To investigate the impact of personalized three dimensional (3D) printed pelvicalyceal system models on patient information before percutaneous nephrolithotripsy surgery. Patients with unilateral complex renal stones with indicatation of percutaneous nephrolithotripsy surgery were selected. Usable data of patients were obtained from CT scans as Digital Imaging and Communications in Medicine (DICOM) format. Mimics software version 16.0  was used for segmentation and extraction of pelvicalyceal systems. DICOM format were converted to Stereolithography file format. Finally, fused deposition modeling was used to create plasticine 3D models of pelvicalyceal systems. A questionnaire was designed for patients to assess personalized 3D models effect on patient’s understanding their conditions before percutaneous nephrolithotripsy surgery (PCNL). The day before surgery, each patient was seen by a urologist to deliver information about surgery. Questionnaire forms were asked to patients complete before and after presentation of 3D models and the results of the questions were compared. Five patient’s anatomically accurate models of the human renal collecting system were successfully generated. After the 3D printed model presentation, patients demonstrated an improvement in their understanding of basic kidney anatomy by 60% (p=0.017), kidney stone position by 50% (p=0.02), the planned surgical procedure by 60% (p=0.017), and understanding the complications related to the surgery by 64% (p=0.015). In addition, overall satisfaction of conservation improvement was 50% (p=0.02). Generating kidney models of PCSs using 3D printing technology is feasible, and understandings of the disease and the surgical procedure from patients were well appreciated with this novel technology. Three-dimensional 3D) printing is a new technology that has developed rapidly in recent years. This new approach holds significant benefits for medical procedures such as maxillofacial reconstruction . It has D printinPercutaneous nephrolithotripsy (PCNL) is a standard, safe, and efficient method for treating renal stones larger than 2cm in size . Access There is now a growing interest in providing information to support patient’s participation in choosing treatments and deciding on strategies for managing their health problems . InformaIn this study, our aim was to assess whether personalized 3D printed models of PCSs can improve patient’s understanding of their conditions before PCNL surgeries.Creating 3D models from medical imaging data is a multi-step process. First, usable data must be obtained from CT scans, magnetic resonance images, or ultrasound images. In our hospital , we used CT (Toshiba Alexion™ multislice CT) scan data from five patients. Data from CT scans in Digital Imaging and Communications in Medicine (DICOM) format are important because low-resolution images can result in inaccurate models .Second, segmentation, or extraction and isolation of the area of interest, of the data must be performed. We sent our DICOM-formatted data to a bioengineer  for segmentation. Numerous software programs are available for use with DICOM datasets. In our case, Mimics software version 16.0  was used.Finally, data must be saved in a file format recognized by the 3D printer software. The most commonly used format is the Stereolithography (stl) file format.Recent advances in 3D printing technology have produced new processes that allow the use of a variety of materials factors. Acrylonitrile butadiene styrene (ABS) has been used for creating 3D models of PCSs. The most important mechanical properties of ABS are impact resistance, toughness, high radiodensity, and low cost. We used fused deposition modeling, an inexpensive technology popular with consumers. These printers use a polymer filament that is heated to a liquid state in a printer head and deposited in predefined locations corresponding to the model shape (Stratasys Inc.) .A survey questionnaire with open ended questions of ordinal 10-point rating scales  was given to patients which consisted of 3 components: a) overall satisfaction of conversation (1 item), b) model assistance in understanding the disease and procedure (3 items) and c) understanding the complications related to the surgery (1 item). The day before surgery, each patient was seen by a urologist, the IVU images and CT scans were used as a teaching aid to deliver information about surgery and the disease. After the conversation, the questionnaire form asked patients to complete and then 3D printed models were presented to patients and they completed the questionnaire form again. Results of the questions were compared before and after the presentation of the 3D models.A total of 25 questions were asked to patients. Median total scores of responses for each category, before and after 3D printed model presentation, was compared (Wilcoxon test). Statistical analyses were performed using SPSS Statistics version 21.0 From June 2015 to January 2016, 5 patients with a unilateral staghorn renal stone and clinical indication of percutaneous nephrolithotomy were selected. Our first aim was to successfully create a 3D model of the pelvicalyceal systems. After two attempts we successfully generated anatomically accurate human renal collecting system. Overall collecting systems were clearly presented with regards to the virtual reconstructions. Fused deposition modeling with Acrylonitrile butadiene styrene (ABS) was used to generate anatomically-correct size and shape renal collecting system. The consumable costs of the model are low, at around 100$, and the print time for the 3D model is approximately 2 hours  and 2.Patients demonstrated an improvement in their understanding of basic kidney anatomy, planned surgical procedure and understanding the complications related to the surgery after viewing their personal 3D kidney models. After the 3D printed model presentation, it was found that the mean improvement rate of total scores was higher. Patients demonstrated an improvement in their understanding of basic kidney anatomy by 60% (p=0.017), kidney stone position by 50% (p=0.02), the planned surgical procedure by 60 % (p=0.017), and understanding the complications related to the surgery by 64% (p=0.015). In addition, overall satisfaction of conservation improvement was 50% (p=0.02).Accurate information which patients find useful has the potential to enhance the quality and appropriateness of health care. However, using only 2D images makes it very difficult for surgeons to inform their patients about the surgery and the complications. This is why new modalities have been developed for patient information . PCNL isIn the present study, we have created 5 physical, patient-specific, anatomically identical human renal collecting system 3D models before operative intervention, based on CT imaging of patients with unilateral staghorn renal stones. 3D models of the PCS can provide not only necessary images but are also feasible in planning collecting system access for planning PCNL surgery in complex staghorn renal stones.Printed models have been used for preoperative planning in complex orthopedic and craniofacial procedures and neurosurgery , 15. TheGenerating anatomically identical 3D models of renal collecting systems allows surgeons and patients to interact with the renal unit in a tangible way than using conventional images. In this study, it has shown that this interaction with models is an effective educational tool for patients resulting in an improvement over conventional imaging. With the 3D models, patients were better able to understand the renal anatomy and renal physiology, and the complications related to the surgery compared to CT scans and IVU images.This study has shown that, physician and patient communication were improved after presentation of personalized physical 3D models. 3D physical models are valuable for patient satisfaction of conservation with a 50% improvement. Despite given detailed information with IVU and CT images about planned surgery and complications that may occur, patient’s initial reference level of understanding was low. It is very difficult for patients to understand IVU and CT images and it can also be difficult for physicians to inform patients about the surgery and associated complications. After the presentation of 3D models, we witnessed how they helped patients raise and ask their own questions, enhancing their understanding. Improving patient education by the use of personalized 3D printed models appears to be a promising way to efficiently enhance the quality of personal exchange between a patient and his surgeon and influence overall patient satisfaction. In addition, patient education level is effective on the disclosure process.Moreover, using this kind of personalized 3D models not only for patient counseling but also for students, residents and fellow’s surgical teaching could help achieve better cost-effectiveness. Indeed, such models, in making easier 3D anatomical understanding, may certainly be useful tools to enhance surgical strategy discussion and improve preoperative planning.Generating kidney models of pelvicalyceal systems with 3D printing technology is feasible; understanding of the disease and the surgical procedure from patients were well appreciated with this novel technology.


