Among mayflies, Baetidae are often considered as easy to recognise at the family level, but difficult to identify at lower level. In several faunistic or ecological studies, the identification remains at the family level; Baetidae are generally considered as widespread and ubiquitous, therefore as poorly informative for ecological studies or bioassessments. Here, a straightforward identification key is offered to larvae of the ten genera of Baetidae reported from Maghreb based on easily observable and understandable characters. The diversity, ecology, and distribution of each taxonomic unit (genera or subgenera) are discussed and the main difficulties for deeper identification are pointed out. Future challenges and remaining taxonomic riddles for Maghrebian Baetidae are detailed.

Algeria, aquatic insects, identification key, mayflies, Morocco, Tunisia

Ephemeroptera (mayflies) is a small order of insects with approximately 3700 species. Baetidae are the most diversified family as they encompass approximately one third of generic and specific mayfly diversity (Jacobus et al. 2019). The family is almost worldwide distributed but is mostly diversified in the tropics (Gattolliat and Nieto 2009). Mayflies are merolimnic insects, the larval stage is strictly linked to freshwater habitats while the winged stages are aerial. Imaginal stages are extremely brief and have no functional mouthparts and digestive system. Mayflies are the only insects having an intermediate winged stage between larva and imago, called subimago (Barber-James et al. 2008; Sartori and Brittain 2015).

Mayflies are widely used to assess freshwater quality and global changes in hydrosystems. They are generally very abundant, sensitive to environmental alterations, sufficiently diversified and can be considered as efficient bioindicators if identified at a relevant systematic level (Jacobus et al. 2019). Most Mediterranean rivers and streams, and especially Maghrebian ones, suffer from several threats directly or indirectly linked to human activities. Water abstraction for agriculture and domestic use, water pollution and eutrophication, dam construction and other water regulation, in addition to climate change, have direct severe negative impact on the river ecosystem and on aquatic community composition (Hafiane et al. 2016; Morghad et al. 2019; Zerrouk et al. 2021).

The term Maghreb (Arabic for "the west") refers to the countries of western North Africa. In its traditional sense, the Maghreb includes Morocco, Algeria, and Tunisia. The Maghreb, a biogeographic unit, is distinct from the "Greater Maghreb" or "Great Maghreb", a political and historical entity that additionally includes Libya and Mauritania. As no data and materials are available for Libya, and data is limited to a single short checklist for Mauritania (Fraser 1952), we refrain from including these countries in our study. However, we can assume that they have a very impoverished fauna, mainly covered by our study. Despite important improvements in the last decades, the knowledge of the mayfly fauna of the Maghreb is still incomplete. Historically, Eaton (1899) was the first to establish a list of mayflies for a Maghrebian country. He reported thirteen species from Algeria, including six species of Baetidae, one of them being new to science (Eaton 1899). For one century, little attention was paid to this fauna (Lestage 1925; Navás 1929; Kimmins 1938; Grandi 1951; Verrier 1952), till Thomas and collaborators gave a new impulse to the study of this fauna. They described new species and provided new reports in almost all families. In addition, Thomas (1998) provided a provisional checklist of the mayflies from the Maghreb including 69 species: 41 from Morocco, 50 from Algeria, and 29 from Tunisia. He listed 17 species of Baetidae and considered the report of nine additional species as needing to be confirmed. The checklist was updated by various subsequent contributions including description of new taxa and new reports (see below for the complete reference per country).

The Moroccan mayflies remained practically unknown until the 1970s, since only a few reports were available: five species inventoried by Lestage (1925), then seven other species listed by Navás (1929) and Kimmins (1938). The first faunistic inventory dedicated to this group was carried out by Dakki and El Agbani (1983) who were able to identify 26 species of Ephemeroptera, distributed in the different Moroccan regions. This list was greatly enriched subsequently through hydrobiological studies carried out on various Moroccan streams and rivers (Dakki 1986, 1987; Ouahsine and Lavandier 1988a, b; Qninba et al. 1988; El Agbani et al. 1992; El Alami and Dakki 1998; El Alami et al. 2000; El Bazi et al. 2017; Khadri et al. 2017; Mabrouki et al. 2017; Guellaf et al. 2021). Species new for science were also described (Dakki and Giudicelli 1980; Peters 1980; Thomas and Mohati 1985; Dakki and Thomas 1986; Sartori and Thomas 1986; Thomas and Bouzidi 1986; Thomas et al. 1987, 1992; Thomas and Vitte 1988; Vitte and Thomas 1988a, b; Vitte 1991), but only a few of them were on Baetidae (Thomas and Bouzidi 1986; Thomas et al. 1992). Thus, after the synthesis of these works, Thomas (1998) was able to list 41 species distributed in the different Moroccan massifs. Subsequent studies enabled the discovery of nine additional new species or new reports for Morocco; the specific richness for this country reaches 50 species, half of them being Baetidae (Alba-Tercedor and El Alami 1999; El Alami et al. 2000; Mabrouki et al. 2017; Zerrouk et al. 2021). A complete checklist including the diversity and distribution of all Moroccan mayflies was recently published (El Alami et al. 2022a). In order to assess the impact of climate change, human disturbances and pollution on aquatic macroinvertebrates, studies have been carried out over the different geographical Moroccan areas; the main goal was to evaluate the evolution of the mayfly community between the 1980s and the present days by prospecting selected stations in Haut Sebou (Zerrouk et al. 2021), Moulouya (Mabrouki et al. 2017), Ourika (Zuedzang Abessolo et al. 2021) and Rifian watersheds (El Bazi et al. 2017; Khadri et al. 2017; Guellaf et al. 2021).These recent studies of the main Moroccan watersheds confirms the presence of some species never reported since their original descriptions and increase the known distribution of others. Moreover, the specific diversity could increase with the verification of some doubtful identifications through genetic analysis, and the discovery of new species such as the recent description of Prosopistoma maroccanum (El Alami et al. 2022b).

