Molecular characterisation of chlamydial isolates from birds

Fifty-one chlamydial isolates from birds collected in Switzerland were classified by amplification and restriction analysis of the 16S-23S rRNA intergenic spacer region as Chlamydophila psittaci. The aim was to characterise a broad panel of chlamydial strains from birds and to apply and verify the methods of classification and differentiation described for chlamydial organisms. Two of the six known avian chlamydial serovars (A and B) were found by serotyping with monoclonal antibodies. One isolate was not typable. Digestion of ompA-PCR amplicons by AluI generated four distinct restriction patterns (genotypes A, B, F and G). Genotypes A and B correlated in most cases to serovars A and B, respectively. One serovar A isolate was verified as genotype B instead of A and one serovar B isolate belonged to genotype A. The non-serotypable isolate was of genotype F and one serovar A generated genotype G. OmpA sequences of one strain of each genotype were determined and compared to data bank entries. Amino acid sequences of genotype A and B strains corresponded well, showing more than 98.0% homology. The homologies of genotypes F and G sequences to genotype A strain were 82.0 and 83.0% respectively.     

Serotyping of US isolates of Chlamydophila psittaci from domestic and wild birds.

The identities of chlamydial strains, which can infect a given host, are important to know for disease prognosis, disease control, and epidemiology. The microimmunofluorescence test (MIFT) was used with a panel of 14 serovar-specific monoclonal antibodies (MAbs) to serotype 150 chlamydial isolates from domestic and wild birds. The isolates were obtained from birds submitted to diagnostic laboratories or during investigation of outbreaks. The 150 US isolates included 96 from the order Psittaciformes, 14 isolates from the order Columbiformes, 2 from the order Passeriformes, 16 from the order Galliformes, 12 from the order Struthioniformes, and 3 from the order Falconiformes. A total of 93, or 97%, of the Psittaciformes isolates were of serovar A; 11, or 79%, of the Columbiformes isolates were of serovar B; 64% of the Galliformes isolates were of serovar D, and all the Struthioniformes isolates were of serovar E. The 3 Falconiformes isolates did not react with any of the MAbs to the avian and mammalian isolates and are presumed to represent a new strain. The results show that specific chlamydial strains are usually associated with certain types of birds and that some serovars may be unusually virulent for certain species of birds. The MIFT using serovar-specific MAbs provides a rapid method to serotype new isolates, making it a useful system for epidemiological studies.     

Antigenic characterization of street rabies virus isolates from Nigeria using monoclonal antibodies

Twenty isolates of street rabies virus were recovered in mouse neutroblastoma cells from 84 rabies suspect brain specimens from Nigeria dogs and a cat. They were characterized with the Tübingen monoclonal antibody panels directed against nucleocapsid and glycoprotein antigens. Antigenic variations were detected both with antinucleocapsid (anti-NC) and antiglycoprotein (anti-GP) monoclonal antibodies (mabs). One isolate reacted positively with anti-NC mab P41 which hitherto has been known to react positively with polar rabies. Another isolate did not react with anti-NC mab 187.5; a reaction normally seen with ERA/SAD strains of rabies virus. With anti-GP mabs it was possible to group the isolates by their area of origin. Isolates from Plateau State were not neutralized by anti-GP mabs ERA 543 and P44.7.2. The isolates studied had glycoprotein antigenic patterns different from the pattern for low egg passage (LEP) Flury strain vaccine virus used in Nigeria for immunization of dogs.     

Antigenic characterization of bovine viral diarrhoea virus isolates from Spain with a panel of monoclonal antibodies

A group of 47 bovine viral diarrhoea virus (BVDV) strains isolated from a variety of bovine tissues from eight different geographical areas of Spain and two BVDV strains isolated from a cell line were characterized antigenically with a panel of 23 monoclonal antibodies (mAbs). The mAbs were directed at one of three viral proteins: E2, Erns and NS2-3. A peroxidase-linked assay was used to test the mAbs for reactivity against infected cell monolayers. The data were analysed by two computational methods: the Antigenic Distance Program (MAP) and the Phylogeny Inference Package (PHYLIP), and compared with those obtained previously using the same mAbs with other pestiviruses, including reference strains and UK field isolates. All the Spanish field strains studied appeared to be broadly similar to reference strains of BVDV and were included in the subgroup of classical BVDV, meanwhile the two strains isolated from a cell line were included in the subgroup of atypical pestiviruses.     

