The use of a machine designed to follow repeatedly and automatically a predetermined sequence of individual operations.
AUTOMATED WASHING
AUTOMATED MEDIA PREPARATORS
AUTOMATED COLLECTION AND
PROCESSING OF SAMPLES
CYTOSPIN
AUTOMATED GRAM STAINING
AUTOMATED STREAKING
SPIRAL PLATER
AUTOMATED ANTIBIOTIC -
SENSITIVITY SYSTEM
AUTOMATIC COLONY COUNTER
AUTOMATED URINE MICROSCOPY -
ANALYSER
Sugar fermentation tests, Cetrimide agar medium, Hugh Leifson medium Shivam kumar Sriwas
Combined presentation on:-
1. Sugar fermentation tests
2. Cetrimide agar
3. Hugh Leifson medium
Covering principle, preparation of the medium, test protocol, result interpretations, test conclusions.
The lecture describes the performance and presentation of the antibiograms by the hospitals based upon recommendations of CLSI and shows experience of some of our MOH hospitals with the advantages and pitfalls in them.
The use of a machine designed to follow repeatedly and automatically a predetermined sequence of individual operations.
AUTOMATED WASHING
AUTOMATED MEDIA PREPARATORS
AUTOMATED COLLECTION AND
PROCESSING OF SAMPLES
CYTOSPIN
AUTOMATED GRAM STAINING
AUTOMATED STREAKING
SPIRAL PLATER
AUTOMATED ANTIBIOTIC -
SENSITIVITY SYSTEM
AUTOMATIC COLONY COUNTER
AUTOMATED URINE MICROSCOPY -
ANALYSER
Sugar fermentation tests, Cetrimide agar medium, Hugh Leifson medium Shivam kumar Sriwas
Combined presentation on:-
1. Sugar fermentation tests
2. Cetrimide agar
3. Hugh Leifson medium
Covering principle, preparation of the medium, test protocol, result interpretations, test conclusions.
The lecture describes the performance and presentation of the antibiograms by the hospitals based upon recommendations of CLSI and shows experience of some of our MOH hospitals with the advantages and pitfalls in them.
Prof.Mr.Kiran K. Shinde (M.Pharm), Assistant professor (VNIPRC)
Pharmaceutical microbiology (Second year b.pharm) (3rd semester)
Introduction
Methods Of Different microbiological assays
Principles of Assays with Procedure
Methods For Standardization of
1. Antibiotics
2. Vitamins
3. Amino Acids
Assessment of new Antibiotic
Microbiological Assay of Vitamin & Amino acid Assessment of a New Antibiotic...Ms. Pooja Bhandare
PHARMACEUTICAL MICROBIOLOGY (BP303T) Unit-IV Part-3
Microbiological Assay of Vitamin & Amino acid Assessment of a New Antibiotic: Introduction:
Principle
Microbiological Assay of Cynocobalamin (Vitamin B12):
Tritrimetric Method.
Turbidimetric Method.
Preparation of Standard Cynocobalmine stock solution:
Preparation of Basal Medium Stock Solution:
Test Solution of the material to be assayed Preparation of inoculum: Procedure of Titrimetric method: Turbidimetric Method: Microbiological assay of Amino acids. Assessment of a New Antibiotic.
Introduction:
MIC of an antibiotic is tested either by one of the following ways,
Liquid Dilution Method.
Solid Dilution Method
The inhibition of growth under standardized conditions may be utilized for demonstrating the therapeutic efficacy of antibiotics.
Any subtle change in the antibiotic molecule which may not be detected by chemical methods will be revealed by a change in the antimicrobial activity and hence microbiological assays are very useful for resolving doubts regarding possible change in potency of antibiotics and their preparations.
principle;
The microbiological assay is based upon a comparison of the inhibition of growth of micro-organisms by measured concentration of the antibiotics to be examined with that produced by known concentrations of a standard preparation of the antibiotic having a known activity.