Malaria in pregnancy has been associated with maternal morbidity, placental malaria, and adverse birth outcomes. However, data are limited on the relationships between longitudinal measures of malaria during pregnancy, measures of placental malaria, and birth outcomes.This is a nested observational study of data from a randomized controlled trial of intermittent preventive therapy during pregnancy among 282 participants with assessment of placental malaria and delivery outcomes. HIV-uninfected pregnant women were enrolled at 12–20 weeks of gestation. Symptomatic malaria during pregnancy was measured using passive surveillance and monthly detection of asymptomatic parasitaemia using loop-mediated isothermal amplification (LAMP). Placental malaria was defined as either the presence of parasites in placental blood by microscopy, detection of parasites in placental blood by LAMP, or histopathologic evidence of parasites or pigment. Adverse birth outcomes assessed included low birth weight (LBW), preterm birth (PTB), and small for gestational age (SGA) infants.The 282 women were divided into three groups representing increasing malaria burden during pregnancy. Fifty-two (18.4%) had no episodes of symptomatic malaria or asymptomatic parasitaemia during the pregnancy, 157 (55.7%) had low malaria burden , and 73 (25.9%) had high malaria burden during pregnancy . Women with high malaria burden had increased risks of placental malaria by blood microscopy and LAMP , compared to the other two groups combined. Compared with women with no malaria exposure during pregnancy, the risk of placental malaria by histopathology was higher among low and high burden groups . Detection of placental parasites by any method was significantly associated with PTB [aRR 5.64 (1.46–21.8)], and with a trend towards increased risk for LBW and SGA irrespective of the level of malaria burden during pregnancy.Higher malaria burden during pregnancy was associated with placental malaria and together with the detection of parasites in the placenta were associated with increased risk for adverse birth outcomes.Trial Registration Current Controlled Trials Identifier NCT02163447The online version of this article (doi:10.1186/s12936-017-2040-4) contains supplementary material, which is available to authorized users. Malaria in pregnancy remains a major public health problem in many parts of sub-Saharan Africa partly because the coverage of malaria control measures is still low. The proportion of the population sleeping under an ITN and protected by IRS was estimated to be 57% in 2015 and only 31% of eligible pregnant women received three or more doses of IPTp in the same year , 5. ApprAs most available data are from cross-sectional studies at the time of delivery, there are limited longitudinal data on malaria during pregnancy, and the impact of gestational malaria burden on measures of placental malaria and birth outcomes. A prospective study in Benin showed an association between microscopic and submicroscopic infections (detected by PCR) measured early in pregnancy with and increased risk of LBW and PTB, respectively , but didThe overall objective of this study was to fill the evidence gap on the relationships between longitudinal measures of malaria burden during pregnancy, placental malaria and birth outcomes. Factors associated with the frequency of symptomatic malaria and asymptomatic parasitaemia during pregnancy were examined, and the associations between malaria burden and measures of placental malaria and adverse birth outcomes assessed.This was a cohort study that utilized data from a randomized controlled trial of IPTp in Tororo, Uganda. Tororo is a rural district in southeastern Uganda with an entomologic inoculation rate estimated at 310 infectious bites per person year in 2012 . From JuAt enrollment, pregnant women underwent a standardized history and physical exam including assessment of wealth status, gravidity, gestational age by ultrasound, and age. Each participant received a long-lasting insecticide-treated bed net (LLIN). Women received all their medical care at a designated study clinic that was open daily. Routine visits were conducted every 4 weeks, including collection of dried blood spots (DBS) for LAMP. Participants with positive LAMP results were not treated for malaria. Women were also encouraged to present to the clinic with any illness. Patients who presented with a documented fever (tympanic temperature ≥ 38.0 °C) or a history of fever in the prior 24 h had peripheral blood collected for a thick blood smear. If the blood smear was positive, the patient was diagnosed with symptomatic malaria and treated with artemether–lumefantrine. At delivery, a standardized assessment was completed including evaluation of infant birth weight and gestational age, and collection of specimens within 1 h of delivery, including placental tissue, placental blood smears, and DBS of placental blood. All women were encouraged to deliver at the hospital adjacent to the study clinic. Women delivering at home were visited by study staff as close to the time of delivery as possible for assessment and sample collection.DBS were tested for the presence of malaria parasites using LAMP as previously described , 14. ForThe following demographic data was collected: maternal age, possession of bed net at enrollment, wealth status, gravidity, gestational age, and IPTp treatment arm. Wealth status was categorized into lowest, middle, and highest tertiles designed as a composite variable using ownership of several household items and land. Gravidity was grouped as primigravidas (1st pregnancy) and multigravidas (≥ 2 pregnancies). Gestational age at enrollment was confirmed by ultrasound measurements by fetal biometry at less than 20 weeks’ gestation. IPTp treatment arm was categorized as SP given every 8 weeks, DP given every 8 weeks and DP given every 4 weeks.Malaria in pregnancy was defined as both symptomatic malaria and asymptomatic parasitaemia. Symptomatic malaria was measured using passive surveillance and defined as fever with positive blood smear. Asymptomatic parasitaemia was measured using active surveillance every 4 weeks and defined as molecular detection of malaria parasites from a DBS by LAMP. Measures of placental malaria at the time of delivery included: the detection of malaria parasites in placental blood by both microscopy and LAMP, and histopathologic evidence of placental malaria (parasites and/or pigment) from placental biopsies. Birth outcomes assessed included: LBW (< 2500 g), PTB , and SGA  . In thisData were double entered into a Microsoft Access database. Data analysis was done using Stata 14 . For baseline characteristics, comparison of proportions was done using the Informed consent was obtained from all study participants. Ethical approval was obtained from the Uganda National Council of Science and Technology, the Makerere University School of Medicine Research and Ethics Committee, the Makerere University School of Biomedical Sciences Research and Ethics Committee, and the University of California, San Francisco, Committee on Human Research.A total of 57/282 (20.2%) women had symptomatic malaria during pregnancy. Of these, 44/282 (15.6%) had only one episode of symptomatic malaria during pregnancy and 13/282 (4.6%) had more than one episode (2–3 episodes), resulting in a total of 72 malaria episodes. Asymptomatic parasitaemia during pregnancy was detected by LAMP on DBS in 230/282 (81.6%) of women; 162/282 (57.4%) had asymptomatic parasitaemia with < 50% of their monthly DBS samples positive for malaria parasites and 68/282 (24.1%) had ≥ 50% of their DBS samples positive for malaria parasites.To investigate the associations between the frequency of malaria infection during pregnancy and placental malaria, a categorical variable of malaria burden during pregnancy was created. Three groups representing increasing malaria burden were defined as follows: 1) “none” = women who had no episodes of symptomatic malaria or asymptomatic parasitaemia during pregnancy; (2) “low” = women with 0–1 episode of symptomatic malaria and < 50% of DBS samples positive for parasites by LAMP during pregnancy; and (3) “high” = women who had ≥ 2 episodes of symptomatic malaria or ≥ 50% of their DBS samples positive for parasites by LAMP during pregnancy  of placental biopsies. Risks of placental malaria were: 8/280 women (2.9%) by microscopy of placental blood smears, 24/280 women (8.6%) by LAMP of placental blood, and 105/282 women (37.2%) by placental histopathology. All 105 placentas positive by histopathology had pigment in fibrin indicative of past infection, but only 7 had parasites indicative of concomitant active infection. There were a total of 25 cases of placental malaria in which parasites were detected by any method  women had LBW infants, 26 (9.2%) women delivered preterm, and 57 (20.2%) had infants that were SGA. A composite indicator of gestational malaria burden and placental malaria was generated to explore associations between malaria in pregnancy with birth weight, and adverse birth outcomes. Women were grouped into the following categories: (1) “none” category with no symptomatic or asymptomatic malaria and no placental malaria, (2) low malaria burden and no placental malaria, (3) high malaria burden and no placental malaria, (4) any malaria burden  and placental malaria with pigment only on histopathology, and (5) any malaria burden and placental parasites detected  , there was a non-significant trend towards increased risk of LBW, PTB, and SGA in category 3  in univariate analysis  from placental biopsies. When evaluating associations between a composite indicator of malaria in pregnancy and adverse birth outcomes , the detection of malaria parasites in the placenta was found to be a statistically significant risk factor for PTB. However, it should be noted that due to the relatively small sample size of some categories of malaria in pregnancy, the lack of statistically significant associations with adverse birth outcomes may have been due to a lack of statistical power.The results presented here are consistent with prior studies that demonstrated an association between peripheral parasitaemia during pregnancy and placental malaria , 10, 18.Plasmodium falciparum infection during pregnancy and adverse pregnancy outcomes, such as LBW, PTB, and SGA [Prior longitudinal studies have also shown associations between peripheral  and SGA , 18, 19.Associations between the presence of placental malaria and an increased risk of LBW and/or PTB have been previously reported in several cross-sectional studies , 20, 21.Low birth weight can be due to prematurity, or SGA , or both. SGA infants have been shown to have twice the mortality risk compared to appropriate for gestational age infants in low and middle income countries, for both preterm and term births , 23. FurAs ultrasound technology has become more widely available in malaria-endemic regions, the importance of distinguishing between these factors has become more apparent. For example, Rijken et al. showed that in a Thai population, even treated malaria in pregnancy was associated with an increased risk of LBW, PTB, and SGA; SGA was present in 5% of malaria-affected pregnancies in non-LBW babies born after 39-weeks of gestation . Thus, bPrimigravidity is a well-established risk factor for increased susceptibility to malaria in pregnancy and adverse clinical outcomes , 28, conOf note, in this study LAMP was used to screen for asymptomatic parasitaemia. LAMP is more sensitive than microscopy, in both asymptomatic and symptomatic cases in pregnancy , 31, andThree methods of detection of placental malaria were directly compared: placental blood smear, LAMP of placental blood, and histopathology. While a high prevalence (37%) of placental malaria with only pigment on histopathology was observed, this measure was not significantly associated with adverse birth outcomes. This is not surprising given that pigment in the placenta represents prior infection. In contrast, the presence of parasites in the placenta, diagnosed by any of the three methods, was significantly associated with PTB and with an increased risk for LBW and SGA. LAMP was the most sensitive of the three, demonstrating that this method may provide the optimal diagnostic test for placental malaria. Additionally, LAMP has clear advantages over the other two methods in low resource settings—it is easy to learn, does not require specialized equipment, and is low cost with rapid results. In contrast, placental blood smear is less sensitive (detecting 1/3 of the cases diagnosed by LAMP in the study cohort), and placental histopathology is resource-intensive and not routinely available in malaria-endemic settings.This study did have limitations. Malaria burden prior to enrollment at 12–20 weeks gestation was not evaluated, which may have influenced both measures of placental malaria and birth outcomes. Other factors that could affect fetal growth, birth weight, and prematurity were not investigated, such as maternal nutritional status, child spacing, maternal anaemia, genetics, other infections, and smoking. Serial ultrasound examinations during the pregnancy were not performed, which could help to distinguish constitutionally SGA (for example from genetic factors) from SGA due to fetal growth restriction. Given the relatively small sample size and low frequency of several of the variables of interest, the lack of statistically significant associations may have been due to insufficient power rather than true negative findings (type II error). However, this study also has several strengths. One is the longitudinal design with frequent (monthly) measurements of parasitaemia by molecular methods, obstetric dating of gestational age by ultrasound prior to 20 weeks gestation, and comprehensive analysis of placental malaria.Increasing malaria burden in pregnancy was associated with primigravidity, lower household wealth, and IPTp treatment regimen . Higher malaria burden in pregnancy was associated with placental malaria, and LAMP was the most sensitive measure of malaria parasites in the placenta compared with placental blood microscopy and histopathology. The detection of malaria parasites in placental samples was associated with an increased risk of adverse birth outcomes.


