EPIC) nuclear loci and mitochondrial cytochrome oxidase subunit I, to investigate the evolutionary history of five medically important and inherently forest dependent mosquito species of the genus Aedes. By testing different demographic hypotheses, we show that Aedes bromeliae and Aedes lilii fit the same model of lineage diversification, admixture, expansion, and recent population structure previously inferred for Aedes aegypti. In addition, analyses of population structure show that Aedes africanus has undergone lineage diversification and expansion while Aedes hansfordi has been impacted by population expansion within Uganda. This congruence in evolutionary history is likely to relate to historical climate‐driven habitat change within Africa during the late Pleistocene and Holocene epoch. We find differences in the population structure of mosquitoes from Tanzania and Uganda compared to Benin and Uganda which could relate to differences in the historical connectivity of forests across the continent. Our findings emphasize the importance of recent climate change in the evolution of African forest biota.The study of demographic processes involved in species diversification and evolution ultimately provides explanations for the complex distribution of biodiversity on earth, indicates regions important for the maintenance and generation of biodiversity, and identifies biological units important for conservation or medical consequence. African and forest biota have both received relatively little attention with regard to understanding their diversification, although one possible mechanism is that this has been driven by historical climate change. To investigate this, we implemented a standard population genetics approach along with Approximate Bayesian Computation, using sequence data from two exon‐primed intron‐crossing ( Although biodiversity has not been characterized for the majority of taxonomic groups, it is clear that the forests of Africa hold a vast amount of species diversity and endemism with important areas including the Guineo‐Congolian rainforest in West and Central Africa, the montane forests of the Albertine rift in Central Africa and the Eastern Arc mountains and coastal forests of Kenya and Tanzania in East Africa , which is a primary vector for several arthropod‐borne viruses (arboviruses) including dengue (DENV), yellow fever (YFV), chikungunya (CHIKV), Venezuelan Equine Encephalitis (VEE), and Zika (ZIKAV)  (YFV), Ae. africanus  (Babanki (BBKV), Bouboui (BOUV), CHIK, Rift Valley Fever (RVFV), YFV, and ZIKAV), and Aedes hansfordi  (RVFV) , which although not currently known to transmit disease to humans, is the sister taxa of Ae. bromeliae (Aedes simpsoni complex)  Huang, , 1986 ye) Huang, .Aedes mosquitoes. We use a comparative phylogeographic approach to investigate the population histories of four species of Aedes (Stegomyia) mosquitoes—Ae. africanus, Ae. bromeliae, Ae. hansfordi, and Ae. lilii—and compare these to that previously reported for Ae. aegypti , Colocasia spp., and Dracaena spp. from Uganda (n = 153), Tanzania (n = 64), and the Republic of Benin (n = 67)  Figure . When po) Figure  and chec) Figure . As Ae. 2.2IDH2 (with primers 5′‐CAGCAGTGCGTTCTTTTTCC‐′3 and 5′‐CAGTATCGATGCCCTTGTGG‐′3) developed in Bennett et al. . The GenElute PCR Clean‐Up Kit  was used to purify PCR products. The BigDye Terminator v3.1 cycle sequencing kit was used to sequence the targeted regions in both forward and reverse directions on a 3,730 automated Sanger sequencer .DNA was extracted from larvae or the legs of the adult using a phenol–chloroform extraction method detailed in Surendran et al. . Mosquitt et al. , and RpLnd Weeks . PCR proenhoek's  universaPCR products were cloned when indels between homologous alleles within an individual obscured the sequence read in the direct sequence of a PCR product and when sequences had low haplotype phase probability. PCR products for cloning were generated with high fidelity MyFi polymerase , 1× MyFi Reaction Buffer, 0.8 μM forward and reverse primer, and 1–10 ng of template DNA. To reduce the formation of chimeras . For all sequence data, compound indels affecting less than 5% of the sequence alignment were removed to eliminate areas with uncertain sequence homology in Geneious v5.4.7   and Neighbour Similarity Score implemented in PhiPack  using the best available substitution models as chosen by jModelTest Figures .2.6w), and the number of segregating sites (S). Statistics to detect demographic expansion, Tajima's D and Fu's Fs, and genetic differentiation, FST, were generated in Arlequin v3.5 , Ae. africanus (n = 1), and Ae. hansfordi (n = 1) because their inclusion increased network complexity  using a Tamura and Nei substitution model with uniform rates across sites (Table Median‐joining networks were constructed for sequence datasets in NETWORK v4.6.1.3 (Fluxus Technology Ltd.). Reticulation within networks was resolved using the recommendations in Crandall and Templeton . Haplotyes Table .2.8COI clades within Ae. bromeliae, Ae. lilii, Ae. africanus, and Ae. aegypti were used in separate analyses. GMRF skyride plots were generated with 1,000,000,000 Markov chains sampled every 100,000 generations. Each sequence set was tested for conformity to a molecular clock model in MEGA6 using a Likelihood Ratio Test  skyride model   between populations. Posterior based error rates were calculated using the scenario choice and parameter values of the closest 500 simulated datasets to the observed data.Tanzanian populations represent East Africa, Ugandan populations represent Central Africa, and the Republic of Benin as West Africa. Maximum effective population sizes of African demes were set to cover a wide range  encompassing values based on estimates of Ɵ calculated in DNAsp v5 and LAMARC as described above. For both t et al. . These s33.1IDH2 for Ae. bromeliae (p = .001), Ae. hansfordi (p = .02), and Ae. africanus (p = .03) and at nuclear Rp30Lb for Ae. africanus (p = .01). These datasets were trimmed by approximately 20–50 bp to remove signals of recombination for phylogenetic analyses, giving a reduced sequence length of 357 bp at IDH2 for Ae. bromeliae with a nonsignificant homoplasy score (p = .28), 501 bp at IDH2 for Ae. hansfordi (p = .37), 426 bp at IDH2 for Ae. africanus (p = .21), and 252 bp at Rp30Lb for Ae. africanus (p = .26).No recombination was detected for the nuclear sequence datasets with the tests implemented in RDP4. However, significant homoplasy compatibility scores (φ) were detected at nuclear 3.2Aedes species have high levels of genetic diversity (Ɵ and π) and effective population sizes within Africa that are generally comparable across species . Genetic diversity (Ɵ and π) was lower in Benin than Tanzania and Uganda for Ae. lilii and Ae. aegypti (the only two species sampled in Benin) for all loci except COI of Ae. lilii. Tajima's D was significant and negative within Africa at one or more nuclear loci for all species except Ae. africanus . By comparison, the IDH2 locus had fewer significantly negative Tajima's D values, but they were present for the Africa‐wide analyses of Ae. lilii and Ae. aegypti as well as for Ugandan Ae. aegypti, Ae. bromeliae, and Ae. hansfordi. With the exception of Ae. lilii in Uganda, all Tajima's D values for mitochondrial COI were nonsignificant. Similarly, significant Fu's Fs values were seen in Africa and regional populations for all species including Ae. africanus  and Rp30Lb  and mitochondrial COI , somewhat higher than the values reported for Ae. aegypti previously . Significant genetic differences were also apparent between populations of Ae. lilii in Benin, West Africa, and Uganda, Central Africa at the COI gene  but not at nuclear loci. Similarly, structure between West and Central Africa was reported previously at Rp30Lb (FST = 0.013) and COI (FST = 0.075) for Ae. aegypti, in addition to structure between East and West African Ae. aegypti at nuclear IDH2 (FST = 0.015), Rp30Lb (FST = 0.036), and mitochondrial COI (FST = 0.252)  and nuclear IDH2  and Rp30Lb  for Ae. bromeliae. In addition, significant IBD was detected at mitochondrial COI for Ae. lilii  and Ae. africanus . All other tests for IBD were nonsignificant, as reported for these loci for Ae. aegypti previously  was detected at the 3.3COI from Ae. bromeliae revealed a strong geographic structure with two major haplogroups representing Uganda, Central Africa, and Tanzania, East Africa  and mitochondrial COI data in the majority of cases. However, a signal of demographic expansion was detected on analysis of COI haplogroup 2 for Ae. lilii and for Ae. africanus with an increase in effective population size from approximately 40,000–2,000,000 from 100,000 years ago and an increase in effective population size from 30,000 to 1,000,000 from 70,000 years ago, respectively.For all species, Bayesian GMRF skyride analysis did not detect a change in population size on analysis of nuclear loci  analysis of nuclear sequence data, that is, 4Ae. aegypti were found to fit a pattern of historical lineage diversification and admixture followed by recent population structuring , few are shared between these and the coastal forests of East Africa and the Eastern Arc mountain ranges (Couvreur et al., Annonaceae flowering plant species of the Guineo‐Congolian and East African forests reveal multiple origins of this rainforest restricted taxon throughout the last 8–33 million years (Couvreur et al., Aedes mosquitoes, including within the Simpsoni Complex, whose geographical distributions are currently divided across the African continent (Bennett et al., Ae. bromeliae from Tanzania and Uganda and Ae. lilii from Benin and Uganda using ABC analysis suggests a more recent connection of forest habitats across the African continent. This is in keeping with pollen and soil data suggesting that forest cover was at its maximum during the early Holocene era beginning 11,000 years ago (Castañeda et al., Aedes mosquitoes throughout their evolutionary history, generating both inter‐ and intraspecific genetic diversity.In relation to this, it has been suggested that cyclic isolation of the east coast and Eastern Arc mountain forests from the Guineo‐Congolian block may be one factor explaining the high levels of endemicity in East Africa (Couvreur et al., 5Aedes species in this and previous studies (Bennett et al., The genetic variation observed within and between Aedes species are characterized by the admixture of genetically divergent lineages and exhibit high effective population sizes. This high level of genetic diversity indicates a high adaptive potential (Facon et al. Aedes mosquitoes will follow in the evolutionary tracks of Ae. aegypti and advance into domestic environments, increasing arboviral disease transmission in the future.In this and previous studies (Bennett et al., None declared.MF183226–MF183441 and MF183443–MF183893. Sampling locations and/or online‐only appendices uploaded as online supplemental material.DNA sequences: GenBank Accession nos COI gene. K.L.B. wrote the manuscript with help from C.W. All authors contributed to comments and approved the final version.C.W. and K.L.B. designed the study. K.L.B., M.K, F.S., R.D., G.M., J.L., Y‐M.L., and C.W. carried out the fieldwork and contributed samples. K.L.B. performed the research and analyzed the data under the supervision of C.W. Y.‐M.L. contributed to sequencing of the  Click here for additional data file. Click here for additional data file. Click here for additional data file. Click here for additional data file. Click here for additional data file. Click here for additional data file. Click here for additional data file.


