rmm product matrix - rapid microbiology and rapid microbiological methods

5/26/2018 RMM Product Matrix - Rapid Microbiology and Rapid Microbiological Methods

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RMMS FOR MICROBIAL IDENTIFICATION

Company

Product

Name

Scientific

Method

App lic ations

Time to

ResultThroughput

Sample Size

or TypeSensitivity

Organism

LibrariesWorkflow

Abbott

Laboratories,

Ibis Biosciences

Division

PLEX-ID System

Note that this

RMM is currently

being redesigned

and is currently

unavailable.

Genotyping by Multi

Loci Base

Composition

(MLBC) analysis

using PCR and

mass spectrometry

Identification

6-8 hr250 every 24

hr

40-160 uL of

nucleic acid

sample

10 copies per

gene

Bacteria, fungi and

viruses. > 750,000

microbial signatures

with detection down to

the strain level

DNA is extracted from cells originating

from colonies or from liquid samples.

The DNA sample is applied to specific

wells of a 96 well-plate, preloaded

with sequence primer pairs. After PCR

amplification, the plate is loaded on

the PLEX-ID system where pico-liter

amounts of the PCR products are

analyzed in an electrospray TOF

mass spectrometer. The base

composition for all amplicons present

are determined and searched against

a database to determine the

organisms present. This multilocus

approach provides the ability to

distinguish between closely related

species as well as single nucleotide

polymorphisms or mutations. There isno requirement for additional staining,

culturing or enrichment steps. Simple

mixtures of organisms can also be

analyzed. Not for use in diagnostic

procedures.

Applied

Biosystems

MicroSEQ

PCR and gene

sequencing

Identification

4-5 hr

80 per day.

Higher with

greater

capacity

capillaryanalyzers.

Cells from

colonyNot applicable

>1800 bacteria

>90 Mycoplasma

>1100 yeast and mold


from isolated colonies. Amplify target

16S rDNA for bacteria or the D2

region of large-subunit rDNA for fungi

using PCR. Perform sequencing of the

PCR amplicons, resulting in DNA

fragments of various sizes ending in

different nucleotides/dyes. A genetic

analyzer separates the fragments by

size and a laser detects thefluorescence color from each dye,

producing a full gene sequence of the

target DNA. The resulting sequence is

compared with an internal database of

known sequences. No Gram staining

is required.

Battelle

REBS

Raman

spectroscopy

Identification and

enumeration

3 min160 every 8

hr

Cells from

colony, liquid

medium,

product/raw

material,

surfaces

1 cell is

identified and

quantified

Alcaligenes,

Pseudomonas,

Brevundimonas,

Candida, E. coli,

Bacillus, Ralstonia,

vegetative and spore

forms.

Sample material is retained on a

supported film. The area is examined

for microscopic particulates using

Raman spectroscopy and a spectral

signature is provided for each

particulate. The spectral signatures

are statistically correlated to a library

of known microorganisms. No need

for Gram staining. Mixed cultures and

be identified and enumerated.

Non-destructive for further analysis.

Biolog

OmniLog;

MicroStation;

MicroLog

Growth-based;

carbohydrate

utilization.

Identification

2-72 hr 50 per dayCells from


>1226 bacteria and

>885 yeast/mold (GEN

II cards)

>1000 bacteria (GEN III

card)

Cells from isolated colonies are used

to prepare a microbial suspension,

which is then added to specific test

cards containing a variety of

carbohydrates and a colorless

tetrazolium violet dye. If growth

occurs, the dye turns violet in color.

The resulting color patterns are

compared with an internal library.

Gram staining is required when using

GEN II test cards for bacteria, yeast

and mold. No Gram stain is required

for the GEN III bacterial card (ID's

both Gram + and - bacteria).

BD Diagnostic

Systems

Phoenix

Growth-based;

biochemical

utilization and

antibiotic

3 hr 100 per dayCells from

colonyNot applicable >225 bacteria




cards containing substrates (in wells)

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susceptibility

Identification

for biochemical utilization. Color or

fluorescence changes in each well are

compared with an internal library.

Gram staining is required to determine

the correct test card to use.

bioMrieux

Vitek 2 Compact

Growth-based;

biochemical and

carbohydrate

utilization.

Identification

2-18 hr 30-60 per dayCells from


>285 bacteria

>48 yeast




cards containing substrates for

enzymatic utilization, carbohydrate

acidification and other tests. Color or

turbidity changes in each well are

measured every 15 minutes and

results are compared with an internal

library. Gram staining is required to

determine the correct test card to use.

bioMrieux

DiversiLab

PCR

Identification

4 hr

13 samples

per chip in 4

hr. Each

additional

chip (13

samples)

every 1 hr.

