Background: The microsporidian Enterocytozoon hepatopenaei was first described from Thailand in 2009 in farmed,
indigenous giant tiger shrimp Penaeus (Penaeus) monodon. The natural reservoir for the parasite is still unknown.
More recently, a microsporidian closely resembling it in morphology and tissue preference was found in Thai-farmed,
exotic, whiteleg shrimp Penaeus (Litopenaeus) vannamei exhibiting white feces syndrome (WFS). Our objective was to
compare the newly found pathogen with E. hepatopenaei and to determine its causal relationship with WFS.
Results: Generic primers used to amplify a fragment of the small subunit ribosomal RNA (ssu rRNA) gene for cloning and
sequencing revealed that the new parasite from WFS ponds had 99% sequence identity to that of E. hepatopenaei,
suggesting it was conspecific. Normal histological analysis using tissue sections stained with hematoxylin and eosin (H&E)
revealed that relatively few tubule epithelial cells exhibited spores, suggesting that the infections were light. However, the
H&E results were deceptive since nested PCR and in situ hybridization analysis based on the cloned ssu rRNA gene fragment
revealed very heavy infections in tubule epithelial cells in the central region of the hepatopancreas in the absence of spores.
Despite these results, high prevalence of E. hepatopenaei in shrimp from ponds not exhibiting WFS and a pond that had
recovered from WFS indicated no direct causal association between these infections and WFS. This was supported by
laboratory oral challenge trials that revealed direct horizontal transmission to uninfected shrimp but no signs of WFS.
Conclusions: The microsporidian newly found in P. vannamei is conspecific with previously described E. hepatopenaei
and it is not causally associated with WFS. However, the deceptive severity of infections (much greater than previously
reported in P. monodon) would undoubtedly have a negative effect on whiteleg shrimp growth and production
efficiency and this could be exacerbated by the possibility of horizontal transmission revealed by laboratory challenge
tests. Thus, it is recommended that the PCR and in situ hybridization methods developed herein be used to identify
the natural reservoir species so they can be eliminated from the shrimp rearing system.