In Algeria, the largest African country, it took decades after the pioneering investigations of Eaton (1899) and Lestage (1925), before significant taxonomical progress was made on mayfly knowledge (Soldán and Thomas 1983a; Gagneur and Thomas 1988; Thomas and Vitte 1988; Thomas 1998). Thomas (1998) provisionally listed 50 species of mayflies from Algeria. This checklist is undeniably valuable, despite confirmatory work remains needed as it likely contained some synonymies and misidentifications. In the last few years, a renewed interest in the taxonomy and ecological determinants of mayfly distribution is noted (Mebarki et al. 2017; Benhadji et al. 2018, 2019, 2020; Kechemir et al. 2020; Samraoui et al. 2021a–d). Systematic surveys by the Laboratoire de Conservation des Zones Humides in Algeria are covering eight regions or river basins (Seybouse River, Rhummel, Wadi El Kebir-East, Collo, Aures, Djurdjura, Tiaret, and the Sahara). Collected data have improved knowledge of the distribution and status of Algerian mayflies and led to the discovery of undescribed species (Samraoui et al. 2021c). In addition, collected data have allowed the elucidation of several mayfly life cycles (Bouhala et al. 2020a, b; Samraoui et al. 2021a, d). Further west, investigations of the mayflies from the Tafna River Basin are still proceeding (Benhadji et al. 2020). With 19 reported species, Baetidae is by far the most diversified but also the most problematic family in Algeria. Recent studies allowed the discovery of several potentially new species of Baetidae, as well as species not previously reported from this country (Benhadji et al. 2020; Samraoui et al. 2021b, c; Kaltenbach et al. 2022).

Mayflies from Tunisia encompass 25 species, 12 of them belonging to Baetidae (Zrelli et al. 2016). Boumaiza and Thomas (1986, 1994, 1995) studied and detailed the distribution and ecology of the different species. Baetis punicus was originally described from northern Tunisia (Thomas et al. 1983), but it is now reported from the whole Maghreb (Thomas 1998). More recently, a long-term survey was carried out in northern Tunisia, allowing the report of five additional species for this country (including three species of Baetidae) and the description a new species of Rhithrogena (Heptageniidae) (Zrelli et al. 2011a, b, 2012, 2016). The most important streams are in northern Tunisia where all the species occur. Despite corresponding to the 4/5 of the territory, the arid southern area only harbours three species. The Tunisian fauna can be considered as relatively well known; recent surveys did not reveal any new taxon or report (Bennas et al. 2018). As in Algeria and Morocco, the main challenges concern the identification of specimens assigned to widely distributed Western European species. Affinities and biogeographical patterns are discussed (Zrelli et al. 2016), but, here again, they need to be updated in the light of new molecular data. As far as we know, after two decades of important surveys, less attentions are paid nowadays on the mayfly systematics, ecology or monitoring in Tunisia (Bennas et al. 2018).

In Baetidae, imaginal stages remain difficult to identify to the species level. Larvae are easier to determine at a finest taxonomic level. Moreover, they are generally present all-year-round while emergence can be more sporadic. Therefore, collecting larvae generally remains the most efficient method to correctly assess the local fauna. Our main aim is to provide a key as easy to use as possible to allow a secure identification of baetid larvae to the most efficient level. The circled alphanumeric codes (1a, 1b, ...) indicated in the dichotomous key refer to the different illustrations of Figs 1, 2. We also summarize the main difficulties and gaps in knowledge.

Figure 1. 

