Transcriptome sequencing data provides a solid base to understand the phylogenetic relationships, biogeography and reticulated evolution of the genus Zamia L. (Cycadales: Zamiaceae).

BACKGROUND AND AIMS: Cycad is a key lineage to understand the early evolution of seed plants and their response to past environmental changes. However, tracing the evolutionary trajectory of cycad species is challenging when the robust relationships at inter- or infrageneric level are not well resolved. METHODS: Here, using 2,901 single-copy nuclear genes, we explored the species relationships and gene flow within the second largest genus of cycads, i.e., Zamia, based on phylotranscriptomic analyses of 90% extant Zamia species. Based on a well-resolved phylogenetic framework, we performed gene flow analyses, molecular dating, and biogeographical reconstruction to examine the spatiotemporal evolution of Zamia. We also performed ancestral state reconstruction (ASR) of a total of 62 traits of the genus to comprehensively investigate its morphological evolution. KEY RESULTS: Zamia is comprised of seven major clades corresponding to seven distinct distribution areas in the Americas, with at least three reticulation nodes revealed in this genus. Extant lineages of Zamia initially diversified around 18.4-32.6 (29.14) million years ago (MA) in the Mega-Mexico, and then expanded eastward into the Caribbean and southward into Central and South America. ASR revealed homoplasy in most of the morphological characters. CONCLUSIONS: This study revealed congruent phylogenetic relationships from comparative methods/datasets, with some conflicts being the result of incomplete lineage sorting and ancient/recent hybridization events. The strong association between the clades and the biogeographic areas suggested that ancient dispersal events shaped the modern distribution pattern, and regional climatic factors may have resulted in the following in-situ diversification. Climate cooling starting during the mid Miocene is associated with the global expansion of Zamia to the tropical South America that have dramatically driven lineage diversification in the New World flora, as well as the extinction of cycad species in the nowadays cooler regions of both hemispheres as indicated by the fossil records.

Eco-evolutionary evidence for the global diversity pattern of Cycas (Cycadaceae).

The evolution of the latitudinal diversity gradient (LDG), characterized by a peak in diversity toward the tropics, has captured significant attention in evolutionary biology and ecology. However, the inverse LDG (i-LDG) mechanism, wherein species richness increases toward the poles, remains inadequately explored. Cycads are among one of the oldest lineages of extant seed plants and have undergone extensive diversification in the tropics. Intriguingly, the extant cycad abundance exhibits an i-LDG pattern, and the underlying causes for this phenomenon remain largely elusive. Here, using 1,843 nuclear genes from a nearly complete sampling, we conducted comprehensive phylogenomic analyses to establish a robust species-level phylogeny for Cycas, the largest genus within cycads. We then reconstructed the spatial-temporal dynamics and integrated global environmental data to evaluate the roles of species ages, diversification rates, contemporary environment, and conservatism to ancestral niches in shaping the i-LDG pattern. We found Cycas experienced decreased diversification rates, coupled with the cooling temperature since its origin in the Eocene from continental Asia. Different regions have distinctively contributed to the formation of i-LDG for Cycas, with the northern hemisphere acting as evolutionary museums and the southern hemisphere serving as cradles. Moreover, water-related climate variables, specifically precipitation seasonality and potential evapotranspiration, were identified as paramount factors constraining Cycas species richness in the rainforest biome near the equator. Notably, the adherence to ancestral monsoonal climates emerges as a critical factor in sustaining the diversity pattern. This study underscores the imperative of integrating both evolutionary and ecological approaches to comprehensively unravel the mechanisms underpinning global biodiversity patterns.

Towards the plastome evolution and phylogeny of Cycas L. (Cycadaceae): molecular-morphology discordance and gene tree space analysis.