We report the first case of severe respiratory failure after thyroid surgery requiring venovenous extracorporeal membrane oxygenation (vvECMO). The patient was a 41-year-old woman with metastatic thyroid cancer. She underwent thyroidectomy, including left lateral and bilateral central neck dissection. During surgery, the patient developed pneumomediastinum with bilateral pneumothoraces. Despite early treatment with bilateral chest tubes and no evidence of a tracheal perforation, the patient developed severe respiratory failure after extubation on the intensive care unit. Because pneumothorax and pneumomediastinum might be more common than reported, and considering increasing cases of thyroid surgery, staff should remain vigilant of pulmonary complications after thyroid surgery. The incidence of thyroid cancer has more than tripled since 1975 and, in women, it is the cancer with the fastest-growing number of new cases . In fact2 of 96% on room air.A 41-year-old, 75 kg, 150 cm (body mass index of 33), woman with thyroid cancer metastatic to the bilateral neck underwent thyroidectomy, including left lateral and bilateral central neck dissection. No chemotherapy or radiation therapy was employed as an adjunctive prior to surgery. Her medical history included well-controlled arterial hypertension and diabetes mellitus type 2. She was a nonsmoker and denied any history of cardiac or pulmonary disease. The physical exam prior to surgery was unremarkable. The initial CT scan of the neck showed no abnormalities of the trachea . The pre2, ETCO2, and NIBP [IBP added after 4 hours of surgery]) was established and maintained throughout the procedure. After intravenous pretreatment with midazolam 2 mg and fentanyl 0.15 mg, general anesthesia was induced using propofol 200 mg, followed by 80 mg succinylcholine to facilitate intubation. We maintained anesthesia using desflurane inhalation and remifentanil infusion (0.2 μg/kg/min). To minimize the risk of recurrent laryngeal nerve damage, we used an electromyogram endotracheal tube , placed using a video laryngoscope and inflated the cuff to a pressure of 25 cm H2O. Then, guaranteed volume, pressure-controlled ventilation was established and we maintained the end-tidal CO2 (ETCO2) levels at 35 mmHg with tidal volumes (TV) of 350 ml and a respiratory rate of 15/min. The adjustable pressure limiting valve was set to 5 cm H2O and average peak inspiratory pressures (PIP) and mean airway pressures (AP mean) were 22 cm H2O and 12 cm H2O, respectively.Standard anesthesia monitoring  and received 50 mg propofol (i.v.). During cervical lymph node dissection and thymus removal, the care team noted a gurgling and bubbling sound radiating from the thorax. We checked the cuff pressure and tube position using a video laryngoscope, but no abnormalities were found. Three hours into the surgery the thyroid was unroofed and the thyroid ligament was dissected. In addition, all the lymphatic tissue from the carotid and innominate artery towards the midline was cleared out. In addition, the esophagus nerve and the side of the trachea from the thyroid cartilage down to the innominate artery just below the sternum were skeletonized. Again, a cycle of gurgling and bubbling of air into the neck occurred and the patient became difficult to ventilate. The patient slightly desaturated (SpO2 91%) and low tidal volume alarms were noticed. The FiO2 was set to 1.0 and the patient was manually ventilated. During manual ventilation decreased resistance was noted and TVs were as low as 42 ml, PIPs were as low as 11 cm H2O, and AP mean was as low as 7 cm H2O. The ETCO2 was 30 mmHg due to increased manual ventilation rates (30/min). To check for an airway leak, the surgical team poured normal saline over the surgical field and observed bubbles in the saline but could not identify any lacerations of the trachea or pleural defects. As ventilation remained difficult with TVs constantly dropping and requiring consecutive recruitment maneuvers over the next 30 minutes, a bronchoscopy was performed. The cuff was deflated and the ET tube was withdrawn until the vocal cords became visible. Ventilation was paused and the bronchoscopy was performed which showed a normal looking tracheal and lobar bronchial mucosa. However, the distal trachea and left mainstem bronchus appeared dynamically collapsed. Subsequent intraoperative chest X-ray revealed a small left-sided pneumothorax and a mild pneumomediastinum  in the absence of cardiac failure, the care team diagnosed a severe respiratory distress syndrome (ARDS) based on the Berlin definition  with SpO2s in the 80s , the patient was placed on venovenous extracorporeal membrane oxygenation (vvECMO). Over the following three days the patient's respiratory function improved. VvECMO therapy was discontinued and the patient was subsequently extubated. Follow-up echocardiography found normal heart function with an EF of 73%. We discharged the patient from the hospital on postoperative day 12 in good physical condition. Due to posttraumatic stress disorder, the missing right lateral neck dissection was postponed but successfully performed eight months later.During positioning, the patient coughed . Direct laryngoscopy and auscultation ruled out a partial extubation . As the ETT cuff maintained the applied pressure, a cuff leak seemed unlikely. In fact, the majority of airway leaks are not associated with cuff defects . Thus, tWhen air enters the thoracic cavity or the mediastinum, pneumothorax and pneumomediastinum develop. This requires a direct communication either between the alveoli and the pleural cavity or between the atmosphere and the pleura . This caIn our case, the thyroid gland was described as “rock” hard and after dissecting the thyroid, the thyroid ligaments, and the deep cervical lymph nodes, gurgling and bubbling of air into the neck and difficult ventilation occurred. The size of the excised thyroid was 7.2 cm from superior to inferior, 6.0 cm from right to left, and 2.0 cm from posterior to anterior and weighing 35 grams. In addition, the papillary thyroid carcinoma had spread to the thymus and several lymph node levels (levels III–V), with the largest metastatic focus at a level V lymph node (1.3 cm). The relative large size and the changed tissue structure of the thyroid indicate a difficult dissection which is also reflected by the time (3 hours) it took to unroof the thyroid. Thus, the difficult “unroofing” of the thyroid in conjunction with a deep cervical lymph node dissection makes traumatic pneumothorax likely.However, in our patient pneumomediastinum and pneumothorax could have also developed unrelated to surgery. During intraoperative bronchoscopy, we found dynamic airway collapse at the distal trachea and the left main stem bronchus. This finding can imply preexisting and undetected airway disease , includiWhy the patient developed ARDS postoperatively is unclear, since common causes such as pneumonia, sepsis, or severe trauma were absent. However, the literature reports one case of unexpected severe respiratory failure in a surgical patient in the presence of dynamic airway collapse undetected prior to surgery . Thus, aIn conclusion, we report the first case of acute respiratory failure with ECMO treatment following thyroid surgery, complicated by pneumomediastinum and bilateral pneumothoraces. It highlights that anesthesiologists need to be aware of the anatomical routes and the pathogenesis of air trapping in patients undergoing thyroid surgery in particular in cancer-related bilateral neck dissection. Even in the absence of an obvious airway perforation, the care team should critically evaluate the airway before early postoperative extubation. As also suggested by others , any une


The size, magnesium (Mg) content and molecular-scale skeletal disorder of CCA patches were assessed at 3.5, 6.5 and 14 months from tile deployment. Despite reductions in their abundance in low pH, the largest CCA from ambient and low pH zones were of similar sizes and had similar Mg content and skeletal disorder. This suggests that the most resilient CCA in low pH did not trade-off skeletal structure to maintain growth. CCA that settled in the extremely low pH, however, were significantly smaller and exhibited altered skeletal mineralogy ), although at present it is unclear if these mineralogical changes offered any fitness benefits in extreme low pH. This field assessment of biological effects of OA provides endpoint information needed to generate an ecosystem relevant understanding of calcifying system persistence.To understand the effects of ocean acidification (OA) on marine calcifiers, the trade-offs among different sublethal responses within individual species and the emergent effects of these trade-offs must be determined in an ecosystem setting. Crustose coralline algae (CCA) provide a model to test the ecological consequences of such sublethal effects as they are important in ecosystem functioning, service provision, carbon cycling and use dissolved inorganic carbon to calcify and photosynthesize. Settlement tiles were placed in ambient pH, low pH and extremely low pH conditions for 14 months at a natural CO Laboratory experiments provide critical insight into the direct effects of environmental change on individual species, but it is often unclear how such effects will be balanced with other responses, or manifest in more complex ecosystems or naturally variable environments. Thus, a better understanding of the trade-offs among sublethal effects of environmental change, as well as how the sublethal effects reported in laboratory experiments materialize in field environments can improve forecasts of the ecological effects of global change.2) into seawater, is projected to have widespread impacts on marine species and ecosystems in the near future [3) are projected to be some of the worst affected, as changes in seawater chemistry may increase the energetic costs of building and maintaining CaCO3 structures [Ocean acidification (OA), caused by the absorption of atmospheric carbon dioxide  [Coralline algae are important in ecosystem functioning and service provision; they create habitats and nursery areas, host high biodiversity, stabilize coral reef systems and contribute to carbon cycling –11. Moreskeleton . The var% MgCO3)  with occ% MgCO3) . The sol% MgCO3) .pCO2 on coralline algae include reduced growth rates [pCO2 conditions stimulated by day-time hypercalcification (rapid deposition of CaCO3) [pCO2 environments have found that some coralline algal species can grow at similar rates in ambient and high pCO2 conditions, and reductions in the abundance of coralline algae in these more acidic conditions may be driven by altered species interactions [The negative effects of increasing th rates ,17, calcth rates , epithelth rates  and distth rates ,20–22. Ef CaCO3) ,23. Fielractions ,25.pCO2 conditions can come at the expense of skeletal structure [pCO2 environments. For example, OA-induced hypercalcification leads to CaCO3 skeletal material with poor molecular-scale ordering [pCO2 induced a change in skeletal mineralogy to less soluble calcium sulfates [Continued skeletal accretion under high tructure ,16,26 ortructure . A tradeordering  and lesssulfates . Such alsulfates ; for exasulfates ,19, trad2 vents create gradients in carbonate chemistry that have been used to study the effects of OA on coralline algae in the context of other co-occurring species. Previous studies have concluded that crustose coralline algae (CCA) from CO2 vents can maintain growth rates in low pH conditions, but it is unknown whether the maintenance of growth resulted in a trade-off with skeletal structure or masked other sublethal effects that limited the abundance of CCA in low pH [pCO2, ecosystem context.To understand the ecological effects of OA on coralline algae, we need to examine whether energetic trade-offs result in sublethal effects on CCA, which may in turn impact their abundance in an ecosystem. Observational field studies using natural gradients in environmental conditions provide opportunities to study the emergent effects of environmental factors on species and ecosystems. Such gradients incorporate environmental variability and species interactions that may mediate the sublethal effects of stressors . In partn low pH . Here, w2.