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Contamination of water (and food) with fecal bacteria is a well known common and diligent problem affecting public health and the local and national economy One of the best indicators of fecal bacterial water pollution is Escherichia coli (E coli) which belongs to the group of enteric bacteria and most often creates diarrheal diseases which can have fatal consequences. Within this study, a comparative analysis using conventional methods (microbiological detection and Real-Time PCR analysis) and Loop mediated isothermal amplification.

In order to disentangle the currently confused interpretations and nomenclature ofMerodon aureusandM. aeneus, we have reviewed all existing type material and species names known to us as assigned synonyms of these taxa. We resolveM. aeneusas being a junior synonym ofM. aureus. We designate a lectotype forM. aureusand a neotype forM. aeneus.Additionally, we provide evidence thatM. aureus, together with two newly discovered taxa (M. calidussp. nov.andM. ortussp. nov.), represent a complex of cryptic species named theM. aureusspecies complex. This complex, together with theM. unicolorspecies complex and the speciesM. pumilus, is part of theM. aureussubgroup. TheM. unicolorspecies complex comprises two cryptic species:M. unicolorandM. albidussp. nov. The new species are described by applying an integrative taxonomic approach using several data types (COIand 28S rRNA genes, geometric morphometry of the wings, ecological and distributional data). Based on theCOIgene sequence analysis and distributional data, the pupa previously described as an immature stage of the speciesM. aureusis redefined as an immature stage of the new speciesM. calidus. Speciation within theM. aureussubgroup is discussed in the context of the phylogeographic history in the studied region.

Considering the threat of pollinator decline to seriously affect ecosystem function and food security, the conservation of pollinating insects has become of utmost importance, entailing the accurate taxa identification in order to establish a long-term monitoring strategy. In addition to pollination service, hoverflies (Diptera: Syrphidae) also have a role as habitat bioindicators, as well as in organic matter recycling and protection of cultivated plants from pests. Representing one of the most species-rich hoverfly genera, Merodon Meigen, 1803 holds numerous taxonomic challenges, especially within the monophyletic Merodon ruficornis species group, which is characterized by incongruence between morphological and molecular data. In the present study, we successfully resolved the taxonomy of the M. ruficornis group using a combination of four genetic markers (3′ and 5′ ends of the COI gene, 28S and ITS2) and demonstrate their potential as molecular tools for resolving taxonomic questions within the Merodon genus. We proved the utility of indels as valuable molecular signals for the taxa delimitation within M. ruficornis group by performing two different indels schemes—(1) excluding indels from analysis as missing data; and (2) coding indels as presence/absence character matrix. In addition to the found congruence with morphologically defined species, our results revealed intraspecific differentiation between Anatolian and Balkan populations within three species—M. loewi, M. trebevicensis and M. armipes. According to the divergence time estimates, lineage diversification within the M. ruficornis group probably took place during Pleistocene climatic changes as a consequence of the species’ range alterations. © 2022, Gesellschaft für Biologische Systematik.

INTRODUCTION: As of lately, loop-mediated isothermal amplification (LAMP) has become a powerful alternative to the polymerase chain reaction (PCR) in the molecular diagnostic world, especially for pathogen detection in clinical and food samples. LAMP is a powerful and new method of nucleic acid amplification, with the ability to detect DNA at a very low level. In comparison to other molecular methods such as PCR and Real-Time PCR, LAMP eliminates the need for sophisticated thermal cyclers. High efficiency of DNA amplification by LAMP significantly shortens the whole amplification process. OBJECTIVES: This study aims to assess the efficacy of two different LAMP kits, WarmStart (Real-Time LAMP) and Colorimetric LAMP in detection of Escherichia coli. We tested and optimized the kits for the detection of E. coli ATCC® 25922 directly from bacterial suspension and indirectly using bacterial genomic DNA (gDNA). LAMP results were then compared with the results of E. coli detection using standard PCR methodology. METHOD / DESIGN: For use in LAMP detection of E. coli directly from the bacterial suspension, we prepared eight bacterial dilutions in concentrations ranging from 1.5x108 - 1.5x101 CFU/mL. All bacterial dilutions were then used for LAMP detection using WarmStart Colorimetric LAMP 2X Master Mix (New England BioLabs) for colorimetric LAMP and WarmStart LAMP Kit (New England BioLabs) for Real-Time LAMP. Real-Time LAMP reactions were done on the Genie® II, an instrument for isothermal nucleic acid amplification (Figure 1). LAMP primers (F3, B3, FIP, BIP, LF, BF) (Figure 2) for the malB gene are taken from the literature. For indirect detection we extracted genomic DNA from E. coli using GeneJET Genomic DNA Purification Kit (Thermo Scientific). Extracted gDNA with the concentration 50 ng/μl was used for preparation of ten-fold dilutions (10-1 - 10-4 ), and these dilutions were used in both Colorimetric and WarmStart LAMP assays in order to compare limits of detection with and without DNA extraction. Finally, we used E. coli genomic DNA serial dilutions for standard PCR, using F3 and B3 primers for malB gene. RESULTS: The results showed that both LAMP methods (WarmStart and Colorimetric LAMP) enabled the detection of E. coli directly, in suspension (without prior DNA isolation) for bacterial concentration of 1.5 x 10 8 CFU/mL (Figure 3, Chart 1). Additionally, the results of Colorimetric LAMP using serial dilutions of E. coli gDNA showed detection in all tested concentrations, except the lowest (10 -4 ) (Figures 4-5). On the other hand, the WarmStart LAMP kit enabled E. coli gDNA detection in all tested gDNA dilutions (Chart 2, Figure 6). The conventional PCR methods failed to detect E. coli malB gene product for the lowest DNA concentration (Figure 7.). Additionally, DNA bends on agarose gels are more intensive in the LAMP products compared to those obtained with PCR products., International Bioscience Conference and the 8th International PSU - UNS Bioscience conference. Novi Sad, Serbia, November 25-26, 2021, Book of Abstracts, 159-160.

