Evolving multicellularity is easy, especially in phototrophs and osmotrophs whose multicells feed like unicells. Evolving animals was much harder and unique; probably only one pathway via benthic ‘zoophytes’ with pelagic ciliated larvae allowed trophic continuity from phagocytic protozoa to gut-endowed animals. Choanoflagellate protozoa produced sponges. Converting sponge flask cells mediating larval settling to synaptically controlled nematocysts arguably made Cnidaria. I replace Haeckel's gastraea theory by a sponge/coelenterate/bilaterian pathway: Placozoa, hydrozoan diploblasty and ctenophores were secondary; stem anthozoan developmental mutations arguably independently generated coelomate bilateria and ctenophores. I emphasize animal origin's conceptual aspects  related to feeding modes, cell structure, phylogeny of related protozoa, sequence evidence, ecology and palaeontology. Epithelia and connective tissue could evolve only by compensating for dramatically lower feeding efficiency that differentiation into non-choanocytes entails. Consequentially, larger bodies enabled filtering more water for bacterial food and harbouring photosynthetic bacteria, together adding more food than cell differentiation sacrificed. A hypothetical presponge of sessile triploblastic sheets (connective tissue sandwiched between two choanocyte epithelia) evolved oogamy through selection for larger dispersive ciliated larvae to accelerate benthic trophic competence and overgrowing protozoan competitors. Extinct Vendozoa might be elaborations of this organismal grade with choanocyte-bearing epithelia, before poriferan water channels and cnidarian gut/nematocysts/synapses evolved.This article is part of the themed issue ‘Evo-devo in the genomics era, and the origins of morphological diversity’. Bacteria and protists greatly exceed vertebrates in different kinds of organism too. Lamarck thought unicells so evolutionarily recent that they had not yet had time to inexorably become multicellular. Not so; they existed billions of years longer than complex multicells and may outlive them. There are hordes of excellent unicellular niches; multicellularity is often selectively disadvantageous. Yeasts evolved multiply from multicellular filamentous ancestors; Myxozoa are parasitic unicells that evolved from animals with nervous systems (early-branching Cnidaria), losing epithelia, connective tissue, nerves and 70% of genes as useless, only their multicellular spores keeping nematocysts . So how Multicellularity evolves in two ways. Naked cells, as in animals and slime moulds, evolve glue to stick together. Walled cells modify wall biogenesis to inhibit the final split that normally makes separate unicells, so daughters remain joined. The ease of blocking that split allowed almost every group of bacteria, fungi and plants (and many chromists) to evolve multicellular walled filaments, more rarely two-dimensional sheets, most rarely three-dimensional tissues. Tissues require more geometric control of daughter wall orientation, as in embryophyte green plants and chromist brown algae; both can grow longer than blue whales. Evolving tissues is selectively harmful to many walled multicells whose filaments are best for reproductive success. Almost all multicells retain unicellular phases , so adhesion is temporally controlled and developmentally reversible—except for purely clonal vegetatively propagating plants or ‘colonial’ invertebrates (evolutionarily transient) the only organisms that are never unicellular.2O, CO2 and minerals) just as does a single cell; so does a saprotrophic bacterial or fungal filament. However, a phagotrophic amoeba could not aggregate into a multicellular body and still locomote and feed the same way. Nor could most other protozoa. Many amoebae have become multicellular, but only temporarily for spore dispersal, not feeding. Aggregative multicellularity has produced multispore fruiting bodies numerous times in fundamentally different protist lineages  to switch from intracellular phagocytosis, as in amoebae or ciliates, to eating with a multicellular mouth and gut, whose cells have novel functions and structures absent in their unicellular ancestors. Animal feeding is effective only if novel cell types cooperate at a higher organizational level; most give up the ability to feed or reproduce, huge selective disadvantages not easily overcome.In 1866, James-Clark discovered choanoflagellate protozoa and their feeding on bacteria trapped by a collar surrounding their undulating cilium that generates the water current that draws them towards it. He noted that sponge collar cells (choanocytes) have the same structure and feeding method, correctly suggesting that sponges evolved from a choanoflagellate . Often sAmong extant animals, only sponges could have evolved directly from protozoa without changing feeding mode. The key problems in understanding animal origins are therefore how and why sponges evolved from a craspedid-like stem choanoflagellate and later generated all other animals. I attempt to explain both after briefly outlining enabling protozoan innovations. I shall emphasize simple conceptual aspects of the choanoflagellate/animal transition, often overlooked but more important than discovering extra protozoan genes suitable as precursors to animal functions. Such ancestral features exist in both choanoflagellates and more distant protozoan relatives of substantially different cell structures and feeding mode .In the light of site-heterogeneous trees using 187 protein sequences ,17, figuSulcozoan flagellates clearly could not have retained their characteristic locomotory or feeding modes had they evolved glue to stick together as a multicellular organism; such mutants would necessarily quickly starve to death. Nor could their immediate ancestors—three successive groups of swimming, not gliding flagellates  collectively called excavates because their ventral groove looks more obviously scooped out ,17. The Knowing the structure and evolutionary potential of the closest relatives and ancestors of animals  and thatDictyostelium has a β-integrin-like adhesion protein [Integrins and associated molecules used for epithelial cell adhesion to extracellular matrix (ECM) were secondarily lost by choanoflagellates and fungi; without full genomes for the deepest branching Sulcozoa, the exact point of origin is unclear : though  protein  and its  protein  —or even why—but defining the selective forces that promoted the fundamental differentiation between sponge feeding cells (choanocytes) and non-feeding cells and between cells that stick together as epithelia and connective tissue cells embedded separately in a gelatinous mesohyl. Did epithelia evolve first or did epithelia and mesenchyme coevolve?Carchesium, Zoothamnion); some algal, e.g. chrysophyte Dinobryon. Mucilaginous multicellular branching structures are formed by Rhipidodendron (cercozoan chromists) or Phalansterium (uniciliate Amoebozoa). As no branching protists evolved a multicellular tissue, similar ‘colonial’ choanoflagellates are probably not directly relevant to animal origins. Nonetheless, they show that various linked flagellates can still feed in the same way as when unicellular, and their frequency suggests that branching stalks advantageously enable them to sweep prey from a much larger water volume than can one sessile cell. Filtering more water by a different sessile body form is, I argue, the selective advantage that made sponges.Four different ways of making multicellular choanoflagellates exist. Many become ‘colonial’ sessile organisms by evolving thin extracellular stalks that join cells together to form branched tree-like structures analogous to corals or plants ,10. OtheProterospongia choanojuncta, but I doubt this had a potential to yield a sponge. Sponge collars also join laterally often by a second mucus mesh to achieve 100% removal of suspended bacteria [More rarely, choanoflagellate multicells arise by linking adjacent cells by their collar microvilli as in bacteria , showingDiaphanoeca sphaerica, where cells often clump in hollow balls with cilia pointing inwards [Diaphanoeca, like other loricates (Acanthoecida), are tiny cells suspended within a much larger lorica of siliceous strips porous to water currents carrying prey. Aggregating porous loricas by connecting longitudinal strips allows colonial feeding despite cilia pointing inwards, as the collar outer surface that traps food still faces outwards. Water and bacteria can pass through the lorica mesh or wide interlorica spaces, so feeding mode is unchanged compared with unicells; cell bodies are not in contact so could not evolve into an epithelium to make a sponge. Acanthoecida are necessarily an evolutionary dead end.The loricate  inwards , exempli inwards  was mislSphaeroeca is a multicellular planktonic craspedid whose colonies are hollow balls with a surface cell monolayer, associated by cell bodies not collars, analogous to the alga Volvox that Hardy [Salpingoeca rosetta reversibly makes little multicellular balls, a capacity influenced by bacteria [Non-loricates (Craspedida) never aggregate with cilium facing inwards like sponges as that would suicidally stop collar-based feeding. at Hardy  invoked bacteria . Numerou4.Willmer emphasized the basic dichotomy between ciliated epithelial and non-ciliated, amoeboid, connective tissue cells as fundamental to animal development . Figure Dictyostelium dead stalk cells.The primary dichotomy between uniciliate choanocyte and non-ciliate pinacocyte is also mirrored by that between sperm and egg. Therefore, part of the same gene switches needed for somatic differentiation could also be used to differentiate gametes. Once a three-layered structure with just two somatic cell types evolved, presponges could become quite large (compared with choanoflagellate unicells); selection for rapid establishment of a large embryo would strongly favour oogamy  by modifying choanocytes, presumably hermaphrodite. The animal bauplan was in place once a selective force for ever-larger filtering structures built from two dissimilar cell types existed: two germ line and two soma cell types. Accidental fragments could also reproduce vegetatively as choanocytes retained pluripotency . There wAnother selective advantage of evolving mesenchyme and massive tissues perhaps gave extra impetus to early animal evolution. Mucilage easily harbours bacterial symbionts potentially able to provide enough extra food to repay a presponge several times over the trophic and reproductive costs of non-feeding cells. Cultivating cyanobacteria in ECM mucilage would make the photophagotrophic consortium an extremely effective competitor with merely branched choanocyte-only colonial choanoflagellates. Lichen fungi can survive solely by cultivating cyanobacteria; a presponge could be even better off, being also a phagotroph able to grow far faster than a lichen in bacteria-rich water. Great Barrier Reef sponges 1–2 m high are often red through being packed with cyanobacteria whose biomass is greater than that of the sponge cells. Lake Baikal giant freshwater sponge tissues cultivate green algae. Both habitats are oligotrophic, making internal algae especially advantageous, but even in habitats rich in particulate food, the majority of sponge species are often photosynthetic . In orgaExtra cell types could be added relatively simply to help presponges to grow bigger and be less susceptible to environmental damage. An individual could grow basally across a rock and erect multiple laminae. Spatial controls evolved to prevent laminae from interfering with each other. Presumably, various morphologies and arrangements and ratios of the two basic cell types were experimented with, giving different compromises between maximizing feeding and mechanical stability. An early innovation necessary for large structures was to increase the ECM-synthesizing cells initially perhaps by evolving a third cell type—the ancestral archaeocyte that left the epithelium, entering the mesohyl for secreting ECM in all directions, making a triploblastic tissue with mesenchyme sandwiched between two epithelia. Nowadays archaeocytes and choanocytes are the demosponge stem cells, expressing PIWI double-strand RNA-binding domain proteins whose short-RNA related functions are associated with germline and stem cell maintenance in higher animals  as well 5.Site-heterogeneous multigene trees  maximally support choanoflagellates being sisters to animals ,33; theySite-heterogeneous multigene trees equally strongly show sponges as a clade, disproving Nielsen's assumption that eumetazoa are more closely related to homoscleromorphs than others, and invalidating his twin assumptions