Genetically encoded multivalent liquid glycan array displayed on M13 bacteriophage.

The central dogma of biology does not allow for the study of glycans using DNA sequencing. We report a liquid glycan array (LiGA) platform comprising a library of DNA 'barcoded' M13 virions that display 30-1,500 copies of glycans per phage. A LiGA is synthesized by acylation of the phage pVIII protein with a dibenzocyclooctyne, followed by ligation of azido-modified glycans. Pulldown of the LiGA with lectins followed by deep sequencing of the barcodes in the bound phage decodes the optimal structure and density of the recognized glycans. The LiGA is target agnostic and can measure the glycan-binding profile of lectins, such as CD22, on cells in vitro and immune cells in a live mouse. From a mixture of multivalent glycan probes, LiGAs identify the glycoconjugates with optimal avidity necessary for binding to lectins on living cells in vitro and in vivo.

Genetically Encoded Fragment-Based Discovery from Phage-Displayed Macrocyclic Libraries with Genetically Encoded Unnatural Pharmacophores.

Genetically encoded macrocyclic peptide libraries with unnatural pharmacophores are valuable sources for the discovery of ligands for many targets of interest. Traditionally, generation of such libraries employs "early stage" incorporation of unnatural building blocks into the chemically or translationally produced macrocycles. Here, we describe a divergent late-stage approach to such libraries starting from readily available starting material: genetically encoded libraries of peptides. A diketone linchpin 1,5-dichloropentane-2,4-dione converts peptide libraries displayed on phage to 1,3-diketone bearing macrocyclic peptides (DKMP): shelf-stable precursors for Knorr pyrazole synthesis. Ligation of diverse hydrazine derivatives onto DKMP libraries displayed on phage that carries silent DNA-barcodes yields macrocyclic libraries in which the amino acid sequence and the pharmacophore are encoded by DNA. Selection of this library against carbonic anhydrase enriched macrocycles with benzenesulfonamide pharmacophore and nanomolar Kd. The methodology described in this manuscript can graft diverse pharmacophores into many existing genetically encoded phage libraries and significantly increase the value of such libraries in molecular discoveries.

Humoral Immunodeficiency and Immune Globulin Replacement Therapy (IGRT) Usage in DiGeorge Syndrome.

PURPOSE: An analysis of patients in the United States Immunodeficiency Network (USIDNET) registry previously described a discordance in the reported prevalence of humoral immune deficiency in patients with DiGeorge syndrome (DGS) and its treatment. The primary purpose of this study is to evaluate the rates of humoral immunodeficiency and immune globulin replacement therapy (IGRT) use in patients with DiGeorge syndrome in the USIDNET registry as of September 2016, and to correlate IGRT use with prior infections and laboratory evidence of immune deficiency. METHODS: Current patients in the USIDNET registry with DGS were identified. Patients who were treated with immune globulin replacement therapy (IGRT) were compared with those who were untreated with respect to their laboratory findings and clinical history. RESULTS: Four hundred seventy-three patients were identified. The use of IGRT in patients with DGS has increased over time from 3 to 6.6%. IGRT was more common in patients with humoral immune deficiency (18.2% of those with hypogammaglobulinemia, 39.1% of those with documented low vaccine titers), but most patients with evidence of humoral immune deficiency remain untreated with IGRT. Patients treated with IGRT were more likely to have experienced episodes of pneumonia, sepsis, and bacterial skin infections (p < 0.01 for all). CONCLUSIONS: Humoral immune deficiencies were more common among patients with DGS than previously reported. IGRT was used most commonly in patients with DGS who demonstrated frequent or severe bacterial infections. There is still a significant deficit between those with DGS who have laboratory evidence of a humoral immune deficiency and those being treated for it.

Hypothermia: A Sign of Sepsis in Young Infants in the Emergency Department?

OBJECTIVES: Diagnosis of sepsis in young infants can be challenging due to the nonspecific signs, which can include hypothermia. Whether the presence of hypothermia in young infants should prompt evaluation for serious infection is unclear. The objectives were to measure the prevalence of serious infection among infants ≤60 days of age with hypothermia in the emergency department (ED) and determine other clinical features of hypothermic infants who have serious infection. METHODS: This is a retrospective analysis of all infants ≤60 days seen in a children's hospital ED from April 2014 to February 2017. Primary outcome was presence of serious infection, defined as urinary tract infection, bacteremia, meningitis, pneumonia, or herpes virus infection. Hypothermia was defined as a rectal temperature of 36.0°C or less. RESULTS: Of 4797 infants ≤60 days of age seen in the ED, 116 had hypothermia. The prevalence of serious infection was 2.6% (3/116) in hypothermic infants compared with 15.2% (61/401) in febrile infants (P < 0.01). Hypothermic infants with serious infections were more likely to have a history of prematurity, apnea, poor feeding, lethargy, ill-appearance, and respiratory signs than hypothermic infants without serious infection. All 3 hypothermic infants with serious infection had other concerning features. CONCLUSIONS: The prevalence of serious infection in hypothermic young infants in the ED is low. Serious infection is unlikely in infants with isolated hypothermia.

