Culture method for isolation of Helicobacter pylori contamination of Endoscopic Units in Basra Hospitals

Background: In this study, the first attempt in Basrah city to isolate and diagnose H. pylori from endoscopic units and workers by culture method. Objective: to isolate and identify H. pylori by culture method and conventional tests from possibly contaminated sources and workers in endoscopy units in five hospitals. Materials and methods: A total of 78 swabs were collected endoscopic units in five hospitals in Basrah city in Iraq namely: Altaalemi, Al-mawani, Al-Basrah general, Al-Shifaa and Al-Fayhaa during the period from September 2011 to June 2012. Results: A total of 78 swabs, of which 12(15.38%) isolates of H. pylori were isolated by culture method and identified by biochemical tests. The highest frequency of H. pylori was in Altaalemi 7(29.16%) followed by AlShifaa 3(25%) and then Al-mawani 2(14.2%) and zero for the rest. The frequency of positive H. pylori in 13 samples from mouth of staff, was 3(23.1%). It was statistically significant (P<0.05) using SPSS program. Of the total 10 samples from forceps, 2(20%) showed positive tests for H. pylori, the frequency of positive H. pylori in 3 samples from sponge, was 2(66.7%), of the total 8 samples from flexible shaft, 2(25%) showed positive tests for H. pylori, 6 specimens from flexible cord transmitting light, air and water, 1(16.7%) showed positive tests for H. pylori, 5 specimens from Petridis for mouth pieces and 5 specimens from opening for biopsy channel, 1(20%) showed positive tests for H. pylori, Of the total 10, 6, 5, 5 and 2 samples from mouth, tables surface, beds, sinks and Pail sterilization respectively, the frequency of H. pylori was (0%). Conclusion: H. pylori can be isolated by culture method from endoscopic units and workers and disinfection technique for manual cleaning and disinfection of the instrument may not remove the risk of H. pylori transmission. مادختسا ةرصبلا ةظفاحم تايفشتسم يف روظانلا تادحول ةثولملا ةيدعملا ةباوبلا ةموثرج لزعل عرزلا ةقيرط م . د . اللهادبعلا اضرلادبع يناما ١ أ و . د . يدامح نيهاش دعس ٢ أ و . د . نيمأ يملسلا رابجلادبع ٣ ١ ةايحلا مولع مسق / مولعلا ةيلك / ةرصبلا ةعماج ، ٢ ينطابلا بطلا عرف / بطلا ةيلك / ةرصبلا ةعماج / ٣ ةايحلا مولع مسق / ةيبرتلا ةيلك / ةرصبلا ةعماج . ةمدقملا : ةموثرج صيخشتو لزعل ةرصبلا ةظفاحم يف ىلولأا ةلواحملا يه ةساردلا هذه دعت H. pylori انلا ةدحو نم ةدعملا ةحرقل ةببسملا ظ رو عرزلا ةقيرطب ةدحولاب نيلماعلاو . فدهلا : ةموثرج صيخشتو لزع H.pylori تادحو يف اهثولت لمتحملا رداصملاو نيلماعلا نم ةيديلقتلا تارابتخلااو عرزلا ةقيرط مادختساب تايفشتسم سمخل روظانلا . لمعلا ةقيرط : عمج مت ٧٨ ةرصبلا ةظفاحم يف تايفشتسم ةسمخ يف روظانلا تادحو نم ةحسم يلي امكو قارعلا : م . م و يميلعتلا . م و ئناوملا . م و ماعلا ةرصبلا . م و ءافشلا . لوليأ رهش نم ةرتفلا للاخ ءاحيفلا ٢٠١١ ناريزح ىلا ٢٠١٢ . جئاتنلا : نم ٧٨ لزع مت ةحسم ١٢ ) ١٥,٣٨ (% ةموثرج نم ةلزع H.pylori ةيويحوميكلا تارابتخلاا مادختساب تصخشو . ترهظأ نإ ةساردلا م يف تناك ةموثرجلل لزع ةبسن ىلعأ . يميلعتلا ٧ ) ٢٩,١٦ (% م اهتلت . ءافشلا ٣ ) ٢٥ (% م مث . ئناوملا ٢ ) ١٤,٢ (% يف لزعت مل نيح يف ةيقبتملا تايفشتسملا . ةموثرج ددرت صيخشتلاو لزعلا جئاتن ترهظأ H. pylori ةبسنب ٣ ) ٢٣,١ (% نم ١٣ روظانلا ةدحوب نيلماعلا نم ةنيع يونعم قرافبو (P<0.05) يئاصحلإا جمانربلا مادختساب SPSS . ةموثرج لزع متو H. pylori ةبسنب ٢ ) ٢٠ (% نم ١٠ طقلاملا نم تانيع و ٢ ) ٦٦,٧ (% نم ٣ نم تانيع أ و طقلاملا لسغ ةجنفس ٢ ) ٢٥ (% نم ٨ و نرملا دومعلا نم تانيع ١ ) ١٦,٧ % ( نم ٦ دومعلا نم تانيع MJBU, VOL. 31, No.2, 2013_____________________________________________________________________________________ 62 ةبسنبو ءاوهلاو ءاملل لقانلا نرملا ١ ) ٢٠ (% نم ٥ و مفلا حتف ةلآ فيظنت ءاعو نم تانيع ٥ طقلملا كلس لاخدإ ةحتف نم تانيع ، لزعت ملو ةموثرجلا ) رفص (% نم ١٠ و ٦ و ٥ و ٥ و لسغلا ضاوحأو ةرسلآاو دضانملا حوطس و مفلا حتف ةلآ نم ةذوخأم تانيع ءانإ لا يلاوتلا ىلع ميقعت . تاجاتنتسلاا : ةموثرج لزع H. pylori ليزت لا امبر زاهجلا ميقعتو فيظنتل ةيوديلا ةقيرطلا ناو اهيف نيلماعلاو روظانلا تادحو نم عرزلا ةقيرطب ةموثرج لاقتنا ةروطخ H. pylori .


