Andrzej Gaweł

Highly Sensitive Detection of Cancer Antigen 15-3 Using Novel Avian IgY Antibodies

Early detection of cancer development is crucial for successful therapy and for monitoring patient outcome. Various immunodiagnostic methods are able to detect pathological changes in the human body ahead of symptomatic manifestation of the disease. Most immunological examinations are based on the detection of specific tumor markers in body fluids. Of the various cancer-specific proteins used for breast cancer diagnostics, one of the most commonly applied is the cancer antigen 15-3 (CA 15-3). An elevation in its serum level (>25-40 U/ml) usually correlates with tumor malignancy. The CA 15-3 antigen is also used for monitoring patients after surgical treatment and for measuring therapeutic efficacy. Herein, we present the generation of polyclonal IgY antibodies isolated from egg yolks of immunized hens and their application for CA 15-3 detection. The developed sandwich ELISA assay showed a detection limit of 0.028 U/ml, thus demonstrating its potential for clinical applications.

The development and evaluation of cross-priming amplification for the detection of avian reovirus.

The aim of this study was to develop and evaluate cross‐priming amplification (CPA) for the detection of avian reovirus (ARV).

Pharmacokinetics of Repeated Sodium Salicylate Administration to Laying Hens: Evidence for Time Dependent Increase in Drug Elimination from Plasma and Eggs

Salicylates were the first non-steroid anti-inflammatory drugs (NSAIDs) to be used in any species and are still widely used in humans and livestock. However, the data on their pharmacokinetics in animals is limited, especially after repeated administration. Evidence exist that in chickens (Gallus gallus) salicylate (SA) may induce its own elimination. The aim of this study was to investigate salicylate pharmacokinetics and egg residues during repeated administration of sodium salicylate (SS) to laying hens. Pharmacokinetics of SA was assessed during 14 d oral administration of SS at daily doses of 50 mg/kg and 200 mg/kg body weight to laying hens. On the 1st, 7th and 14th d a 24 h-long pharmacokinetic study was carried out, whereas eggs were collected daily. Salicylate concentrations in plasma and eggs were determined using high-performance liquid chromatography with ultraviolet detection and pharmacokinetic variables were calculated using a non-compartmental model. Mean residence time (MRT), minimal plasma concentration (Cmin, C16h) and elimination half-life (T1/2el) of SA showed gradual decrease in layers administered with a lower dose. Total body clearance (ClB) increased. Layers administered with the higher dose showed a decrease only in the T1/2el. In the low dose group, SA was found only in the egg white and was low throughout the experiment. Egg whites from the higher dose group showed initially high SA levels which significantly decreased during the experiment. Yolk SA levels were lower and showed longer periods of accumulation and elimination. Repeated administration of SS induces SA elimination, although this effect may differ depending on the dose and production type of a chicken. Decreased plasma drug concentration may have clinical implications during prolonged SS treatment.

Generation and application of polyclonal IgY antibodies specific for full-length and nicked prostate-specific antigen.

BACKGROUND The prostate-specific antigen (PSA) is considered an important serum marker for prostate cancer detection, monitoring and staging. The purpose of this study was to generate IgY class antibodies that recognize native PSA and selected epitopes. METHODOLOGY Hens immunized with either full-length human PSA or its peptidyl fragment-conjugates produced specific antibodies that were isolated from egg yolks. We developed a monoclonal/IgY sandwich ELISA with a PSA detection limit of 50 pg/ml and a linear range of 0.05-1.0 ng/ml. CONCLUSION Because the signal observed for the PSA-specific IgY antibodies by ELISA and the reactivity profile of the epitope-derived IgYs were comparable to those of mouse monoclonal IgG antibodies, avian antibodies may be a cost-effective alternative to mammalian antibodies for prostate cancer diagnostics.

Hemorrhagic Nephritis and Enteritis in a Goose Flock in Poland—Disease Course Analysis and Characterization of Etiologic Agent

