Laboratory of Epidemiology, Biostatistics and Animal Health Economics, University of Thessaly, 224 Trikalon st, 43100, Karditsa, Greece
Veterinary Research Institute, National Agricultural Research Foundation, Nagref Campus, PO Box 60272, 57001, Thermi, Greece
Abstract
Background
Piglet isosporosis is one of the most common parasitic diseases in modern pig production. To prevent clinical disease, prophylactic treatment of piglets with toltrazuril (BAYCOX® 5%, Bayer HealthCare, Animal Health, Monheim, Germany) is widely practiced in the past 20 years. There are only very few reports documenting the likely effect of managerial practices, such as hygiene measures, allinallout management of farrowing facilities and piglet manipulations, and/or farmspecific environment  i.e. design and materials of the farrowing pen and room  in the risk of disease occurrence and transmission. Therefore, in this crosssectional study, we identified litter and herdlevel factors associated with the odds and the level of
Results
Factors associated with lower odds of oocyst excretion were: use of toltrazuril treatment, allin allout management of the farrowing rooms, no crossfostering or fostering during the first 24 hours after farrowing, plastic flooring in the farrowing pens, farrowing rooms with more than fourteen farrowing pens and employment of more than two caretakers in the farrowing section. Factors associated with lower oocyst excretion level were: use of toltrazuril treatment and caretakers averting from entering into farrowing pens.
Conclusion
Apart from prophylactic treatment with toltrazuril, the risk and the level of
Background
Several aspects of the
Methods
Pilot study
A pilot study, including ten herds, randomly selected from the country’s national registry, was conducted before the initiation of the study, to obtain prior information for estimation of the required sample sizes. From each of these ten herds, ten randomly selected litters in their second week of life were sampled and tested as described below (section ‘Parasitological methods’). Based on the data collected from the pilot study, we estimated an intraherd correlation coefficient (ICC) for oocyst excretion of 0.4, and detected excretion in 20% and 80% of the litters in herds treated (six herds) or not (four herds) with toltrazuril (BAYCOX® 5%, Bayer HealthCare, Animal Health, Monheim, Germany), respectively.
Sample size determination
Based on the results of the pilot study a minimum difference of 60% was expected in the proportion of positive litters between those treated and not treated with toltrazuril. The minimum required sample size for comparing two proportions, based on the standard sampling formulae, equals to 26 litters, 13 for each group. Sample size calculations were done with the Piface java applet
A twostage sampling design was used. Initially, herds were randomly selected from the country’s national registry, after excluding herds with less than 20 sows and those located on the Greek islands. Consent for participation in the study was obtained after personal communication of the owner or the manager of the farm with the primary author. When they denied participation, the farm located closest was contacted and, if agreed to participate, was sampled. Subsequently, litters were randomly sampled within the selected herds.
Collection of questionnaire data
To collect data on factors likely affecting the risk of oocyst excretion we developed a questionnaire (see Additional file
Piglet isosporosis questionnaire.
Click here for file
The data included information on herd size, production parameters, housing conditions, managerial strategies, disease prevention, hygiene practices, cleaning and disinfection procedures, farrowing room and pen design as well as application of toltrazuril treatment. Fiftythree questions were closed (e.g. yes/no, always/frequently/seldom/never or preset options), twentyone were semiclosed (e.g. information on number of days, farrowing pens and rooms, application frequencies of certain procedures) and the remaining were openended (e.g. product names, descriptions). The interviewer checked the accuracy of some data, such as size and location or flooring of the farrowing pen, by inspecting the farrowing facilities. Questionnaires were filledin, before sampling, with personal interview of either the owner or manager of the herd by the first author (VS). The interviewer had no prior knowledge of the
Parasitological methods
From each of the selected herds five to ten litters in their second week of life (from day 8 to 14 post farrowing (p.f.)) were sampled. The crosssectional type of the study and the fact that all the studied farms were managed with continuous farrowing, justified the variable number of sampled litters. Evidently, at the herd visit we could find more litters in the second week of life in larger than in smaller herds. Each sampled litter represented a different number of litters in small than in large herds. Faecal samples were collected, using a swab, from the rectum
Each pooled faecal sample was considered positive for oocyst excretion if the minimum concentration of 20 OPG was detected and the respective litter was classified as one that excreted oocysts. When all litters from a herd were negative, we used a standard protocol of resampling the same litters one week later to minimize the probability of misclassifying an infected herd
Statistical analysis
Assessing the odds and the level of oocyst excretion
OPG counts were semi continuous data (Figure
Frequency distribution of the natural logarithm of
Frequency distribution of the natural logarithm of
We employed a twopart model with the presence or absence of oocyst excretion at the litterlevel as the response variable in the first part of the model, while the natural logarithm of the nonzero OPG was the response variable in the second part. Randomeffect terms were incorporated in the model to account for the within herd correlation of observations for both the odds and level of oocyst excretion. Furthermore, to capture the biologically plausible fact that herds with higher rates of oocyst excretion may also have higher OPG counts, we adopted a structure that adjusted for the crossequation correlation of the randomeffects terms between the logistic and linear part of the model
For model building, all candidate variables were initially screened, onebyone, using a bivariable approach
Goodness of fit
To check the adequate fit of the model to the observed data, we simulated the infection status (oocyst excretion or not) and the amount of oocysts excreted for each litter, under the final model. The simulated data under each part of the model were then compared to the observed data
Statistical software
Estimation of the twopart random effects model was performed using the freely available software aML (
Results
All but one of the selected farmers consented to participate in the study. Data from five herds were not considered in the analyses because the farmers failed to adequately fillin several questions. Thus, the data analysed comprised of fiftyfive herds, totalling 314 sampled litters. Only two out of the ten herds which were initially found negative, were found positive at resampling while the remaining eight herds tested negative. From the herds found infected, 146 of the sampled litters tested positive (Table
Definition of factor
Level
Number of litters
Proportion of positive litters
Mean (SD) ln(OPG)
Dual Pvalues correspond to factors that were offered to both parts of the model. Data are from 314 litters in 55 farrowtofinish Greek pig herds.
^{a}
Part I + Part II
Use of toltrazuril
Yes
200
0.30
6.6 (3.1)
0.001^{a}
No
114
0.65
8.2 (2.7)
0.002^{b}
Piglet age at weaning
c