Two general method are usually employed:-
The cylinder-plate (or cup-plate) method.
The turbidimetric (or tube assay) method.
Assessment of microbial contamination and spoilage. PHARMACEUTICAL MICROBIOLO...Ms. Pooja Bhandare
PHARMACEUTICAL MICROBIOLOGY (BP303T)Unit-VPart-2
Assessment of microbial contamination and spoilage.
Assessment of microbial contamination and spoilage
1. Physical and chemical changes:
2. Assessment of viable microorganisms in non-sterile products:
3. Sterility test:
4. Estimation of pyrogens:
Microbial Limit Tests:
Total Aerobic Microbial Count:
Membrane Filtration.
Plate Count Methods.
Pour Plate Method.
Surface spread Method.
Most Probable Number(MPN)
A scientific presentation that describes the steps to perform antimicrobial sensitivity assay for water insoluble compounds and the type of data that can be obtained.
Similar to Broth microdilution reference methodology (20)
Small ruminant keepers’ knowledge, attitudes and practices towards peste des ...ILRI
Presentation by Guy Ilboudo, Abel Sènabgè Biguezoton, Cheick Abou Kounta Sidibé, Modou Moustapha Lo, Zoë Campbell and Michel Dione at the 6th Peste des Petits Ruminants Global Research and Expertise Networks (PPR-GREN) annual meeting, Bengaluru, India, 28–30 November 2023.
Small ruminant keepers’ knowledge, attitudes and practices towards peste des ...ILRI
Poster by Guy Ilboudo, Abel Sènabgè Biguezoton, Cheick Abou Kounta Sidibé, Modou Moustapha Lo, Zoë Campbell and Michel Dione presented at the 6th Peste des Petits Ruminants Global Research and Expertise Networks (PPR-GREN) annual meeting, Bengaluru, India, 29 November 2023.
A training, certification and marketing scheme for informal dairy vendors in ...ILRI
Presentation by Silvia Alonso, Jef L. Leroy, Emmanuel Muunda, Moira Donahue Angel, Emily Kilonzi, Giordano Palloni, Gideon Kiarie, Paula Dominguez-Salas and Delia Grace at the Micronutrient Forum 6th Global Conference, The Hague, Netherlands, 16 October 2023.
Milk safety and child nutrition impacts of the MoreMilk training, certificati...ILRI
Poster by Silvia Alonso, Emmanuel Muunda, Moira Donahue Angel, Emily Kilonzi, Giordano Palloni, Gideon Kiarie, Paula Dominguez-Salas, Delia Grace and Jef L. Leroy presented at the Micronutrient Forum 6th Global Conference, The Hague, Netherlands, 16 October 2023.
Food safety research in low- and middle-income countriesILRI
Presentation by Hung Nguyen-Viet at the first technical meeting to launch the Food Safety Working Group under the One Health Partnership framework, Hanoi, Vietnam, 28 September 2023
Presentation by Hung Nguyen-Viet at the first technical meeting to launch the Food Safety Working Group under the One Health Partnership framework, Hanoi, Vietnam, 28 September 2023
Reservoirs of pathogenic Leptospira species in UgandaILRI
Presentation by Lordrick Alinaitwe, Martin Wainaina, Salome Dürr, Clovice Kankya, Velma Kivali, James Bugeza, Martin Richter, Kristina Roesel, Annie Cook and Anne Mayer-Scholl at the University of Bern Graduate School for Cellular and Biomedical Sciences Symposium, Bern, Switzerland, 29 June 2023.
Assessing meat microbiological safety and associated handling practices in bu...ILRI
Presentation by Patricia Koech, Winnie Ogutu, Linnet Ochieng, Delia Grace, George Gitao, Lily Bebora, Max Korir, Florence Mutua and Arshnee Moodley at the 8th All Africa Conference on Animal Agriculture, Gaborone, Botswana, 26–29 September 2023.
