The use of a pediatric appendicitis pathway in a large integrated health system reduced computed tomography imaging in the ED.

BACKGROUND: Appendicitis is the most common cause of an acute surgical abdomen in children. Diagnosis is often challenging as few pediatric patients present with classic symptoms. Clinicians are thus dependent on imaging to reach an accurate diagnosis. Although computerized tomography (CT) has high sensitivity and specificity, it has the disadvantage of imparting ionizing radiation. Ultrasound (US) is readily available and has comparable accuracy to CT when performed by experienced sonographers. We sought to examine the impact of a system-wide process improvement plan on CT use and other metrics in pediatric patients who presented to the Emergency Department (ED) with suspected appendicitis. METHODS: This is a retrospective study of the impact of a Pediatric Appendicitis Pathway (PAP) within a large integrated hospital system with 12 EDs including 3 designated hub EDs. Patients were placed in an initial risk category utilizing the Pediatric Appendicitis Score (PAS), and received US of the appendix at a hub ED if indicated by the PAS. Patients presenting to community EDs who required US appendix were transferred to hub EDs for imaging. Patients presenting in the 6-month pre-implementation period were compared to patients presenting in a 14-month post-implementation period on CT and US utilization, negative and missed appendectomy rates, and ED length of stay (LOS). RESULTS: 1874 patients (401 pre-PAP and 1473 post-PAP) were included in the study. At the hub EDs the rate of CT imaging for suspected appendicitis was reduced from 31% to 17% with a resultant increase in US utilization from 83% (333/401) to 90% (1331/1473) (p < 0.001). At community general EDs (404 pre-PAP and 449 post-PAP), the rate of CT was decreased from 45% (181/404) to 32%(144/449) (p < 0.001)) There was no significant change in the negative appendectomy rate pre-PAP (1/59 = 1.7%) and post-PAP (4/168 = 2.4%) (p = 0.99) at the hub EDs. There were no missed appendicitis cases after PAP implementation compared to 1 case in the pre-PAP period. Overall LOS was similar pre and post-PAP, however LOS was longer in patients that required transfer from community general EDs to hub EDs (median 264 vs 342 min, p < 0.001). CONCLUSIONS: A PAP that stratified patients into risk groups using the PAS and encouraged the use of US as a first line imaging modality, reduced the number of CT performed in a large integrated health system without significant changes to clinical outcomes. Furthermore, transferring select patients for an US as opposed to obtaining an initial CT in community general EDs was feasible and reduced CT use in the pediatric population.

A radiolabeled oligonucleotide ligation assay demonstrates the high frequency of nevirapine resistance mutations in HIV type 1 quasispecies of NVP-treated and untreated mother-infant pairs from Uganda.

This study explores the levels of NVP and AZT resistance mutations in untreated, NVP- or AZT-treated mother-infant pairs in Uganda. PCR-amplified reverse transcriptase (RT) gene fragments derived from PBMC samples of 85 mothers (10 AZT treated, 35 NVP treated, and 40 untreated) and their 52 infected infants (5 AZT, 9 NVP, and 38 untreated) were classified as subtype A (59%), D (29%), C (3%), and recombinant forms (9%) by population sequencing. Only 16% of the NVP-treated infected mothers and infants harbored either the K103N or the Y181C at 6 weeks postdelivery. The majority of these samples (n = 107) were then analyzed using a radiolabeled oligonucleotide ligation assay (OLA) specific for K70R, K103N, and Y181C, using nonstandard bases to accommodate sequence heterogeneity. By OLA, 43% of the NVP-treated group had K103N and/or Y181C mutations in their HIV-1 population, using >0.6% cutoff based on a comparative clonal analysis of clinical isolates. Surprisingly, an equal fraction of the untreated and NVP-treated mother-infant group had the K103N mutation in their HIV-1 population in the range of 0.6-5%. These findings suggest a relatively high frequency of K103N mutation in the drug-naive, subtype A and D infected Ugandan population as compared to the very low frequency of the Y181C and K70R mutation (<0.6%). The prevalence of the K103N mutations may be related to its low fitness cost and high genetic stability. The persistence of these mutations may reduce the effectiveness of subsequent NVP use in treatment or prevention of perinatal transmission.

Changes in human immunodeficiency virus type 1 fitness and genetic diversity during disease progression.