0.116^{a}
0.190^{b}
Number of pens per farrowing room^{*}
>14
154
0.31
7.0 (3.3)
0.035^{a}
≤14
160
0.52
7.6 (2.8)
0.067^{b}
Remaining faeces after cleaning
Yes
88
0.52
8.0 (3.0)
0.056^{a}
No
226
0.37
7.1 (2.9)
0.145^{b}
Size of farrowing pen in m^{2*}



0.042^{a}
0.113^{b}
Part I
Crossfostering of piglets
Within the 1st day p.f.^{**} no fostering
60
0.26
6.8 (2.5)
0.061
After 1st day p.f.
254
0.44
7.4 (3.1)
Cleaning entire farrowing room
Yes
193
0.34
7.4 (2.7)
0.077
No
121
0.63
7.3 (3.5)
Disinfection of farrowing unit on a regular basis
Yes
290
0.40
7.4 (3.0)
0.050
No
24
0.56
7.6 (3.3)
Flooring of farrowing pen in creep area^{*}
Perforated
91
0.27
6.3 (3.1)
0.070
Solid
223
0.51
7.8 (2.9)
Number of caretakers in farrowing unit
>2
55
0.24
6.3 (2.7)
0.186
≤2
259
0.48
7.6 (3.0)
Farrowing pen flooring^{*}
Plastic
89
0.38
7.1 (3.3)
0.160
Metal
225
0.41
7.5 (2.9)
Slatted portion of farrowing pen’s flooring^{*}
30%
55
0.27
8.5 (2.3)
0.171
60%
111
0.48
7.7 (2.6)
100%
148
0.40
6.7 (3.3)
Part II
Use of drying substances
Yes
115
0.31
7.3 (2.9)
0.170
No
199
0.48
7.4 (3.1)
Caretakers entering pens
Yes
294
0.41
7.5 (3.0)
0.110
No
20
0.34
5.7 (2.4)
Mechanical ventilation in farrowing unit
Yes
207
0.39
7.5 (2.6)
0.169
No
107
0.45
7.0 (3.8)
Washing water
Cold
255
0.40
7.4 (3.2)
0.002
Hot
59
0.38
7.1 (2.5)
After questionnaire data compilation, fiftythree variables (18 litter and 35 herdlevel) were initially screened as candidates for inclusion in the final twopart model. Those with
Part I
* Litterlevel variables. ** Post farrowing.
Variable
Parameter
OR
95% C.I.
Use of toltrazuril
Yes
1

No
3.7
1.6; 9.1
0.005
Clean entire farrowing room
Yes
1

No
3
1.0; 9.0
0.045
Crossfostering of piglets
Within the 1st day p.f.^{**} or no fostering
1

After 1st day p.f.
4.2
1.2; 15.6
0.039
Farrowing pen flooring^{*}
Plastic
1

Metal
2.8
1.2; 6.5
0.022
Number of pens per farrowing room^{*}
>14
1

≤14
3.7
1.8; 7.7
0.001
Number of caretakers in farrowing unit
>2
1

≤2
2.7
1.3; 5.9
0.015
Part II
Variable
Parameter
Coeff.
95% C.I.
Use of toltrazuril
Yes
−1.9
−0.8; 3.1
0.002
No
0