Ecological factors associated with abundance and distribution of mosquito vec...ILRI
Poster by Max Korir, Joel Lutomiah and Bernard Bett presented the 8th All Africa Conference on Animal Agriculture, Gaborone, Botswana, 26–29 September 2023.
Practices and drivers of antibiotic use in Kenyan smallholder dairy farmsILRI
Poster by Lydiah Kisoo, Dishon M. Muloi, Walter Oguta, Daisy Ronoh, Lynn Kirwa, James Akoko, Eric Fèvre, Arshnee Moodley and Lillian Wambua presented at Tropentag 2023, Berlin, Germany, 20–22 September 2023.
(May 29th, 2024) Advancements in Intravital Microscopy- Insights for Preclini...Scintica Instrumentation
Intravital microscopy (IVM) is a powerful tool utilized to study cellular behavior over time and space in vivo. Much of our understanding of cell biology has been accomplished using various in vitro and ex vivo methods; however, these studies do not necessarily reflect the natural dynamics of biological processes. Unlike traditional cell culture or fixed tissue imaging, IVM allows for the ultra-fast high-resolution imaging of cellular processes over time and space and were studied in its natural environment. Real-time visualization of biological processes in the context of an intact organism helps maintain physiological relevance and provide insights into the progression of disease, response to treatments or developmental processes.
In this webinar we give an overview of advanced applications of the IVM system in preclinical research. IVIM technology is a provider of all-in-one intravital microscopy systems and solutions optimized for in vivo imaging of live animal models at sub-micron resolution. The system’s unique features and user-friendly software enables researchers to probe fast dynamic biological processes such as immune cell tracking, cell-cell interaction as well as vascularization and tumor metastasis with exceptional detail. This webinar will also give an overview of IVM being utilized in drug development, offering a view into the intricate interaction between drugs/nanoparticles and tissues in vivo and allows for the evaluation of therapeutic intervention in a variety of tissues and organs. This interdisciplinary collaboration continues to drive the advancements of novel therapeutic strategies.
Slide 1: Title Slide
Extrachromosomal Inheritance
Slide 2: Introduction to Extrachromosomal Inheritance
Definition: Extrachromosomal inheritance refers to the transmission of genetic material that is not found within the nucleus.
Key Components: Involves genes located in mitochondria, chloroplasts, and plasmids.
Slide 3: Mitochondrial Inheritance
Mitochondria: Organelles responsible for energy production.
Mitochondrial DNA (mtDNA): Circular DNA molecule found in mitochondria.
Inheritance Pattern: Maternally inherited, meaning it is passed from mothers to all their offspring.
Diseases: Examples include Leber’s hereditary optic neuropathy (LHON) and mitochondrial myopathy.
Slide 4: Chloroplast Inheritance
Chloroplasts: Organelles responsible for photosynthesis in plants.
Chloroplast DNA (cpDNA): Circular DNA molecule found in chloroplasts.
Inheritance Pattern: Often maternally inherited in most plants, but can vary in some species.
Examples: Variegation in plants, where leaf color patterns are determined by chloroplast DNA.
Slide 5: Plasmid Inheritance
Plasmids: Small, circular DNA molecules found in bacteria and some eukaryotes.
Features: Can carry antibiotic resistance genes and can be transferred between cells through processes like conjugation.
Significance: Important in biotechnology for gene cloning and genetic engineering.
Slide 6: Mechanisms of Extrachromosomal Inheritance
Non-Mendelian Patterns: Do not follow Mendel’s laws of inheritance.
Cytoplasmic Segregation: During cell division, organelles like mitochondria and chloroplasts are randomly distributed to daughter cells.
Heteroplasmy: Presence of more than one type of organellar genome within a cell, leading to variation in expression.
Slide 7: Examples of Extrachromosomal Inheritance
Four O’clock Plant (Mirabilis jalapa): Shows variegated leaves due to different cpDNA in leaf cells.
