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Background The emergence of insecticide resistance is a major threat to malaria control programmes in Africa, with many different factors contributing to insecticide resistance in its vectors, Anopheles mosquitoes. CYP6M2 has previously been recognized as an important candidate in cytochrome P450-mediated detoxification in Anopheles. As it has been implicated in resistance against pyrethroids, organochlorines and carbamates, its broad metabolic activity makes it a potential agent in insecticide cross-resistance. Currently, allelic variation within the Cyp6m2 gene remains unknown. Methods Here, Illumina whole-genome sequence data from Phase 2 of the Anopheles gambiae 1000 Genomes Project (Ag1000G) was used to examine genetic variation in the Cyp6m2 gene across 16 populations in 13 countries comprising Anopheles gambiae and Anopheles coluzzii mosquitoes. To identify whether these alleles show evidence of selection either through potentially modified enzymatic function or by being linked to variants that change the transcriptional profile of the gene, hierarchical clustering of haplotypes, linkage disequilibrium, median joining networks and extended haplotype homozygosity analyses were performed. Results Fifteen missense biallelic substitutions at high frequency (defined as > 5% frequency in one or more populations) are found, which fall into five distinct haplotype groups that carry the main high frequency variants: A13T, D65A, E328Q, Y347F, I359V and A468S. Despite consistent reports of Cyp6m2 upregulation and metabolic activity in insecticide resistant Anophelines, no evidence of directional selection is found occurring on these variants or on the haplotype clusters in which they are found. Conclusion These results imply that emerging resistance associated with Cyp6m2 is potentially driven by distant regulatory loci such as transcriptional factors rather than by its missense variants, or that other genes are playing a more significant role in conferring metabolic resistance.

Ancient DNA sequencing has shed significant light upon our knowledge of archaic and modern humans during the Middle and Upper Palaeolithic. Many Palaeolithic sites contain large numbers of bones, but due to the combination of post-depositional influences and carnivore processing of bone remains, many of them lack the diagnostic features required for identification of bone to specific taxon. Human remains dating to this period are, therefore, very rare. We have been applying a method of collagen fingerprinting to screen Palaeolithic bone fragments to identify the species/taxon of the bone, and importantly, to identify bone which has characteristic unique human peptides. The method utilizes mass spectrometry (MALDI-ToF-ToF) to produce a spectrum of peptide masses. Different species disclose small differences in the sequence of these peptides that enable them to be identified. Thus far we have found 4 new human fossil bone fragments from >4, 000 undiagnostic bone remains, ranging down to as small as 23 mm in length. We have screened bone from the Palaeolithic archaeological sites of Denisova Cave (Russia) and Vindija Cave (Croatia). We then carried out DNA sequencing to the bone fragments to identify into which human group they fall. At Denisova Cave, we previously identified a tiny bone as a hominin and showed, using DNA methods, that its mitochondrial DNA was of the Neanderthal type (Denisova 11). We have identified 2 additional bones which have been genetically analysed. At Vindija Cave we identified a new bone fragment from the G1 level and found that it had the same mtDNA sequence as another Neanderthal from the same site but a deeper level. We also found evidence for human processing on this bone in the form of cut-marks. We directly dated the bones using single amino acid dating of hydroxyproline. We will describe the significance of the new fossil finds in terms of the archaeological sequences at both sites. Collagen fingerprinting has immense potential for identifying hominin remains in highly fragmentary archaeological assemblages. Coupled with DNA analysis and direct dating, this method should be widely applied to previously excavated archaeological materials.

The period between ~45, 000 and 35, 000 14C years BP in Europe witnessed the “biocultural” transition from the Middle to early Upper Palaeolithic, when anatomically modern humans from Africa displaced Neanderthals across the continent. Significant questions remain regarding how this transition happened, for example to what extent Neanderthals and modern humans overlapped temporally and spatially, if modern humans or Neanderthals were responsible for various ‘transitional’ early Upper Palaeolithic industries, and when the last Neanderthals disappeared. Previous dating of Neanderthal remains from Vindija Cave (Croatia) led to the suggestion that Neanderthals survived there as recently as 28, 000-29, 000 14C years BP. Subsequent dating of Neanderthal specimens Vi-207 and Vi-208 from level G1 yielded older dates, interpreted as being at least ~32, 500 BP, but probably older. We have redated these same specimens using a different approach, developed at the ORAU, which is based on the extraction of the amino acid hydroxyproline that occurs in mammalian collagen using preparative high performance liquid chromatography. This method is more efficient than other methods in eliminating modern carbon contamination. We also applied Zooarchaeology by Mass Spectrometry (ZooMS) on 383 unidentified bone samples to find additional hominin remains and we identified one bone as Neanderthal based on its mitochondrial DNA. We also attempted to date some of the early Upper Palaeolithic bone points from stratigraphic units G1, Fd/d+G1, Fd/d, Fd, with mixed success owing to low levels of surviving collagen. In the current presentation we report all the new radiocarbon dates and DNA results. In contrast to previous suggestions, these data show that there is no reason to assume that the Neanderthals in Vindija Cave survived substantially later than at other places in Europe. Rather, they seem to pre- date the arrival of anatomically modern humans in Eastern Europe.

