Detection of Bartonella schoenbuchensis (sub)species DNA in different louse fly species in Saxony, Germany: The proof of multiple PCR analysis necessity in case of ruminant-associated bartonellae determination.

BACKGROUND: Hippoboscid flies are bloodsucking arthropods that can transmit pathogenic microorganisms and are therefore potential vectors for pathogens such as Bartonella spp. These Gram-negative bacteria can cause mild-to-severe clinical signs in humans and animals; therefore, monitoring Bartonella spp. prevalence in louse fly populations appears to be a useful prerequisite for zoonotic risk assessment. METHODS: Using convenience sampling, we collected 103 adult louse flies from four ked species (Lipoptena cervi, n = 22; Lipoptena fortisetosa, n = 61; Melophagus ovinus, n = 12; Hippobosca equina, n = 8) and the pupae of M. ovinus (n = 10) in the federal state of Saxony, Germany. All the samples were screened by polymerase chain reaction (PCR) for Bartonella spp. DNA, targeting the citrate synthase gene (gltA). Subsequently, PCRs targeting five more genes (16S, ftsZ, nuoG, ribC and rpoB) were performed for representatives of revealed gltA genotypes, and all the PCR products were sequenced to identify the Bartonella (sub)species accurately. RESULTS AND CONCLUSIONS: The overall detection rates for Bartonella spp. were 100.0%, 59.1%, 24.6% and 75.0% in M. ovinus, L. cervi, L. fortisetosa and H. equina, respectively. All the identified bartonellae belong to the Bartonella schoenbuchensis complex. Our data support the proposed reclassification of the (sub)species status of this group, and thus we conclude that several genotypes of B. schoenbuchensis were detected, including Bartonella schoenbuchensis subsp. melophagi and Bartonella schoenbuchensis subsp. schoenbuchensis, both of which have previously validated zoonotic potential. The extensive PCR analysis revealed the necessity of multiple PCR approach for proper identification of the ruminant-associated bartonellae.

A multicenter evaluation of pediatric emergency department injury visits during the COVID-19 pandemic.

BACKGROUND: Injuries, the leading cause of death in children 1-17 years old, are often preventable. Injury patterns are impacted by changes in the child's environment, shifts in supervision, and caregiver stressors. The objective of this study was to evaluate the incidence and proportion of injuries, mechanisms, and severity seen in Pediatric Emergency Departments (PEDs) during the COVID-19 pandemic. METHODS: This multicenter, cross-sectional study from January 2019 through December 2020 examined visits to 40 PEDs for children < 18 years old. Injury was defined by at least one International Classification of Disease-10th revision (ICD-10) code for bodily injury (S00-T78). The main study outcomes were total and proportion of PED injury-related visits compared to all visits in March through December 2020 and to the same months in 2019. Weekly injury visits as a percentage of total PED visits were calculated for all weeks between January 2019 and December 2020. RESULTS: The study included 741,418 PED visits for injuries pre-COVID-19 pandemic (2019) and during the COVID-19 pandemic (2020). Overall PED visits from all causes decreased 27.4% in March to December 2020 compared to the same time frame in 2019; however, the proportion of injury-related PED visits in 2020 increased by 37.7%. In 2020, injured children were younger (median age 6.31 years vs 7.31 in 2019), more commonly White (54% vs 50%, p < 0.001), non-Hispanic (72% vs 69%, p < 0.001) and had private insurance (35% vs 32%, p < 0.001). Injury hospitalizations increased 2.2% (p < 0.001) and deaths increased 0.03% (p < 0.001) in 2020 compared to 2019. Mean injury severity score increased (2.2 to 2.4, p < 0.001) between 2019 and 2020. Injuries declined for struck by/against (- 4.9%) and overexertion (- 1.2%) mechanisms. Injuries proportionally increased for pedal cycles (2.8%), cut/pierce (1.5%), motor vehicle occupant (0.9%), other transportation (0.6%), fire/burn (0.5%) and firearms (0.3%) compared to all injuries in 2020 versus 2019. CONCLUSIONS: The proportion of PED injury-related visits in March through December 2020 increased compared to the same months in 2019. Racial and payor differences were noted. Mechanisms of injury seen in the PED during 2020 changed compared to 2019, and this can inform injury prevention initiatives.

