**4. Discussions**

The enthusiasm beginning with the isolation of *H. pylori* from gastric biopsies by Warren and Marshall in 1982 has increasingly continued after the important role of this agent in the aetiology of gastric cancer has been established; consequently, the interest for *H. pylori* has increased. The association of gastric cancer, one of the most frequent causes of death worldwide, with a treatable aetiological factor has led to a profound impact on researchers [7, 8].

in the stomach. The antrum has been found to be the most affected part of the stomach by PPIs as *H. pylori* almost disappears from this niche. To avoid false-negative results, it is recom-

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87

In our study, exclusion criteria were of inhibitory proton pump or H2-receptor antagonists and antibiotics 2 weeks before the beginning of the study, similarly with another study and

The recent Maastricht V/Florence Consensus Report recommends discontinuation of antibiot-

The number of biopsies necessary to diagnose *H. pylori* infection is a subject of controversy. A single biopsy specimen taken from the antrum (2 cm from the pylorus) gives good sensitivity, but it is not sufficient for a reliable diagnosis. Indeed, *H. pylori* may have a patch distribution, and the more biopsy specimens analysed, the higher the chance of detecting the organism. There are some rare cases where the infection lies only in the corpus, but usually, *H. pylori* is present in all sites. After consumption of antisecretory drugs, as pointed out before, the

We took two biopsies from the antrum for *H. pylori* culture, and maybe this could be adjusted by obtaining another one/two samples from the gastric body as well in order to try to improve

The usual recommendation derived from the Sydney system is to obtain two biopsy specimens from the antrum and two specimens from the corpus. Bacteria are usually present at both sites even if the lesions occur essentially in the antrum. When topographic studies of *H. pylori* distribution and gastritis were performed, the best site suitable from diagnosis was the lesser curvature of the midantrum, while for the corpus, there was a discrepancy between greater and lesser curvatures [15, 16]. Others showed that two antral biopsies only were suf-

We took the biopsy specimens for culture before specimens for histology, and we used an

We analysed the correlation between densities of *H. pylori* in histological exam and positive *H. pylori* culture: in 14.28% the *H. pylori* culture was positive in mild *H. pylori* density in histology, 18.18% in moderate *H. pylori* density and 33.33% in marked *H. pylori* score. We have not had a significant statistics correlation between densities of *H. pylori* in histological exam and positive *H. pylori* culture (p = 0.7). Similar results were reported by other authors [8].

We analysed the correlation between gastritis activity and density of *H. pylori* in histological

The results of this study are in agreement with published work, suggesting that a strain of the organism may be a more important factor than the density of infection in determining the

In our study, we had four children with bleeding, and all of them had negative *H. pylori*

appropriate commercially transport medium, according to the recommendations.

exam, and we have not had a significant statistics correlation (p = 0.30).

gastric inflammatory response to *H. pylori* [18].

culture.

ics 4 weeks before the study to allow an increase of detectable bacterial load [2].

corpus may be the only site that remains positive [15].

the culture success rate, but this option is difficult to apply in children.

mended not to consume these drugs 2 weeks prior to endoscopy [14, 15].

recommendations [15].

ficient to detect *H. pylori* [17].

*H. pylori* infection is generally acquired in childhood, and it persists throughout life. Spontaneous resolution is rare, and a targeted therapy is needed [9].

The correct diagnosis and effective treatment of *H. pylori* gastric infection are essential. The recent guidelines for the management of *H. pylori* in children and adolescents recommend the initial diagnosis of *H. pylori* infection to be performed using invasive gastric biopsy methods including the following: obtaining a positive bacterial culture or demonstrating *H. pylori* gastritis on histopathology; using the updated Sydney classification, with at least one other positive test such as RUT or molecular-based assays where available; and including polymerase chain reaction or fluorescent in situ hybridisation. The initial diagnosis of *H. pylori* infection should not be based on noninvasive tests (i.e. 13C-UBT and *H. pylori* stool antigen test) or other noninvasive methods. A positive noninvasive test, however, supports the diagnosis in cases in which positive histology is the only available invasive test [4].

76.31% of patients enrolled in the study were positive for *H. pylori* infection. The diagnosis was made in 96.55% by histology. In 17.25% of cases, both histology and bacterial culture were positive.

