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Conventional means of data extraction using optical fiber interrogators are not adequate for fast-paced detection of a target parameter. In this instance, the relationship between the critical meniscus heights (CMH) of several liquids to the extraction speed of a rod submerged in them, have been analyzed. A limitation of a previous interrogator used for the purpose had been light absorption by the liquid due to the used bandwidth of the readily-available light source, i.e., C-band. The newly proposed technique addresses this limitation by utilizing a broadband light source instead, with a Si-photodetector and an Arduino. In addition, the Arduino is capable of extracting data at a relatively faster rate with respect to the conventional optical interrogator. The use of a different operational wavelength (850 nm instead of 1550 nm) increased the r2 and the sensitivity of the sensor. The new setup can measure surface chemistry properties, with the advantage of being comparatively cheaper than the conventionally available interrogator units, thereby providing a suitable alternative to conventional measurement techniques of liquid surface properties, while reducing material waste, i.e., in terms of the required volume for detection of a target parameter, through the use of optical fiber.
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Proxy measures of genome-wide heterozygosity based on approximately 10 microsatellites have been used to uncover heterozygosity fitness correlations (HFCs) for a wealth of important fitness traits in natural populations. However, effect sizes are typically very small and the underlying mechanisms remain contentious, as a handful of markers usually provides little power to detect inbreeding. We therefore used restriction site associated DNA (RAD) sequencing to accurately estimate genome-wide heterozygosity, an approach transferrable to any organism. As a proof of concept, we first RAD sequenced oldfield mice (Peromyscus polionotus) from a known pedigree, finding strong concordance between the inbreeding coefficient and heterozygosity measured at 13,198 single-nucleotide polymorphisms (SNPs). When applied to a natural population of harbor seals (Phoca vitulina), a weak HFC for parasite infection based on 27 microsatellites strengthened considerably with 14,585 SNPs, the deviance explained by heterozygosity increasing almost fivefold to a remarkable 49%. These findings arguably provide the strongest evidence to date of an HFC being due to inbreeding depression in a natural population lacking a pedigree. They also suggest that under some circumstances heterozygosity may explain far more variation in fitness than previously envisaged.
Assuntos
Aptidão Genética/genética , Variação Genética , Heterozigoto , Endogamia , Peromyscus/genética , Phoca/genética , Animais , Genética Populacional , Sequenciamento de Nucleotídeos em Larga Escala , Illinois , Mar do Norte , Phoca/parasitologia , Polimorfismo de Nucleotídeo Único/genética , Mapeamento por RestriçãoRESUMO
BACKGROUND: Selective plane illumination microscopy (SPIM a type of light-sheet microscopy) involves focusing a thin sheet of laser light through a specimen at right angles to the objective lens. As only the thin section of the specimen at the focal plane of the lens is illuminated, out of focus light is naturally absent and toxicity due to light (phototoxicity) is greatly reduced enabling longer term live imaging. OpenSPIM is an open access platform (Pitrone et al. 2013 and OpenSPIM.org) created to give new users step-by-step instructions on building a basic configuration of a SPIM microscope, which can in principle be adapted and upgraded to each laboratory's own requirements and budget. Here we describe our own experience with the process of designing, building, configuring and using an OpenSPIM for our research into the early development of the polyclad flatworm Maritigrella crozieri - a non-model animal. RESULTS: Our OpenSPIM builds on the standard design with the addition of two colour laser illumination for simultaneous detection of two probes/molecules and dual sided illumination, which provides more even signal intensity across a specimen. Our OpenSPIM provides high resolution 3d images and time lapse recordings, and we demonstrate the use of two colour lasers and the benefits of two color dual-sided imaging. We used our microscope to study the development of the embryo of the polyclad flatworm M. crozieri. The capabilities of our microscope are demonstrated by our ability to record the stereotypical spiral cleavage pattern of M. crozieri with high-speed multi-view time lapse imaging. 3D and 4D (3D + time) reconstruction of early development from these data is possible using image registration and deconvolution tools provided as part of the open source Fiji platform. We discuss our findings on the pros and cons of a self built microscope. CONCLUSIONS: We conclude that home-built microscopes, such as an OpenSPIM, together with the available open source software, such as MicroManager and Fiji, make SPIM accessible to anyone interested in having continuous access to their own light-sheet microscope. However, building an OpenSPIM is not without challenges and an open access microscope is a worthwhile, if significant, investment of time and money. Multi-view 4D microscopy is more challenging than we had expected. We hope that our experience gained during this project will help future OpenSPIM users with similar ambitions.