(a)2 vents (0.5–3 m depth) occurring on the north and south side of the small islet, Castello Aragonese at Ischia island . Gas was emitted at 0.7 × 106 l day−1 in the north vent (area approx. 2000 m2) and 1.4 × 106 l day−1 in the south site (area approx. 3000 m2) with no evidence of seasonal, tidal or diurnal variation in gas flow rates [in situ monitoring of seawater pH delineated a pH gradient with three carbonate chemistry zones , caused by spatial variability in CO2 venting intensity [North,South = 8.0, 8.1; mean saturation state of calciteNorth,South = 4.8, 5.1), whereas the low pH zones are comparable with values predicted for 2100  [in situ pH sensors and discrete water samples across May–June 2010 [Research was conducted at the shallow volcanic CO.0, 3.5)   growing on the tiles were identified under a dissecting scope using morphological features. All species showed the same responses  so were pooled for subsequent analyses to enable statistically meaningful replication and interpretation.Nineteen recruitment stone tiles (15 × 15 × 1 cm) were attached to the substrate in each pH zone  in both sites (north and south) at 1.5 m depth as part of a previous study  figure . The til(b)Tiles were photographed , and coralline algal size was determined using Image J . The 50 (c)−1 grating via a 20× objective. Raman spectra were taken from individual CCA on six randomly selected tiles, ensuring all the genera were represented where possible. To remove organic material from the CCA surface, they were exposed to laser light prior to analysis for up to 60 s. However, fluorescence interference still caused some spectra to be rejected. Owing to the low abundance of CCA in the extreme low pH zone, only 20 Raman spectra were collected for extreme low pH . Raman spectroscopy was carried out using Renishaw inVia Raman microscope equipped with a Leica DM 2500 M  microscope at the School of Geographical and Earth Sciences at the University of Glasgow . CCA were analysed on the tiles and were excited using a 785 nm laser with a 1200 mm(d)4 · 2H2O): ID: R040029) minerals are included in electronic supplementary material, figure S1, because of their presence in the samples.Raman spectroscopy was used to identify the calcium mineral present by comparing the sample spectra with calcium minerals in the RRUFF library . Calcite(e)−1 peak in spectra of biogenic calcite is primarily controlled by the Mg content (positive relationship) [−1 peak increases with Mg concentration due to Mg ions being smaller than calcium ions, thereby causing a decrease in the interatomic distances and an increase in vibrational frequencies [−1 peak (a proxy of relative Mg concentration) for each individual was extracted from the corresponding spectrum for analysis. Higher frequencies indicate more Mg within the high Mg skeleton. Where calcium sulfate was present as the skeletal mineral, this peak was replaced by a peak at approximately 1008 cm−1.The exact frequency of the 1089 cmionship) ,34–36. Tquencies . The exa(f)−1 Raman spectrum peak is due to the positional disorder of the carbonate ions in the calcite structure, which is characterized by increased rotation of the planar carbonate group out of the basal plane [a-axis in the direction of the c-axis [−1 peak for each individual was extracted from the corresponding Raman spectrum for analysis.Variation observed in bandwidth (FWHM) of the 1089 cmal plane . Specifie c-axis . With Mge c-axis ,34. Disoe c-axis . The FWH(g)10 transformed to meet general linear model assumptions. Time was not treated as a repeated measure as at each time point separate tiles were collected and removed, thus individuals were not re-sampled.At 14 months, coralline algae were not observed growing on the tiles in the extreme low pH zone at the north site which prevented a balanced analytical design. Similarly, in the extreme low pH treatment, high Mg calcite was only present in CCA at 3.5 months. At all other time points where individuals were present the skeletal calcium carbonate mineralogy changed to gypsum, preventing direct comparison of the mineralogy. Thus, extreme low pH zones were excluded from statistical analysis for Mg composition, carbonate ion positional disorder and area. Analyses of variance  with Tukey's pairwise comparisons were used to determine differences in Mg composition, carbonate ion disorder and areas between time and pH zone (both fixed) with site as a random variable with interactions being examined. Area was log3.(a)F1,100 = 3.41, p = 0.068; F2,100 = 177.33, p < 0.001 with Tukey's HSDs p < 0.002 were significant). These differences generated significant interactions between month and site  as well as month and pH zone  but not site and pH . At 6.5 months, CCA in the north site were significantly larger than those in the south site (Tukey's HSD p = 0.01). Few coralline algae were present in the extreme low pH zones During all sampling time points at both ambient and low pH zones, the skeletal material of all CCA was composed of calcite (high Mg) or a mix of calcite (high Mg) and aragonite . However, in the extreme low pH zones, CCA were only composed of high Mg calcite in the north site at 3.5 months. CCA in the extreme low pH zones of the north site at 6.5 months and south site at 3.5, 6.5 and 14 months had skeletal material composed of a gypsum mineral. In the north extreme low pH sites, CCA were not present on the tiles at 14 months .−1 Raman spectrum peak) of CCA calcium carbonate did not differ between ambient and low pH locations . In the north site, Mg concentrations were significantly lower at 6.5 months than 3.5 months . There were no significant interactions between month and site , month and pH  or site and pH .Mg concentrations (peak frequency of the 1089 cm(d)−1 Raman spectrum peak) of CCA calcium carbonate did not differ significantly between ambient and low pH locations . In the north site, positional disorder was lower at 3.5 and 14 months than 6.5 months and in the south site positional disorder was lower at 3.5 months than 14 months .Positional disorder (FWHM of the 1089 cm4.pCO2 conditions similar to those projected for 2100 (i.e. the low pH zones of the Castello vents) can maintain growth and calcification without detectable trade-offs in skeletal structure. The size of the largest CCA patches did not differ between ambient and low pH conditions over 14 months, suggesting growth of some individuals can be maintained in low pH (mean 7.8). This unexpected result is in agreement with growth measurements taken during the early stages of succession using repeated measurements on discrete patches of CCA over 3.5 months [in situ results indicate that the maintenance of growth of these individual CCA patches did not come at the expense of sublethal changes in skeletal structure, suggesting that these individuals may be already acclimated/adapted to these low pH conditions. The reduced CCA per cent cover observed in [Individual CCA patches growing in low pH/high 5 months , as well5 months . While r5 months  found therved in  could sierved in , or thaterved in  and otheerved in ,40.The calcium mineral composing the CCA skeleton in the ambient and low pH zones was calcite or calcite with some aragonite. Similarly, there was a general trend of no difference in the Mg content and skeletal molecular-scale structural disorder of the CCA grown in ambient and low pH zones, suggesting these temperate CCA can continue to accrete high Mg calcite at low pH levels projected for the turn of this century. This also occurs in the surface of tropical CCA, which maintain their Mg concentrations in reduced pH . The preet al. [et al. [This study, Diaz-Pulido et al.  and Nash [et al.  suggest  [et al. ,16,44 in [et al. ,36 have  [et al. , and pul [et al. ), as opppCO2 conditions in the diffusive boundary layer due to (i) respiration at night, and (ii) conversely, it is possible that turf algae in close association with CCA could locally elevate seawater pH in the diffusive boundary layer during photosynthesis to buffer the CCA from the acidic waters in the low pH zone [Although high Mg calcite becomes less stable in low pH, it is unknown if incorporating and maintaining Mg in the calcite lattice is more energetically costly in low pH conditions. Moreover, little is known about the seawater chemistry in the diffusive boundary layer of CCA at this site. It is thus possible that CCA regularly experience variable pH/high  pH zone ,47. If t pH zone  could poPadina pavonica, which changes from aragonite to gypsum in low pH [pCO2 [In contrast with the CCA in low pH, those found in extreme low pH were rare and very small. This suggests they are (i) selected against by critically low pH levels, (ii) grow more slowly or (iii) are affected by other post-settlement processes (including faster dissolution) and therefore play a reduced ecological role in these conditions. Their presence, however, allowed us to better understand how the mineralogy, Mg content and structural disorder of CCA can change in extreme pH conditions. In the extreme low pH zone, the skeletal mineral changed from calcite or calcite and aragonite in the ambient and low pH zones to gypsum by either 3.5 (South) or 6.5 (North) months. Gypsum contains no Mg and is more stable than calcite or aragonite in low pH . As we dn low pH . Other on low pH ,49, medun low pH ,51 and mn low pH . So alonn low pH  and thuspH [pCO2 ,54. DespPadina pavonica [The variability in the responses of extreme low pH CCA between sites and among time points restricts our ability to make robust conclusions about the impacts of extreme low pH, but it also provides an opportunity to develop hypotheses about the potential drivers of variability. We cannot determine the physiological condition of the CCA from the extreme low pH at the time of collection . If the CCA were dead, they may have not survived in the extreme low pH treatment despite the mineralogical changes. However, there is evidence that live calcifying algae can change their calcium mineral . Thus, ipavonica  are copiIn the extreme low pH zone in the south site, the gypsum is present throughout the time series. By contrast, the gypsum only appears in the extreme low pH zones in the north site at 6.5 months. If the CCA were alive, this inconsistency among sites could potentially be due to