Preventive Veterinary Medicine 157 (2018) 152–161

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Preventive Veterinary Medicine

journal homepage: www.elsevier.com/locate/prevetmed

Mycoplasma bovis and bovine respiratory disease: A risk factor study in T

Australian feeder cattle
⁎
M.L. Schibrowskia, J.S. Gibsonb, K.E. Haya, T.J. Mahonya, , T.S. Barnesa,b
a
The University of Queensland, Queensland Alliance for Agriculture and Food Innovation, St Lucia, 4072, Queensland, Australia
b
The University of Queensland, School of Veterinary Science, Gatton, 4343, Queensland, Australia

A R T I C LE I N FO A B S T R A C T

Keywords: Mycoplasma bovis can be a bacterial inhabitant of the upper respiratory tract of healthy bovines. In body regions
Mycoplasma bovis other that the upper respiratory tract however, M. bovis is associated with a number of clinical syndromes such as
Sero-prevalence bovine respiratory disease (BRD). This study used two enzyme-linked immunosorbent assays to assess the sero-
Risk factors status of M. bovis-specific antibodies in Australian feeder cattle at the time of feedlot induction and at ap-
Bovine respiratory disease
proximately 42 days on feed (follow-up). The apparent sero-prevalence of M. bovis-specific antibody at induction
Feedlot
was estimated to be 3.5% (95% confidence interval [CI] 2.0–5.0%, 47/1354) and 25.3% (95% CI 21.9–28.8%,
343/1354) at follow-up. Exposure to M. bovis between induction and follow-up as demonstrated by an increase
in serum antibodies was estimated to be 19.4% (95% CI 16.2–22.6%, 261/1349).
Risk factors associated with sero-positivity at feedlot induction included the region where animals were 28
days prior to induction and saleyard exposure at least 27 days prior to induction. Risk factors associated with a
sero-increase between induction and follow-up included breed, source region and access to water shared with an
adjoining pen of animals. Of these, shared pen water was considered the most important (odds ratio [OR] 3.3,
95% CI 1.5–7.4, p = 0.003). Animals exposed to M. bovis between induction and follow-up were at a sub-
stantially increased risk of BRD (OR 2.2, 95% CI 1.4–3.4, p = 0.001).
This is the first Australian study that has identified risk factors for M. bovis sero-positivity and sero-increase
and shown an association between sero-increase and the risk of BRD in the feeder cattle population. These
findings suggest that M. bovis is a significant pathogen in the Australian feeder cattle population. In addition,
identification of defined risk factors associated with an increased risk of exposure to M. bovis can assist in the
development of targeted control measures to reduce the economic impact of M. bovis associated disease and BRD
in feeder cattle.

1. Introduction
Mycoplasma bovis can be a bacterial inhabitant of the upper re-
spiratory tract of healthy bovines. However, under certain circum-
stances and conditions that remain incompletely understood, M. bovis
can be associated with a number of clinical syndromes. One other
Australian study to date, has assessed the sero-prevalence of M. bovis-
specific antibody among Australian beef cattle populations (Wawegama
et al., 2016). This previous study used an enzyme-linked im-
munosorbent assay (ELISA) based on the recently identified antigen
MilA. The test is currently not commercially available (Wawegama
et al., 2014). In addition, this ELISA has not been used in cattle popu-
lations in countries other than Australia, so comparisons with interna-
tional M. bovis research cannot be made.
At the time of the current study, two ELISAs for the detection of M.
bovis-specific antibody were available commercially in Australia. Both
ELISAs were produced by the same manufacturer, Bio-X Diagnostics®
(Belgium), but were marketed for different purposes. The BIO K302
requires a single serum sample and is marketed to determine the im-
mune status of an animal to M. bovis at the time the sample was col-
lected. The BIO K260 however, is marketed to assess recent exposure of
the animal to M. bovis. This ELISA requires two serum samples collected
from the same animal, two to three weeks apart.
Bovine respiratory disease (BRD) is the major cause of clinical dis-
ease and death in feeder cattle worldwide (Edwards, 2010). In Aus-
tralian feedlots it has been estimated to cause between 60 and 70% of
all clinical cases and deaths (Sackett et al., 2006). The disease is mul-
tifactorial and in recent years there has been increased interest in the

⁎
Corresponding author.
E-mail address: t.mahony@uq.edu.au (T.J. Mahony).