According to literature data on the distribution of water frogs in Croatia, the area south of the Sava River is inhabited by Pelophylax ridibundus, while P. ridibundus, Pelophylax lessonae and their hybridogenetic complex (Pelophylax kl. esculentus) can be observed in the area north of the Sava River. Since the morphological determination of species and hybrids of water frogs is not reliable, the aim of this study was to determine the taxonomical status of water frogs in Croatia using molecular markers (length variation of serum albumin gene intron‐1 and the mitochondrial gene for NADH dehydrogenase subunit 3). The study was performed on water frog populations from northern part and Adriatic coast of Croatia (Crna Mlaka fish ponds, Lika, Istra, Dalmatia, ). Results confirm the presence of species P. ridibundus, P. lessonae and their associated klepton P. kl. esculentus, and show an unexpected presence of species Pelophylax kurtmuelleri as well as new hybrid complexes. The presence of P. kurtmuelleri in the Lika region and Crna Mlaka fish ponds, and the discovery of P. lessonae in the Poštak Mountain (Dalmatia), point to the importance of revising the distribution data of water frog populations in Croatia.

Familie Trachinidae (Weeverfish) consists of 2 genera: Trachinus and Echiichthys. Trachinidae are known for their venomous spines on the gill covers and first few dorsal fins. All species live in sandy or sandy-muddy habitat buried in the ground of benthic, coastal or open ocean area. In the Adriatic 4 species of spiders are present: Trachinus draco, T. radiatus , T. araneus and Echiichthys Vipera. Due to great morphological diversity among species of genus Trachinus it is not easy to identify them unequivocally based only on biometric traits. The goal of this research was to determine population structure among species of genus Trachinus from Southern and Central part of Adriatic Sea based on three different mitochondrial phylogenetic markers. In total we examined 87 samples of Trachinus draco, Trachinus radiatus and Echyiichthys vipera species. Upon DNA extraction and PCR amplification data for analyses was obtained by standard Sanger sequencing of following markers: COI, 12S rDNA and 16S rDNA. Results of phylogenetic analyses on all gene markers have shown the same identification pattern of population structure. Therefore our research proved that used genetic markers and applied methods of molecular phylogenetics reconstruction are excellent for resolving population structure quandary inside genus Trachinus. It is necessary to conduct further detailed morphological and meristic studies in order to determine the differences observed by means of molecular phylogenetic analyses

Due to great morphological diversity among species of genus Trachinus it is not easy to identify them unequivocally based only on biometric traits. Three different mitochondrial phylogenetic markers, (COI, 12S rDNA, 16S rDNA) were examined on total of 87 samples. Results of phylogenetic analyses on all gene markers have managed to distinguish successfully different species among genus Trachinus.

