Esophageal Surveillance Practices in Esophageal Atresia Patients: A Survey by the Eastern Pediatric Surgery Network.

INTRODUCTION: Endoscopic surveillance guidelines for patients with repaired esophageal atresia (EA) rely primarily on expert opinion. Prior to embarking on a prospective EA surveillance registry, we sought to understand EA surveillance practices within the Eastern Pediatric Surgery Network (EPSN). METHODS: An anonymous, 23-question Qualtrics survey was emailed to 181 physicians (surgeons and gastroenterologists) at 19 member institutions. Likert scale questions gauged agreement with international EA surveillance guideline-derived statements. Multiple-choice questions assessed individual and institutional practices. RESULTS: The response rate was 77%. Most respondents (80%) strongly agree or agree that EA surveillance endoscopy should follow a set schedule, while only 36% claimed to perform routine upper GI endoscopy regardless of symptoms. Many institutions (77%) have an aerodigestive clinic, even if some lack a multi-disciplinary EA team. Most physicians (72%) expressed strong interest in helping develop evidence-based guidelines. CONCLUSIONS: Our survey reveals physician agreement with current guidelines but weak adherence. Surveillance methods vary greatly, underscoring the lack of evidence-based data to guide EA care. Aerodigestive clinics may help implement surveillance schedules. Respondents support evidence-based protocols, which bodes well for care standardization. Results will inform the first multi-institutional EA databases in the United States (US), which will be essential for evidence-based care. LEVEL OF EVIDENCE: This is a prognosis study with level 4 evidence.

Gain-of-Function Lrp5 Mutation Improves Bone Mass and Strength and Delays Hyperglycemia in a Mouse Model of Insulin-Deficient Diabetes.

High fracture rate and high circulating levels of the Wnt inhibitor, sclerostin, have been reported in diabetic patients. We studied the effects of Wnt signaling activation on bone health in a mouse model of insulin-deficient diabetes. We introduced the sclerostin-resistant Lrp5A214V mutation, associated with high bone mass, in mice carrying the Ins2Akita mutation (Akita), which results in loss of beta cells, insulin deficiency, and diabetes in males. Akita mice accrue less trabecular bone mass with age relative to wild type (WT). Double heterozygous Lrp5A214V /Akita mutants have high trabecular bone mass and cortical thickness relative to WT animals, as do Lrp5A214V single mutants. Likewise, the Lrp5A214V mutation prevents deterioration of biomechanical properties occurring in Akita mice. Notably, Lrp5A214V /Akita mice develop fasting hyperglycemia and glucose intolerance with a delay relative to Akita mice (7 to 8 vs. 5 to 6 weeks, respectively), despite lack of insulin production in both groups by 6 weeks of age. Although insulin sensitivity is partially preserved in double heterozygous Lrp5A214V /Akita relative to Akita mutants up to 30 weeks of age, insulin-dependent phosphorylated protein kinase B (pAKT) activation in vitro is not altered by the Lrp5A214V mutation. Although white adipose tissue depots are equally reduced in both compound and Akita mice, the Lrp5A214V mutation prevents brown adipose tissue whitening that occurs in Akita mice. Thus, hyperactivation of Lrp5-dependent signaling fully protects bone mass and strength in prolonged hyperglycemia and improves peripheral glucose metabolism in an insulin independent manner. Wnt signaling activation represents an ideal therapeutic approach for diabetic patients at high risk of fracture. © 2021 The Authors. Journal of Bone and Mineral Research published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research (ASBMR).

Survival and Growth in vitro of Paramoeba perurans Populations Cultured Under Different Salinities and Temperatures.

