The identification of microorganisms, especially detection of antibiotic resistant bacteria, is of great importance in the medical field. It is well known that health care facilities invest large efforts to prevent patients from being infected with secondary diseases caused by environmental bacteria and especially those due to antibiotic resistant bacteria.
The commonly used method to distinguish between antibiotic resistant bacteria and antibiotic sensitive bacteria is to use PCR directly on a sample or after culturing the sample. Such a method is disclosed, for example, in U.S. Pat. No. 4,683,202. Another method is by detecting the proteome, i.e., different proteins expressed by a genome.
DNA-based methods for universal bacterial detection by detection of common bacterial pathogens are also known in the art, for example, as disclosed in U.S. Pat. Appl. Pub. No. 2005/0042606. Detection of viable bacteria in biological samples by exposing bacterial cultures obtained from the samples to transducing particles having a known host range has been disclosed in PCT Pub. No. WO90/04041.
A problem with these methods is that they generally take a significant amount of time (typically at least an hour) to produce a result, and can only be performed by a qualified professional technician. One possible approach to solving these problems might be the use of spectroscopic techniques, which are inherently faster than these methods. Some spectroscopic methods for identifying bacteria, not specific to antibiotic resistant strains, are already known in the art.
For example, PCT Pub. No. WO98/41842 discloses a system for the detection of bacteria antibody complexes by Raman spectroscopy. The sample to be tested for the presence of bacteria is placed in a medium which contains antibodies attached to a surface for binding to specific bacteria to form an antigen—antibody complex. Similarly, Resonance Raman backscattering is disclosed as a method for identification of a bacterium in U.S. Pat. No. 4,847,198. In these methods, the presence in the Raman spectrum of markers associated with particular bacteria is taken as an indication of the presence of the bacterium.
U.S. Pat. No. 6,379,920 discloses a spectroscopic method for detecting and identifying specific bacteria in a biologic sample, for which it is claimed that culturing is not required. The method includes obtaining spectra of a biological sample from a non-infected patient for use as a reference, subtracting the reference from the spectra of a possibly infected sample, and comparing the fingerprint regions of the resulting difference spectrum with reference spectra of known bacteria.
Naumann et al. (Encyclopedia of Analytical Chemistry, R. A. Meyers (Ed.) pp. 102-131, John Wiley & Sons Ltd, Chichester, 2000) have reported the use of FTIR spectroscopy for detection and classification of bacteria in dried samples. Live microbes have been identified by using FTIR and near-infrared FT-Raman spectroscopies. Other methods involve the use of fluorescence spectroscopy or a combination of the above spectroscopic techniques.
None of the prior art literature discloses means and methods that can quickly (less than one hour) and without the need for a professional technician detect and distinguish antibiotic resistant bacteria and antibiotic sensitive bacteria. Furthermore, none of the prior art literature discloses means and method that can eliminate the interference of water contained in a sample on the experimental signal in order to provide more sensitive and accurate detection of bacteria in general and antibiotic resistant bacteria in particular.
Thus, there is a long felt need for means and methods for rapid, sensitive, and accurate detection and identification of microorganisms from a primary culture plate sample without the use of additional reagents or complicated sample preparation, in particular, means and methods that can differentiate antibiotic-sensitive bacteria from antibiotic-resistant bacteria.