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Magnetic ratcheting cytometry is a promising approach to separate magnetically-labeled cells and magnetic particles based on the quantity of magnetic material. We have previously reported on the ability of this technique to separate magnetically-labeled cells. Here, with a new chip design, containing high aspect ratio permalloy micropillar arrays, we demonstrate the ability of this technique to rapidly concentrate and collect superparamagnetic iron oxide particles. The platform consists of a mechatronic wheel used to generate and control a cycling external magnetic field that impinges on a "ratcheting chip." The ratcheting chip is created by electroplating a 2D array of high aspect ratio permalloy micropillars onto a glass slide, which is embedded in a thin polymer layer to create a planar surface above the micropillars. By varying magnetic field frequency and direction through wheel rotation rate and angle, we direct particle movement on chip. We explore the operating conditions for this system, identifying the effects of varying ratcheting frequency, along with time, on the dynamics and resulting concentration of these magnetic particles. We also demonstrate the ability of the system to rapidly direct the movement of superparamagnetic iron oxide particles of varying sizes. Using this technique, 2.8 μm, 500 nm, and 100 nm diameter superparamagnetic iron oxide particles, suspended within an aqueous fluid, were concentrated. We further define the ability of the system to concentrate 2.8 μm superparamagnetic iron oxide particles, present in a liquid suspension, into a small chip surface area footprint, achieving a 100-fold surface area concentration, and achieving a concentration factor greater than 200%. The achieved concentration factor of greater than 200% could be greatly increased by reducing the amount of liquid extracted at the chip outlet, which would increase the ability of achieving highly sensitive downstream analytical techniques. Magnetic ratcheting-based enrichment may be useful in isolating and concentrating subsets of magnetically-labeled cells for diagnostic automation.

Magnetic ratcheting cytometry is a promising approach to separate magnetically-labeled cells and magnetic particles based on the quantity of magnetic material. We have previously reported on the ability of this technique to separate magnetically-labeled cells. Here, with a new chip design, containing high aspect ratio permalloy micropillar arrays, we demonstrate the ability of this technique to rapidly concentrate and collect superparamagnetic iron oxide particles. The platform consists of a mechatronic wheel used to generate and control a cycling external magnetic field that impinges on a "ratcheting chip." The ratcheting chip is created by electroplating a 2D array of high aspect ratio permalloy micropillars onto a glass slide, which is embedded in a thin polymer layer to create a planar surface above the micropillars. By varying magnetic field frequency and direction through wheel rotation rate and angle, we direct particle movement on chip. We explore the operating conditions for this system, identifying the effects of varying ratcheting frequency, along with time, on the dynamics and resulting concentration of these magnetic particles. We also demonstrate the ability of the system to rapidly direct the movement of superparamagnetic iron oxide particles of varying sizes. Using this technique, 2.8 μm, 500 nm, and 100 nm diameter superparamagnetic iron oxide particles, suspended within an aqueous fluid, were concentrated. We further define the ability of the system to concentrate 2.8 μm superparamagnetic iron oxide particles, present in a liquid suspension, into a small chip surface area footprint, achieving a 100-fold surface area concentration, and achieving a concentration factor greater than 200%. The achieved concentration factor of greater than 200% could be greatly increased by reducing the amount of liquid extracted at the chip outlet, which would increase the ability of achieving highly sensitive downstream analytical techniques. Magnetic ratcheting-based enrichment may be useful in isolating and concentrating subsets of magnetically-labeled cells for diagnostic automation.

A shift in the Neonatal Resuscitation Program (NRP) guidelines occurred in 2015 from routine intubation and endotracheal suctioning of all meconium-stained non-vigorous infants towards less aggressive interventions based on response to initial resuscitation. This study aims to examine the impact of this change on outcomes of non-vigorous infants born through meconium-stained amniotic fluid at a level III academic NICU encompassing years before and after the change in guideline. This single-center retrospective study compared NICU therapies and clinical outcomes of 117 non-vigorous newborns pre-guideline implementation to 106 non-vigorous newborns post-guideline implementation. Nearly two thirds of infants in the pre-guideline cohort received endotracheal suctioning with recovery of meconium compared to less than a third of infants in the post-guideline cohort (p