BACKGROUND: Plastid genomes (plastomes) present great potential in resolving multiscale phylogenetic relationship but few studies have focused on the influence of genetic characteristics of plastid genes, such as genetic variation and phylogenetic discordance, in resolving the phylogeny within a lineage. Here we examine plastome characteristics of Cycas L., the most diverse genus among extant cycads, and investigate the deep phylogenetic relationships within Cycas by sampling 47 plastomes representing all major clades from six sections. RESULTS: All Cycas plastomes shared consistent gene content and structure with only one gene loss detected in Philippine species C. wadei. Three novel plastome regions (psbA-matK, trnN-ndhF, chlL-trnN) were identified as containing the highest nucleotide variability. Molecular evolutionary analysis showed most of the plastid protein-coding genes have been under purifying selection except ndhB. Phylogenomic analyses that alternatively included concatenated and coalescent methods, both identified four clades but with conflicting topologies at shallow nodes. Specifically, we found three species-rich Cycas sections, namely Stangerioides, Indosinenses and Cycas, were not or only weakly supported as monophyly based on plastomic phylogeny. Tree space analyses based on different tree-inference methods both revealed three gene clusters, of which the cluster with moderate genetic properties showed the best congruence with the favored phylogeny. CONCLUSIONS: Our exploration in plastomic data for Cycas supports the idea that plastid protein-coding genes may exhibit discordance in phylogenetic signals. The incongruence between molecular phylogeny and morphological classification reported here may largely be attributed to the uniparental attribute of plastid, which cannot offer sufficient information to resolve the phylogeny. Contrasting to a previous consensus that genes with longer sequences and a higher proportion of variances are superior for phylogeny reconstruction, our result implies that the most effective phylogenetic signals could come from loci that own moderate variation, GC content, sequence length, and underwent modest selection.

Not that young: combining plastid phylogenomic, plate tectonic and fossil evidence indicates a Palaeogene diversification of Cycadaceae.

BACKGROUND AND AIMS: Previous molecular dating studies revealed historical mass extinctions and recent radiations of extant cycads, but debates still exist between palaeobotanists and evolutionary biologists regarding the origin and evolution of Cycadaceae. METHODS: Using whole plastomic data, we revisited the phylogeny of this family and found the Palawan endemic Cycas clade was strongly related to all lineages from Southeast Eurasia, coinciding with a plate drift event occurring in the Early Oligocene. By integrating fossil and biogeographical calibrations as well as molecular data from protein-coding genes, we established different calibration schemes and tested competing evolutionary timelines of Cycadaceae. KEY RESULTS: We found recent dispersal cannot explain the distribution of Palawan Cycas, yet the scenario including the tectonic calibration yielded a mean crown age of extant Cycadaceae of ~69-43 million years ago by different tree priors, consistent with multiple Palaeogene fossils assigned to this family. Biogeographical analyses incorporating fossil distributions revealed East Asia as the ancestral area of Cycadaceae. CONCLUSIONS: Our findings challenge the previously proposed Middle-Late Miocene diversification of cycads and an Indochina origin for Cycadaceae and highlight the importance of combining phylogenetic clades, tectonic events and fossils for rebuilding the evolutionary history of lineages that have undergone massive extinctions.

Phylotranscriptomics reveal the spatio-temporal distribution and morphological evolution of Macrozamia, an Australian endemic genus of Cycadales.

BACKGROUND AND AIMS: Cycads are regarded as an ancient lineage of living seed plants, and hold important clues to understand the early evolutionary trends of seed plants. The molecular phylogeny and spatio-temporal diversification of one of the species-rich genera of cycads, Macrozamia, have not been well reconstructed. METHODS: We analysed a transcriptome dataset of 4740 single-copy nuclear genes (SCGs) of 39 Macrozamia species and two outgroup taxa. Based on concatenated (maximum parsimony, maximum likelihood) and multispecies coalescent analyses, we first establish a well-resolved phylogenetic tree of Macrozamia. To identify cyto-nuclear incongruence, the plastid protein coding genes (PCGs) from transcriptome data are extracted using the software HybPiper. Furthermore, we explore the biogeographical history of the genus and shed light on the pattern of floristic exchange between three distinct areas of Australia. Six key diagnostic characters are traced on the phylogenetic framework using two comparative methods, and infra-generic classification is investigated. KEY RESULTS: The tree topologies of concatenated and multi-species coalescent analyses of SCGs are mostly congruent with a few conflicting nodes, while those from plastid PCGs show poorly supported relationships. The genus contains three major clades that correspond to their distinct distributional areas in Australia. The crown group of Macrozamia is estimated to around 11.80 Ma, with a major expansion in the last 5-6 Myr. Six morphological characters show homoplasy, and the traditional phenetic sectional division of the genus is inconsistent with this current phylogeny. CONCLUSIONS: This first detailed phylogenetic investigation of Macrozamia demonstrates promising prospects of SCGs in resolving phylogenetic relationships within cycads. Our study suggests that Macrozamia, once widely distributed in Australia, underwent major extinctions because of fluctuating climatic conditions such as cooling and mesic biome disappearance in the past. The current close placement of morphologically distinct species in the phylogenetic tree may be related to neotenic events that occurred in the genus.

