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WO2025022409A1 - Kit permettant d'identifier les bactéries et de déterminer leur sensibilité aux antibiotiques - Google Patents

Kit permettant d'identifier les bactéries et de déterminer leur sensibilité aux antibiotiques Download PDF

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Publication number
WO2025022409A1
WO2025022409A1 PCT/IN2024/050303 IN2024050303W WO2025022409A1 WO 2025022409 A1 WO2025022409 A1 WO 2025022409A1 IN 2024050303 W IN2024050303 W IN 2024050303W WO 2025022409 A1 WO2025022409 A1 WO 2025022409A1
Authority
WO
WIPO (PCT)
Prior art keywords
kit
tubes
water
antibiotic
media
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Pending
Application number
PCT/IN2024/050303
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English (en)
Inventor
Roshan Ratnakar Naik
Lakshmi Swaroopa Naik
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Individual
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Individual
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Filing date
Publication date
Application filed by Individual filed Critical Individual
Publication of WO2025022409A1 publication Critical patent/WO2025022409A1/fr
Pending legal-status Critical Current
Anticipated expiration legal-status Critical

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Classifications

    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/02—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving viable microorganisms
    • C12Q1/18—Testing for antimicrobial activity of a material

Definitions

  • Urinary tract infections account for 150 million cases annually worldwide with a prevalence rate of 33.54% in India.
  • the most common UTI pathogens are Escherichia coli, Klebsiella pneumoniae, Proteus mirabilis, Enterococcus faecalis and Staphylococcus saprophyticus and all are nitrite positive. Amongst these, 90% of all UTI infections are caused by E. coli.
  • the test has a sensitivity of 25-75% and specificity of 94-100% but exact identification of microorganisms and the right antibiotics is not possible. Patients often try home remedies or take over the counter medication given by pharmacists or doctor prescription based on symptoms but not based on urine culture/microbiology analysis. Use of wrong antibiotics contributes to antibiotic resistance, recurrent UTIs and worsening of symptoms. When symptoms persist beyond 1-2 days then the doctors recommend urine culture that usually involves a series of different media compositions to isolate single colonies using differential media, to identify bacteria using Indole, Methyl Red, Voges Proskauer and Citrate (IMViC), Triple Sugar Iron (TSI) test or similar tests and antibiotic susceptibility tests, leading to a wait of nearly 72 hours to identify bacteria and the correct antibiotics.
  • IMViC Indole, Methyl Red, Voges Proskauer and Citrate
  • TSI Triple Sugar Iron
  • the present invention discloses a kit for identification of bacteria and determining antibiotic sensitivity, comprising: a differential media formulation; distilled water; a color developer; a set of antibiotics; a control test; a set of Eppendorf tubes and a dropper.
  • the differential media formulation includes Peptone- 03 gm by weight%; Meat Extract- 02 gm by weight%; and A nitrate source- 0. 6gm by weight%.
  • the nitrate source is sodium nitrate or potassium nitrate.
  • the differential media formulation is dispersed in 10 ml of water wherein the water is clean tap water or distilled water.
  • the color developer in the kit is Griess reagent.
  • the present invention discloses a method for determination of antibiotic sensitivity having the following steps: dissolving the media formulation in 10 ml of water using the dropper; distributing 1ml of dissolved media into individual Eppendorf tubes and inoculate with 2 drops of urine sample; adding 100 ul of antibiotics into individual tubes along with a negative control tube (no antibiotic) and incubating for 6-8 hours at 37°C; and adding 2 drops of color developer into the Eppendorf tubes and observing the tubes for color change at end of 1 minute.
  • a negative control tube no antibiotic
  • FIG. 1 depicts the methodology of conventional urine culture and testing of antibiotic sensitivity.
  • FIG. 2a and b show graphs indicating the growth curve of gram positive and gramnegative bacteria in media composition of varying concentrations.
  • FIG. 4 depicts the test results of antibiotic susceptibility using an alternative embodiment of the kit of the present invention.
  • references in the specification to "one embodiment” or “an embodiment” means that a particular feature, structure, characteristic, or function described in connection with the embodiment is included in at least one embodiment of the invention.
  • the appearances of the phrase “in one embodiment” in various places in the specification are not necessarily all referring to the same embodiment.
  • the present invention relates to kit for the quantitative or qualitative susceptibility testing for the most clinically significant urinary tract infection causing microorganisms.
  • the kit of the present invention includes the following components:
  • the differential media formulation includes: i) peptone- 03 gm by weight%; ii) meat extract- 02 gm by weight%; and iii) a nitrate source- 0. 6gm by weight%;
  • the above components of the media formulation are added to 100 ml of clean tap water or distilled water.
  • the nitrate source is sodium nitrate or potassium nitrate.
  • the color developer in the kit is Griess reagent.
  • the set of antibiotics included in the kit are Nitrofurantoin (NFN), Trimethoprim (TMP) and Sulfamethoxazole (SMZ).
  • the predefined amount of water is 10 ml wherein the water is clean tap water or distilled water.
  • the predefined amount of dissolved media added into individual Eppendorf tubes is 1 ml.
  • the predefined amount of urine sample from suspected UTI patient is 2 drops (approx. 50-100 ul)
  • the tubes are incubated for predefined period of 6 hrs at a predefined temperature of 37°C.
  • the predefined amount of color developer is 2 drops.
  • a yellow, yellowish orange, pink to red color precipitate indicates presence of S. aureus, MRSA, E. coli and P. aeruginosa respectively that helps in bacterial identification.
  • the kit of the present invention enables bacterial identification and determining antibiotic susceptibility in lesser time than that of the conventional tests thereby reducing the time required in the treatment of urinary tract infection and thus reducing the incidences of antibiotic resistance.
  • the use of Eppendorf tubes reduces headspace leading to less oxygen availability for microbes in nutrient broth and urine. Thus, the reaction process is completed in less time unlike 24 hrs in glass tubes.