This study examined the relationship between ex vivo human immunodeficiency virus type 1 (HIV-1) fitness and viral genetic diversity during the course of HIV-1 disease. Primary HIV-1 isolates from 10 patients at different time points were competed against control HIV-1 strains in peripheral blood mononuclear cell (PBMC) cultures to determine relative fitness values. Patient HIV-1 isolates sequentially gained fitness during disease at a significant rate that directly correlated with viral load and HIV-1 env C2V3 diversity. A loss in both fitness and viral diversity was observed upon the initiation of antiretroviral therapy. A possible relationship between genotype and phenotype (virus replication efficiency) is supported by the parallel increases in ex vivo fitness and viral diversity during disease, of which the correlation is largely based on specific V3 sequences. Syncytium-inducing, CXCR4-tropic HIV-1 isolates did have higher relative fitness values than non-syncytium-inducing, CCR5-tropic HIV-1 isolates, as determined by dual virus competitions in PBMC, but increases in fitness during disease were not solely powered by a gradual switch in coreceptor usage. These data provide in vivo evidence that increasing HIV-1 replication efficiency may be related to a concomitant increase in HIV-1 diversity, which in turn may be a determining factor in disease progression.

Differences in the fitness of two diverse wild-type human immunodeficiency virus type 1 isolates are related to the efficiency of cell binding and entry.

The ability of one primary human immunodeficiency virus type 1 (HIV-1) isolate to outcompete another in primary CD4+ human lymphoid cells appears to be mediated by the efficiency of host cell entry. This study was designed to test the role of entry on fitness of wild-type HIV-1 isolates (e.g., replicative capacity) and to examine the mechanism(s) involved in differential entry efficiency. The gp120 coding regions of two diverse HIV-1 isolates (the more-fit subtype B strain, B5-91US056, and less-fit C strain, C5-97ZA003) were cloned into a neutral HIV-1 backbone by using a recently described yeast cloning technique. The fitness of the primary B5 HIV-1 isolates and its env gene cloned into the NL4-3 laboratory strain had similar fitness, and both were more fit than the C5 primary isolate and its env/NL4-3 chimeric counterpart. Increased fitness of the B5 over C5 virus was mediated by the gp120 coding region of the env gene. An increase in binding/fusion, as well as decreased sensitivity to entry inhibitors (PSC-RANTES and T-20), was observed in cell fusion assays mediated by B5 gp120 compared to C5 gp120. Competitive binding assays using a novel whole virus-cell system indicate that the primary or chimeric B5 had a higher avidity for CD4/CCR5 on host cells than the C5 counterpart. This increased avidity of an HIV-1 isolate for its cell receptors may be a significant factor influencing overall replicative capacity or fitness.

Relationships between infectious titer, capsid protein levels, and reverse transcriptase activities of diverse human immunodeficiency virus type 1 isolates.

Most studies on human immunodeficiency virus type 1 (HIV-1) replication kinetics or fitness must rely on a particular assay to initially standardize inocula from virus stocks. The most accurate measure of infectious HIV-1 titers involves a limiting dilution-infection assay and a calculation of the dose required for 50% infectivity of susceptible cells in tissue culture (TCID(50)). Surrogate assays are now commonly used to measure the amount of p24 capsid, the endogenous reverse transcriptase (RT) activity, or the amount of viral genomic RNA in virus particles. However, a direct comparison of these surrogate assays and actual infectious HIV-1 titers from TCID(50) assays has not been performed with even the most conserved laboratory strains, let alone the highly divergent primary HIV-1 isolates of different subtypes. This study indicates that endogenous RT activity, not p24 content or viral RNA load, is the best surrogate measure of infectious HIV-1 titer in both cell-free supernatants and viruses purified on sucrose cushions. Sequence variation between HIV-1 subtypes did not appear to affect the function or activity of the RT enzyme in this endogenous assay but did affect the detection of p24 capsid by both enzyme immunoassays and Western blots. Clear groupings of non-syncytium-inducing (NSI), CCR5-tropic (R5), and SI/CXCR4-tropic (X4) HIV-1 isolates were observed when we compared the slopes derived from correlations of RT activity with infectious titers. Finally, the replication efficiency or fitness of both the NSI/R5 and SI/X4 HIV-1 isolates was not linked to the titers of the virus stocks.