Nucleotide sequences which hybridize specifically with a Campylobacter jejuni genomic nucleic sequence

A purified nucleotide sequence consisting of 12-20 contiguous nucleotides of a nucleic acid sequence selected from the group consisting of:

• a nucleotide sequence of SEQ ID NO:1;
• a nucleotide sequence of SEQ ID NO:2;
• a nucleotide sequence fully complementary to SEQ ID NO:1;
• a nucleotide sequence fully complementary to SEQ ID NO:2;
• a nucleotide sequence which hybridizes with SEQ ID NO:1 under stringent conditions;
• a nucleotide sequence which hybridizes with a nucleotide sequence fully complementary to SEQ ID NO:1 under stringent conditions;
• a nucleotide sequence which hybridizes with SEQ ID NO:2 under stringent conditions;
• or
• a nucleotide sequence which hybridizes with a nucleotide sequence fully complementary to SEQ ID NO:2 under stringent conditions;
• wherein stringent conditions are
• hybridizing for 16-18 hours at 65.degree. C. in 6X SSC buffer containing 10% dextran sulfate, a 5X concentrated Denhardt's solution 10 mM EDTA, 0.05% SDS, 100 .mu.g/ml denatured salmon sperm DNA;
• washing twice for 10 minutes at 65.degree. C. in 2X SSC, once for 30 minutes at 65.degree. C. in 2X SSC+0.1% SDS and once for 10 minutes at 65.degree. C. in 0.1X SSC;
• and
• detecting hybridization to C. jejuni but not to other Campylobacter species.

A cloning vector comprising a nucleotide sequence selected from the group consisting of:

• a nucleotide sequence of SEQ ID NO:1;
• a nucleotide sequence of SEQ ID NO:2;
• a nucleotide sequence fully complementary to SEQ ID NO:1;
• a nucleotide sequence fully complementary to SEQ ID NO:2;
• a nucleotide sequence which hybridizes with SEQ ID NO:1 under stringent conditions,
• a nucleotide sequence which hybridizes with a nucleotide sequence fully complementary to SEQ ID NO:1 under stringent conditions;
• a nucleotide sequence which hybridizes with SEQ ID NO:2 under stringent conditions;
• or
• a nucleotide sequence which hybridizes with a nucleotide sequence fully complementary to SEQ ID NO:2 under stringent conditions;
• wherein stringent conditions are
• hybridizing for 16-18 hours at 65.degree. C. in 6X SSC buffer containing 10% dextran sulfate, a 5X concentrated Denhardt's solution 10 mM EDTA, 0.05% SDS, 100 .mu.g/ml denatured salmon sperm DNA;
• washing twice for 10 minutes at 65.degree. C. in 2X SSC, once for 30 minutes at 65.degree. C. in 2X SSC+0.1% SDS and once for 10 minutes at 65.degree. C. in 0.1X SSC;
• and
• detecting hybridization to C. jejuni but not to other Campylobacter species.

A probe specific for a genomic nucleotide sequence of Campylobacter jejuni, comprising at least 12 contiguous nucleotides of a nucleic acid sequence selected from the group consisting of:

• a nucleotide sequence of SEQ ID NO:1;
• a nucleotide sequence of SEQ ID NO:2;
• a nucleotide sequence fully complementary to SEQ ID NO:1;
• a nucleotide sequence fully complementary to SEQ ID NO:2;
• a nucleotide sequence which hybridizes with SEQ ID NO:1 under stringent conditions;
• a nucleotide sequence which hybridizes with a nucleotide sequence fully complementary to SEQ ID NO:1 under stringent conditions;
• a nucleotide sequence which hybridizes with SEQ ID NO:2 under stringent conditions;
• or
• a nucleotide sequence which hybridizes with a nucleotide sequence fully complementary to SEQ ID NO:2 under stringent conditions;
• wherein stringent conditions are
• hybridizing for 16-18 hours at 65.degree. C. in 6X SSC buffer containing 10% dextran sulfate, a 5X concentrated Denhardt's solution 10 mM EDTA, 0.05% SDS, 100 .mu.g/ml denatured salmon sperm DNA;
• washing twice for 10 minutes at 65.degree. C. in 2X SSC, once for 30 minutes at 65.degree. C. in 2X SSC+0.1% SDS and once for 10 minutes at 65.degree. C. in 0.1X SSC;
• and
• detecting hybridization to C. jejuni but not to other Campylobacter species.