Petite Mutants in Yeast: Result from mutations in mitochondrial DNA affecting respiration.
Slide 8: Importance of Extrachromosomal Inheritance
Evolution: Provides insight into the evolution of eukaryotic cells.
Medicine: Understanding mitochondrial inheritance helps in diagnosing and treating mitochondrial diseases.
Agriculture: Chloroplast inheritance can be used in plant breeding and genetic modification.
Slide 9: Recent Research and Advances
Gene Editing: Techniques like CRISPR-Cas9 are being used to edit mitochondrial and chloroplast DNA.
Therapies: Development of mitochondrial replacement therapy (MRT) for preventing mitochondrial diseases.
Slide 10: Conclusion
Summary: Extrachromosomal inheritance involves the transmission of genetic material outside the nucleus and plays a crucial role in genetics, medicine, and biotechnology.
Future Directions: Continued research and technological advancements hold promise for new treatments and applications.
Slide 11: Questions and Discussion
Invite Audience: Open the floor for any questions or further discussion on the topic.
Salas, V. (2024) "John of St. Thomas (Poinsot) on the Science of Sacred Theol...Studia Poinsotiana
I Introduction
II Subalternation and Theology
III Theology and Dogmatic Declarations
IV The Mixed Principles of Theology
V Virtual Revelation: The Unity of Theology
VI Theology as a Natural Science
VII Theology’s Certitude
VIII Conclusion
Notes
Bibliography
All the contents are fully attributable to the author, Doctor Victor Salas. Should you wish to get this text republished, get in touch with the author or the editorial committee of the Studia Poinsotiana. Insofar as possible, we will be happy to broker your contact.
The ability to recreate computational results with minimal effort and actionable metrics provides a solid foundation for scientific research and software development. When people can replicate an analysis at the touch of a button using open-source software, open data, and methods to assess and compare proposals, it significantly eases verification of results, engagement with a diverse range of contributors, and progress. However, we have yet to fully achieve this; there are still many sociotechnical frictions.
Inspired by David Donoho's vision, this talk aims to revisit the three crucial pillars of frictionless reproducibility (data sharing, code sharing, and competitive challenges) with the perspective of deep software variability.
Our observation is that multiple layers — hardware, operating systems, third-party libraries, software versions, input data, compile-time options, and parameters — are subject to variability that exacerbates frictions but is also essential for achieving robust, generalizable results and fostering innovation. I will first review the literature, providing evidence of how the complex variability interactions across these layers affect qualitative and quantitative software properties, thereby complicating the reproduction and replication of scientific studies in various fields.
I will then present some software engineering and AI techniques that can support the strategic exploration of variability spaces. These include the use of abstractions and models (e.g., feature models), sampling strategies (e.g., uniform, random), cost-effective measurements (e.g., incremental build of software configurations), and dimensionality reduction methods (e.g., transfer learning, feature selection, software debloating).
I will finally argue that deep variability is both the problem and solution of frictionless reproducibility, calling the software science community to develop new methods and tools to manage variability and foster reproducibility in software systems.
Exposé invité Journées Nationales du GDR GPL 2024
Nutraceutical market, scope and growth: Herbal drug technologyLokesh Patil
As consumer awareness of health and wellness rises, the nutraceutical market—which includes goods like functional meals, drinks, and dietary supplements that provide health advantages beyond basic nutrition—is growing significantly. As healthcare expenses rise, the population ages, and people want natural and preventative health solutions more and more, this industry is increasing quickly. Further driving market expansion are product formulation innovations and the use of cutting-edge technology for customized nutrition. With its worldwide reach, the nutraceutical industry is expected to keep growing and provide significant chances for research and investment in a number of categories, including vitamins, minerals, probiotics, and herbal supplements.
