands of species that are still awaiting discovery. Among the groups with a particularly large suspected number of unknown species are the Diptera (flies) and the Hymenoptera (in particular, the parasitoid wasps). With almost 10,000 known species each, these two insect orders account for two-thirds of the German insect fauna, underlining their importance.

Bar Graph depicting availability of taxonomic expertise for major insects orders in Germany.

“Dark taxa” are, as a rule, small-sized and rich in species, and have therefore been largely ignored by taxonomists. This is reflected by the number of undescribed species in these taxa, combined with a low chance to get specimens identified by specialists.

The insight that there are not only a few but many unknown species in Germany is a result of the earlier German Barcode of Life projects GBOL I and II, both supported by the Federal Ministry of Education and Research (Bundesministerium für Bildung und Forschung, BMBF) and the Bavarian Ministry of Science (project Barcoding Fauna Bavarica). The projects aimed at making all German species reliably, quickly and inexpensively identifiable by DNA barcodes. Since the first project was launched about ten years ago, more than 25,000 animal species have been barcoded, in collaboration with national and international partners. Among them are mostly well-known groups such as butterflies, moths, beetles, grasshoppers, spiders, bees and wasps.

Two scatterplots demonstrating relationship between body size (top) and species richness (bottom) in German Diptera

Relationship between body size (top) and species richness (bottom) in German Diptera1

Despite their popularity, these groups represent only a fraction of the total inventory of German insects. In Germany there are 170 butterfly species, 81 dragonfly and damselfly species, 87 species of grasshoppers, katydids and crickets, and 580 species of ground beetles, all of which are well-studied. Taken together, these 918 species stand for only a small fraction (2.8%) of the German insect fauna. They are morphologically well identifiable, have manageable species numbers, can easily be monitored during daytime and are therefore regarded as relevant in nature conservation and often used for monitoring species diversity. Conversely, however, this means that the vast majority of the native species diversity has been largely ignored in nature conservation and in general and applied research.

Circular tree depicting Nematocera (midges) and Brachycera (flies)
A circular neighbour‐joining tree for the two suborders of flies, Nematocera and Brachycera1. Each line in the tree corresponds to a distinct Barcode Index Number (BIN). Whereas for two of the “big four” insect orders, the Lepidoptera and Coleoptera, the number of German species are very precisely known, the numbers for the Diptera and Hymenoptera must rely on rough estimates. 

This applies in particular to the Diptera (flies). The observation that estimates of the number of species of native Diptera have been far too low was not only a result of the DNA barcoding projects at the ZSM, but became clear in a recent study by Paul Hebert and his team2. In this large-scale study, DNA barcodes of about one million insects were analyzed. Based on this study, Canada’s gall midges alone are estimated to include about 16,000 species, suggesting the existence of at least two million species on earth. That would be more species of gall midges worldwide than all previously described animal species combined.

The little-known or unknown species, referred to as ‘dark taxa’, are the subject of another BMBF-funded DNA barcoding project that is being carried out at the ZSM in collaboration with other German natural history museums and institutions. The project focuses on Diptera and Hymenoptera (in particular, parasitoid wasps), each with a large proportion of ‘dark taxa’. The new project, funded by a grant of 5.3 million Euro, starts July 1st 2020, with 12 PhD students at three major natural history institutions in Bonn (Zoological Research Museum Alexander Koenig), Munich (SNSB – Zoologische Staatssammlung München) and Stuttgart (State Museum of Natural History Stuttgart), to address a range of questions related to the taxonomy of German ‘dark taxa’, targeting selected groups of Diptera and parasitoid Hymenoptera.

Detailed photo of a Eulophidae specimen
Yellow Mymaridae specimen

Small parasitoid wasps of the families Eulophidae (top) and Mymaridae (bottom), both group with possibly hundreds of new species in Germany that still await discovery.

Among the major aims of GBOL III is assessing of the performance of DNA barcoding for species identification of ‘dark taxa’, and assessing the species detection ability of DNA barcodes in mass samples that are obtained from metabarcoding studies. Other aims of the project include the development of a platform for managing OTU-based taxonomic data, developing a pipeline for reliable and fast barcoding of small and poor-quality samples, and training of the next generation of taxonomists.

GBOL III is designed to make an important contribution to the global BIOSCAN initiative of the Centre for Biodiversity Genomics. It helps to lay the foundations for a global biomonitoring system to record the biodiversity of our planet on a large geographical scale in times of rising temperatures, increasing weather extremes and receding ice, and to track its changes as a result of global environmental changes.

References:

1. Morinière J, Balke M, Doczkal D, Geiger MF, Hardulak LA, Haszprunar G, Hausmann A, Hendrich L, Regalado L, Rulik B, Schmidt S, Wägele J, Hebert PDN (2019) A DNA barcode library for 5,200 German flies and midges (Insecta: Diptera) and its implications for metabarcoding‐based biomonitoring. Molecular Ecology Resources 19: 900–928. https://doi.org/10.1111/1755-0998.13022

2. Hebert PDN, Ratnasingham S, Zakharov EV, Telfer AC, Levesque-Beaudin V, Milton MA, Pedersen S, Jannetta P, deWaard JR (2016) Counting animal species with DNA barcodes: Canadian insects. Philosophical Transactions of the Royal Society B: Biological Sciences 371: 20150333. https://doi.org/10.1098/rstb.2015.0333

Written by

Axel Hausmann

Axel Hausmann

SNSB - Zoologische Staatssammlung München, Munich, Germany

Lars Krogmann

Lars Krogmann

State Museum of Natural History Stuttgart, Stuttgart, Germany

Ralph S. Peters

Ralph S. Peters

Zoological Research Museum Alexander Koenig, Bonn, Germany

Vera Rduch

Vera Rduch

Zoological Research Museum Alexander Koenig, Bonn, Germany

Stefan Schmidt

Stefan Schmidt

SNSB - Zoologische Staatssammlung München, Munich, Germany

July 10, 2020

doi: 10.21083/ibol.v10i1.6242

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