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  • Chemical & Material Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Health & Medical Sciences (AREA)
  • Organic Chemistry (AREA)
  • Engineering & Computer Science (AREA)
  • Zoology (AREA)
  • Wood Science & Technology (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Analytical Chemistry (AREA)
  • Toxicology (AREA)
  • Immunology (AREA)
  • Microbiology (AREA)
  • Molecular Biology (AREA)
  • Biophysics (AREA)
  • Physics & Mathematics (AREA)
  • Biotechnology (AREA)
  • Biochemistry (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • General Engineering & Computer Science (AREA)
  • General Health & Medical Sciences (AREA)
  • Genetics & Genomics (AREA)
  • Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)

Abstract

La présente invention concerne un kit permettant d'identifier les bactéries et de déterminer leur sensibilité aux antibiotiques. Le kit de la présente invention comprend les éléments suivants : une formulation de milieu différentiel ; de l'eau distillée ; un révélateur de couleur ; un ensemble d'antibiotiques ; un test témoin ; un ensemble de tubes Eppendorf et un compte-gouttes. La formulation du milieu différentiel comprend du peptone - 0,3 g en % du poids, de l'extrait de viande - 0,2 g en % du poids et une source de nitrate - 0,6 g en % du poids. Le kit de la présente invention permet d'identifier les bactéries et de déterminer leur sensibilité aux antibiotiques en moins de temps qu'avec les tests conventionnels, ce qui réduit le temps nécessaire au traitement de l'infection urinaire et, par conséquent, l'incidence de la résistance aux antibiotiques.
PCT/IN2024/050303 2023-07-21 2024-03-23 Kit permettant d'identifier les bactéries et de déterminer leur sensibilité aux antibiotiques Pending WO2025022409A1 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
IN202321049369 2023-07-21
IN202321049369 2023-07-21

Publications (1)

Publication Number Publication Date
WO2025022409A1 true WO2025022409A1 (fr) 2025-01-30

Family

ID=94374466

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/IN2024/050303 Pending WO2025022409A1 (fr) 2023-07-21 2024-03-23 Kit permettant d'identifier les bactéries et de déterminer leur sensibilité aux antibiotiques

Country Status (1)

Country Link
WO (1) WO2025022409A1 (fr)

Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US3785929A (en) * 1971-11-30 1974-01-15 Warner Lambert Co Diagnostic composition for the detection of nitrite
WO1996028570A1 (fr) * 1995-03-09 1996-09-19 Xechem, Inc. Methode rapide et trousse de diagnostic pour la detection de microorganismes
WO1998050578A1 (fr) * 1997-05-02 1998-11-12 Kocagoez Zuehtue Tanil Test de sensibilite antibacterien
IN2012CH00115A (fr) * 2012-01-11 2013-07-12

Patent Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US3785929A (en) * 1971-11-30 1974-01-15 Warner Lambert Co Diagnostic composition for the detection of nitrite
WO1996028570A1 (fr) * 1995-03-09 1996-09-19 Xechem, Inc. Methode rapide et trousse de diagnostic pour la detection de microorganismes
WO1998050578A1 (fr) * 1997-05-02 1998-11-12 Kocagoez Zuehtue Tanil Test de sensibilite antibacterien
IN2012CH00115A (fr) * 2012-01-11 2013-07-12

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
"07773 Bacteriuria Test Kit (Nitrate Reagent Test Kit; Urine NitriteTest Kit; Nitrite Indicator Strips Kit", SIGMA ALDRICH, 12 December 2015 (2015-12-12), pages 1 - 2, XP009560584 *

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