Assuntos
Microscopia de Fluorescência/instrumentação , Platelmintos/crescimento & desenvolvimento , Animais , Processamento de Imagem Assistida por Computador , Luz , Software , Interface Usuário-ComputadorRESUMO
We estimated the spontaneous mutation rate in Heliconius melpomene by genome sequencing of a pair of parents and 30 of their offspring, based on the ratio of number of de novo heterozygotes to the number of callable site-individuals. We detected nine new mutations, each one affecting a single site in a single offspring. This yields an estimated mutation rate of 2.9 × 10(-9) (95% confidence interval, 1.3 × 10(-9)-5.5 × 10(-9)), which is similar to recent estimates in Drosophila melanogaster, the only other insect species in which the mutation rate has been directly estimated. We infer that recent effective population size of H. melpomene is about 2 million, a substantially lower value than its census size, suggesting a role for natural selection reducing diversity. We estimate that H. melpomene diverged from its Müllerian comimic H. erato about 6 Ma, a somewhat later date than estimates based on a local molecular clock.
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Borboletas/crescimento & desenvolvimento , Genoma de Inseto , Taxa de Mutação , Análise de Sequência de DNA/métodos , Animais , Borboletas/classificação , Borboletas/genética , Drosophila melanogaster/genética , Evolução Molecular , Feminino , Masculino , Filogenia , Densidade Demográfica , Seleção GenéticaRESUMO
Most speciation events probably occur gradually, without complete and immediate reproductive isolation, but the full extent of gene flow between diverging species has rarely been characterized on a genome-wide scale. Documenting the extent and timing of admixture between diverging species can clarify the role of geographic isolation in speciation. Here we use new methodology to quantify admixture at different stages of divergence in Heliconius butterflies, based on whole-genome sequences of 31 individuals. Comparisons between sympatric and allopatric populations of H. melpomene, H. cydno, and H. timareta revealed a genome-wide trend of increased shared variation in sympatry, indicative of pervasive interspecific gene flow. Up to 40% of 100-kb genomic windows clustered by geography rather than by species, demonstrating that a very substantial fraction of the genome has been shared between sympatric species. Analyses of genetic variation shared over different time intervals suggested that admixture between these species has continued since early in speciation. Alleles shared between species during recent time intervals displayed higher levels of linkage disequilibrium than those shared over longer time intervals, suggesting that this admixture took place at multiple points during divergence and is probably ongoing. The signal of admixture was significantly reduced around loci controlling divergent wing patterns, as well as throughout the Z chromosome, consistent with strong selection for Müllerian mimicry and with known Z-linked hybrid incompatibility. Overall these results show that species divergence can occur in the face of persistent and genome-wide admixture over long periods of time.
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Borboletas/genética , Fluxo Gênico , Genes de Insetos , Especiação Genética , Simpatria , Alelos , Animais , Borboletas/classificação , Evolução Molecular , Variação Genética , Genoma , Genoma de Inseto , Desequilíbrio de Ligação , Fenótipo , Filogenia , Cromossomos Sexuais/genéticaRESUMO
Atraumatic splenic rupture is a complex surgical pathology owing to its rarity, non-specificity of symptoms and gravity of possible outcomes. This case outlines the investigation and management of a patient with atraumatic splenic rupture secondary to undiagnosed hairy cell leukaemia. While the patient was initially managed conservatively, they went on to have a splenectomy owing to ongoing transfusion requirements. A review of the literature has also been performed and presented to highlight the potential causes of atraumatic splenic rupture and the various options for confirming diagnosis and definitive management.
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Haemobilia, or bleeding within the biliary tree, is rare. It can cause biliary obstruction secondary to blood clots. A comorbid 87-year-old was admitted to hospital with acute cholecystitis, choledocholithiasis, and an Escherichia coli bacteremia. He had a partial pancreatectomy and gastrojejunostomy 35 years prior for severe pancreatitis. He was treated with antibiotics and a percutaneous cholecystostomy. He developed atrial fibrillation and was subsequently commenced on warfarin. He re-presented 5 days after discharge with abdominal pain and fevers. Liver function tests revealed cholestasis and a supratherapeutic international normalised ratio. Imaging showed cholecystitis, biliary obstruction, and extensive biliary blood clots. He improved with antibiotics, vitamin K, and alteplase flushes through the percutaneous cholecystostomy. Repeat cholangiogram demonstrated dissolution of the biliary clots. Due to altered anatomy and comorbidities, alteplase flushes were utilized to relieve this patient's biliary obstruction. Thrombolytics may assist in treating biliary clots when first-line options are not possible or favourable.
