Point-of-Care Ultrasound of Post-acute COVID-19 Syndrome: A Prospective Cohort Study.

Introduction Acute COVID-19 patients can suffer from chronic symptoms known as post-acute sequelae of SARS-CoV-2 infection (PASC). Point-of-care ultrasound (POCUS) is established in acute COVID, but its utility in PASC is unclear. We sought to determine the incidence of cardiac and pulmonary abnormalities with POCUS in patients with PASC in a COVID-19 recovery clinic. Methods This prospective cohort study included adults (>18 years old) presenting with cardiopulmonary symptoms to the COVID-19 recovery clinic. A lung ultrasound and standard bedside echocardiogram were performed by ultrasound-trained physicians. Images were interpreted in real time by the performing sonographer and independently by a blinded ultrasound faculty member. Discrepancies in interpretation were addressed by consensus review. A modified Soldati score was calculated by the sum of the scores in each of the 12 lung zones, with each zone score ranging from 0 to 3 (maximum score of 36). The score was then compared to clinical outcomes and outpatient testing.  Results Between April and July 2021, 41 patients received POCUS examinations, with 24 of those included in the study. In all, 15 out of 24 (62.5%) had a normal lung ultrasound. Of the nine subjects with lung abnormalities, the median modified Soldati score was 2. Three patients had trivial pericardial effusions, and all had normal left and right ventricular size and function. Conclusion The majority (62.5%) of patients presenting to the PASC clinic had a normal pulmonary ultrasound, and the vast majority (87.5%) had normal cardiac ultrasounds. These findings suggest that cardiopulmonary symptoms in PASC may be from etiologies not well evaluated by POCUS.

Design and validation of Dolosigranulum pigrum specific PCR primers using the bacterial core genome.

Dolosigranulum pigrum-a lactic acid bacterium that is increasingly recognized as an important member of the nasal microbiome. Currently, there are limited rapid and low-cost options for confirming D. pigrum isolates and detecting D. pigrum in clinical specimens. Here we describe the design and validation of a novel PCR assay targeting D. pigrum that is both sensitive and specific. We designed a PCR assay targeting murJ, a single-copy core species gene identified through the analysis of 21 D. pigrum whole genome sequences. The assay achieved 100% sensitivity and 100% specificity against D. pigrum and diverse bacterial isolates and an overall 91.1% sensitivity and 100% specificity using nasal swabs, detecting D. pigrum at a threshold of 1.0 × 104 D. pigrum 16S rRNA gene copies per swab. This assay adds a reliable and rapid D. pigrum detection tool to the microbiome researcher toolkit investigating the role of generalist and specialist bacteria in the nasal environment.

Class switching is differentially regulated in RBC alloimmunization and vaccination.

BACKGROUND: Studies of human patients have shown that most anti-RBC alloantibodies are IgG1 or IgG3 subclasses, although it is unclear why transfused RBCs preferentially drive these subclasses over others. Though mouse models allow for the mechanistic exploration of class-switching, previous studies of RBC alloimmunization in mice have focused more on the total IgG response than the relative distribution, abundance, or mechanism of IgG subclass generation. Given this major gap, we compared the IgG subclass distribution generated in response to transfused RBCs relative to protein in alum vaccination, and determined the role of STAT6 in their generation. STUDY DESIGN AND METHODS: WT mice were either immunized with Alum/HEL-OVA or transfused with HOD RBCs and levels of anti-HEL IgG subtypes were measured using end-point dilution ELISAs. To study the role of STAT6 in IgG class-switching, we first generated and validated novel STAT6 KO mice using CRISPR/cas9 gene editing. STAT6 KO mice were then transfused with HOD RBCs or immunized with Alum/HEL-OVA, and IgG subclasses were quantified by ELISA. RESULTS: When compared with antibody responses to Alum/HEL-OVA, transfusion of HOD RBCs induced lower levels of IgG1, IgG2b, and IgG2c but similar levels of IgG3. Class switching to most IgG subtypes remained largely unaffected in STAT6 deficient mice in response to HOD RBC transfusion, with the one exception being IgG2b. In contrast, STAT6 deficient mice showed altered levels of all IgG subtypes following Alum vaccination. DISCUSSION: Our results show that anti-RBC class-switching occurs via alternate mechanisms when compared with the well-studied immunogen alum vaccination.