Richard's aventures in two entangled wonderlandsRichard Gill
Since the loophole-free Bell experiments of 2020 and the Nobel prizes in physics of 2022, critics of Bell's work have retreated to the fortress of super-determinism. Now, super-determinism is a derogatory word - it just means "determinism". Palmer, Hance and Hossenfelder argue that quantum mechanics and determinism are not incompatible, using a sophisticated mathematical construction based on a subtle thinning of allowed states and measurements in quantum mechanics, such that what is left appears to make Bell's argument fail, without altering the empirical predictions of quantum mechanics. I think however that it is a smoke screen, and the slogan "lost in math" comes to my mind. I will discuss some other recent disproofs of Bell's theorem using the language of causality based on causal graphs. Causal thinking is also central to law and justice. I will mention surprising connections to my work on serial killer nurse cases, in particular the Dutch case of Lucia de Berk and the current UK case of Lucy Letby.
Observation of Io’s Resurfacing via Plume Deposition Using Ground-based Adapt...Sérgio Sacani
Since volcanic activity was first discovered on Io from Voyager images in 1979, changes
on Io’s surface have been monitored from both spacecraft and ground-based telescopes.
Here, we present the highest spatial resolution images of Io ever obtained from a groundbased telescope. These images, acquired by the SHARK-VIS instrument on the Large
Binocular Telescope, show evidence of a major resurfacing event on Io’s trailing hemisphere. When compared to the most recent spacecraft images, the SHARK-VIS images
show that a plume deposit from a powerful eruption at Pillan Patera has covered part
of the long-lived Pele plume deposit. Although this type of resurfacing event may be common on Io, few have been detected due to the rarity of spacecraft visits and the previously low spatial resolution available from Earth-based telescopes. The SHARK-VIS instrument ushers in a new era of high resolution imaging of Io’s surface using adaptive
optics at visible wavelengths.
This presentation explores a brief idea about the structural and functional attributes of nucleotides, the structure and function of genetic materials along with the impact of UV rays and pH upon them.
2. Standardisation of AST
• Results change with changed parameters.
– It is crucial to adhere to the methodology to get
reproducible and reliable results!
• Standardisation of:
– Potency of antimicrobial agent/disk potency
– Media
• Type of media, supplements, pH, agar depth etc.
– Inoculum
– Incubation
– Reading of results
3. MIC – the gold standard
Minimum Inhibitory Concentration
• “The lowest concentration that, under defined in
vitro conditions, prevents visible growth of
bacteria within a defined period of time” (ISO
20776-1).
• The MIC recorded as the lowest concentration of
the agent that completely inhibits visible growth.
6. Broth microdilution methodology
Inoculum 5 x 105 CFU/mL
Media MH broth (non-fastidious organisms)
MH-F broth (Fastidious organisms)
Incubation 35°C in air, 16-20 h (sealed panels)
Reading Read the MIC as the lowest
concentration of antimicrobial agent that
completely inhibits growth of the
organism as detected by the unaided eye
ISO 20776-1, 2019
EUCAST media for fastidious organisms
7.
8. Broth microdilution (BMD)
ISO 20776-2, 2019
• Rapidly growing aerobic bacteria
• Fresh or frozen panels (<-60°C)
• Microdilution: ≤200 µL per well
• Standardisation of
– Antimicrobial agents
• Solvents, diluents, 2-fold dilutions
– Inoculum
– Incubation time and temperature
• Referral to EUCAST and CLSI documents for
– Testing of fastidious organisms
– Reading of MIC endpoints
9. EUCAST recommendations for BMD
• Perform BMD according to the ISO standard
– Final inoculum of 5×105 CFU/mL
– Sealed panels
– Incubation in air, at 35°C for 16-20 h
• 24 h incubation for glycopeptides
• Commercial panels with freeze-dried antimicrobials
are accepted by EUCAST if they produce expected
results
– BUT many commercial products include few dilutions,
resulting in:
• Truncated MICs in both the lower and upper end
• Products difficult to verify and control
Non-fastidious and
fastidious organisms
10. EUCAST media for BMD
• Broth media
–Non-fastidious organisms
• Un-supplemented Mueller-Hinton broth
(ISO 20776-1)
–Fastidious organisms
• MH-F broth: Mueller-Hinton broth with 5%
lysed horse blood and 20 mg/L β-NAD
11. Preparation of MH-F broth
• Prepare and autoclave cation-adjusted MHB according to the
manufacturer’s instructions, but with 100 mL less deionized
water per litre to allow for the addition of lysed horse blood.