Proteini MAB sadrţe dvije funkcionalno bitne interakcijske domene: MATH i BTB. Do sada je pokazano da su oni dio sustava eliminacije proteina koji stanici više nisu potrebni. Neki od istraženih proteina MAB (CeMEL-26, HsSPOP) dio su multiproteinskih kompleksa kulin 3 E3 ligaza i sluţe za prepoznavanje supstrata koje treba razgraditi. Tijekom razvoja pšenice ekspresija gena TaMAB2 prisutna je samo u oplođenoj jajnoj stanici, u zigoti te u dvostaničnom embriju. U ovom radu napravljena je heterologna ekspresija gena TaMAB2 iz pšenice u uročnjaku. Utvrđeno je da transgenične biljke proizvode cjeloviti protein TaMAB2 koji se u stanicama nakuplja. Sam protein TaMAB2 je ubikvitiniran, te vjerojatno stvara homo- i heterodimere. Rast i razvoj transgeničnih biljaka je značajno fenotipski promijenjen u odnosu na biljke divljeg tipa ali im fototropizam i gravitropizam nisu poremećeni. Analizom unutarstanične lokalizacije proteina TaMAB2 u stanicama BY-2 i pokožici luka utvrđeno je nakupljanje velikih agregata oko jezgrine ovojnice i u blizini stanične membrane. Ovi rezultati ukazuju da bi protein TaMAB2 mogao imati bitnu ulogu u eliminaciji proteina i uspostavljanju polarnosti tijekom rane embriogeneze pšenice. Water frogs (genus Rana, subgenus Pelophylax) present an interesting research subject for ecology, physiology and genetics. Rana kl. esculenta hybridogenetic complex, which consists of two parental species (Rana ridibunda and Rana lessonae) and their hybrid species (Rana kl. esculenta), is of special interest. Hybrids often show morphological traits similar to either parent, making them harder to identify on the field. Because of that, more relevance is given to molecular and phylogenetic methods in the identification of water frog species. Identification by morphological traits has shown that in central Croatia both parental populations of R. ridibunda and R. lessonae can inhabit the same location and thus lead to the formation of hybrids. The goal of this project was to determine the phylogenetic relationships of water frog populations from Crna Mlaka based on 16S rRNA, cytochrome b and ITS2sequence analysis, and to examine the efficiency of a simple RFLP method for identifying parental species and hybrids based on the digestion of a conserved ITS2 fragment with three restriction enzymes (KpnI, HaeII and SmaI). Results of this phylogenetic analysis have shown that both parental species R. ridibunda and R. lessonae, and thus potential hybrids, inhabit Crna Mlaka. Phylogenetic analysis of mitochondrial 16S rRNA and cytb genes proved to be the best method for species and haplotype identification, while the RFLP method and phylogenetic analysis of nuclear ITS2 regions, which could help differentiate hybrid samples, need further protocol optimization.

Water frogs (genus Rana, subgenus Pelophylax) present an interesting research subject for ecology, physiology and genetics. Rana kl. esculenta hybridogenetic complex, which consists of two parental species (Rana ridibunda and Rana lessonae) and their hybrid species (R. kl. esculenta), is of special interest. Hybrids often show morphological traits similar to either parent, making them harder to identify on the field. Because of that, more relevance is given to molecular and phylogenetic identification methods. In central Croatia both R. ridibunda and R. lessonae parental populations can inhabit the same location and thus lead to the formation of hybrids. The goal of this project was to determine the phylogenetic relationships of water frog populations from Crna Mlaka based on 16S rRNA, cytochrome b and ITS2 sequence analysis, and to examine the efficiency of a simple RFLP method for identifying parental species and hybrids based on the digestion of a conserved ITS2 fragment with three restriction enzymes (KpnI, HaeII and SmaI). Results of this phylogenetic analysis have shown that both parental species R. ridibunda and R. lessonae, and thus potential hybrids, inhabit Crna Mlaka. Phylogenetic analysis of mitochondrial 16S rRNA and cytb genes proved to be the best method for species and haplotype identification, while the RFLP method and phylogenetic analysis of nuclear ITS2 regions, which could help differentiate hybrid samples, need further protocol optimization.

Zelene žabe (rod Rana, podrod Pelophylax) često su predmet ekoloških, fizioloških i genetičkih istraživanja. Najviše je istražen hibridogenetički kompleks koji se sastoji od roditeljskih vrsta Rana ridibunda i Rana lessonae te njihovog hibrida Rana kl. esculenta. Hibridi pokazuju obilježja oba roditelja, što otežava determinaciju isključivo na temelju morfoloških osobina. Stoga se u determinaciji sve veću važnost pridaje molekularno-filogenetskim metodama. Kontinentalna Hrvatska je moguće područje stvaranja hibrida zbog preklapanja areala populacija R. ridibunda i R. lessonae. Cilj ovog istraživanja bilo je utvrditi molekularno-filogenetske odnose jedinki zelenih žaba R. kl esculenta kompleksa prikupljenih na močvarnom lokalitetu Crna Mlaka analizom sekvenci za 16S rRNA, citokrom b i jezgrinu ITS2 regiju, te provjeriti valjanost određivanja roditeljskih vrsta i hibrida jednostavnom RFLP metodom koja se temelji na restrikciji konzerviranog ITS2 fragmenta restrikcijskim enzimima KpnI, HaeII i SmaI. Rezultati analize pokazali su prisutnost R. ridibunda i R. lessonae jedinki te moguću prisutnost hibrida na području Crne Mlake. Za determinaciju na razini vrste i haplotipa najuspješnijom se pokazala filogenetska analiza mitohondrijskih 16S i cytb gena, dok je za RFLP metodu i filogenetsku analizu molekularnim biljegom ITS2, pomoću kojih bi bilo moguće razlikovati hibridne jedinke od roditelja, potrebna optimizacija protokola.