Congruence between ocean-dispersal modelling and phylogeography explains recent evolutionary history of Cycas species with buoyant seeds.

Ocean currents play a significant role in driving the long-distance dispersal (LDD), spatial distribution and phylogeographic patterns of many organisms. Integrating phylogeographic analyses and mechanistic ocean current modelling can provide novel insights into the evolutionary history of terrestrial littoral species but has been rarely applied in this context. We focused on a group of Cycas that have buoyant seeds and occupy coastal habitats. By integrating evidence from mechanistic simulations and whole plastomic data, we examined the role of ocean circulation in shaping the phylogeography of these Cycas species. Plastomes of the studied Cycas species showed extreme conservatism, following a post-Pleistocene divergence. Phylogenies revealed three subclades, corresponding to the Pacific Ocean, Sunda Shelf and Indian Ocean. The ocean modelling results indicate that hotspots of seed stranding coincide well with the contemporary distribution of the Cycas species and that drifting trajectories from the three subclades are largely confined to separate regions. These findings suggest that ocean current systems, by driving long-distance dispersal, have shaped the distribution and phylogeography for Cycas with buoyant seeds. This study highlights how the combination of genomic data and ocean drift modelling can help explain phylogeographic patterns and diversity in terrestrial littoral ecosystems.

Large outbreak of tularaemia, central Sweden, July to September 2019.

On 31 of July 2019, the Public Health Agency of Sweden was alerted about an increasing number of tularaemia cases in Gävleborg, a county in central Sweden. The number of cases increased thereafter peaking at about 150 reports of illnesses every week. As at 6 October, a total of 979 cases (734 laboratory-confirmed) have been reported, mainly from counties in central Sweden. The outbreak is now considered over (as at 14 October).

Phylogeny of the gymnosperm genus Cycas L. (Cycadaceae) as inferred from plastid and nuclear loci based on a large-scale sampling: Evolutionary relationships and taxonomical implications.

The gymnosperm genus Cycas is the sole member of Cycadaceae, and is the largest genus of extant cycads. There are about 115 accepted Cycas species mainly distributed in the paleotropics. Based on morphology, the genus has been divided into six sections and eight subsections, but this taxonomy has not yet been tested in a molecular phylogenetic framework. Although the monophyly of Cycas is broadly accepted, the intrageneric relationships inferred from previous molecular phylogenetic analyses are unclear due to insufficient sampling or uninformative DNA sequence data. In this study, we reconstructed a phylogeny of Cycas using four chloroplast intergenic spacers and seven low-copy nuclear genes and sampling 90% of extant Cycas species. The maximum likelihood and Bayesian inference phylogenies suggest: (1) matrices of either concatenated cpDNA markers or of concatenated nDNA lack sufficient informative sites to resolve the phylogeny alone, however, the phylogeny from the combined cpDNA-nDNA dataset suggests the genus can be roughly divided into 13 clades and six sections that are in agreement with the current classification of the genus; (2) although with partial support, a clade combining sections Panzhihuaenses + Asiorientales is resolved as the earliest diverging branch; (3) section Stangerioides is not monophyletic because the species resolve as a grade; (4) section Indosinenses is not monophyletic as it includes Cycas macrocarpa and C. pranburiensis from section Cycas; (5) section Cycas is the most derived group and its subgroups correspond with geography.

Macrocheles species (Acari: Macrochelidae) associated with human corpses in Europe.