• Cool medium to 42-45°C.
• Aseptically add 100 mL 50% lysed horse blood and 1 mL β-
NAD stock solution per litre medium and mix well.
• Dispense MH-F broth in sterile containers with screw caps.
• Store MH-F broth at 4-8°C.
– Storage conditions and shelf life should be determined as part of
the laboratory quality assurance programme. A shelf life of 3
months can be expected.
https://www.eucast.org/ast_of_bacteria/media_preparation
12. Preparation of 50% lysed horse
blood for MH-F broth
• Aseptically dilute mechanically defibrinated horse blood with
an equal amount of sterile deionized water.
• Freeze the blood at -20°C overnight and thaw. Repeat the
cycle until the cells are completely lysed (three cycles is
usually sufficient but the ISO standard 20776-1 suggests that
up to seven cycles may be required).
• Clarify the 50% lysed horse blood by centrifugation and
discard the pellet. A clear solution is essential for reading.
Repeating the centrifugation may improve the clarity of the
solution.
• The stock solution may be stored at -20°C in aliquots and
defrosted as required. Do not refreeze unused solution.
https://www.eucast.org/ast_of_bacteria/media_preparation
13. Preparation of inoculum
• Final inoculum of 5×105 CFU/mL
• Start with McF 0.5 (~ 1-2 x 108 CFU/mL for
E. coli ATCC 25922)
– Dilutions depend on:
• Volume of broth in wells (Fresh or freeze-dried
panels)
• Volume to be dispensed in each well
14. Preparation of inoculum
• Example for freeze-dried Sensititre panels with
100 µl dispensing (reconstitution) volume
Organism McF Transfer volume to 11 mL
broth
Non-fastidious Gram-positive
and Gram-negative bacteria
0.5 50 µL
Proteus, Morganella and
Providencia spp.
0.5 10 µL (to reduce problems
with swarming)
Haemophilus influenzae 0.5 50 µL
Streptococcus spp. and other
fastidious Gram-positive cocci
0.5 100 µL
Check manufacturer’s recommendations for commercial plates!
15. Control of inoculum / Viable counts
• Remove 10 μL from the growth control well
immediately after inoculation
• Dilute in 10 mL of broth or saline
• Spread 100 μL of this dilution over the
surface of a suitable agar plate, which is then
incubated overnight
• 20-80 colonies are expected from an
acceptable test suspension
ISO 20776-1, 2019
16. Inoculation of BMD plates
• Manually, preferably with a multi-channel
pipette
• Using a dispensing robot
AIM: part of the Sensititre system
from Thermo Scientific
17. Incubation
• Seal panels with a tight lid or adhesive seal to avoid
evaporation
• To avoid uneven heating, micro-dilution trays should not be
stacked more than four high
– By placing an empty tray at the top of each stack, condensation
on the inside of the adhesive cover is reduced, which facilitates
reading.
• Incubate at 35°C in air for 16-20 h
– Glycopeptides should be read after 24 h incubation
– Prolonged incubation is only allowed when specified in the
EUCAST Breakpoint Table
• Corynebacterium, Aerococcus, Kingella kingae and Campylobacter
18. Reading BMD endpoints
• Results should be read manually. The use of a mirror
may facilitate reading.
– If an automated reader or camera system is used, it must be
calibrated to manual reading.
19. Reading BMD endpoints
• Read the MIC as the lowest concentration of
antimicrobial agent that completely inhibits
growth of the organism as detected by the
unaided eye.
