High frequency and high genetic diversity of Pneumocystis sp. in foals.

Overview

• This study investigated the presence and genetic diversity of the fungus Pneumocystis sp. in foals, revealing a high frequency of infection and the existence of multiple strains, including a possible new species infecting horses.

Study Subjects and Samples

• 22 foals aged between one and seven months were examined post-mortem.
• 15 foals had died from interstitial pneumonia while 7 died from unrelated causes.
• A total of 52 lung tissue specimens were collected from these foals for analysis.

Detection and Identification of Pneumocystis

• Pneumocystis DNA was detected in 21 out of 22 foals and in 48 out of 52 lung specimens using a PCR assay targeting the mitochondrial small subunit rRNA (mtSSUrRNA) gene specific to horse-related Pneumocystis species.
• The PCR products were successfully sequenced in 34 of the 48 positive specimens.

Genetic Diversity Findings

• Sequencing revealed:
  • Seven specimens (from five foals) had sequences identical to a known horse-associated Pneumocystis reference sequence.
  • 24 specimens (from 16 foals) showed single nucleotide polymorphisms (SNPs), indicating variation.
  • Three distinct alleles were identified based on these SNPs.
  • Allele identification was clear in 10 specimens (7 foals), but 21 specimens (14 foals) contained mixed alleles, suggesting co-infection with multiple Pneumocystis strains.
• Three sequences from two foals did not align well with the reference sequence, showing only 86.12% identity, hinting at a possible different Pneumocystis species.

Advanced Genetic Analysis

• Ultradeep sequencing of the mitochondrial large subunit rRNA (mtLSUrRNA) gene was performed on 5 selected specimens.
• The sequences from mtLSUrRNA were combined with mtSSUrRNA sequences for detailed phylogenetic analysis.
• Phylogenetic results suggested:
  • The Pneumocystis strains from foals cluster closely with a Pneumocystis species known from pigs (Sus scrofa domesticus).
  • This closeness implies the existence of another, previously uncharacterized Pneumocystis species infecting foals in addition to the known horse-associated species.

Conclusions and Implications

• Pneumocystis infection is common in young foals, as indicated by the high detection rate.
• The fungus exhibits significant genetic diversity within the foal population, including multiple strains infecting the same individual.
• There may be at least one other distinct Pneumocystis species capable of infecting foals, broadening understanding of Pneumocystis host species range.
• These findings contribute novel insights into Pneumocystis epidemiology and genetic diversity in equine hosts, potentially influencing diagnosis and treatment strategies in veterinary medicine.