microbiology @ the speed of light

GRAMRAY in Beverage Microbiology

Methode

Speed = Security The growth of bacteria, yeast and mold nega-

tively impacts the sensory properties of bever-

ages and influences the safety of the product

for the consumer. An accurate and fast deter-

mination of microorganisms gives the water,

beer, wine or juice producer the long-term se-

curity they need to guarantee the highest qual-

ity at all times.

Targeted development GRAMRAY has been designed to provide fast,

cost-effective and reliable information on all

spoilage microorganisms in beverage produc-

tion. The early warning system reports the

number and species of all living microbes, ide-

ally before problems can arise. Release deci-

sions for produced batches can be made within

the same day instead of 2-7 days.

methods used for antimicrobial susceptibility

testing

Streamline beverage quality

control with confidence GRAMRAY can be integrated into the company's

HACCP system to identify and control microbial

contamination of raw ingredients, in-process

beverages, and finished goods more quickly

with less hands-on labor than with conven-

tional microbiology methods. Finished goods

release decisions are accelerated and compli-

ance with customer specifications can be easily

demonstrated. The system provides the neces-

sary information for the implementation of

quality standards such as IFS 6.1 quickly and

without additional effort.

Sample Prep in 10 Minutes

Simple and fast sample prep

without culturing or amplifica-

tion steps. GRAMRAY can be elegantly integrated into the

traditional beverage sample prep processes.

Low-cost sample substrate 1X, 4 X, 12X up to 96 X sample carriers are avail-

able. The high-quality smooth metal surface

guarantees a high recovery rate and optimal

spectral quality.

Automated sample preparation mibic is developing a fully automated sample

prep solution to enable processing of up to In

288 GRAMRAY samples per day with minimal

hands on time.

Fully automatic measurement All microbes on each of the up to 96 points of a

well plate are counted and identified. The re-

sults are summarized and displayed in a way

that is easy to understand.

Step 1: Concentration Like traditional methods, fil-

tration takes place on a 47

mm 0.45µm membrane.

Step 2: Matrix removal After concentration, the sam-

ple is resuspended, washed

and centrifuged.

Step 3: Dispensing The concentrated and

washed sample is applied to

an inexpensive specialized

sample substrate.

Easy to Use

All information at a glance GRAMRAY quantifies the number of viable mi-

crobial cells and provides phenotypic charac-

terization with species-level classification,

making validation easy by direct comparison to

CFUs recovered by classical microbiology tech-

niques.

Unparalleled sensitivity &

accuracy The combination of single cell analysis and the

high recovery rate (depending on the sample

matrix) makes the sensitivity of the method

better than that of classical methods. GRAMRAY

is independent of the cultivation of bacteria

and is therefore superior to classical methods

in the ability to detect viable but non-cultura-

ble cells. The high accuracy (specificity) of spe-

cies classification is ensured by integrated and

standardized modelling and machine-based

decisions (artificial intelligence).

Simplest display of results The easy-to-use intuitive software turns any-

one into a successful microbial detective after

just 3 days of training. The measurement result

displays each species present and number of vi-

able cells of each. A quality number indicates

the degree of agreement with the species iden-

tification model, allowing for simple assess-

ment of the classification results.

Open microbiological concept In contrast to molecular biological methods,

the GRAMRAY system is open and can be

adapted to meet the needs of specific indus-

tries, processes and products. "House flora"

can be easily integrated into the species classi-

fication model by automated machine learning

and are reliably assigned in future and past

measurements.

Sensitivity + Specificity

Performance Validation To validate the sensitivity of GRAMRAY studies

were done to test recovery rates of yeasts from

red wine samples. Accuracy of species identifi-

cation was tested with a panel of common bev-

erage spoilage yeasts.

Recovery rate > 95%1* • Preparation eliminates disturbing wine ma-

trix residues on > 80% of all cells

• Removed > 99% of all Debris particles

• Resolved cellular aggregates into measura-

ble single cells

Detection limit - 1 cell • Samples were each inoculated with a nom-

inal 2-3 cells of Saccharomyces cerevisiae.

• GRAMRAY has on average 2.5 cells found

per sample

• Perfect match with results from plate culti-

vation

1 The recovery rate reflects the ratio between a measured value and a reference value. The refer-ence value is the number of colony-forming units by classical cultivation on agar plates.

Species Identification Accuracy Specificity > 88% (true positive)

Selectivity > 97% (true negative) • Yeast cells were isolated from white wine,

red wine and water samples using

GRAMRAY.

• 9 genera, 13 species, 19 strains of yeasts

• Cross-validation of the database: sensitiv-

ity and specificity > 99.6%2.

• Strain identification within a species is also

possible.

• GRAMRAY measurement is not subject to

any preconditions - all microbes present in

the model are identified.

2 This value reflects the correctly identified data-base entries from PASTEURX build in model cross validation

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Viable Cell Counts - GRAMRAY

Single Cell Analysis on GRAMRAY • GRAMRAY scans the entire sample surface

and provides an overview image.

• Target images of the single microbes spec-

tral recording allow a precise examination.

• The Raman spectrum is automatically com-

pared with the stored data model.

• The quality of the spectrum and the

correspondence with the data model is

evaluated.

• Number and species of microorgan-

isms is displayed on results.

GRAMRAY – System overview

Control and software package -

LISTER • Simple recipe creation and execution of

measurements

• Optimized high-throughput sample analy-

sis

• User hierarcy-levels

• Highest start-up accuracy and speed in im-

age analysis and Raman spectroscopy

through automated exposure, illumination

modes, autofocus and binarization control

Identification and artificial intel-

ligence - PASTEURX • Software package for modelling for easy in-

tegration of new microorganisms

• Integrated automated spectra comparison

and result by chemotaxono-metric deci-

sion algorithms

Microbial counting and detec-

tion • Measuring interval for image recognition

of individual microbes: 0.5 µm - 50 µm

• Identification for Gram +/- bacteria, yeasts,

fungi and mycobacteria

• Image Analysis for Dark Field Illumination /

Fluorescence

Spectroscopy • Raman system (≤ 20 mW) - optimized for

single cell identification

• Duration per microbe: 1-100 s

• Laser power - selectable from 5% to 100%.

• Spectral resolution - 5 cm-1.

• Wavelength range 200-3.200 cm-1

• Laser safety - Class 1.

• Lens - 100x, 0.9 NA, 1.0 mm WD.

Documentation and results • Original and binarized image data, Raman

spectrum, database match with quality

number

• Number of Viable Cell Counts

Image analysis and sample unit • 2 MP camera for target evaluation and doc-

umentation

• Feedback-controlled motorized x,y,z table

with a restart accuracy of < 100 nm

• Lighting - dark field and fluorescence

• Magnification - 200x, 500x 1000x.

Calibration and self-tests • Integrated for autocalibration and self-

tests

Environmental parameters • Mains voltage + power - 110-230 V, 500 W

• Weight - 52 kg

• Dimensions - 67x58x35 cm

• Temperature range 17-29 ºC,

• Humidity range 10-75 % non-condensing at

29°C

mibiC GmbH & Co. KG | Ostendstrasse 25| Haus 6 | 12459 Berlin ||||||| ||| ||||||| ||||

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mibi-C.com ||||| fast microbiological classification || spoilage organisms || wild yeast | detection |

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