Factors associated with US caregivers' uptake of pediatric COVID-19 vaccine by race and ethnicity.

OBJECTIVES: To assess differences in willingness to vaccinate children against COVID-19, and factors that may be associated with increased acceptance, among US caregivers of various racial and ethnic identities who presented with their child to the Emergency Department (ED) after emergency use authorization of vaccines for children ages 5-11. STUDY DESIGN: A multicenter, cross-sectional survey of caregivers presenting to 11 pediatric EDs in the United States in November-December 2021. Caregivers were asked about their identified race and ethnicity and if they planned to vaccinate their child. We collected demographic data and inquired about caregiver concerns related to COVID-19. We compared responses by race/ethnicity. Multivariable logistic regression models served to determine factors that were independently associated with increased vaccine acceptance overall and among racial/ethnic groups. RESULTS: Among 1916 caregivers responding, 54.67% planned to vaccinate their child against COVID-19. Large differences in acceptance were noted by race/ethnicity, with highest acceptance among Asian caregivers (61.1%) and those who did not specify a listed racial identity (61.1%); caregivers identifying as Black (44.7%) or Multi-racial (44.4%) had lower acceptance rates. Factors associated with intent to vaccinate differed by racial/ethnic group, and included caregiver COVID-19 vaccine receipt (all groups), caregiver concerns about COVID-19 (White caregivers), and having a trusted primary provider (Black caregivers). CONCLUSIONS: Caregiver intent to vaccinate children against COVID-19 varied by race/ethnicity, but race/ethnicity did not independently account for these differences. Caregiver COVID-19 vaccination status, concerns about COVID-19, and presence of a trusted primary provider are important in vaccination decisions.

Should COVID-19 vaccines be mandated in schools? - an international caregiver perspective.

OBJECTIVES: Caregiver attitudes toward mandating COVID-19 vaccines for their children are poorly understood. We aimed to determine caregiver acceptability of COVID-19 vaccine mandates for schools/daycares and assess if opposition to mandates would result in removal of children from the educational system. STUDY DESIGN: Perform a cross-sectional, anonymous survey of adult caregivers with children ≤ 18 years presenting to 21 pediatric emergency departments in the United States, Canada, Israel, and Switzerland, November 1st through December 31st, 2021. The primary outcome was caregiver acceptance rates for school vaccine mandates, and the secondary outcomes included factors associated with mandate acceptance and caregiver intention to remove the child from school. RESULTS: Of 4,393 completed surveys, 37% of caregivers were opposed to any school vaccine mandate. Caregiver acceptance was lowest for daycare settings (33%) and increased as the child's level of education increased, college (55%). 26% of caregivers report a high likelihood (score of 8-10 on 0-10 scale) to remove their child from school if the vaccine became mandatory. Child safety was caregivers' greatest concern over vaccine mandates. A multivariable model demonstrated intent to vaccinate their child for COVID-19 (OR = 8.9, 95% CI 7.3 to 10.8; P < 0.001) and prior COVID-19 vaccination for the caregiver (OR = 3.8, 95% CI 3.0 to 4.9; P < 0.001) had the greatest odds of increasing mandate acceptance for any school level. CONCLUSIONS: Many caregivers are resistant to COVID-19 vaccine mandates for schools, and acceptance varies with school level. One-fourth of caregivers plan to remove their child from the educational system if vaccines become mandated.

Multimedia Evaluation of EMT-Paramedic Assessment and Management of Pediatric Respiratory Distress.