BackgroundThe evidence for an increased incidence of sexually transmitted infections (STIs) among patients utilizing HIV pre-exposure prophylaxis (PrEP) has been inconsistent. We assessed the risk of incident STI while on PrEP compared to periods off PrEP among military service members starting PrEP.MethodsIncidence rates of chlamydia, gonorrhea, syphilis, hepatitis C virus, and HIV were determined among military service members without HIV prescribed daily oral tenofovir disoproxil fumarate and emtricitabine for HIV PrEP from February 1, 2014 through June 10, 2016. Hazard ratios for incident STIs were calculated using an Anderson-Gill recurrent event proportional hazard regression model.ResultsAmong 755 male service members, 477 (63%) were diagnosed with incident STIs (overall incidence 21.4 per 100 person-years). Male service members had a significantly lower risk of any STIs (adjusted hazard ratio (aHR) 0.21, 95% CI 0.11-0.40) while using PrEP compared to periods off PrEP after adjustment for socio-demographic characteristics, reasons for initiating PrEP, surveillance period prior to PrEP initiation, and the effect of PrEP on site and type of infection in multivariate analysis. However, when stratifying for anatomical site and type of infection, the risk of extragenital gonorrhea infection (pharyngeal NG: aHR 1.84, 95% CI 0.82-4.13, p = 0.30; rectal NG: aHR 1.23, 95% CI 0.60-2.51, p = 1.00) and extragenital CT infection (pharyngeal CT: aHR 2.30, 95% CI 0.46-11.46, p = 0.81; rectal CT: aHR 1.36, 95% CI 0.81-2.31, p = 0.66) was greater on PrEP compared to off PrEP although these values did not reach statistical significance.ConclusionsThe data suggest entry into PrEP care reduced the overall risk of STIs following adjustment for anatomical site of STI and treatment. Service members engaged in PrEP services also receive more STI prevention counseling, which might contribute to decreases in STI risk while on PrEP.

IntroductionWe performed a longitudinal SARS-CoV-2 seroepidemiological study in healthcare personnel of the two largest tertiary COVID-19 referral hospitals in Mexico City.MethodsAll healthcare personnel, including staff physicians, physicians in training, nurses, laboratory technicians, researchers, students, housekeeping, maintenance, security, and administrative staff were invited to voluntarily participate, after written informed consent. Participants answered a computer-assisted self-administered interview and donated blood samples for antibody testing every three weeks from October 2020 to June 2021.ResultsA total of 883 participants (out of 3639 registered employees) contributed with at least one blood sample. The median age was 36 years (interquartile range: 28-46) and 70% were women. The most common occupations were nurse (28%), physician (24%), and administrative staff (22%). Two hundred and ninety participants (32.8%) had a positive-test result in any of the visits, yielding an overall adjusted prevalence of 33.5% for the whole study-period. Two hundred and thirty-five positive tests were identified at the baseline visit (prevalent cases), the remaining 55 positive tests were incident cases. Prevalent cases showed associations with both occupational (institution 2 vs. 1: adjusted odds ratio [aOR] = 2.24, 95% confidence interval [CI]: 1.54-3.25; laboratory technician vs. physician: aOR = 4.38, 95% CI: 1.75-10.93) and community (municipality of residence Xochimilco vs. Tlalpan: aOR = 2.03, 95% CI: 1.09-3.79) risk-factors. The incidence rate was 3.0 cases per 100 person-months. Incident cases were associated with community-acquired risk, due to contact with suspect/confirmed COVID-19 cases (HR = 2.45, 95% CI: 1.21-5.00).ConclusionsWe observed that between October 2020 and June 2021, healthcare workers of the two largest tertiary COVID-19 referral centers in Mexico City had similar level of exposure to SARS-CoV-2 than the general population. Most variables associated with exposure in this setting pointed toward community rather than occupational risk. Our observations are consistent with successful occupational medicine programs for SARS-CoV-2 infection control in the participating institutions but suggest the need to strengthen mitigation strategies in the community.