that ancestral animals were lecithotrophic and eumetazoa secondarily lost lecithotrophy . His sug6.This presponge was not a sponge, for it lacked an aquiferous system (AS) with incurrent pores (ostia) and larger excurrent osculum or oscula. AS architecture has two advantages: (i) it increases food supply by pumping much larger water volumes past the choanoderm; (ii) compared with the essentially ‘free-living gills’ of the presponge, placing the choanoderm inside a globular or encrusting body protects choanocytes from damage by sand and other things swept against them by vigorous water currents and from damage by the currents themselves. Essential innovations making a sponge were (i) controlled formation of ostia of appropriate size, frequency and distribution; (ii) rearrangement of pinacocytes and choanocytes to internalize the latter, make a more compact less easily damaged body, and optimize water flow through internal choanoderm-lined channels. Ostia are intercellular in all Homoscleromorpha and most demosponges, but are formed by channels through specialized porocytes in Calcarea and not obviously homologous contractile porocytes in a few haplosclerid demosponges. I suspect they originated not by evolving a new cell type but by spatially controlling pinacocyte contacts and geometry; porocytes evolved later independently in Calcarea and haplosclerids. If so, ostia arose as part of the supracellular rearrangements that made an axially polarized water channel system. This major innovation almost certainly depended on prior evolution of morphogen gradients and homeobox and other spatially controlled switch genes that sponges share with Eumetazoa ,38. BeneChlamydomonas and could be a general property of eukaryote cilia that evolved during the origin of two structurally and behaviourally dissimilar cilia in the eukaryote cenancestor [Ephydatia, oscular sensory cilia lack the centre-pair microtubules as in eumetazoan sensory cilia [Making AS development and functioning more efficient probably entailed differentiating pinacocyte subtypes: specialization of some as myocytes to exert some control on oscular and ostial opening; and multiplication of non-epithelial mesohyl cell types. The branched mesohyl cells that synthesize a variety of neurotransmitters are obvious candidates for precursors of eumetazoan nerve cells, requiring only the origin of electrosensitve channels to cause action potentials and synapses to make a nerve net. The syncytial body form and calcium/potassium action potentials of hexactinellid glass sponges are secondary, not the ancestral condition for sponges, as hexactinellids are related to demosponges not the deepest lineage . They arancestor . In the ry cilia . Early sry cilia  with a hThe greater complexity of true sponges over presponges required planktonic ciliated larvae for dispersal to new fixed sites that grew big enough to transform immediately into a tiny triploblastic sponge with internal choanoderm able to feed at once. Abundant egg yolk enabled more rapid development than feeding by surface choanocytes, making sponge larvae lecithotrophic unlike planktotrophic presponge and ancestral eumetazoan ciliated larvae. Larvae evolved phototaxis using cryptochromes , not rhoThe phrase ‘from amoeba to man’ epitomizing Haeckel's early phylogenetic views doubly misleads. Amoebae are not primitive but arose from zooflagellate ancestors independently in each of the three ancestrally biciliate eukaryotic supergroups . Epithel7.Hydra [Trichoplax (unlike sponges) has numerous presynaptic protein precursors as well as gap junctions, chemical and electrical synapses probably both originated after the pre-cnidarian lineage diverged from placozoa yielding an anthozoan-like stem coelenterate. Thus, neither muscular [1The larger larvae of true sponges provided a novel, hitherto unexploited, food for predators. One stem sponge lineage, I suggest, evolved nematocysts to catch and digest them, thereby becoming the ancestor of coelenterates , a clade on the best multigene trees . NematocHydra ), and thHydra , and synHydra ; very feHydra . As Tricmuscular  nor cilimuscular  initiateKey to neurogenesis was a multicellular precursor with neurotransmitter-making cells and already adjacent receptor and effector cells linkable by evolving synapses under a strong selective advantage, exactly as this flask cell to nematocyte transition postulates without missing links or improbable events. Thus, improving the sessile zoophyte lifestyle by increasing survival (e.g. against waves tearing settling larvae from rocks) at the crucial, but uniquely vulnerable, pelagic–larval/benthic–adult transition was, I contend, the selective force for evolving synapses, ultimately leading to brains, culture and science. Synapses evolved to make ciliated larval settlement faster and more effective by neural coordination of concerted banks of nematocysts under the control of ciliated sensors that selected the best sites. Flask cell precursors concentrate at the aboral pole. Nematocysts remain there to mediate settlement but concentrated also around the osculum (making it a mouth) and along ancestral anthozoan protosepta to trap food.+ and K+ channels (both originating in bacteria) were modified to generate sodium/potassium action potentials in longer nerve cell branches for distant coordination of feeding responses, making eating more efficient—a selective advantage an automatic corollary of this explanation of synaptic origin. Action potentials evolved many times, thus easily—not only in hexactinellids, but also filamentous fungi, plants and ciliate protozoa [Adding synaptic junctions not only between sensory cells and nematocysts, but between sensory cells and branched pre-existing branched transmitter-making cells  and myocytes, would establish local neuromuscular control by a nerve net. This speeded oscular contraction making it an effective mouth, its reversible closure plus adhaerens junctions being key innovations for initial extracellular digestion of larger prey caught by oral and septal nematocysts. Having established neuromuscular synapses, pre-existing voltage-dependent Naprotozoa . Axons eper se does not make nerves.Before tentacles evolved, partially redirected ciliary currents (importing food and exporting waste through the mouth) likely made an asymmetric single-siphonoglyph protopharynx; and eight functionally complementary nematocyst-rich septa  and Ctenophora.Without giving reasons, Nielsen unjustifiably asserted ‘it seems impossible to derive eumetazoans from an adult sponge’ , p. 148.8.I have argued that the ancestral coelenterate was a bilateral octomerous stem anthozoan that lost choanocyte microvilli as neurally controlled nematocyst/tentacle feeding on larger prey improved, its mouth evolving from the osculum, and ostia closed  suppressing water channels, yielding a single body cavity, the coelenteron. Choanoderm and endoderm are homologous , as are Contrary to dogma, Anthozoa, Scyphozoa and Cubozoa are mostly triploblastic with true mesoderm –74. The Probably before Medusozoa originated, a stem coelenterate switched completely from benthic to planktonic life by evolving multiaxonemal macrocilia and comb plates  and losing nematocysts no longer required for settlement and accelerating oral and sexual development. This radical shift in adaptive zone and developmental fate of the ancestral planula larva entailed numerous unique innovations giving Ctenophora such a different body form from crown cnidarian adults, and unique embryology. In Cnidaria, the larval nervous system is concentrated largely aborally but degenerates during metamorphosis after settlement, being replaced by an oppositely polarized adult system with an oral focus . UnsurprPolypodium, a tentaculate triploblastic polypoid cnidarian (class Polypodiozoa) whose highly modified planula endoparasites sturgeon oocytes [Polypodium triploblasty supports treatment as a separate class outside diploblastic Hydrozoa [Polypodium clade might be sister to Hydrozoa, as some trees indicate [Ideas that the nervous system evolved twice or was lost by sponges –56 are u oocytes . PolypodHydrozoa , their aindicate . Myxozoaindicate  might haindicate , as expeBeroe [Others advocate one neural origin and invoke tree artefacts, giving more supporting details . Saying Beroe ). The beBeroe  with no Rapid divergence of Anthozoa (benthic nematocystous adults) and Ctenophora  neatly partitioned the Early Cambrian adaptive zone for predating larger prey. By not settling, ctenophores could evolve anal pores at the statocyst pole, enabling more efficient unidirectional ingestion and defecation currents independently of unianal bilateria, allowing secondary biradial gut symmetry by losing the siphonoglyph . Like ctenophores, early adult anthozoa probably relied on ciliary feeding (often helped by mucus secretion as in scleractinia). More complex barbed nematocysts  and toxins evolved divergently only after bilateria arose and became cnidarian prey.9.Renilla) or by apical invagination (Alcyonium); the stomodaeal cavity/invagination joins the coelenteron secondarily when the two separating epithelia at the stomodaeal base degenerate, making a novel opening [Renilla to fuse basally with the side of the developing coelenteron wall before the breakthrough, would immediately connect the pharyngeal cavity not with the coelenteron but through the body wall to the outside  develops separately from the coelenteron cavity by an apical inwardly projecting tissue mass that secondarily develops an inner cavity  almost without further modification, and separated its mechanical functions from those of a gut. The new through gut could retain digestive and absorptive functions, likely improved by modifying their positional control to regionally differentiate the former pharynx and be suppressed in the former coelenteron, now coelom. By focusing on burrowing and processing ingested sediment, nematocysts were lost and tentacles modified in function to simple mouthparts (or lost in some lineages). Such a radical change would necessarily dramatically affect embryology; unsurprisingly, thereupon two different ways immediately arose to stabilize mouth/anus formation in this protobilaterian: the proterostome/deuterostome bifurcation.Sequence phylogeny makes it virtually certain that the deuterostome ancestor was non-cephalized, whether a burrower like acorn worm or tentaculate like pterobranchs, possibly colonial like tunicate and salp. All these could readily have arisen from this tentaculate/burrowing intermediate. Lophotrochozoa also appear primitively to have had non-cephalized tentaculate or burrowing forms. The common ancestor of both groups can be argued to have been a tentaculate form, retaining pharyngeal ciliary currents that Anthozoa use in feeding, but a better burrower than burrowing sea anemones. This protobilaterian would be preadapted as ancestor of all major deuterostome and lophotrochozoan groups; acquiring ecdysis and very different mouthparts was more radical, yielding a priapulid-like ecdysozoan ancestor. Site-heterogeneous multigene analyses show that deuterostome acoels lost gut ,85 and pSarcophyton mitochondrial genome [Although lacking synapses, sponge tissues and embryology are as complex as in Cnidaria ,88–91; Hl genome  confirmel genome . Radial Drosophila eyes; these eyes evolved independently by modifying eukaryote cells (not strictly homologous with the bacteria that invented rhodopsin) and arranging them into contrasting supracellular structures. It is too often overlooked that structural homologies like those of tetrapod limb bones are at a higher level of organization than are transcription factors or building blocks such as collagen that they may share with morphologically non-homologous arthropod or annelid limbs and can often be recognized unambiguously entirely independently of gene sequences; there is almost certainly no ‘pentadactyl-limb gene’. Non-homologous structures  are often built partly of homologous components.Hyman's assertion  that non10.Haootia might be a muscular cnidarian impression [Acetabularia being superficial. Acetabularia is not syncytial; its form requiring cell walls is adapted for photosynthesis. Habitat proves that Vendozoa were not generally phototrophs [Codium are never quilted. Absorptive feeding by filamentous syncytial fungi like zygomycetes would cease if they evolved that body form. Large fungal fruiting bodies are non-trophic for spore dispersal. Syncytial sponges evolved secondarily from cellular ancestors. The largest protozoan syncytia (myxogastrid Mycetozoa) are naked phagotrophs with no architectural potential to evolve a vendozoan body form, unassignable to any protist group. Vendozoan