Majority of the patients with H. pylori infection were in the age range of 18-39 years, while H. felis and H. heilmannii positive patients did not show this distribution. (Table ). There was no difference in the gender, ethnicity of patients, crowding index (CI) and source of water distribution among the patients with H. pylori and non-H. pylori infections (Table ). All patients had abdominal pain with endoscopic gastritis as the predominant finding. The false positive and false negative results obtained with RUT were 15(36%) and 6(12%), respectively while with histology the false positive and false negative results obtained were 20(30%) and 10(11%), respectively (Table -).
Demography and clinical features of patients enrolled
PCR results for Helicobacter species
PCR for Helicobacter genus specific 16S rDNA was positive in 167/250 (67%), glmM (H. pylori) in 142/250 (57%), H. heilmanii in 17/250 (6%) and H. felis in 10/250 (4%), respectively (Table ).
PCR was positive for both H. pylori and H. heilmannii in 17(6%) and for H. pylori and H. felis in 10(4%), respectively (Table ). All the H. heilmannii and H. felis positive patients were also positive for H. pylori glmM PCR amplification (Table ).
26% (66 out of 250) were exposed to pets either cats or dogs. Most H. heilmannii positive patients did not have pet contact. Only one out of 66 exposed to pets was positive for H. heilmannii and two for H. felis (Table ).
Association of Helicobacter species with pets
A higher degree of bacterial density was associated with H. pylori infection alone (p < 0.001) (Table ). Chronic active inflammation was observed in 53(56%) cases with H. pylori alone infection (p = 0.001) compared to 3(37%) in H. heilmannii (p = 0.73) and 3(60%) in H. felis positive patients coinfected with H. pylori (p = 0.66) (Table ). Intestinal metaplasia (IM) was present in 3(3%) out of 94 cases with H. pylori infection alone compared to 2(25%) out of 8 cases of H. Heilmannii and H. pylori coinfection, and 1(20%) out of 5 cases of H. felis and H. pylori coinfection in which histology has been performed.
PCR product sequences were compared to the sequences of ureaseB of different H. heilmannii and H. felis strains. The H. heilmannii sequences had 100% similarity to 'Candidatus Helicobacter heilmannii' strains GenBank: AF508012 and L25079; while it was 99% to GenBank: AY139170, AF507996, AY139172, AY139173 and 98% to GenBank: AY139171, respectively. The H. felis sequences had 100% similarity to H. felis strains GenBank: FQ670179 and X69080; while it was 99% to H. felis GenBank: AY368267, AY368261 and 98% to GenBank: DQ865138, respectively.
Among our patients, the cohort exposed to pet animals was limited to 26%. There were more patients with H. pylori infection who were in the 18-39 years age range. Such age distribution was not seen in cases with H. felis and H. heilmannii infection. There was no difference in the gender, ethnicity of patients, crowding index (CI) and source of water distribution among the patients with H. pylori and non-H. pylori helicobacter species infections. There were no statistically significant differences in the endoscopic findings in patients with H. pylori infection alone or with coinfection of H. pylori and non-H. pylori Helicobacter species. Chronic active inflammation was associated with H. pylori infection compared to H. heilmannii or H. felis coinfections with H. pylori (Table ). However, the histology was not obtained in all the cases that showed H. heilmannii and H. felis infection. Intestinal metaplasia was present in 2(25%) out of 8 cases of H. heilmannii coinfection with H. pylori and in 1(20%) out of 5 cases of H. felis coinfection with H. pylori as compared to 3(3%) of 94 cases with H. pylori infection alone who had undergone the histological study. Although it was not possible to draw a conclusion that IM was significantly associated with the coinfection of either of the species and H. pylori, a tendency in that way would be likely, as it has also been reported by other authors.
PCR positives at the species level were also positive for the Helicobacter genus specific 16S rDNA and all the H. heilmannii and H. felis positive patients were also positive for H. pylori glmM PCR (Table ). PCR product sequences of ureaseB gene of H. heilmannii had shown 100% similarity to 'Candidatus H. heilmannii strains' GenBank: AF508012 and L25079; while H. felis sequences had shown 100% similarity to strains GenBank: FQ670179 and X69080.
In this study, we used urease gene-based PCR method developed by Nieger et al that detected only 'Candidatus
H. heilmannii' DNA from pure in vitro cultures of other non-H. pylori
helicobacter species [14
]. This method was also used by other investigators to demonstrate the presence of Candidatus
H. heilmannii DNA in gastric biopsies from patients with dyspepsia [11
]. The limitations of our study include the small number of patients who had non-H. pylori
helicobacter infection and the presence of H. pylori
co-infection which precluded assessment of the histological effect of these species under consideration. Also, the significance of coinfection in terms of disease development could not be determined. We could have identified few more cases of non-H. pylori
helicobacter species by other reported methods used to study non H. pylori
helicobacter species including fluorescent in situ hybridization (FISH), transmission electron microscopy (TEN) and partial 16S ribosomal sequencing for analyses of the amplified products [12
The implications of this study are that non-H. pylori
helicobacter species infection occurs in patients with abdominal pain or discomfort similar to H. pylori
infection. Most of our H. heilmannii
infections were not associated with contact with animals. This is in contrast to a previous analysis of 125 patients with confirmed H. heilmannii
infection that showed some 70.3% of the 111 patients had a history of contact with one or more animals [17
]. All of our patients with non-H. pylori
infection had endoscopic gastritis though their association with peptic ulcer is well known [19
]. The prevalence of coinfection of H. felis
with H. pylori
in our population is less than what has been reported from South Africa among African population but is certainly higher than that for H. heilmannii
and H. pylori
from the northern Europe which showed that only 1.6% had concomitant infection with H. pylori
]. The coinfection in our patients demonstrated severe gastric pathology, as intestinal metaplasia was present in 25% of H. heilmannii
coinfection with H. pylori
while in 20% of H. felis
coinfection with H. pylori
. This was also reported in previous studies [22
]. In this study, the difference was not statistically significant due to the number of subjects in each group. The routine transmission of H. pylori
appears to be human-human whereas non-H. pylori
helicobacter species are transmitted by cats, dogs, etc [22
]. Consequently, the prevalence of H. heilmannii
is expected to be significantly higher in environment with less hygiene and higher physical exposure to animals. However, in our study there was a negative association with pet contact as the patients reported limited exposure to these animals. There is a need to look into other modes of transmission of these infections.