Photoconvertible proteins are powerful tools widely used in cellular biology to study cell dynamics and organelles. Over the past decade, photoconvertible proteins have also been used for developmental biology applications to analyze cell lineage and cell fate during embryonic development. One of these photoconvertible proteins called Kaede, from the stony coral Trachyphyllia geoffroyi, undergoes irreversible photoconversion from green to red fluorescence when illuminated with UV light. Undertaking a cell tracing approach using photoconvertible proteins can be challenging when using unconventional animal models. In this protocol, we describe the use of Kaede to track specific cells during embryogenesis of the cephalochordate Branchiostoma lanceolatum. This protocol can be adapted to other unconventional models, especially marine animals.

The cephalochordates amphioxus or lancelets are benthic marine animals representing the earliest divergent evolutionary lineage within chordates. Although amphioxus are present in most of the world's tropical and temperate oceans, only about thirty different species grouped into three different genera, Branchiostoma, Epigonichthys and Asymmetron have been described. In the genus Asymmetron, only two species have been characterized, although for one of them, A. lucayanum, several cryptic lineages exist. In this work we have sequenced and analyzed the mitogenome of an A. lucayanum population previously described in the Red Sea. The phylogenetic study using this complete mitogenome as well as the analysis of COI gene sequences of several individuals of this Red Sea population show that the Red Sea population is a new cryptic species. We propose to call this new species Asymmetron rubrum.

Holothuria leucospilota (Echinodermata: Holothuroidea) is a widespread tropical sea cucumber with strong value for the ecological restoration of coral reefs. Therefore, some studies regarding the artificial breeding and cultivation of H. leucospilota have been undertaken recently. However, the biological functions of the digestive system of this species have not been elucidated. In this study, a cDNA coding for α-amylase, an indicator of digestive maturity in animals, was identified from H. leucospilota and designated Hl-Amy. The full-length cDNA of the Hl-Amy gene, which is 1734 bp in length with an open reading frame (ORF) of 1578 bp, encodes a 525 amino acid (a.a.) protein with a deduced molecular weight of 59.34 kDa. According to the CaZy database annotation, Hl-Amy belongs to the class of GH-H with the official nomenclature of α-amylase (EC 3.2.1.1) or 4-α-D-glucan glucanohydrolase. The Hl-Amy protein contains a signal peptide at the N-terminal followed by a functional amylase domain, which includes the catalytic activity site. The mRNA expression of Hl-Amy was abundantly exhibited in the intestine, followed by the transverse vessel with a low level, but was hardly detected in other selected tissues. During embryonic and larval development, Hl-Amy was constitutively expressed in all stages, and the highest expression level was observed in the blastula. By in situ hybridization (ISH), positive Hl-Amy signals were observed in different parts of the three different intestinal segments (foregut, midgut and hindgut). The Hl-Amy recombinant protein was generated in an E. coli system with codon optimization, which is necessary for Hl-Amy successfully expressed in this heterogenous system. The Hl-Amy recombinant protein was purified by immobilized metal ion affinity chromatography (IMAC), and its activity of starch hydrolysis was further detected. The optimal temperatures and pH for Hl-Amy recombinant protein were 55°C and 6.0, respectively, with an activity of 62.2 U/mg. In summary, this current study has filled a knowledge gap on the biological function and expression profiles of an essential digestive enzyme in sea cucumber, which may encourage future investigation toward rationalized diets for H. leucospilota in artificial cultivation, and optimized heterogenous prokaryotic systems for producing recombinant enzymes of marine origins.

BACKGROUND:Cellulitis is a frequent cause of hospital admission of adult patients. Increasing prevalence of multiresistant microorganisms, comorbidities, predisposing factors and medical and surgical therapies might affect cellulitis response and recurrence rate. METHODS:Prospective and observational study of 606 adult patients with cellulitis admitted to several Spanish hospitals. Comorbidities, microbiological, clinical, diagnostic, treatment (surgical and antibiotic) data were analyzed according to the cellulitis response. Good response implied cure. Poor response implied failure to cure or initial cure but relapse within 30 days of hospital discharge. RESULTS:Mean age was 63.3 years and 51.8% were men. Poor responses were significantly associated with age, previous episodes of cellulitis, prior wounds and skin lesions, venous insufficiency, lymphedema, immunosuppression and lower limbs involvement. No differences in ESR or CRP blood levels, leukocyte counts, pus or blood cultures positivity or microbiological or imaging aspects were observed in those with good or poor responses. Regarding antimicrobials, no differences in previous exposition before hospital admission, treatment with single or more than one antibiotic, antibiotic switch, days on antimicrobials or surgical treatment were observed regarding good or poor cellulitis response. Prior episodes of cellulitis (P = 0.0001), venous insufficiency (P = 0.004), immunosuppression (P = 0.03), and development of sepsis (P = 0.05) were associated with poor treatment responses, and non-surgical trauma (P = 0.015) with good responses, in the multivariate analysis. CONCLUSIONS:Prior episodes of cellulitis, non-surgical trauma, venous insufficiency, sepsis and immunosuppression were independently associated with treatment response to cellulitis, but not the causative microorganism, the number of antimicrobials administered or its duration.

