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PCR-based genotyping of Helicobacter pylori of Gambian children and adults directly from biopsy specimens and bacterial cultures.

Secka O, Antonio M, Tapgun M, Berg DE, Bottomley C, Thomas V, Walton R, Corrah T, Adegbola RA, Thomas JE - Gut Pathog (2011)

Bottom Line: Virulence genes were amplified in 127 of 190 cases tested (121 adults and 6 children); each of 60 bacterial cultures, and 116 from DNA extracted directly from biopsies.In order to detect the range of bacterial genotypes harbored by individual patients, direct PCR proved slightly superior to isolation of H. pylori by biopsy culture, but the techniques were complementary, and the combination of both culture and direct PCR produced the most complete picture.The seemingly higher virulence of strains from adult than infant infections in The Gambia merits further analysis.

View Article: PubMed Central - HTML - PubMed

Affiliation: Bacterial Diseases Programme, Medical Research Council Laboratories, The Gambia. osecka@mrc.gm.

ABSTRACT

Background: Helicobacter pylori is an important agent of gastroduodenal disease in Africa and throughout the world. We sought to determine an optimum method for genotyping H. pylori strains from children and adults in The Gambia, West Africa.

Results: Virulence genes were amplified in 127 of 190 cases tested (121 adults and 6 children); each of 60 bacterial cultures, and 116 from DNA extracted directly from biopsies. The proportion of biopsies that were cagA+, the ratio of vacAs1/s2, and vacAm1/m2, and the proportion of mixed strain populations in individual subjects changed with age. Strains lacking virulence cagA and vacA genes and with apparently homogeneous (one predominant strain) infections were more common among infants than adults.

Conclusions: In order to detect the range of bacterial genotypes harbored by individual patients, direct PCR proved slightly superior to isolation of H. pylori by biopsy culture, but the techniques were complementary, and the combination of both culture and direct PCR produced the most complete picture. The seemingly higher virulence of strains from adult than infant infections in The Gambia merits further analysis.

No MeSH data available.


Related in: MedlinePlus

PCR inferred results of signal region of vacA gene. 1.5% gel electrophoresis of H. pylori genotypes showing PCR results of s1 and s2 allelles of vacA gene. Lane M is a 100 bp ladder (Biolabs, UK), lane 1 and 2 showed the presence of s1 (259 bp), lane 3 is both s1 and s2 positive and lane 4 and 5 are s2 (289 bp) positive.
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Figure 3: PCR inferred results of signal region of vacA gene. 1.5% gel electrophoresis of H. pylori genotypes showing PCR results of s1 and s2 allelles of vacA gene. Lane M is a 100 bp ladder (Biolabs, UK), lane 1 and 2 showed the presence of s1 (259 bp), lane 3 is both s1 and s2 positive and lane 4 and 5 are s2 (289 bp) positive.

Mentions: Virulence gene amplification was successful in 127 (121 adults and 6 children) cases. A comparison of the products that were indicative of cagA (Figure 1), cag emptysite (Figure 2), vacAs alleles (Figure 3), vacAm alleles (Figure 4), iceA1 (Figure 5) and iceA2 (Figure 6) between both methods for detecting H. pylori is summarized for 60 samples for which sufficient amplified DNA was obtained for this further analysis. The proportion of samples that were cagA+ve with DNA from biopsies and from culture was similar, 58.3% and 61.7% respectively. The success in amplification of vacAs1/s2 (s1 = toxigenic vs s2 = non-toxigenic), vacAm1/m2 and iceA1 alleles was similar from cultures and corresponding biopsies, and agreements between genotypes inferred using DNAs directly from these two sources was good for both cagA and m1, m2 alleles of vacA, moderate for s1, s2 alleles of vacA, and poor for iceA. The poor agreement in the iceA analysis stemmed from the many classified only as iceA2 by PCR from bacterial culture but iceA1 and iceA2 by biopsy which could have been due to the fact that certain bacterial strains in a mixed infection grew much better than others in culture. In direct PCR up to 16.7% of culture positive biopsies failed to amplify DNA for individual alleles.