Use of Procalcitonin and a Respiratory Polymerase Chain Reaction Panel to Reduce Antibiotic Use via an Electronic Medical Record Alert.

BACKGROUND: Respiratory tract infections are often viral and but are frequently treated with antibiotics, providing a significant opportunity for antibiotic de-escalation in patients. We sought to determine whether an automated electronic medical record best practice alert (BPA) based on procalcitonin and respiratory polymerase chain reaction (PCR) results could help reduce inappropriate antibiotic use in patients with likely viral respiratory illness. METHODS: This multisite, pre-post, quasi-experimental study included patients 18 years and older with a procalcitonin level <0.25 ng/mL and a virus identified on respiratory PCR within 48 hours of each other, and 1 or more systemic antibiotics ordered. In the study group, a BPA alerted providers of the diagnostic results suggesting viral infection and prompted them to reassess the need for antibiotics. The primary outcome measured was total antibiotic-days of therapy. RESULTS: The BPA reduced inpatient antibiotic-days of therapy by a mean of 2.2 days compared with patients who met criteria but did not have the alert fire (8.0 vs 5.8 days, respectively, P < .001). The BPA also reduced the percentage of patients prescribed antibiotics on discharge (20% vs 47.8%, P < .001), whereas there was no difference in need for antibiotic escalation after initial discontinuation (7.6% vs 4.3%, P = .198). CONCLUSIONS: The automated antimicrobial stewardship BPA effectively reduced antibiotic use and discharge prescribing rates when diagnostics suggested viral respiratory tract infection, without a higher rate for reinitiation of antibiotics after discontinuation.

Success and complications by team composition for prehospital paediatric intubation: a systematic review and meta-analysis.

BACKGROUND: Clinical team composition for prehospital paediatric intubation may affect success and complication rates. We performed a systematic review and meta-analysis to determine the success and complication rates by type of clinical team. METHODS: We searched MEDLINE, EMBASE, and CINAHL for interventional and observational studies describing prehospital intubation attempts in children with overall success, first-pass success, and complication rates. Eligible studies, data extraction, and assessment of risk of bias were assessed independently by two reviewers. We performed a random-effects meta-analysis of proportions. RESULTS: Forty studies (1989 to 2019) described three types of clinical teams: non-physician teams with no relaxants (22 studies, n = 7602), non-physician teams with relaxants (12 studies, n = 2185), and physician teams with relaxants (12 studies, n = 1780). Twenty-two (n = 3747) and 18 (n = 7820) studies were at low and moderate risk of bias, respectively. Non-physician teams without relaxants had lower overall intubation success rate (72%, 95% CI 67-76%) than non-physician teams with relaxants (95%, 95% CI 93-98%) and physician teams (99%, 95% CI 97-100%). Physician teams had higher first-pass success rate (91%, 95% CI 86-95%) than non-physicians with (75%, 95% CI 69-81%) and without (55%, 95% CI 48-63%) relaxants. Overall airway complication rate was lower in physician teams (10%, 95% CI 3-22%) than non-physicians with (30%, 95% CI 23-38%) and without (39%, 95% CI 28-51%) relaxants. CONCLUSION: Physician teams had higher rates of intubation success and lower rates of overall airway complications than other team types. Physician prehospital teams should be utilised wherever practicable for critically ill children requiring prehospital intubation.

T-Cell Receptor Excision Circles in Newborns with Congenital Heart Disease.

