Improving Outcomes in Infants With Neonatal Abstinence Syndrome With the Eat, Sleep, Console Method.

BACKGROUND: Neonatal abstinence syndrome (NAS) is a significant public health concern. A quality improvement project was executed in a neonatal intensive care unit at a large urban hospital. The aim was to address the prolonged hospitalization of infants and exposure to medications to treat NAS. PURPOSE: The goal was to determine whether the eat, sleep, console (ESC) method decreases the length of stay (LOS) and morphine usage when compared with the Finnegan Neonatal Abstinence Scoring System (FNASS). METHODS: The inclusion criteria were 36 weeks' or longer gestation and exposure to opiates in utero. The FNASS method was replaced by the ESC method with a refocus on nonpharmacologic care. Data were collected for 6 months during implementation of the ESC method and compared with the 6 months prior to implementation. RESULTS: The results of the project include: the average LOS decreased from 25.9 days to 13.7 days, a 47% reduction; the rate of scheduled morphine initiation decreased from 58% to 7%, an 88% reduction; as-needed morphine initiation decreased from 33% to 7%, a 79% reduction; and the rate of adjunctive medication initiation decreased from 17% to 0%, a 100% reduction. IMPLICATIONS FOR PRACTICE AND RESEARCH: The outcomes of LOS and rate of morphine usage were significantly improved when using the ESC method when compared with the FNASS at this facility. The results support future implications including expanding the ESC program to the well newborn population at this facility and other similar units. Further research needs to be done on long-term neurodevelopmental outcomes.

Extensive Allelic Diversity of MHC Class I in Wild Mallard Ducks.

MHC class I is critically involved in defense against viruses, and diversity from polygeny and polymorphism contributes to the breadth of the immune response and health of the population. In this article, we examine MHC class I diversity in wild mallard ducks, the natural host and reservoir of influenza A viruses. We previously showed domestic ducks predominantly use UAA, one of five MHC class I genes, but whether biased expression is also true for wild mallards is unknown. Using RT-PCR from blood, we examined expressed MHC class I alleles from 38 wild mallards (Anas platyrhynchos) and identified 61 unique alleles, typically 1 or 2 expressed alleles in each individual. To determine whether expressed alleles correspond to UAA adjacent to TAP2 as in domestic ducks, we cloned and sequenced genomic UAA-TAP2 fragments from all mallards, which matched transcripts recovered and allowed us to assign most alleles as UAA Allelic differences are primarily located in α1 and α2 domains in the residues known to interact with peptide in mammalian MHC class I, suggesting the diversity is functional. Most UAA alleles have unique residues in the cleft predicting distinct specificity; however, six alleles have an unusual conserved cleft with two cysteine residues. Residues that influence peptide-loading properties and tapasin involvement in chicken are fixed in duck alleles and suggest tapasin independence. Biased expression of one MHC class I gene may make viral escape within an individual easy, but high diversity in the population places continual pressure on the virus in the reservoir species.

Expression of duck CCL19 and CCL21 and CCR7 receptor in lymphoid and influenza-infected tissues.

Ducks are the natural host and reservoir of influenza viruses. We are interested in their immune responses to these viruses, to understand host-pathogen interactions and to develop effective agricultural vaccines. We identified duck homologues of the chemokines CCL19 and CCL21 and cloned their cognate receptor, CCR7. Conservation of key features, and expression in lymphoid tissues suggests that these chemokines are the direct orthologues of their mammalian counterparts. Mammalian CCL19 and CCL21 are responsible for the homing of dendritic cells and naïve lymphocytes to secondary lymphoid tissues. The contribution of local tertiary lymphoid tissues may be important during influenza infection in ducks. Consistent with leukocyte recruitment, CCL19 and CCL21 transcripts are abundant in lung tissues at 1 day post-infection with highly pathogenic avian influenza A/Vietnam/1203/04 (H5N1) (VN1203). In contrast, expression in lung or intestine tissues infected with low pathogenic A/mallard/BC/500/05 (H5N2) (BC500) is not significant. Recruitment and aggregation of leukocytes is visible in the vicinity of major airways 3 days after infection with VN1203. Chemokine gene expression may serve as a useful marker to evaluate duck immune responses to natural infections and vaccine strains.

Dendritic cell inhibitory and activating immunoreceptors (DCIR and DCAR) in duck: Genomic organization and expression.

C-type lectin immunoreceptor genes encoding DC inhibitory and activating receptors (DCIR and DCAR) were identified in a spleen EST library of duck (Anas platyrhynchos). These receptors are of interest for their potential as regulators of antigen presenting cells. A genomic clone was isolated and fully sequenced, containing one DCIR gene and two DCAR genes arranged in tandem order. Duck DCIR encodes an inhibitory receptor that features an immunoreceptor tyrosine-based inhibitory motif (ITIM) in the cytoplasmic domain. DCAR1 is a pseudogene. DCAR2 encodes an activating receptor with a positively charged residue in the transmembrane region. Full-length and alternatively spliced forms of both DCIR and DCAR2 are apparent. Duck DCIR and DCAR transcripts are preferentially expressed in immune and mucosal tissues including spleen, bursa of Fabricius, intestine and lung. Targeting these receptors on dendritic cells holds potential for breaking tolerance or for enhancing immune responses, relevant to the duck model for hepatitis B and vaccination against avian influenza.