A pair of oligonucleotide primers specific for the amplification of Campylobacter jejuni, wherein each primer consists of 18 to 30 contiguous nucleotides of a nucleic acid sequence selected from the group consisting of:

• a nucleotide sequence of SEQ ID NO:1;
• a nucleotide sequence of SEQ ID NO:2;
• a nucleotide sequence fully complementary to SEQ ID NO:1;
• a nucleotide sequence fully complementary to SEQ ID NO:2;
• a nucleotide sequence which hybridizes with SEQ ID NO:1 under stringent conditions;
• a nucleotide sequence which hybridizes with a nucleotide sequence fully complementary to SEQ ID NO:1 under stringent conditions;
• a nucleotide sequence which hybridizes with SEQ ID NO:2 under stringent conditions;
• or
• a nucleotide sequence which hybridizes with a nucleotide sequence fully complementary to SEQ ID NO:2 under stringent conditions;
• wherein stringent conditions are
• hybridizing for 16-18 hours at 65.degree. C. in 6X SSC buffer containing 10% dextran sulfate, a 5X concentrated Denhardt's solution 10 mM EDTA, 0.05% SDS, 100 .mu.g/ml denatured salmon sperm DNA;
• washing twice for 10 minutes at 65.degree. C. in 2X SSC, once for 30 minutes at 65.degree. C. in 2X SSC+0.1% SDS and once for 10 minutes at 65.degree. C. in 0.1X SSC;
• and
• detecting hybridization to C. jejuni but not to other Campylobacter species.

A method of detecting the presence of Campylobacter jejuni in a biological sample, comprising the steps of:

• (a) contacting said biological sample with a pair of primers, wherein each primer consists of 18 to 30 contiguous nucleotides of a nucleic acid sequence selected from the group consisting of:
• a nucleotide sequence of SEQ ID NO:1;
• a nucleotide sequence of SEQ ID NO:2;
• a nucleotide sequence fully complementary to SEQ ID NO:1;
• a nucleotide sequence fully complementary to SEQ ID NO:2;
• a nucleotide sequence which hybridizes with SEQ ID NO:1 under stringent conditions within a temperature range of (Tm--15.degree. C.) to (Tm--20.degree. C.);
• a nucleotide sequence which hybridizes with a nucleotide sequence fully complementary to SEQ ID NO:1 under stringent conditions;
• a nucleotide sequence which hybridizes with SEQ ID NO:2 under stringent conditions;
• or
• a nucleotide sequence which hybridizes with a nucleotide sequence fully complementary to SEQ ID NO:2 under stringent conditions;
• wherein stringent conditions are
• hybridizing for 16-18 hours at 65.degree. C. in 6X SSC buffer containing 10% dextran sulfate, a 5X concentrated Denhardt's solution 10 mM EDTA, 0.05% SDS, 100 .mu.g/ml denatured salmon sperm DNA;
• washing twice for 10 minutes at 65.degree. C. in 2X SSC, once for 30 minutes at 65.degree. C. in 2X SSC+0.1% SDS and once for 10 minutes at 65.degree. C. in 0.1X SSC;
• and
• detecting hybridization to C. jejuni but not to other Campylobacter species;
• (b) amplifying any genomic nucleic acid sequences of Campylobacter jejuni in said sample;
• and
• (c) detecting the presence of any amplified genomic nucleic acid sequences.

A method of detecting the presence of Campylobacter jejuni in a food sample, comprising the steps of:

