VNTR analysis reveals unexpected genetic diversity within Mycoplasma agalactiae, the main causative agent of contagious agalactia

Laura Mcauliffe,Guido Loria,Joanna R Lawes,Colin P Churchward,Roger D Ayling,Robin Aj Nicholas

doi:10.1186/1471-2180-8-193

Abstract

BackgroundMycoplasma agalactiae is the main cause of contagious agalactia, a serious disease of sheep and goats, which has major clinical and economic impacts. Previous studies of M. agalactiae have shown it to be unusually homogeneous and there are currently no available epidemiological techniques which enable a high degree of strain differentiation.ResultsWe have developed variable number tandem repeat (VNTR) analysis using the sequenced genome of the M. agalactiae type strain PG2. The PG2 genome was found to be replete with tandem repeat sequences and 4 were chosen for further analysis. VNTR 5 was located within the hypothetical protein MAG6170 a predicted lipoprotein. VNTR 14 was intergenic between the hypothetical protein MAG3350 and the hypothetical protein MAG3340. VNTR 17 was intergenic between the hypothetical protein MAG4060 and the hypothetical protein MAG4070 and VNTR 19 spanned the 5' end of the pseudogene for a lipoprotein MAG4310 and the 3' end of the hypothetical lipoprotein MAG4320.We have investigated the genetic diversity of 88 M. agalactiae isolates of wide geographic origin using VNTR analysis and compared it with pulsed field gel electrophoresis (PFGE) and random amplified polymorphic DNA (RAPD) analysis. Simpson's index of diversity was calculated to be 0.324 for PFGE and 0.574 for VNTR analysis. VNTR analysis revealed unexpected diversity within M. agalactiae with 9 different VNTR types discovered. Some correlation was found between geographical origin and the VNTR type of the isolates.ConclusionVNTR analysis represents a useful, rapid first-line test for use in molecular epidemiological analysis of M. agalactiae for outbreak tracing and control.

Highlights

Mycoplasma agalactiae is the main cause of contagious agalactia, a serious disease of sheep and goats, which has major clinical and economic impacts
variable number tandem repeat (VNTR) profiles were recorded as character data using allelic profiles and dendrograms were constructed with PHYLIP hosted at http://pubmlst.org/analysis/ using neighbor-joining
VNTR analysis was far more effective and enabled us to detect variation between M. agalactiae strains with the 88 isolates being divided into 9 VNTR types showing some correlation between geographical origin and the VNTR profiles of the isolates

Summary

Introduction

Mycoplasma agalactiae is the main cause of contagious agalactia, a serious disease of sheep and goats, which has major clinical and economic impacts. In most cases, infected hosts spontaneously recover from acute clinical signs within a few weeks but develop a chronic infection accompanied by shedding of M. agalactiae in milk and/or other body secretions for years without presenting any clinical signs [1]; these (asymptomatic) carriers can transmit the bacteria to other susceptible animals and cause acute disease [2]. This has seriously hindered the eradication of CA despite research into vaccines [3,4], identification of effective antibiotics [5,6] and improved diagnostic testing [7]. One of the reasons for this antigenic divergence is the presence of gene families of variable proteins such as Avg and VmpA; these are similar to the Vsp gene family in Mycoplasma bovis (M. bovis) [12,13,14]

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