Baetidae: larval characters A claw: Baetis sp. B claw: Cloeon sp. C lateral margin of cercus: Procloeon sp. D labial palp: Cloeon peregrinator E labial palp: Cheleocloeon dimorphicum F labial palp: Labiobaetis cf. neglectus G labial palp: Baetis (Rhodobaetis) sp. H abdominal gill IV: Cloeon peregrinator I abdominal gill IV: Centroptilum sp. J abdominal gill IV: Procloeon sp. K abdominal gill IV: Similicloeon simile L lateral margin of abdominal segments VII to IX: Cloeon peregrinator.

Figure 2. 

Baetidae: larval characters A right mandible: Alainites sadati B right mandible: Nigrobaetis rhithralis C right mandible: Baetis (Rhodobaetis) sp. D right mandible: Baetis (Patites) sp. E head frontal view: Alainites sp. F head frontal view: Baetis sp. G costal margin of gill IV: Baetis (Baetis) sp. H gill IV: Baetis (Rhodobaetis) sp. I costal margin of gill IV: Baetis (Rhodobaetis) sp. J paraproct: Nigrobaetis rhithralis K paraproct: Alainites sadati L distal margin tergite IV: Baetis (Rhodobaetis) sp. M distal margin tergite IV: Baetis (Patites) sp. N distal margin tergite IV: Baetis (Baetis) pavidus O foretibia: Acentrella cf. sinaica P foretibia: Baetis (Rhodobaetis) sp.

Mayfly larvae can be sampled using a Surber net or a dipnet, then stored in alcohol ideally at 80% to 95%. To preserve DNA, they must not be fixed in 5% formaldehyde. Adults can be collected with handnets or attracted by light traps. For the association of the ontogenic stages or just obtaining imagoes, rearing can be made in the field (following detailed instructions presented in http://www.insecta.bio.spbu.ru/z/rearing.htm). Rearing larvae in the laboratory until emergence generally requests equipment for water oxygenation. Association of the different stages can be also securely made by using molecular barcodes (Gattolliat and Monaghan 2010; Gattolliat et al. 2012, 2018).

Identification at the family or generic levels can be generally made under an efficient stereo microscope. In most cases, specific identification request slide mounting and observation under a compound microscope. Dissection can be made in alcohol or in Cellosolve (2-Ethoxyethanol), in adequation with subsequent mounting liquid (Canada balsam or Euparal). Identification to the species level based on mesoscopic characters such as abdominal pattern, shape and setation on legs, relative length of cerci and paracercus or tergite ornamentation should be restricted to the case of well-known fauna of a restricted watershed with examiners possessing important skills and training.

DNA can be extracted from the whole specimens or just from a small part, such as leg or thorax; specimens must be stored in alcohol at high concentration, without denaturant. Long term storage under inappropriate conditions (high temperature or temperature variations) may fragment DNA and inhibit the gene amplification. Non-destructive methods allowing subsequent morphological analysis should be preferentially used (see Vuataz et al. (2011) for details). For routine procedure, most effective results are obtained by amplifying the 658 bp fragment of the mitochondrial gene cytochrome oxidase subunit 1 (COI) using the primers LCO 1490 and HCO 2198 (Folmer et al. 1994, see Kaltenbach et al. (2020) for details).

Acentrella almohades Alba-Tercedor & El-Alami, 1999

Acentrella cf. sinaica Bogoescu, 1931

Alainites cf. muticus (Linnaeus, 1758)

Alainites oukaimeden (Thomas & Sartori, 1992)

Alainites sadati Thomas, 1994

Baetis (Baetis) cf. fuscatus (Linnaeus, 1760)

Baetis (Baetis) pavidus Grandi, 1949

Baetis (Patites) berberus Thomas, 1986

Baetis (Patites) maurus Kimmins, 1938

Baetis (Patites) punicus Thomas, Boumaiza & Soldán, 1983

Baetis (Rhodobaetis) atlanticus Soldán & Godunko, 2006

Baetis (Rhodobaetis) chelif Soldán, Godunko & Thomas, 2005

Baetis (Rhodobaetis) sinespinosus Soldán & Thomas, 1983

Centroptilum alamiae Kaltenbach, Vuataz & Gattolliat, 2022

Centroptilum samraouii Kaltenbach, Vuataz & Gattolliat, 2022

Cheleocloeon dimorphicum (Soldán & Thomas, 1985)

Cloeon peregrinator Gattolliat & Sartori, 2008

Cloeon saharense Soldán & Thomas, 1983

Labiobaetis cf. neglectus (Navàs, 1913)

Nigrobaetis numidicus (Soldán & Thomas, 1983)

Nigrobaetis rhithralis (Soldán & Thomas, 1983)

Procloeon algiricum (Eaton, 1899)

Procloeon cf. pennulatum (Eaton, 1870)

Procloeon stagnicola Soldán & Thomas, 1983

Similicloeon simile (Eaton, 1870)