The *H. pylori* culture was positive in only six cases (21.42%). Our results are less significant if compared to other published data in which higher percentages of positive cultures were obtained, except one. Kaya et al. reported the sensitivity for *H. pylori* culture as 22.5%, and the specificity as 97.1% [10].

The specificity for *H. pylori* culture, in our study, was 90.90%, but the sensitivity was low, 20.68%.

A recent Israeli study, conducted in the paediatric population, reported 57.8% of positive *H. pylori* culture in 154 children with positive RUT [3]. In another study conducted on children and adolescents, the sensitivity for *H. pylori* culture was 79.3%, and the specificity was 100% [11].

The sensitivity of culture method to detect *H. pylori*, in adult population, ranges from 62.7% to 96.3% in the studies performed [8, 10, 12].

Although the culture method is accepted as a "gold standard" for the diagnosis, it is difficult to use alone as a routine diagnostic method. As the sensitivity of culture method is low, *H. pylori* positivity can be detected in case of growth. The absence of growth does not indicate *H. pylori* negativity.

Despite its long use, culture remains a challenge because of the fastidious nature of the bacterium, with particular growth requirements regarding environment and atmosphere [13]. Altering pH, the proton pump inhibitors (PPIs) indirectly interfere with *H. pylori* distribution in the stomach. The antrum has been found to be the most affected part of the stomach by PPIs as *H. pylori* almost disappears from this niche. To avoid false-negative results, it is recommended not to consume these drugs 2 weeks prior to endoscopy [14, 15].

**4. Discussions**

86 Histology

were positive.

20.68%.

specificity as 97.1% [10].

*H. pylori* negativity.

to 96.3% in the studies performed [8, 10, 12].

The enthusiasm beginning with the isolation of *H. pylori* from gastric biopsies by Warren and Marshall in 1982 has increasingly continued after the important role of this agent in the aetiology of gastric cancer has been established; consequently, the interest for *H. pylori* has increased. The association of gastric cancer, one of the most frequent causes of death worldwide, with a treatable aetiological factor has led to a profound impact on researchers [7, 8]. *H. pylori* infection is generally acquired in childhood, and it persists throughout life.

The correct diagnosis and effective treatment of *H. pylori* gastric infection are essential. The recent guidelines for the management of *H. pylori* in children and adolescents recommend the initial diagnosis of *H. pylori* infection to be performed using invasive gastric biopsy methods including the following: obtaining a positive bacterial culture or demonstrating *H. pylori* gastritis on histopathology; using the updated Sydney classification, with at least one other positive test such as RUT or molecular-based assays where available; and including polymerase chain reaction or fluorescent in situ hybridisation. The initial diagnosis of *H. pylori* infection should not be based on noninvasive tests (i.e. 13C-UBT and *H. pylori* stool antigen test) or other noninvasive methods. A positive noninvasive test, however, supports the diagnosis in cases

76.31% of patients enrolled in the study were positive for *H. pylori* infection. The diagnosis was made in 96.55% by histology. In 17.25% of cases, both histology and bacterial culture

The *H. pylori* culture was positive in only six cases (21.42%). Our results are less significant if compared to other published data in which higher percentages of positive cultures were obtained, except one. Kaya et al. reported the sensitivity for *H. pylori* culture as 22.5%, and the

The specificity for *H. pylori* culture, in our study, was 90.90%, but the sensitivity was low,

A recent Israeli study, conducted in the paediatric population, reported 57.8% of positive *H. pylori* culture in 154 children with positive RUT [3]. In another study conducted on children and adolescents, the sensitivity for *H. pylori* culture was 79.3%, and the specificity was 100% [11].

The sensitivity of culture method to detect *H. pylori*, in adult population, ranges from 62.7%

Although the culture method is accepted as a "gold standard" for the diagnosis, it is difficult to use alone as a routine diagnostic method. As the sensitivity of culture method is low, *H. pylori* positivity can be detected in case of growth. The absence of growth does not indicate

Despite its long use, culture remains a challenge because of the fastidious nature of the bacterium, with particular growth requirements regarding environment and atmosphere [13]. Altering pH, the proton pump inhibitors (PPIs) indirectly interfere with *H. pylori* distribution

Spontaneous resolution is rare, and a targeted therapy is needed [9].

in which positive histology is the only available invasive test [4].