complexity required extensive connective tissue to make quilt seams as struts supporting two outward facing trophic epithelia. Broken frondule internal structure [Rejecting the then prevalent idea that Ediacaran macrofossils antedating the Cambrian explosion included bilateria , I arguepression . Howeverpression , his anatotrophs . Syncytitructure  suggestsThat Vendozoa were osmoheterotrophs  is implaPhalansterium and spongomonads are possibilities that in principle might retain their feeding mechanism after evolving a multicellular differentiated tissue. But I strongly doubt any did, as evolving a multicellular phagotroph with tissues is difficult (see above) except via a flagellate/sponge pathway, and vendozoan timing just before sponges and eumetazoa can hardly be mere coincidence. Vendozoa flourished 580–541 Ma, becoming extinct at the Cambrian explosion approximately 541 Ma. Their reduced disparity and diversity 5 Ma before the Cambrian explosion [It is theoretically possible that Vendozoa arose independently of sponges by evolving a connective tissue in another colonial flagellate group—xplosion  I attribFunisia [Eocyathispongia [Several simpler, seemingly non-quilted, sessile Ediacaran fossils could also be presponges, e.g. the tubular Funisia . The 1 mispongia  is more ispongia . Crown sispongia . Unique Kimberella may belong here [Eocyathispongia) new subhylum Varisarca: Diagnosis: extinct macroscopic sessile multicells inferred to be ciliary filter feeding phagotrophs with epithelial/ECM organization; body form: variable arrangements of thin sheets, neither arranged in a quilted array , nor having ostia and internal water channels (unlike Porifera); non-mobile as adults. Etymology Vari variable sarco Gk flesh signifies variable body forms of epithelioid/ECM presponge fleshy organization.I regard Vendozoa as the oldest phylum of kingdom Animalia, distinct from Porifera, Placozoa and Eumetazoa. I divide it into subphylum Petalonamae  for petaong here ) and forTrichoplax be direct descendants of presponges.Vendozoa likely had Wnt/catenin axial patterning and ciliated planktonic larvae for dispersal, as without an AS they could not have easily brooded larvae as most sponges do (perhaps secondarily as protection after coelenterates evolved). If Placozoa are sisters of Eumetazoa as most multigene trees suggest, Placozoa were secondarily simplified by AS or coelenteron loss, evolving neotenously by prolonging the usual larval presettling benthic creeping phase by evolving extracellular digestion of benthic microbes and losing metamorphosis. Only if nested within Eumetazoa (Coelenterata plus bilateria) as some unconvincing trees suggest, need they have lost neurons also, like Myxozoa. Only if branching deeper than sponges and Eumetozoa, which multigene trees mostly exclude, could 11.The Cambrian explosion is the most striking animal example of ultrarapid origins of novel body forms: Simpson's quantum evolution, convincingly attributed to the invasion of previously unexploited major adaptive zones . Some leThere truly was an Early Cambrian explosion of animal (and protist) phyla, now ecologically and evolutionarily quite easy to understand. Such an explosion is expected for the very reasons that Darwin and Simpson convincingly explained. When a bilaterian with through gut and coelom arose, it created a new competitor-free adaptive zone and was bound to diversify rapidly into all body plans developmentally readily made by simple modifications and able to survive ecologically . It woulA widespread explanatorily empty speculation that many groups originated long before their objective fossil dates is fuelled by deep uniformitarian prejudices about evolutionary rates that palaeontology long ago refuted, and three other prejudices/biases that synergistically led to the notion of a ‘slow burning fuse’—a journalistic slogan, not critical evolutionary thought, evaluation or synthesis. First is excessive confidence in the certainly false idea of a ‘molecular clock’ and in the reliability of current implementations of oxymoronic ‘relaxed clock’ computer programs . Second 12.The best way to understand megaevolutionary events is by a coherent synthesis unifying data of every kind using explicit reasoning and well-tested explanatory principles. Haeckel's idea that animals evolved from a protozoan ancestor directly via a gastraea with triploblastic body, mouth, gut and anus, and that the animal archetype was a flatworm-like bilateral mobile predator like us minus coelom and anus must be wrong. A gastraea is far too complicated to evolve in one step. Instead, a choanoflagellate became a triploblastic sponge (arguably in two separate stages), a sponge became an anthozoan cnidarian, stem anthozoa generated pelagic ctenophores and independently an ancestral sessile bryozoan-like bilaterian, whose headless zoophyte descendants independently evolved morphologically contrasting heads through inventing burrowing, crawling or swimming, in annelids, molluscs, arthropods and vertebrates; all acoelomate bilateria arose secondarily by coelom occlusion. Nematocyst-triggered origin of neurons and zoophyte origin of bilateria adumbrated here put sessile headless animals central to eumetazoan and bilateria origins, just as they are to the already widely accepted choanoflagellate-sponge transition . All three problems are more deeply illuminated by a unifying zoophyte perspective than by Haeckel's anthropomorphic, self-mobile adult bias. Sessile presponge headless zoophytes with dispersive ciliated larvae were the first animals; muscle-driven mobility is secondary. Heads followed rather than led basic animal innovations. Can a simpler path fit the facts?


Persea americana Miller (Lauraceae), is an important fruit crop cultivated by small-holder farmers along Afrotropical highlands of Taita Hills in South-eastern Kenya and Mount Kilimanjaro in Northern Tanzania. The small-holder farmers in these East African regions generate substantial food and cash from avocado fruits. However, the avocado crop is faced with challenges of infestation by insect pests such as the common blossom thrips (Frankliniella schultzei Trybom) which feeds on pollen and floral tissue thereby reducing productivity of the trees. Moreover, there is no information describing distribution patterns of Frankliniella schultzei and associated weather in East African avocado orchards despite the fact that small-scale farming is dependent on rainfall. This article was, therefore, initiated to provide dataset on abundance of Frankliniella schultzei from the avocado plants that relates with monthly rainfall and air temperatures at Taita Hills and Mount Kilimanjaro. Frankliniella schultzei was collected using white coloured beating tray and camel brush whereas air temperatures (°C) and rainfall (mm) was recorded daily using automatic data loggers and rain gauge, respectively. The survey at the two transects commenced during peak flowering season of avocado crop in August up to end of harvesting period in July of the following year. Temporal datasets were generated by Kruskal-Wallis Chi-square test. Current temporal datasets presents strong baseline information specifically for Kenya and Tanzania government agencies to develop further agricultural strategies aimed at improving avocado farming within Taita Hills and Mount Kilimanjaro agro-ecosystems.Avocado,  Specifications TableValue of the data•Frankliniella schultzei presents baseline information specifically for Kenya and Tanzania government agencies, scientists and avocado farmers to enhance agricultural strategies.Abundance datasets of •Weather datasets contributes to precise climate information of Taita Hills and Mount Kilimanjaro.•The weather variables may be used to relate with avocado plant phenology  and also seasonal abundance of pest or beneficial insect species.1Frankliniella schultzei in avocado orchards at Taita Hills and Mount Kilimanjaro, respectively. Frankliniella schultzei, rainfall, maximum temperature (Tmax), and minimum temperature (Tmin) in avocado orchards at Taita Hills and Mount Kilimanjaro, respectively. Frankliniella schultzei Trybom) specimen and 2This study was carried out in small-scale avocado farmlands at Taita Hills in South-eastern Kenya and at the South-eastern slopes of Mount Kilimanjaro in Northern Tanzania as described by Frankliniella schultzei Trybom, was sampled monthly from avocado trees using a white coloured beating tray and camel brush for two years as described by Palmer Air temperature (°C) of avocado orchards was recorded daily along the study areas using data loggers. The data loggers were hang on the lower canopy of avocado trees at a height of 2 m. Rainfall records was obtained from rain gauge for two years between August 2012 and July 2014. Geographical coordinates and elevation of study areas was verified using Geographical position system (GPS) Garmin model eTrex 30. The common blossom thrips,


A 46-year-old woman had a delayed breast reconstruction using a free deep inferior epigastric perforator (DIEP) flap raised on a single medial row deep inferior epigastric perforator, complicated by intraoperative pedicle avulsion. Following a 1-cm excision of the intimally damaged vessel, the pedicle was repaired and the flap was salvaged using a supermicrosurgical technique.What is supermicrosurgery?What levels of pedicle injury can occur?What are the intraoperative strategies to salvage the free flap from pedicle avulsion?What scope is there for future advancements in free flap reconstruction using supermicrosurgery?Supermicrosurgery was first used in the 1980s, with successful replantation of distal fingertips.2Mishandling of the pedicle by the surgeon. This can result in intimal injury to an intact vessel.3Avulsion of the vascular pedicle by excessive traction. Single-vessel avulsion involves either arterial or venous supply to the flap.4Double-vessel avulsion of both arterial and venous supplies to the flap.5Diathermy injuries to the pedicle, from direct cautery or local thermal spread.6Anastomotic aneurysmal rupture, due to a pressure gradient created by mismatched donor and recipient vessels or a poorly constructed anastomosis.7A vascular pedicle injury is potentially disastrous in free flap surgery. Although uncommon, pedicle injury is an important surgical complication and understanding how it arises is essential to effective management. Different levels of pedicle injury are as follows:Before flap inset: If avulsion occurs while raising the flap, use an alternative perforator if available or repair the injured vessels if none available.During flap inset: If avulsion occurs during the flap transplant or microsurgical anastomosis with recipient vessels, reattempt a primary anastomosis. However, if the pedicle is now too short to reach the recipient vessel, use an interposition vein graft to lengthen the vascular pedicle and perform a tension-free anastomosis.3After flap inset: If avulsion occurs after the flap is inset, identify and perform an anastomosis with another shorter recipient vessel or repair and reanastomose the avulsed vessel.8Salvage of free flaps where smaller diameter vessel injury has occurred is now possible using supermicrosurgical techniques. Importantly, the surgeon must excise the zone of trauma prior to repair. Selecting the appropriate operative strategy to then repair the avulsed pedicle will depend on the nature of injury. We therefore propose an algorithm to support this selection:double-vessel avulsion, repairing the artery first allows the vein to fill and become more prominent for repair. In DIEP breast reconstruction specifically, repairing only the artery produces flap congestion and backpressure that expands the superficial inferior epigastric vein, which can then be used instead.For Free flap vascular pedicle injury remains uncommon but potentially devastating. However, free flap salvage is possible by use of innovative supermicrosurgical techniques and the operative strategies proposed. Supermicrosurgery has already brought a new frontier to free tissue transfer. Autologous transfer of tissue segments for congenital, oncological, and trauma-related defects is now possible. Perforator-to-perforator and free-style flaps, raised on anatomically variable vessels, have increased the diversity of reconstructive options, especially when smaller diameter vessels can be exploited using supermicrosurgical techniques. This will lead to spare-part surgery where disposable tissues with minimal donor morbidity can be used instead of locally destructive reconstructive options. Supermicrosurgical transfer of tissue and organ segments for facial allotransplantation and end-stage organ failure is being explored.