Ichthyoelephas longirostris (Characiformes: prochilodontidae) is a short-distance migratory species endemic to Colombia. This study developed for the first time a set of 24 polymorphic microsatellite loci by using next-generation sequencing to explore the population genetics of this commercially exploited species. Nineteen of these loci were used to assess the genetic diversity and structure of 193 I. longirostris in three Colombian rivers of the Magdalena basin. Results showed that a single genetic stock circulates in the Cauca River, whereas other single different genetic stock is present in the rivers Samaná Norte and San Bartolomé-Magdalena. Additionally, I. longirostris was genetically different among and across rivers. This first insight about the population genetic structure of I. longirostris is crucial for monitoring the genetic diversity, the management and conservation of its populations, and complement the genetic studies in Prochilodontidae.The Neotropical freshwater fish  Ichthyoelephas longirostris  is a Colombian endemic fish of importance in the commercial and subsistence fisheries. This freshwater fish is a member of the family Prochilodontidae, which comprises three genera  and 21 species that occur in the major river basins of South America , infrequent catches, disappearance in some floodplain lakes and rivers (Ranchería) and the habitat degradation by anthropogenic activities , and the Samaná Norte and San Bartolomé rivers (Porvenir II project). All these samples were provided by Integral S.A., through two scientific cooperation agreements . In the Cauca River, the samples came from three out of eight sections sampled .DNA isolations were performed with the commercial kit GeneJET DNA purification (Thermo Scientific) following the manufacturer’s instructions. To identify microsatellite regions and develop primers to amplify them, low-coverage genome sequencing was carried out with the Illumina MiSeq v2 instrument using the Nextera library preparation kits. This sequencing process generated paired-end reads of 250 bases that were cleaned using Prinseq-lite v0.20.4  to elimiI. longirostris. Preliminary tests of standard PCR conditions  run at 100 volts for 45 min and visualized by silver-stain. Polymorphic loci were selected based on criteria of amplification in all samples, band resolution, specificity, size (from 100 to 400 bp) and ability to detect heterozygotes in the different samples analyzed. Then, a set of 24 polymorphic microsatellite loci that met these criteria and amplified consistently were selected and fluorescently labelled for further genotyping of 28 samples of I. longirostris. Finally, a subset of 19 loci were selected to evaluate the genetic diversity and genetic structure in 193 samples from three Colombian rivers.A total of 40 microsatellites were selected for optimization and polymorphism analysis in nditions  were car2. The PCR amplifications were performed on a thermocycler T100 (BioRad) with an initial denaturation step of 95°C for 3 min, followed by 32 cycles consisting of a denaturation step of 90°C for 22 s and an annealing step of 57°C for 16 s. The extension step and a final elongation were absent in this thermal profile. Finally, the PCR products were submitted to electrophoresis on an automated sequencer ABI 3730 XL (Applied Biosystems) using LIZ500 (Applied Biosystems) as internal molecular size. Allelic fragments were denoted according their molecular size and scored using GeneMapper 4.0 (Applied Biosystems).PCR reactions in volumes of 10 μl containing final concentrations of 1×buffer (Invitrogen), 2–4 ng/μl of template DNA, 2.5% formamide (Sigma) 0.35 pmoles/μl labelled forward primer , 0.5 pmoles/μl reverse primer (Macrogen), 0.2 mM dNTPs (Thermo Scientific), 0.05 U/μl Platinum™ Taq DNA Polymerase (Invitrogen) and 2.5 mM MgClHO) and expected (HE) heterozygosities were performed using Arlequin v.3.5.2.2  was calculated using ΔK ad hoc statistic (F’ST (Dest (http://www.ipez.es/RWizard/).Genetic diversity was estimated by calculating the average number of alleles per locus, observed and expected average heterozygosities and fixation index. Non-geographical genetic differentiation among samples was tested the Bayesian analysis of population partitioning using Structure v.2.3.4 . This antatistic  with STRtatistic . Resultstatistic , settingtatistic . In addiic  of 0.669. Additionally, allelic frequencies of 20 loci were concordant with Hardy-Weinberg and linkage equilibria after sequential Bonferroni correction and no evidence of null alleles or scoring errors were detected by Micro-checker. Moreover, the PIC values ranged from 0.375 to 0.871 (average: 0.733) indicating that these markers are highly informative  and Samaná Norte  followed by Cauca S1 , Cauca S6  and San Bartolomé-Magdalena .In the rivers, 3 of the 19 loci  departed from Hardy-Weinberg equilibrium expectations in all samples evaluated . The aveK = 2), one predominantly in the Cauca River and the other one in the rivers San Bartolomé and Samaná Norte  was examined to compare it with the other approaches (I. longirostris among (FST(0.001) = 0.010; P = 0.000) rivers. In addition, within the Cauca River, the discriminant analysis of principal components  or five stocks .The three approaches for measuring population genetic structure generated different, but non-excluding results. STRUCTURE revealed only two stocks related to the highest hierarchical grouping, which is concordant with other studies that show that this software is limited for the nested fine substructure detection  and for P. magdalenae  stocks in the examined sites. This information, previous to the hydropower station construction, is crucial for monitoring the genetic diversity for management and conservation of this species as well as for complementing the genetic studies in Prochilodontidae.In summary, this study developed the first set of polymorphic microsatellite loci for population genetics of 10.7717/peerj.2419/supp-1Figure S1Click here for additional data file.10.7717/peerj.2419/supp-2Data S1Click here for additional data file.


ALS) is increasingly recognized as a genetic disease. There is no consensus, however, as to the role of genetic testing in the care of the ALS patient.Amyotrophic lateral sclerosis   were offered testing, while a minority of respondents with sporadic ALS (sALS)  were offered testing (p = .00001). Comparison of mean test experience scores between groups revealed that respondents with fALS were no more likely to report a favorable experience with genetic testing than those with sALS (p = .51). Respondents who saw a genetic counselor did not have significantly different test experience scores, compared to those who did not (p = .14). In addition, no differences in test experience scores were observed between those who received positive or negative genetic test results (p = .98).Among 449 survey respondents, 156 (34.7%) were offered testing and 105 of 156 (67.3%) completed testing. The majority of respondents with familial ALS found value in clinical genetic testing.These data indicate that patients with  However, there is no consensus as to the role of genetic testing in the care of the ALS patient, and US practice guidelines do not provide recommendations for the offer of genetic testing  is increasingly recognized as a genetic disease. The genetic basis of ~70% of familial ALS (fALS) and ~10% of sporadic ALS (sALS) is now understood  from The Ohio State University Wexner Medical Center, Columbus, Ohio. Specific, separate ethics committee review and/or approval for this study was not indicated per the IRB review.®. Patients affected with ALS were eligible to participate; caregivers were permitted to assist, if needed. One survey was completed per respondent. Eligible participants were identified and contacted through the United States Centers for Disease Control–Agency for Toxic Substance and Disease Registry (CDC–ATSDR). The CDC–ATSDR advertised and distributed the survey link via a one‐time email announcement, after approval from the ATSDR Committee. All results were tabulated and exported in .xls and/or .csv format for analysis.Study subjects completed an anonymous online survey using the survey engine SurveyMonkeyThe survey instrument was described in detail previously . Associations between groups and binary characteristics were studied using proportions and Fisher's exact test. A two‐tailed 3p = .00001). Approximately half of respondents with a positive family history of ALS  underwent genetic testing; 79 of 404 (19.5%) of all sALS respondents underwent testing (p = .05). A minority of respondents (12.5%) indicated contact with a genetic counselor, and these respondents were much more likely to be offered genetic testing (p = .00001) than those who did not indicate contact with a genetic counselor. Respondents with fALS were more likely to have seen a genetic counselor than respondents with sALS (p = .0082).A positive family history of ALS (fALS) was reported by 45 of 449 (8.5%) of respondents; 341 of 449 (75.9%) reported a negative family history of ALS (sALS); and 63 of 449 (14.0%) indicated they did not know their family history status for ALS. Among 449 survey respondents who completed the portion of the survey addressing personal experience with genetic testing, 156 (34.7%) were offered testing, and 105 of 156 (67.3%) completed testing. The majority of fALS respondents  were offered testing, while a minority of sALS respondents  were offered testing  was recalled and reported by 75 respondents (see Table n = 68) and their families  with 83.3% (n = 80) agreeing that other persons with ALS should consider genetic testing  revealed that respondents who answered negatively to one item were also very likely to answer negatively to other items, confirming that a small group of respondents had a generally negative genetic testing experience. Of these six respondents, none knew the outcome of their testing: Four reported that they were not informed of their result, one indicated that they did not know their result and did not know if they were told, and one was informed of the result but could not recall it.An overall mean test experience score was calculated for each respondent. Only six respondents  had negative or neutral mean test experience scores. Rank correlation analysis of individual respondents who completed at least two Likert scale items (p = .51). Respondents who indicated contact with a genetic counselor had a similar test experience as those respondents who did not indicate contact with a genetic counselor (p = .14). In addition, no differences in test experience scores were observed between those who received positive or negative genetic test results (p = .98).Comparison of mean Likert scale scores between groups revealed that respondents with fALS or sALS reported similar experiences with genetic testing, including those sALS respondents who tested positive despite no family history  of respondents had negative or neutral mean test experience scores. Each of these six reported that they were not told or could not recall the outcome of their testing, suggesting that receipt and recollection of results is a necessary component of a positive genetic testing experience. The majority of respondents indicated receiving results during an office visit, suggesting a traditional face‐to‐face results disclosure with a clinician such as a neurologist or genetic counselor may in fact support a positive test experience, although specific methods  of results disclosure were not directly compared in this survey.Mean test experience scores of respondents with a family history of ALS were not statistically different than those without a family history, indicating that sALS patients were equally satisfied with genetic testing as fALS patients. This satisfaction score also includes the few respondents (5) who tested positive for a mutation despite no family history of the disease. Although the proportion of sALS cases attributable to identifiable, highly penetrant mutations is increasing with continued gene discovery, the practice of offering genetic testing to sALS patients remains inconsistent. The European Federation of Neurological Societies (EFNS) issued guidelines indicating that genetic testing should not be performed in “cases with sporadic ALS with a typical classical