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ABSTRACTInsertion sequence IS900is used as a target for the identification of Mycobacterium aviumsubsp. paratuberculosis.Previous reports have revealed single nucleotide polymorphisms within IS900. This study, which analyzed the IS900sequences of a panel of isolates representing M. aviumsubsp. paratuberculosisstrain types I, II, and III, revealed conserved type-specific polymorphisms that could be utilized as a tool for diagnostic and epidemiological purposes.

The intraerythrocytic protozoans Theileria equiand Babesia caballiare the causative agents of equine piroplasmosis (EP), one of the most important equine tick-borne diseases due to its significant impact on global international horse trade. Although EP is known to be endemic in Spain, previous phylogenetic studies have only been conducted for limited geographical regions. Therefore, the objective of this study was to evaluate the genetic diversity and distribution of these parasite species nationwide. This was performed by amplification of the 18S small subunit (SSU) rRNA gene from 100 EP positive equine blood samples using a nested PCR protocol, and sequencing the obtained amplicons. Seventy-seven T. equiand six B. caballiisolates were successfully sequenced and phylogenetic analysis revealed that the T. equiisolates grouped into the previously described clades A (n = 21/77), D (n = 1/77) and E (n = 55/77), while B. caballiisolates were placed into clades A (n = 5/6) and B (n = 1/6). Isolates from T. equiclade D and B. caballiclade B have not previously been reported in Spain. A greater intra-clade diversity (97.3–98.3 % identity) was observed between T. equiclade E isolates compared to those within clade A (99.7–100 % identity). Additionally, a multivariable logistic regression model was used to analyse associations between the clade of T. equiinfection and available epidemiological data. Horses residing in Spanish northern regions were statistically more likely to be infected with T. equiclade E (p = 0.01). We conclude that while extensive sequence variation of equine piroplasms exists in Spanish infected horses, a requirement for increased equine movement controls between Spain and EP-endemic countries should be considered.