In the Maghreb, Baetidae are the most diversified family of mayflies; they encompass ten genera, and three subgenera. We offer a straightforward dichotomic key to separate this family in twelve taxonomic units corresponding either to genera or subgenera. In the future, these taxonomic units should represent the standard identification level for environmental studies and water quality assessment. Among the 25 species of Baetidae reported from Maghreb, at least fourteen species are endemic to this area, underlying the diversity and uniqueness of this fauna. With eight species, Baetis is by far the most species-rich genus; other genera only contain one or two species. Links between European and Maghrebian faunas exist (Thomas 1998; Zrelli et al. 2016) but are less important than previously thought (Benhadji et al. 2020). Only six species unambiguously occur in the Maghreb and in Central Europe. Similicloeon simile is widely distributed in West Palearctic and in the Maghreb; Cloeon peregrinator is reported from Macaronesia and the Maghreb; Baetis atlanticus was originally described from Madeira, it is now reported from the Maghreb to Sweden including the British Islands; Baetis pavidus is extremely common in the whole Maghreb but seems rare in South of France and Italy; Acentrella almohades and Baetis maurus occur in the Maghreb and the Iberian Peninsula (Alba-Tercedor and El Alami 1999; Gattolliat et al. 2008; Benhadji et al. 2020; Samraoui et al. 2021b; El Alami et al. 2022a). Five taxa are tentatively attributed to Central European species (indicated as cf. in the list). Preliminary studies indicated that Acentrella cf. sinaica, Alainites cf. muticus and Labiobaetis cf. neglectus morphologically and/or genetically differ between these two regions (Benhadji et al. 2020; Samraoui et al. 2021c). These three taxa may represent endemic species to the Maghreb, closely related to their European sister species. The case of Cloeon cf. dipterum is more problematic as the species concept itself remains unclear. This complex of species encompasses at least six different lineages in the West Palearctic based on molecular evidence only, representing the same number of potential species (Rutschmann et al. 2014, 2017). According to our present knowledge, it remains impossible to decide which lineage corresponds to Cloeon dipterum sensu stricto. For the moment, only one lineage is reported with certainty from the Maghreb and it corresponds to Cloeon peregrinator (Benhadji et al. 2020). According to the diversity of habitats colonized by the larvae and the important morphological differences observed between populations, we could assume that more than one species occurs in the Maghreb. Species identification within Cloeon remains impossible without a broad scale study based on an integrative approach.

We consider as dubious, or at least requiring confirmation, the reports of seven species in the Maghreb, all of them having a European distribution. Baetis rhodani, Cloeon dipterum, and Cloeon cognatum belong to complexes of very close species; other species from these complexes are already reported from the Maghreb. However, the presence of these three species cannot be completely excluded. At least an important part of their reports corresponds to old identifications and are based on inappropriate concepts. According to preliminary results (El Alami et al. 2022a), some specimens of Baetis from Morocco cannot be assigned to any reported species. They clearly belong to the subgenus Baetis, their paracercus is not reduced and they do not exhibit spines on the margin of gills. They cannot be assigned to any species of the checklist; they are reported either as Baetis gr. fuscatus or Baetis gr. lutheri in El Alami et al. (2022a). As mentioned above, beside P. stagnicola, other species of Procloeon need a complete revision and extensive morphological and molecular comparisons with Iberian and Central European populations.

All these problematic cases clearly indicate the need of an extensive taxonomic revision in some taxa including specimens from Maghrebian, Mediterranean and Central European populations. Only an integrative approach involving at least morphology and molecular evidence can solve these taxonomic riddles.

In conclusion, Baetidae is the most diverse family of mayflies in the Maghreb. This family encompasses on the one hand common species with large ecological valence (e.g., Baetis pavidus, Baetis atlanticus, Cloeon peregrinator) and, on the other hand, rare species with very specific ecological requirements (e.g., Nigrobaetis rhithralis, Nigrobaetis numidicus, Alainites sadati). Therefore, identification to the family level may completely hide important environmental information as key conservation values. By offering a reasonably simple key to generic or subgeneric level, the main goal is to open the identification of this family to a wide range of scientists, and not only to a restricted set of experts. We hope that further ecological or environmental studies will confirm the high potential of this group for bioindication when working at lower taxonomic level. A better understanding of the distribution and ecology of the members of this family is an essential step for the conservation of these species and of the endangered freshwater habitat in general.

We want to thank Marion Podolak (MZL: Museum of Zoology, Lausanne) for the pictures of the different genera. The molecular study of Maghrebian Baetidae is still in progress; we want to express our gratitude to Céline Stoffel (MZL) and Marion Podolak for their dedicated work with the molecular lab. Further, we are thankful to Thomas Kaltenbach and Laurent Vuataz (MZL) for their constant interest and support for our projects in link with the study of Maghrebian mayflies. Lastly, the authors are grateful to the reviewers, Pavel Sroka and Zohar Yanai, for their valuable recommendations and comments on the manuscript.