ALS phenotype” (Andersen et al., We also examined whether seeing a genetic counselor was associated with a more positive test experience. Survey data collected on ALS patient attitudes indicated that patients who reported contact with a genetic counselor had more positive attitudes regarding the potential benefits of genetic testing for patients, the medical community, and society (Wagner et al., Respondents who tested positive for an ALS mutation did not have significantly lower test experience scores, indicating that “getting bad news” did not negatively impact their perception of the testing process. This finding appears to be in concordance with the published literature suggesting that increased post‐test distress in persons undergoing genetic testing is usually transient and not clinically significant (Lerman, Croyle, Tercyak, & Hamann, The lowest test satisfaction scores were observed in items related to implications for family members, including “My doctor/care team explained what my result means for my children/family members,” and “the results of my genetic testing were useful to my family members.” More thorough discussion of the implications of test results for family members may be beneficial for ALS patients and their families, as genetic testing results have implications for family members and may result in psychological and emotional effects for the extended family (Clarimon & Kulisevsky, 4.1The study survey was sent to ATSDR registrants via a one‐time email announcement per CDC protocol. Previous survey research utilizing the registry for survey distribution reported a similar response rate of 11.5% (Malek et al., Responses relied on patient recall of discussions by healthcare providers. We had no way of objectively assessing the quality of information that respondents received during the course of genetic testing and genetic counseling. As ALS patients may rely solely or primarily on the healthcare team for information, it seems possible that they “don't know what they don't know” and therefore may tend report satisfaction with poor and/or incomplete information. This could have biased test experiences in the positive direction.p‐value was utilized during data analysis.Finally, numerous univariate tests were performed without correction for family‐wise error rate, and some statistically significant findings may be due to chance alone. However, due to the exploratory nature of this research, a less stringent 4.2The finding that patients with both fALS and sALS report satisfaction and find value in genetic testing supports the practice of offering testing to all ALS patients. Patient‐reported outcomes are increasingly recognized as a critical component in shaping medical management policy (Stefanou & Amygdalos, 5Current US best practice guidelines for ALS care do not provide recommendations for the offer of genetic testing. Available data suggest that neurologists are likely to offer genetic testing to patients with familial ALS, but the offer of genetic testing to patients with sporadic ALS is inconsistent. Our survey of a national ALS registry indicates that ALS patients find value in genetic testing, whether or not they have a family history of the condition. Genetic testing may help ALS patients to understand the cause of their condition, allow more accurate risk assessment and testing of family members, and facilitate clinical trial inclusion, including genotype‐specific treatments. As the genetic basis of ALS is further elucidated, genetic testing and counseling will become an increasingly vital component of ALS care.Authors Karin N. Wagner, Dr. Haikady Nagaraja, Dawn C. Allain, Dr. Adam Quick, Dr. Stephen Kolb, and Jennifer Roggenbuck report no disclosures. Click here for additional data file.


According to that, mental imagery could lead to (1) a superior performance in object recognition and associative learning if these objects are imagery-congruent  and to (2) an inferior performance if these objects are imagery-incongruent. In the first experiment, we used a static environment and tested associative learning. In the second experiment, subjects encoded object information in a dynamic environment by means of a virtual sewer system. Our results demonstrate that subjects who received a role adoption task  performed better when imagery-congruent objects were used and worse when imagery-incongruent objects were used. We finally discuss our findings also with respect to alternative accounts and plead for a multi-methodological approach for future research in order to solve this issue.The role of mental imagery has been puzzling researchers for more than two millennia. Both positive and negative effects of mental imagery on information processing have been discussed. The aim of this work was to examine how mental imagery affects object recognition and associative learning. Based on different perceptual and cognitive accounts we tested our  By systematically manipulating landmark types and role adoption tasks, we will show that the effect of mental imagery can influence subsequent encoding and recognition processes of landmarks. Whether this effect facilitates or impedes these cognitive abilities depends on the semantic congruence between landmark and adopted role, as we shall later see. Our findings suggest that we need to be cautious and that more research is needed in order to understand when detrimental effects of mental imagery occur.“You are what you eat”–this well-known proverb (directed to our dietary habits), in a more abstract sense, stands for the idea that our actions influence our body and our personality. At the same time, we see that our thoughts also have an influence on us such as it is the case with guided imagery. It is a guided form of mental imagery that can, for example, be found in clinical settings. So, what if you become as good as you are told you are? What if imagining of being a rat in the sewer, improves the way you perceive and recognize conceptually related objects? Focusing on spatial cognition, this work examines the role of mental imagery and its influence on the recognition of salient objects along a route, so-called In the following, we will first present our theoretical foundation before we derive our hypothesis from that. For readability purposes, we will divide them into three subsections: (1) mental imagery & perception, (2) mental imagery & semantic salience and (3) mental imagery & memory.Theaetetus, Plato developed the wax tablet metaphor which described our memories as the imprinted information on a wax tablet  This very idea can also be found in Neisser’s theoretical framework who, however, opposed both Kosslyn and Pylyshyn with his alternative approach . He insiNow it is time to mention the other side: mental imagery can also hinder subsequent object recognition. Neisser pointed to studies that reported facilitated identification of capital letters after imagining them and hindered identification when these letters were presented in lower case . PetersoThe problem with the aforementioned findings is that all of them were conducted with a small spatio-temporal window in mind. However, it was Neisser who already emphasized (1) the importance of movement and space during the exploration phase of the perceptual cycle model and (2) the abstractness of schemata . TherefoSpatial orientation and wayfinding has been well studied , 14. In In our current study, we want to extend the line of research, which aims to understand how mental imagery affects landmark information processing. Landmarks, in general, are defined as objects , which stand out from the environment and can therefore act as reference points . Of courIn principle, the extent to which a landmark stands out from its environment is called landmark salience. However, it should not only be seen as the property of certain spatial features. Based on a trilateral relation between the observer’s point of view, the feature itself and the surrounding environment, Caduff et al.  describevisual salience describes visual object properties . Accordingly, the salience of a bright shining big white sphere in a dark environment would be classified as high. Second, the term structural salience stands for navigation-related features . For example, a statue in the center of a roundabout or an object in front of an intersection in direction of turn would be considered as having a high structural salience  ANOVA.F = 110.909, p < .001, M = 82.78%, SEM = 1.37) could more easily be recognized than imagery-congruent landmarks .The ANOVA revealed a significant main effect of landmark type on recognition performance, F < 1), there was a significant role adoption task x landmark type interaction, F = 5.757, p = .005, M = 65.83%, SEM = 3.05) performed significantly better than the control condition  when imagery-congruent landmarks were used, p = .023. Additionally, rats  were almost on the same level as squobbles. All groups were significantly better than chance level (50%).While there was no significant main effect of role adoption task , = 11.663, p = .001, p = .006) and hindered the recognition of imagery-incongruent landmarks (p = .051).Afterwards, since there were no differences between ‘rats’ and ‘squobbles’, we pooled the data and compared the performance between those who received a role adoption task and those who did not (same as in Experiment 1). The results  of the 2F = 12.497, p = .001. Subjects in the imagery-congruent landmark condition  were significantly slower than subjects in the imagery-incongruent landmark condition .Lastly, we analyzed the response time data. However, there was only a significant main effect of landmark type, In Experiment 2, we let subjects encode landmark information in a dynamic environment . After that, we tested whether subjects could recognize the landmarks they previously had seen. Our results show that guided mental imagery leads to superior recognition of imagery-congruent landmarks and worse recognition of imagery-incongruent landmarks. These findings are in line with our hypothesis and further strengthen our results of the previous experiment. In fact, it shows that even in a more difficult environment (dark dynamic scene) where people can freely look around, the role adoption task can facilitate or impair landmark recognition performance.In contrast to Experiment 1, we did not find support for our hypothesis in terms of subject’s response times. Subjects were generally faster in recognizing imagery-incongruent landmarks . One simple reason for that might have been that the content of these landmarks were clearer. For example, it might be easier to detect an elephant compared to a valve. Nevertheless, our results demonstrate the interaction effect in object recognition despite these facts.Assessing recognition performance can be a sensitive tool for investigating the role of landmarks in wayfinding in general. They are valuable because in environments which feature landmarks, associations between movement decisions and visual cues can be learned , 33. TheIn addition to the role adoption task of Experiment 1, we created another one that consisted of a fictitious character. We did so in order to control for prior knowledge. This is also critically discussed by Shapiro , who conImagery-Induced Interaction Hypothesis (3IH). On a general level, this hypothesis states that mental imagery influences the encoding of object information positively or negatively, depending on the semantic congruence of object and imagined concept. We tested this in a series of two experiments. The first experiment consisted of a static scene. The second experiment was conducted in a dynamic scene by using a virtual environment. In both experiments, a role adoption task (by means of guided imagery) was given to subjects of the experimental group. Additionally, we used objects (landmarks) that were either imagery-congruent or imagery-incongruent. Summarized, our results corroborate the 3IH.The current study sought to examine the role of mental imagery within spatial cognition. Based on our theoretical foundation we proposed the How can we explain this effect? In the beginning of this paper we presented the controversy around mental imagery and discussed three different aspects that built our theoretical foundation. First, mental imagery can change how we perceive our