https://doi.org/10.1016/j.prevetmed.2018.06.005
Received 18 October 2017; Received in revised form 14 May 2018; Accepted 19 June 2018
0167-5877/ © 2018 Elsevier B.V. All rights reserved.
M.L. Schibrowski et al.
role M. bovis may play in its development. With improvements in the
isolation and detection of M. bovis, it is becoming increasingly clear that
M. bovis infections are major primary or contributory causes of the
development of BRD in intensive cattle production systems (Nicholas
et al., 2008; Caswell et al., 2010). Respiratory disease associated with
M. bovis can occur in cattle of any age (Maunsell et al., 2011) however,
the nature of the association between M. bovis and respiratory disease in
cattle more than six months of age is not well understood. Some evi-
dence to support M. bovis as a predisposing factor in the infectious
process for the development of BRD in cattle more than six months of
age has been provided by Divers (2008), Horwood et al. (2014) and
Poumarat et al. (2001). However, in Australia the association between
M. bovis infections and the development of BRD in the early feeding
period of feeder cattle has not been assessed.
The aims of the current study were to utilise commercially available
diagnostic tests used in cattle populations both in Australia and inter-
nationally to; (i) estimate the apparent sero-prevalence of M. bovis-
specific antibody at the time of feedlot induction (Day 0) and at ap-
proximately 42 days on feed (follow-up); (ii) assess and quantify pu-
tative risk factors associated with sero-positivity for M. bovis-specific
antibody at Day 0; (iii) determine the level of sero-increase to M. bovis
that occurs between Day 0 and follow-up; (iv) assess and quantify pu-
tative risk factors associated with a sero-increase between Day 0 and
follow-up, and (v) assess sero-positivity and sero-increase to M. bovis-
specific antibody as risk factors for BRD.
2. Materials and methods
2.1. Study population and structure of the data
The study population consisted of a subset of animals randomly
selected from those enrolled in the National Bovine Respiratory Disease
Initiative (NBRDI). The NBRDI study design and population have been
described elsewhere by Hay et al. (2014). The study was conducted
under animal ethics approvals SVS/383/07/MLA, SVS/495/08/MLA,
SVS/125/10/MLA and AEC09/027. Serum samples were collected from
all animals enrolled in the NBRDI at the time of feedlot induction (Day
0) and at approximately 42 days on feed (follow-up). In the NBRDI for
animals that were not pre-assembled at the feedlot, Day 0 was defined
as the day of induction (when animals were individually identified and
enrolled into the study). For animals that were pre-assembled, Day 0
was the day the animals were moved into the feedlot pen. Pre-assembly
is a management practice whereby animals from different farms are
assembled on pasture close to the feedlot for various periods of time
prior to being placed in a feedlot pen.
In the current study, animals were eligible for selection if records in
the NBRDI database indicated an adequate serum sample was available
for both the Day 0 and follow-up time points. Of the 35,131 animals in
the NBRDI dataset, 29,512 animals were eligible (Fig. 1). Sample size
was chosen a priori based on prevalence estimation and determined
using an online sample size calculator (Sergeant, 2013). Input values
were an estimated apparent prevalence of 50%, an infinite study po-
pulation, 95% confidence level and a desired precision of 0.03. The
sample size estimate was inflated using a design effect of 1.26 based on
an estimated average of two animals from each purchase group in the
dataset and an intraclass correlation co-efficient (ICC) of 0.26. The ICC
was estimated by fitting a null mixed effects logistic regression model
with a random effect for purchase group and an outcome of sero-status
at the time of induction using the results from a subset of NBRDI ani-
mals tested using a multiplex ELISA, the Bio-X Diagnostics® BIO K284,
during the early stages of the NBRDI analyses. A retrospective assess-
ment of the sample size of 1354 indicated more than 80% power to
identify a risk factor with an odds ratio of two or greater, assuming a
significance level of 5%, prevalence of the risk factor in the population
of 20%, prevalence/incidence of the outcome of 10% and a design ef-
fect of 1.26. All eligible animals were assigned a random number and
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Preventive Veterinary Medicine 157 (2018) 152–161
samples from these animals were collated in that order until a sufficient
number of samples were obtained from the serum bank (Fig. 1).
The NBRDI data had a nested hierarchical structure with four levels:
feedlot, cohort, group and animal as described in detail by Hay et al.
(2014). A ‘cohort’ comprised all animals in a single feedlot pen fol-
lowing the induction process while a ‘group’ comprised all animals in
the same cohort that were together on the same property at a specified
time point prior to Day 0. The two higher levels of clustering (feedlot
and cohort) were clearly defined by the data. However, there was no
single definition of a group as typically animals did not remain in stable
groups from birth through to arrival at the feedlot. In this study, group-
13 (a group of animals in the same cohort that were together on the
same property 13 days prior to Day 0) was chosen as the lowest level of
clustering. Thus group-13 s were nested within cohorts which were
nested within feedlots. This was the main hierarchy used in the model
building process. The property where a group-13 cluster of animals was
located 13 days prior to Day 0 was referred to as the ‘PIC-13’ where PIC
stands for ‘Property Identification Code’. Thus group-13 s were also
separately nested within PIC-13 s. PIC-13 s were only considered when
exploring the partitioning of the variance in sero-status at Day 0.
2.2. Serological testing and sero-prevalence estimation
All sera were tested for the presence of M. bovis-specific antibody
using both commercially available Bio-X Diagnostics® BIO K302 and
BIO K260 M. bovis ELISAs. Testing and plate validation were carried out
as per the manufacturer’s instructions ensuring both the Day 0 and
follow-up sera from the same animal were tested on the same test plate.
Samples were retested if an ELISA plate failed the manufacturer vali-
dation criteria. Results were interpreted using the manufacturer’s in-
structions whereby samples with a BIO K302 sample coefficient ≥ 37%
were considered sero-positive. Sero-prevalence estimates were adjusted
for clustering by group-13 at Day 0 and by cohort at follow-up. There
were six result categories for the BIO K260 ELISA ranging from ‘0’ to ‘+
++++’ based on delta values of ≤ 37, > 37–≤ 60, > 60–≤
83, > 83–≤ 106, > 106–≤ 129 and > 129%. Animals with a Day 0
serum result of ‘0’, ‘+’, ‘++’ or ‘+++’ and a follow-up result of at
least two categories greater (i.e. ‘++’, ‘+++’, ‘++++’ or ‘++++
+’) were considered to have sero-increased. Animals with a Day 0 re-
sult in the ‘++++’ or ‘+++++’ category were classified as being
high initially and consequently did not have the potential to increase
two or more categories between Day 0 and follow-up. These animals
were excluded from the sero-increase analyses. Estimates for sero-in-
crease were adjusted for clustering by cohort.
2.3. Outcome and exposure variables for risk factor analyses
There were three outcomes of interest for the risk factor analyses: (i)
the presence or absence of serum antibody specific for M. bovis at Day 0
based on BIO K302 results, (ii) sero-increase between Day 0 and follow-
up based on the BIO K260 results and (iii) the occurrence of BRD within
the first 50 days following induction. The case definition for BRD was
based on examination of hospitalised animals by feedlot staff and has
been described in detail elsewhere (Hay et al., 2014).
The NBRDI aimed to identify risk factors associated with BRD and as
such, a series of variables were available in the NBRDI database for
each animal. Analysis of putative risk factors for M. bovis sero-positivity
at Day 0 or a sero-increase at follow-up, were restricted to those for
which data were available in the NBRDI database and where prior
hypotheses suggested an association with sero-positivity or sero-in-
crease. Mycoplasma bovis sero-positivity and sero-increase were then
separately considered as risk factors for BRD.
Exposure variables of interest are described in detail by Hay et al.
(2014, 2016a,b,c,d) and were categorised as putative animal-level,
group-level or cohort-level risk factors. Several putative animal-level
risk factors were considered. ‘Breed’ consisted of seven categories and
M.L. Schibrowski et al.
Fig. 1. Flow diagram depicting the selection of animals from those enrolled in the National Bovine Respiratory Disease Initiative (NBRDI) and sample collation
process.
‘Dentition’ was used as a crude estimate of age and comprised three
categories (‘0’, ‘2’, ‘4 or 6’ permanent incisors). Live weight at the time
of induction was defined as ‘Induction weight’ and described using four
categories. Two mixing variables were considered: (i) ‘Mixing prior to
day -27’ indicating an animal had been mixed with animals from a
different PIC at any time point > 27 days prior to Day 0 was considered
as a risk factor for M. bovis sero-positivity at Day 0 and (ii) ‘Mixing
summary’ was considered as a risk factor for sero-increase at follow-up.
This was a composite variable that categorised animals based on whe-
ther or not they had been mixed with animals from other properties at
any time point > 27 days prior to Day 0 and the number of groups
defined 28 days prior to Day 0 that formed the animal’s cohort (i.e.
yes, < 4 [the animal had been mixed with other animals at some
point > 27 days prior to Day 0 and had gone into a cohort with < 4
groups of animals that were together 28 days prior to Day 0]; yes ≥ 4;
no, < 4; and no, ≥ 4). Two binary saleyard variables were also con-
sidered. ‘Saleyard exposure prior to day -27’ indicated whether an an-
imal had been exposed to the saleyard at any time point > 27 days
prior to Day 0 and ‘Saleyard exposure between day -27 to 0’ indicated
exposure to the saleyard at some time point between 27 days prior to
Day 0 and Day 0.
Putative group-level risk factors were ‘Source region’, ‘Number of
animals in group-13’, ‘Feedlot move timing’ and ‘BVDV exposure’.
Source region was determined by the geographical co-ordinates of the
animals PIC location 28 days prior to Day 0. It comprised six broad
regions based on proximity, similarity in geography, ecology, weather
patterns and the density of animals as described in detail by Hay et al.
(2016d). ‘Feedlot move timing’ indicated if an animal was moved to the
feedlot at a time point > 27 days prior to Day 0 or between day -27 and
Day 0. ‘BVDV exposure’ was a composite variable identifying the pre-
sence or absence of active infection with bovine viral diarrhoea virus 1
(BVDV-1) in any animal in the cohort and the presence of a persistently
infected animal (PI) in the animal’s group-28 (the -28 day equivalent of
group-13). There were three categories: ‘active infection, no PI’; ‘active
infection, PI’ and ‘no active infection, no PI’ (Barnes et al., 2015).
Putative cohort-level risk factors were ‘Induction year’, ‘Number of
animals in cohort’ (< 200, ≥ 200), ‘Induction season’ (i.e., spring,
summer, autumn, winter) and pen characteristics. Pen characteristics
were derived for the pen where the cohort spent the majority of time
during their first 50 days on feed as described by Hay et al. (2016c).
These included the binary variable ‘Shared pen water’ which indicated
whether or not the pen’s water trough(s) were accessible to animals in
an adjoining pen, the number of immediately adjoining pens (‘Pens
joining’: 1, 2), the provision of shade in the pen (‘Pen shade’: yes (i.e.
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Preventive Veterinary Medicine 157 (2018) 152–161
any shade), no) and the stocking density (‘Pen density’), a four-category
variable describing the number of square meters per standard cattle
unit (equivalent to an animal weighing 600 kg).
2.4. Causal diagrams and risk factor analyses
As recommended by the updated STROBE guidelines for the re-
porting of observational studies, the putative causal structure among
variables was represented using causal diagrams (Sargeant et al., 2016).
In a similar manner to the other analyses within the NBRDI, these were
then used to inform the model-building process to estimate the total
effects for all putative risk factors of interest and direct effects for the
saleyard variables as they were identified a priori as being of interest
(Hay et al., 2014). These diagrams, one for each set of analyses, were
constructed based on a priori postulated causal pathways, represented
by unidirectional arrows, linking exposures of interest with each other
and with each outcome (Figs. 2–5). Thus, each arrow in each causal
diagram depicts a hypothesised causal pathway in which one variable
(the variable from which the arrow starts) might at least partly de-
termine another (the variable to which the arrow points). A pathway
going directly from a variable to the outcome depicts a direct effect of
that variable. Indirect pathways are those where a variable is linked to
another via one or more intervening variables; this is depicted as a
sequence of arrows, so the pathway can be traced passing through these
intervening variables by following the sequence of arrows in the correct
direction. There may be multiple indirect pathways from any one par-
ticular variable to any other particular variable. Effects mediated in this
way are known as indirect effects. The total effect of a variable on the
outcome is the sum of the direct and all the indirect effects for that
variable on the outcome. The total effect is usually of greatest interest
as this estimate describes the expected change in the risk of the out-
come if that exposure were changed. In some cases, the direct effect is
also of practical interest as it relates to the expected change in outcome
that is not mediated by the intervening variable(s).
Backwards elimination methods are commonly used in large ob-
servational studies in veterinary epidemiology where data are gathered
on a large number of putative risk factors with the aim of identifying a
parsimonious set of risk factors associated with the outcome of interest.
However, using this approach, the effect estimates for the variables
included may be total, direct or partial and these estimates may be
biased due to inadequate control of confounding (Westreich and
Greenland, 2013). For these reasons, Westreich and Greenland speci-
fically recommend the use of separate models based on pathways in a
plausible causal diagram to obtain total effect estimates for variables of
M.L. Schibrowski et al. Preventive Veterinary Medicine 157 (2018) 152–161