PCR-based genotyping of Helicobacter pylori of Gambian children and adults directly from biopsy specimens and bacterial cultures.

Secka O, Antonio M, Tapgun M, Berg DE, Bottomley C, Thomas V, Walton R, Corrah T, Adegbola RA, Thomas JE - Gut Pathog (2011)

PCR inferred results of signal region of vacA gene. 1.5% gel electrophoresis of H. pylori genotypes showing PCR results of s1 and s2 allelles of vacA gene. Lane M is a 100 bp ladder (Biolabs, UK), lane 1 and 2 showed the presence of s1 (259 bp), lane 3 is both s1 and s2 positive and lane 4 and 5 are s2 (289 bp) positive.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3107793&req=5

Figure 3: PCR inferred results of signal region of vacA gene. 1.5% gel electrophoresis of H. pylori genotypes showing PCR results of s1 and s2 allelles of vacA gene. Lane M is a 100 bp ladder (Biolabs, UK), lane 1 and 2 showed the presence of s1 (259 bp), lane 3 is both s1 and s2 positive and lane 4 and 5 are s2 (289 bp) positive.
Mentions: Virulence gene amplification was successful in 127 (121 adults and 6 children) cases. A comparison of the products that were indicative of cagA (Figure 1), cag emptysite (Figure 2), vacAs alleles (Figure 3), vacAm alleles (Figure 4), iceA1 (Figure 5) and iceA2 (Figure 6) between both methods for detecting H. pylori is summarized for 60 samples for which sufficient amplified DNA was obtained for this further analysis. The proportion of samples that were cagA+ve with DNA from biopsies and from culture was similar, 58.3% and 61.7% respectively. The success in amplification of vacAs1/s2 (s1 = toxigenic vs s2 = non-toxigenic), vacAm1/m2 and iceA1 alleles was similar from cultures and corresponding biopsies, and agreements between genotypes inferred using DNAs directly from these two sources was good for both cagA and m1, m2 alleles of vacA, moderate for s1, s2 alleles of vacA, and poor for iceA. The poor agreement in the iceA analysis stemmed from the many classified only as iceA2 by PCR from bacterial culture but iceA1 and iceA2 by biopsy which could have been due to the fact that certain bacterial strains in a mixed infection grew much better than others in culture. In direct PCR up to 16.7% of culture positive biopsies failed to amplify DNA for individual alleles.

Bottom Line: Virulence genes were amplified in 127 of 190 cases tested (121 adults and 6 children); each of 60 bacterial cultures, and 116 from DNA extracted directly from biopsies.In order to detect the range of bacterial genotypes harbored by individual patients, direct PCR proved slightly superior to isolation of H. pylori by biopsy culture, but the techniques were complementary, and the combination of both culture and direct PCR produced the most complete picture.The seemingly higher virulence of strains from adult than infant infections in The Gambia merits further analysis.

View Article: PubMed Central - HTML - PubMed

Affiliation: Bacterial Diseases Programme, Medical Research Council Laboratories, The Gambia. osecka@mrc.gm.

ABSTRACT

Background: Helicobacter pylori is an important agent of gastroduodenal disease in Africa and throughout the world. We sought to determine an optimum method for genotyping H. pylori strains from children and adults in The Gambia, West Africa.

Results: Virulence genes were amplified in 127 of 190 cases tested (121 adults and 6 children); each of 60 bacterial cultures, and 116 from DNA extracted directly from biopsies. The proportion of biopsies that were cagA+, the ratio of vacAs1/s2, and vacAm1/m2, and the proportion of mixed strain populations in individual subjects changed with age. Strains lacking virulence cagA and vacA genes and with apparently homogeneous (one predominant strain) infections were more common among infants than adults.

Conclusions: In order to detect the range of bacterial genotypes harbored by individual patients, direct PCR proved slightly superior to isolation of H. pylori by biopsy culture, but the techniques were complementary, and the combination of both culture and direct PCR produced the most complete picture. The seemingly higher virulence of strains from adult than infant infections in The Gambia merits further analysis.

No MeSH data available.


Related in: MedlinePlus