BACKGROUND: The prehospital care of asthma, bronchiolitis and croup is directed by evidence-based Emergency Medical Services (EMS) protocols. Determining the appropriate intervention for these conditions requires Emergency Medical Technicians-Paramedics (EMT-Ps) to correctly differentiate asthma/bronchospasm, bronchiolitis, and croup. The diagnostic accuracy of EMT-Ps for these pediatric respiratory distress conditions is unknown. OBJECTIVE: We hypothesized increasing provider age, years of provider experience, higher volume of pediatric cases, self-reported comfort with pediatric patients, and having children of one's own would be associated with increased accuracy in diagnosis on a validated multimedia questionnaire. METHODS: This is a cross-sectional study of paramedics from a single EMS agency who completed a validated, case-based questionnaire between July and September 2018. The multimedia questionnaire consisted of four cases, each of which included patient videos and lung sound recordings. Paramedics were asked to assess the severity of distress and ascribe the correct diagnosis and prehospital intervention for each case. Each paramedic completed the questionnaire independently. We defined high questionnaire performance a priori as correctly identifying the diagnosis for ≥75% of cases and used multivariate regression to assess factors associated with high questionnaire performance. Provider age and EMS experience were reported in years and analyzed as continuous variables. Volume of pediatric cases was dichotomized to <1 and ≥1 case per shift and having children was dichotomized to either having children or not having children. RESULTS: Of 514 paramedics, 420 (82%) completed the questionnaire. Overall, paramedics correctly assessed the severity of respiratory distress 92% of the time. However, they only ascribed the correct diagnosis 50% and selected the correct intervention(s) 38% of the time. Increasing age, years of experience, higher volume of pediatric cases, self-reported comfort with pediatric patients, and having children of their own were not associated with questionnaire performance. CONCLUSION: Paramedics accurately assessed severity of distress in multimedia cases of asthma/bronchospasm, bronchiolitis and croup in children, but showed significant room for improvement in correctly identifying the diagnosis and in selecting appropriate intervention(s). Age, years of EMS experience, higher volume of clinical pediatric cases, self-reported comfort with pediatric patients, and having children of their own were not associated with questionnaire performance.

Paramedic-Identified Enablers of and Barriers to Pediatric Seizure Management: A Multicenter, Qualitative Study.

Background: Seizures have the potential to cause significant morbidity and mortality, and are a common reason emergency medical services (EMS) are requested for a child. An evidence-based guideline (EBG) for pediatric prehospital seizures was published and has been implemented as protocol in multiple EMS systems. Knowledge translation and protocol adherence in medicine can be incomplete. In EMS, systems-based factors and providers' attitudes and beliefs may contribute to incomplete knowledge translation and protocol implementation. Objective: The purpose of this study was to identify paramedic attitudes and beliefs regarding pediatric seizure management and regarding potential barriers to and enablers of adherence to evidence-based pediatric seizure protocols in multiple EMS systems. Methods: This was a qualitative study utilizing semi-structured interviews of paramedics who recently transported actively seizing 0-17 year-old patients in 3 different urban EMS systems. Interviewers explored the providers' decision-making during their recent case and regarding seizures in general. Interview questions explored barriers to and enablers of protocol adherence. Two investigators used the grounded theory approach and constant comparison to independently analyze transcribed interview recordings until thematic saturation was reached. Findings were validated with follow-up member-checking interviews. Results: Several themes emerged from the 66 interviewed paramedics. Enablers of protocol adherence included point-of-care references, the availability of different routes for midazolam and availability of online medical control. Systems-level barriers included equipment availability, controlled substance management, infrequent pediatric training, and protocol ambiguity. Provider-level barriers included concerns about respiratory depression, provider fatigue, preferences for specific routes, febrile seizure perceptions, and inaccurate methods of weight estimation. Paramedics suggested system improvements to address dose standardization, protocol clarity, simplified controlled substance logistics, and equipment availability. Conclusions: Paramedics identified enablers of and barriers to adherence to evidence-based pediatric seizure protocols. The identified barriers existed at both the provider and systems levels. Paramedics identified multiple potential solutions to overcome several barriers to protocol adherence. Future research should focus on using the findings of this study to revise seizure protocols and to deploy measures to improve protocol implementation. Future research should also analyze process and outcome measures before and after the implementation of revised seizure protocols informed by the findings of this study.