The biology of macrochelid mites might offer new venues for the interpretation of the environmental conditions surrounding human death and decomposition. Three human corpses, one from Sweden and two from Spain, have been analysed for the occurrence of Macrochelidae species. Macrocheles muscaedomesticae (Scopoli) females were associated with a corpse that was found in a popular beach area of southeast Spain. Their arrival coincides with the occurrence of one of their major carrier species, the filth fly Fannia scalaris, the activity of which peaks during mid-summer. Macrocheles glaber (Müller) specimens were collected from a corpse in a shallow grave in a forest in Sweden at the end of summer, concurrent with the arrival of beetles attracted by odours from the corpse. Macrocheles perglaber Filipponi and Pegazzano adults were sampled from a corpse found indoors in the rural surroundings of Granada city, south Spain. The phoretic behaviour of this species is similar to that of M. glaber, but it is more specific to Scarabaeidae and Geotrupidae dung beetles, most of which favour human faeces. Macrocheles muscaedomesticae is known from urban and rural areas and poultry farms, M. glaber from outdoors, particularly the countryside, whereas M. perglaber is known from outdoor, rural, and remote, potentially mountainous locations. Macrocheles muscaedomesticae and M. perglaber are reported for the first time from the Iberian Peninsula. This is the first record of M. perglaber from human remains.

From Knowing Nothing to Knowing What, How and Now: Parents' Experiences of Caring for their Children With Congenital Adrenal Hyperplasia.

Objective: This study investigates various kinds of knowing that European parents use when caring for their children with congenital adrenal hyperplasia (CAH). Methods: Semi-structured qualitative interviews with 20 parents of 22 children with CAH. Results: Parents emphasized the importance of knowing what CAH is and what support their child needs, but also knowing how to cope and make sense of the new situation, how to attend to their child's medical needs as well as how to talk to their child. Parents also reported challenges related to connecting with their social network, experiences of emergency care, and how to help their children become independent. These challenges require knowing now : being able to respond appropriately to unique circumstances. Conclusions: Parents experience diverse challenges that may moderate the effects of the diagnosis on children's well-being. Parenting children with CAH requires the development of knowing that goes beyond medical information.

First report of spinose ear tick, Otobius megnini (Acari, Argasidae), in Sweden.

A dog that had travelled to Sweden was inspected by a veterinarian. In one ear canal a tick was found. It was later identified as a spinose ear tick, Otobius megnini. In this case report we also review the previously known reports of O. megnini in Europe and the recent introduction and spread in Turkey.

Identification of European mosquito species by MALDI-TOF MS.

MALDI-TOF MS profiling has proved to be efficient for arthropod identification at the species level. However, prior to entomological monitoring, the reference spectra database should cover relevant species. Here, 74 specimens were field-collected from 11 mosquito species captured in two distinct European areas and used either to increment our database or for blind tests. Misidentification was not noted, underlining the power of this approach. Nevertheless, three out of the 26 specimens used for the blind test did not reach the significant identification threshold value set, attributed to lower spectral quality. In the future, the quality control spectra parameters need to be defined to avoid not achieving significant threshold identification.

UV-B exposure of indoor-grown Picea abies seedlings causes an epigenetic effect and selective emission of terpenes.

Terpenoids are involved in various defensive functions in plants, especially conifers. Epigenetic mechanisms, for example DNA methylation, can influence plant defence systems. The purpose of the present study was to investigate the influence of UV-B exposure on the release of terpenoids from spruce seedlings and on needle DNA methylation. Ten-week-old seedlings grown indoors were exposed to UV-B radiation during 4 h, and the volatile compounds emitted from the seedlings were analysed. Analysis of the volatiles 1, 3, and 22 d after this UV-B exposure showed that bornyl acetate, borneol, myrcene, and limonene contents increased during the first 3 days, while at day 22 the level of emission had returned to the control level. UV-B exposure decreased the level of DNA methylation in needles of young seedlings, reflected in methylation changes in CCGG sequences. Exposure of young seedlings to UV-B radiation might be a way to potentiate the general defensive capacity, improving their ability to survive in outdoor conditions. UV-B-induced defence is discussed in the light of epigenetic mechanisms.