environment. For example, priming can facilitate object recognition, at the same time anticipating one object can lead to overlooking another one . Second, mental imagery can change the extent to which objects become important and meaningful to us . Finally, mental imagery can influence to what extent information is being encoded and retrieved later on . Obviously, all these three elements interact with each other and cannot account for our results separately. The reason for introducing this trichotomy was to structure this paper in a manner that corresponds to findings in different research areas. Nevertheless, these findings underline our hypothesis.One could also think of an analogy in the human visual system. After a while in the dark, rods and even more cones increase their sensitivity (dark adaption) due to visual pigment regeneration . TurningOther accounts can be derived from neuroimaging studies. For example, the retrosplenial/parietal-occipital sulcus region plays an important role in integrating path and landmark information . The samWe would like to encourage other researchers to further investigate this topic with respect to perceptual and cognitive accounts in order to understand the mechanisms behind such an interaction to the fullest.theory of mind, self-efficacy, or embodiment cannot explain our findings very well. The term theory of mind stands for our ability to attribute certain mental states to others as well as oneself and is, for example, impaired in children with autism [Alternative constructs such as h autism . This abThis leads to the idea that self-efficacy could have played a role. Self-efficacy describes the case where expectations of personal efficacy can predict performance . HoweverCould one of those mechanisms be related to embodiment, which is the way we experience a certain body as ours? We believe that this is not the case. In the well-known rubber-hand illusion, for example, synchronous visuo-tactile stimulation of an artificial hand (rubber-hand) and one’s own hand leads to tactile sensations referred to the alien limb . TechnolIt is possible that all three, theory of mind, self-efficacy and embodiment, are crucial when it comes to the role adoption tasks in spatial cognition. However, there is hardly any research available in the spatial cognition literature and not much is known about the respective contributions to such an effect, especially in the context of wayfinding.There is one noteworthy aspect that, at first glance, could be seen as a limitation. Imagery-incongruent landmarks had a higher visual salience . The reason for that is that subjects rated such landmarks as imagery-congruent that were “sewer-themed” . This should not be a problem in general as long as they can really focus on the landmark. However, in Experiment 2 they were virtually guided through the sewer system which makes it more difficult to get a grasp of the exact object properties. This can explain why there was a main effect of landmark type only in Experiment 2. Despite this “visual strength” or visual salience of imagery-incongruent landmarks, our results demonstrated that mental imagery can overcome such object properties. Subjects who used mental imagery (role adoption task) were still better in the recognition of imagery-congruent landmarks and worse in imagery-incongruent landmarks compared to subjects who did not conduct a role adoption task. Therefore, we believe that this strengthens our findings even more.Nevertheless, this being one of the first studies in the field, future studies could link mental imagery more closely to spatial cognition. For example, experiments investigating survey knowledge could shed light on other aspects of spatial cognition that were not in the center of this study. Another aspect of mental imagery can be investigated with respect to mental simulation of routes. For example, one line of research could show that such mental simulations are temporally compressed . The autLastly, it may be seen as a limitation that our experiment was conducted in a virtual environment. Given the fact that walking is inherently linked to bodily sensations [i.e., during locomotion; In sum, our study showed that mental imagery could indeed play a role in landmark recognition. Still, a lot of research has to be conducted in order to understand the mechanisms behind such an effect. We suggest that the effect we found in our study is somehow linked to processes involving perceptual and higher cognitive mechanisms as well as working memory. Therefore, we plead for a multi-methodological approach for future research.S1 Fig(PDF)Click here for additional data file.S1 Text(PDF)Click here for additional data file.S1 Data files(ZIP)Click here for additional data file.


FST) and haplotype networks to infer intraspecific population connectivity between vent sites. Furthermore, we evaluate contrasting scenarios of demographic population expansion/decline versus constant population size . Our results indicate high diversity, population expansion and high connectivity of all copepod populations in all oceans. For example, haplotype diversity values range from 0.89 to 1 and FST values range from 0.001 to 0.11 for Stygiopontius species from the Central Indian Ridge, Mid Atlantic Ridge, East Pacific Rise, and Eastern Lau Spreading Center. We suggest that great abundance and high site occupancy by these species favor high genetic diversity. Two scenarios both showed similarly high connectivity: fast spreading centers with little distance between vent fields and slow spreading centers with greater distance between fields. This unexpected result may be due to some distinct frequency of natural disturbance events, or to aspects of individual life histories that affect realized rates of dispersal. However, our statistical performance analyses showed that at least 100 genomic regions should be sequenced to ensure accurate estimates of migration rate. Our demography parameters demonstrate that dirivultid populations are generally large and continuously undergoing population growth. Benthic and pelagic species abundance data support these findings.Communities in spatially fragmented deep-sea hydrothermal vents rich in polymetallic sulfides could soon face major disturbance events due to deep-sea mineral mining, such that unraveling patterns of gene flow between hydrothermal vent populations will be an important step in the development of conservation policies. Indeed, the time required by deep-sea populations to recover following habitat perturbations depends both on the direction of gene flow and the number of migrants available for re-colonization after disturbance. In this study we compare nine dirivultid copepod species across various geological settings. We analyze partial nucleotide sequences of the mtCOI gene and use divergence estimates ( Deep-sea hydrothermal vents are island habitats that occur globally along mid-ocean ridges, back-arc basins and island arcs. Tectonic events and volcanic eruptions make them unstable in space and time. The emergence of hot, sulfide- and mineral-rich hydrothermal fluids also characterizes this extreme ecosystem. Most vent macrofauna and several meiofauna species are restricted to the vent environment, where the sulfide-rich fluids nourish chemolithoautotrophic bacteria –3. TheseThe rich sulfide mineral deposits at deep-sea hydrothermal vents could make this unique ecosystem a target area for the mining industry in the near future . Seabed FST values) can provide indirect estimates of the extent to which the evolution of subpopulations is influenced by migration (genetic connectivity) [Genetic analyses of so far undisturbed populations can greatly enhance our understanding of vent species population biology, and can allow predictions of recolonization and recovery. Recolonization potential can be described using the conceptual framework of population connectivity, a concept that describes the extent to what subpopulations exchange migrants. While a direct assessment of migration rates requires model-based demographic analyses of population genomic datasets –11, divectivity) . In the ctivity)  and theiGenetic analyses have been previously applied for various macrofaunal and megafaunal species representing different superordinate taxa  in distinct geographical settings, including the Mid Atlantic Ridge, East Pacific Rise, Eastern Lau Spreading Centre, and Central Indian Ridge . These sThe underlying mechanisms of high connectivity (migration capability), high genetic diversity and expansive population growth across different species and taxa are not fully understood. On one hand, life history traits must play an important role, since they allow for rapid population growth and/or guarantee long-term planktonic larval duration as connectivity between invertebrate populations is often achieved by passive larval drift. On the other hand, passive larval drift is influenced by abiotic characteristics such as plume height or ocean currents . FurtherVolcanic eruption and vent field frequency differ among hydrothermal vents at distinct spreading centers along mid-ocean ridges, back-arc basins and at volcanic arcs. Vent field frequency increases with increased spreading rate . At the Disentangling to what extent geographic and geologic characteristics of vent fields  can shape population genetic structure of invertebrate species, such as diversity, demography, connectivity, is indeed important for conservation plans; however, this remains challenging, not in the least because endemicity level per biogeographic province among vent macrofauna species is typically high (95%) and no vent species occurs circumglobally . If suchAlvinella pompejana), they are the most abundant animals. They are typically free-living among aggregations of foundation species such as Pompeii worms, tubeworms, snails, or bivalves [Stygiopontius and Aphotopontius occur at vents worldwide [In the absence of a circumglobally occurring model species that allow analyses of population structure at different vent regions, this study instead comparatively explores patterns of genetic diversity, demography, and divergence in populations of very closely related copepod species with similar life history traits across different oceans. We investigate nine species belonging to two genera within the vent endemic family Dirivultidae, a taxon with more than 50 described species. Dirivultidae is the one of the most species-rich invertebrate family at hydrothermal vents . They arbivalves , 34. Somorldwide ; they tyorldwide , 37. Nauorldwide , 38.We have selected 9 dirivultid species from two genera from four areas: fast-spreading Eastern Lau Spreading Center (ELSC) and East Pacific Rise (9°N EPR), and slow-spreading Mid Atlantic Ridge , and Central Indian Ridge (CIR). We morphologically identify the species and use partial nucleotide sequences of the mitochondrial gene Cytochrome Oxidase I  to estimate genetic diversity and demographic characteristics within populations. In addition, divergence between subpopulations is assessed to provide an estimate of genetic connectivity, i.e., describing to what extent gene flow affects evolutionary processes within populations. We further link genetic data to species abundance data in the benthos and in the pelagial, to the end of discussing our genetic results in the light of actual species population size and dispersal potential. Our analyses provide a basis for discussing whether the frequencies of vent fields and volcanic eruptions can shape the genetic composition of populations. Furthermore, based on observed population genetic patterns in dirivultid species in different oceans, we carry out statistical performance analyses and lay out a conceptual basis for future population genomic surveys in these populations. Finally, we use our data to estimate potential mining impact on this faunal group.Alvin on the East Pacific Rise (EPR) and the Guaymas Basin (GB) in the East Pacific, by the submersible Nautile on the Mid-Atlantic Ridge (MAR) in the Atlantic, by the ROV Jason the Eastern Lau Spreading Center (ELSC) in the West Pacific, and by the ROV Kiel 6000 on the Central Indian Ridge (CIR) in the Indian Ocean  and the interactive map from the interridge data base (http://vents-data.interridge.org/maps). Information on dive number, longitude, latitude and depth of