An enduring theme in microbial ecology is the interdependence of microbial community members. Interactions between community members include provision of cofactors, establishment of redox gradients, and turnover of key nutrients to drive biogeochemical cycles. An enduring theme in microbial ecology is the interdependence of microbial community members. Interactions between community members include provision of cofactors, establishment of redox gradients, and turnover of key nutrients to drive biogeochemical cycles. Pathways canonically conducted by isolated organisms in laboratory cultures are instead collective products of diverse and interchangeable microbes in the environment. Current sequence-based methods provide unprecedented access to uncultivated microorganisms, allowing prediction of previously cryptic roles in biogeochemical cycles and interactions within communities. A renewed focus on cultivation-based methods is required to test predictions derived from environmental sequence data sets and to address the exponential increase in genes lacking predicted functions. Characterization of enriched microbial consortia to annotate hypothetical proteins and identify previously unknown microbial functions can fundamentally change our understanding of biogeochemical cycles. As we gain understanding of microbial processes and interactions, our capacity to harness microbial activities to address anthropogenic impacts increases. Microorganisms represent the majority of life on earth, both in cell numbers and in diversity. They conduct critical ecosystem services, including biogeochemical cycling and contaminant transformations. Interactions between microbial community members are essential for maintenance of these ecosystem services. In most cases, microorganisms live within complex communities containing diverse populations, with key biogeochemical cycle steps distributed between community members. With significant heterogeneity across environments, characterization of microbial interactions is an ongoing endeavor within microbial ecology. High-throughput sequencing approaches, including total community DNA sequencing, or metagenomics, have provided a torrent of information on microbial distribution, diversity, and metabolic potential. This perspective argues that sequencing and enrichment cultivation together represent a powerful combination of new and old tools for examining microbial interactions and for identifying currently uncharacterized microbial activities.High-throughput sequencing combined with reconstruction of high-quality draft metagenome-associated genomes (MAGs) provides detailed information on taxonomy, metabolic potential, and organism abundance. Where initial metagenome studies were able to reconstruct genomes for the most abundant organisms in low-diversity environments, such as acid mine drainage or the oligotrophic ocean, genome binning from metagenome sequencing now routinely results in tens to hundreds, and even thousands, of MAGs for organisms within environments as complex as sediments and soils –4. This –2 . The majority of organisms implicated in denitrification contained a single pathway reaction encoded on their genomes, with the complete pathway dispersed across the microbial community. These trends held true for sulfur cycling as well, with the complete pathway from sulfide to sulfate present in only 3% of genomes for the organisms with predicted roles in sulfur oxidation, with single pathway reactions more commonly identified (2 formation is dependent on a functionally diverse community in many environments (With hundreds or thousands of MAGs available from a single environment, organismal roles in nutrient cycling can now be examined from a whole-community perspective, within the framework of partial genomes for the majority of populations. Deep metagenomic sequencing of a terrestrial sediment system identified a striking number of broken pathways within the thousands of reconstructed microbial genomes . The vas2 12 of 30, 3.6%.ronments . From thronments , alongsiThe current biggest challenge in environmental microbiology is annotating the thousands of hypothetical proteins identified from each new genome and metagenome—the known unknowns. Reconstructed genomes can have upward of 40 to 50% of their genes as predicted open reading frames with no known function . The datHow best to identify unknown functions within organisms whose obligate interactions with community members preclude isolate cultivation? Enrichment cultures, originally developed by pioneering environmental microbiologist Martinus Beijerinck at the turn of the 19th century, remain a relevant and powerful tool. Enrichment cultures streamline the environmental community to a subset tractable for experimentation. The enriched consortia are frequently more robust than isolate cultures, as beneficial microbial interactions are maintained (Nitrospira species capable of conducting complete nitrification used enrichment cultures to simplify an environmental community without the need for isolation. Metagenome sequencing of the enrichments to reconstruct complete or high-quality draft genomes proved cooccurrence of ammonia oxidation genes with the nitrite oxidoreductase for nitrite oxidation (Candidatus Methylomirabilis oxyfera,” the first genome for the NC10 phylum (Ca. Methylomirabilis oxyfera,” where oxygen is produced intracellularly by metabolizing nitrite via nitric oxide into oxygen and dinitrogen gas (Combinations of metagenome sequencing and enrichment cultivation have already dramatically impacted our understanding of biogeochemical cycles. A major revision to the nitrogen cycle came with the discovery of comammox , 11. Botxidation , 11. Simxidation . Metagen0 phylum . Stable ogen gas . While togen gas , furtherParcubacteria, a CPR lineage of otherwise-uncultivated phyla (Parcubacteria in culture will allow experimentation to optimize further cultivation attempts. Enrichment cultures containing Dehalococcoides mccartyi sp. have been developed as bioremediation tools for chlorinated solvents, recalcitrant and common groundwater contaminants. Metagenome sequencing of D. mccartyi enrichment cultures identified functionally conserved nondechlorinating microbes responsible for maintenance of a reducing environment, provision of cofactors including cobalamin, and carbon turnover (D. mccartyi. Notably, the organisms conducting these roles within different consortia varied, preventing prediction of interactions from taxonomic information alone (Beyond explorations of biogeochemical cycles, enrichment cultivation coupled with metagenomics can identify interactions present within microbial consortia that contribute to a population’s survival or that support a microbial activity of interest. An enrichment culture developed to examine benzene degradation contains the only known cultivated member of the ed phyla . Having turnover , activiton alone . EnrichmStudies connecting in-depth metagenomic characterization of environments with cultivation experiments to confirm predicted functions are the future of environmental microbiology . Bolster


Radiation-induced alopecia after fluoroscopically guided procedures is becoming more common due to an increasing use of endovascular procedures. It is characterized by geometric shapes of nonscarring alopecia related to the area of radiation. We report a case of a 46-year-old man presenting with asymptomatic, sharply demarcated rectangular, nonscarring alopecic patch on the occipital scalp following cerebral angiography with fistula embolization under fluoroscopy. His presentations were compatible with radiation-induced alopecia. Herein, we also report a novel scalp dermoscopic finding of blue-grey dots in a target pattern around yellow dots and follicles, which we detected in the lesion of radiation-induced alopecia. Radiation-induced alopecia after fluoroscopically guided procedures, although infrequently reported in the dermatological literature, is possibly becoming more common due to an increasing use of endovascular procedures. It is characterized by geometric shapes of nonscarring alopecia confined to the area of radiation, usually asymptomatic without signs of scalp inflammation. We report a case of a 46-year-old man presenting with asymptomatic, sharply demarcated rectangular, nonscarring alopecic patch on the occipital scalp, 1 month after cerebral angiography with fistula embolization under fluoroscopy. His distinct clinical presentations together with a history of endovascular embolization under fluoroscopy were compatible with radiation-induced alopecia.A 46-year-old Thai man presented with asymptomatic sharply demarcated rectangular balding patch on the occipital scalp of 2 weeks' duration. Ten months earlier, he had a car accident and was diagnosed as having traumatic right carotid-cavernous fistula and optic neuropathy. Cerebral angiography revealed direct carotid-cavernous fistula from the C1 segment of cavernous part of the right internal carotid artery to the right cavernous sinus. Six months later, he was treated with transarterial balloon embolization without any complications. A month earlier, a follow-up magnetic resonance imaging and magnetic resonance angiography of the brain showed residual shunt. He underwent a cerebral angiogram with transvenous coil and glue embolization under fluoroscopy. The total procedure duration and the total radiation exposure time were 150 minutes and 67 minutes, respectively. The peak skin dose was 2.9 Gy. Two weeks after the procedure, he presented at the outpatient clinic with asymptomatic balding patch on his occipital scalp. Dermatologic examination revealed a sharply demarcated rectangular nonscarring alopecic patch, measuring 10 × 12 centimeters in size, on the occipital scalp, without erythema or scaling Figures . A 4 mm Fluoroscopically guided endovascular procedures are becoming more common in medical practice with an increasing use of minimally invasive techniques for various medical conditions. In general, radiation-induced alopecia is common and develops following radiation therapy of neoplasms. However, alopecia secondary to radiation exposure from fluoroscopically guided procedures is increasingly being reported due to an increasing use of endovascular procedures.Alopecia after fluoroscopically guided endovascular procedures is rarely reported in the dermatological literature, although this complication seems to be relatively common. Given that the procedures often require prolonged and repeated fluoroscopic imaging, doses above a deterministic threshold can potentially react with the skin and hair follicles , 2. The Pathogenesis of radiation-induced alopecia involves acute damage to actively dividing matrix cells of anagen follicles causing immediate loss of dystrophic anagen hairs (anagen effluvium) and premature entry of some anagen hair follicles into catagen and then into telogen phase, resulting in delayed onset of hair shedding (telogen effluvium) . RadiatiClinical presentation of radiation-induced alopecia includes geometric shapes of nonscarring alopecic patch confined to the area of radiation, usually asymptomatic without signs of scalp inflammation. Occipital, parietal, and temporal scalp are commonly affected. Occasionally, the alopecic area is clinically similar to other nonscarring alopecia cases such as alopecia areata. However, important clues to diagnosis are a history of fluoroscopically guided procedure and alopecic area which is confined to the radiation site. Hair loss mostly occurs within 1–3 weeks after radiation exposure with spontaneous regrowth of hair within 2–4 months –14. Cho Diagnosis of radiation-induced alopecia is primarily based on distinct clinical presentations and history of radiation exposure. Investigations including trichogram, dermoscopy, and histopathology are