OBJECTIVES: To determine if children with congenital heart disease (CHD) have lower newborn T-cell receptor excision circles (TREC) levels than the general population and to evaluate if low TREC levels in newborns with CHD are associated with clinical complications such as hospitalization for infection. STUDY DESIGN: The Connecticut Newborn Screening Program reported TREC levels for newborns with CHD delivered between October 2011 and September 2016 at 2 major Connecticut children's hospitals. TREC levels for children with CHD were compared with the general population. TREC levels and outcome measures, including hospitalization for infection, were compared. RESULTS: We enrolled 575 participants with CHD in the study. The median TREC level for newborns with CHD was lower than the general population (180.1 copies/µL vs 312.5 copies/µL; P < .01). patients with CHD requiring hospitalization for infection had lower median TREC levels than their counterparts (143.0 copies/µL vs 186.7 copies/µL; P < .01). The combination of prematurity and low TREC level had a strong relationship to hospitalization for infection (area under the receiver operative characteristic curve of 0.89). There was no association between TREC level and CHD severity. CONCLUSIONS: Newborns with CHD demonstrated lower TREC levels than the general population. Low TREC levels were associated with hospitalization for infection in preterm children with CHD. Study limitations include that this was a retrospective chart review. These findings may help to identify newborns with CHD at highest risk for infection, allowing for potential opportunities for intervention.

Physician-staffed helicopter emergency medical services augment ground ambulance paediatric airway management in urban areas: a retrospective cohort study.

OBJECTIVES: Paediatric intubation is a high-risk procedure for ground emergency medical services (GEMS). Physician-staffed helicopter EMS (PS-HEMS) may bring additional skills, drugs and equipment to the scene including advanced airway management beyond the scope of GEMS even in urban areas with short transport times. This study aimed to evaluate prehospital paediatric intubation performed by a PS-HEMS when dispatched to assist GEMS in a large urban area and examine how often PS-HEMS provided airway intervention that was not or could not be provided by GEMS. METHODS: We performed a retrospective observational study from July 2011 to December 2016 of a PS-HEMS in a large urban area (Sydney, Australia), which responds in parallel to GEMS. GEMS intubate without adjuvant neuromuscular blockade, whereas the PS-HEMS use neuromuscular blockade and anaesthetic agents. We examined endotracheal intubation success rate, first-look success rate and complications for the PS-HEMS and contrasted this with the advanced airway interventions provided by GEMS prior to PS-HEMS arrival. RESULTS: Overall intubation success rate was 62/62 (100%) and first-look success was 59/62 (95%) in the PS-HEMS-treated group, whereas the overall success rate was 2/7 (29%) for the GEMS group. Peri-intubation hypoxia was documented in 5/65 (8%) of the PS-HEMS intubation attempts but no other complications were reported. However, 3/7 (43%) of the attempted intubations by GEMS were oesophageal intubations, two of which were unrecognised. CONCLUSIONS: PS-HEMS have high success with low complication rates in paediatric prehospital intubation. Even in urban areas with rapid GEMS response, PS-HEMS activated in parallel can provide safe and timely advanced prehospital airway management for seriously ill and injured children beyond the scope of GEMS practice. Review of GEMS airway management protocols and the PS-HEMS case identification and dispatch system in Sydney is warranted.

Genetically-encoded fragment-based discovery of glycopeptide ligands for DC-SIGN.

We have employed genetically-encoded fragment-based discovery to identify novel glycopeptides with affinity for the dendritic cell receptor DC-SIGN. Starting from libraries of 108 mannose-conjugated peptides, we identified glycopeptides that exhibited up to a 650-fold increase in multivalent binding affinity for DC-SIGN, which is also preserved in cells. Monovalently, our most potent glycopeptides have a similar potency to a Man3 oligosaccharide, representing a 15-fold increase in activity compared to mannose. These compounds represent the first examples of glycopeptide ligands that target the CRD of DC-SIGN. The natural framework of glycopeptide conjugates and the simplicity of orthogonal conjugation to make these glycopeptides anticipates a promising future for development of DC-SIGN-targeting moieties.

Silicone Oil Migration to the Lateral Rectus Muscle: An Unusual Finding.

We report a case of silicone oil migration to the lateral rectus muscle discovered during otherwise uncomplicated strabismus surgery in a patient with multiple previous vitreoretinal surgical interventions. A 58-year-old female underwent strabismus surgery to correct a long-standing sensory exotropia. This patient underwent numerous previous vitreoretinal surgical interventions due to a complex history of retinal detachments with subsequent repairs, including inserting and removing intraocular silicone as an endotamponade. During this procedure, silicone oil cysts were discovered firmly adhered within the substance of the lateral rectus muscle. These cysts were subsequently removed either by rupture or whole excision, and specimens were sent for pathological examination. Microscopy confirmed sections of fat necrosis of the lateral rectus muscle with areas incorporating silicone oil. Following this, a lateral rectus recession and medial rectus resection were performed, and the exotropia was satisfactorily corrected. This report highlights an unusual complication of otherwise unchallenging strabismus surgery. Intraocular silicone oil introduced during vitreoretinal surgical procedures may present as cysts embedded on extraocular muscle, thus presenting a unique finding of strabismus surgery.

