Effect of intestinal microbiota on duck short-beak and dwarf syndrome caused by novel goose parvovirus.

Short-beak and dwarf syndrome (SBDS) is caused by infection with novel goose parvovirus (NGPV), which leads to intestinal dysbiosis, developmental delay, short beak, lameness, and paralysis in ducks and is the cause of skeletal health problems. NGPV infection can cause intestinal microbial disturbances, but it is still unclear whether the intestinal microbiota affects the pathogenicity of NGPV. Here, the effects of intestinal microbiota on NGPV-induced SBDS in Cherry Valley ducks were assessed by establishing a duck model for gut microflora depletion/reestablishment through antibiotics (ABX) treatment/fecal microbiota transplanted (FMT). By measuring body weight, beak length, beak width and tarsal length, we found that SBDS clinical symptoms were alleviated in ducks treated with ABX, but not in FMT ducks. Next, we conducted a comprehensive analysis of bone metabolism, gut barrier integrity, and inflammation levels using quantitative real-time PCR (qPCR), enzyme linked immunosorbent assay (ELISA), biochemical analysis and histological analysis. The results showed that ABX treatment improved bone quality reduced bone resorption, mitigated tissue lesions, protected intestinal barrier integrity, and inhibited systemic inflammation in NGPV-infected ducks. Moreover, cecal microflora composition and short-chain fatty acids (SCFAs) production were examined by bacterial 16S rRNA sequencing and gas chromatography. The results revealed that ABX treatment mitigated the decreased abundance of Firmicutes and Bacteroidota in NGPV-infected ducks, as well as increased SCFAs production. Furthermore, ABX treatment reduced the mucosa-associated lymphoid tissue lymphoma translocation protein 1 (Malt1) and nuclear factor κB (NF-κB) expression, which are correlated with systemic inflammation in SBDS ducks. These findings suggested that intestinal microflora depletion alleviated NGPV-induced SBDS by maintaining intestinal homeostasis, inhibiting inflammatory response and alleviating bone resorption. These results provide evidence for the pivotal role of intestinal microbiota in the process of SBDS and contribute a theoretical basis for the feasibility of microecological preparation as a method to control SBDS.

Preparation of Monoclonal Antibodies against the Capsid Protein and Development of an Epitope-Blocking Enzyme-Linked Immunosorbent Assay for Detection of the Antibody against Porcine Circovirus 3.

Porcine circovirus type 3 (PCV3) is endemic in swine worldwide and causes reproductive disorders, dermatitis and nephrotic syndrome, and multi-organ inflammation. Currently, there is a growing need for rapid and accurate diagnostic methods in disease monitoring. In this study, four monoclonal antibodies (mAbs) against PCV3 capsid proteins were prepared (mAbs 2F6, 2G8, 6E2, and 7E3). MAb 7E3, which had the highest binding affinity for the Cap protein, was chosen for further investigation. A novel B cell epitope 110DLDGAW115 was identified using mAb 7E3. An epitope-blocking (EB) enzyme-linked immunosorbent assay (ELISA) was successfully developed using horseradish-peroxidase-labeled mAb 7E3 to detect PCV3 antibodies in porcine sera. Moreover, the EB-ELISA showed no specific reaction with other porcine disease sera, and the cut-off value was defined as 35%. Compared with the commercial ELISA, the percentage agreement was 95.59%. Overall, we have developed a novel EB-ELISA method that accurately and conveniently detects PCV3 in serum, making it a valuable tool for the clinical detection of PCV3 infection.

Genetics etiologies and genotype phenotype correlations in a cohort of individuals with central conducting lymphatic anomaly.