INTRODUCTION
here are reports of a high prevalence of H. pylori infections in institutions for people with intellectual disability and health care workers in these institutions because of their close contact.
[1] H. pylori inhabits the gastric mucosa, consequently, the endoscopes used to perform gastroscopies on patients affected become contaminated.Infection can be passed on to other patients via these endoscopes.[4] The first recorded nosocomial infection with H. pylori was reported in 17 of 37 healthy subjects who took part in a study on acid concentrations in the stomach developed gastritis after an endoscopy. [5]Study of Nurnberg, et al., [5] concluded that endoscopes are frequently contaminated with H. pylori immediately after gastro duodenal endoscopy in H. pylori-infected patients.Although, appropriate decontamination can certainly prevent transmission via this route. [5,6]Iatrogenic transmission is possible, as H. pylori can survive manual cleaning and disinfection with 2% glutaraldehyde, particularly when there is ineffective cleaning before disinfection.De Schryver and Van Winckel, [7] make survey for a literature search was performed using Pubmed (January 1990 to May 2001), various pathways of agent transmission, favoring person-to-person mode of transmission early in life.Faeco-oral, oraloral and gastro-oral transmissions are proposed and may be of different relevance among various populations.Screening for colorectal cancer (CRC) by gastrointestinal endoscopy (flexible sigmoidoscopy or colonoscopy) has been recommended for the average-risk population. [8]The endoscopes have to be cleaned manually and/or by using automatic washing machines (best) especially designed for this purpose as the design and material of the endoscopes do not allow sterilization.Cleaning and disinfection of the devices has been a subject of concern, as transmission of infectious material cannot be entirely dismissed. [9]H. pylori can be identified by various methods such as culture, urease breath test, and histological studies of biopsy specimens; however most of these methods are invasive and non-invasive include Serologic methods and PCR.Aim: the objective of the present study was to isolate and identify H. pylori by culture method and conventional tests from possibly contaminated sources and workers in endoscopy units in five hospitals.