A multimodal antimicrobial stewardship intervention to improve antibiotic prescribing in patients with COVID-19.

This study assessed outcomes prior to and after electronic medical record-based clinical decision support implementation combined with prospective audit in patients with COVID-19. This multimodal stewardship intervention was associated with a decrease in antibiotic exposure for patients with COVID-19 (44.4% vs 61.8%, p = 0.002) within the first 7 days of hospitalization.

Cryo-electron microscopy of the f1 filamentous phage reveals insights into viral infection and assembly.

Phages are viruses that infect bacteria and dominate every ecosystem on our planet. As well as impacting microbial ecology, physiology and evolution, phages are exploited as tools in molecular biology and biotechnology. This is particularly true for the Ff (f1, fd or M13) phages, which represent a widely distributed group of filamentous viruses. Over nearly five decades, Ffs have seen an extraordinary range of applications, yet the complete structure of the phage capsid and consequently the mechanisms of infection and assembly remain largely mysterious. In this work, we use cryo-electron microscopy and a highly efficient system for production of short Ff-derived nanorods to determine a structure of a filamentous virus including the tips. We show that structure combined with mutagenesis can identify phage domains that are important in bacterial attack and for release of new progeny, allowing new models to be proposed for the phage lifecycle.

Unrecognized viral respiratory tract infections in premature infants during their birth hospitalization: a prospective surveillance study in two neonatal intensive care units.

OBJECTIVE: To determine the frequency and effects of nosocomial respiratory viral infections (RVIs) in premature neonates, including those who may be asymptomatic. STUDY DESIGN: We performed a year-long surveillance for RVIs in infants <33 weeks gestational age admitted to 2 Syracuse neonatal intensive care units. Infants were enrolled within 3 days of neonatal intensive care unit admission and were sampled for RVIs until discharge using a multiplex polymerase chain reaction assay capable of detecting 17 different respiratory viruses or subtypes. RESULTS: Twenty-six of 50 prematurely born infants (52%) tested positive for a respiratory virus at least once during their birth hospitalization. Testing positive for a respiratory virus was significantly associated with longer length of stay (70 days vs 35 days, P = .002) and prolonged ventilatory support (51 vs 13 days, P = .002). Infants who tested positive for a respiratory virus during their birth hospitalization had more than twice the rate of developing bronchopulmonary dysplasia (P < .05). CONCLUSION: Nosocomial RVIs were frequent in our study population, despite the absence of clinical indicators of illness. Length of hospital stay was significantly longer and a diagnosis of bronchopulmonary dysplasia was more common in infants who had respiratory viruses detected.

Diastereoselective alkylation reactions of 1-methylcyclohexa-2,5-diene-1-carboxylic acid.

The deprotonation and alkylation of 1-methylcyclohexa-2,5-diene-1-carboxylic acid has been investigated under a range of conditions. In all cases, the formation of compounds 14 was found to be completely stereoselective, although compound 14c was formed as an impurity when alkyl iodides were used as electrophiles, and doubly-alkylated compounds 17 were formed in some cases when alkyl bromides were used.

Ceftriaxone versus cefazolin for the treatment of methicillin-susceptible Staphylococcus aureus bacteraemia.

Few studies have evaluated the use of ceftriaxone (CRO) in the treatment of methicillin-susceptible Staphylococcus aureus (MSSA) infections. The purpose of this study was to compare the safety and efficacy of CRO versus cefazolin (CZO) for patients with MSSA bacteraemia. This was a multicentre, single health-system, retrospective study. Adult inpatients were included if they had a primary episode of MSSA bacteraemia and received CRO or CZO as definitive therapy. The primary endpoint was clinical cure at 28 days or at discharge, whichever came first. Secondary endpoints included treatment failure at 90 days, time to treatment failure, re-admission due to recurrent MSSA bacteraemia, duration of bacteraemia, discontinuation of treatment due to adverse drug events, and Clostridioides difficile infection. A total of 248 patients were included, of which 87 (35.1%) received CRO and 161 (64.9%) received CZO. There was no difference in the primary outcome of clinical cure at 28 days or at discharge between the CRO and CZO groups [75 (86.2%) vs. 145 (90.1%); P = 0.359], even after adjusting for Charlson comorbidity index and Pitt bacteremia score (adjusted OR = 1.35, 95% CI 0.58-3.12; P = 0.49). There were no differences in time to clinical cure, treatment failure at 90 days or safety events between the two groups. In conclusion, our findings suggest no clinical difference between CRO and CZO for the definitive treatment of MSSA bacteraemia. Further prospective studies are needed to confirm these findings.