sites where specimens were collected, as well as inter-site distances, is provided in A map of the sites was created using GeoMapApp , and for S. hispidulus (EPR) from Gollner et al. (2011) [In this study, we morphologically identified and produced new sequences from the following species: . (2011)   and HCOI (50-TAA ACT TCA GGG TGA CCA AAA AAT CA-30) [http://www.technelysium.com.au). Contaminations were extracted using BLAST against a non-copepod COI sequence database. Contig assembly of forward and reverse reads was completed using the Cap-Contig assembly function in BioEdit. All GenBank accession numbers, including newly generated and published ones from Gollner et al. [The same animals that were morphologically identified were used for our mtCOI study. We applied the same methods for all specimens, published sequences and newly analyzed specimens alike. DNA was extracted from single specimens in 40 microliters of chelex  or 100 microl DNeasy Tissue Extraction Kit . We discovered no difference in PCR amplification success rate between these two extraction methods. Partial cytrochrome c oxidase subunit I (mtCOI) was PCR-amplified using universal primers LCOI (T CA-30)  and GE HT CA-30)  and Chror et al.  are giveAmeira species). Species-specific alignments where then used to estimate p-distances between COI nucleotide sequences using the software MEGA 5.1 [http://popart.otago.ac.nz) and the statistical parsimony network implementation that uses TCS [The dataset used was a combination of the newly produced sequences and those from Gollner et al. 2011 . SpeciesMEGA 5.1 . This wauses TCS , 47. PopPopulation specific parameters reflecting diversity and demography at the mtCOI gene were estimated using the software DnaSPv5  for popuDemographic history of populations was analyzed using the software DnaSPv5  to estimfastsimcoal2 [-8 events/bp/generation. For each model and for each number of simulated fragments we simulated 20 pseudo-observed datasets that were used as input to the maximum-likelihood parameter estimation procedure implemented in fastsimcoal2 [2 of Stygiopontius hispidulus, Aphotopontius mammillatus and A. limatulus (see 2. Time of divergence (TDIV) was set to 1000 and 100000 generations. TDIV of 1000 is an estimate based on presumably high generation time of copepods and recent bottleneck events due to major volcanic eruptions. For TDIV = 1000 we assume ~33 copepod generations per year in an environment with temperatures of 20°C [DIV = 100 000 is an estimate based on lower generation time . Simulations were completed with simcoal2  under twsimcoal2 . One frasimcoal2 . In both of 20°C , and maj of 20°C . TDIV = of 10°C; ) and majStygiopontius, and from 23 to 29% within the genus Aphotopontius. Intraspecific p-distance estimates were low for Stygiopontius spec.  and Aphotopontius spec. (0.8–1.2%)  from a cluster. S. sp. 1 nov., S. sp.2 nov. from CIR from a cluster with S. pectinatus from MAR . Haplotype networks of Stygiopontius spp.  revealed the same entities and there was no evidence for cryptic species . Uncorre.8–1.2%) . The maxius spp.  and Aphoius spp.  indicatec. 0.5–1,% and Aph0 bp reveStygiopontius spp., ranging from 0.893 to 1.00 and nucleotid diversity ranged from <0.001 to 0.01   . The ratStygiopontius spp. from all biogeographic regions. Some of the more extreme values were seen for S. pectinatus from MAR , for S. brevispina from ELSC , and for S. hispidulus from EPR . Similarly, Fu’s F values were significantly negative, and R2 and raggedness index (rg) were low for Stygiopontius species, indicating sudden expansion. Similar values  were observed for Aphotopontius species  for S. pectinatus from MAR sites (sites > 300 km apart). At ELSC sites, FST was 0.02 (p = 0.15) for S. lauensis and 0.11 (p = 0.02) for S. brevispina; this includes samples from three vent sites separated by 80 and 140 km. In contrast, EPR vent sites were separated by just a few hundred meters and at most by 5 km, so estimates for S. hispidulus were only -0.001 (p = 0.47)  also reflected by the absence of a relationship between network topology and sampling locality.AMOVA-based estimates of divergence between populations were made for species that where found at more than one locality . We esti = 0.47) . Low div7 p = 0.0 for S. pDIV) of 100000 and 1000 generations. Results for TDIV = 100000 show that accurate estimates of the migration rate and the population size can be achieved when the number of simulated fragments is at least 100 both for the symmetrical and asymmetrical migration models. The reason for the low performance in re-estimating the age of the split between the two populations in the asymmetrical migration rate model can be explained by the fact that the time to the most recent common ancestor in the whole sample is probably consistently younger than the age of the population split (100000 generations). Analyses with a divergence time of 1000 generations provided similar results: using at least 100 fragments gives good estimates on migration rate although precision in the re-estimation of the migration rate decreased compared to the case where TDIV = 100000 . S. pectinatus from MAR is known to occur in high abundances in close association with the shrimp Rimicaris exoculata (pers. obs. Florence Pradillion IFREMER). To conclude, Dirivultids maintain numerous colonies with locally abundant populations, characterized genetically by high haplotype diversity.High haplotype diversity of dirivultid copepods corresponds to high site occupancy, similar to what was observed for macrofauna species from EPR . Copepodr et al. ), and thLarge dirivultid population sizes might be a result of high resource availability at hydrothermal vents. Most dirivultid copepods are deposit feeders and/or graze on the abundant bacterial mats associated with hydrothermal vents , 60, 61.S. pectinatus is found associated with the shrimp Rimicaris exoculata. It lives around and inside the gills of the shrimp and likely grazes on the bacteria farmed there [R. exoculata ranged from 0.69 to 0.82 for sample populations with more than 11 individuals, and was similarly high as in S. pectinatus, with values of with 0.94 to 1 [S. hispidulus (hd 0.92–1) is typically found in very high abundance and in association with the pompei worm Alvinella pompejana [S. lauensis (Hd 0.98–1) was very abundant amongst the snail Ifremeria nautilei (pers. obs. Sabine Gollner) with Hd ranging from 0.61 to 1 [I. nautilei), larval nutrition (lecitotrophic Dirivultid larvae versus planctotrophic R. exoculata larvae), or number of offspring per individual  [The copepod species studied here live in close association with highly abundant megafauna that show similarly high haplotype diversity. For example, the copepod ed there , 67. Hap.94 to 1 . The copompejana . Haplotyompejana . S. lauegafauna) .ST values are generally low and non-significant indicating weak or no divergence between populations at different vent sites. In addition, haplotype networks showed no correlation between topology and distribution of haplotypes, and haplotype sharing among localities, pointing to similar phylogeography of species and unrestricted gene-flux during colonization processes. A priori, we had expected that genetic connectivity within dirivultids having similar life history traits is higher at fast-spreading centers with many vent fields (one every 10–20 km) than at slow-spreading centers with few vent fields (one every 100 km) [We observed similar genetic connectivity within the same genus regardless of distinct field frequency on EPR, ELSC, and MAR. This conclusion was based on divergence estimates derived from our AMOVA analyses, i.e., F 100 km) . High ve 100 km) . We specOcean current regimes also play a crucial role in vent fauna connectivity and dispersal, since larvae typically drift passively. Dirivultids have lectitotrophic nauplii , 37 and Connectivity between active vent sites might be also accomplished through the use of intermediate habitats as stepping stones. Vent copepods have been observed, although not frequently, up to 1 kilometer away from the nearest vent . NematodRimicaris exoculata, even along the slow spreading MAR, was likely related to several phenotypic and life history traits: its very large lecitotrophic eggs, planktotrophic larvae that feed in photic zones, delayed metamorphosis, and active, directed migration [Alvinella pompejana from fast-spreading EPR likely experiences high gene flow rates due to its lecitotrophic larvae (which arrest development in cold bottom waters), prolonged larval duration and increasing dispersal potential [Biological traits may also influence species connectivity. High gene flow in the shrimp igration , 15, 20.otential , 68, 72.The negative Tajima's D values observed in our dataset are consistent with population size expansion, or could reflect the effects of background selection acting on a non-recombining genome . PopulatOur comparative study suggests that populations of dirivultid copepods show a pattern of generally high haplotype diversity, high connectivity and population expansion, regardless of vent region. On the EPR, high species abundance values in the benthos and in the pelagial support our genetic data. Benthic copepod abundance data from CIR and ELSC are currently being analyzed by TK and SG, with first results (not yet published) implying that these also support our genetic data. Dirivultids maintain numerous colonies in close proximity, with locally abundant populations, allowing them to quickly colonize nascent vents. This could make these copepods rather robust against threats of mineral mining, such as destruction of active vent sites. However, it should be acknowledged that Dirivultids have different, important characteristics compared to other vent copepods and meiofauna taxa . For exaOur mtDNA results also raised questions that need to be addressed in the future in order to better understand the underlying processes of connectivity and population expansion. Genomic approaches analyzing 100 fragments, as well as current regime studies and studies on life history traits such as larval duration can help to further unravel mechanisms leading to high genetic connectivity in Dirivultids in areas of different vent field frequencies. These more detailed results will be necessary to the design of deep-sea preserves in areas that are targeted for future mineral mining operations.S1 FigBoth models are characterized by a population size of 10000 diploid individuals, a time of divergence of 100000 and 1000 generations respectively, and a migration rate of 0.0001. The model on the left allows migration to occur at rate 0.0001 in both directions whereas the model on the right only allows migration to occur in a single direction.(TIF)Click here for additional data file.S1 Tablea priori classification based on their morphological characters. Sequences that did not fulfill these four criteria were completely excluded. Species name, individual identification number, locality, source, phylogenetic assignment, and usage in population genetics are given. Genera: A.–Aphotopontius; S.–Stygiopontius.All these samples passed i) quality check of Sanger Sequence-Electropherograms, ii) Blast search if reads for artifacts (e.g. human COI), iii) can be unambiguously translated into a COI-amino acid sequence, and iv) clustered in reciprocally monophyletic clades according to their (PDF)Click here for additional data file.S2 Table-8 events/per bp/ per generation (m), number of individuals per population (Ne), number of migrants per generation (migr./gen.), length of loci in base pairs (bp), mjgration type, number of pseudo observed data (pseu.obs.), and number of replicates per pseudo observed sample (repl.pseu.obs.).Identification number of simulation (id), divergence time in generations (tau), mutation and recombination rates of 1x10(PDF)Click here for additional data file.S3 Table(PDF)Click here for additional data file.S4 TableSite, year of collection, number of samples (n), and species are given. Abundance data from sites Sketchy, P-Vent and Tica were extracted from raw data from Gollner et al. 2015. Data from sites Bio9 and Eastwall are here published for the first time. Information on sampling strategies and methods in Gollner et al. 2013 and Gollner et al. 2015.(PDF)Click here for additional data file.