Sample collection
A total of 78 swabs were collected from endoscopic units in five hospitals in Basrah city south Iraq namely: Altaalemi, Al-mawani, Al-Basrah general, Al-shifaa and Al-Fayhaa during the period from September 2011 to June 2012.These samples include 10 from forceps, 3 from sponge for washing, 8 from flexible shaft, 6 from flexible cord transmitting light, air and water, 10 from mouth piece, 5 from petridish for mouth pieces, 5 from opening for biopsy channel, 13 from mouth of staff, 6 from tables surface, 5 from beds, 5 from sink and 2 from sterilization pail.All swabs were cultured by streaking on chocolate and Modified Columbia Urea Agar (MCUA) medium [10] and incubated in microaerophillic condition in anaerobic jar at 37 C o for 5-7 days.

T
The ways of disinfection in Al-Mawania, Al-Basrah and Al-Shifaa hospitals using medical gauze to clean forcipes once and then thrown it away except Al-sadr Altaalemi and Al-Fayhaa hospital using sponge instead of gauze .

Isolation and identification
The suspected purified colonies were chosen according to Gram staining and cultural characteristics and tested for the production of catalase, oxidase and urease.Statistical analysis: Data were analyzed using SPSS program for window (version 10).

RESULTS
Culture result: This is the first attempt in Basrah city to isolate and diagnose H. pylori from endoscopic units and workers by culture method.From a total of 78 swabs, of which 12(15.38%) isolates of H. pylori were isolated by culture method and identified by biochemical tests.From 24 samples from Altaalemi hospital, H.pylori was isolated in 7(29.16%),14 samples from Al-mawani hospital, H. pylori was isolated in 2(14.2%), 12 samples from Al-Shifaa hospital, H. pylori was isolated in 3(25%), 13 and 15 samples from Al-Basrah and Al-Fayhaa hospital respectively, H. pylori was not isolated (0%) (table-1).were on MCUA medium small to middle in size, rounded and creamy in color, H. pylori on chocolate agar were translucent, rounded and tiny size.All H. pylori isolates were Gram negative, spiral, rods, or curved in shape.The biochemical tests performed to confirm the identity of H.pylori includ: Oxidase, catalase and urease which were positive in all.Frequency of H. pylori according to the source of swab Of the total 10 samples from forcipes, 2(20%) showed positive tests for H. pylori, but the difference was statistically not significant (P>0.05).The frequency of positive H. pylori in 3 samples from sponge for washing, was 2(66.7%) but it was statistically not significant (P>0.05).Of the total 8 samples from flexible shaft, 2(25%) showed positive tests for H.
pylori, but the difference was statistically not significant (P>0.05).6 specimens from flexible cord transmitting light ,air and water, 1(16.7%)showed positive tests for H. pylori, but the difference was statistically not significant (P>0.05).5 specimens from Petridis for mouth pieces and 5 specimens from opening for biopsy channel, 1(20%) showed positive tests for H. pylori, but the difference was statistically not significant (P>0.05).The frequency of positive H. pylori in 13 samples from staff, was 3(23.1%).It was statistically significant (P<0.05).Of the total 10, 6, 5, 5 and 2 samples from mouth piece, table surface, beds, sink and Pail sterilization respectively, the frequency of H. pylori was (0%) and the results show no significant (P>0.05)(table 2).