Enhanced presentation of MHC class Ia, Ib and class II-restricted peptides encapsulated in biodegradable nanoparticles: a promising strategy for tumor immunotherapy.

BACKGROUND: Many peptide-based cancer vaccines have been tested in clinical trials with a limited success, mostly due to difficulties associated with peptide stability and delivery, resulting in inefficient antigen presentation. Therefore, the development of suitable and efficient vaccine carrier systems remains a major challenge. METHODS: To address this issue, we have engineered polylactic-co-glycolic acid (PLGA) nanoparticles incorporating: (i) two MHC class I-restricted clinically-relevant peptides, (ii) a MHC class II-binding peptide, and (iii) a non-classical MHC class I-binding peptide. We formulated the nanoparticles utilizing a double emulsion-solvent evaporation technique and characterized their surface morphology, size, zeta potential and peptide content. We also loaded human and murine dendritic cells (DC) with the peptide-containing nanoparticles and determined their ability to present the encapsulated peptide antigens and to induce tumor-specific cytotoxic T lymphocytes (CTL) in vitro. RESULTS: We confirmed that the nanoparticles are not toxic to either mouse or human dendritic cells, and do not have any effect on the DC maturation. We also demonstrated a significantly enhanced presentation of the encapsulated peptides upon internalization of the nanoparticles by DC, and confirmed that the improved peptide presentation is actually associated with more efficient generation of peptide-specific CTL and T helper cell responses. CONCLUSION: Encapsulating antigens in PLGA nanoparticles offers unique advantages such as higher efficiency of antigen loading, prolonged presentation of the antigens, prevention of peptide degradation, specific targeting of antigens to antigen presenting cells, improved shelf life of the antigens, and easy scale up for pharmaceutical production. Therefore, these findings are highly significant to the development of synthetic vaccines, and the induction of CTL for adoptive immunotherapy.

Antigenic profiling of glioma cells to generate allogeneic vaccines or dendritic cell-based therapeutics.

PURPOSE: Allogeneic glioma cell lines that are partially matched to the patient at class I human leukocyte antigen (HLA) loci and that display tumor-associated antigens (TAA) or antigenic precursors [tumor antigen precursor proteins (TAPP)] could be used for generating whole tumor cell vaccines or, alternatively, for extraction of TAA peptides to make autologous dendritic cell vaccines. EXPERIMENTAL DESIGN: Twenty human glioma cell lines were characterized by molecular phenotyping and by flow cytometry for HLA class I antigen expression. Twelve of the 20 cell lines, as well as analyses of freshly resected glioma tissues, were further characterized for protein and/or mRNA expression of 16 tumor antigen precursor proteins or TAA. RESULTS: These 20 human glioma cell lines potentially cover 77%, 85%, and 78% of the U.S. Caucasian population at HLA-A, HLA-B, and HLA-C alleles, respectively. All cells exhibited multiple TAA expressions. Most glioma cells expressed antigen isolated from immunoselected melanoma-2 (Aim-2), B-cyclin, EphA2, GP100, beta1,6-N-acetylglucosaminyltransferase V (GnT-V), IL13Ralpha2, Her2/neu, hTert, Mage, Mart-1, Sart-1, and survivin. Real-time PCR technology showed that glioblastoma specimens expressed most of the TAA as well. Tumor-infiltrating lymphocytes and CD8(+) CTL killed T2 cells when loaded with specific HLA-A2(+) restricted TAA, or gliomas that were both HLA-A2(+) and also positive for specific TAA (Mart-1, GP100, Her2/neu, and tyrosinase) but not those cells negative for HLA-A2 and/or lacking the specific epitope. CONCLUSIONS: These data provide proof-in-principle for the use of allogeneic, partially HLA patient-matched glioma cells for vaccine generation or for peptide pulsing with allogeneic glioma cell extracts of autologous patient dendritic cells to induce endogenous CTL in brain tumor patients.