The taxonomy of Merodon dobrogensis Bradescu, 1982 (Diptera: Syrphidae) species subgroup was reviewed. Multiple data sources (morphology, geometric morphometry of wings and surstylus, molecular data, and distributional data) were used to investigate the species subgroup in the manner of integrative taxonomy. Merodon dobrogensis Bradescu, 1982 and M. puniceus Vujić, Radenković, and Pérez-Bañón, 2011 are supported as distinct species belonging to the M. dobrogensis species complex within the M. dobrogensis species subgroup. Additionally, evidence is presented for the description of a new species, M. rojoi Radenković and Vujić new species, with a distribution in mainland Greece, the Greek island Euboea, and the Peloponnese. A short diagnosis is provided for the M. aureus species group, the M. dobrogensis species subgroup, and the newly defined M. dobrogensis species complex, in addition to a description of the new species, with drawings and photographs of adult morphology.

© 2019 Blackwell Verlag GmbH In this study, we explored intraspecific genetic differentiation of hoverfly species of the genus Eumerus with regard to landscape discontinuities (due to paleogeological events), isolation-by-distance, evolutionary processes, and Quaternary climatic oscillations. We unveil genetically diverging regions and discuss the potential driving forces that gave rise to these spatial genetic patterns. We generated mitochondrial DNA (mtDNA) barcodes for 274 individuals of nine Eumerus species, sampled from 58 localities in the Mediterranean and Balkans. Spatially explicit Bayesian clustering, correlation tests between geographic and genetic distances (presence of isolation-by-distance), median neighbor-joining haplotype networks, and landscape shape interpolation analyses were employed to investigate spatial genetic patterns. Bayesian clustering generated one to three genetic clusters with high posterior probability values. We also observed high mtDNA haplotype diversity consisting of unique and shared haplotypes, as well as starlike mtDNA haplotype patterns. The mtDNA haplotype network was consistent with species distributions and Bayesian clustering for four tested species. The Mantel tests confirmed the absence of isolation-by-distance in seven species. We identified genetically diverging areas through our landscape shape interpolation analyses. Five species displayed neither spatial genetic patterns nor evidence of isolation-by-distance, indicative of relict taxa. Our study is the first broad- and large-scale study of Eumerus species in the Mediterranean and Balkans; it reveals spatial genetic clusters in four species and identifies the potential factors driving those patterns.

The taxonomy of the syrphid genus Chrysotoxum Meigen, 1803 (Diptera: Syrphidae), is complex and currently under scrutiny. Two new species allied with Chrysotoxum vernale, one from the Western Mediterranean, Chrysotoxum hispanicum sp. n. and the other from the Eastern Mediterranean, Chysotoxum anatolicum sp. n., are described and illustrated. Chrysotoxum hispanicum sp. n. is distinguished from the similar C. vernale Loew by the size of the yellow abdominal fasciae and shape of surstyli. Chrysotoxum anatolicum sp. n., known only from females, possesses an almost entirely yellow-pigmented wing, unusual amongst the other studied species of the C. vernale group. Additionally, C. hispanicum sp. n. and C. anatolicum sp. n. are separated from each other, as well as from other species of the C. vernale group by COI and ITS2 gene markers. An identification key to the West Palaearctic species of the C. vernale group is provided. This study was funded by the Ministry of Education, Science and Technological Development of the Republic of Serbia (Research Project No: OI173002), the H2020 project "ANTARES" (No: 664387), and the ‘Vicerrectorado de Investigación y Transferencia de Conocimiento’ of the University of Alicante (Ref. UATAL05).