SUMMARY Hemorrhagic nephritis enteritis of geese (HNEG) is an epizootic viral disease caused by infection with goose hemorrhagic polyomavirus (GHPV) that affects domestic geese. This study describes the epizootic analysis, laboratory diagnosis, and molecular characterization of GHPV isolates associated with HNEG cases in Poland. HNEG symptoms persisted in infected flocks for 2 wk with a 32% mortality rate. Primary gross lesions included hemorrhaging of the kidneys, intestines, and lungs. Histopathologic examination confirmed HNEG and identified that the causative agent was similar to other GHPV isolates and identical to the Toulouse 2008 isolate. RESUMEN Nefritis y enteritis hemorrágica en una parvada de gansos en Polonia – Análisis del curso de la enfermedad y caracterización del agente etiológico. La nefritis y enteritis de los gansos (con las siglas en inglés HNEG) es una enfermedad viral epizoótica causada por la infección con poliomavirus hemorrágico del ganso (GHPV) que afecta a los gansos domésticos. Este estudio describe el análisis epizoótico, el diagnóstico de laboratorio y la caracterización molecular de los aislamientos del virus de la nefritis y enteritis de los gansos asociados a casos de la enfermedad en Polonia. Los signos de la nefritis y enteritis de los gansos persistieron en las parvadas infectadas por dos semanas con una tasa de mortalidad del 32%. Lesiones macroscópicas primarias incluyeron hemorragia de los riñones, los intestinos y los pulmones. El examen histopatológico confirmó la enfermedad y se identificó que el agente causante era similar a otros aislamientos del virus de la nefritis y enteritis de los gansos e idéntico al aislamiento de Toulouse del año 2008.

Ascaridia galli isolates with ITS1-5.8rRNA-ITS2 fragment homologous to Ascaridia columbae

Ascaridia (A.) galli is one of the most commonly occurring nematodes in poultry worldwide, often in hens and broiler chickens. The infection with Ascaridia galli in free-range chickens was even 70%. There is not much information about A. galli genetic features. The present study was conducted to assess the genetic diversity of A. galli isolated from hens in Poland by analyzing the nucleotide sequence of the region ITS1-5.8rRNA-ITS2 and to define its homology within the family Ascaridiidae. Adult A. galli were collected from the intestines of naturally infected hens from two flocks of free-run laying hens from the Wielkopolska region in Poland. From all parasites an identical ITS1-5.8rRNA-ITS2 sequence was obtained, which was homologous in 99% with A. columbae (JQ995321.1) sequence. The high homology sequences of A. galli (KX683286) from Poland and A. columbae (JQ995321.1) isolate from the USA, support the observations of other authors suggesting that A. galli and A. columbae might be closely related. It is the first whole ITS1-5.8rRNA-ITS2 of A. galli in the GenBank database, so there is not enough data for detailed phylogenetic analysis of A. galli. Detailed genetic analysis is necessary to get better insight into the birds’ Ascaridia species.

Adjuvant-dependent immunogenicity of Staphylococcus aureus Efb and Map proteins in chickens

The avian IgY antibodies generated in hens and isolated from egg yolk have gained in popularity as they present an alternative source of antibodies for diagnostic as well as therapeutic applications. One of the advantages of IgY technology are the large amounts of produced antibodies from a single animal combined with their high reactivity representing an attractive alternative for mammalian antibodies. Despite many known protocols for the immunization of chickens, the administration of new antigens often requires additional modification such as antigen dose or use of an adjuvant in order to elicit a significant immune response. We investigated the immunogenicity of three Staphylococcus aureus antigens including two extracellular proteins Map and Efb and one selected Efb105-124 epitope conjugated to KLH that were administered to the animals. Additionally, the immunization protocol included two adjuvant systems: Freunds complete adjuvant and Emulsigen-D. The results demonstrated a high immunostimulatory potency of Freunds complete adjuvant, especially in case of Efb compared to the immune response elicited by Emulsigen-D. However, after immunization with the KLH-Efb105-124 conjugate, the obtained antibodies showed similar reactivity regardless of adjuvant system used with the only exception being their avidity.

Erysipelas Outbreaks in Flocks of Geese in Poland—Biochemical and Genetic Analyses of the Isolates

SUMMARY Erysipelothrix rhusiopathiae causes erysipelas in many vertebrate species. Severe disease outbreaks have been noted in many poultry species—chickens, ducks, emus, pheasants, pigeons, and geese. This article describes the biochemical and genetic analyses of six E. rhusiopathiae strains isolated from geese for meat production. The isolates came from birds kept in different poultry houses on one farm, and were collected during two erysipelas outbreaks. We analyzed and compared the isolates by random amplified polymorphic DNA with the use of NK6 primer and pulsed-field gel electrophoresis, with the restriction enzyme SmaI. Biochemical examination was performed with API Coryne test. Analyses showed that the three strains isolated during the first outbreak differed, whereas the three isolates from the second outbreak were identical to one another, but distinct from the isolates from the first outbreak. The results of biochemical and genetic analyses of E. rhusiopathiae strains isolated from geese suggest different sources of infection for the erysipelas outbreaks.