Fig. 2. Causal diagram based on a priori knowledge and biologically plausible pathways interlinking the measured exposure variables with each other, as appropriate,
and with the occurrence of Mycoplasma bovis sero-positivity at Day 0.

interest and to repeat this process to obtain direct effect estimates when
they are specifically of interest. A disadvantage of the causal diagram
approach is that it can result in uncontrolled confounding if the causal
diagram does not accurately capture causal pathways or important
confounders are missing from the diagram or the directionality of as-
sociations is not captured correctly. However, Shrier and Platt (2008)
strongly advocate the approach as not following it favours chance over
rational deliberation.
Several methods have been proposed to determine which covariates
to include in models to estimate total and direct effects (Greenland
et al., 1999; Shrier and Platt, 2008; Dohoo et al., 2010; Textor and
Liskiewicz, 2011). Of these, the use of the DAGitty® software described
by Textor et al. (2011) was preferred as this automated approach is less
subject to human error with more complex diagrams. The causal dia-
grams were reproduced using this software to identify minimal suffi- Fig. 4. Causal diagram interlinking Mycoplasma bovis sero-positivity at Day 0
cient adjustment sets (a set of variables for each risk factor of interest with the outcome of bovine respiratory disease within the first 50 days on feed
that appropriately controls confounding) to separately assess the total and possible confounding variables.
effects of each risk factor on the outcome of interest and the direct
effects for the saleyard variables (Textor et al., 2011).
regression models were fitted with group-13 fitted as a single random
Stata/SE 12.1® (Stata Statistical Software, Stata Corporation,
effect. To assess risk factors for sero-increase at follow-up and to assess
College Station, TX, USA) was used for all data management and ana-
sero-positivity and sero-increase to M. bovis as risk factors for BRD,
lyses. To assess risk factors for sero-positivity at Day 0, logistic
models were fitted with feedlot, cohort and group-13 as random effects.

Fig. 3. Causal diagram based on a priori knowledge and biologically plausible pathways interlinking the measured exposure variables with each other, as appropriate,
and with the occurrence of sero-increase of two or more BIO K260 categories between Day 0 and follow-up.