• (a) low speed centrifuging said food sample to remove coarse organic food debris,
• (b) contacting the food sample with a pair of primers, wherein each primer consists of 18 to 30 consecutive nucleotides of a nucleic acid sequence selected from the group consisting of:
• a nucleotide sequence of SEQ ID NO:1;
• a nucleotide sequence of SEQ ID NO:2;
• a nucleotide sequence fully complementary to SEQ ID NO:1;
• a nucleotide sequence fully complementary to SEQ ID NO:2;
• a nucleotide sequence which hybridizes with SEQ ID NO:1 under stringent conditions;
• a nucleotide sequence which hybridizes with a nucleotide sequence fully complementary to SEQ ID NO:1 under stringent conditions;
• a nucleotide sequence which hybridizes with SEQ ID NO:2 under stringent conditions;
• or
• a nucleotide sequence which hybridizes with a nucleotide sequence fully complementary to SEQ ID NO:2 under stringent conditions;
• wherein stringent conditions are
• hybridizing for 16-18 hours at 65.degree. C. in 6X SSC buffer containing 10% dextran sulfate, a 5X concentrated Denhardt's solution 10 mM EDTA, 0.05% SDS, 100 .mu.g/ml denatured salmon sperm DNA;
• washing twice for 10 minutes at 65.degree. C. in 2X SSC, once for 30 minutes at 65.degree. C. in 2X SSC+0.1% SDS and once for 10 minutes at 65.degree. C. in 0.1X SSC;
• and
• detecting hybridization to C. jejuni but not to other Campylobacter species;
• (c) amplifying any genomic nucleic acid sequences of Campylobacter jejuni in said sample;
• and
• (d) detecting the presence of any amplified genomic nucleic acid sequences.

A kit for detecting the presence of Campylobacter jejuni in a biological or food sample, comprising:

• (a) a pair of primers, wherein each primer consists of 18 to 30 contiguous nucleotides of a nucleic acid sequence selected from the group consisting of:
• a nucleotide sequence of SEQ ID NO:1;
• a nucleotide sequence of SEQ ID NO:2;
• a nucleotide sequence fully complementary to SEQ ID NO:1;
• a nucleotide sequence fully complementary to SEQ ID NO:2;
• a nucleotide sequence which hybridizes with SEQ ID NO:1 under stringent conditions;
• a nucleotide sequence which hybridizes with a nucleotide sequence fully complementary to SEQ ID NO:1 under stringent conditions;
• a nucleotide sequence which hybridizes with SEQ ID NO:2 under stringent conditions;
• or
• a nucleotide sequence which hybridizes with a nucleotide sequence fully complementary to SEQ ID NO:2 under stringent conditions;
• wherein stringent conditions are
• hybridizing for 16-18 hours at 65.degree. C. in 6X SSC buffer containing 10% dextran sulfate, a 5X concentrated Denhardt's solution 10 mM EDTA, 0.05% SDS, 100 .mu.g/ml denatured salmon sperm DNA;
• washing twice for 10 minutes at 65.degree. C. in 2X SSC, once for 30 minutes at 65.degree. C. in 2X SSC+0.1% SDS and once for 10 minutes at 65.degree. C. in 0.1X SSC;
• and
• detecting hybridization to C. jejuni but not to other Campylobacter species;
• and
• (b) reagents necessary for amplifying genomic nucleic acid sequences of Campylobacter jejuni.

A method for the epidemiological locating and classification of identical Campylobacter jejuni strains and the establishment of links as to their source and the propagation of an infection, said method comprising measuring the repetitiveness in said Campylobacter jejuni strain's genome of a nucleic acid sequence corresponding to the sequence of a probe comprising at least 12 contiguous nucleotides of a nucleic acid sequence selected from the group consisting of:

• a nucleotide sequence of SEQ ID NO:1;
• a nucleotide sequence of SEQ ID NO:2;
• a nucleotide sequence fully complementary to SEQ ID NO:1;
• a nucleotide sequence fully complementary to SEQ ID NO:2;
• a nucleotide sequence which hybridizes with SEQ ID NO:1 under stringent conditions;
• a nucleotide sequence which hybridizes with a nucleotide sequence fully complementary to SEQ ID NO:1 under stringent conditions;
• a nucleotide sequence which hybridizes with SEQ ID NO:2 under stringent conditions;
• or
• a nucleotide sequence which hybridizes with a nucleotide sequence fully complementary to SEQ ID NO:2 under stringent conditions;
• wherein stringent conditions are
• hybridizing for 16-18 hours at 65.degree. C. in 6X SSC buffer containing 10% dextran sulfate, a 5X concentrated Denhardt's solution 10 mM EDTA, 0.05% SDS, 100 .mu.g/ml denatured salmon sperm DNA;
• washing twice for 10 minutes at 65.degree. C. in 2X SSC, once for 30 minutes at 65.degree. C. in 2X SSC+0.1% SDS and once for 10 minutes at 65.degree. C. in 0.1X SSC;
• and
• detecting hybridization to C. jejuni but not to other Campylobacter species .