more likely to exclude other causes of nonscarring alopecia rather than confirming the diagnosis. Trichogram performed during an early phase usually shows dystrophic anagen hairs. However, in our patient, the presence of telogen hairs without dystrophic anagen in trichogram was a result of prolonged time from radiation exposure to the examination. All dystrophic anagen hairs shed off in the first few weeks after radiation exposure. A study of 10 patients with postangioembolization alopecia by Cho et al. reported that dermoscopic findings demonstrated both yellow dots and black dots (60%), short vellus hair (50%), peripilar sign (20%), broken hair (10%), coiled hair (10%), and white dots (10%), whereas histopathology showed increased numbers of catagen and telogen hairs without peribulbar inflammatory cell infiltrate, unlike alopecia areata, which usually shows peribulbar inflammation . AlthougOther possible mechanisms of alopecia after fluoroscopy-guided endovascular embolization including alopecia secondary to an impairment of the external carotid blood supply of the scalp and pressure-induced alopecia should be also considered. The impairment of the external carotid blood supply of the scalp may result from therapeutic embolization of blood vessels or radiation-induced luminal fibrosis . Owing tTreatment of radiation-induced alopecia is usually unnecessary due to its benign and self-limiting nature. Complete hair regrowth generally occurs within 2–4 months after irradiation –14. In aIn conclusion, we report the case of radiation-induced temporary alopecia after endovascular embolization under fluoroscopy. In addition to other nonscarring alopecia cases, physicians should be aware of and include this condition in the differential diagnosis when individuals presenting with nonscarring alopecia had a history of prolonged fluoroscopic endovascular procedures. Although the condition is benign and self-limiting, awareness, monitoring, and limitation of radiation exposure to the patient are the most important strategies to prevent hair loss.


Intracellular bacterial pathogens replicate within eukaryotic cells and display unique adaptations that support key infection events including invasion, replication, immune evasion, and dissemination. From invasion to dissemination, all stages of the intracellular bacterial life cycle share the same three-dimensional cytosolic space containing the host cytoskeleton. For successful infection and replication, many pathogens hijack the cytoskeleton using effector proteins introduced into the host cytosol by specialized secretion systems. A subset of effectors contains eukaryotic-like motifs that mimic host proteins to exploit signaling and modify specific cytoskeletal components such as actin and microtubules. Cytoskeletal rearrangement promotes numerous events that are beneficial to the pathogen, including internalization of bacteria, structural support for bacteria-containing vacuoles, altered vesicular trafficking, actin-dependent bacterial movement, and pathogen dissemination. This review highlights a diverse group of obligate intracellular bacterial pathogens that manipulate the host cytoskeleton to thrive within eukaryotic cells and discusses underlying molecular mechanisms that promote these dynamic host-pathogen interactions. Obligate intracellular bacteria comprise a group of highly infectious human pathogens that cause a spectrum of life threatening diseases. These bacteria demonstrate remarkable adaptations that support intracellular lifestyles comprised of unique challenges and advantages. To support diverse infection events, intracellular pathogens have evolved numerous methods to subvert host molecular signaling machinery that controls essential cellular functions. Intracellular pathogens display tropism for specific eukaryotic cell types and attach to these cells to initiate internalization, allowing intracellular growth using energy rich host metabolic products. Successful immune evasion is essential for productive infection, and the intracellular environment provides temporary cover from detection by immune cells and antibody/compliment-mediated immunity. Following replication, these pathogens exit the infected cell and disseminate to additional sites of infection.in vitro. These human pathogens include Chlamydia , Rickettsia , Anaplasma (human granulocytic anaplasmosis), and Ehrlichia (human monocytic ehrlichiosis) species. We also include the Q fever agent Coxiella burnetii, which replicates exclusively within host cells during natural infection, but can be cultured in specialized axenic media in vitro. The interaction of these pathogens with the host cytoskeletal network has been studied for many years, but detailed molecular mechanisms are lacking due to difficulty in genetically manipulating these bacteria. However, recent genetic breakthroughs provide tractable systems by which bacteria-actin interactions can now be defined for this intriguing group of pathogens -actin that hydrolyzes ATP to provide energy for F-actin polymerization Reisler, . Actin nHost cell entry is the first step in the intracellular pathogen invasion process. Bacterial cell wall components facilitate attachment to specific host cells via distinct receptors. Pathogens use the rearranging abilities of actin to facilitate rapid entry into the host cell, with phagocytic and non-phagocytic cells internalizing bacteria by different mechanisms Figure . BacteriC. burnetii preferentially infects phagocytic human macrophages via binding to CR3 receptors, triggering reorganization of filamentous actin at the attachment site  is essential for invasion -anchored protein DNase X located on the cell surface. This interaction triggers cytoskeletal rearrangement and filopodia formation  domain that binds to actin monomers and WASP proteins. RickA ultimately activates the Arp2/3 complex, promoting actin polymerization and intracellular motility  in macrophages. F-actin assembles around the PV and is required for optimal vacuole formation  signaling using a type IV secretion system, and this process is essential for PV formation  observed in infected patients. Typhus group Rickettsia do not form actin tails (Heinzen et al., Spotted Fever group E. chaffeensis uses two different exit strategies during early and late stages of infection (Thomas et al., E. chaffeensis is transported into filopodia that extend into neighboring cells, providing a direct path for bacterial entry into nearby cells without immune detection. Actin rearrangement in infected cells is essential for filopodia formation and bacterial localization into filopodia. However, during late stages of infection, bacteria exit following host cell lysis (Thomas et al., Anaplasma, it has been suggested that A. phagocytophilum exits host granulocytes by exocytosis and host cell lysis. However, specific roles for cytoskeletal components during pathogen exit have not been characterized (Rikihisa, Chlamydia can exit host cells by host cell lysis or extrusion of the inclusion. Extrusion requires actin rearrangement and is considered an exocytosis-like process. Extrusion initiates with formation of inclusion-containing protrusions from infected cells. Extrusions are then pinched into separable compartments located at the cell periphery, allowing inclusion release without triggering cell death (Todd and Caldwell, Obligate intracellular bacteria include a unique group of human pathogens that have adapted to thrive within the host cell environment. These pathogens hijack host cell signaling by secreting bacterial effector proteins into the host cytosol to promote formation of a replication-permissive niche. Intracellular bacteria constantly encounter the dynamic host cytoskeletal network and have evolved methods to use actin and related proteins to facilitate infection. Intracellular pathogens rearrange the actin cytoskeleton during internalization by phagocytic and non-phagocytic host cells. Some intracellular bacteria replicate within a specialized membrane-bound vacuole, while others replicate free in the cytosol. Host cytoskeletal proteins often form a filamentous cage around bacteria-containing vacuoles, providing structural support and allowing vesicle fusion with the vacuole. Other intracellular bacteria stimulate actin polymerization and form actin tails that aid bacterial movement within the host cytosol and propulsion into neighboring cells. Intracellular bacteria further manipulate the actin cytoskeleton to exit host cells by lysis or vacuole extrusion. In conclusion, major events in bacterial lifecycles require host cytoskeletal components, and blocking these interactions negatively impacts bacterial replication. However, we clearly have more to learn about the molecular mechanisms controlling pathogen interaction with the host cytoskeleton. Targeting these interactions may provide novel approaches to develop antibacterial therapeutics targeting obligate intracellular bacterial pathogens.All authors listed, have made substantial, direct and intellectual contribution to the work, and approved it for publication.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Eutrophication and climate warming are profoundly affecting fish in many freshwater lakes. Understanding the specific effects of these stressors is critical for development of effective adaptation and remediation strategies for conserving fish populations in a changing environment. Ecological niche models that incorporated the individual effects of nutrient concentration and climate were developed for 25 species of fish sampled in standard gillnet surveys from 1,577 Minnesota lakes. Lake phosphorus concentrations and climates were hindcasted to a pre-disturbance period of 1896–1925 using existing land use models and historical temperature data. Then historical fish assemblages were reconstructed using the ecological niche models. Substantial changes were noted when reconstructed fish assemblages were compared to those from the contemporary period (1981–2010). Disentangling the sometimes opposing, sometimes compounding, effects of eutrophication and climate warming was critical for understanding changes in fish assemblages. Reconstructed abundances of eutrophication-tolerant, warmwater taxa increased in prairie lakes that experienced significant eutrophication and climate warming. Eutrophication-intolerant, warmwater taxa abundance increased in forest lakes where primarily climate warming was the stressor. Coolwater fish declined in abundance in both ecoregions. Large changes in modeled abundance occurred when the effects of both climate and eutrophication operated in the same direction for some species. Conversely, the effects of climate warming and eutrophication operated in opposing directions for other species and dampened net changes in abundance. Quantifying the specific effects of climate and eutrophication will allow water resource managers to better understand how lakes have changed and provide expectations for sustainable fish assemblages in the future. Climate change and eutrophication are potent environmental stressors that are altering the fundamental ecological processes that structure freshwater communities throughout the world ,2. SigniAs climate changes the thermal structure of lakes, fish are influenced through a number of pathways and processes ,9. EpiliEutrophication has substantially altered fish assemblages around the world  and can Eutrophication and climate warming have affected many lakes in Minnesota ,26. NatiEcological niche modeling that incorporates specific effects of individual stressors has proven useful for disentangling multiple-stressor impacts on biotic communities ,33. EachUnderstanding the specific effects of eutrophication and climate warming is of critical importance for water resource managers and policy makers. Minnesota’s lake fish