155
M.L. Schibrowski et al.
Fig. 5. Causal diagram interlinking the occurrence of sero-increase of two or
more BIO K260 categories between Day 0 and follow-up with the outcome of
bovine respiratory disease within the first 50 days on feed and possible con-
founding variables.
The overall fit of the models was not assessed as the standard tools for
assessing model fit are not readily available for multi-level logistic
models (Dohoo et al., 2010). Model assumptions of normal distribution
of random effects and homoscedasticity of residuals at the level of each
random effect were assessed graphically.
Partitioning of the variance in sero-status at Day 0 was assessed
using the latent variable threshold approach by fitting a null mixed
effects logistic regression model with PIC-13 and group-13 as random
effects with the BIO K302 Day 0 result as the outcome variable (Snijders
and Bosker, 2012). The ICC for group-13 was calculated as the pro-
portion of total variance that was at either the PIC-13 or the group
level. The design effect of clustering at the group-13 level was then
estimated using the mean number of animals in a group-13 that were
included in the analysis (Dohoo et al., 2010). An equivalent analysis to
assess the partitioning of the variance in sero-increase between Day 0
and follow-up at the feedlot, cohort, group and animal levels and design
effect at the cohort level was conducted with sero-increase based on the
BIO K260 results as the outcome variable.
3. Results
3.1. Sero-prevalence and sero-increase estimates
Of the 29,512 animals eligible for selection in this study, 91 animals
were excluded at the time of sample selection from the NBRDI serum
bank as either a Day 0 or follow-up sample was unavailable (Fig. 1).
The timing of the follow-up sample collection for the selected animals
(n = 1,354) ranged from Day 31 to Day 75, with 1,074 (79.3%) re-
corded as having their follow-up sample collected between Day 40 and
Day 49. Of the 14 feedlots, 170 cohorts and 1077 group-13 s in the full
NBRDI dataset, 14, 158 and 525 were represented in these analyses,
respectively.
The estimated sero-prevalences of M. bovis-specific antibody were
3.5% (95% confidence interval [CI] 2.0–5.0%, 47/1,354) and 25.3%
(95% CI 21.9–28.8%, 343/1354) at Day 0 and follow-up, respectively
using the BIO K302 ELISA. Based on the Day 0 results from the BIO
K260 ELISA, 1,349 animals had the potential to sero-increase at follow-
up and of these 19.4% (95% CI 16.2–22.6%, 261/1,349) did sero-in-
crease (Table 1).
3.2. Risk factors for sero-positivity at day 0
The proportion of the study population in each putative risk factor
category was similar to those reported by Hay et al. (2014, 2016d). The
distribution and estimated odds ratios (OR) for total effects (and direct
where applicable) of each putative risk factor on the presence of M.
bovis-specific antibody at Day 0 are provided in Table 2 (covariates for
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Preventive Veterinary Medicine 157 (2018) 152–161
each model are listed in the footnotes). Exposure to a saleyard > 27
days prior to Day 0 was associated with an increased risk of sero-po-
sitivity at Day 0 (OR 2.9, 95% CI 1.1–7.7, p = 0.03). The direct effect
estimate was similar, indicating that the effect of exposure to a saleyard
was not mediated through mixing prior to day -27, the only postulated
intervening variable. Source region was associated with variability in
risk of sero-positivity at Day 0. Most notably animals from the Darling
Downs and New England regions were at reduced risk compared to the
reference group, Central and Southern Tablelands of New South Wales
(OR 0.2, 95% CI 0.0–1.0, p = 0.05). Within induction years, animals
inducted in 2011 were at lower risk than those inducted in 2009 (OR
0.3, 95% CI 0.1–0.8, p = 0.04). There was also weak evidence of an
association with the number of animals in the group-13, with animals in
groups of at least 100 possibly being at higher risk of sero-positivity at
Day 0 compared to those in groups of less than 50 (OR 3.0, 95% CI
0.8–11.0, p = 0.11). Estimates for the association between each of
breed, induction weight, dentition and mixing > 27 days prior to Day
0, and the risk of sero-positivity to M. bovis at Day 0 were imprecise so
no conclusion could be reached (Table 2).
3.3. Risk factors for sero-increase between day 0 and follow-up
The proportion of the study population in each risk factor category
was similar to those reported in Hay et al. (2014, 2016a,b,c,d). The
distribution and estimated odds ratios for total (and direct where in-
dicated) effects of each putative risk factor and sero-change at follow-
up are shown in Table 3 (covariates for each model are listed in the
footnotes). Breed category was associated with variability in risk of
sero-increase between Day 0 and follow-up, with British cross cattle and
European cattle and their crosses being at reduced risk compared to
Angus cattle, (British cross: OR 0.4, 95% CI 0.2 – 0.8, p = 0.01, Eur-
opean/European cross: OR 0.3, 95% CI 0.1–1.0, p = 0.05). There was
also some evidence of reduced risk among tropical cattle and their
crosses. Source region was associated with variability in risk of sero-
increase between Day 0 and follow-up. Most notably animals from the
Darling Downs and New England, western New South Wales, Queens-
land and Northern Territory, and South and Western Australia regions
were at reduced risk compared to the reference group, Central and
Southern Tablelands of New South Wales and (Table 3). There was
some evidence that the risk of sero-increase between Day 0 and follow-
up differed across the years in which cattle were inducted into the
study, with animals inducted in 2011 at higher risk (OR 2.6, 95% CI
1.1–6.0, p = 0.03) compared to animals inducted in 2009 (Table 3).
There was strong evidence that the risk of sero-increase between Day 0
and follow-up was increased when animals were in pens where the
water troughs could be accessed by animals in adjoining pens (OR 3.3,
95% CI 1.5–7.4, p = 0.003). Estimates for the association between each
of induction weight, dentition, mixing summary, exposure to a saleyard
during either time period, feedlot move timing, the number of animals
in the group-13, cohort size, other pen characteristics, season or BVDV
activity and the risk of sero-increase between Day 0 and follow-up were
imprecise so no conclusion could be reached (Table 3).
3.4. M. bovis sero-status as a risk factor for BRD
Estimates for the association between sero-positivity to M. bovis and
the risk of BRD were imprecise but the crude incidence risk of BRD was
lower among the small number that were sero-positive at Day 0 (8.5%)
compared to those that were sero-negative (15.5%) (Table 4). There
was however strong evidence that the risk of BRD by Day 50 was in-
creased among animals that sero-increased to M. bovis between Day 0
and follow-up (OR 2.2, 95% CI 1.4–3.4, p = 0.001). This was consistent
with the higher crude incidence risk of BRD among animals that sero-
increased between Day 0 and follow-up (25.2%) compared to those that
did not (12.9%) (Table 4).
M.L. Schibrowski et al. Preventive Veterinary Medicine 157 (2018) 152–161

Table 1
Comparison of BIO K260 ELISA results at Day 0 and follow-up. (For interpretation of the references to color in this table, the reader is referred to the web version of
this article).
Follow-up BIO K260 category
Day 0 BIO K260 category Total
0 + ++ +++ ++++ +++++

Potential to sero-increase

0 773 254 143 64 31 11 1,276

+ 17 11 10 10 2 0 50

++ 4 3 2 4 0 0 13

+++ 2 4 2 1 1 0 10

Initially high

++++ 0 1 1 0 0 1 3

+++++ 0 0 0 0 0 2 2

Total 796 273 158 79 34 14 1,354

The blue shading represents those animals classified as having sero-increased to Mycoplasma bovis based on the BIO K260 ELISA category results between Day 0 and
follow-up.