Phylotranscriptomics Shed Light on Intrageneric Relationships and Historical Biogeography of Ceratozamia (Cycadales).

Ceratozamia Brongn. is one of the species-rich genera of Cycadales comprising 38 species that are mainly distributed in Mexico, with a few species reported from neighboring regions. Phylogenetic relationships within the genus need detailed investigation based on extensive datasets and reliable systematic approaches. Therefore, we used 30 of the known 38 species to reconstruct the phylogeny based on transcriptome data of 3954 single-copy nuclear genes (SCGs) via coalescent and concatenated approaches and three comparative datasets (nt/nt12/aa). Based on all these methods, Ceratozamia is divided into six phylogenetic subclades within three major clades. There were a few discrepancies regarding phylogenetic position of some species within these subclades. Using these phylogenetic trees, biogeographic history and morphological diversity of the genus are explored. Ceratozamia originated from ancestors in southern Mexico since the mid-Miocene. There is a distinct distribution pattern of species through the Trans-Mexican Volcanic Belt (TMVB), that act as a barrier for the species dispersal at TMVB and its southern and northern part. Limited dispersal events occurred during the late Miocene, and maximum diversification happened during the Pliocene epoch. Our study provides a new insight into phylogenetic relationships, the origin and dispersal routes, and morphological diversity of the genus Ceratozamia. We also explain how past climatic changes affected the diversification of this Mesoamerica-native genus.

Curcumamaxwellii and C.rubroaurantiaca (Zingiberaceae, Zingiberoideae), two new species from Thailand.

Curcumamaxwelliisp. nov. and Curcumarubroaurantiacasp. nov. (Zingiberaceae, Zingiberoideae, Zingibereae), two new red-orange-flowered species from Thailand, are described. They are compared to the morphologically closest species from the Curcumasubgen.Ecomatae and detailed descriptions, colour plates and information on their distribution, ecology, phenology and uses are provided. Preliminary IUCN conservation assessments for both of these species are proposed as Least Concern.

The Original Scientific Description of the Lone Star Tick (Amblyomma americanum, Acari: Ixodidae) and Implications for the Species' Past and Future Geographic Distributions.

Amblyomma americanum L. is an important vector in North America originally described by Linnaeus based on Pehr Kalm's 1754 report. While Kalm's 'Travels into North America' is well known, his 1754 report remains obscure. Some authors were skeptical that Kalm referred to A. americanum because he encountered them at sites farther north outside of the species' range. However, the details in 1754 report leave no doubt that Kalm described lone star ticks. In this historical review, we provide support for Kalm's identification using a modern translation of his 1754 report and other sources. We also delineate distributional changes of lone star ticks from the pre-colonization era to the present and interpret them in the context of large-scale anthropogenic changes in the landscape. In this framework, the lone star tick's current northward expansion is a recolonization of their former range. Extensive deforestation and extirpation of their principal host species, white-tailed deer, led to A. americanum's disappearance from the northern parts of its range by the 20th century. Subsequent recolonization by second-growth forest and increases in white-tailed deer populations by the mid-20th century is now allowing A. americanum to reclaim its former range. These changes in the land appear to be the driving force behind A. americanum's present expansion. Understanding this species' history and the factors contributing to its current expansion will enable better predictions about its future distribution and potential to transmit human pathogens.

The Cycas genome and the early evolution of seed plants.

Cycads represent one of the most ancient lineages of living seed plants. Identifying genomic features uniquely shared by cycads and other extant seed plants, but not non-seed-producing plants, may shed light on the origin of key innovations, as well as the early diversification of seed plants. Here, we report the 10.5-Gb reference genome of Cycas panzhihuaensis, complemented by the transcriptomes of 339 cycad species. Nuclear and plastid phylogenomic analyses strongly suggest that cycads and Ginkgo form a clade sister to all other living gymnosperms, in contrast to mitochondrial data, which place cycads alone in this position. We found evidence for an ancient whole-genome duplication in the common ancestor of extant gymnosperms. The Cycas genome contains four homologues of the fitD gene family that were likely acquired via horizontal gene transfer from fungi, and these genes confer herbivore resistance in cycads. The male-specific region of the Y chromosome of C. panzhihuaensis contains a MADS-box transcription factor expressed exclusively in male cones that is similar to a system reported in Ginkgo, suggesting that a sex determination mechanism controlled by MADS-box genes may have originated in the common ancestor of cycads and Ginkgo. The C. panzhihuaensis genome provides an important new resource of broad utility for biologists.