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The evolutionary origins of Bilateria remain enigmatic. One of the more enduring proposals highlights similarities between a cnidarian-like planula larva and simple acoel-like flatworms. This idea is based in part on the view of the Xenacoelomorpha as an outgroup to all other bilaterians which are themselves designated the Nephrozoa (protostomes and deuterostomes). Genome data can provide important comparative data and help to understand the evolution and biology of enigmatic species better. Here we assemble and analyse the genome of the simple, marine xenacoelomorph Xenoturbella bocki, a key species for our understanding of early bilaterian evolution. Our highly contiguous genome assembly of X. bocki has a size of ~111 Mbp in 18 chromosome like scaffolds, with repeat content and intron, exon and intergenic space comparable to other bilaterian invertebrates. We find X. bocki to have a similar number of genes to other bilaterians and to have retained ancestral metazoan synteny. Key bilaterian signalling pathways are also largely complete and most bilaterian miRNAs are present. Overall, we conclude that X. bocki has a complex genome typical of bilaterians, which does not reflect the apparent simplicity of its body plan that has been so important to proposals that the Xenacoelomorpha are the simple sister group of the rest of the Bilateria.
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This report describes a patient with destructive multivalvular endocarditis with multiorgan dysfunction. Intraoperatively, severe unexpected adhesions due to pancarditis precluded the standard Commando operation. The approach of combining autotransplantation with a Commando operation was used to facilitate adequate débridement of infected tissues and reconstruction in a reasonable time and with satisfactory prosthetic valvular function. This approach may be considered in other cases of challenging destructive endocarditis with severe adhesions. We speculate that this approach may also be useful in redo surgeries.
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Endocardite Bacteriana , Endocardite , Doenças das Valvas Cardíacas , Humanos , Transplante Autólogo , Endocardite Bacteriana/complicações , Endocardite Bacteriana/cirurgia , Endocardite/complicações , Endocardite/cirurgia , Valva Aórtica/cirurgia , Doenças das Valvas Cardíacas/cirurgiaRESUMO
Iatrogenic diaphragmatic herniation is rare. This case is an example of herniation of the liver into the pericardial space post-transdiaphragmatic pericardial window formation for recurrent pericarditis. This case highlights that transdiaphragmatic herniation of intra-abdominal organs should be considered in patients presenting with gastrointestinal or cardiorespiratory symptoms with history of iatrogenic diaphragmatic defect.
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Pelagic larval stages are widespread across animals, yet it is unclear whether larvae were present in the last common ancestor of animals or whether they evolved multiple times due to common selective pressures. Many marine larvae are at least superficially similar; they are small, swim through the beating of bands of cilia, and sense the environment with an apical organ. To understand these similarities, we have generated single-cell atlases for marine larvae from two animal phyla and have compared their cell types. We found clear similarities among ciliary band cells and between neurons of the apical organ in the two larvae pointing to possible homology of these structures, suggesting a single origin of larvae within Spiralia. We also find several clade-specific innovations in each larva, including distinct myocytes and shell gland cells in the oyster larva. Oyster shell gland cells express many recently evolved genes that have made previous gene age estimates for the origin of trochophore larvae too young.
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Evolução Biológica , Neurônios , Animais , Larva/fisiologiaRESUMO
PURPOSE: Peri-operative blood transfusion has been identified as a risk factor for anastomotic leak in recent studies, but little is known about which patients are at risk for blood transfusion. This study aims to assess the relationship between blood transfusion and anastomotic leak and factors predisposing to leak in patients undergoing colorectal cancer surgery. METHODS: This retrospective cohort study was conducted in a tertiary hospital in Brisbane, Australia, between 2010 and 2019. A total of 522 patients underwent resection of colorectal cancer with primary anastomosis with no covering stoma and the prevalence of anastomotic leak was compared between those who had had perioperative blood transfusion(s) and those who had not. RESULTS: A total of 19 of 522 patients undergoing surgery for colorectal cancer had developed an anastomotic leak (3.64%). 11.3% of patients who had had a perioperative blood transfusion developed an anastomotic leak whereas 2.2% of patients who had not had a blood transfusion developed an anastomotic leak (p = 0.0002). Patients undergoing procedure on their right colon had proportionally more blood transfusions and this approached statistical significance (p = 0.06). Patients who received a greater quantity of units of blood transfusion prior to their diagnosis of anastomotic leak were more likely to develop an anastomotic leak (p = 0.001). CONCLUSION: Perioperative blood transfusions are associated with a significantly increased risk of an anastomotic leak following bowel resection with primary anastomosis for colorectal cancer.
