The objective of this study was to evaluate the use of real-time TaqMan PCR assays for detection of coinfections with “ Candidatus Mycoplasma haemominutum” (Mhm), and Mycoplasma haemofelis (Mhf), in vitro and over time in experimentally infected cats. First, the ability of each real-time PCR assay to detect and quantify mixed infections was determined in vitro by testing mixtures of plasmids containing Mhm and Mhf 16S rDNA with each assay. Subsequently, 4 specific pathogen-free (SPF) cats, 2 of which were splenectomized, were inoculated with blood from a cat infected with both Mhm and Mhf. Sixteen blood samples were then collected from each cat over a 55-day period. Each of the 64 postinoculation samples was tested using both conventional polymerase chain reaction (cPCR) and real-time PCR for the 16S rRNA gene of each organism. When applied to mixtures of plasmid DNA from each species, the results of quantitation with each of the real-time PCR assays approximately reflected the number of plasmid copies present. Forty-nine of 64 post-inoculation samples (77%) were positive using both cPCR and real-time PCR, 4 (6%) were positive using cPCR only, and 3 (5%) were positive using real-time PCR only. Both organisms were detected in 23 samples using real-time PCR. Mixed infections were not detected using cPCR. The size of the corresponding cPCR products suggested infection with Mhm in 4 and Mhf in 18 of these samples. The use of multiple separate real-time PCR assays rather than cPCR alone should thus be considered for epidemiologic studies of hemoplasmosis in cats.
The aim of this study was to evaluate by PCR the presence of Helicobacter spp. in gastric mucus from the fundic region of the stomach and to investigate its role in oesophagogastric ulcers in swine bred and regularly slaughtered in Piedmont (Northern Italy). Stomachs from 595 regularly slaughtered swine were subjected to gross pathological examination in order to evaluate the presence of gastric ulcers (revealed in 75 cases, 12.6%). Histopathological examination was performed to better characterise erosions and ulcers. DNA extracted from gastric mucus collected from all the ulcer-affected and from 25 normal stomachs was submitted to PCR using Helicobacter spp. 16S rRNA gene target primers. Sixty-three percent (47/75) of the affected stomachs was positive as well as 24% (6/25) of the non-affected ones. Sequence analysis from 5 positive samples showed 99% homology with Helicobacter candidatus suis 16S ribosomal RNA gene.