Multiplex polymerase chain reaction assays for detecting a bacteria (e.g. Helicobacter pylori) in a specimen use multiple oligonucleotide primer pairs based on the sequences of multiple loci of the bacteria. In one application, up to five loci in the genomic DNA sequences of Helicobacter pylori were amplified. Two fragments of H. pylori were amplified from each locus, wherein a second fragment was an internal fragment of the first fragment.