DISCUSSION
Many studies have been performed about the contamination of endoscpic units by H. pylori, but no study has been done in Iraq specially in Basrah up to our knowledge, so the results of the present study will be compared with studies done in other countries.H. pylori is distributed worldwide and is found in developing countries in particular. [11]Although there is much information about H. pylori infection, several aspects of the pathogenesis and epidemiology of this organism remains unclear. [12]The transmission route of H. pylori infection has been the topic of several studies.Most infections are probably acquired in childhood, mainly via oral-oral or fecal-oral routes. [13]atients harboring H. pylori was reported by Al-Sulami, et al [14] and it was also isolated from drinking water by culture method only [15] and another study of Al-Sulami, et al [16] showed H. pylori has been isolated and diagnosed from drinking water by culture method and a combination of biochemical and PCR test.In present study, H. pylori was isolated by culture method from different sources in endoscopic units and workers.Contamination of endoscopes may be due to inappropriate procedure of cleaning hygiene and sterilization or individual failure to adhere to the procedures.The frequency of H. pylori isolates was different from one hospital to another, the high percentage of H. pylori was 7(29.16%) in Altaalemi, 3(25%) in Al-shifaa, 2(14.2%) in Almawani, but there were no isolates from Al-Basrah and Al-Fayhaa hospital respectively, because of the use of more hygienic way of cleaning of the scope after use by disposable gauze instead of the same dish and sponge for all patients.The frequency of positive H. pylori in 3 samples from sponge in Al-sadr Altaalemi and Al-Fayhaa hospital, was2(66.7%),reflecting the use of clean and forcipes sponge which represent an important source of pollution by H. pylori.Not taken samples of the sponge in Al-Mawania, Al-Basrah and Al-Shifaa hospitals because they are using medical gauze to clean forcipes once and then thrown it away.The frequency of H. pylori was 0(0%) from mouth pieces, tables surface, bed, sink and Pail sterilization and the results showed no significant (P>0.05).Staff was an important source for contamination, the frequency of positive H. pylori in 13 samples from mouth of staff in Altaalemi hospital, was 3(23.1%), a statistically significant difference (P<0.05).
Metanat, et al., [17] found the total frequency of H. pylori infection was 34% among healthcare workers in Zahedan, that is lower when compared to studies performed in other cities in Iran.Mastromarino, et al., [18] concluded that nurses had a significant higher prevalence of H. pylori infection (P<0.01).Of the total 10 samples from forceps, 2(20%) showed positive tests for H. pylori, the frequency of positive H. pylori in 3 samples from sponge for washing, was 2(66.7%), of the total 8 samples from flexible shaft, 2(25%) showed positive tests for H. pylori, 6 specimens from flexible cord transmitting light, air and water, 1(16.7%)showed positive tests for H. pylori, 5 specimens from Petridis of mouth and 5 specimens from opening for biopsy channel, 1(20%) showed positive tests for H. pylori.The frequency of positive H. pylori in 13 samples from mouth of staff, was 3(23.1%).Matysiak-Budnik [19] showed an association between occupational exposure and an increased risk of infection.
Williams, [20] too, stated that there were increased occupational risks for endoscopy personnel.However, the evidence in this review appears contradictory, since the findings varied between no risk and a five times greater risk.De Schryver et al., [21] were able to show in their reviews increased risks for gastroenterologists and endoscopy personnel.Magalhaes Queiroz [22] found controversial data on the occupational risk, but they considered only some of the studies also included here for gastroenterological personnel.Hildebrand, et al., [23] concluded that gastroenterologists have a high risk of becoming infected with Helicobacter pylori, claiming the potential role of saliva or gastric juice droplets during endoscopy.Endoscopes are often cleaned and disinfected only manually in between the collection of gastric tissue specimens, [24] on the other hand, gastric biopsy specimens may have been contaminated with H. pylori DNA remaining behind in the biopsy channels of the fiberoptic endoscopes.Indeed, it has been reported that patients can be infected with H. pylori through gastrointestinal equipment which is not properly disinfected [25] H. pylori DNA has been detected by PCR in fluid flushed from endoscopes channels after cleaning and disinfection with 2% glutaraldehyede. [26]urnberg, et al, [5] showed one of the 128 rinsing samples (0.8%) was found to be contaminated with H. pylori even after routine manual cleaning and disinfection-indicating that these cleaning and disinfection procedures may be insufficient to eradicate H.pylori from endoscopes completely.

Conclusions
1. H. pylori can be isolated by culture method from endoscopic units and workers .

Gastroenterological personnel are exposed
to an increased risk of H. pylori infection.

Disinfection technique for manual
cleaning and disinfection of the instrument may not remove the risk of H. pylori transmission and may be other bacteria and viruses.Recommendations -Use of the international way of cleaning of endoscopic and its accessories by washing machine after cleaning with medical gauze instead of sponge for washing forceps.

Table 1 . Number and percentage of H. pylori isolates from twelve different sources in endoscopic units in five hospitals. Hp+: H. pylori positive Al-Basrah general and Al-Shifaa hospitals not using sponge for cleaning.
The highest frequency of H. pylori was in Altaalemi 7(29.16%)followed by Al-Shifaa3(25%) and then Al-Mawani 2(14.2%) and zero for the rest.The colonies of H. pylori

-
Using mask and glasses to protect against H. pylori infection for this and other viral infection of medical personnel and prevent it transmission to the patients.