Central conducting lymphatic anomaly (CCLA) is a heterogenous disorder caused by disruption of central lymphatic flow that may result in dilation or leakage of central lymphatic channels. There is also a paucity of known genetic diagnoses associated with CCLA. We hypothesized that specific genetic syndromes would have distinct lymphatic patterns and this would allow us to more precisely define CCLA. As a first step toward "precision lymphology", we defined the genetic conditions associated with CCLA by performing a retrospective cohort study. Individuals receiving care through the Jill and Mark Fishman Center for Lymphatic Disorders at the Children's Hospital of Philadelphia between 2016 and 2019 were included if they had a lymphangiogram and clinical genetic testing performed and consented to a clinical registry. In our cohort of 115 participants, 26% received a molecular diagnosis from standard genetic evaluation. The most common genetic etiologies were germline and mosaic RASopathies, chromosomal abnormalities including Trisomy 21 and 22q11.2 deletion syndrome, and PIEZO1-related lymphatic dysplasia. Next, we analyzed the dynamic contrast magnetic resonance lymphangiograms and found that individuals with germline and mosaic RASopathies, mosaic KRASopathies, PIEZO1-related lymphatic dysplasia, and Trisomy 21 had distinct central lymphatic flow phenotypes. Our research expands the genetic conditions associated with CCLA and genotype-lymphatic phenotype correlations. Future descriptions of CCLA should include both genotype (if known) and phenotype to provide more information about disease (gene-CCLA). This should be considered for updated classifications of CCLA by the International Society of Vascular Anomalies.

Liver lymphatic anatomy and role in systemic lymphatic disease.

OBJECTIVES: To characterize hepatic to systemic lymphatic connections in patients with systemic lymphatic disease using intra-hepatic lymphangiography and to compare outcomes after lymphatic intervention. METHODS: In this retrospective study, patients with intra-hepatic lymphangiography from May 2014 - April 2019 at our institution were included. Imaging review was performed and hepatic lymphatic connections and flow patterns were characterized. Clinical data were reviewed and comparisons between patients undergoing lymphatic intervention with or without abnormal hepatic lymphatics were performed. RESULTS: During the study period, 105 patients underwent intra-hepatic lymphangiography. Primary clinical presentation included ascites (19/105), chylothorax (27/105), plastic bronchitis (PB) (17/105), and protein losing enteropathy (PLE) (42/105). Five categories of hepatic lymphatic connections and flow patterns were identified (%): normal (25%, 26/105), hepatoperitoneal (12%, 13/105), hepatopulmonary (10.5%, 11/105), hepatomesenteric (7.5%, 8/105), and hepatoduodenal (41%, 43/105) with four patients having more than one abnormal pattern. A comparison between clinical presentation and imaging category revealed an increased likelihood of having ascites with hepatoperitoneal (p < .0001), chylothorax/PB with hepatopulmonary (p = .01), and PLE with hepatoduodenal (p < .001) connections. Seventy-six patients had a lymphatic intervention, 24% with normal, and 76% with abnormal liver lymphatics. There was no difference in length of hospital stay or mortality between the two groups, but there was a prolonged time to symptom resolution (p = .006) and persistent symptoms after 6 months (5% vs 44%, p = .002) in the group with abnormal liver lymphatics. CONCLUSION: We identified five liver lymphatic imaging categories with a substantial correlation to presenting lymphatic disease. Abnormal imaging patterns correlated with increased morbidity. Evaluation of liver lymphatics should be considered in patients with a systemic lymphatic disease if central lymphatic imaging is normal. KEY POINTS: • We identified five liver lymphatic imaging patterns: normal, hepatoperitoneal, hepatomesenteric, hepatopulmonary, and hepatoduodenal. • Imaging patterns were correlated with disease presentation (normal - chylothorax/PB, hepatoperitoneal - ascites/chylothorax, hepatopulmonary - chylothorax/PB, hepatoduodenal - PLE). • Abnormal imaging patterns correlated with increased morbidity.

Neonatal lymphatic flow disorders: impact of lymphatic imaging and interventions on outcomes.