We sought to examine sex-related differences in the characteristics and outcome in patients presenting with acute symptomatic pulmonary embolism (PE).We conducted a retrospective cohort study of 2,096 patients diagnosed with acute PE. The characteristics were recorded at presentation. Treatment was at the discretion of patients' physicians. The primary study outcome, all-cause mortality, and the secondary outcomes of PE-specific mortality, recurrent venous thromboembolism, and major bleeding were assessed during the first month of follow-up after PE diagnosis.Overall, the women were older than the men and had significantly higher rates of immobilization. They had significantly lower rates of chronic obstructive pulmonary disease and cancer. Women had a higher prevalence of syncope and elevated brain natriuretic peptide levels. Thirty-day all-cause mortality was similar between women and men (7.1% versus 6.2%; P = 0.38). Male gender was not independently significantly associated with PE-related death (adjusted odds ratio [OR] 1.02; 95% CI, 0.50 to 2.07; P = 0.96). Restricting the analyses to haemodynamically stable patients (n = 2,021), female gender was an independent predictor of all-cause (adjusted OR 1.56; 95% CI, 1.07 to 2.28; P = 0.02) and PE-specific mortality (adjusted OR 1.85; 95% CI, 1.02 to 3.33; P = 0.04). Compared with men, women were 2.05 times more likely to experience a major bleed.Women and men with PE had different clinical characteristics, presentation, and outcomes. Women receiving anticoagulation have a significantly higher risk of major bleeding, suggesting the need for careful monitoring of anticoagulant intensity in women.

During vertebrate development, the paraxial mesoderm becomes segmented, forming somites that will give rise to dermis, axial skeleton and skeletal muscles. Although recently challenged, the "clock and wavefront" model for somitogenesis explains how interactions between several cell-cell communication pathways, including the FGF, RA, Wnt and Notch signals, control the formation of these bilateral symmetric blocks. In the cephalochordate amphioxus, which belongs to the chordate phylum together with tunicates and vertebrates, the dorsal paraxial mesendoderm also periodically forms somites, although this process is asymmetric and extends along the whole body. It has been previously shown that the formation of the most anterior somites in amphioxus is dependent upon FGF signalling. However, the signals controlling somitogenesis during posterior elongation in amphioxus are still unknown. Here we show that, contrary to vertebrates, RA and FGF signals act independently during posterior elongation and that they are not mandatory for posterior somites to form. Moreover, we show that RA is not able to buffer the left/right asymmetry machinery that is controlled through the asymmetric expression of Nodal pathway actors. Our results give new insights into the evolution of the somitogenesis process in chordates. They suggest that RA and FGF pathways have acquired specific functions in the control of somitogenesis in vertebrates. We propose that the "clock and wavefront" system was selected specifically in vertebrates in parallel to the development of more complex somite-derived structures but that it was not required for somitogenesis in the ancestor of chordates.

Insulin is one of the most studied proteins since it is central to the regulation of carbohydrate and fat metabolism in vertebrates and its expression and release are disturbed in diabetes, the most frequent human metabolic disease worldwide. However, the evolution of the function of the insulin protein family is still unclear. In this study, we present a phylogenetic and developmental analysis of the Insulin Like Peptide (ILP) in the cephalochordate amphioxus. We identified an ILP in the European amphioxus Branchiostoma lanceolatum that displays structural characteristics of both vertebrate insulin and Insulin-like Growth Factors (IGFs). Our phylogenetic analysis revealed that amphioxus ILP represents the sister group of both vertebrate insulin and IGF proteins. We also characterized both temporal and spatial expression of ILP in amphioxus. We show that ilp is highly expressed in endoderm and paraxial mesoderm during development, and mainly expressed in the gut of both the developing embryo and adult. We hypothesize that ILP has critical implications in both developmental processes and metabolism and could display IGF- and insulin-like functions in amphioxus supporting the idea of a common ancestral protein.