Svjetlosno onečišćenje je pojam koji se koristi kako bi se opisalo negativne posljedice prekomjerne, neadekvatne i nametljive umjetne noćne rasvjete koja u konačnici dovodi do gubitka mraka i noći kao važnog biološkog čimbenika. Razlikujemo „astronomsko svjetlosno onečišćenje“ koje onemogućuje astronomska opažanja zbog porasta sjaja neba čime se smanjuje kontrast sa objektima opažanja (zvijezde, asteroidi i sl.) te „ekološko svjetlosno onečišćenje“ koje mijenja ili interferira sa prirodnom noćnom svjetlosti. Preko ovog seminara dan je kratak pregled do sada otkrivenih negativnih posljedica svjetlosnog onečišćenja, sa naglaskom na taksonomske skupine poput morskih kornjača, ptica selica, noćnih leptira i šišmiša, kod kojih su primijećene najteže posljedice. Posebno su obrađene negativne posljedice na biljke, kod kojih je rađeno vrlo malo konkretnih istraživanja, te negativne posljedice na čovjeka, zbog toga što je nedavno otkriven cijeli niz zdravstvenih problema povezanih uz izlaganje svjetlu za vrijeme noći, od kojih je najteži povećan rizik od raka dojke. Iako se na prvi pogled čini kako je posljednjih godina napravljeno puno posla, veći dio ipak ostaje neistražen, a pravi omjer problema počeo se tek nedavno naslućivati. Light pollution is a term used to describe the negative consequences of excessive, scattered and intrusive artificial night light, which ultimately leads to the loss of darkness and the night as an important biological factor. We distinguish “astronomical light pollution” which prevents astronomical observations because of increased night sky brightness that lowers the contrast with the observed objects (stars, asteroids and such), and “ecological light pollution” which alters or interferes with natural light regimes during the night. The aim of this seminar is to give a short review of the negative consequences of light pollution that were discovered so far, with emphasis on taxa such as sea turtles, migrating birds, moths and bats, because they showed the most severe side effects. The negative consequences on plants and humans were described separately, mostly because there was little research on plants, and because the full list of health problems associated with exposure to light during nighttime was just recently discovered, the most severe being the increased risk of breast cancer development. Even though it may seem that a lot of work has been done these previous years, there is a lot that still needs to be researched, and so far we have only gotten a glimpse of the real extent of the problem.

A model for analysis of light pollution effects on the brightnes of the night sky is presented. The model is based on Walker's law of LP propagation and statistical data about population of the area of interest, including all surroundig areas closer than 200 km. The situation in Croatia is modelled for the first time. The model shows that the whole area of it is at least moderately light-polluted, with exception of a few islands in southern Adriatic, and some places in Lika and Gorski kotar. The model can be very useful in assesing the effect of light-pollution in cases where ecological impact studies are required for new or existing structures. Field measurements support the model predictions rather well and will be described for a few darkest places in Croatia.

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Additional file 7: Fig. S5. Hierarchical clustering and missense mutations for Cyp6m sub cluster. Top: a dendrogram showing hierarchical clustering of haplotypes across the Cyp6m sub cluster of genes containing Cyp6m2, Cyp6m3 and Cyp6m4. The genes are located at position 6928858 to 6935721. The colour bar indicates the population of origin for each haplotype. Bottom: high frequency (> 5%) alleles identified within each haplotype (white = reference allele; black = alternative allele).

Additional file 9: Fig. S7. Extended haplotype homozygosity across all populations. A rapid decay of EHH in comparison to other haplotypes implies absence of positive selection.

Additional file 6: Fig. S4. Hierarchical clustering and missense mutations for SH2. Top: a dendrogram showing hierarchical clustering of haplotypes across the SH2 gene. The gene is located at position 8,176,778 to 8,183,084: 1,246,231 bases downstream of Cyp6m2. The colour bar indicates the population of origin for each haplotype. Bottom: high frequency (> 5%) alleles identified within each haplotype (white = reference allele; black = alternative allele).