differences in the timing of CCA recruitment to the tiles at each site. For example, recruitment may have happened earlier in the south than the north. Differences in the age of the recruits would correspond to different lengths of CCA exposure to extreme low pH, which could explain the site differences. Future studies with higher temporal resolution of the Mg content of CCA recruits can help test this hypothesis.It is important to note that the reactivity of calcium minerals to their environment is not solely controlled by the concentration of Mg in their skeletons. Rather, the reactivity of the calcium minerals post deposition is driven by a combination of (i) the saturation state of the surrounding fluid or water, (ii) the microstructural complexity of the biomineral, and (iii) the mineral stabilities . It is tEnvironmental variability between the north and south could contribute to the differences seen in the responses of area, calcite mineral, Mg content and positional disorder to pH between the north and south sites at different time points . For example, the north and south sites are characterized by different exposures to light intensity, dominant winds, wave action and fetch. Generally, the south site is more sheltered and the pH gradient is more consistent . BecausepCO2 enrichment is a more recent process [We observed no differences in response between the different genera of CCA  to individual site pH zone-time points. It may be that this apparent equivalency among taxa is due to the relatively low numbers of available individuals from each genus  causing low statistical power. However, if this pattern holds across larger populations of individuals from particular genera, it is possible that the estimated millennial longevity of the venting systems at Castello Aragonese  has been process  and adap5.pCO2 enrichment projected for 2100, although their abundance is probably limited by their interactions with other species at these pCO2 levels [CCA present in a naturally acidified ecosystem appear to be skeletally robust to the 2 levels . In more


Renal transplant recipients have increased risk for developing malignant diseases because of immunosuppression or donor-to-recipient transmission. Malignant rhabdoid tumor (MRT) is a rare, highly aggressive and lethal tumor primarily affecting the kidney of infants and young children. MRT has not been reported in the renal allograft of an adult recipient after kidney transplantation.In this report, a 47-year-old woman who received a kidney transplantation from an infant donor and developed a mass in the transplanted kidney is presented. Pathological examinations revealed a malignant tumor with rhabdoid cells morphologically and the loss of INI1 expression immunohistochemically. The diagnosis of malignant rhabdoid tumor in the transplanted kidney was made. We confirmed that donor-to-recipient malignancy transmission was the cause of MRT in the transplanted kidney by fluorescence in situ hybridization (FISH) and short tandem repeat (STR) analysis.To our knowledge, this is the first case of MRT in an adult renal allograft recipient. This report highlights the importance of the criteria for selection of donors to screen possible malignant tumors transmission. SMARCB1/INI1 gene located on chromosome 22q 11.2 genetically [Graft survival has improved considerably in kidney transplantation thanks to potent immunosuppression. On the other hand, the overall incidence of malignancy after renal transplantation is much higher than in the general population . The majetically . Here, wApproval was obtained from Ethics committee of Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology. Written informed consent was obtained from the patient in this study.nifedipine for hypertension for three years, but her health conditions worsened. A renal function examination showed an elevated serum creatinine (Scr) level (342 μmol/l), and symptomatic treatment did not improve renal function before dialysis. After six months of dialysis, the patient underwent kidney transplant operation since a 56-day-old infant who died from a central nerve system (CNS) tumor  came to the hospital as a kidney donor by his parents and was found to be suitable for kidney donation. There was no history of rhabdoid tumors in the donor family history.A female patient was diagnosed with thrombocytopenia due to persistent gum bleeding and lower extremity congestion at the age of 40 years in a local hospital. Subsequently, she was detected to have hypertension. She took Dual kidneys were transplanted in the right iliac fossa of the recipient  methanol solution treatment was done for 30 min to eliminate nonspecific reaction with proteins. The primary antibodies applied are given in Table The removed transplanted kidney measured 9.0 × 7.9 × 7.5 cm. The cut surface showed a 7.4 × 6.3 × 6.0 cm mass without a capsule located in the upper pole, replacing almost the entire kidney Fig. . The samMicroscopic examination showed patternless sheets or nests of noncohesive, uniform, round and oval tumor cells having eccentric nuclei with macro-nucleoli and abundant eosinophilic cytoplasm Fig. . Some ofcarbonic anhydrase IX(CAIX), CD34, CD56, chromogranin A (CgA), synaptophysin (Syn), GATA binding protein 3 (GATA-3), CD10, CK5/6, P63, CD117, P504s, transcription factor binding to IGHM enhancer 3 (TFE-3), S-100, HMB45, MelanA, CD38, and CD138) were not expressed in the tumor cells. The Ki67 labeling index was approximately 80%  and cytokeratin, but negative for SMARCB1/INI1Fig. . All oth80% Fig. .SMARCB1/INI1 and sex chromosomes, paraffin-embedded 5-μm sections were deparaffinized. A probe specific for SMARCB1/INI1  and a dual-color interphase FISH probe set for the X centromere (CEP X) and Y centromere (CEP Y)  were used to detect any abnormality of SMARCB1/INI1 and male (XY) donor or female (XX) recipient cells present in the kidney tumor specimen according to the manufacturers’ protocols. The fluorescence signals were analyzed using an Olympus BX51 fluorescence microscope  equipped with appropriate filters and imaged using Vysis software. At least 200 cells were scored.For FISH analysis of SMARCB1/INI1 was deleted . Multiplex genotyping was performed with the EX22 STR kit  and the AmpFLSTR® Identifiler® PCR Amplification kit  on the case samples according to the manufacturers’ recommendations. The experimental procedures followed internal laboratory control standards and kit controls.For the short tandem repeat (STR)-based concordance study, genomic DNA was extracted from tumor tissue, normal renal tissue samples of the transplanted kidney and The STR results showed that of the 22 microsatellite markers tested, differences were found between tumor cells Fig.  and skelPrimary tumors in the kidney allograft recipients have often been reported since the advent of potent immunosuppression. The most common malignant tumors of organ transplant recipients are post-transplant lymphoproliferative diseases and skin cancer . To our SMARCB1 (hSNF5/INI1), which can be identified by a lack of staining with INI1 immunohistochemically. Fewer than 10 cases of MRTs are reported in adult patients in the English-language literature [MRT is a rare neoplasm that occurs mainly in the kidney in children less than 1 year of age, with an aggressive clinical course . Some caterature , 9, and INI1 deficiency. Epithelioid sarcoma is a malignant tumor affecting an older age group than MRT, with prominent rhabdoid cells and loss of nuclear INI1 in most cases, and it often shows CD34-positive staining, mainly located in the extremities. Other malignancies, including rhabdomyosarcoma, myoepithelioma in soft tissue, epithelioid angiosarcoma, and epithelioid malignant peripheral nerve sheath tumor (MPNST), can be distinguished from MRT by their specific markers and INI1 expression. Loss of INI1 is also helpful to differentiate MRT from other primary renal neoplasms with rhabdoid differentiation, including clear cell renal carcinoma, chromophobe cell renal carcinoma, and other malignances in the urinary system [Other primary renal neoplasms and certain epithelioid or rhabdoid cell lesions arising from the kidney and urinary tract should be distinguished from MRT upon diagnosis. Nephroblastoma (Wilms tumor), a common solid tumor of childhood, typically shows a distinct triphasic pattern with blastema, stromal, and epithelial components and without y system , 12.Due to the rarity of MRT in transplanted kidney, FISH and STR analysis were used to determine the tumor origin in this case. The results showed a Y chromosome in the tumor cells, which was consistent with donor origin. In general, malignant tumors in allografts can develop in two ways: de novo development in the allograft of an immunosuppressed recipient  and donoThere was no history of rhabdoid tumors in the donor family history.Although we cannot refute the diagnosis of astrocytoma and prove the presence of AT/RT, we strongly suspect the CNS tumor of the infant donor was an AT/RT, a tumor that predominantly affects infants and young children and has histological features similar to MRT in kidney, with poor prognosis. FISH and STR analysis of the recipient’s tumor was consistent with donor origin. Therefore, we consider donor-to-recipient malignancy transmission likely in this case, making this the first report of MRT in a renal allograft of an adult recipient following renal transplantation.Donor-to-recipient malignancy transmission could occur in two ways. First, there could be some metastatic cells in the transplanted kidney that come from a primary tumor in another organ and then transfer in the organ recipient. Second, post-transplant lymphoproliferative disorder, the most common post-transplant malignancy, could come from malignant change of passenger leukocytes in the transplanted organ. We favor the former explanation in this case, as it was likely that the tumor cells from AT/RT in the CNS of the pediatric donor metastasized to the kidney, which was transplanted to the adult female recipient.According to strategy and rules for organ screening and acknowledgment before transplantation, donors with a history of lung cancers, sarcomas and grade IV CNS neoplasms are considered unacceptable, but donors with a past with grade I-II CNS tumors are acceptable. In the present case, the donor, a 56-day-old infant who died of a CNS tumor with a suspected diagnosis of astrocytoma, was not reasonable because AT/RT should have been considered in the differential diagnosis before the transplantation.Since the number of patients requiring transplantation therapy is growing, there is increasing demand for donor organs. The expansion of the general donor pool has led to the inclusion of donors of extreme ages and donors who may possibly transmit disorder to the recipients. In any case, the peripheral donors’ organ utility is frequently connected with a more serious danger of undiagnosed disease including malignant tumors and some infections . Thus, tWe report a unique case of MRT in an adult renal allograft recipient after kidney transplantation with characteristic histologic features and loss of INI1 expression. This report highlights the importance of the criteria for selection of donors to screen possible malignant tumors transmission in living and dead donor transplants.