Growth rates of Paramoeba perurans cultures under different temperature and salinity conditions were investigated in vitro over a 15day period. Optimal population growth, under the experimental conditions, was observed at 15°C and a salinity of 35‰, with amoebae populations doubling every 14h. Positive P. perurans populations growth was observed at 15°C between salinities of above 20‰ and 50‰, and at 8°C, 11°C and 18°C at salinities between 25‰ and 50‰, 50‰ being the maximum salinity tested. Amoebae numbers were sustained at 4°C. Therefore, lower temperature and salinity thresholds for P. perurans population growth lie between 4 to 8°C, and salinities of 20 to 25‰, respectively. Upper limits were not determined in this study. The populations remained relatively stable at 4°C and 2°C at permissive salinities with respect to numbers of viable amoebae over the 15day exposure period.

Individual Monitoring of Immune Response in Atlantic Salmon Salmo salar following Experimental Infection with Infectious Salmon Anaemia Virus (ISAV).

Monitoring the immune response in fish over the progression of a disease is traditionally carried out by experimental infection whereby animals are killed at regular intervals and samples taken. We describe here a novel approach to infectiology for salmonid fish where blood samples are collected repeatedly in a small group of PIT-tagged animals. This approach contributes to the reduction of animals used in research and to improved data quality. Two groups of 12 PIT-tagged Atlantic salmon (Salmo salar) were i.p infected with Infectious Salmon Anaemia Virus (ISAV) or culture medium and placed in 1 m3 tanks. Blood samples were collected at 0, 4, 8, 12, 16, 21 and 25 days post infection. The viral load, immune and stress response were determined in individual fish by real-time quantitative PCR (QPCR) on the blood cells, as well as the haematocrit used as an indicator of haemolysis, a clinical consequence of ISAV infection. "In-tank" anaesthesia was used in order to reduce the stress related to chase and netting prior to sampling. The data were analysed using a statistical approach which is novel with respect to its use in fish immunology. The repeated blood collection procedure did not induce stress response as measured by HSP70 and HSP90 gene expression in the un-infected animals. A strong increase in viraemia as well as a significant induction of Mx and γIP gene expression were observed in the infected group. Interleukin 10 was found induced at the later stage of the infection whereas no induction of CD8 or γ IFN could be detected. These results and the advantages of this approach are discussed.

Low virulent infectious salmon anaemia virus (ISAV) replicates and initiates the immune response earlier than a highly virulent virus in Atlantic salmon gills.

Observations from the field and experimental evidence suggest that different strains of infectious salmon anaemia virus (ISAV) can induce disease of varying severity in Atlantic salmon. Variation in host mortality and dissemination of ISAV isolates with high and low virulence was investigated using immersion challenge; from which mortality, pathological, immunohistochemical and preliminary molecular results have been previously published. Here, real-time RT-PCR analysis and statistical modelling have been used to further investigate variation in virus load and the response of four select immune genes. Expression of type I and II interferon (IFN), Mx and γIFN induced protein (γIP) to high and low pathogenic virus infection were examined in gill, heart and anterior kidney. In addition, a novel RNA species-specific assay targeting individual RNA types was used to investigate the separate viral processes of transcription and replication. Unexpectedly, the low virulent ISAV (LVI) replicated and transcribed more rapidly in the gills compared to the highly virulent virus (HVI). Subsequently LVI was able to disseminate to the internal organs more quickly and induced a more rapid systemic immune response in the host that may have offered some protection. Contrary to this, HVI initially progressed more slowly in the gills resulting in a slower generalised infection. However HVI ultimately reached a higher viral load and induced a greater mortality.

Multilocus variable-number tandem-repeat genotyping of Renibacterium salmoninarum, a bacterium causing bacterial kidney disease in salmonid fish.