The MHC of the duck (Anas platyrhynchos) contains five differentially expressed class I genes.

MHC class I proteins mediate a variety of functions in antiviral defense. In humans and mice, three MHC class I loci each contribute one or two alleles and each can present a wide variety of peptide Ags. In contrast, many lower vertebrates appear to use a single MHC class I locus. Previously we showed that a single locus was predominantly expressed in the mallard duck (Anas platyrhynchos) and that locus was adjacent to the polymorphic transporter for the Ag-processing (TAP2) gene. Characterization of a genomic clone from the same duck now allows us to compare genes to account for their differential expression. The clone carried five MHC class I genes and the TAP genes in the following gene order: TAP1, TAP2, UAA, UBA, UCA, UDA, and UEA. We designated the predominantly expressed gene UAA. Transcripts corresponding to the UDA locus were expressed at a low level. No transcripts were found for three loci, UBA, UCA, and UEA. UBA had a deletion within the promoter sequences. UCA carried a stop codon in-frame. UEA did not have a polyadenylation signal sequence. All sequences differed primarily in peptide-binding pockets and otherwise had the hallmarks of classical MHC class I alleles. Despite the presence of additional genes in the genome, the duck expresses predominantly one MHC class I gene. The limitation to one expressed MHC class I gene may have functional consequences for the ability of ducks to eliminate viral pathogens, such as influenza.

The dominant MHC class I gene is adjacent to the polymorphic TAP2 gene in the duck, Anas platyrhynchos.

We are investigating the expression and linkage of major histocompatibility complex (MHC) class I genes in the duck ( Anas platyrhynchos) with a view toward understanding the susceptibility of ducks to two medically important viruses: influenza A and hepatitis B. In mammals, there are multiple MHC class I loci, and alleles at a locus are polymorphic and co-dominantly expressed. In contrast, in lower vertebrates the expression of one locus predominates. Southern-blot analysis and amplification of genomic sequences suggested that ducks have at least four loci encoding MHC class I. To identify expressed MHC genes, we constructed an unamplified cDNA library from the spleen of a single duck and screened for MHC class I. We sequenced 44 positive clones and identified four MHC class I sequences, each sharing approximately 85% nucleotide identity. Allele-specific oligonucleotide hybridization to a Northern blot indicated that only two of these sequences were abundantly expressed. In chickens, the dominantly expressed MHC class I gene lies adjacent to the transporter of antigen processing ( TAP2) gene. To investigate whether this organization is also found in ducks, we cloned the gene encoding TAP2 from the cDNA library. PCR amplification from genomic DNA allowed us to determine that the dominantly expressed MHC class I gene was adjacent to TAP2. Furthermore, we amplified two alleles of the TAP2 gene from this duck that have significant and clustered amino acid differences that may influence the peptides transported. This organization has implications for the ability of ducks to eliminate viral pathogens.

THE EVOLUTION OF PARASITES FROM THEIR HOSTS: A CASE STUDY IN THE PARASITIC RED ALGAE.

Morphological similarities of many parasites and their hosts have led to speculation that some groups of plant, animal, fungal, and algal parasites may have evolved directly from their hosts. These parasites, which have been termed adelphoparasites in the botanical literature, and more recently, agastoparasites in the insect literature, may evolve monophyletically from one host and radiate secondarily to other hosts or, these parasites may arise polyphyletically, each arising from its own host. In this study we compare the internal transcribed spacer regions of the nuclear ribosomal repeats of species and formae specialis (host races) included in the red algal parasite genus Asterocolax with its hosts, which all belong to the Phycodrys group of the Delesseriaceae and with closely related nonhost taxa of the Delesseriaceae. These analyses reveal that species of Asterocolax have evolved polyphyletically. Asterocolax erythroglossi from the North Atlantic host Erythroglossum laciniatum appears to have evolved from its host, whereas taxa included in the north Pacific species Asterocolax gardneri have had two independent origins. Asterocolax gardneri from the host Polyneura latissima probably arose directly from this host. In contrast, all other A. gardneri formae specialis appear to have originated from either Phycodrys setchellii or P. isabelliae and radiated secondarily onto other closely related taxa of the Phycodrys group, including Nienburgia andersoniana and Anisocladella pacifica. Gamete crossing experiments confirm that A. gardneri from each host is genetically isolated from both its host, and from other A. gardneri and their hosts. Cross-infection experiments reveal that A. gardneri develops normally only on its natural host, although some abberrant growth may occur on alternate hosts. The ability of red algal parasites to radiate secondarily to other red algal taxa, where they may become isolated genetically and speciate, suggests that this process of speciation is not a "genetic dead end" but one that may give rise to related clusters of parasite species.