*2.2.2 CagA protein and the cag pathogenicity island*

Cag pathogenicity island is a chromosomal region with about 37,000 bp and 29 genes. Four genes are similar to the components of the type IV secretion system. Proteins encoded by the island are involved in two major processes: the induction of interleukin-8 production by gastric epithelial cells and the translocation of CagA

#### **Figure 2.**

*The protein VacA. From Atherton et al [10].*

#### **Figure 3.** *The Cag pathogenicity Island. Modified by Suerbaum et al [11].*

*Virulence Markers, Genotypic versus Phenotypic Resistance and New Treatment Strategies... DOI: http://dx.doi.org/10.5772/intechopen.97026*

from the bacterium into the host cell by forming a syringe and a needle apparatus that delivers CagA protein (a protein of about 1200 amino-acids whose size varies between strains). Some genes are essential for the induction of interleukin-8; other genes are required for the translocation of CagA. The cagA-positive strain increases the risk of athrophic gastritis development and mucosal inflammation [16]. (see **Figure 3**).

## *2.2.3 Heat shock proteins (Hsps) or stress proteins*

Hsps are families of highly conserved proteins serving as a strong antigenic target for the immune response linked with pathology. *H. pylori* produces 2 Hsps: a gro Es-like HspsA (size 13 kd) and a gro E1-like HspsB (size 54–60 kd). Hsp60 is shared by *H.pylori* and eukariotic cells (autoimmune response) [17, 18]. The role of heatshock proteins in immune reactions is complex especially for the cellular effects of this proteins during the recognition processes by innate immunity. Heat-shock proteins (HSPs) are expressed at high levels by bacterial pathogens during adaptation to intracellular survival. The Hsps of both pathogens and hosts are involved in the activation of the receptors in innate immune response and in the presentation of antigens for the adaptive immune response [17, 18].

#### *2.2.4 Bab A adhesions*

The microorganism needs of adhesins for beginning its infective process. The presence of bacterial adhesins devoted to the attachment to human gastric epithelium, is essential [19]. *Helicobacter pylori* adherence to the human gastric mucosa involves specific bacterial adhesins and cognate host receptors. *babA* gene codes for the blood group antigen-binding adhesin BabA whereas the babB product is associated with a non-binding phenotype. BabA major adhesin is directed to the fucosylated Lewis b blood group antigen not present in all kinds of gastric cells [19].

#### *2.2.5 Urease virulence determinants*

The enzymatic activity of urease manages to break the urea molecule in bicarbonate ion and ammonia so that it is able to neutralize the gastric acid and is also correlated with the dual function of adhesivity and immunogenicity. The urease binds to the CD74 receptors of gastric epithelial cells [20]. It can be suggested that urease is specifically important for the attachment of *Hp* and for the pro-inflammatory immune response initiated by the bacterium. The binding of the subunit urease B to CD74 expressed on gastric cells may therefore contribute to increase the bacterial virulence during infection [20].

## **3. Molecular diagnostic of** *Helicobacter pylori*

*Helicobacter pylori* is a fastidious bacterium difficult to grow in the common media culture. Essential conditions for *Hp* culture were the following: microaerophilic atmosphere, temperature of 37° (range 33°-40°), presence of 0.5% glycine. The appropriate culture medium such as Pylori Selective Agar (bio-Merieux, Marcy L'Etoile France) contains 5% sheep blood and antibiotics (amphotericin, vancomycin and trimethoprim). The incubation lasts 10 days under CO2 atmosphere. This method is complicated, expensive and time-consuming so that when the isolation of the strain is unnecessary, the molecular method is very useful for its rapidity and appropriateness.

*Helicobacter pylori* presence in gastric specimens can be assessed by various methods including molecular PCR assay (GenoType HelicoDR kit) [21]. PCR is a more sensitive method (84.3% sensitivity, 75.0% specificity), with a higher *H. pylori* detection rate compared to culture [22].

Tissue obtained from gastroscopic biopsy was minced using a sterile scalpel, lysed by tissue lysis buffer and proteinase-K enzyme (Bioneer, Daejeon, Korea), and incubated for 10 min at 60°C. Ten total DNA was extracted with an AccuPrep Genomic DNA Extraction Kit (Bioneer, Daejeon, Korea). This kit contains a glass filter in a column tube that can bind efficiently to DNA in the presence of salts. Additional washing steps were performed for proteins and salt removal. Aliquots of 50 μL were used for PCR amplification as reported elsewhere [23].

Molecular methods such as PCR offer marginal improvements when done on biopsy material, but have the advantage of being able to accurately identify *H. pylori* in areas outside the stomach where cultures usually fail. PCR can detect low numbers of organisms in gastric juice, bile, stool and oral secretions. Because of its high sensitivity it can also be used for epidemiologic investigations of environmental sources [24]. Molecular methods have been used in variable specimens other than gastric mucosa [25].