3.5. Partitioning of variance
There was a high level of clustering of sero-positivity to M. bovis at
Day 0, with 10.8% variance at the PIC-13 level, 50.9% at the group-13
level and 38.3% at the animal level. The group-level ICC was 0.62 and
the design effect was 2.0. There was also a high level of clustering of
sero-increase to M. bovis between Day 0 and follow-up, with 6.6%
variance at the feedlot level, 14.5% at the cohort level, 15.6% at the
group-13 level and 63.3% at the animal level. The group-level ICC was
0.46, the cohort-level ICC was 0.30 the corresponding design effect was
3.3.
3.6. Model checking
Normal plots of random effects indicated that in all models where
they were included cohort level random effects were approximately
normally distributed. The distributions of feedlot level random effects
was hard to assess given the small number of feedlots, and the dis-
tributions of group-13 level random effects typically did not follow a
normal distribution. This is a typical finding for random effects in multi-
level logistic models with small numbers of observations per group
(Dohoo et al., 2010). Furthermore, there is evidence that model esti-
mates are usually robust to mis-specification of the random effects
(McCulloch and Neuhaus, 2011). There were three extreme values for
group-13 random effects from the model used to estimate the total ef-
fect of saleyard on sero-positivity at Day 0, but this pattern was not
observed in the equivalent direct effects model. Heteroscedasticity plots
did not show any particular patterns.
4. Discussion
This study included animals sourced throughout the beef cattle
producing regions of Australia that were then fed at different feedlots
over a three year timeframe. Hence it should be appropriate to extra-
polate study findings to the Australian feeder cattle population. Cattle
were commonly exposed to M. bovis during their time at study feedlots
and this exposure was strongly associated with an increased risk of BRD
(OR 2.2, 95% CI 1.4–3.4, p = 0.001).
The apparent sero-prevalence estimates in this study of 3.5% at Day
0 and 25.3% at follow-up, were considerably lower than the respective
estimates of 13.1% and 73.5% reported by Wawegama et al. (2016)
using an ELISA based on the recently identified MilA antigen
(Wawegama et al., 2014). As the populations of both studies were de-
rived from the NBRDI population the true prevalence would be ex-
pected to be similar. The differences in the estimates is likely to be
largely driven by differences in test performance, in particular the
lower sensitivity of the BIO K302. The sensitivity and specificity of the
MilA ELISA were estimated to be 87% and 90% using sera from animals
experimentally exposed to a strain of M. bovis known to express this
antigen and the sensitivity and specificity of the BIO K302 as 37% (95%
CI 22–54%) and 95% (95% CI 83–99%) using the same sera
(Wawegama et al., 2016). However, the aim of this study was to utilise
commercially available tests as they were the only diagnostic methods
readily available to potential users not only in Australia, but worldwide.
Sero-prevalence studies elsewhere have used commercially avail-
able diagnostic tests, including tests in the Bio-X Diagnostics® range.
Bednarek et al. (2012) reported that 78.2% (n = 3670) of clinically
healthy cattle from 361 herds covering 16 Polish provinces were sero-
positive for M. bovis using a BIO K162 M. bovis Bio-X Diagnostics® ELISA
kit. Hanzlicek et al. (2011) reported that 26.6% (n = 304) of male
crossbred beef calves purchased from a livestock auction market in
Georgia, USA were sero-positive at feedlot arrival using an unspecified
Bio-X Diagnostics® ELISA kit. It has been suggested that intensive
breeding and production are associated with M. bovis sero-positivity
(Bednarek et al., 2012) and it is possible that the intensity of farming in
Australia is less than that seen in Europe and some parts of the USA.
This factor could provide one possible explanation for the apparent
sero-prevalence in this study being at the low end of the range. Few
studies report the apparent sero-prevalence of M. bovis antibody after
feedlot arrival. Hanzlicek et al. (2011) found that 98.2% (n = 281) of
male crossbred calves purchased from a livestock auction in Georgia,
USA were sero-positive for M. bovis antibody using an unspecified Bio-X
Diagnostics® ELISA at Day 42 after feedlot entry. This estimate is much
higher than the apparent sero-prevalence reported in the current study
(25.3%, 95% CI 21.9–28.8%).
The incidence of sero-increase to M. bovis seen in this study (19.4%)
is below that reported in other studies which utilised Bio-X Diagnostics®
ELISA tests (Hanzlicek et al., 2011) or other another commercially
available test, Chekit M. bovis, Bommeli AG, Liebefeld-Bern, Switzer-
land (Tschopp et al., 2001; Arcangioli et al., 2008) and report M. bovis
sero-conversion incidences of 54.7–100%. Hanzlicek et al. (2011), used
the same criteria for sero-increase as the current study. Tschopp et al.
(2001) and Arcangioli et al. (2008) defined seroconversion as a change
from a negative result to a positive one (OD% > 80%). Feedlot