Additional file 5: Fig. S3. Hierarchical clustering and missense mutations for HAM. Top: a dendrogram showing hierarchical clustering of haplotypes across the HAM gene. The gene is located at position 7,435,306 to 7,485,012: 504,759 bases downstream of Cyp6m2. The colour bar indicates the population of origin for each haplotype. Bottom: high frequency (> 5%) alleles identified within each haplotype (white = reference allele; black = alternative allele).

Additional file 7: Fig. S5. Hierarchical clustering and missense mutations for Cyp6m sub cluster. Top: a dendrogram showing hierarchical clustering of haplotypes across the Cyp6m sub cluster of genes containing Cyp6m2, Cyp6m3 and Cyp6m4. The genes are located at position 6928858 to 6935721. The colour bar indicates the population of origin for each haplotype. Bottom: high frequency (> 5%) alleles identified within each haplotype (white = reference allele; black = alternative allele).

Additional file 4: Fig. S2. Hierarchical clustering and missense mutations for ODR2. Top: a dendrogram showing hierarchical clustering of haplotypes across the ODR2 gene. The gene is located at position 7,059,422 to 7,119,244: 128,875 bases downstream of Cyp6m2. The colour bar indicates the population of origin for each haplotype. Bottom: high frequency (> 5%) alleles identified within each haplotype (white = reference allele; black = alternative allele).

Additional file 8: Fig. S6. Hierarchical clustering and missense mutations for Cyp6 supercluster. Top: a dendrogram showing hierarchical clustering of haplotypes across the Cyp6 supercluster of 14 P450 genes containing Cyp6s2, Cyp6s1, Cyp6r1, Cyp6n2, Cyp6y2, Cyp6y1, Cyp6m1, Cyp6n1, Cyp6m2, Cyp6m3, Cyp6m4, Cyp6z3, Cyp6z2 and Cyp6z1. The genes are located at position 6903106 to 6978142. The colour bar indicates the population of origin for each haplotype. Bottom: high frequency (> 70%) alleles identified within each haplotype (white = reference allele; black = alternative allele).

Additional file 9: Fig. S7. Extended haplotype homozygosity across all populations. A rapid decay of EHH in comparison to other haplotypes implies absence of positive selection.

Additional file 3: Fig. S1. Linkage disequilibrium (D′) between non-synonymous variants. A value of 1 shows perfect linkage between the alleles. A value of − 1 shows that the alleles are never found conjointly. The bar plot indicates allele frequencies within the Ag1000G phase 2 cohort.

Additional file 3: Fig. S1. Linkage disequilibrium (D′) between non-synonymous variants. A value of 1 shows perfect linkage between the alleles. A value of − 1 shows that the alleles are never found conjointly. The bar plot indicates allele frequencies within the Ag1000G phase 2 cohort.

Additional file 5: Fig. S3. Hierarchical clustering and missense mutations for HAM. Top: a dendrogram showing hierarchical clustering of haplotypes across the HAM gene. The gene is located at position 7,435,306 to 7,485,012: 504,759 bases downstream of Cyp6m2. The colour bar indicates the population of origin for each haplotype. Bottom: high frequency (> 5%) alleles identified within each haplotype (white = reference allele; black = alternative allele).

Additional file 4: Fig. S2. Hierarchical clustering and missense mutations for ODR2. Top: a dendrogram showing hierarchical clustering of haplotypes across the ODR2 gene. The gene is located at position 7,059,422 to 7,119,244: 128,875 bases downstream of Cyp6m2. The colour bar indicates the population of origin for each haplotype. Bottom: high frequency (> 5%) alleles identified within each haplotype (white = reference allele; black = alternative allele).

Additional file 8: Fig. S6. Hierarchical clustering and missense mutations for Cyp6 supercluster. Top: a dendrogram showing hierarchical clustering of haplotypes across the Cyp6 supercluster of 14 P450 genes containing Cyp6s2, Cyp6s1, Cyp6r1, Cyp6n2, Cyp6y2, Cyp6y1, Cyp6m1, Cyp6n1, Cyp6m2, Cyp6m3, Cyp6m4, Cyp6z3, Cyp6z2 and Cyp6z1. The genes are located at position 6903106 to 6978142. The colour bar indicates the population of origin for each haplotype. Bottom: high frequency (> 70%) alleles identified within each haplotype (white = reference allele; black = alternative allele).