Templated microbiology comments with candiduria to enhance antimicrobial stewardship.

Objective: To evaluate the effect of templated microbiology reporting comments on antifungal utilization in patients with candiduria. Design: In this retrospective, quasi-experimental study, we evaluated a preimplementation cohort (June 2018-January 2019) compared with a postimplementation cohort (June 2019-January 2020). Setting: A multisite health system including 1 academic hospital and 4 community hospitals. Patients: Patients were aged ≥18 years, were hospitalized, and had candiduria documented at least once during their admission. The study included 156 patients in the preimplementation period and 141 patients in the postimplementation period. Methods: In June 2019, Saint Luke's Health System implemented the use of templated comments for urine cultures with Candida spp growth. When Candida is isolated, the following comment appears in the microbiology result section: "In the absence of symptoms, Candida is generally considered normal flora. No therapy indicated unless high risk (pregnant, neonate, or neutropenic) or undergoing urologic procedure. If Foley catheter present, remove or replace when able." The primary outcome was rate of antifungal prescribing. Results: Antifungal administration within 72 hours of a culture identifying a Candida spp occurred in 75 patients in the preimplementation group and 48 patients in the postimplementation group (48.1% vs 34.0%; P = .02). We did not detect a difference between groups in antifungal administration between 73 and 240 hours (1.3% vs 3.5%; P = .26), nor did we detect a difference in median antifungal duration (4 vs 3 days; P = .43). Conclusion: Using a templated comment with urine cultures reduced antifungal prescription rates in hospitalized patients with candiduria. This strategy is a low-resource technique to improve antimicrobial stewardship.

Filamentous bacteriophage: biology, phage display and nanotechnology applications.

Filamentous bacteriophage, long and thin filaments that are secreted from the host cells without killing them, have been an antithesis to the standard view of head-and-tail bacterial killing machines. Episomally replicating filamentous phage Ff of Escherichia coli provide the majority of information about the principles and mechanisms of filamentous phage infection, episomal replication and assembly. Chromosomally- integrated "temperate" filamentous phage have complex replication and integration, which are currently under active investigation. The latter are directly or indirectly implicated in diseases caused by bacterial pathogens Vibrio cholerae, Pseudomonas aeruginosa and Neisseria meningitidis. In the first half of the review, both the Ff and temperate phage are described and compared. A large section of the review is devoted to an overview of phage display technology and its applications in nanotechnology.

Pneumocystis pneumonia.

Pneumocystis (carinii) jiroveci pneumonia can occur in immunocompromised individuals, especially hematopoietic stem and solid organ transplant recipients and those receiving immunosuppressive agents, and is the most common opportunistic infection in persons with advanced human immunodeficiency virus (HIV) infection. The Pneumocystis genus was initially mistaken as a trypanosome and later as a protozoan. Genetic analysis identified the organism as a unicellular fungus. Pneumocystis jiroveci is the species responsible for human infections. A slow indolent time course with symptoms of pneumonia progressing over weeks to months is characteristic in HIV-infected patients. Fulminant respiratory failure associated with fever and dry cough is typical in non-HIV-infected patients. Definitive diagnosis relies on histopathological testing of sputum, induced or sampled by fiberoptic bronchoscopy with bronchoalveolar lavage. The first-line drug for treatment and prevention is trimethoprim-sulfamethoxazole.

Corynebacterium Bovis: A Rare Case of Persistent Bacterial Keratitis and Corneal Perforation.

We report a rare case of severe, non-contact lens-related Corynebacterium bovis corneal infection on a background of viral keratitis, resulting in corneal abscess formation with subsequent corneal perforation. An 89-year-old Caucasian lady presented with a significant epithelial defect and a dense stromal infiltrates on a background of herpes zoster keratitis, ultimately resulting in corneal perforation. Enrichment culture obtained from corneal scraping isolated the unusual organism Corynebacterium bovis. This was treated with a combination of culture-directed, targeted course of antibiotics and surgical interventions. To the best of our knowledge, this is the first reported case of profuse bacterial keratitis secondary to Corynebacterium bovis infiltration, on a background of viral keratitis, resulting in corneal abscess formation and subsequent perforation. This report highlights this rare bacterium's characteristics including its pathogenicity in causing severe corneal disease, particularly in immunosuppressed environments such as in this case, apparent antibiotic sensitivities & resistance, and potential transmission route.