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Table 2 Table 3
Distribution of risk factors and estimated odds ratios for their total effects (and Distribution of risk factors and estimated odds ratios for their total effects (and
direct effect where indicated) on the presence of Mycoplasma bovis serum an- direct effect where indicated) on sero-increase to Mycoplasma bovis between
tibody at Day 0 based on BIO K302 ELISA results. OR: Odds ratio; CI: Day 0 and follow-up based on BIO K260 results. OR: Odds ratio; CI: Confidence
Confidence interval; Individual p values for each category are shown in the interval; Individual p values for each category are shown in the same row as the
same row as the risk factor category while the overall Wald p value is shown in risk factor category while the overall Wald p value is shown in the risk factor
the risk factor category row (bold and italics). DD: Darling Downs; Euro.: category row (bold and italics). DD: Darling Downs; Euro.: European; NSW:
European; NSW: New South Wales; NT: Northern Territory; QLD: Queensland; New South Wales; NT: Northern Territory; QLD: Queensland; SA: South
SA: South Australia; Sth: Southern; TAS: Tasmania; VIC: Victoria; WA: Western Australia; Sth: Southern; TAS: Tasmania; VIC: Victoria; WA: Western Australia;
Australia; West.: Western. West.: Western.
Risk Factor and No. of No. positive OR (95% CI) p value Putative risk factor No. with No. OR (95% CI) P value
associated categories animals (%) (%) and associated potential seroincreased
categories (%)
Breeda 1,353 47 (3.5) 0.95
Angus 761 (56.2) 24 (3.2) Ref. Breeda 1,348 261 (19.4) – 0.06
British Cross 164 (12.1) 5 (3.1) 0.6 (0.2–2.1) 0.42 Angus 758 178 (23.5) Ref. –
Hereford 58 (4.3) 2 (3.5) 0.9 (0.1–7.5) 0.94 British Cross 164 22 (13.4 0.4 (0.2–0.8) 0.01
Shorthorn 56 (4.2) 1 (1.8) 0.5 (0.0–7.8) 0.59 Hereford 58 13 (22.4) 1.1 (0.4–2.9) 0.80
Murray Grey 33 (2.4) 5 (15.2) 1.5 (0.3–7.3) 0.64 Shorthorn 56 11 (19.6) 1.0 (0.4–2.5) 0.92
Euro/Euro Cross 49 (3.6) 2 (4.1) 1.4 (0.1–15.1) 0.77 Murray Grey 33 3 (9.1) 0.4 (0.1–1.7) 0.21
Tropical/Tropical Cross 232 (17.2) 8 (3.5) 1.3 (0.3–5.7) 0.70 Euro./Euro. Cross 48 6 (12.5) 0.3 (0.1–1.0) 0.05
Tropical/Tropical 231 28 (12.1) 0.5 (0.2–1.1) 0.09
Induction weight (kg)b 1,354 47 (3.5) 0.41
Cross
190– < 400 215 (15.9) 19 (8.8) 2.5 (0.7–9.4) 0.17
400– < 440 434 (32.1) 13 (3.0) 1.2 (0.4–3.2) 0.60 Induction weight 1,349 261 (19.4) – 0.28
440– < 480 499 (36.8) 12 (2.4) Ref. (kg)b
≥480 206 (15.2) 3 (1.5) 0.8 (0.2–3.6) 0.73 190–≤ 400 214 24 (11.2) 0.7 (0.4–1.4) 0.33
400–≤ 440 432 87 (20.1) 1.2 (0.8–1.8) 0.46
Dentitionc 1,342 47 (3.5) 0.43
440–≤ 480 497 102 (20.5) Ref. –
Milk tooth 1,055 (78.6) 40 (3.8) Ref.
≥480 206 48 (23.3) 1.4 (0.8–2.3) 0.20
Two tooth 240 (17.9) 5 (2.1) 0.5 (0.2–1.6) 0.22
Four and six tooth 47 (3.5) 2 (4.3) 1.3 (0.2–9.8) 0.81 Dentitionc – 261 (19.4) – 0.50
Milk tooth 1,053 216 (20.5) Ref. –
Mixing prior to day 1,342 46 (3.4) 0.66
Two tooth 237 35 (14.8) 0.8 (0.5–1.3) 0.39
-27d
Four and six tooth 47 9 (19.1) 1.4 (0.5–3.5) 0.52
No 417 (31.1) 11 (2.6) Ref.
Yes 925 (68.9) 35 (3.8) 1.3 (0.4–4.6) 0.66 Mixing summaryd 1,337 261 (19.5) – 0.38
No : Low 58 13 (22.4) 1.5 (0.5–4.4) 0.49
Saleyard exposure 1,340 46 (3.4) 0.03
No : High 359 82 (22.8) 1.3 (0.8–2.2) 0.35
prior to day -27a
Yes : Low 283 35 (12.4) 0.7 (0.3–1.4) 0.29
No 838 (62.5) 25 (3.0) Ref.
Yes : High 637 130 (20.4) Ref. –
Yes 502 (37.5) 21 (4.2) 2.9 (1.1–7.7) 0.03
Saleyard exposure 1,335 260 (19.5) – 0.71
Saleyard exposure prior to day -27 – 0.05
prior to day -27a
(direct effect)e
No 834 157 (18.8) Ref. –
No – – Ref.
Yes 501 103 (20.6) 0.9 (0.6–1.4) 0.71
Yes – – 3.0 (1.0–8.7) 0.05
Saleyard exposure 1,349 261 (19.3) – 0.52
Source regionf 1,354 47 (3.5) 0.04
between day -27
Central & Sth 219 (16.2) 11 (5.0) Ref.
and day 0a
Tablelands NSW
No 1,274 247 (19.4) Ref. –
Coastal NSW or QLD 47 (3.5) 2 (4.3) 1.7 (0.2–15.3) 0.66
Yes 75 14 (18.7) 1.4 (0.5–3.4) 0.52
DD/New England 386 (28.5) 3 (0.8) 0.2 (0.0–1.0) 0.05
West. NSW/QLD / NT 331 (24.5) 11 (3.3) 1.1 (0.3–4.6) 0.91 Source regione 1,349 261 (19.3) – 0.01
Riverina/VIC/TAS 209 (15.4) 4 (1.9) 0.5 (0.1–2.5) 0.37 Central & Sth 218 63 (28.9) Ref. –
SA/WA 162 (12.0) 16 (9.9) 3.8 (0.8–18.9) 0.10 Tablelands NSW
Coastal NSW or QLD 47 13 (27.7) 0.8 (0.3–2.0) 0.58
Induction yearg 1,354 47 (3.5) 0.11
DD/New England 386 67 (17.4) 0.4 (0.2–0.7) 0.002
2009 163 (12.0) 11 (6.7) Ref.
West. NSW/QLD/NT 329 44 (13.4) 0.4 (0.2–0.8) 0.01
2010 458 (33.9) 22 (4.8) 0.5 (0.1–2.0) 0.33
Riverina/VIC/TAS 208 55 (26.4) 1.0 (0.5–2.1) –
2011 733 (54.1) 14 (1.9) 0.3 (0.1–0.8) 0.04
SA/WA 161 19 (11.8) 0.4 (0.1–1.0) 0.04
Number of animals in 1,354 47 (3.5) 0.06
Feedlot move timingf 1,349 261(19.3) – 0.62
group-13h
Move between day 1,294 256 (19.8) Ref. –
< 50 543 (40.1) 10 (1.8) Ref.
-27 and day 0
50–99 384 (28.4) 10 (2.6) 1.2 (0.3–4.2) 0.81
Move prior to day -27 55 5 (9.1) 0.7 (0.2–2.6) 0.62
≥100 427 (31.5) 27 (6.3) 3.0 (0.8–11.0) 0.11
Number of animals in 1,349 261 (19.3) – 0.33
a
Covariates: Source region. group-13g
b
Covariates: Source region, Induction year, Breed and Dentition. < 50 542 121 (22.3) Ref. –
c
Covariates: Induction year and Breed. 50–99 383 85 (22.2) 1.1 (0.7–1.7) 0.69
d ≥100 424 55 (13.0) 0.7 (0.4–1.3) 0.21
Covariates: Source region, Induction year, Dentition, Saleyard exposure
prior to day -27. Number of animals in 0.69
e group-13 (Direct
Covariates: Source region, Induction year, Dentition, Mixing prior to day
-27. effect)h
f < 0 Ref.
Covariates: Induction year.
g 50–99 1.1 (0.7–1.8) 0.61
No covariates.
h ≥100 0.9 (0.4–1.7) 0.63
Covariates: Source region, Induction year.
Induction yeari 1,349 261 (19.3) – 0.10
2009 163 19 (11.7) Ref. –
(continued on next page)