Caretakers entering pens
Yes
1.3
0.2; 2.5
0.024
No
0

Litters in herds using toltrazuril treatment had 3.70 [95% Confidence Interval (CI): 1.56; 9.10] times lower odds of being positive compared to litters in herds not treating. Application of cleaning and disinfection procedures to the entire farrowing room, after all litters were weaned, compared to splitweaning of the room and cleaning only the pens of the weaned litters, decreased the odds of litter positivity by 3.00 (1.03; 9.00) times. Litters in which crossfostering of piglets was performed after the first day p.f. had 4.20 (1.15; 15.64) times higher odds of positivity, compared to either not applying crossfostering of piglets or performing this practice during the first 24 hours p.f. Litters in farrowing pens with metal perforated floor had 2.81 (1.19; 6.52) times higher odds of excreting oocysts, compared to litters in pens with plastic perforated floor. Litters in farrowing rooms with more than fourteen pens had 3.70 (1.81; 7.69) times decreased odds in excreting oocysts. Finally, in the herds where more than two caretakers were employed in the farrowing section, litters had 2.70 (1.28; 5.88) times lower odds of being identified as positive.
From the linear part, litters in herds using toltrazuril treatment had 7.03 (2.23; 22.20) times lower mean OPG compared to herds not using toltrazuril treatment. Further, litters in herds where caretakers of the farrowing section entered in the farrowing pens, in order to perform the necessary manipulations in piglets, had 3.85 (1.22; 11.94) times higher mean OPG compared to herds where caretakers avoided entering the farrowing pens.
The herdlevel variance was 0.82 (standard error: 0.19) and 1.49 (0.33) for the logistic and the linear part, respectively. The cross equation correlation between the herdlevel random effects of the logistic and the linear part was 0.95, revealing that herds at higher risk of excretion were more likely to also experience higher excretion levels (
Discussion
We performed this crosssectional study in order to identify litter and herdlevel factors affecting the odds of occurrence and the level of
Risk factors
Application of early routine treatment with toltrazuril reduced both the odds and the level of oocyst excretion. The efficacy of this treatment against piglet coccidiosis has been previously demonstrated in experimentally infected piglets
Herds practising crossfostering of piglets after day one p.f., had litters with higher odds of oocyst excretion compared to litters in herds where crossfostering was not applied or done within the first 24 hours p.f. Late crossfostered piglets may shed earlier and may be responsible for litter overcontamination. Early fostering reduces exposure of piglets to the stress of fostering. The severity and duration of diarrhoea are greater and earlier in crossfostered piglets compared with resident counterparts despite that fact that the former excrete less oocysts
Cleaning and disinfecting the entire farrowing room after all litters were weaned reduced the odds of oocyst excretion. The proposed approach is an indication of a relatively high standard in the practiced hygiene procedures in a farm, as part of an integrated allinallout management. Reduced environmental contamination, by thorough cleaning, can be effective in preventing or delaying initial
We failed to identify a significant association of the perforated proportion of pen flooring with the odds or the level of oocyst excretion. However, a significant effect was observed for the type of perforated flooring: plastic flooring decreased the risk of oocyst excretion compared to metal flooring. Likely, this is because plastic flooring is a material that can be easily and effectively cleaned, thus, leading to a reduced number of ingested oocysts. Metal flooring, compared with plastic, is harder to maintain clean
Litters in farrowing rooms with large number of farrowing pens had decreased odds of occurrence of oocyst excretion. An important route of
The presence of more than two caretakers in the farrowing section resulted in decreased odds of oocyst excretion. The existence of more caretakers could provide adequate manpower to maintain a relatively constant and high level of hygiene. Moreover, hygiene is sometimes neglected during workintensive periods
Conclusions
We have assessed the impact of several managerial factors on the odds and the level of
Abbreviations
(ICC): Intraherd correlation coefficient; (VIF): Variance inflation factor; (p.f.): Post farrowing; (OPG): Oocysts per gram of faeces.
Competing interests
The authors declare that they have no competing interests.
Authors’ contributions
VS carried out the collection of the samples, parasitological testing, statistical analysis and drafted the manuscript. SS participated in the design of the study, the parasitological testing of the samples and the drafting of the manuscript. PK participated in the design of the study, the development of the statistical models and the drafting of the manuscript. LL conceived and coordinated the carryout of the study and was involved in the statistical analysis, the drafting and the critical review of the manuscript. All authors have read and approved the final manuscript.
Acknowledgements
This study was financially supported by a FP6 Marie Curie ERG (contract nr: MERCCT2004516482).