taxa differ in their thermal and eutrophication tolerances ,29 and rEmpirical ecological niche models were developed for 25 species of fish commonly captured in Minnesota Department of Natural Resources (MDNR) lake netting assessments . The MinTaxa-specific ecological niche models were developed using generalized additive models (GAMs). The effects of several environmental and ecological stressor variables on the relative abundance of each species were quantified with the models. GAMs are particularly useful for modeling the ecological niche of biological organisms because they directly incorporate nonlinear smoothing to describe the effects of predictors on the response variable  and are http://prism.oregonstate.edu, retrieved 3 Dec 2015). Air temperatures were estimated for the geographical center of each lake for the period from 1981–2010 using the geoknife package in R [Mean annual air temperature (MAT), which integrates a number of ecological effects across all seasons, was used as a proxy variable that captured the effects of climate on fish assemblages. Lake-specific air temperature data were developed from high resolution spatial interpolations of historic air temperatures available from the Oregon State University PRISM Climate Group .Phosphorus was selected as a proxy variable that captured the effects of eutrophication because it is the primary limiting nutrient in many lakes, including Minnesota ,30. Meanmgcv  and by z-scores (subtracting the species-specific mean and dividing by the species-specific standard deviation) to allow for standardized comparisons among species. Lake size (ha) and depth (m) were log transformed and total alkalinity was square root transformed to normalize their distributions. Response surfaces of relative abundances were estimated from joint smooths of MAT and TP. The joint smooths allowed for the visualization of how responses of each variable changed as a function of the other variable. Individual responses can be visualized as slices through the response surface at specific values of the other variable. These interaction of MAT and TP can also be visualized as the shape of the slices change as a function of the other variable. Environmental variables  were entered into the model as single variable smooth terms. A Gaussian family, with an identity link function, was used for each GAM and a dimension basis of k = 4, was used to limit the degrees of freedom for each smooth term [The  package  in the soth term .Pre-disturbance lake conditions were estimated for climate and eutrophication variables which provided inputs into species-specific ecological niche models. Pre-disturbance climate was calculated from historical air temperatures retrieved from the PRISM data set using identical methods as the contemporary climate data for comparison to present-day conditions. Annual lake-specific air temperatures from 1896–1925 (a thirty year period of the same duration as the contemporary period) were processed using the geoknife package in R , and meaTP = mean summer epilimnetic total phosphorus concentration (μg/l), depth = maximum lake depth (m), outwash = proportion of glacial outwash soils in watershed, disturbance = proportion of disturbed land uses  in watershed. Assuming depth and proportion of outwash soils remained constant over both periods, estimation of pre-disturbance lake productivity collapses to:TPp = pre-disturbance mean summer epilimnetic phosphorus concentration (μg/l), TPc = contemporary, observed mean summer epilimnetic phosphorus concentration (μg/l), and disturbance = proportion disturbed land uses  currently in watershed. Watershed delineations were available for 1,236 lakes in the analysis. Contemporary values of land uses were calculated with the 2001 National Land Cover Database [Pre-disturbance land use corresponded to the late 1800s when prairies were first converted to agriculture and urban areas started to develop in Minnesota. Pre-disturbance eutrophication status was estimated by hindcasting the lake phosphorus model developed by Cross and Jacobson  to undisDatabase . Level 1Pre-disturbance fish assemblages were estimated by using the derived historical conditions as inputs to taxa-specific ecological niche models. Lake-specific CPEs were calculated from the fitted GAM models for each species to explore changes in abundance from pre-disturbance to contemporary conditions. The effects of climate warming and eutrophication on the relative abundance of each taxa were examined independently by using pre-disturbance conditions for one stressor while holding the other at contemporary levels, and then collectively by using pre-disturbance levels for both stressors.Generalized additive models described between 1.2 and 57.6% of deviance in fish relative abundance for the 25 species . Joint fResponse shapes of the joint effects of climate and productivity varied considerably between species . Some haMean annual air temperatures increased from pre-disturbance (1896–1925) to contemporary (1981–2010) times, although the magnitude of increase varied substantially between lakes and ecoregions . TemperaHindcasted total phosphorus concentrations indicated that lakes in the transition and prairie ecoregions have become more productive . Those eTrajectories of individual lakes within a 2-dimensional climate and productivity niche space  illustraStressor-specific changes in modeled fish relative abundance varied considerably by species and ecoregion . The dirFor most taxa, hindcast changes in abundance were largest in prairie lakes, where both climate and productivity shifts were substantial. All nine eutrophication-tolerant warmwater species were estimated to have increased in abundance from increased phosphorus and temperature, while six intolerant species  declined over the period. Stressors operated in opposite directions for a total of eight species, several of which experienced little change in abundance. Eutrophication-tolerant coolwater yellow perch benefited from increased total phosphorus, but warmer temperatures were detrimental. Conversely, intolerant warmwater bluegill benefited from warming temperatures, but gains in abundance were overwhelmed by detrimental effects of increased phosphorus. In general, eutrophication produced larger effects on relative abundance than climate warming for most taxa in the prairie ecoregion.Fish assemblages in the transition ecoregion also experienced significant changes after disturbance. Taxa-specific responses were similar to the prairie ecoregion with eutrophication-tolerant warmwater species benefitting most from increased temperatures and phosphorus concentrations. Increased phosphorus concentrations were detrimental to a number of eutrophication-intolerant species and increased temperatures were beneficial to warmwater taxa. Tolerant coolwater yellow perch also benefited from increased lake productivities, but increased temperatures were detrimental. Intolerant warmwater bluegill gains in abundance from increased temperatures were completely offset by losses from increased phosphorus concentrations and resulted in no net change. A total of 12 species in the transition ecoregion were affected by stressors that operated in opposite directions.Fishes in the forested ecoregion experienced changes primarily driven by temperature increases in the past century. Warmwater fishes such as black crappie, bluegill, largemouth bass, pumpkinseed, and yellow bullhead increased in abundance. The greatest abundance declines were seen within coolwater species such as white sucker, walleye, yellow perch, and smallmouth bass. The number of species affected by stressors that operated in opposite directions was large (15), but the net effect of increased phosphorus was very small in all cases.Grouping of taxa by thermal preference and eutrophication tolerance summarized fish assemblage changes that have occurred in the past century . WarmwatA century of eutrophication and climate warming profoundly affected lakes in Minnesota and consequently, their fish assemblages. Conversions of native prairies and forests to agricultural and urban uses increased nutrient concentrations in the prairie and transition ecoregions of the state. In response, abundances of eutrophication-tolerant fish likely increased. Climate also warmed in the past century throughout the entire state and reconstructed warmwater fish abundances increased accordingly. Trajectories of lake-specific changes spanned an extraordinary range of the 2-dimensional niche space represented by these two stressors in Minnesota. Together, eutrophication and climate warming likely drove an expansion of tolerant warmwater fish abundance at the expense of intolerant coolwater taxa. Conversely, climate and productivity shifts produced opposing effects on abundance of several species. For example, tolerant coolwater yellow perch likely benefitted from lake productivity increases, but climate warming was detrimental. Intolerant warmwater bluegill benefitted from climate warming, but gains in abundance were overwhelmed by negative effects of eutrophication. Disentangling the sometimes opposing, sometimes compounding, effects of these two important ecological stressors was critical for understanding changes in fish assemblages. Taxa-specific ecological niche models and explicit identification of stressor-specific tolerance guilds  were valEutrophication drove changes in modeled lake fish assemblages in the prairie and transition ecoregions of Minnesota more than climate. Vegetated fish habitat in the many shallow lakes of the prairie was likely reduced by nutrient enrichment . Shifts Climate drove modeled fish assemblage change in forested ecoregion lakes more than eutrophication. Watersheds of these lakes remained largely undisturbed and forested  resultinAlthough sequence and timing of individual stressor effects was not determined in this study, evidence exists that eutrophication was most important during the first part of the century and climate became more significant in recent years. Increases in mean annual temperatures in the state have accelerated since 1980 . Many laThe effects of climate warming on fish are expected to continue in all three ecoregions. Lakes in the forested ecoregion are likely to maintain suitable thermal habitat for cool- and warmwater species even under extreme warming scenarios ,55. ThusAlthough ecological niche modeling was useful for disentangling the effects of climate and eutrophication, significant uncertainty remains for predicting fish assemblage responses. Model errors propagated through the phosphorus hindcasting model and fish assemblage reconstruction were likely present. Modeled niches were assumed to remain unchanged from historic to contemporary conditions . PredictUnderstanding the specific effects of climate and land use change on lakes and fish assemblages is critical for developing management and adaptation strategies for effective conservation ,69. For S1 TableP-values and percent deviance explained for generalized additive models of the relative abundance of 25 fish species predicted by mean annual temperature (MAT) and mean summer epilimnetic total phosphorus concentrations (TP), depth, area, and alkalinity sampled in 1,577 Minnesota lakes.(XLSX)Click here for additional data file.S2 TableLake specific data used for ecological niche models developed from 1,577 Minnesota lakes. Metadata describing each data field are described in the first portion of the Table.(XLSX)Click here for additional data file.S1 FigGeneralized additive model responses of mean annual temperature (MAT °C) and mean summer epilimnetic total phosphorus concentrations (TP μg/l), depth (m), area (ha), and alkalinity on the relative abundance of 25 fish species sampled in 1,577 Minnesota lakes. Species codes are defined in (PDF)Click here for additional data file.