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The link between abdominal surgery and venous thromboembolism (VTE) has been well established with recent evidence exploring the optimal VTE risk reducing strategy. However, despite these strategies pulmonary embolisms (PEs) do occur, which in the immediate post-operative setting creates a dilemma; to treat the VTE with anticoagulation but balance against the risk of hemorrhage. Treatment guidelines often do not include post-operative patients leaving the decision up to the treating physician to weigh the relative risks on an individual basis. We present a 59-year-old lady who developed a life-threatening submassive PE within 9 h of an emergency laparotomy for a perforated rectal cancer. She was treated with systemic thrombolysis after alternative interventions had been excluded. She responded well to therapy with no major bleeding. She was successfully discharged home after a short period of inpatient rehabilitation.
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Synchronous malignancies of the bile duct and the gallbladder are rare. These cases are often associated with pancreaticobiliary maljunction which is characterized by a long common shared pancreatobiliary channel leading to the Sphincter of Oddi. This predisposes the biliary epithelium to pancreatic enzyme reflux and makes the development of neoplasia more likely. We describe the case of a 64-year-old Caucasian female who presented with new jaundice and severe cholecystitis secondary to an impacted gallstone which was seen on ultrasound. Magnetic resonance cholangiopancreatography was organized with suspicion of a possible Mirizzi syndrome. This revealed a mid-distal bile duct cancer in addition to cholecystitis from an impacted gallstone. She was treated with intravenous antibiotics for her cholecystitis and underwent an urgent endoscopic retrograde cholangiopancreatography procedure for biliary decompression and stenting for her obstructive jaundice. The patient proceeded to pancreaticoduodenectomy with final histopathology revealing a synchronous primary gallbladder malignancy in addition to the known bile duct cancer.
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(1) Background: The intra-tumoural heterogeneity (ITH) of hepatocellular carcinoma (HCC) and its microenvironment (TME) across primary and secondary disease is poorly characterised. (2) Methods: Intra-tumoural (IT) and peri-tumoural (PT) staining of matched primary and secondary samples was conducted to evaluate the distribution of CD4+/FOXP3+ and CD8+/PD1+ T-cells. Samples underwent PD-L1/2 immunostaining, tumour mutational burden (TMB) evaluation, and high-resolution T-cell receptor (TCR) sequencing to derive T-cell clonality and targeted transcriptomics. (3) Results: We analysed 24 samples from matched primary (n = 11) and secondary (n = 13; 5 synchronous, 6 metachronous) deposits, 11 being extrahepatic (84.6%). IT CD8+ density was lower than PT in both primary (p = 0.005) and secondary deposits (p = 0.01), consistent with immune exclusion. PD-L1+ tumours displayed higher IT and PT CD8+/PD1+ cell density compared to PD-L1- (p < 0.05), and primary IT infiltrate was enriched in CD4+/FOXP3+ cells, compared to PT regions (p = 0.004). TCR-sequencing demonstrated enrichment of the top T-cell clonotype in secondary versus primary HCC (p = 0.02), without differences in overall productive clonality (p = 0.35). TMB was similar across primary versus secondary HCC (p = 0.95). While directed gene set analysis demonstrated the uniformity of transcriptional signatures of individual immune cell types, secondary deposits demonstrated higher COLEC12 (p = 0.004), CCL26 (p = 0.02), CD1E (p = 0.02) and CD36 (p = 0.03) expression with downregulation of CXCL1 (p = 0.03), suggesting differential regulation of innate immunity. (4) Conclusion: Immune exclusion is a defining feature of the HCC TME. Despite evidence of homogeneity in somatic TMB, secondary HCC is characterised by the expansion of a distinct T-cell clonotype and differential regulation of innate immune pathways.
