Influence of Selective Agents (EMJH-STAFF), Sample Filtration and pH on Leptospira interrogans Serovar Icterohaemorrhagiae Cultivation and Isolation from Swine Urine

Abstract

Leptospira spp. cause the zoonotic disease leptospirosis, which occurs in numerous mammalians worldwide. Isolation is still important for serotyping and genotyping of Leptospira, which in turn is essential for epidemiological surveillance of leptospirosis and the development of diagnostic tests and vaccines. However, isolation of Leptospira from clinical specimens is inherently insensitive. This study was conducted to examine the influence of selective agents, sample filtration, sample pH and the use of phosphate buffered saline (PBS) buffer for sample storage to improve the success of cultivation and isolation of Leptospira interrogans serovar Icterohaemorrhagiae from swine urine. EMJH (Ellinghausen McCullough, Johnson and Harris) medium including the selective agents sulfamethoxazole, trimethoprim, amphotericin, fosfomycin and 5-fluorouracil (STAFF) increased the success of Leptospira isolation from spiked swine urine samples. Sample filtration yielded only negative results. Isolation in EMJH-STAFF was successful from swine urine with a density as low as 104 Leptospira/mL, and urine with pH ≤ 7 impaired the cultivation rate. Cultivation and isolation were not improved by the addition of PBS to spiked urine samples prior to storage for 24 h at 4 °C. The results of the study demonstrate that cultivation and isolation of leptospires from swine urine can be improved by enhanced methods.

Leptospira interrogans Serovar Icterohaemorrhagiae Failed to Establish Distinct Infection in Naïve Gilts: Lessons Learned from a Preliminary Experimental Challenge

“…Furthermore, the sera, urine pellets, vaginal swabs, tissue samples, and manure were tested for Leptospira spp. by real-time PCR targeting the leptospiral outer membrane lipoprotein gene lipL32 [ 37 ] following sample type-specific pre-treatment and nucleic acid extraction; no pre-treatment was performed for serum samples. Urine pellets were dissolved in 140 µL PBS.…”