We revise the Merodon constans species group of the genus Merodon Meigen, 1803 (Diptera: Syrphidae), provide morphological diagnoses and descriptions, as well as an illustrated key and a discussion on the different taxonomic characters used. In total, 15 species were studied, their geographic distributions are presented on maps, and nine new species are described. Two species are redefined and neotypes are designated, lectotypes are designated for five species, and one species is reinstated as valid. Following a detailed study of type material in different entomological collections, the status of several species is revised and three new synonymies are proposed. The M. constans species group was resolved as being monophyletic within the M. albifrons lineage based on molecular analyses using COI and 28S rRNA gene sequences. Three species morphologically similar to M. constans (Rossi, 1794) but occurring outside its distributional range were supported as being valid and distinct species on the basis of molecular data, but they were not distinguishable based on morphological characters. By contrast, continental populations of M. analis Meigen, 1822 could not be separated from Mediterranean M. constans based on differences in COI or 28S rRNA genes. The same molecular markers could not discriminate between two other species pairs. We conclude that these molecular markers only partially resolve species within the M. constans group. Geometric morphometry of wing shape successfully separated M. analis and M. constans, as well as M. spineus Vujić, Šašić Zorić & Likov, sp. nov. in both species and population analyses.

Background: The black soldier fly (Diptera: Stratiomyidae, Hermetia illucens) is renowned for its bioconversion ability of organic matter, and is the worldwide most widely used source of insect protein. Despite varying extensively in morphology, it is widely assumed that all black soldier flies belong to the same species, Hermetia illucens. We here screened about 600 field-collected and cultured flies from 39 countries and six biogeographic regions to test this assumption based on data for three genes (mitochondrial COI, nuclear ITS2 & 28S rDNA) and in order to gain insights into the phylogeography of the species. Results: Our study reveals a surprisingly high level of intraspecific genetic diversity for the mitochondrial barcoding gene COI (divergences up to 4.9%). This level of variability is often associated with the presence of multiple species, but tested nuclear markers (ITS2 and 28S rDNA) were invariant and fly strain hybridization experiments under laboratory conditions revealed reproductive compatibility. COI haplotype diversity is not only very high in all biogeographic regions (56 distinct haplotypes in total), but also in breeding facilities and research centers from six continents (10 haplotypes: divergences up to 4.3%). The high genetic diversity in fly-breeding facilities is mostly likely due to many independent acquisitions of cultures via sharing and/or establishing new colonies from field-collected flies. However, explaining some of the observed diversity in several biogeographic regions is difficult given that the origin of the species is considered to be New World (32 distinct haplotypes) and one would expect severely reduced genetic diversity in the putatively non-native populations in the remaining biogeographic regions. However, distinct, private haplotypes are known from the Australasian (N = 1), Oriental (N = 4), and the Eastern Palearctic (N = 4) populations. We reviewed museum specimen records and conclude that the evidence for introductions is strong for the Western Palearctic and Afrotropical regions which lack distinct, private haplotypes. Conclusions: Based on the results of this paper, we urge the black soldier fly community to apply molecular characterization (genotyping) of the fly strains used in artificial fly-breeding and share these data in research publications as well as when sharing cultures. In addition, fast-evolving nuclear markers should be used to reconstruct the recent invasion history of the species. This work was funded by the H2020 Research and Innovation Staff Exchange Programme of the European Commission (RISE), project 645636: ‘Insect-plant relationships: insights into biodiversity and new applications’ (FlyHigh), and the grants R-154-000-B46–114 and R-154-000-A94–592 to RM, as well as a grant of the Swiss Federal Office for Agriculture to CS (‘Efficiency and sustainability of the production and feeding of insect based feedstuff to fish and poultry’).

© 2018, © 2018 Taylor & Francis. A new species of the genus Chrysotoxum (Diptera: Syrphidae), Chrysotoxum bozdagensis Nedeljković, Vujić & Hayat sp. n., collected from Bozdağ Mt., İzmir Province, Turkey is described and illustrated. Chrysotoxum bozdagensis sp. n. can be distinguished from the similar Ch. octomaculatum Curtis, 1837 by the presence of a black pile on the mesonotum and the colour of the legs. It is also similar to Ch. elegans Loew, 1841, from which it can be distinguished by the colour of the pile on the vertical triangle and scutellum, as well from the structure of the male genitalia. Additionally, Ch. bozdagensis sp. n. can be clearly distinguished from the latter two species by DNA sequence data. This new species increases the total number of Chrysotoxum species in Turkey to 18. http://www.zoobank.org/urn:lsid:zoobank.org:pub:7465AAEA-21E6-4F0F-984C-CD48BE75436F.