Neonatal chylothorax (NCTx) and central lymphatic flow disorder (CLFD) are historically challenging neonatal disorders with high morbidity and mortality. METHODS: We conducted a retrospective study of 35 neonates with pulmonary lymphatic abnormalities at our institution who underwent lymphatic evaluation between December 2015 and September 2018. Patients with only pulmonary lymphatic perfusion syndrome were classified as NCTx and those with multiple flow abnormalities were classified as CLFD. Demographics, clinical characteristics, and outcomes were compared using t-tests/Wilcoxon rank sum tests and Fisher's exact tests. RESULTS: All 35 patients had intranodal MR lymphangiography and 14 (40%) also had conventional fluoroscopic lymphangiography. Fifteen (42.8%) patients were diagnosed with NCTx and 20 (57.1%) were diagnosed with CLFD. Thirty-four (97.1%) patients had pleural effusions. None of the NCTx group had ascites, anasarca, or dermal backflow compared to 17 (85%) (p < 0.001), 8 (42.1%) (p: 0.004), and 20 (100%) (p < 0.001) of the CLFD group, respectively. In the NCTx group, 11 (73.3%) had ethiodized oil embolization and 4 (26.7%) received conservative therapy. Ten (50%) of the CLFD patients had an intervention; of those, two (10%) had ethiodized oil-only embolization. Eight had non-ethiodized oil embolizations (two (25%) had embolization with glue, three (37.5%) underwent surgical lymphovenous anastomosis, two (25%) underwent thoracic duct (TD) externalization, and one (12.5%) had a non-TD lymphatic channel drain placed). Complete resolution of pleural effusions was achieved in all 15 NCTx patients, whereas 9 (45%) of 20 CLFD patients had resolution of chylothorax (p: 0.001). CONCLUSIONS: Establishing a diagnosis of NCTx or CLFD is paramount in selecting treatment options and providing prognostic information. Development of lymphatic interventions represents a paradigm shift in our understanding of neonatal lymphatic flow disorders and may be associated with improved survival.

Immunogenicity and protective efficacy of enterotoxigenic Escherichia coli (ETEC) total RNA against ETEC challenge in a mouse model.

Enterotoxigenic Escherichia coli (ETEC), an essential cause of post-weaning diarrhea (PWD) in piglets, leads to significant economic losses to the pig industry. The present study aims to identify the role of ETEC total RNA in eliciting immune responses to protect animals against ETEC infection. The results showed that the total RNA isolated from pig-derived ETEC K88ac strain effectively stimulated the IL-1ß secretion of porcine intestinal epithelial cells (IPEC-J2). The mouse model immunized with ETEC total RNA via intramuscular injection (IM) or oral route (OR) was used to evaluate the protective efficiency of the ETEC total RNA. The results suggested that 70 µg ETEC total RNA administered by either route significantly promoted the production of the serum IL-1ß and K88ac specific immunoglobulins (IgG, IgM, and IgA). Besides, the ETEC RNA administration augmented strong mucosal immunity by elevating K88ac specific IgA level in the intestinal fluid. Intramuscularly administered RNA induced a Th1/Th2 shift toward a Th2 response, while the orally administered RNA did not. The ETEC total RNA efficiently protected the animals against the ETEC challenge either by itself or as an adjuvant. The histology characterization of the small intestines also suggested the ETEC RNA administration protected the small intestinal structure against the ETEC infection. Particularly of note was that the immunity level and protective efficacy caused by ETEC RNA were dose-dependent. These findings will help understand the role of bacterial RNA in eliciting immune responses, and benefit the development of RNA-based vaccines or adjuvants.

Use of Contrast-Enhanced Ultrasound to Determine Thoracic Duct Patency.

The aim of this study was to determine the feasibility of using contrast-enhanced ultrasound (CEUS) evaluation to determine thoracic duct (TD) outlet patency. Nine patients referred for lymphatic imaging and intervention underwent percutaneous intranodal ultrasound contrast injection and conventional lymphangiography (CL). Eight of 9 patients had a patent TD by CEUS and CL. One patient did not have a patent TD. There was 100% agreement between CEUS and CL. These results suggest that CEUS is an imaging modality that might be as accurate as CL in determining TD patency.