In our study, exclusion criteria were of inhibitory proton pump or H2-receptor antagonists and antibiotics 2 weeks before the beginning of the study, similarly with another study and recommendations [15].

The recent Maastricht V/Florence Consensus Report recommends discontinuation of antibiotics 4 weeks before the study to allow an increase of detectable bacterial load [2].

The number of biopsies necessary to diagnose *H. pylori* infection is a subject of controversy. A single biopsy specimen taken from the antrum (2 cm from the pylorus) gives good sensitivity, but it is not sufficient for a reliable diagnosis. Indeed, *H. pylori* may have a patch distribution, and the more biopsy specimens analysed, the higher the chance of detecting the organism. There are some rare cases where the infection lies only in the corpus, but usually, *H. pylori* is present in all sites. After consumption of antisecretory drugs, as pointed out before, the corpus may be the only site that remains positive [15].

We took two biopsies from the antrum for *H. pylori* culture, and maybe this could be adjusted by obtaining another one/two samples from the gastric body as well in order to try to improve the culture success rate, but this option is difficult to apply in children.

The usual recommendation derived from the Sydney system is to obtain two biopsy specimens from the antrum and two specimens from the corpus. Bacteria are usually present at both sites even if the lesions occur essentially in the antrum. When topographic studies of *H. pylori* distribution and gastritis were performed, the best site suitable from diagnosis was the lesser curvature of the midantrum, while for the corpus, there was a discrepancy between greater and lesser curvatures [15, 16]. Others showed that two antral biopsies only were sufficient to detect *H. pylori* [17].

We took the biopsy specimens for culture before specimens for histology, and we used an appropriate commercially transport medium, according to the recommendations.

We analysed the correlation between densities of *H. pylori* in histological exam and positive *H. pylori* culture: in 14.28% the *H. pylori* culture was positive in mild *H. pylori* density in histology, 18.18% in moderate *H. pylori* density and 33.33% in marked *H. pylori* score. We have not had a significant statistics correlation between densities of *H. pylori* in histological exam and positive *H. pylori* culture (p = 0.7). Similar results were reported by other authors [8].

We analysed the correlation between gastritis activity and density of *H. pylori* in histological exam, and we have not had a significant statistics correlation (p = 0.30).

The results of this study are in agreement with published work, suggesting that a strain of the organism may be a more important factor than the density of infection in determining the gastric inflammatory response to *H. pylori* [18].

In our study, we had four children with bleeding, and all of them had negative *H. pylori* culture.

Peptic ulcer bleeding and atrophic gastritis decreased the accuracy of *H. pylori* diagnostic test. The histology was found to be a reliable test in the presence of bleeding [19].

as histology (90.90%), while its sensitivity was 20.68%. We did not find out a statistically significant positive correlation between *H. pylori* density observed at the histological exam and positive bacterial culture. This result may have been influenced by the limited number of patients and by the few cases with positive bacterial culture. Larger studies are needed in

A Study of the Correlation between Bacterial Culture and Histological Examination in Children…

http://dx.doi.org/10.5772/intechopen.80257

89

The authors thank Dr. Violeta Cristea and Dr. Augustina Enculescu for their laboratorial and

, Cătălin Boboc<sup>2</sup>

1 University of Medicine and Pharmacy Carol Davila, Bucharest, Romania

2 Maria Sklodowska Curie Children's Emergency Hospital, Bucharest, Romania

[1] Sanders CJ, Yu Y, Moore DA, Williams IR, Gewirtz AT. Humoral immune response to flagellin requires T cells and activation of innate immunity. Journal of Immunology.