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M.L. Schibrowski et al. Preventive Veterinary Medicine 157 (2018) 152–161

Table 3 (continued) Table 4

Distribution of Mycoplasma bovis sero-status variables and estimated odds ratios
Putative risk factor No. with No. OR (95% CI) P value for their total effects on the occurrence of BRD by day 50. OR: Odds ratio; CI:
and associated potential seroincreased
Confidence interval; Individual p values for each category are shown in the
categories (%)
same row as the risk factor category while the overall Wald p value is shown in
2010 455 90 (19.8) 2.2 (0.9–5.3) 0.09 the risk factor category row (bold and italics).
2011 731 152 (10.8) 2.6 (1.1–6.0) 0.03 Risk Factor and associated No. of No. (%) with OR (95% CI) p
Number of animals in 1,349 261 (19.3) – 0.99 categories animals (%) BRD by day
cohorti 50
< 200 444 78 (17.6) 1.0 (0.5–1.9) 0.99
≥200 905 183 (20.2) Ref. – Sero-status to M. bovis at 1,354 207 (15.3) 0.87
Day 0a
Pens joiningi 1,344 259 (19.3) – 0.55 Positive 47 (3.5) 4 (8.5) 0.8 (0.1–6.8) 0.87
1 428 91 (21.3) Ref. – Negative 1,307 (96.5) 203 (15.5) Ref.
2 916 168 (18.3) 0.8 (0.5–1.5) 0.55
Sero-increase to M. bovis 1,349 206 (15.3) 0.001
Shared pen wateri 1,349 261 (19.3) – 0.003 between Day 0 and
No 247 23 (9.3) Ref. – follow-upb
Yes 1,102 238 (21.6) 3.3 (1.5–7.4) 0.003 Positive 261 (19.3) 66 (25.2) 2.2 (1.4–3.4) 0.001
Pen shadej 1,349 261 (19.3) – 0.31 Negative 1,088 (80.7) 140 (12.9) Ref.
No 411 62 (15.1) Ref. –
a
Yes 938 199 (21.2) 1.5 (0.7–3.0) 0.31 Covariates: source region, group size, mixing at least 27 days prior to Day 0
and history of a saleyard transfer at least 27 days prior to Day 0.
Pen density (m2/ 1,349 261 (19.3) – 0.15 b
SCU)k
covariates: for source region, breed, group size, mixing summary, shared
11– < 13 234 53 (22.6) 1.6 (0.6–4.5) 0.35 water and exposure to a persistently infected bovine viral diarrhoea virus an-
13– < 15 512 96 (18.7) Ref. – imal.
15– < 17 249 66 (26.5) 1.6 (0.8–3.4) 0.20
≥17 354 46 (13.0) 0.7 (0.3–1.6) 0.37 identified, targeted animal management strategies could be im-
Induction seasoni 1,349 261 (19.3) – 0.29 plemented to reduce the BRD risk.
Spring 405 77 (19.0) Ref. – The risk factor most strongly associated with sero-increase was
Summer 301 50 (16.6) 0.8 (0.4–1.5) 0.45
shared pen water (Table 3). This higher risk was detected when the
Autumn 280 75 (26.8) 1.4 (0.7–2.9) 0.28
Winter 363 59 (16.2) 0.8 (0.4–1.5) 0.51 water source could be accessed by animals from adjacent pens and
likely reflects the increased potential and frequency of indirect/direct
BVDV exposurel – 261 (19.3) – 0.24
Active infection, no PI 756 170 (22.5) 1.5 (0.8–2.6) 0.21
transmission of M. bovis from infected animals to susceptible animals,
Active infection, PI 143 22 (15.4) 0.9 (0.4–2.1) 0.80 coupled with the capacity for M. bovis to survive in water for extended
No active infection, 450 69 (15.3) Ref. – periods (Pfützner, 1984). This risk factor was also identified by Hay
no PI et al. (2016c), who suggested the higher risk of BRD resulting from
a water trough placement to be due to more frequent pathogen trans-
Covariates: Source region.
b mission. Importantly, this risk factor has the potential for industry to
Covariates: Source region, Induction year, Dentition, Induction season and
Breed. address, as changing the placement of water troughs to ensure they can
c
Covariates: Induction year, Induction season and Breed. only be accessed by animals within the same pen could reduce BRD.
d
Covariates: Source region, Induction year, Dentition, Number of animals in The impact of the imperfect performance of the BIO K302 and BIO
group-13, Feedlot move timing, Saleyard exposure prior to day -27 and K260 assays on sero-prevalence and sero-increase estimates and on the
Saleyard exposure between day -27 and Day 0. identification of risk factors is hard to assess. Several studies have re-
e
Covariates:. Induction year. ported highly variable estimates for the BIO K302 test performance.
f
Covariates: Source region, Induction year, Dentition and Weight. Wawegama et al. (2016) estimated the sensitivity and specificity as
g
Covariates: Source region and Induction year. 37% (95% CI 22–54%) and 95% (95% CI 83–99%) respectively, using
h
Covariates: Source region, Induction year, Dentition, Saleyard exposure
sera from an experimental challenge with a single M. bovis strain.
prior to day -27, Saleyard exposure between day -27 and Day 0, Mixing sum-
Schibrowski et al. (2018) reported sensitivity and specificity estimates
mary and Feedlot move timing.
i as 47% (95% CI 10–87%) and 96% (95% CI 87–99%) respectively,
No covariates.
j
Covariate: Induction season. using sera from experimentally challenged cattle sourced from Australia
k
Covariate: Number of animals in cohort. (a subset of animals from Wawegama et al., 2016), Canada and Eng-
l
Covariates: Source region and Number of animals in cohort. land, with each source using a different challenge strain(s). Nielsen
et al. (2015) used latentclass analyses to estimate sensitivity and spe-
management practices in Australia may differ compared to other cificity as 60.4% (95% Credible Interval [CrI] 37.5–96.2–95%) and
countries. In addition, the age of cattle at the time of feedlot entry in 97.3% (95% CrI 94.0–99.8%) respectively, using bulk milk samples
Australia (approximately 24 months of age) may have some effect on from a large number of Danish dairy herds. The wide variation and lack
colonisation of the lower respiratory tract and in turn the development of precision in these estimates across the three studies indicate that it is
of antibody. Alternatively, a low prevalence of the organism in the difficult to infer the likely sensitivity and specificity of the BIO K302 in
environment and few animals actively shedding M. bovis could provide the study population of Australian feeder cattle. Thus, although
possible explanations for the lower incidence of sero-increase seen in methods to estimate true prevalence and adjust for bias due to im-
this study. perfect test results in logistic regression analyses do exist (Rogan and
Sero-positivity at Day 0 and sero-increase by follow-up were highly Gladen, 1978; Valle et al., 2015), they were not considered useful in
clustered at the group level (ICCs of 0.62 and 0.46, respectively), in- this case. However, the consistently low sensitivity but relatively high
dicating that some groups had a high prevalence of sero-positivity and/ specificity estimates suggest that the true prevalence estimates are
or a high incidence of sero-increase. This level of clustering is greater likely to be higher than the apparent sero-prevalence estimates reported
than that typically observed (usually less than 0.2) for infectious dis- in the current study. Furthermore, if the misclassification due to im-
eases at the herd level (Otte and Gumm, 1997). This is an important perfect sensitivity (and to a lesser extent specificity) is non-differential
finding from an industry perspective, as if high risk groups could be (i.e. are independent of exposure status), the odds ratio estimates for

