Detection of spore forming bacteria

A technology of spores and bacteria, which is applied in the field of primers for the detection of spore-forming bacteria, and can solve problems such as inability to kill bacterial populations and severe infections

Inactive Publication Date: 2004-06-23
HERCULES INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Sometimes, for example, in the treatment of bacterial infections (such as bronchitis, upper respiratory tract infections, earaches, etc.), the antibiotic chosen is effective against the microorganism causing the infection but not for example a strain of Clostridium (a spore-forming bacteria) group of bacteria
Although Clostridia do not usually cause problems, in the absence of competition from other microbes that have been killed by an initial round of antibiotics, Clostridia can multiply, causing potentially serious infections

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0071] Example 1 - Primer Set No. 1

[0072] A set of spoOA sequences were retrieved from GenBank and aligned using the CLUSTAL sequence alignment program. By sequence alignment, the oligonucleotide initiation sites were screened and a preliminary primer set was selected. A forward primer, 5-AAAAAAGCAGTTGACT-3 (SEQ ID NO. 2), and a reverse primer, 5-CGGCTTGCCGTTGTATT-3 (SEQ ID NO. 3) were synthesized. A PCR product of approximately 300-400 base pairs is expected using this primer set - thermal optimization was performed using "Ready to Go" PCR beads (Pharmacia Biotech, Piscataway, N.J.) and varying annealing temperatures. Reaction conditions for cycling program PCR. This step can use any PCR instrument, non-limiting examples of which are MiniCycler and Delta Cycler. The characteristic SFB used in this test and a set of non-characterized SFB were isolated from one paper mill.

[0073] Uncharacterized samples (samples from paper mills) that did not test positive were subsequ...

Embodiment 2

[0076] Example 2 - Primer Set No. 2

[0077] Based on the results of Example 1 above, a second primer set was generated by comparison with a larger data set. Table 2 shows the data of the forward primer of the improved primer set, while Table 3 shows the data of the reverse primer.

[0078] Bacillus cereus

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Bacillus megaterium

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Bacillus stearothermophilus

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...

Embodiment 3

[0104] Example 3 - Primer Set No. 3

[0105] The primer set in Example 2 may not be consistent in the detection of Bacillus sphaericus. To solve this problem, a new primer set was prepared. The data considered for the preparation of this improved primer set are shown in Table 6 (forward primers) and Table 7 (reverse primers).

[0106] Bacillus cereus

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Bacillus megaterium

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Bacillus stearothermophilus

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C

G ...

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PUM

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Abstract

Disclosed are methods, probes, and nucleic acid sequences for the systematic identification of sporulation genes in spore forming bacteria. A probe comprises a nucleic acid sequence able to form a detectable hybrid with highly conserved regions of the spoOA gene of certain spore forming bacteria Bacillus and Clostridium bacteria species, said nucleic acid sequence being unable to form a detectable hybrid with genetic material of non-spore forming bacteria. Amplification of a portion of the spoOA gene from the cellular DNA of a spore forming bacteria by a polymerase chain reaction using such probe, primer or nucleic acid sequence as one member of a primer set results in the generation of a detectable 346-365 nucleotide long DNA product. One disclosed method comprises a) combining a tagged or labeled probe with a sample, b) hybridizing the probe to the target spore forming bacteria spoOA gene; and c) detecting the hybridized product.

Description

technical field [0001] The present invention relates to a method for the detection of spore forming bacteria. The invention is particularly suitable for bacteria detection of paper products and papermaking processes. Detection of spore-forming bacteria according to the invention includes methods involving polymerase chain reaction (PCR). The invention also discloses primers which are especially suitable for detecting spore-forming bacteria. Background technique [0002] Paper products used for food packaging should not contain various microorganisms that are not conducive to food hygiene. The manufacturing process of paper products is the most common way for this type of microbial contamination to occur, and microorganisms can grow and multiply in paper products. For this kind of pollution, usually take the use of biocide (biocide) or by heating to deal with. However, possible hazards at the paper mill and in the final paper product limit the use of biocides. Furthermor...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C12Q1/689
CPCC12Q1/689
Inventor 亚历山大・W・布林弗雷迪・L・辛格尔顿
Owner HERCULES INC
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