Class switching is differentially regulated in RBC alloimmunization and vaccination.

Background: Studies of human patients have shown that most anti-RBC alloantibodies are IgG1 or IgG3 subclasses, though it is unclear why transfused RBCs preferentially drive these subclasses over others. Though mouse models allow for the mechanistic exploration of class-switching, previous studies of RBC alloimmunization in mice have focused more on the total IgG response than the relative distribution, abundance, or mechanism of IgG subclass generation. Given this major gap, we compared the IgG subclass distribution generated in response to transfused RBCs relative to protein in alum vaccination, and determined the role of STAT6 in their generation. Study Design and Methods: WT mice were either immunized with Alum/HEL-OVA or transfused with HOD RBCs and levels of anti-HEL IgG subtypes were measured using end-point dilution ELISAs. To study the role of STAT6 in IgG class-switching, we first generated and validated novel STAT6 KO mice using CRISPR/cas9 gene editing. STAT6 KO mice were then transfused with HOD RBCs or immunized with Alum/HEL-OVA, and IgG subclasses were quantified by ELISA. Results: When compared to antibody responses to Alum/HEL-OVA, transfusion of HOD RBCs induced lower levels of IgG1, IgG2b and IgG2c but similar levels of IgG3. Class switching to most IgG subtypes remained largely unaffected in STAT6 deficient mice in response to HOD RBC transfusion, with the one exception being IgG2b. In contrast, STAT6 deficient mice showed altered levels of all IgG subtypes following Alum vaccination. Discussion: Our results show that anti-RBC class-switching occurs via alternate mechanisms when compared to the well-studied immunogen alum vaccination.

The lysophospholipid-binding molecule CD1D is not required for the alloimmunization response to fresh or stored RBCs in mice despite RBC storage driving alterations in lysophospholipids.

BACKGROUND: Despite the significant adverse clinical consequences of RBC alloimmunization, our understanding of the signals that induce immune responses to transfused RBCs remains incomplete. Though RBC storage has been shown to enhance alloimmunization in the hen egg lysozyme, ovalbumin, and human Duffy (HOD) RBC alloantigen mouse model, the molecular signals leading to immune activation in this system remain unclear. Given that the nonclassical major histocompatibility complex (MHC) Class I molecule CD1D can bind to multiple different lysophospholipids and direct immune activation, we hypothesized that storage of RBCs increases lysophospholipids known to bind CD1D, and further that recipient CD1D recognition of these altered lipids mediates storage-induced alloimmunization responses. STUDY DESIGN AND METHODS: We used a mass spectrometry-based approach to analyze the changes in lysophospholipids that are induced during storage of mouse RBCs. CD1D knockout (CD1D-KO) and wild-type (WT) control mice were transfused with stored HOD RBCs to measure the impact of CD1D deficiency on RBC alloimmunization. RESULTS: RBC storage results in alterations in multiple lysophospholipid species known to bind to CD1D and activate the immune system. Prior to transfusion, CD1D-deficient mice had lower baseline levels of polyclonal immunoglobulin (IgG) relative to WT mice. In response to stored RBC transfusion, CD1D-deficient mice generated similar levels of anti-HOD IgM and anti-HOD IgG. CONCLUSION: Although storage of RBCs leads to alteration of several lysophospholipids known to be capable of binding CD1D, storage-induced RBC alloimmunization responses are not impacted by recipient CD1D deficiency.

Cognitive ecology in hummingbirds: the role of sexual dimorphism and its anatomical correlates on memory.

In scatter-hoarding species, several behavioral and neuroanatomical adaptations allow them to store and retrieve thousands of food items per year. Nectarivorous animals face a similar scenario having to remember quality, location and replenishment schedules of several nectar sources. In the green-backed firecrown hummingbird (Sephanoides sephanoides), males are territorial and have the ability to accurately keep track of nectar characteristics of their defended food sources. In contrast, females display an opportunistic strategy, performing rapid intrusions into males territories. In response, males behave aggressively during the non-reproductive season. In addition, females have higher energetic demands due to higher thermoregulatory costs and travel times. The natural scenario of this species led us to compared cognitive abilities and hippocampal size between males and females. Males were able to remember nectar location and renewal rates significantly better than females. However, the hippocampal formation was significantly larger in females than males. We discuss these findings in terms of sexually dimorphic use of spatial resources and variable patterns of brain dimorphisms in birds.