Bronchopulmonary dysplasia (BPD) is a chronic lung disease most commonly seen in premature infants who required mechanical ventilation and oxygen therapy for acute respiratory distress. While advances in neonatal care have resulted in improved survival rates of premature infants, limited progress has been made in reducing rates of BPD. Lack of progress may in part be attributed to the limited therapeutic options available for prevention and treatment of BPD. Several lung-protective strategies have been shown to reduce risks, including use of non-invasive support, as well as early extubation and volume ventilation when intubation is required. These approaches, along with optimal nutrition and medical therapy, decrease risk of BPD; however, impacts on long-term outcomes are poorly defined. Characterization of late outcomes remain a challenge as rapid advances in medical management result in current adult BPD survivors representing outdated neonatal care. While pulmonary disease improves with growth, long-term follow-up studies raise concerns for persistent pulmonary dysfunction; asthma-like symptoms and exercise intolerance in young adults after BPD. Abnormal ventilatory responses and pulmonary hypertension can further complicate disease. These pulmonary morbidities, combined with environmental and infectious exposures, may result in significant long-term pulmonary sequalae and represent a growing burden on health systems. Additional longitudinal studies are needed to determine outcomes beyond the second decade, and define risk factors and optimal treatment for late sequalae of disease. Bronchopulmonary dysplasia (BPD) is a chronic lung disease most commonly seen in premature infants who required mechanical ventilation and oxygen therapy for acute respiratory distress but can also occur in neonates that had a less severe respiratory course ,2,3. BPDThe incidence of BPD in surviving infants less than or equal to 28 weeks gestational age has been relatively stable at approximately 40% over the last few decades ,10,11,12Criteria to define BPD have historically lacked uniformity. The earliest clinical definition of BPD was limited to oxygen requirement at 28 days with consistent radiologic changes. These were originally modified to include continuing need for oxygen therapy at 36 weeks corrected gestational age (CGA). However, this definition inadequately addresses highly variable clinical practices as well as the wide range of disease, leading to further modification to include a severity assessment at 36 weeks gestational age . The defAt greatest risk of poor long-term outcome are the approximately 25% of infants with BPD who are identified to have elevated pulmonary pressures or BPD-associated pulmonary hypertension (PH), resulting in mortality rates as high as 14%–38% with one series noting only 25% survival at 2–3 years of age with severe BPD-associated PH ,22,23,24As the population of NICU survivors grow, long-term manifestations of chronic lung injury with BPD is likely to represent a greater burden to health systems. This paper seeks to review the pathophysiology of BPD, recent management strategies and what is currently known about long-term pulmonary outcomes in survivors.The phenotype seen with BPD is the end result of a complex multifactorial process in which various pre- and postnatal factors compromise normal development in the immature lung.  The specBPD occurs almost exclusively in preterm infants that have received positive pressure ventilation suggesting that mechanical lung over-distension and alveolar stretch play a critical role in the pathogenesis of BPD. Ineffective pulmonary mechanics results in need for ventilatory assistance at birth. Recent data suggests that 65% of preterm infants born at 22 to 28 weeks gestational age are intubated in the delivery room, which has decreased since 1993 when 80% of this population was intubated immediately following birth . The preStudies in numerous animal models have identified that exposure to supraphysiologic oxygen alone induces a phenotype comparable to that seen with BPD, including compromised alveolar development and pulmonary vascular remodeling . ClinicaAnimal models suggest that even brief exposures to high concentrations of oxygen can result in long-term morphologic and functional changes in the lung . In addiEscherichia Coli endotoxin to pregnant ewes resulted in amplified inflammation with ventilation of the exposed preterm lambs including evidence of cellular apoptosis and compromised alveolar development [Controversy exists regarding the contribution of chorioamnionitis and prenatal inflammation to the risk of developing BPD . Clinicaelopment ,45. Whilelopment . This paelopment . Controvelopment .Less controversy exists regarding the contribution of postnatal inflammation or nosocomial infection to the increased risk of developing BPD ,48. NoviPreterm infants that are small for gestational age (SGA) at birth or with intrauterine growth restriction (IUGR) are at increased risk for adverse pulmonary outcomes ,54. StudExtremely premature infants are at additional risk for postnatal growth restriction secondary to challenges of delivering optimal nutrition. Despite significant advances in the content and use of both enteral and parental nutritional support, 55% of infants born less than 27 weeks gestation demonstrate growth failure with weight less than 10th percentile at 36 weeks postmenstrual age ,26,53. PWhile BPD results from cumulative exposures to both the pre- and postnatal factors noted above, there is a growing interest in the heritable contributions to development of BPD. Twin studies provide insight into genetic predispositions as monozygotic twins share 100% of their genetic information while dizygotic twins are 50% concordant . A totalMore recently, several genome-wide association studies (GWAS) have been conducted to identify candidate single nucleotide polymorphisms associated with BPD. The largest evaluated over 1700 infants and failed to identify genomic loci or pathways that accounted for the previously described heritability for BPD . A seconManagement strategies are aimed at protecting against lung injury and the development of BPD. As the pathogenesis of disease is multifactorial, diverse approaches have been adopted including both ventilation and medical strategies. Interestingly, both antenatal steroids and surfactant reduce rates of RDS and improve survival; however, neither has been shown to reduce incidence of BPD .2 targets (>52 versus <48 mmHg) [Evidence of lung injury induced by volutrauma has led to efforts to promote “gentle ventilation,” in part through the use of permissive hypercapnia. However, the data to support this strategy has been inconsistent and long-term neurodevelopmental outcomes remain unknown ,69. None48 mmHg) ,71. Gent48 mmHg) ,73,74. R48 mmHg) ,76; howe48 mmHg) . As manySignificant efforts have been made to move away from use of invasive ventilation over the past two decades . Meta-anUse of alternative non-invasive modalities including non-invasive positive pressure ventilation (NIPPV), bilevel nasal CPAP (biPAP) and high-flow nasal cannula (HFNC) is also increasing with some evidence to suggest these modes may also be effective in managing neonatal respiratory disease ,82,83. SAdditional controversy exists surrounding the use of non-synchronized NIPPV. Synchronization can be achieved and has been described in premature infants using an abdominal pneumatic (or Graseby) capsule to detect diaphragmatic descent. This approach has the theoretical advantage of ensuring glottis patency before flow is triggered. While randomized trials using SNIPPV have not been performed, a retrospective study suggested comparable clinical outcomes, including rates of BPD, in infants managed with synchronized as compared to non-synchronized NIPPV . More reFinally, for those infants who do require intubation, an early trial of extubation is encouraged to potentially reduce risks of ventilator-induced lung injury. Studies have identified that an early attempt at extubation alone may decrease the risk of BPD, regardless of need for reintubation or duration of ventilation ,89. ThesExtensive efforts have been made to define optimal saturation targets for premature infants with ongoing concerns for the quality of evidence available . A meta-Specific to the outcome of BPD, the Surfactant, Positive Pressure and Oxygenation Randomization Trial (SUPPORT) conducted in the US found slightly lower rates of BPD (38% vs. 41.7%) in the low-saturation group without statistical significance . The CanPostnatal glucocorticoids are recognized to reduce rates of BPD via reduced inflammation as well as the induction of lung maturational changes. However, the potential benefits of systemic steroids are often outweighed by concerns for long-term neurodevelopmental sequelae including increased risk of cerebral palsy (CP) ,99. RateIn attempt to more reliably deliver corticosteroids directly to alveoli of infants at risk of early lung injury, co-administration of budesonide with surfactant has been considered. Yeh, et al. have reported significant decrease in the incidence of BPD or death in ventilated infants with severe RDS who received endotracheal budesonide . AdditioIn the recent randomized, multicenter Caffeine for Apnea of Prematurity (CAP) trial, early initiation of caffeine was found to result in lower incidence of BPD as well as a shorter course of respiratory support as compared to controls . The speVitamin A deficiency may predispose to chronic lung disease as it plays a critical role in maintaining the integrity of respiratory tract epithelium and is a key regulator of normal lung growth ,111. WhiInhaled nitric oxide (iNO) for prevention of bronchopulmonary dysplasia deserves mention as it has been explored in numerous studies with inconsistent findings ,116. WhiLong-term outcomes of BPD remain difficult to characterize as adult populations currently available to study represent survivors of outdated care. While BPD tends to improve with ongoing lung development, data available from follow-up studies identify concerns for persistent pulmonary sequelae of disease. Notably, the impact on late pulmonary health as well as the consequences of additional infectious or environmental exposures remain poorly characterized.It is important to recognize the tremendous emotional, medical, and financial efforts invested into care of extremely premature infants. The mean length of hospitalization for those born under 1000 grams is approximately 60 days with high rates of need for additional medical support including rehospitalization after discharge . During Numerous studies have evaluated long-term pulmonary function after premature birth alone. A meta-analysis including 59 articles identified that percent of predicted forced expiratory volume in 1 second (FEV1) is decreased in preterm-born survivors, even in patients who did not have a history of BPD . EvaluatAdditional studies have raised concerns for the persistence of pulmonary disease in patients with BPD. Correlation between maximal flow at functional residual capacity in infancy and forced expiratory flow at 8 years raises concerns that limited recovery occurs during late stages of alveolarization . In addiExposure to environmental insults including respiratory infections, tobacco and pollution may complicate resolution of BPD and prolong risks of pulmonary morbidity ,142. PreBeyond environmental insults, pulmonary morbidity in ex-preterm infants can be complicated by chronic reflux and microaspiration with risk of aspiration pneumonia and/or chronic inflammation. While the evidence is variable, improvement of respiratory status has been reported after Nissen fundoplication of infants with severe chronic lung disease with some programs advocating for this intervention in severe cases ,147,148.Many survivors of BPD demonstrate a component of reactive airway disease. Long-term follow-up of infants born <26 weeks gestation identified that 25% had an asthma diagnosis at 11 years of age while over twice