The immature stages of species of the phytophagous genus Merodon Meigen (Diptera: Syrphidae), as well as their host plants, are mostly unknown. All known immature stages of Merodon feed on underground storage organs (bulbs, rhizomes and corms) of geophytes of the families Asparagaceae, Iridaceae and Amaryllidaceae. Of 160 known Merodon species, to date, the pupal stages have been described for only four: M. equestris (Fabricius), M. bombiformis Hull, M. luteihumerus Marcos-García, Vujić & Mengual, and M. geniculatus Strobl. During field investigations in Đerdap National Park, Serbia, Merodon puparia were found in the ground near the bulbs of Ornithogalum umbellatum L. (Asparagaceae). DNA barcoding revealed that they belonged to the species M. aureus Fabricius and M. avidus (Rossi). Analysis of museum material from the Bavarian State Collection of Zoology in Germany revealed the puparium of an additional species, M. rufus Meigen. In our study we provide for the first time descriptions of the puparia of these three Merodon species. The main diagnostic morphological characters of the pupal spiracles and posterior respiratory processes are described using scanning electron microscopy, and cephalopharyngeal skeletons using binocular microscopy. In addition, puparium morphology of M. aureus, M. avidus and M. rufus is compared with known puparia of four other Merodon species and with the third larval stage of M. hurkmansi Marcos-García, Vujić & Mengual. The study was supported by the Ministry of Education, Science and Technological Development, Republic of Serbia, Grant No. 173002.

© 2017 Blackwell Verlag GmbH The Merodon aureus group is characterized by high endemism and the presence of morphologically cryptic species. Within one of its subgroups, M. bessarabicus, seven species and four more species complexes have been described to date. One of these complexes, the M. luteomaculatus, comprises new taxa that are the subject of the present study. Its members have allopatric ranges restricted to the Balkan Peninsula and Aegean islands. This complex exhibits morphological variability that could not be characterized using a traditional morphological approach. Thus, we used integrative taxonomy with independent character sets (molecular, geometric morphometric, distributional) to delimit species boundaries. Data on three molecular markers (COI, 28S rRNA, and ISSR) and geometric morphometry of the wing and male genitalia, together with distributional data, enabled recognition of six cryptic species within the complex: M. andriotes sp. n., M. euri sp. n., M. erymanthius sp. n., M. luteomaculatus sp. n., M. naxius sp. n., and M. peloponnesius sp. n. We discuss the possible influence of Aegean paleogeographical history on the speciation of this complex.

A new species of the genus Chrysotoxum (Diptera: Syrphidae), Chrysotoxum bozdagensis Nedeljković, Vujić & Hayat sp. n., collected from Bozdağ Mt., İzmir Province, Turkey is described and illustrated. Chrysotoxum bozdagensis sp. n. can be distinguished from the similar Ch. octomaculatum Curtis, 1837 by the presence of a black pile on the mesonotum and the colour of the legs. It is also similar to Ch. elegans Loew, 1841, from which it can be distinguished by the colour of the pile on the vertical triangle and scutellum, as well from the structure of the male genitalia. Additionally, Ch. bozdagensis sp. n. can be clearly distinguished from the latter two species by DNA sequence data. This new species increases the total number of Chrysotoxum species in Turkey to 18.

Examination of 122 specimens of Xanthogramma Schiner, 1861 (Diptera: Syrphidae)from varied localities in Europe (+ Turkey) resulted in the description of two new species (X. aeginaeRicarte, Nedeljković, and Vujić new species and X. pilosum Nedeljković, Ricarte, and Vujić newspecies), as well as new data on six other species. Most of the examined material originated from theBalkan Peninsula and Greek islands. New species concepts were supported by morphological andmolecular evidence. Relationships among the eight studied species were analysed and discussed basedon the data of nuclear (ITS2) and mitochondrial (COI) genes sequences. An identification key to theEuropean species of Xanthogramma is provided. Lectotypes are designated for Doros decoratumZetterstedt, 1843, Lasiophthicus novus Rondani, 1857, Syrphus laetus Fabricius, 1794, Syrphusornatus Meigen, 1822, and Xanthogramma nobilitatum Frey, 1946.

