PCR Detection of Mycoplasma hyopneumoniae in Processing Fluids: a Diagnostic Error or a Significant Finding? - Dr. Carlos Vilalta, from the 2018 Allen D. Leman Swine Conference, September 15-18, 2018, St. Paul, Minnesota, USA. More presentations at http://www.swinecast.com/2018-leman-swine-conference-material

1. PCR detection of Mycoplasma
hyopneumoniae in processing
fluids: A diagnostic error or a
significant finding?
Carles Vilalta, Juan Sanhueza, Deb Murray, Maria Pieters

2. Clinical importance of Mycoplasma
hyopneumoniae (MH)
• Enzootic pneumonia (EP).
• Primary agent of porcine
respiratory disease complex
(PRDC).
• Economic impact.

3. Colonization of pigs with MH
• Initial group colonization with MH determines downstream
clinical presentation and disease severity (Fano et al., 2007; Sibila
et al., 2007).
• The influence of the duration of lactation on piglet
colonization.
– Proof of concept - segregated early weaning (SEW) to obtain MH-free pigs born to
positive dams (Alexander et al., 1980; Pieters et al., 2014).
• Transmission of MH among pen-mates is slow (Meyns et al.,
2004; Villarreal et al., 2011; Roos et al., 2016)

4. MH epidemiology in the breeding herd
• Older sows may transmit MH to naive gilts in MH positive herds.
• Infected gilts:
– Maintain the pathogen within the farm
– Are responsible for the majority of bacterial shedding to
newborn pigs
• Adequate gilt acclimation to reduce the bacterial shedding at first
farrowing is needed (Pieters and Fano, 2016)

5. Where do we look for MH?
• MH is primarily identified on the mucosal
surface of the trachea, bronchi and
bronchioles.
• Can be found in the nasal cavity and lower
respiratory tract (laryngeal and tracheal
swabs).
• Lung lesions.

6. MH in other organs
• Friis et al (1974): nasal cavity and brain. Experimentally infected
pigs.
• Le Carrou et al (2006): liver, spleen, kidneys and bronchial
lymph nodes. Experimental inoculation.
• Marois et al (2006): liver and spleen. Infected and contact pigs.
• Wooley et al (2012): spleen, kidney and liver. Experimentally
infected pigs.
• Seems to be a strain related and transitory phenomenon.

7. Processing fluids
• Fluids that derive from
the testicles at
castration and clipping
of the tails and
accumulate at the
bottom of the container.
Vilalta et al., Vet
Microbiology (2018)
• MH not expected BUT DETECTED!!!

8. Objective
• Investigate the anecdotal
reporting of rt-PCR
detection of MH on PF in a
positive farm.

9. Material and methods - Farm
• Filtered farm
located in southern
MN, NW IA
• Two-site sow farm
(3000 sows/site)
Continuous flow
GDU off-site.
1 mile
3000 feet
Site 1
Site 2

10. Material and methods - Farm
• Animals PRRSV negative at the collection point. PRRSV
positive on the second collection set (wild type
introduction into the farm)
• MH positive flow, with no activity in the breeding herd and
with very little downstream activity.
• Clinical signs/lesions in the downstream population is
minimal. Occasional clinical signs in barns, but not many

11. Material and methods - Farm
Antibiotic
treatment
MH VX Circo/MH VX
MH gilts
Nursery
AI-AO
Continuous
flow breeder
Continuous
flow GDU
6 w 4-8 w
3 w

12. Material and methods - Samples
• 21 litters in the same room were sampled:
– Testicles and tails were bagged separately.
– 80 incision swabs from males.
– Sow feces.
– Sow laryngeal swabs and parity.
– Four previously daily PF were collected.
• Collection of several samples of daily PF

13. Results
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21
Testicles
Tails
Swabs NC NC NC NC NC NC NC NC
Laryngeal NC NC NC NC NC
Feces NT NT NT NT NT NT NT NT NT NT NT NT NT
Orange: positive Blue: negative Green: gilts NC: not collected NT: not tested
8/21 (38%) of testicle fluids were positive.
Gilts Odds Ratio 3.2 (4/3)/(4/10)
1/21 (4.8%) of tails were positive.
2/80 (2.5%) of incision swabs were positive

14. Results

15. 14/32 pos PF
25
30
35
40
45
Sep Oct Nov Dec Jan Feb
CtValues
Month
MH Ct Results of daily aggregated PF
Neg
Results
2/4 pos PF

16. Results
MH confirmed
by nested
PCR and
whole
genome
M. Suis was
ruled out.

17. Discussion
• Can Mycoplasma spp. be found in testicles?
– Yes. Myco spp colonizes mucosal surfaces in oral cavity, the
respiratory tract, the eyes, the urogenital tract and the synovial lining
of joints.
– Mycoplasma genitalium has been isolated from
human respiratory tract. Mycoplasma
pneumoniae has been isolated from human
urogenital tract in males.

18. Discussion
• These results raise a lot of questions on the
epidemiology of MH.
– How MH got to the testicles?
– When are the piglets colonized?
• In utero?
• At birth?
• After birth?
– What is the role of gilts in MH epidemiology?

19. Conclusions
• Mycoplasma hyopneumoniae can be detected
in the processing fluids. More specifically, the
testicles seem to be the source of bacterium.
• These findings open the door to new questions
about the epidemiology of MH.

20. Acknowledgements
• Albert Rovira
• Stephanie Rossow
• Fabio Vanucci Funding

21. cvilalta@umn.edu
Thanks for your
attention