• Read MICs only when there is sufficient growth,
i.e. obvious button or definite turbidity, in the
positive growth control.
20. Growth appearance
• Growth appears as turbidity or as a deposit of cells at the
bottom of the well (i.e. as a pellet). The appearance of
growth differs depending on the microorganism and the
antimicrobial agent tested.
• For round-bottom wells, growth will most often appear as
a button/pellet centered in the middle. For flat-bottom
wells, growth may be scattered.
• Growth in antibiotic-containing wells may differ from
growth seen in the positive growth control, even for pure
cultures.
21. Trailing endpoints
• Most antimicrobial agent-organism combinations give
distinct endpoints.
• Some agent-organism combinations may give trailing
endpoints with a gradual fading of growth over 2 to 3
wells.
• Unless otherwise stated, endpoints should be read at
complete inhibition of growth.
22. Reading of BMD panels
Positive
control
Antimicrobial concentration
Unless otherwise stated, endpoints should
be read at complete inhibition of growth.
23. Turbidity without pellet
• Haze or turbidity without a pellet is often seen for
Pseudomonas spp. and Acinetobacter spp. This should
be regarded as growth and the endpoint read at the first
well with complete inhibition (clear broth).
24. Haemolysis
• For fastidious organisms tested in MH-F broth,
haemolysis of the blood can be seen. This is often
accompanied by turbidity or a deposit of growth (pellet).
• Haemolysis with turbidity or pellet should be regarded as
growth when determining endpoints.
25. Skipped wells
• Occasionally a skip may be seen, i.e. a well showing no
growth bordered by wells showing growth. There are
several possible explanations including incorrect
inoculation, contaminations, heterogenous resistance
etc.
• When a single skipped well occurs, retest the isolate or
read the highest MIC value to avoid reporting isolates as
false susceptible.
• Do not report results for antimicrobial agents for which
there is more than one skipped well.
27. Specific reading instructions
• The following antimicrobial agents require specific
reading instructions:
• Bacteriostatic antimicrobial agents, both with Gram-
positive and Gram-negative organisms
• Trimethoprim and trimethoprim-sulfamethoxazole
• Cefiderocol
31. Summary of BMD methodology
Inoculum 5 x 105 CFU/mL
Media MH broth (non-fastidious organisms)
MH-F broth (Fastidious organisms)
Incubation 35°C in air, 16-20 h (sealed panels)
Reading Read the MIC as the lowest
concentration of antimicrobial agent that
completely inhibits growth of the
organism as detected by the unaided eye
34. MIC determination (broth microdilution according to ISO standard 20776-1 except for fosfomycin where
agar dilution is used)
Medium: Mueller-Hinton broth
Inoculum: 5x10
5
CFU/mL
Incubation: Sealed panels, air, 35±1ºC, 18±2h
Reading: Unless otherwise stated, read MICs at the lowest concentration of the agent that completely inhibits
visible growth. See ”EUCAST Reading Guide for broth microdilution” for further information.
Quality control: Staphylococcus aureus ATCC 29213. For agents not covered by this strain, see EUCAST QC
Tables.
MIC determination (broth microdilution according to ISO standard 20776-1)
Medium: Mueller-Hinton broth + 5% lysed horse blood and 20 mg/L β-NAD (MH-F broth)
Inoculum: 5x105
CFU/mL
Incubation: Sealed panels, air, 35±1ºC, 18±2h
Reading: Unless otherwise stated, read MICs at the lowest concentration of the agent that completely inhibits
visible growth. See ”EUCAST Reading Guide for broth microdilution” for further information.
Quality control: Streptococcus pneumoniae ATCC 49619. For agents not covered by this strain, see
EUCAST QC Tables.
Information in EUCAST Breakpoint Tables
Staphylococcus spp.
Streptococcus pneumoniae
36. Quality control
• Reference methods don’t automatically give
the correct results just because they are
reference methods.
