The development and application of aMycoplasma gallisepticumsequence database

Abstract

Molecular analysis was conducted on 36 Mycoplasma gallisepticum DNA extracts from tracheal swab samples of commercial poultry in seven South African provinces between 2009 and 2012. Twelve unique M. gallisepticum genotypes were identified by polymerase chain reaction and sequence analysis of the 16S-23S rRNA intergenic spacer region (IGSR), M. gallisepticum cytadhesin 2 (mgc2), MGA_0319 and gapA genetic regions. The DNA sequences of these genotypes were distinct from those of M. gallisepticum isolates in a database composed of sequences from other countries, vaccine and reference strains. The most prevalent genotype (SA-WT#7) was detected in samples from commercial broilers, broiler breeders and layers in five provinces. South African M. gallisepticum sequences were more similar to those of the live vaccines commercially available in South Africa, but were distinct from that of F strain vaccine, which is not registered for use in South Africa. The IGSR, mgc2 or MGA_0319 sequences of three South African genotypes were identical to those of the ts-11 vaccine strain, necessitating a combination of mgc2 and IGSR targeted sequencing to differentiate South African wild-type genotypes from ts-11 vaccine. To identify and differentiate all 12 wild-types, mgc2, IGSR and MGA_0319 sequencing was required. Sequencing of gapA was least effective at strain differentiation. This research serves as a model for the development of an M. gallisepticum sequence database, and illustrates its application to characterize M. gallisepticum genotypes, select diagnostic tests and better understand the epidemiology of M. gallisepticum.

Assessment of Mycoplasma gallisepticum vaccine efficacy in a co-infection challenge model with QX-like infectious bronchitis virus

“…MG field strain B2159/13 was isolated in October 2013 from a flock showing typical signs of mycoplasmosis. The strain was confirmed by Illumina ultradeep sequencing to be a pure culture, and is typical one of the unique South African "wild types" described by Armour et al (2013) (Bwala, 2017). B2159/13 was cultured to a titre of 3.125 x 10 9 CFU/ml and stored at -80 o C until use.…”

“…The presence of genetically unique MG strains in South Africa (Armour et al, 2013;Bwala, 2017), and field reports of sub-optimal vaccine protection prompted us to investigate the protective efficacy of ts-11 and 6/85 MG vaccines against a local virulent MG field strain, B2159/13. To enhance clinical symptoms and mimic a typical field situation, we co-infected birds with a virulent strain of QX-like IBV.…”

“…MG infection usually produces mild disease, but mycoplasmas' ability to synergize with other poultry respiratory pathogens such as infectious bronchitis virus (IBV), Newcastle disease virus (NDV) and Escherichia coli (E. coli) to exacerbate infections under intensive poultry management conditions makes avian mycoplasmosis a disease of economic significance (Rodriguez & Kleven, 1980;Nakamura et al, 1994). Generally, poultry farmers have depended on vaccination in combination with antimicrobials and biosecurity to control mycoplasmosis (Armour et al, 2013). Among the live vaccines available for MG control, strains ts-11 (Whithear 1996) and 6/85 (Evans & Hafez, 1992) have long been applied in South Africa, whereas the F strain vaccine (Adler et al, 1960;Luginbuhl et al, 1967) was not registered for use until the first quarter of 2015.…”

“…MG vaccine strains ts-11 and 6/85 are both attenuated and avirulent and have been reported to be safe and efficacious (Evans & Hafez, 1992;Whithear, 1996). The use of live-attenuated MG vaccines in some parts of the world has, however, been reported to have variable efficacy in challenge experiments (Abd-El-Motelib & Kleven, 1993), and in South Africa, field reports have suggested suboptimal protection of flocks against infection with local MG isolates, which were reported to be genetically unique (Armour et al, 2013).…”

Evaluation of the egg transmission and pathogenicity of Mycoplasma gallisepticum isolates genotyped as ts-11

“…PCR amplification, sequencing and sequence analysis of the 16S-23S rRNA IGSR genetic target were performed on selected M. gallisepticum isolates using previously described primer sequences, thermocycler programmes, PCR amplification methods and sequence analysis software (Raviv et al, 2007;Armour et al, 2013).…”