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Attempts by biogeographers to understand biotic diversification in the Amazon have often employed contemporary species distribution patterns to support particular theories, such as Pleistocene rainforest refugia, rather than to test among alternative hypotheses. Suture zones, narrow regions where multiple contact zones and hybrid zones between taxa cluster, have been seen as evidence for past expansion of whole biotas that have undergone allopatric divergence in vicariant refuges. We used coalescent analysis of mutilocus sequence data to examine population split times in 22 pairs of geminate taxa in ithomiine and heliconiine butterflies. We test a hypothesis of simultaneous divergence across a suture zone in NE Peru. Our results reveal a scattered time course of diversification in this suture zone, rather than a tight cluster of split times. Additionally, we find rapid diversification within some lineages such as Melinaea contrasting with older divergence within lineages such as the Oleriina (Hyposcada and Oleria). These results strongly reject simple vicariance as a cause of the suture zone. At the same time, observed lineage effects are incompatible with a series of geographically coincident vicariant events which should affect all lineages similarly. Our results suggest that Pleistocene climatic forcing cannot readily explain this Peruvian suture zone. Lineage-specific biological traits, such as characteristic distances of gene flow or varying rates of parapatric divergence, may be of greater importance.
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Borboletas/genética , Especiação Genética , Genética Populacional , Animais , Teorema de Bayes , Núcleo Celular/genética , DNA Mitocondrial/genética , Geografia , Funções Verossimilhança , Modelos Genéticos , Dados de Sequência Molecular , Tipagem de Sequências Multilocus , Peru , Filogeografia , Densidade Demográfica , Análise de Sequência de DNARESUMO
Animals detect light using opsin photopigments. Xenopsin, a recently classified subtype of opsin, challenges our views on opsin and photoreceptor evolution. Originally thought to belong to the Gαi-coupled ciliary opsins, xenopsins are now understood to have diverged from ciliary opsins in pre-bilaterian times, but little is known about the cells that deploy these proteins, or if they form a photopigment and drive phototransduction. We characterized xenopsin in a flatworm, Maritigrella crozieri, and found it expressed in ciliary cells of eyes in the larva, and in extraocular cells around the brain in the adult. These extraocular cells house hundreds of cilia in an intra-cellular vacuole (phaosome). Functional assays in human cells show Maritigrella xenopsin drives phototransduction primarily by coupling to Gαi. These findings highlight similarities between xenopsin and c-opsin and reveal a novel type of opsin-expressing cell that, like jawed vertebrate rods, encloses the ciliary membrane within their own plasma membrane.
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Peptídeos/metabolismo , Células Fotorreceptoras de Invertebrados/fisiologia , Platelmintos/fisiologia , Células Fotorreceptoras Retinianas Bastonetes/metabolismo , Proteínas de Xenopus/metabolismo , Animais , Encéfalo , Membrana Celular/metabolismo , Evolução Molecular , Olho/citologia , Olho/metabolismo , Subunidades alfa de Proteínas de Ligação ao GTP , Humanos , Larva , Transdução de Sinal Luminoso/fisiologia , Opsinas/classificação , Opsinas/genética , Opsinas/metabolismo , Células Fotorreceptoras/citologia , Células Fotorreceptoras/fisiologia , Células Fotorreceptoras de Vertebrados/fisiologia , Filogenia , Células Fotorreceptoras Retinianas Bastonetes/citologia , Alinhamento de Sequência , Análise de Sequência de ProteínaRESUMO
Type 1 interferons (IFN) are an antiviral cytokine family, important in juvenile onset systemic lupus erythematosus (jSLE) which is more common in females, around puberty. We report that plasmacytoid dendritic cells (pDC) from healthy females produced more type 1 IFN after toll like receptor (TLR) 7 signaling than males, even before puberty, but that puberty itself associated with increased production of type 1 IFN. A unique human model allows us to show that this was related to X chromosome number, and serum testosterone concentration, in a manner which differed depending on the number of X chromosomes present. In addition, we have showed that pDC were more activated in females overall, and immune cell TLR7 gene expression was higher in females after puberty. Therefore, sex hormones and X chromosome number were associated individually and interactively with the type 1 IFN response, which contributes to our understanding of why females are more likely to develop an IFN mediated disease like jSLE after puberty.