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M.L. Schibrowski et al.
the risk factors will have be biased towards the null (Copeland et al.,
1977).
The power to detect associations between putative risk factors and
the outcomes of interest in this study was limited because of the small
sample size and the relatively poor performance of the diagnostic as-
says, particularly the sensitivity. Although the sample size was suffi-
cient for the purpose of apparent sero-prevalence estimation, it was
inadequate for the evaluation of many risk factors largely because the
design effect for both group and cohort levels was higher than the es-
timated design effect for purchase group based on early analyses in the
NBRDI. A larger sample size was not possible in the current study due to
time and budgetary constraints. The combined impact of the small
sample size, the larger than expected design effect and the limited
ability of the assays to detect true positives, is reflected by the wide
confidence intervals for the odds ratio estimates. In this context, risk
factors assessed and found to have no significant association with sero-
positivity to M. bovis or sero-increase at follow-up should not be com-
pletely dismissed, particularly if the point estimate for the odds ratio is
well away from one (i.e. < 0.8 or > 1.5). Rather, a larger risk factor
study, using a more sensitive test is warranted.
Bovine respiratory disease is multi-factorial with many physical,
physiological (Taylor et al., 2010) and infectious stressors combining to
predispose susceptible animals to the development of clinical disease.
There are many viral and bacterial pathogens that are consistently
implicated in BRD. Internationally, it has been suggested that M. bovis
can play an important role in the development of BRD (ter Laak et al.,
1992; Burnens et al., 1999; Le Grand et al., 2002; Tenk et al., 2004;
Arcangioli et al., 2008). Despite this, the role of M. bovis in the devel-
opment of BRD in Australia has received little attention perhaps due the
cattle entering feedlots in this country being older compared to else-
where in the world (Horwood et al., 2014). Preliminary investigations
by Horwood et al. (2014) demonstrated that M. bovis was present in
both clinical and fatal cases of BRD in Australian feedlots. The study
assessed lung (n = 2) and tracheal samples (n = 12) from animals that
had died from BRD for the presence of M. bovis using quantitative real-
time PCR. Mycoplasma bovis was the most frequently detected pathogen
in the tissue samples, and in most cases, was detected in combination
with bovine herpesvirus 1. Although preliminary in nature, this study
did provide evidence for the inclusion of M. bovis to the list of patho-
gens implicated in BRD in Australia. The current study, has linked ex-
posure to M. bovis and BRD thus adding further evidence to support an
important role for this pathogen in Australian feeder cattle.
Unexpectedly, the current study did not show a significant decrease
in the risk of BRD among cattle that were sero-positive at feedlot in-
duction (Table 4). In contrast, Hay et al. (2016c) reported that sero-
positivity to the viruses commonly associated with BRD at induction
reduced the risk of BRD and it is reasonable to expect that prior ex-
posure to a pathogen would reduce the BRD risk. While the point es-
timate for the association between sero-positivity at entry and BRD risk
suggest a protective effect, it was imprecise (Table 4). The relative small
sample size of the current study and the lower crude incidence risk of
BRD among cattle that were sero-positive at Day 0 suggest that a pos-
sible association cannot be dismissed and further research is required.
Sero-increase to M. bovis-specific antibody between Day 0 and
follow-up was found to be strongly associated with an increased risk of
BRD in the first 50 days on feed (OR 2.2, 95% CI 1.4–3.4, p = 0.001).
This association is stronger than the association reported between sero-
increase to bovine herpesvirus 1 (OR 1.4, 95% CrI 1.2–1.6), BVDV-1
(OR 1.3, 95% CrI 1.1–1.6), bovine respiratory syncytial virus (OR 1.5,
95% CrI 1.3–1.7) and bovine parainfluenza virus 3 (OR 1.4, 95% CrI
1.1–1.6) (Hay et al., 2016b). Of highest priority on the Bradford Hill
criteria for assessing causality is the strength of association (Hill, 1965).
The strength of association between sero-increase to M. bovis and the
occurrence of BRD seen in this study, especially when compared to that
seen with commonly implicated viruses, provides support for a causal
association of M. bovis in the development of BRD. Further
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Preventive Veterinary Medicine 157 (2018) 152–161
investigations into possible synergistic interactions between M. bovis,
BVDV-1 and other viruses implicated in BRD are warranted.
5. Conclusions
This study provides evidence that exposure to M. bovis is relatively
common among cattle in Australian feedlots and increases the risk of
BRD. These findings can be used to develop targeted control measures
and help reduce the economic impact of M. bovis-associated disease and
BRD. Eliminating the use of shared water sources between feedlot pens
has the potential to reduce the BRD risk of cattle. Additional research is
required to investigate possible synergistic interactions between M.
bovis and viruses in the pathogenesis of BRD.
Acknowledgements
This study was supported by grant B.FLT.0224 from Meat and
Livestock Australia with matching funds provided by the Australian
Government. MLS was supported by an Australian Postgraduate Award
from the Australian Government.
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