Coturnix coturnix) in the intact egg were exposed to hypoxia (~15% O2) during “early” , “middle” (D6-D10) or “late” (D11-D15) incubation or for their entire 16–18 day incubation (“continuous hypoxia”) to determine critical windows for viability and growth. Viability, body mass, beak and toe length, heart mass, and hematology (hematocrit and hemoglobin concentration) were measured on D5, D10, D15 and at hatching typically between D16 and D18 Viability rate was ~50–70% immediately following the exposure period in the early, middle and late hypoxic groups, but viability improved in the early and late groups once normoxia was restored. Middle hypoxia groups showed continuing low viability, suggesting a critical period from D6-D10 for embryo viability. The continuous hypoxia group experienced viability reaching <10% after D15. Hypoxia, especially during late and continuous hypoxia, also inhibited growth of body, beak and toe when measured at D15. Full recovery to normal body mass upon hatching occurred in all other groups except for continuous hypoxia. Contrary to previous avian studies, heart mass, hematocrit and hemoglobin concentration were not altered by any hypoxic incubation pattern. Although hypoxia can inhibit embryo viability and organ growth during most incubation periods, the greatest effects result from continuous or middle incubation hypoxic exposure. Hypoxic inhibition of growth can subsequently be “repaired” by catch-up growth if a final period of normoxic development is available. Collectively, these data indicate a critical developmental window for hypoxia susceptibility during the mid-embryonic period of development.Hypoxia during embryonic growth in embryos is frequently a powerful determinant of development, but at least in avian embryos the effects appear to show considerable intra- and inter-specific variation. We hypothesized that some of this variation may arise from different protocols that may or may not result in exposure during the embryo’s critical window for hypoxic effects. To test this hypothesis, quail embryos ( Normal morphological and physiological development of the avian embryo, as well as successful hatching, depends on appropriate temperature as well as ambient partial pressures of oxygen, carbon dioxide, and water vapor–for an entry into the voluminous, long-standing literature see ,2. HypoxMost of these studies on the effect of hypoxia on avian development have exposed embryos to chronic hypoxia throughout the entire incubation prior to measurements or during the last half of incubation—see  for discGallus gallus, as the model for studies of hypoxic effects on development has led to a considerable assemblage of data for this species. Whether the response of the chicken embryo is, in fact, actually broadly representative of all developing birds is unknown–though this is generally assumed in the absence of data to the contrary. Yet, experiments on, for example, cardiovascular development in birds suggest that there are both profound quantitative and qualitative differences in chickens compared with emus ) was determined on another subsample of 30 μl of blood injected into a Radiometer OSM2 Hemoximeter. Duplicate measurements were made and averaged for both hemoglobin and hematocrit.Assessment for normality of distributions  and equality of variances was followed by within- and between-group testing for statistical significance with parametric ANOVAs and Student's t-test (SPSS software). Holmes-Sidak viability data were assessed with a Kaplan-Meier logrank test (SigmaPlot software). All statistical decisions were made with a 0.05 level of significance, and are provided in A total of 154 eggs were used in these experiments, with individual n values for each incubation condition provided in All embryos survived all treatments through D5. Viability of the normoxic (control) group of quail embryos was ~75% and ~60% on D10 and D15, respectively, with approximately 55% of the originally incubated eggs successfully hatching . The earSpecific statistical comparisons of viability data are provided in the inset of Morphologically, quail embryos in the early, middle, and late hypoxic treatment groups appeared identical to the normoxia group on all three measurement days. However, the survivors of the continuous hypoxia treatment group displayed many abnormal physiological and anatomical characteristics. At both D10 and 15, the overall appearance of the embryo was edematous, especially in the cephalic region. In addition to this profound ascites, there were also notable beak and eye deformations in the continuous hypoxic group. Deformities included incomplete or absent lower or upper beak or eye. Eyes were open by D10 in some cases, as in the other groups, but remained closed at D15 in some of the embryos from continuous hypoxia. The full length of the wing was attached to the embryonic body in some continuous hypoxia embryos, and feather development was delayed up until D15 in the continuous hypoxic group. The third toe was attached to the adjacent toes in some cases.Mean wet and dry body mass of control embryos increased significantly as a function of development , as anticipated , Fig 3. On Day 10 wet body mass of all groups that experienced hypoxia at some point during their development  were not statistically different from each other. HoweverDespite having a lower body mass at D15, quail embryos exposed to early, middle or late hypoxia exhibited “catch-up” growth such that there were no significant differences between these groups and the control group at hatching.The relationships described above for wet mass generally also held for dry mass , Fig 3B,At D10, beak length averaged 2.2 ± 0.1 mm and toe lengths averaged 6.5 ± 0.2 mm , Fig 4. Similar absolute sizes and rates of growth in toe and beak length occurred in the early, middle and late hypoxic groups on D10 , Table 2At D10, the average wet heart mass of normoxic embryos was about 33 mg, while dry heart mass was ~3.6 mg, yielding a heart water content of 89% . The dryWet and dry heart mass increased slightly between D10 and D15 and then to a greater extent at hatching in all hypoxic incubation groups, as a consequence of overall developmental progression , Table 3Hematocrit in all groups of quail embryos was 22–23% at D10, increasing to 25–29% at hatching . HemogloGrowth during normoxic incubation was relatively modest through the first half of incubation, thereafter increasing more rapidly, with more than 3/4 of body mass accumulation occurring in the last 50% of incubation. This aligns with the embryos of other galliform birds –60, inclEmbryonic mortality in quail embryos was greatly influenced by patterns of hypoxic exposure during incubation. Early (D0-D5) hypoxic exposure had little effect on survivability in any treatment group, and eggs hatched at approximately the same rate as the normoxic controls . More in2) affect weight at hatching but otherwise have little morphological effect in chicken embryos, whereas the aerobic energy shortfall associated with severe hypoxia (10% O2) actually affects embryonic viability [Not only was lack of viability progressive and highest in the continuous hypoxic group , but numiability .Hypoxic incubation in chicken embryos typically indicates, as for the present study on quail, that continuous hypoxia, even relatively mild, is among the most disruptive of hypoxic exposure protocols ,25,58. LIn the case of early hypoxic exposure, the early growth restriction induced by hypoxia can be reversed by the restoration of normoxia in incubation, so-called “catch-up growth” ,64. In tIt is important to note that despite early incubation being the most sensitive in chicken embryos regarding viability (see above), the metabolic responses to early, middle and late hypoxic exposure occurs in the chicken embryo appear to align more closely with the viability data for quail from the present study. Thus, hypoxic exposure in the middle of incubation in chickens leads to the most profound and lasting adjustments to oxygen consumption . Apparen2) when measured in middle of incubation in chicken embryos, but additional incubation in normoxia enabled catch-up growth [2 [Reflecting the greatly reduced ultimate hatching success, continuous hypoxic exposure had the most debilitating effect on beak and toe development, with inhibited growth already evident by D10 and even more pronounced by D15, after which low viability occurred . While np growth . Howeverrowth [2 .Heart mass (wet or dry) in quail, unlike beak and toe length, was unaffected by any combination of hypoxic exposure, including continuous exposure, an example of “organ sparing” also evident for the hearts of chicken embryos exposed to hypoxia or malnutrition ,26. It hInterestingly, the ratio of wet heart mass to body mass in the present study on quail showed some significant changes with hypoxia incubation. Most striking was the larger heart mass to body mass ratio evident in the continuous hypoxic group on D15  comparedDespite protocols that included intervals of 5 day hypoxic incubation as well as continuous hypoxic incubation, no significant changes in hematology were measured in any of the quail embryo groups. During early development chicken embryos are similarly unable to counter hypoxic incubation by mounting an increased production of red blood cells and/or the expansion of total blood volume , despite2 affinity is produced earlier in development, and is replaced gradually by hemoglobin with lower affinity [2 transport in quail embryos could also be enhanced by changes in blood O2 affinity due to alterations in red blood cell organophosphates such as ATP, as has been documented in chicken and turkey embryos [2 affinity in chicken embryos exposed to hypoxia [2 carrying characteristics of quail blood were temporarily or permanently modified by hypoxic incubation.Though an adaptive polycythemic response is lacking, potentially reduced oxygen transport during hypoxic incubation in quail embryos could be offset by production of hemoglobin(s) with a greater affinity for oxygen, as occurs in other birds ,81–84. Caffinity ,79,85. B embryos ,86. Decr hypoxia ,87,88. FGallus gallus) is indisputably the main avian model for developmental studies–for recent reviews see [The domestic chicken  and non-domesticated wild birds with respect to the reduction of metabolism, hatchability, growth rate, and hatching mass upon exposure to moderate hypoxia related to altitude [Dromiceius novaehollandiae) in the pattern of onset of key cardiovascular control mechanism–see [Coturnix coturnix) reports additional key differences in responses compared to chicken embryos, in viability rates during normal development, and also in the primary period of vulnerability  for growth and morphology during incubation.The use of model organisms (or organisms assumed to be like another model organism) has greatly advanced our understanding of development within every area of biology. However, an unduly narrow focus on models to the exclusion of exploring species diversity can also lead to a distorted perception of what is “representative” ,95. For altitude . Similarnism–see ,38,96. AContinuous hypoxia is clearly not well tolerated in quail embryos, leading to high rates of mortality. Yet, embryos experiencing a mixture of hypoxia and normoxia at various times during incubation were capable of sufficient self-repair and/or catch-up growth over time so as to appear morphologically identical at hatching, even though they followed significantly different developmental trajectories in arriving at this common phenotype.Collectively, these findings for avian embryonic development suggests that avian embryos exposed to hypoxia during incubation at various stages in development are pushed into abnormal developmental trajectories, but once hypoxic incubation is replaced by normoxic incubation, development of surviving embryos not only progresses, but follows a novel developmental trajectory leading back towards the normal hatchling phenotype for quail. Though the developmental trajectory is not a normal one, it may provide the additional time to repair earlier hypoxia-induced damage.Future experiments should be directed at determining dose-response relationships, which will reveal potentially differential susceptibilities to hypoxia by different tissues and organ systems. Additionally, such studies may additionally reveal “soft edges” to the critical windows that are not readily revealed by a simple two dimensional approach to susceptible periods ,97. A moS1 File(DOCX)Click here for additional data file.S2 File(DOCX)Click here for additional data file.S3 File(DOCX)Click here for additional data file.S4 File(DOCX)Click here for additional data file.S5 File(DOCX)Click here for additional data file.