In this paper we analyse the degree of concordance in species richness and taxonomic distinctness (diversity) patterns among different freshwater taxonomic groups in order to test three long held patterns described in Mexican freshwater biogeography: 1. The aquatic biota of Mexico includes two distinct faunas, a rich Neotropical component in the south and a south-eastern region and a less rich Nearctic component towards central and northern latitudes of the country. 2. A hotspot of species richness and diversity has been recorded in the Usumacinta, including the Yucatan Peninsula. 3. The presence of two distinct biotas in Mexico, an eastern one distributed along the Gulf of Mexico slope, and a western one associated to the Pacific versant. We use species richness and taxonomic distinctness to explore patterns of diversity and how these patterns change between zoogeographical regions. This paper points out a clear separation between Neotropical and Nearctic drainage basins but also between eastern (Gulf of Mexico) and western (Pacific) drainage basins. Present data gives additional empirical support from freshwater biota for three long held beliefs regarding distributional patterns of the Mexican biota. The neotropical basins of Mexico are generally host to a richest and more diversified fauna, that includes more families, genera and species, compared to the less rich and less diverse fauna in the nearctic basins.

BACKGROUND: The basally divergent phylogenetic position of amphioxus (Cephalochordata), as well as its conserved morphology, development and genetics, make it the best proxy for the chordate ancestor. Particularly, studies using the amphioxus model help our understanding of vertebrate evolution and development. Thus, interest for the amphioxus model led to the characterization of both the transcriptome and complete genome sequence of the American species, Branchiostoma floridae. However, recent technical improvements allowing induction of spawning in the laboratory during the breeding season on a daily basis with the Mediterranean species Branchiostoma lanceolatum have encouraged European Evo-Devo researchers to adopt this species as a model even though no genomic or transcriptomic data have been available. To fill this need we used the pyrosequencing method to characterize the B. lanceolatum transcriptome and then compared our results with the published transcriptome of B. floridae. RESULTS: Starting with total RNA from nine different developmental stages of B. lanceolatum, a normalized cDNA library was constructed and sequenced on Roche GS FLX (Titanium mode). Around 1.4 million of reads were produced and assembled into 70,530 contigs (average length of 490 bp). Overall 37% of the assembled sequences were annotated by BlastX and their Gene Ontology terms were determined. These results were then compared to genomic and transcriptomic data of B. floridae to assess similarities and specificities of each species. CONCLUSION: We obtained a high-quality amphioxus (B. lanceolatum) reference transcriptome using a high throughput sequencing approach. We found that 83% of the predicted genes in the B. floridae complete genome sequence are also found in the B. lanceolatum transcriptome, while only 41% were found in the B. floridae transcriptome obtained with traditional Sanger based sequencing. Therefore, given the high degree of sequence conservation between different amphioxus species, this set of ESTs may now be used as the reference transcriptome for the Branchiostoma genus.

A methodology for inducing spawning in captivity of the lancelet Branchiostoma lanceolatum has been developed recently with animals collected at the Racou beach, in the southern coast of France. An increasing amount of laboratories around the world are now working on the evolution of developmental mechanisms (Evo-Devo) using amphioxus collected in this site. Thus, today, the development of new aquaculture techniques for keeping amphioxus in captivity is needed and the study of the natural conditions at which amphioxus is exposed in the Racou beach during their spawning season becomes necessary. We have investigated the amphioxus distribution, size frequency, and population structure in the Racou beach during its natural spawning season using multivariate methods (redundancy analysis and multiple regression). We found a clear preference of amphioxus for sandy sites, something that seems to be a general behaviour of different amphioxus species around the world. We have also estimated the amphioxus growth rate and we show how the animals are preferentially localized in shallow waters during April and June.