Deuterocohnia longipetala (Bromeliaceae) to investigate species and subspecies boundaries within the genus and the genetic diversity of natural populations.Microsatellite markers were developed in D. longipetala. Of 64 loci selected for primer design, 15 were polymorphic among 23 individuals of D. longipetala and 76 individuals of the heterologous subspecies D. meziana subsp. meziana and D. meziana subsp. carmineo‐viridiflora. Twelve and 13 of these loci were also polymorphic in one population each of D. brevispicata and D. seramisiana, respectively. Numbers of alleles per locus varied from two to 14 in D. longipetala, two to 12 in D. meziana, one to nine in D. brevispicata, and one to 10 in D. seramisiana. STRUCTURE analyses clearly separated the taxa from each other.We used 454 pyrosequencing to isolate 835 microsatellite loci in Deuterocohnia species.The 15 new microsatellite markers are promising tools for studying population genetics in  Deuterocohnia Mez (Bromeliaceae) includes 17 species that are mainly distributed in the Andes of central South America  Mez that was collected along the road from Bermejo to Limal (Bolivia) and that is now cultivated in the greenhouse of the University of Kassel . From those sequences, we identified 835 perfect repeats with a minimum of seven units for di‐, six for tri‐, five for tetra‐, and four for penta‐ and hexanucleotide repeats, respectively. Sixty‐four microsatellite loci with sufficient flanking sequence and high repeat numbers were selected for PCR primer construction  and D. brevispicata Rauh & L. Hrom. , as well as in one individual each of D. seramisiana R. Vásquez, Ibisch & E. Gross (NiSch_06‐045K) and D. longipetala (NiSch_06‐068 as a positive control). PCRs were conducted in 12.5‐μL volumes in a T‐Gradient thermocycler  following a touchdown protocol . Fragment sizes were scored with the help of an external size standard as described by Wöhrmann et al.  and expected (He) heterozygosity values were determined with ARLEQUIN version 3.11  and deviations from Hardy–Weinberg equilibrium (HWE) were calculated with GENEPOP  and 0.66 (He). A total of 68 alleles were detected in 76 individuals of D. meziana, represented by D. meziana subsp. carmineo‐viridiflora  and D. meziana subsp. meziana , and the overall number of alleles ranged from two to 12. In the D. brevispicata population (n = 13), mean heterozygosity values of 0.39 (Ho) and 0.50 (He) over all loci were obtained. Finally, one to 10 alleles per locus were found in the 17 samples of the D. seramisiana population. Mean heterozygosity values in this species were 0.47 (Ho) and 0.51 (He). Mean FIS values ranged from a minimum of 0.11 for D. meziana subsp. carmineo‐viridiflora to a maximum of 0.32 for D. longipetala , the analysis was run for 1,000,000 generations in the burn‐in period and for 100,000 generations in the Markov chain Monte Carlo simulation analyses after burn‐in. Ten repetitions for each K (1 ≤ K ≤ 10) were performed, and the admixture level for each individual (Q) was also inferred. By calculating the ΔK statistic using STRUCTURE HARVESTER version 0.6.94 , there is a clear division among one cluster composed of all D. meziana subsp. meziana samples (Fig. K = 4, there is a second cluster containing all D. meziana subsp. carmineo‐viridiflora plants, a third cluster combining all samples from D. brevispicata and D. seramisiana, and a fourth containing all samples from D. longipetala (Fig. K = 5, D. brevispicata and D. seramisiana also become clearly separated from each other (Fig. To evaluate the potential of microsatellite markers for distinguishing between closely related taxa, a Bayesian cluster analysis was performed on a set of 129 plants comprising all samples from the two subspecies of D. longipetala revealed moderate levels of genetic diversity in the source species as well as in three heterologous Deuterocohnia taxa investigated. Whereas the two subspecies of D. meziana were surprisingly well separated from each other, the distinction between D. brevispicata and D. seramisiana was less pronounced, suggesting some ongoing gene flow among populations of these two species. The microsatellite markers developed here are promising tools for the study of population genetics, phylogeography, and the cohesion and delimitation of species and subspecies in Deuterocohnia. Genetic data generated by these markers will also provide important guidelines for designing management and conservation strategies in endangered species like D. meziana.The 15 microsatellite markers developed from 454 sequences of


The leakage of large plastic litter (macroplastics) into the ocean is a major environmental problem. A significant fraction of this leakage originates from coastal cities, particularly during extreme rainfall events. As coastal cities continue to grow, finding ways to reduce this macroplastic leakage is extremely pertinent. Here, we explore why and how coastal cities can reduce macroplastic leakages during extreme rainfall events. Using nine global cities as a basis, we establish that while cities actively create policies that reduce plastic leakages, more needs to be done. Nonetheless, these policies are economically, socially and environmentally cobeneficial to the city environment. While the lack of political engagement and economic concerns limit these policies, lacking social motivation and engagement is the largest limitation towards implementing policy. We recommend cities to incentivize citizen and municipal engagement with responsible usage of plastics, cleaning the environment and preparing for future extreme rainfall events. An exact estimate for the amount of plastic in the ocean is unknown, but the amount floating at the surface of the ocean is at least 93 thousand tons . HoweverThis ocean plastic is harmful to marine life, through deformation, maiming, suffocation and death e.g. . FurtherMarine plastic not only harms marine life, but also impacts our ability to interact with the coast. Potential decreases in marine biota would directly affect the fisheries economy. The tourism industry is also influenced by the presence of coastal plastic on beaches. The shoreline does not act as an unlimited plastic sink. Plastic may degrade on the beaches due to chemical and physical erosion and be washed back into the oceans . PlasticHuman behavior and policies on land affect the amount of plastic in the oceans. Littering and poor waste management, stormwater discharge, and extreme events such as floods and landslides are large terrestrial sources of marine debris . From inIn cities, street litter is not only unsightly but also expensive to cleanup. England spends £1 billion annually to clean up the 30 million tons of litter generated  and the Globally, better waste management is a focal point in reducing plastic in the environment. However, there are instances when day-to-day waste management will not suffice in stopping plastic leaking into the ocean. For example, in the immediate aftermath of a tropical storm, resource management is focused on human health, toxic spills and air quality  as opposWhy and how could coastal cities reduce urban plastic leakages due to extreme rainfall events?”As urban environments grow, stormwater management will become more important. In addition, it is projected that extreme rainfall events will increase due to climate change . An incr•To evaluate the current policy towards urban plastic management across the globe•To establish the co-benefits of reducing urban plastic in extreme rainfall events•To evaluate the limitations towards implementing these policies•To make recommendations for cities experiencing a projected increase in extreme rainfall eventsThis study aims to establish how coastal cities reduce plastic leakages and why these initiatives are beneficial to the city system using the following nine case cities as a basis for discussion: Vancouver, New York City, Miami Beach, Sydney, Singapore, Hong Kong, Rio de Janeiro, Copenhagen and Mumbai. It attempts the following four objectives:In 22.1Since the late 20th century there have been international efforts to curb the amount of marine debris in the world's oceans. These agreements underline crucial global issues and efforts and often lead to the creation and enforcement of policy at the national and local levels. In the 2015 G7 summits, the health of the ocean was one of the three key issues of global importance . Table 1If signatories of MARPOL Annex V suspect foreign ships of breaking the agreement in their territorial waters, there arelimited legal options for them to pursue, which are costly and lengthy. It is difficult to witness intentional dumping across all territorial waters and as debris moves with the currents, the presence of debris near a vessel is not condemning evidence . The enf2.2There are efforts to reduce marine plastic on regional levels. However, The EU has the only legally binding approach to reducing marine debris at the regional level . As a re2.3There are further management levels for controlling urban plastic spillage into the oceans. Practices can extend from national policies, regional initiatives, municipal ordinances to community action. National governments are responsible for establishing national frameworks for managing the environment as well as setting national quotas and targets. Regional, state or provincial governments set local targets under the national framework. The municipal level acts as a vector for the regional targets and is able to set further local targets. However, there is often tension between these varying levels of governance in managing the environment . The sma3Human settlements concentrate along the world's coast lines. Roughly half of the global population live within 60 km of the coast and nearly three quarters of all large cities are located here . Coastal3.1Urban settlements can control the release of plastic during extreme rainfall through systems management. Stormwater systems are put in place to channel rainfall through the city to avoid flooding. However, rainfall in cities often flows directly into adjacent water bodies, either as direct runoff or due to channeling. Stormwater can carry plastic directly into the oceans in coastal cities. There is more than one type of stormwater management. However, the most commonplace are traditional combined sewer overflow (CSO) systems, where stormwater is joined with sewage. CSO can lead to the direct flow of this mixed water into the oceans, bringing pollutants with it . AlternaWaste management systems manage the amount of plastic in the urban environment. Landfilling, especially open landfilling near the coast, can allow for plastic leakages into the ocean. However, recycling schemes can control the amount of street plastic. There are both municipal and individual recycling schemes. For example, in Sweden there are municipal recycling schemes in place to collect mixed recyclables including plastic for reprocessing . Bottle 3.2Outside of systems management, there are policy management techniques to minimize the amount of plastic leakages to the oceans during extreme rainfall events. Economic incentives deter people from intentionally littering. Cities such as Miami Beach and countries such as the UK have littering fines in place to deter street litter . Fines cTaxes are instrumental in changing consumer behavior towards plastic. A 15 euro cent tax on plastic bags in Ireland led to a 90% reduction of plastic bag usage in Ireland in the early 2000s . The taxEducation is a key policy area for reducing the amount of plastic easily available to oceans in coastal cities. A plastic consumer will make smarter decisions if they're aware of the consequences of their actions . Hartley4The research presented here can be separated into four components: data collection and city analysis, the co-benefit of policy, limitations to policy, and recommendations for policy makers. Here, we briefly present the methodology followed for each of these components. The full methodology is summarized in 4.1To establish a global snapshot of urban coastal policies towards street plastics, this study examined a selection of nine globally spread cities: Vancouver, New York City, Miami Beach, Rio de Janeiro, Copenhagen, Mumbai, Singapore, Hong Kong and Sydney . These cData was extracted from the nine global cities and tabulated into the following sections: stormwater management, rainfall management, municipal managed recycling, individual recycling, littering laws, smoking laws, taxes on materials, bans on materials, beach management and marine management. The policies and laws were extracted from the respective city's governmental websites as well as