Serotyping of Mannheimia haemolytica isolates from bovine pneumonia: 1987-2006

Over a period of 20 years, a total of 207 Mannheimia haemolytica samples were isolated from calves affected with pneumonic pasteurellosis and serotyped by the indirect haemagglutination test. Serotypes A1 (102 isolates), A2 (47 isolates) and A6 (42 isolates) were most common; in addition, 16 isolates were serotypes A7, A13, A14 or untypable. The relative prevalence of serotype A6 has increased recently in Japan, as has been reported from other countries. The results of this study provide useful information towards the design of efficient vaccines for the prevention of M. haemolytica infection in Japan.     

Complement resistance-related traits among Escherichia coli isolates from apparently healthy birds and birds with colibacillosis

In this study, 294 Escherichia coli isolates from birds with colibacillosis were collected from disease outbreaks throughout the United States and were compared with 75 fecal E. coli isolates of apparently healthy chickens by their possession of several purported virulence genes, resistance to rough-lipopolysaccharide-specific bacteriophages (rLPSr), and elaboration of capsule. Traits were selected for study on the basis of their association with complement resistance. The genes targeted in this study included those encoding colicin V (cvaC) and the outer membrane proteins TraT (traT), OmpA (ompA), and Iss (iss). No significant differences were found between the two groups of isolates in the occurrence of cvaC-, traT-, or ompA-homologous sequences or in rLPSr. Only a few isolates were encapsulated, and the isolates of healthy birds were significantly more likely to be encapsulated than were the isolates of sick birds. However, iss, whether detected through hybridization or amplification, was found in more of the disease-associated isolates than in those of healthy birds. This difference was highly significant. Further, iss sequences were widely distributed among isolates of different serotypes from various avian host species and sites within these hosts. Such results suggest that possession of the iss sequence by an avian E. coli isolate may be a good indicator of that isolate's potential to cause disease. This association warrants further study because iss and the protein it encodes may be useful targets of future colibacillosis control efforts.     

Serotyping of Haemophilus paragallinarum isolates from Mexico by the Kume hemagglutinin scheme

A total of 42 isolates of Haemophilus paragallinarum from Mexico were serotyped by the Kume hemagglutinin scheme. Serovars A-1, A-2, B-1, and C-2 were recognized among 11 (26.2%), 7 (16.6%), 4 (9.5%), and 14 (33.3%) isolates, respectively. A further six isolates (14.3%) showed hemagglutinating activity but could not be classified into any serovar. Commercial vaccines containing Kume serovars A-1, A-2, B-1, and C-2 may provide better protection than those bi- or trivalent infectious coryza vaccines currently used in Mexico.     

鸡传染性支气管炎病毒广西分离株血清型的分析

应用病毒感染的鸡胚材料免疫新西兰兔的方法制备抗鸡传染性支气管炎病毒(IBV)单因子血清,然后在鸡胚气管环培养(Tracheal organ cultures,TOC)上对广西分离的7个IBV代表性毒株和3个常用疫苗株进行交叉病毒中和试验。结果显示,10个毒株被分为6个血清型。根据试验所得的R值,应用聚类分析法分析了各血清型毒株之间的亲缘关系,显示目前在广西流行的IBV野毒株之间以及其与疫苗株间的抗原性存在很大程度的差异,分属不同的血清型。同时还对IBV基因分型和血清分型之间的关系进行了探讨。     

Two types of Newcastle disease viruses isolated from feral birds in Western Australia detected by monoclonal antibodies

Thirteen viruses isolated from feral birds and one isolated from a domestic duck, obtained in 1979-1980 during a survey of birds in Western Australia, were shown to be Newcastle disease viruses of low virulence for chickens. The binding of mouse monoclonal antibodies, raised against NDV-Ulster 2C to MDBK cells infected with the isolates was assessed using an indirect immunoperoxidase test. Five viruses caused binding of all 9 monoclonal antibodies tested, whereas the other 9 isolates induced binding of only 4 monoclonal antibodies.     

Distinguishing between ovine abortion and ovine arthritis Chlamydia psittaci isolates with specific monoclonal antibodies

Monoclonal antibodies to Chlamydia psittaci were prepared by both in vivo and in vitro immunization methods, using an abortion strain of C psittaci as the immunizing antigen. Seven of the 8 monoclonal antibodies produced were genus-specific by the enzyme-linked immunosorbent assay and immunofluorescence test. The genus-specific antibodies were reactive with a protease-resistant, periodate-sensitive antigen of less than 14 kilodaltons. The remaining monoclonal antibody, 10D7, was specific for ovine abortion strains of C psittaci and nonreactive with 2 strains isolated from the joints of lambs with polyarthritis. The type-specific antigen was protease sensitive, but could not be detected in the immunoblot assay.     