[2] Malfertheiner P, Megraud F, O'Morain C, Gisbert JP, Kuipers EJ, Axon AT, Bazzoli F, Gasbarrini A, Atherton J, Graham DY, Hunt R, Moayyedi P, Rokkas T, Rugge M, Selgrad M, Suerbaum S, Sugano K, El-Omar EM. Management of *Helicobacter pylori* infection—The Maastricht V/Florence consensus report. Gut. 2017;**66**:6-30. DOI: 10.1136/

[3] Pastukh N, Peretz A, Brodsky D, Isakovich N, Azrad M, On A. Antimicrobial susceptibility of *Helicobacter pylori* strains isolated from children in Israel. Journal of Global

Antimicrobial Resistance. 2018;**12**:175-178. DOI: 10.1016/j.jgar.2017.10.004

, Cristina Coldea1,2, Mălina Anghel<sup>2</sup>

,

order to obtain relevant conclusions.

histological support, respectively.

, Gabriela Năstase2†

† These two authors contributed equally to this work.

2006;**177**:2810-2818. DOI: 10.4049/jimmunol.177.5.2810

\*Address all correspondence to: felicia\_galos@yahoo.com

Anca Orzan1,2 and Mihaela Bălgrădean1,2

**Acknowledgements**

**Conflict of interest**

None declared.

**Author details**

Felicia Galoș1,2\*†

**References**

gutjnl-2016-312288

When atrophic changes occur in the gastric mucosa, a high percentage of endoscopic biopsy samples become negative at bacterial histology [20, 21]. During atrophy progression the density of *H. pylori* in the stomach mucosa decreases and may disappear completely during the late stages of atrophy [22]. This may explain the lower sensitivity of biopsy-based tests in the presence of atrophy: RUT, histology and culture. UBT and antigen stool detection can also give false-negative results in this situation. Serology is the only diagnostic method not influenced by a lower density of microorganism, being reliable even in advanced gastric body atrophy. Maastricht guidelines updates have reserved serology for special situations, including extensive atrophy of the stomach mucosa, in conditions in which the other tests may be negative because of low bacterial density.

In childhood, advanced gastric atrophy is rare. We found only one case with atrophy, but the *H. pylori* culture was positive. It was an adolescent, 17 years and 8 months old, with a long history of illness (2 years) and with previous treatment failure. We could not find, in this case, antibiotic resistance to amoxicillin, clarithromycin, metronidazole and levofloxacin, and we suppose that the noncompliance to therapy is the cause of failure for bacterial eradication. Successful eradication is important to prevent the development of antibiotic resistance, as well as to reduce the number of treatments and procedures. Among children receiving the triple standard therapy regimen, eradication rate is declining [23]. In part, this decrease can be attributed to increased antibiotic resistance. Other reasons for treatment failure are, among others, host genetic factors, *H. pylori* virulent factors, inadequate compliance to therapy or insufficient duration of therapy and, not in the last row, smoking and household crowding [23].

The sensitivity of histology in our study was 96.55%, while the specificity was 90.90%.

The sensibility and specificity of haematoxylin and eosin stain have been reported as 69–93% and 87–90%, respectively. The specificity can be improved to 90–100% by using special stains such as Giemsa stain, Warthin-Starry silver stain, Genta stain and immunohistochemical stain [15, 20, 21]. Immunohistochemical stain has a particular advantage in patients partially treated for *H. pylori* gastritis, a setting can result in atypical (including coccoid) forms, which may mimic bacteria or cell debris on haematoxylin and eosin preparation. The major advantages of immunohistochemical stain include shorter screening time and high specificity because it can exclude other similar-shaped organisms [15, 21].

In our study, we analysed the sensibility and specificity of histology in *H. pylori* infection, not focusing on the ones specific for the different types of stain. We used in all cases haematoxylin-eosin stain and in a few cases Giemsa stain.

## **5. Conclusions**

Histology is an excellent method for detecting *H. pylori* infection. Haematoxylin-eosin, with or without Giemsa stains, is usually adequate. It is difficult to use culture method alone as routine diagnostic method. In our study, the specificity of histology in identifying *H. pylori* infection was 90.90%, and its sensitivity was 96.55%. Bacterial culture had the same specificity as histology (90.90%), while its sensitivity was 20.68%. We did not find out a statistically significant positive correlation between *H. pylori* density observed at the histological exam and positive bacterial culture. This result may have been influenced by the limited number of patients and by the few cases with positive bacterial culture. Larger studies are needed in order to obtain relevant conclusions.