this percentage (56%) had evidence of abnormal spirometry . While c2 consistent with alveolar hypoventilation [Survivors of BPD may experience exacerbation of pulmonary morbidities with exercise or exposure to hypoxia. Significant risk of exercise-induced bronchoconstriction has been demonstrated in children with BPD consistent with concerns for reactive airway disease noted above . Howevertilation .Chemoreceptor function in preterm infants is dysmature, resulting in abnormal ventilatory responses to alterations of oxygen content in these patients. Specifically, normal responses of increased ventilation with hypoxia as well as decreased ventilation with hyperoxia may be altered. Persistent chemoreceptor dysfunction has been documented in survivors of BPD ,157. InaDysmorphic pulmonary vasculature and compromised angiogenesis with BPD results in risk of elevated pulmonary pressures or BPD-associated pulmonary hypertension (PH). Retrospective studies of infants with BPD suggested that 25%–37% of infants with BPD develop associated PH, but these data are limited by inconsistent definition and screening protocols ,22,23. EAs previously noted, patients with BPD and elevated pulmonary pressures are at high risk for PH crisis and early mortality. Those who survive may ultimately demonstrate resolution of disease with additional lung growth. However, recent data suggests that subclinical right ventricular dysfunction can be detected in children assumed to have recovered from BPD-associated PH . Longer-While severity of BPD does not predict likelihood of associated pulmonary hypertension, sicker infants with prolonged ventilator and oxygen requirements are at greatest risk ,161. AddHowever, the threshold for intervention and optimal treatment of BPD-associated PH remains elusive with limited evidence to guide care . While rSome guidance for treatment has been provided in the literature. However, none of these to date are evidence based ,167. KulWhile the past decade has been notable for increasing use of vasodilator therapy in the treatment of BPD-associated PH , randomiThe definition, pathophysiology, and management of bronchopulmonary dysplasia (BPD) has evolved significantly since first described by Northway almost 50 years ago. Advances in neonatal care have resulted in increased rates of survival of extremely premature infants leading to both a new set of management challenges as well as an emerging population of long-term survivors of BPD. Interdisciplinary care to address the complex pulmonary, nutritional and developmental needs of these patients is critical and may itself influence outcomes of severe BPD .Randomized therapeutic studies in addition to longitudinal evaluation of these patients remains essential to optimize care and further discern risk factors for morbidity. While these studies require significant resources, they are much in need as evidence for optimal treatment is lacking. In addition, little is known regarding pulmonary outcomes of BPD beyond the second decade of life. Of concern are data highlighting the potential for increased risk of subclinical right ventricular dysfunction, obstructive lung disease, exercise intolerance, and asthma-like symptoms in survivors. Abnormal response to hypoxia and central airway disease may further exacerbate illness with risks of sleep disordered breathing. These baseline morbidities, complicated by environmental and infectious exposures, may represent a significant challenge for the aging cohort of BPD survivors.As trends demonstrate increasing survival of extremely premature infants, nearly half of whom will be diagnosed with BPD, it is imperative that future studies investigate mechanisms and risk factors influencing long-term morbidity with an overall goal of reducing the burden of disease.


We administered neuropsychological measures considered sensitive to prefrontal dysfunction  to obsessive-compulsive disorder (OCD) patients and control subjects. OCD subjects exhibited performance deficits, in comparison to community controls, on three measures sensitive to orbitofrontal neocortex dysfunction. Contrary to expectation, OCD patients also exhibited performance deficits on measures sensitive to dorsolateral prefrontal neocortex dysfunction. However, distinct neurocognitive profiles emerged when we examined the impact of comorbid schizotypal personality features on neuropsychological test performance. Primary OCD patients displayed impaired performance on measures sensitive to orbitofrontal dysfunction; however, they did not differ from control subjects on tests of dorsolateral function. OCD subjects presenting with schizotypal personality features performed poorly not only on tests sensitive to orbitofrontal dysfunction, but also on tests sensitive to dorsolateral dysfunction. Findings suggest that OCD can be subdivided into clinical subtypes, and distinct prefrontal subsystems may be differentially involved in these subtypes.


Abnormalities in GABA signaling in the forebrain at all stages - prenatal, postnatal and adult have been implicated with a wide range of neuropsychiatric illnesses ,2. AdditA receptors and GABA function with defective behaviors have been vital for understanding the pathobiology of neuropsychiatric illnesses, but all of these models were systemic or region-specific knockouts that could not serve to establish a cause - effect relationship between neuronal versus endothelial cells. Our observation of expression of GABAA receptor subunits and GAD65/67 in forebrain endothelial cells signified that forebrain angiogenesis has its own intrinsic GABA signaling mechanisms. To discover the functional significance of this endothelial GABA signaling pathway in vivo, we designed strategies to specifically turn off GABA release from endothelial cells or render endothelial GABAA receptors dysfunctional using cre-lox technology during early embryonic development. Both approaches first affected forebrain angiogenesis and in turn impaired neuronal migration. As endothelial GABA’s roles were delineated individually for angiogenesis, neurogenesis, long distance GABAergic neuronal migration and radial migration of projection neurons, so too was the realization that neuronal GABA could not serve to compensate these roles. The data laid the foundation for a novel positive feedback signaling pathway in endothelial cells that functions via GABAA receptor mediated GABA release [Though there have been reports of GAD immunoreactivities in endothelial cells of cerebral arteries  and GAD6A receptors, NKCC1-KCC2 expression, and chloride concentration in forebrain endothelial cells and alterations in disease scenarios. And most importantly, additional questions begin to emerge about the significance of this vascular GABA pathway and novel mechanisms specifically in the postnatal, adult and aging brain (As we now know that a common GABA pathway operates in both endothelial cells and GABAergic neurons of the embryonic telencephalon, it is essential to gain further mechanistic insights by segregating this pathway in individual cell types. It is important for future work to understand how the endothelial GABA signaling pathway specifically influences angiogenesis related genes and specific processes like tight junction formation, tip cell functions, vascular sprouting and migration. Also, it is important to see how components of the neuronal GABA pathway are affected in the absence of endothelial GABA signaling. For instance, how does endothelial GABA modulate interneuron migration - does it affect receptor-mediated signaling that transduces extracellular information or directly the transcription factors that provide interneurons with their intrinsic migratory ability or both? It also brings up several new questions with respect to roles of GABAng brain . New und


BRCA1 or BRCA2 is recognized for its strong familial risk .Genome Wide Association Studies (GWAS) serve as a powerful approach for discovering common risk variants underlying disease etiology. Often these variants reside within the non-coding region of the genome making the determination of functional mechanisms driving susceptibility a challenging task . Differecis-associations (cis-eQTL) were interrogated for association with breast cancer risk. Three SNPs, rs11099601, rs656040 and rs738200 were significantly associated  with an increased risk of breast cancer. The most significant association with increased risk for both ER-positive and ER-negative breast cancer was rs110099601  at 4q21, which constitutes a novel breast cancer susceptibility locus [In the study, Hamdi et al. evaluated 313 SNPs in 175 genes related to cancer for association with breast cancer risk in 46,451 breast cancer cancers and 42,599 controls of Caucasian ancestry participating in the Breast Cancer Association Consortium (BCAC) genotyped using the custom Illumina Infinium array iCOGS  with rs11099601 were functionally annotated using ENCODE chromatin features such as overlapping with Histone epigenetic marks associated with promoters and enhancers or DNAse I hypersensitive sites , which may be particularly effective for tumor types with limited sample sets.In summary, Hamdi et al. reported a novel association to breast cancer risk likely to be related to changes in expression of multiple genes. Importantly, although the approach used by Hamdi et al. may exclude the discovery of genes not previously implicated in cancer, it provides an example of an approach likely to identify variants undetectable under stringent GWAS multiple testing threshold (typically


Optical internal urethrotomy (OIU) is the most common procedure performed for short segment bulbar urethral stricture worldwide. This procedure most commonly performed using Sachse’s cold knife. Various perioperative complications of internal urethrotomy have been described in literature including bleeding, urinary tract infection, extravasation of fluid, incontinence, impotence, and recurrence of stricture. Here we report a unique complication of breakage of Sachse knife blade intraoperatively and its endoscopic management. Optical internal urethrotomy (OIU) is the most common procedure performed for short segment bulbar urethral stricture worldwide . HoweverA 30 year-old male presented with complaint of lower urinary tract symptoms for the last six months. Uroflowmetry voiding pattern was suggestive of urethral stricture disease. Retrograde urethrography (RGU) revealed a short segment bulbar urethral stricture (<1.5cms). Optical internal urethrotomy was performed. Intraoperatively blade of Sachse’s urethrotome accidently broken and fell proximal to the stricture which was confirmed on fluoroscopy . We compOptical internal urethrotomy became popularized after the work of Sachse in 1971  and now This complication should be kept in mind and instruments should be checked properly by the operative surgeon prior to start the procedure. Retained sharp objects like knife blade in urethra as a result of breakage of Sachse knife blade can be managed endoscopically.


Many real-world networks contain highly connected nodes called hubs. Hubs are often crucial for network function and spreading dynamics. However, classical models of how hubs originate during network development unrealistically assume that new nodes attain information about the connectivity (for example the degree) of existing nodes. Here, we introduce hub formation through nonlinear growth where the number of nodes generated at each stage increases over time and new nodes form connections independent of target node features. Our model reproduces variation in number of connections, hub occurrence time, and rich-club organization of networks ranging from protein–protein, neuronal and fibre tract brain networks to airline networks. Moreover, nonlinear growth gives a more generic representation of these networks compared with previous preferential attachment or duplication–divergence models. Overall, hub creation through nonlinear network expansion can serve as a benchmark model for studying the development of many real-world networks.