The ultimate test of whether two diverged populations are, in fact, good species is whether they will maintain their distinctness in sympatry , 2. ThisMimulus has long served as a model for the study of reproductive isolation [Mimulus guttatus and M. nasutus. These recently diverged species have broadly overlapping geographic ranges, and—when sympatric—M. nasutus’ ancestry frequently introgresses into M. guttatus [M. nasutus × M. guttatus hybrids.The genus solation , 6. Muchguttatus –9. Becauguttatus  map the Mimulus  and hl14 (Migut.O00467). These genes are duplicates of plastid transcriptionally active chromosome 14 (pTAC14)―a gene known to be essential for proper chloroplast development in Arabidopsis thaliana [pTAC14 is found on chromosome 14 (hl14) of both the M. guttatus reference genome and the M. guttatus studied samples, but the gene is missing from M. nasutus’ chromosome 14. In contrast, both species have a syntenic copy of pTAC14 (hl13) on chromosome 13, but the M. guttatus allele includes a frameshift mutation and is not expressed. As a result, some F2 hybrids and advanced backcrossed seedlings will be homozygous for the nonfunctional M. guttatus’ copy of pTAC14 at hl13 and M. nasutus’ null allele on chromosome 14; these individuals will lack a functional pTAC14 gene, leaving them unable to photosynthesize and therefore inviable  and Cape Verde Island (Cvi)—the histidinol-phosphate aminotransferasegene exists on chromosome 1 but not chromosome 5 in Cvi, and on chromosome 5 but not chromosome 1 in Col, and F2s lacking histidinol-phosphate aminotransferasegene are inviable [M. guttatus and M. nasutus, none of these species pairs hybridize in nature.Previous studies also have lent support for this model. In the id males . A similinviable . Unlike M. nasutus’ ancestry is elevated around hl14 and depleted around hl13 in sympatric M. guttatus, as would be expected if incompatible alleles are selected against upon introgression [hl13/hl14 acts in concert with the other reproductive isolating mechanisms and mapped incompatibilities  to prevpTAC14 and nonfunctionalization are plausibly neutral, suggesting that reproductive isolation may have arisen by a neutral process. In fact, Zuellig and Sweigart find no strong evidence that natural selection is responsible for either pTAC14‘s duplication or degeneration, and as such the authors suggest that this incompatibility has a neutral origin. While this hypothesis is plausible, it clearly needs more scrutiny. Further study of the effects of the alternative functional copies, and additional population genomic studies of the history of selection on hl13 and hl14, could inform the evolutionary question of how and why genes relocate.Second, both the process of duplication of M. guttatus mean that the hl13/hl14 incompatibility provides an excellent opportunity to identify the factors shaping the alternative resolution of alternative paralogs. This question is particularly interesting because of the evolutionary history of the species’ split; the M. guttatus population studied by Zuellig and Sweigart is more closely related to M. nasutus than it is to the M. guttatus reference genome with which it shares hl13 and hl14 alleles [Finally, the recency of the split between these species and the extensive natural variation in  alleles . This ob alleles .Overall, Zuellig and Sweigart’s results provide empirical evidence that the evolution of gene duplicates might be involved in reproductive isolation in young species that hybridize in nature. Systematic efforts like Zuellig and Sweigart’s will reveal to what extent gene movement is a prevalent force in the formation of species. More generally, their study provides a map route to understand what is the role of hybrid incompatibilities in keeping naturally co-occurring―and potentially hybridizing―species apart.


Metal accumulation in sediments threatens adjacent ecosystems due to the potential of metal mobilization and the subsequent uptake into food webs. Here, contents of heavy metals  and trace elements  were determined for river waters and bed sediments that received sewage discharged from traditional and semiconductor industries. We used principal component analysis (PCA) to determine the metal distribution in relation to environmental factors such as pH, EC, and organic matter (OM) contents in the river basin. While water PCA categorized discharged metals into three groups that implied potential origins of contamination, sediment PCA only indicated a correlation between metal accumulation and OM contents. Such discrepancy in metal distribution between river water and bed sediment highlighted the significance of physical-chemical properties of sediment, especially OM, in metal retention. Moreover, we used Se XANES as an example to test the species transformation during metal transportation from effluent outlets to bed sediments and found a portion of Se inventory shifted from less soluble elemental Se to the high soluble and toxic selenite and selenate. The consideration of environmental factors is required to develop pollution managements and assess environmental risks for bed sediments. Heavy metal distribution has been one of the critical concerns in natural environments due to their toxicity and biomagnification attributes. Lots of regulations have been established to avoid heavy metal concentrations in waters, sediments, and soils exceed quality criteria for environmental protection. Anthropogenic activities have discharged significant amounts of heavy metals into rivers. By accumulated in sediments, heavy metals could enter the food chain as bottom sediments in river basin or reservoirs serve as habitats and food sources for benthic fauna. Subsequently, heavy metals may directly or indirectly threaten the aquatic flora and fauna. Thereby, metal contamination in sediments has been an ongoing issue worldwide2According to chemical and geological conditions, heavy metals discharged into surface waters were rapidly partitioned onto particulate matter and incorporated in sediments. Based on the matrix composition, sediments generally show a superior capacity for metal retention6Our study site locates at the basin of the Fazih and the Wu River in Taichung, Taiwan . This ba6891011Here, we aimed to determine how environmental factors such as pH, electrical conductivity (EC), oxidation-reduction potential (ORP), OM, amorphous  aluminum (Al) and (Fe) dominate the accumulation of heavy metals and trace elements in bed sediments. Data sets on environmental factors and metal contents in river water and bed sediments were subjected to principal component analysis (PCA) to ascertain factors that were responsible for the variation in metal loading in sediments. In particular, Se speciation was performed as an example to examine the species transformation in relation to environmental changes by X-ray absorption near edge structure (XANES) analysis during the transportation from effluent outlet to bed sediments. Recognition of the metal enrichment factors could lead to a better estimation of the dynamics and mass balance of heavy metals and trace elements in contaminated sediments and thereby an improved strategy for wastewater discharge management.2 but not reduced SO44. For the EC, the sewage effluent samples (S1–S5) generally showed the relatively higher EC than that of the Fazih River (F1–F6) and most of the Wu River (W1–W4) samples . The enhancement in EC for the W6 sample might be also caused by the seawater intrusion as the sampling site of W6 locates close to the river mough.Our study sites locate at the basin of the Fazih and the Wu River in Taichung, Taiwan . This 26 samples . The mea−1, that for W2, W4, W5, and W6 sediments were less than 10 mg kg−1.Regarding chemical parameters of solid phases, we determined the contents of TOC and amorphous Fe and Al in sediments. For the amorphous  Fe (0.71 to 1.71%) and Al (0.17 to 0.38%), no particular pattern could be identified according to the sampling geographical locations . However−1), Cu (116.35 μg L−1), Ni (237.60 μg L−1), Pb (0.09 μg L−1), Zn (98.01 μg L−1), Ga (1.81 μg L−1), In (4.56 μg L−1), Mo (226.64 μg L−1), and Se (1.09 μg L−1) was found in the site of S3 or S5, the highest mean amount of Cr (17.89 μg L−1) was found in the W1 site. Given that S3 and S5 were sewage effluents discharged mainly from the traditional and semiconductor industries, it is not surprising to observe substantial amounts of heavy metals in these samples. However, the occurrence of Cr in the W1, which exceeded the CCC, implying a peculiar source to Cr discharges.The maximum, mean, median, and minimum concentration of heavy metals  and trace elements  in sediments were summarized in 153/s) is less than that in the Wu River (114.43 m3/s), resulting in a relatively limited mobilization of suspended particles and thereby a net deposition and accumulation of particulate metals in the Fazih RiverGenerally, sediments in the Fazih River seemed contain relatively higher amounts of heavy metals and trace elements than that in the Wu River. Plausible explanations include: (1) the Fazih River has contained the discharged sewage effluent from hospitals (S1 and S1) and traditional industrial activities . Similar trend was found for the Zn distribution. Only F2, F3, and F6 sites were exceeded in mean and median concentrations of Zn.Although distribution of heavy metals and trace elements varied essentially among individual sites along the Fazih and the Wu River, the contamination of Cr and Zn is generally critical while compared with the PEL. For Cr, the highest level of 357.35 mg kg−1 20−1 is more robust due to the more comprehensive computation for Se aquatic hazard assessments−1 in the F3 sample.Given that the PEL for Ni, Ga, In, Mo, and Se has not been established yetRI) is defined as Equation (1).Given that our study basin received pollution from multiple heavy metals, we used the quantitative approach developed by HakansonTi is the toxic-response factor for a given substance 23Ci is the metal concentration in sediments; C0 is the regional background concentration of heavy