state/regional and federal government websites. Policies and plans that were proposed but not enacted were not included. Stormwater and rainfall management plans were examined for future plans against rainfall inundation and plastic leakages. Recycling, littering and smoking laws as well as taxes and bans were examined for their policies towards reducing the amount of plastic on the street. Beach and marine management were examined for active policies towards cleaning up the environment.We recognize that the majority of these policies are not created explicitly to reduce plastic leakages to the ocean. Nonetheless, they are influential in reducing plastic leakages and help tackle marine plastics. The ten categories provide an overall snapshot of urban policies towards the street plastic problem and the policies were discussed in the context of the nine global cities.4.2Following the initial discussion of the nine cities and their policies a secondary analysis was performed. The policies are discussed under political, economic, social and environmental co-benefits. Three case examples were used to highlight specific co-benefits; beach tourism for economic benefits, smoking and health for social benefits and CSO for environmental benefits. There was no case example used for political co-benefits due to the lack of politically specific co-benefits of reducing urban street plastic leakages into the ocean. As policy is frequently aimed at tackling multiple problems, the co-benefits discussed are those in which by reducing urban street plastics and leakages there is a direct consequence on the urban environment.4.3In the third stage of the analysis, this paper discusses the limitations of policies and policy making in the area of managing urban coastal plastic leakages. Five people were interviewed across four cities: Vancouver, Sydney, New York City and Miami Beach. The interviewees were selected due to their positions in city government, academic work or involvement in non-profit/NGO advocacy. The interviewees' names were altered for the discussion . A semi-The interviews identify key limitations towards policy making across the global cities. The discussion is structured similarly into four areas , basing its argument on the opinions presented by the interviewees. This study recognizes that the limitations to policymaking vary at different sized jurisdictions. Therefore, this discussion focuses on the limitations to policymaking at the urban municipal scale. It does not focus on limitations faced by state/regional or federal governments.4.4This study presents a list of recommendations for policy makers in formulating future policy towards reducing urban plastic leakages to the marine environment. The recommendations are based on the existing policies, co-benefits and successes of these policies, and the limitations to formulating policy. The list intends to be used a guidance for cities to make future policy alongside additional research for a city's unique location and structuring.5The following section presents the findings of the city analysis across the nine global cities referring to 5.1For coastal cities, managing their water is integral to the functioning of the city. All of the nine cities have a plan in place for stormwater management and rainfall management . Rio de Although the plans for stormwater and rainfall management are important for the future of the cities, the legal framework and structuring of urban governance weakens the policies. New York City's Sustainable Stormwater Management Plan is no longer being actively implemented but instead is being reincorporated under OneNYC due to the legislative changes made at the state and local level . These pThe water management plans are often well placed within larger policy contexts. For example, Singapore's Active, Beautiful, Clean Water program incorporates cleaning waterways not only for reduced pollution but also for community access to greenspace and city beautification. Furthermore, Vancouver, New York City, Rio de Janeiro, Copenhagen and Sydney explicitly link their water management with climate change or general sustainable management. This places good stormwater and rainfall management in the discourse of a sustainable, healthy city for the future, encouraging further developments.A few of the policies target niche components of rainfall and stormwater management; extreme downpours and flooding. Copenhagen's Cloudburst Management Plan  “outlineincluding extreme rainfall. With this plan, “[the city has] taken decisive steps forward to protect Copenhagen against high-intensity rain like the ones witnessed in August 2010 and again in July and August 2011” (p.5). The plan recognizes that despite future financial investments in expanding traditional sewer systems, the cost from rainfall inundation would be too high . InsteadBoth Mumbai and Miami Beach have specific plans for disaster management with rainfall and flooding, respectively . While b22). Sydney has also made large efforts to remove CSO systems from the city. So, while these systems are still present, their influence in handling stormwater has decreased with new water management techniques. Vancouver is working towards British Columbia's goal of eliminating CSO by 2050  going up to US$7426  on subsequent fines. Singapore can also issue Corrective Work Orders to litterers forcing them to clean public areas. In 2014, 688 of these orders were issued . FurtherEvery city has a ban on smoking . All theThe most practical measures towards reducing street plastics, bans and taxes, are the most inconsistent across the cities. These ordinances would target the top of the waste pyramid, prevention. Only two cities have a tax on plastics, specifically plastic bags; Copenhagen and Hong Kong. The remaining seven cities have no explicit consumer tax on using plastics. Unlike deposit schemes, the tax on plastic bags is non-refundable and is a direct added cost to the consumer.5.3Beach management and marine management are active ways in which the city can spread education and clean up the environment. Of the nine cities, only Mumbai was not explicit in the government's role in these management areas. Three cities; Rio de Janeiro, Hong Kong and Sydney were explicitly involved in both marine and beach management.Beach cleanup campaigns appeared differently across the cities. Rio de Janeiro, again influenced by the 2016 Olympics, has localized cleanup projects sponsored by the government in the beaches and coastal margins of the city. Similarly, Singapore has localized cleanup campaigns, including those aimed at children, to maintain beaches as well as standardized government cleanup of beaches. However, Singapore does not denote a single agency to be in charge of managing the amount of plastic that ends up in the marine environment and the beaches. In Sydney, the NSW government actively cleans the harbor on behalf of the city. Hong Kong and Miami Beach have the most comprehensive beach cleanup campaigns. Hong Kong has a Clean Shorelines initiative where a working group sponsored by the government attempts to study the marine plastics, identify their sources and identify potential cleanup methods while helping coordinate community action on cleaning up the beaches. In contrast, Miami Beach has partnered with the Environmental Coalition of Miami and the Beaches, a county wide organization that attempts to protect all the beaches in the Miami area. Through this community sponsored adopt-a-beach programmes as well as cleanup events and education campaigns are utilized in Miami Beach as well as standardized for the surrounding area. Of the existing beach management policies, the cities differ on the extent and scope of their involvement.While the city government may not be active in beach management campaigns, NGOs, non-profits and community action often fills the gap left by the city government. In addition, other governments may be charged in handling beach cleanup campaigns. In Miami Beach, the state of Florida has statewide cleanups to increase awareness of the marine pollution problems . In Vanc3 of rubbish is collected from the water and beaches (73). In New York City, just over 1000 m3 of debris was collected in 2012 (65). Furthermore, there may be individual cleanup projects for specific waterways within a city. In New York City, there is a specific plan in place to prevent debris in the Newtown Creek , there can be barriers when creating policy; both from within and from outside the governmental system. These barriers hinder both the formation of new policy as well as the implementation of existing policy. It is important to understand these limitations in order to formulate more effective policy in the future. This discussion of limitations to policy is organized under the following headings: Political, Economic, Social and Environmental. The main limitations are summarized in The four cities represented; Vancouver, Sydney, Miami Beach and New York City all have different demographics, environment and political management; as represented during the interviews. However, the cities were mostly in agreement when it came to the limitations faced by policy in reducing urban plastic leakages to the environment . All fiv7.1Within the political system there are large barriers towards forming policy to prevent plastic leakages. Often, there is little support from the regional or national government. All five interviewees state that the existing policy is mostly driven by the cities themselves. Cities have to interact with regional policies; which has both benefits and limitations. Citizens can be confused between the role of the city and the state in areas such as waste management . While aCity governments often face pressure from outside the government to formulate policy. The plastics industry has a direct stake in this policy and therefore is a powerful lobbying group when the legislation is drafted . While tA large political limitation to policy is that, once the comprehensive plans are passed, the goals set forward are not legally binding. This limits the accountability for cities if they fail to reduce plastic leakages. “It's great to set goals and milestones you want to hit but again its surface level. [Policy makers] don't explain how they're going to do things, what this is going to look like. Let's reduce X by Y amount but [policy makers] don't explain further. And really I think a lot of people don't ask those questions because once you go to the average person they're usually pretty excited that progress is being made on that front but they don't really have the knowledge or the background or the experience of working with the elected officials and city council to understand that… [Policy makers] also know that by the time anything is revealed, they're going to be gone. It transfers the responsibility.”Without legal backing to the policy, the city is under no pressure to adhere to their targets. Once the targets come into place in 2025, 2030 or 2050 for example, the current policy makers are likely to be out of office thus passing the responsibility of the targets to future generations. This is compounded by the citizens' surface level understanding of how the policy targets will be met. The policy targets for plastic leakage reduction are often more statements of public relations than clear cut goals.The city is not always limited politically to create legislation for plastics. Often targets would be impractical for a city to implement. It is difficult to permit and license for stormwater as it's difficult to control its quantity and quality . However7.2Policy that protects urban coastal waters from plastic leakages is not only limited by the political structure but also the cost. Not every plastic material is easily recyclable which inhibits the motivation to develop comprehensive recycling schemes. Despite their presence, “it costs a lot of money to collect lightweight materials that are worth very little” . MateriaThe cost of upgrading existing infrastructure often limits a city's ability to further push through legislation on the control of plastic on the streets. End of pipe solutions for stormwater and waste management facilities are a costly investment that takes years of planning and millions in funding to operate . In addiUncertainty in the future creates an economic doubt on the influence of policy. With a fast-changing climate and fluid demographics it's difficult to plan beyond the 20–50 year targets current policy outlines. Therefore, plans and programmes that continue into the years beyond are questioned because it may be unnecessary to invest for a future that won't appear in reality. While this reasoning