Antigenic characterization of IBDV field isolates by their reactivity with a panel of monoclonal antibodies

Recently Infectious Bursal Disease Virus isolates have been described in USA displaying an antigenic drift. Many of the new isolates were very virulent for chickens. In several European countries severe outbreaks of Gumboro disease have also been reported from vaccinated and non-vaccinated flocks. Since vaccinated SPF birds were shown to be protected against challenge infection with the new isolates under laboratory conditions, a more detailed investigation of the European isolates is wanted. The similarity between the European and US field situation got us to use a panel of monoclonal antibodies (MCAs) previously applied to characterize US strains for testing European isolates. An antigen capture ELISA has been carried out directly on bursa homogenates of chickens form the field. One European (F52/70) and two US (Var. E and GLS-5) strains have been included as reference viruses. From the results presented here it can be concluded that the European isolates (Netherlands, France, UK, Germany, Jugoslavia and Spain) did not undergo the same antigenic drift as the US strains. A more extensive analysis of the isolates will be done to elucidate their role for disease outbreaks.     

Antigenic comparisons of hog cholera virus isolates from Europe, America and Asia using monoclonal antibodies

Nineteen monoclonal antibodies (MAbs) with specificity for hog cholera virus (HCV) were prepared. They were used in an immune binding (peroxidase linked) assay to determine the reaction patterns of HCV isolates from Europe, Brazil, USA, Japan and Malaysia, as well as laboratory reference strains of the virus. A further panel of 17 MAbs raised against bovine virus diarrhoea virus (BVDV) was included in the study, together with 5 MAbs raised against a non-HCV pestivirus of porcine origin. All the MAbs were also tested against representative strains of BVDV and border disease virus. Six MAbs were HCV-specific, reacting with all isolates of HCV and none of the ruminant viruses. Among the other HCV MAbs geographical variation in reaction patterns was observed. There was evidence of antigenic distinction between recent European isolates, and archive material originally isolated more than 10 years ago.     

Serological classification of Japanese isolates of Haemophilus paragallinarum using two serovar-specific monoclonal antibodies

A serological classification of 106 Japanese isolates of Haemophilus paragalinarum recovered from 1960 to 1984 was performed by dot-blotting and hemagglutination-inhibition (HI) tests using two serovar-specific monoclonal antibodies (MAbs), E5C12D10 and F2E6. By the dot-blotting test, 49 of the isolates were serovar A and 55 isolates were serovar C, and the two remaining isolates did not react with either MAb. These two nontypable strains had no hemagglutinating activity against chicken erythrocytes and were nonpathogenic to chickens. Although 49 serovar A isolates were serotyped by the HI test, only 23 of the 55 serovar C isolates could be serotyped. The remaining 32 isolates could not be serotyped because no or low hemagglutinating activity could be detected. Our results indicate that H. paragallinarum serovars A and C have both been present in Japan since 1960, with serovar A isolates being dominant before 1970 and serovar C isolates more prevalent than serovar A since 1970.     

Identification of hog cholera viral isolates by use of monoclonal antibodies to pestiviruses

A collection of 90 field isolates of hog cholera virus (HCV) was used to test the specificity of four hybridoma cell lines secreting monoclonal antibodies against pestiviruses. Reaction of virus isolates and monoclonal antibodies was controlled by an indirect immunofluorescence assay (IFA). Two monoclonal antibodies which had been generated against HC virus strain "Alfort 187" were reactive only with HCV field isolates and an HCV reference strain but not with bovine viral diarrhoea virus (BVDV) reference strains. Two other monoclonal antibodies (generated against BVDV, strain NADL) reacted only with BVDV reference strains but not with HCV field isolates, although with 3 of these strains focal reactions involving only a few cells were detected. The ability to discriminate between both viruses is a diagnostic need which may be fulfilled by these monoclonal antibodies.     

Serotyping of Mannheimia (Pasteurella) haemolytica isolates from the upper Midwest United States.

Mannheimia (Pasteurella) haemolytica biotype A serotype1 (A1) is the primary bacterial agent responsible for the clinical signs and pathophysiologic events in bovine pneumonic pasteurellosis. The goal of this study was to determine the prevalence of other serotypes of M. haemolytica biotype A organisms obtained from the upper Midwest diagnostic laboratories. A total of 147 M. haemolytica isolates were collected from Minnesota, South Dakota, and Michigan. Isolates were tested against M. haemolytica antisera obtained from the National Animal Disease Center, Ames, Iowa. Results indicated that M. haemolytica serotype 1 represented approximately 60%, serotype 6 represented 26%, and serotype 2 represented 7% of the total examined isolates. In addition, 7% of the isolates were serotype 9, 11, or untypable. This finding suggests that M. haemolytica serotypes other than serotype 1 can be isolated from the lung lesions of diseased cattle and seem to be capable of causing the pathologic changes observed in the lung with pneumonic pasteurellosis.