metals, wherein we used the metal concentration obtained from the W2 site as the background because W2 received relatively less industrial and domestic sewage.where RI of heavy metals in sediment samples were categorized into three zones as shown in RI valueThe RI values for the Wu River sediments were generally lower than 300. However, four out of six sediment samples in the Fazih River showed the RI values ranging from 300 to 600, suggesting a considerable ecological risk in this basin. Given that the toxic response factor for trace elements such as Ga, In, Mo, and Se have not been developed well yet, the potential ecological risk of such trace elements were excluded here.In our studied basin, the nd component that accounted for 15.4% of variance, metals of Pb, Ni, and Ga were grouped into the environmental factor of EC. However, the other environmental factors of pH and ORP did not correlate with any metals (the 3rd component with 12.0% of variance). Most noteworthily, Cr was isolated from all other metals and environmental factors (the 4th component with 8.2% of variance). Such results implied that discharged heavy metals and trace elements could be categorized into three groups: (1) Cd, In, Mo, Cu, Se, Zn; (2) Pb, Ni, Ga; (3) Cr. In addition, the environmental factors of pH and ORP showed no correlation with the metal effluent. Only EC might influence the distribution of Pb, Ni, and Ga in river water.The PCA has been applied to determine the correlation between the partition of discharged metals and environmental factors such as pH, EC, ORP, TOC, and amorphous Fe and Al782526For sediment samples, we added the contents of three more environmental factors including TOC, oxalate-extractable Fe and Al of individual sediment samples to process the PCA. According to By coupling the PCA results of water and sediment samples, the noteworthy finding for the metal distribution in our study river basin was that the correlation between discharged metals suggested by the PCA of water samples vanished in sediment samples. The disagreement in metal correlation between water and sediment samples is of interest to us in terms of (1) the rationale to use the metal contents in sediments for the identification of pollution sources672-OH), and carbonyl (R-C = O-R-R/H/OR)a) for carboxyl and phenolic groups ranges from 3–5 and 5–7, surfaces of natural OM are generally covered with negative charges while pH > 33435(s)) or oxyanions (HSeO3− and MnO42−) under the environmental conditions of our study sites  or hydrolyzed species (Cr(OH)2+, Pb(OH)+, etc.)29303839et al.et al.Widespread evidences have existed for the incorporation and retention of metals into OM and Fe-(hydr)oxides in soils and bed sediments525282930dy sites , other mPrior to the retention onto sediments, metals discharged from sewage effluents might transport in the form of soluble species in solutions and/or insoluble species in suspended solids. Taken into account that the metal concentrations in suspended particles were generally higher than that in bed sediments and/or even exceeded the PEL guidelines for the protection of aquatic life−1, and that for the Wu River sediments (W4 and W5) were lower than 1 mg kg−1. However, in suspended solids (S5), the Se content was up to 65.2 mg kg−1, which was 20 to 60 times higher than that of sediments in the river basin. XANES data in Selenium speciation in selected suspended solids and sediments was performed using XANES analysis. Tested suspended solids and sediments were collected in January, 2015. Selenium concentrations for the Fazih River sediments  ranged from 2.17 to 3.18 mg kg0, selenomethionine, selenocystine, trimethylselenonium, selenite, and selenate. Results of LCF shown as solid lines in 0 contributed about 38% of the total Se besides the FeSe. However, in the samples of F3, F6, and W4, not only inorganic FeSe but organic Se of trimethylselenonium was found. The most concerned point is that Se oxyanions (selenite and selenate) was observed at the W5 site, wherein the sediment received a massive amount of Se from the suspended solids of S5. Selenite and selenate are highly soluble, hence more mobile than the less soluble forms such as selenide and elemental Se0 0The presence and proportion of Se species in suspended solids and sediments were determined using LCF with spectra obtained from reference materials including iron selenide, elemental Se0 in suspended solids collected from sewage effluents. However, selenite and selenate was found in the sediment received a massive amount of Se from suspended solids. This is of environmental concern as selenite and selenate could bioaccumulate in organisms and thus incorporate into food webs more readily than selenide and elemental Se0.One of the major environmental impacts for the metal accumulation in sediments was the metal uptake by primary producers and the subsequent access into the food webs of aquatic ecosystems. In this study, the distribution and accumulation of heavy metals and trace elements in relational to environmental factors in the basin of the Fazih and the Wu River that received sewage effluents from traditional and semiconductor industrial activities was determined by PCA. The PCA results pointed out the discrepancy in metal distributions between water and sediment samples. Although PCA for water samples proposed three potential origins of contamination for heavy metals and trace elements, such metal distribution was not observed while metals accumulated in sediments. Among all tested environmental factors, the metal distribution in sediments only correlated to OM contents. In spite of other environmental factors such as pH and ORP did not directly influenced the metal distribution in sediments, such physical-chemical properties in environments may control the species transformation for metals and further change their mobility, bioavailability, and toxicity. Our XANES results indicated that Se inventory mainly comprised selenide and elemental SeCollectively, our results clearly suggest that OM must be taken into account to assess the metal behavior in sediments. Recognition of the roles of environmental factors in metal distribution could lead to a better quantification in dynamics and mass balances for metals in contaminated ecosystems. Given that bed sediments serve both as a sink and source of metals, the prediction of relative abundance of metals in sediments could improve strategies for the management and remediation of metal contamination. Conclusively, the consideration of environmental factors, especially OM, is required while monitoring metal accumulation in sediments. Such comprehensive information can help to develop an adequate protection and restoration plan.The water and sediment samples were collected from 12 sampling sites scattered in the Fazih (F1–F6) and the Wu (W1–W6) River . Sewage 3 background, and preserved in acid-washed polyethylene bottles at 4 °C. For sediments, approximately 100 g of solids were collected using a plastic scoop from the top 2 cm of bottom sediments and preserved in acid-washed polyethylene containers at 4 °C. Sediment samples were air-dried inside a laminar flow chamber, passed through a 2-mm nylon sieve, and ground in an agate mortar. For elemental analysis, 1 g of pretreated sediment was acid digested using 10 mL of aqua regia at 180 °C for 4 h. Metal concentrations in acid digestates were determined by inductively coupled plasma mass spectrometry  with methane as reaction gas to reduce interatomic interferences. A soil standard reference material  was also digested to check the analysis recovery. Three replicates were performed in acid digestion for each sample including blank and SRM for quality control. The recoveries of the SRM ranged from 80 to 120% in all metals. Contents of OM in sediments were determined by amounts total organic carbon (TOC), which was determined using the TOC analyzer . Sediment samples for TOC analysis were pretreated using 10% phosphoric acid overnight to reduce the interference from inorganic carbon49Water samples were collected at river surfaces or from sewage effluents. The pH, EC, and ORP data of water samples were determined using a multi-parameter water quality checker Horiba U-50) immediately after the collection. Once in the laboratory, the other portion of water samples was filtered through a 0.2 μm membrane, acidified to a 2% HNO immediat®) for the Windows. To interpret the results more precisely, the rotation method - Varimax with Kaiser Normalization - was applied to maximize variances of the squared normalized factor loadings across variables for each factor. All principal factors extracted from variables were those whose eigenvalues is higher than 1Principal component analysis (PCA) has been often used to interpret the multivariate, complex, redundant, and not direct data52530, selenomethionine, selenocystine, trimethylselenonium, selenite (H2SeO3), and selenate (H2SeO4). See Liu, et al.Selenium speciation in selected sediments and suspended solids was determined using Se K-edge XANES analysis. Suspended solids were collected by filtering the sewage effluent at the S5 site thought a mixed cellulose esters membrane with a calibrated porosity of 0.2 μm (Millipore). The filter membrane was placed on a standing stainless steel filter holder  and the filtration was operated under an air pressure of 65 psi. All samples for Se-XANES analysis were prepared by mounting approximately 0.2 g solids in acrylic sample holders, sealing with Kapton tape to avoid desiccation. Spectra were acquired at the Beamline BL17C1 at the National Synchrotron Radiation Research Center (NSRRC), Hsinchu, Taiwan. The linear combination fitting (LCF) of Se-XANES data across from 30 eV below to 40 eV above the Se absorption edge was used to determine the Se species. Reference materials used in LCF included iron selenide (FeSe), elemental (gray) SeHow to cite this article: Hsu, L.-C. et al. Accumulation of heavy metals and trace elements in fluvial sediments received effluents from traditional and semiconductor industries. Sci. Rep.6, 34250; doi: 10.1038/srep34250 (2016).