Phenotypic and Molecular Characterization of V-Factor (NAD)-Independent Haemophilus paragallinarum

Abstract

In South Africa from early 1989 onward, strains of Haemophilus paragallinarum not requiring nicotinamide adenine dinucleotide (NAD) have been isolated from commercial chickens suffering from infectious coryza. Fifteen of these field isolates were characterized by biochemical typing, serotyping, restriction endonuclease analysis (REA), and ribotyping. The chosen isolates represented diversity in geographic location, time of disease outbreak, and type of flock. All were typical of the species in biochemical properties, except that they were NAD-independent, and all were Page serovar A. REA was performed with three enzymes: HindIII, HpaII, and SspI. All isolates gave identical REA profiles with all three enzymes. Ribotyping was performed using a probe that consisted of the plasmid pUC19 into which the 16S rRNA operon of H. paragallinarum had been inserted. All 15 isolates gave the same ribotyping profile using each of the three enzymes. As a group, the NAD-independent strains gave REA profiles and ribotypes that were very different from a range of classic South African strains isolated before 1989. Our results strongly suggest that the NAD-independent isolates are clonal in nature.

Reclassification of Pasteurella gallinarum, [Haemophilus] paragallinarum, Pasteurella avium and Pasteurella volantium as Avibacterium gallinarum gen. nov., comb. nov., Avibacterium paragallinarum comb. nov., Avibacterium avium comb. nov. and Avibacterium volantium comb. nov.

“…The available DNA-DNA hybridization data of Mouahid et al (1992), who reported 89 % DNA binding between strain ATCC 29545 T and the biovar 2 strain SA 4461, indicate that the biovar 1 and 2 forms of Avibacterium ([Haemophilus]) paragallinarum can be accommodated in one species. The DNA fingerprinting work of Miflin et al (1995) has provided evidence that the South African NAD-independent isolates of Avibacterium ([Haemophilus]) paragallinarum are clonal, indicating that the DNA-DNA hybridization data for strain SA 4461 are applicable for all of the biovar 2 strains in this study. As well, all South African biovar 2 isolates of Avibacterium ([Haemophilus]) paragallinarum tested to date have yielded a positive reaction for the [Haemophilus] paragallinarum-specific PCR (Chen et al, 1996;Miflin et al, 1999).…”

“…As well, all South African biovar 2 isolates of Avibacterium ([Haemophilus]) paragallinarum tested to date have yielded a positive reaction for the [Haemophilus] paragallinarum-specific PCR (Chen et al, 1996;Miflin et al, 1999). Furthermore, all of the South African biovar 2 isolates that have been serotyped (Miflin et al, 1995) have been allocated to the existing Page serotyping scheme, which was originally created using typical biovar 1 strains (Page, 1962). Hence, there are a number of separate bodies of evidence -the DNA-DNA hybridization data of Mouahid et al (1992), the clonality of the South African biovar 2 isolates as shown by Miflin et al (1995), the positive reaction in the [Haemophilus] paragallinarum PCR (Chen et al, 1996, Miflin et al, 1999, the ability to assign the South African isolates of biovar 2 to the existing [Haemophilus] paragallinarum serotyping scheme (Miflin et al, 1995) and the 16S rRNA gene sequencing results of the current study -that all support the inclusion of the biovar 2 Table 1.…”

“…Furthermore, all of the South African biovar 2 isolates that have been serotyped (Miflin et al, 1995) have been allocated to the existing Page serotyping scheme, which was originally created using typical biovar 1 strains (Page, 1962). Hence, there are a number of separate bodies of evidence -the DNA-DNA hybridization data of Mouahid et al (1992), the clonality of the South African biovar 2 isolates as shown by Miflin et al (1995), the positive reaction in the [Haemophilus] paragallinarum PCR (Chen et al, 1996, Miflin et al, 1999, the ability to assign the South African isolates of biovar 2 to the existing [Haemophilus] paragallinarum serotyping scheme (Miflin et al, 1995) and the 16S rRNA gene sequencing results of the current study -that all support the inclusion of the biovar 2 Table 1. Key characters for differentiation of genera within the family Pasteurellaceae Genera: 1, Avibacterium gen. nov.; 2, Pasteurella; 3, Actinobacillus; 4, Haemophilus; 5, Lonepinella; 6, Mannheimia; 7, Phocoenobacter; 8, Gallibacterium; 9, Histophilus; 10, Volucribacter.…”

Identification of a Plasmid-Encoded Gene from Haemophilus ducreyi Which Confers NAD Independence

“…These include strains of Actinobacillus pleuropneumoniae, which causes pleuropneumonia in swine (22); Haemophilus paragallinarum, which causes fowl choryza (1,15); Haemophilus parainfluenzae, which can cause pneumonia and meningitis in humans (7); and Haemophilus ducreyi, which causes the sexually transmitted disease chancroid in humans (26,27). In H. parainfluenzae, H. paragallinarum, and H. ducreyi, V-factor independence has been shown to be encoded on a plasmid (1,27,28).…”

Molecular characterization of isolates ofHaemophilus paragallinarumfrom China by ribotyping

“…A recent study has successfully utilized ribotyping to demonstrate that growth factor-independent isolates of H. paragallinarum recently reported in South Africa have little genetic diversity, unlike the classic H. paragallinarum strains also present in that country which show considerable diversity (Miflin et al, 1995). When developing ribotyping methods for a particular species, it may be necessary to investigate the usefulness of a number of different restriction endonucleases.…”