BACKGROUND: Bacterial kidney disease (BKD), caused by Renibacterium salmoninarum, is a bacterial disease of fish, which is both geographically widespread and difficult to control. Previously, application of various molecular typing methods has failed to reliably discriminate between R. salmoninarum isolates originating from different host species and geographic areas. The current study aimed to utilize multilocus variable number tandem repeats (VNTR) to investigate inter-strain variation of R. salmoninarum to establish whether host-specific populations exist in Atlantic salmon and rainbow trout respectively. Such information would be valuable in risk assessment of transmission of R. salmoninarum in a multispecies aquaculture environment. RESULTS: The present analysis utilizing sixteen VNTRs distinguished 17 different haplotypes amongst 41 R. salmoninarum isolates originating from Atlantic salmon and rainbow trout in Scotland, Norway and the US. The VNTR typing system revealed two well supported groups of R. salmoninarum haplotypes. The first group included R. salmoninarum isolates originating from both Atlantic salmon and rainbow trout circulating in Scottish and Norwegian aquaculture, in addition to the type strain ATCC33209T originating from Chinook salmon in North America. The second group comprised isolates found exclusively in Atlantic salmon, of mainly wild origin, including isolates NCIB1114 and NCIB1116 associated with the original Dee disease in Scotland. CONCLUSIONS: The present study confirmed that VNTR analysis can be successfully applied to discriminate R. salmoninarum strains. There was no clear distinction between isolates originating from Atlantic salmon and rainbow trout as several haplotypes in group 1 clustered together R. salmoninarum isolates from both species. These findings indicate a potential exchange of pathogens between Atlantic salmon and rainbow trout in Scottish and Norwegian aquaculture during the last 20 years. In a scenario of expansion of rainbow trout farming into the marine environment, appropriate biosecurity measures to minimize disease occurrence are advised. The present results also suggest that R. salmoninarum isolates circulating in European aquaculture over the last 20 years are genetically distant to the wild strains originally causing BKD in the rivers Dee and Spey.

Development of an in vitro system to measure the sensitivity to the antiviral Mx protein of fish viruses.

Mx is a structural protein, induced by type I interferon (IFN), with direct antiviral properties. In fish the inherent contribution of Mx protein to viral protection is unknown. The transgenic Chinook salmon embryonic (CHSE)-TOF cell line was genetically modified to express the rainbow trout Mx (rbtMx1) protein under the control of the tetracycline derivative, doxycycline (DOX). Two clones CHSE-TOF-MX8 and CHSE-TOF-MX10 were isolated and characterised by qPCR. The level of resistance to Infectious Pancreatic Necrosis Virus (IPNV), Salmon Alphavirus (SAV), Infectious Haematopoietic Necrosis Virus (IHNV) and Epizootic Haematopoietic Necrosis Virus (EHNV) of the CHSE-TOF, CHSE-TOF-MX8 and CHSE-TOF-MX10 cell lines cultivated with and without DOX was measured. A novel method was established to measure accurately the level of sensitivity of any given viral isolate to Mx protein. IPNV and SAV viruses were highly sensitive to the presence of rbtMx1 in the cells whereas IHNV and EHNV showed partial resistance suggesting contrasting viral evasion strategies between these categories of viruses.

Modelling management strategies for a disease including undetected sub-clinical infection: bacterial kidney disease in Scottish salmon and trout farms.

Disease is a major constraint on animal production and welfare in agriculture and aquaculture. Movement of animals between farms is one of the most significant routes of disease transmission and is particularly hard to control for pathogens with subclinical infection. Renibacterium salmoninarum causes bacterial kidney disease (BKD) in salmonid fish, but infection is often sub-clinical and may go undetected with major potential implications for disease control programmes. A Susceptible-Infected model of R. salmoninarum in Scottish aquaculture has been developed that subdivides the infected phase between known and undetected sub-clinically infected farms and diseased farms whose status is assumed to be known. Farms officially known to be infected are subject to movement controls restricting spread of infection. Model results are sensitive to prevalence of undetected infection, which is unknown. However, the modelling suggests that controls that reduce BKD prevalence include improve biosecurity on farms, including those not known to be infected, and improved detection of infection. Culling appears of little value for BKD control. BKD prevalence for rainbow trout farms is less sensitive to controls than it is for Atlantic salmon farms and so different management strategies may be required for the sectors.