Examination of 122 specimens of Xanthogramma Schiner, 1861 (Diptera: Syrphidae) from varied localities in Europe (+Turkey) resulted in the description of two new species (X. aeginae Ricarte, Nedeljković, and Vujić new species and X. pilosum Nedeljković, Ricarte, and Vujić new species), as well as new data on six other species. Most of the examined material originated from the Balkan Peninsula and Greek islands. New species concepts were supported by morphological and molecular evidence. Relationships among the eight studied species were analysed and discussed based on the data of nuclear (ITS2) and mitochondrial (COI) genes sequences. An identification key to the European species of Xanthogramma is provided. Lectotypes are designated for Doros decoratum Zetterstedt, 1843, Lasiophthicus novus Rondani, 1857, Syrphus laetus Fabricius, 1794, Syrphus ornatus Meigen, 1822, and Xanthogramma nobilitatum Frey, 1946. Financial support was provided by the Serbian Ministry of Education and Science (projects OI173002 and III43002), the Provincial Secretariat for Science and Technological Development (project “Genetic resources of agro-ecosystems in Vojvodina and sustainable agriculture”), the FP7 EU project, Innosense, and the H2020 project “ANTARES” (664387), Antonio Ricarte’s position at the University of Alicante is funded by the Vicerrectorado de Investigación y Transferencia de Conocimiento).

In this study, we examined the morphology, genetics and distribution of the members of the Merodon chalybeus subgroup (M. aureus species group): M. chalybeus Wiedemann in Meigen, 1822, M. minutusStrobl, 1893 and M. robustusVeselić, Vujić & Radenković, 2017. Two of the species, M. chalybeus and M. minutus, are morphologically very similar and often misidentified in the literature. Here, by employing an integrative taxonomic approach we provide strong evidence for the separation of M. chalybeus and M. minutus. Our results show their clear allopatric distribution: M. minutus on the Balkan Peninsula, Sicily, Sardinia and Corsica, while M. chalybeus is a western Mediterranean species distributed on the Iberian Peninsula and northwest Africa. Data on the distribution of M. robustus were updated, with new records from Cyprus, Israel and Turkey, besides its type locality (Samos in Greece). We provide evidence for M. chalybeus and M. minutus representing a species complex, named the M. chalybeus complex, which together with M. robustus constitute the M. chalybeus subgroup. © 2021 Finnish Zoological and Botanical Publishing Board.

Merodon is the richest genus of hoverflies in Europe with 120 recognized species. The adult species of this genus are important pollinators of diverse plants, while larvae are phytophagous and they develop in geophytes. Species discovery within genus is facilitated by the application of DNA barcoding approach in almost all recent studies. DNA barcoding is based on the analysis of sequence divergence of the short fragment on 5’ end of mitochondrial cytochrome c oxidase subunit I gene (COI), although, in hoverflies, both 5’ and 3’ COI sequences are equally used and often combined. The aim of this study was to identify hoverfly specimens collected in Morocco, Italy, Turkey and Georgia to a species level within the M. aureus species group. In order to achieve this, we analysed 5’ COI sequences of aforementioned specimens. The sequences were blasted against the NCBI nucleotide database and used for Maximum parsimony and Maximum likelihood trees construction. Three new candidate species are discovered: M. sp. nova 1, M. sp. nova 2 and M. aff. bessarabicus. The three species are resolved as reciprocally monophyletic clades with strong branch support on both trees. In order to verify and describe the three species, additional examination of morphological character states is needed., This study was supported by the Ministry of Education, Science and Technological Development, Republic of Serbia, Grant No. 173002, III43002, 451-03-68/2020-14/ 200358 and 451-03-68/2020-14/ 200125, and the European Commission, Horizon 2020 project 'ANTARES' (Grant No. 739570).

Integrative taxonomy is a multisource approach that takes advantage of complementarities among disciplines and tends to make progress in species delimitation and diagnosis of cryptic diversity. Single-method approaches in taxonomic and systematic studies have many limitations, especially for diagnosis of cryptic species that can be overlooked. Cryptic species are morphologically indistinguishable (or almost), so a combination of molecular, ecological and subtle morphological characters, as well as phylogeographic and population genetic analyses have been proposed as a framework to diagnose and distinguish cryptic species. Our overview had three objectives: (1) to present potential limitations of recently used concept of integrative taxonomy in genera Merodon and Cheilosia; (2) to discuss further methods which can be applied to clarify the species borders in these two genera, beside morphology, geometric morphometrics of wings and male surstylus shape, and molecular data; (3) to underline potential taxa where integration of all available data can help to discover cryptic diversity.