Biomedical subjects

D H Ley

Publications and source records attributed to D H Ley.

40 records · Page 3Linked to original sources

PubMedDate not supplied

Identification of Mycoplasma synoviae immunogenic surface proteins and their potential use as antigens in the enzyme-linked immunosorbent assay.

Specific immunogenic proteins of Mycoplasma synoviae (MS) strain WVU-1853 were purified and characterized by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and by Western transfer using anti-MS, anti-M. gallisepticum, and non-immune chicken sera. A cluster of prominent immunoreactive proteins with molecular masses from 46 to 52 kDa (p46-52) and less reactive single proteins of approximately 22 and 92 kDa were shown to partition into the detergent phase of Triton X-114 MS lysates, suggesting that these amphiphilic polypeptides are integral membrane proteins. Monoclonal antibodies, produced by immunizing mice with MS whole cell proteins and shown to bind species-specific determinants, reacted strongly with p46-52 and less intensely with the 22-kDa protein, but they did not react with the 92-kDa protein. A protein fraction extracted from the Triton X-114 detergent phase and further purified by ion-exchange chromatography was found to be highly enriched in p46-52 and was used as antigen in a prototype enzyme-linked immunosorbent assay (ELISA) to detect MS antibodies. Seventy serum samples were taken variously from specific-pathogen-free chickens experimentally infected with MS or MG and from commercial broiler breeder chickens vaccinated for MS and MG. All samples were tested by both rapid serum agglutination and a prototype ELISA described herein; the two tests were strongly correlated (r = 0.776). The results indicate that the ELISA antigen used--the immunodominant cell surface proteins in the p46-52 cluster--appears to be a good candidate for use in the development of improved rapid diagnostics of MS infections in birds.

Animals↗

PubMedDate not supplied

Mycoplasma gallisepticum isolated from house finches (Carpodacus mexicanus) with conjunctivitis.

An epornitic of conjunctivitis in free-flying house finches (Carpodacus mexicanus) occurred in several mid-Atlantic and eastern states of the USA in 1994. Clinical signs and gross lesions ranged from mild to severe unilateral or bilateral conjunctival swelling with serous to mucopurulent drainage and nasal exudate. Microscopic lesions consisted of chronic lymphoplasmacytic conjunctivitis, rhinitis, and sinusitis. Notably slow-growing mycoplasmas were isolated from conjunctival and/or infraorbital sinus swabs from clinically affected birds. Isolates were identified as Mycoplasma gallisepticum (MG) by direct immunofluorescence and DNA probe-based polymerase chain reactions. These findings suggest that MG is the likely etiology for this epornitic of conjunctivitis in house finches.

Animals↗

PubMedDate not supplied

Antibody responses of chickens to inoculation with Mycoplasma gallisepticum membrane proteins in immunostimulating complexes.

Membrane proteins of Mycoplasma gallisepticum (MG) strain R were extracted with the detergent Mega-10 and incorporated into immunostimulating complexes (ISCOMs). A membrane protein of approximately 64 kD (p64) molecular weight was a major component of MG ISCOMs. Six-week-old specific-pathogen-free leghorn chickens were inoculated by various routes (subcutaneous; combined intranasal and eyedrop; and combined subcutaneous, intranasal, and eyedrop) with 10 micrograms MG proteins in ISCOMs, or inoculated subcutaneously with 10 micrograms MG proteins in Freund's adjuvant. Subcutaneous inoculation of MG ISCOMs, or MG Freund's adjuvant resulted in higher sero-positive rates (detected by enzyme-linked immunosorbent assay) in serum and respiratory tract washings, compared to combined routes of MG ISCOM inoculation. In chickens inoculated subcutaneously with MG ISCOMs antibodies were first detected at 31 days postinoculation (PI) and the sero-positive rate peaked at 56 days PI. Sero-positive rates started to decline at day 64 PI. In the Freund's adjuvant group, MG antibodies were first detected at day 21 PI, and the sero-positive rate peaked at day 39 PI and did not decline. MG antibodies were detected by ELISA in upper respiratory tract and tracheal washes from chickens inoculated subcutaneously with MG ISCOMs and MG Freund's adjuvant. Immunoblots to MG strain R whole cell proteins showed that respiratory tract washings and sera from chickens inoculated subcutaneously with MG ISCOMs contained immunoglobulins to MG proteins, with a prominent reaction to p64.

Animals↗

PubMedDate not supplied

Transmissibility of live Mycoplasma gallisepticum vaccine strains ts-11 and 6/85 from vaccinated layer pullets to sentinel poultry.

In separate trials, layer pullets were vaccinated with Mycoplasma gallisepticum (MG) strain 6/85 or strain ts-11 commercially produced live vaccines. For a 15-wk postvaccination (PV) period, vaccinates were commingled with unvaccinated pullets and were in indirect contact with sentinel groups of pullets, broiler breeders, turkey breeders, or meat turkeys in adjoining pens. Infectivity and transmissibility of vaccine strains were determined by tracheal culture and serology at 1 wk followed by 3-wk intervals PV. Strain 6/85 was recovered from 0%-20% of vaccinates, but not from commingled pullets or sentinel birds. Strain ts-11 was recovered from 60%-90% of vaccinates and 0%-40% of commingled pullets but not from any of the sentinel birds. No birds in the 6/85 vaccine trial tested positive for MG antibodies by serology. MG enzyme-linked immunosorbent assays detected positive responses in ts-11 vaccinates (range = 10%-70%) at 42, 63, 84, and 105 days PV, and commingled pullets (10%) at 84 and 105 days PV. MG serum plate agglutination tests detected positive responses in 90% and 20% of ts-11 vaccinates at 42 and 105 days PV, respectively, and commingled pullets (10%) at day 42 PV. Clinical signs, morbidity, or mortality suggestive of pathogenic MG infection were not observed in any bird during either trial, and no gross lesions were observed at necropsy. Random amplified polymorphic DNA analysis was capable of distinguishing each of the vaccinal strains 6/85 and ts-11 from each other by their distinct DNA banding patterns.

Agglutination Tests↗