can be logical for economic decisions, the climate models predict future extreme rainfall events are likely to be more severe, and therefore the damages brought with them are also likely to get more severe. If a system isn't put into place for the potentially destructive future, economic losses will be higher than they are today. Cities work economically in a more practical than precautionary way. In this they'll gratefully partner with NGO and non-profit organizations for education campaigns and cleanup campaigns but will often neglect funding regular cleanup activities themselves . It is e7.3Policy will ultimately not be effective if the public do not take responsibility for their share of solving the plastic leakage problem. All five interviewees agreed that cities try to pass the torch of responsibility onto citizens because “we can't be everywhere; we can't do everything” . By engaCities hope to rely on social policing to alter consumer behavior with plastic. People are more likely to clean their litter, recycle their water bottles, reuse a plastic bag, etc. if they believe it to be the social norm and that their peers are observing and potentially judging their behavior . HoweverBeyond informing the city about legislation, the city struggles to turn that knowledge into active engagement. Cities have higher response rates in areas where they have direct control, such as for example single family homes . EnforciIn daily life, citizens may be ignorant to the problem of coastal marine plastic. Many people in coastal cities don't venture out to the beach and witness the problem first hand . If one Finally, a large social limit to policy is the citizens' commitment to the environment in their day-to-day lives. Plastic bag charges are good at reducing plastic bag usage, but over time citizens may incorporate the tax into their expected spending and that could cause a rebound of plastic bag use . HomelesWhile the environment may not directly limit policy making, the future environment does influence our decision making. The rainfall projections are not completely certain, they are not fact. While models can project the likelihood of whether rainfall increases or decreases, these models will always have an uncertainty range when quantifying the increase in rainfall or frequency of storms. Therefore, cities may be over or under preparing themselves for future rainfall. Coastal cities are not only susceptible to rainfall, but also coastal flooding. Coastal flooding, sea level rise and storm surges can flush cities of street plastic and create further debris if they are destructive. Furthermore, rainfall changes are only one consequence of future climate change. Cities must also prepare for changes in drought, hotter or colder temperatures and other changes to the physical system.Within the city, environmentalism runs throughout city issues. It is often the poorest citizens that live in the least environmentally secure areas . Yet all8Policy that targets urban plastic leakages to the ocean is highly beneficial for a city and its environment; yet the political, economic, social and environmental barriers often hinder the extent this policy to tackle litter is implemented. This paper attempts to put forward suggestions for coastal cities in how to create future policy that overcomes the limitations in the context of existing legislation. Cities should continue to advocate for both stronger international and national laws and agreements on marine plastics , however8.1•To overcome weak regional and national level support for laws targeting the reduction of plastic usage, cities can create a regional committee representing the city and neighboring communities to create policy. In this the entire metropolitan region is formulating a single policy and citizens living in the entire region follow the same criteria whether it be bans, fines, taxes or deposits.•To overcome weak targets set in policy, cities should place their forward-looking plans under an overarching plan for the environment and sustainability. Thereby, each individual strategy for example for waste management or water management is placed in the context of the general city's environmental goals. It also allows for the strategies to be specific and targeted to areas of concern while working towards a common goal for the city. This also encourages different city departments to work closer together to achieve these goals closing any gaps in management.•To overcome other weak targets set in policy, cities should create targets alongside the community. City governments can work towards 50% of the target while the remaining 50% is left up to communities. For example, in increasing recycling rates, the city can increase recycling infrastructure and alter plastic collection methods while districts within the city are responsible to reduce plastic waste being disposed of improperly. In this the city is more likely to meet their half of the target as they are not required to do everything towards the goal. Furthermore, the city districts can develop strategies tailored to their specific community to reduce mismanaged waste.•To overcome citizens' lack of engagement with the plastics problem, cities should create and further education and awareness campaigns. These campaigns can target public areas where citizens are most likely to interact with the campaign such as public transit stations as well as schools which can help shape future citizens' behavior. The increased presence of these campaigns can slowly permeate daily life and decisions.•To overcome citizens' lack of engagement with the plastic problem, cities should allow for non-profit advocacy to engage in the policy formation stages. The non-profit advocacy groups have specific knowledge in more niche fields and their practical knowledge is valuable. Furthermore, these groups have experience with public engagement in taking action against marine plastics and have valuable experience in the application of policy. As opposed to lobbying the NGOs should be present during the drafting of legislation as contributors in contrast to persuading legislators.8.2•To better control extreme rainfall events, cities should develop a rainfall management plan that examines how rainfall collects over the city, its current effects on the environment and the effects of future rainfall projections. The city will be better prepared to respond to extreme rainfall events, have a better understanding of how rainfall interacts in the city and understand how plastic is moved through the inundated environment.•To better control extreme rainfall events and capture street litter, cities should focus on developing green infrastructure. The infrastructure is often cheaper than traditional methods, can occupy both large and small spaces, and has numerous benefits towards the health and environment of the city. Green infrastructure may also be cheaper to maintain and is easier to change in the future. This infrastructure will also help manage water and plastic during normal dry and wet events. Funds should also be set aside to maintain and clean the infrastructure.•To better develop green infrastructure, cities should establish a tax credit system for property owners who invest in developing green infrastructure. As the base cost of introducing green infrastructure projects may not be as large to cities, depevolment of green infrastructure can be unattainable for small businesses and home owners without a financial incentive. The tax credit can be dictated by density zones, with the densest areas receiving the biggest credit.•To better control plastic leakages to the marine environment, cities should focus efforts on reducing Combined Sewer Overflows (CSO). While reducing the flow of water to CSO can be done with green infrastructure, closing the CSO will ultimately prevent sewage from entering the waterways. The infrastructure improvements should also reroute stormwater to water treatment plants as opposed to direct spillage into the environment.8.3•To better control the use of plastic in the city's streets, cities should create strict laws and steep fines on littering. These laws and fines should be monitored closely immediately after implementation and strictly enforced to encourage citizens to change habits. Once the new habits take hold, enforcement can be scaled back with increased social policing.•To better control the use of plastic in the city's streets, cities can incorporate better street recycling vessels. These include recycling containers within proximity of waste bins, separate containers for different materials including plastic, separate containers for items with deposits, or bins that can accept deposited materials and return the value of the deposit to the user. The convenience of disposal of plastics will encourage citizens to dispose of plastic correctly on the streets.•To better control the overall use of plastic, cities should identify necessary and unnecessary plastic products for daily life. Afterwards, the city should apply a tax or deposit on their ‘necessary’ plastic products such as plastic bags and bans on their ‘unnecessary’ products such as Styrofoam cups. The lists should be tailored to each city as for example plastic bags may be viewed as necessary in one but unnecessary in another depending on the residents. Environmentally friendly cities are becoming more enticing for investments. Cities push each other towards higher environmental standards to maintain a competitive edge.•To better control cigarette butts in cities, cities should create more community areas where smoking is not permitted. Public parks, beaches, and commercial areas should be areas of priority for cities seeking to reduce both cigarette butts and second hand smoke hazards.•To better control cigarette butts in cities, cities can implement a deposit or further tax on the cigarettes. The tax can offset costs of collecting cigarette butts in the street while a deposit encourages smokers to not dispose of their waste on the street.8.4•To better cleanup the coastline and local waterways, cities should create a new department or sub-department/committee that is dedicated towards marine and coastal cleanup. By doing this, cities would be able to create a direct strategy for cleanup and allocate specific resources to this. It will also be able to direct citizens concern and involvement towards a specific department.•To better cleanup the coastline and local waterways, cities should partner with non-profits for better environmental management. In the actual cleanup, cities should subcontract and fund the responsibility to these non-profits who can dedicate their time to the specific problem of marine plastics.•To better cleanup the coastline and local waterways, cities should establish tax credits for companies and start-ups pursuing technologies that clean up the debris. This will encourage citizen engagement with clean-up as well as reduce the burden to the city governments to search for solutions.•To better cleanup the coastline and local waterways, cities should establish programmes that allow for citizens or businesses to take ownership of the beaches such as adopt-a-beach programmes. This will further encourage citizen engagement with the environment and reduce the cleanup costs to cities.Most cities have policy in place that control plastic leakages. There is no standard in management, however, and often the policies are conceptual in nature and lack legal follow up in the city and regional governments. Reducing urban plastic leakages can have numerous benefits to the city beyond solving the plastic problem. The city can experience a stronger economy, a cleaner environment and a healthier society while removing plastic leakages. While cities struggle to politically formulate policy or find the appropriate funding for new measures, a lack of social engagement will ultimately limit a policy's effectiveness.By implementing a selection of the recommendations, this paper believes that cities can effectively overcome the limitations towards policy targeting plastic and enjoy the direct and co-benefits. However, each city is unique and this paper should be taken as a guiding document. Certain coastal cities continue to face a threat of extreme rainfall events and further research should be conducted on an individual city level to create comprehensive plans to reduce plastic leakages to the ocean during these events. Cities should continue to focus on reducing the amount of plastic waste produced. This way additional plastic is removed from the waterways and the management of the water systems and cleanup can be more effective in ensuring a greener environment.