• For reproducible and reliable results,
methodology must be strictly adhered to.
• Quality control must be performed for all
materials used (BMD panels, broth etc).
– Commercial BMD panels must often be tested
with several strains to cover all agents
39. Disk diffusion as control method
• Advantages of performing disk diffusion in
parallel for at least some agents
representing relevant antimicrobials:
– Purity test
– Control of the inoculum
– A second method that can be used to interpret
difficult BMD tests
40. Caveats for BMD
• BMD can only be used for organisms that
grow sufficiently well in broth (MH or MH-F
broth)
– Agar dilution is recommended for anaerobic
bacteria and N. gonorrhoeae and will be
recommended for Nocardia spp.
• Some agents cannot be tested with BMD due
to poor reproducibility
– Mecillinam and fosfomycin should be tested with
agar dilution (ISO 20776-1)
41. Implementation of BMD
• ISO 20776-2 (2021): To guide manufacturers
in the conduct of performance evaluation
studies
• Testing scheme:
– QC each day of testing
• ≥95% within range
– Reproducibility (10 isolates x 3)
• MICs within ± one two-fold dilution of the mode/median
for ≥ 95 % of the results
– Clinical isolates (n=300)
• Essential agreement and bias
42. ISO 20776-2:
Data analysis clinical isolates
• Essential agreement (EA)
– MIC within ± one two-fold dilution step from
the reference MIC (≥ 90%)
• Bias
– Whether the results that differ from the
reference method are significantly skewed or
predominantly in one direction (less than ±
30% bias)
44. Implementation of an MIC method
in a clinical laboratory
• No official EUCAST criteria
• Guidance based on EDL experience:
– Relevant QC strains
• At least three repeats per QC strain
– Strains representing target organism
– Beta-lactamase producing strains must be added
for inhibitor-combination agents
– Relevant resistant QC strains if available (e.g. E.
coli NCTC 13846 for colistin)
• Analyse vs. target and range in EUCAST QC
tables
Use QC strains to practise reading of results!
45. Implementation of an MIC method
in a clinical laboratory
• Guidance based on EDL experience:
– Clinical isolates with known MICs
• 25-100 isolates representing target organisms
(depending on product and agents)
– Wild-type isolates and isolates with elevated MICs
– EUCAST/CCUG reference collections for
S. pneumoniae and P. aeruginosa
– UKNEQAS specimens
– Resistant QC strains
46. EUCAST/CCUG reference collections
• To be used to evaluate or implement MIC methods in
laboratories
• Can be ordered from CCUG, https://ccug.se/
• S. pneumoniae
– 10 strains with varying levels of susceptibility to beta-lactam
agents
– Reference MIC values for beta-lactams and agents from other
classes
• P. aeruginosa
– 9 strains with varying levels of susceptibility to agents from
different classes
– Reference MIC values and genomic resistance profile (WGS)
https://www.eucast.org/ast_of_bacteria/strains_with_defined_susceptibility
51. Example of implementation of BMD
using UKNEQAS isolates
Escherichia coli specimen 3253
Antimicrobial agent
Reference
MIC
(mg/L)
MIC
(mg/L)
Reference
categorisation
(SIR)
Categorisation
(SIR)
Comments
Amikacin 8 16 S I Results close to breakpoint. MIC within ± 1 dilution.
Amoxicillin ≥128 >32 R R
Amoxicillin-clavulanic acid 16 16 R R
Ampicillin ≥128 >32 R R
Cefotaxime 32 >8 R R
Ceftazidime 64 >16 R R
Ceftriaxone 32 >4 R R
Cefuroxime ≥128 >16 R R
Ciprofloxacin 0.25 0.25 S S
Ertapenem 0.06 0.06 S S
Gentamicin 0,5-1 1 S S
Imipenem 0.25 0.25 S S
Meropenem 0.03 0.03 S S
Piperacillin-tazobactam 4 8 S S
Tobramycin 8-16 8 R R