Different Hatching Rates of Floodwater Mosquitoes Aedes sticticus, Aedes rossicus and Aedes cinereus from Different Flooded Environments.

In the lower Dalälven region, floodwater mosquitoes cause recurring problems. The main nuisance species is Aedes (Ochlerotatus) sticticus, but large numbers of Aedes (Aedes) rossicus and Aedes (Aedes) cinereus also hatch during flooding events. To increase understanding of which environments in the area give rise to mosquito nuisance, soil samples were taken from 20 locations from four environmental categories: grazed meadows, mowed meadows, unkept open grassland areas and forest areas. In each location 20 soil samples were taken, 10 from random locations and 10 from moisture retaining structures, such as tussocks, shrubs, piles of leaves, logs, and roots. The soil samples were soaked with tap water in the lab, and mosquito larvae were collected and allowed to develop to adult mosquitoes for species identification. Fewer larvae hatched from mowed areas and more larvae hatched from moisture retaining structure samples than random samples. The results showed that Aedes cinereus mostly hatch from grazed and unkept areas and hatched as much from random samples as from structures, whereas Aedes sticticus and Aedes rossicus hatched from open unkept and forest areas and hatch significantly more from structure samples. When the moisture retaining structures in open unkept areas where Aedes sticticus hatched were identified it was clear that they hatched predominantly from willow shrubs that offered shade. The results suggest that Ae. sticticus and Ae. cinereus favor different flooded environments for oviposition.

Diel patterns of stem CO2 efflux vary among cycads, arborescent monocots, and woody eudicots and gymnosperms.

The diel patterns of stem carbon dioxide efflux (Es) were determined for cycads, monocots, and woody eudicot and gymnosperm tree species. Stem Es at a height of 30-40 cm was measured every 2 h throughout 31-h campaigns. Our range of Es was 1.5-4.0 µmol·m-2·s-1 for cycads, 1.0-3.5 µmol·m-2·s-1 for arborescent monocots, and 1.5-4.5 µmol·m-2·s-1 for woody eudicot and gymnosperm trees species. Time of day did not influence Es of cycads or monocots. In contrast, the woody stems of eudicots and gymnosperms exhibited diurnal Es that was 36% to 40% greater than nocturnal Es. The established literature based on Es of woody tree species cannot be used to estimate habitat carbon cycles in habitats which contain cycad or monocot trees. Time of day must be included for accuracy of research on Es of woody tree species. Failures to account for the spatiotemporal differences of Es may explain some of the disparity in outcomes of published stem respiration studies.

The Human Biting Culex pipiens Bioform molestus Detected in Several Areas in Southern Sweden.

Background: The mosquito species Culex pipiens is a known vector of several pathogens and occurs in two distinct bioforms, pipiens and molestus. The bioform molestus thrives in urban environments where there are below-ground habitats; it can mate in confined spaces and feed on mammals as well as birds. In contrast, the bioform pipiens is found above ground, is thought to require more space for mating, and mainly feeds on birds. The pipiens bioform is present in large parts of Sweden but the molestus bioform has previously only been found in major cities. Materials and Methods: People experiencing mosquito nuisance in southern Sweden submitted mosquito samples as part of a citizen science project, and these samples were analyzed to determine the geographical distribution of the molestus bioform of Cx. pipiens. Mosquito specimens were identified to the species level by DNA barcoding of the cytochrome C oxidase subunit I (COI) gene, and the bioforms were determined through the CQ11 microsatellite marker. Results:Culex pipiens f molestus was observed to be spread across large parts of Gothenburg as well as in the suburbs. This bioform was found both in urban and rural areas at several sites across southern Sweden. In one site, hybrids between the two bioforms were found. Conclusions: The detection of Cx. pipiens f molestus in several rural areas was surprising, indicating that it may be more widely spread than urban areas alone, where it has been previously reported.