Cells from


>168 bacteria

>1100 strain or

sub-species patterns


from isolated colonies. Amplify target

DNA (noncoding repetitive DNA

sequences) using PCR. PCR products

are then separated using

electrophoresis in a microfluidics chip

and the resulting patterns are

compared to an internal library.

bioMrieux

Vitek MS

MALDI TOF mass

spectrometry

Identification

2 min480 every 8

hr

Cells from


508 bacteria

78 fungi

Cells from isolated colonies or liquid

medium are added to a stainless steel

target plate and allowed to dry. A

UV-absorbing matrix is added and the

cells are ionized by a laser. The

ionized particles are accelerated in an

electric field and enter the time of

flight (TOF) tube, where protein and

peptide molecules are separated

according to their mass to charge

ratio. The resulting MALDI-TOF mass

spectrum is compared with an internal

database. Gram staining is not

required. Mixed culture ID possible.

Bruker Daltonics

MALDI Biotyper

MALDI-TOF mass

spectrometry

Identification

1-2 min 30-60 per hr

Cells from

colony or liquid

sample

Not applicable

> 2,000 species and >

4,010 strains of

bacteria, yeast and

mold

Cells from isolated colonies or liquid

medium are added to a stainless steel

target plate and allowed to dry. A

UV-absorbing matrix is added and the

cells are ionized by a laser. The

ionized particles are accelerated in an

electric field and enter the time of

flight (TOF) tube, where protein and

peptide molecules are separated

according to their mass to charge

ratio. The resulting MALDI-TOF mass

spectrum is compared with an internal

database. Gram staining is not

required. Mixed culture ID possible.

Bruker Daltonics

TENSOR

27+HTS-XT

Fourier TransformInfrared (FT-IR)

Spectrometry

Identification

Minutes

Continuous

sampling on

96 and 384

plate formats

Cells from


Yeast, bacilli,

Coryneforms,

Micrococci,

Staphylococci, Bifido,

Clostridia,

Pseudomonads,

Lactobacilli,

Acetobactereaceae

Microorganisms are harvested from

the cultivation medium, suspended in

water and then transferred on a

special IR-transparent, reusable

sample plate. The measurement is

performed after drying of the samples.

The dried biofilm is then analyzed by

the FT-IR microplate reader. The

FT-IR spectrum is then compared with

an internal library.

ceeram

ceeramTools

Genotyping Kits

LP

MLVA (Multi Locus

VNTR Analysis)

Genotyping

2 days100 samples

in 2 days

DNA or culture

cell

Not applicable

Legionella

pneumophila,

Staphylococcus aureus,

Pseudomonas

aeruginosa

MLVA is a PCR based typing method

that relies on the inherent variability

found in many regions of repetitive

DNA called VNTR (Variable Number

Tandem Repeat) which represent

sources of polymorphisms. The MLVA

assay for L. pneumophila examines

12 loci and the assay for S. aureus

and P. aeruginosa examines 16 loci.

Thus, each isolate is defined by a 12

or 16-digit numeric code

corresponding to the number of

repeats at each VNTR. Efficient

amplification of the markers is


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performed in a single or 2 multiplex

PCR reactions. An ABI sequencer is

required to analyze the amplicons.

Dupont Qualicon

Riboprinter

Ribotyping of DNA

fragments

Identification



>1,440 bacteria

>8,500 strain or

sub-species patterns


from isolated colonies. The DNA is cut

into fragments using a restriction

enzyme, which are then separated

according to size. The DNA is

immobilized on a nylon membrane,

denatured to produce single-stranded

DNA, and then hybridized with a DNA

probe (derived from an E. colirRNA

operon). An antibody-enzyme

conjugate is bound to the probe and a

chemiluminescent agent is added,

resulting in a banding pattern that is

compared with an internal database.

Fully automated; no Gram staining

required.

Greiner Bio-One

CytoInspect

PCR and microarray

analysis

Mycoplasma

detection and

identification

5 hr 100 per day

Detection of > 90

species of Mycoplasmaand identification of 40

Mycoplasma species

A microarray based test kit for the

detection and identification of

mycoplasma species in cell cultures

and other biological materials. DNA is

extracted and PCR performed using

primers specific for conserved and

species-specific regions of the16S-23S rRNA intergenic transcribed

spacer (ITS) of Mycoplasma DNA.

The fluorescently labeled fragments

are then hybridized to the microarray

chip. The chip contains probes for

both species-specific targets and a

universal probe for all Mycoplasma.

MIDI

Sherlock MIS

Detection of fatty

acids.

Identification

Standard

method (2 hr);

Instant FAME

method (1,200 bacteria

>200 yeast/mold

Fatty acids are extracted from cells

originating from isolated colonies. The

fatty acids are purified and analyzed

using gas chromatography. The

resulting GC chromatogram is

compared with an internal library. No

Gram staining is required.

Pathogenetix, Inc.

Genome

Sequence

Scanning

Fluorescent tagging

of single molecules

of genomic DNA

Identification,

analysis of microbial

mixtures,

epidemiology,

contamination

source tracing

3.5 - 4.5 hr50 samples

per 24 hr

107- 10

9cells

in 0.1 - 5 mL of

culture or from

colony

0.1% of

microbial target

mixed with other

bacteria

1,000+ organisms

including bacteria,

yeast, mold, and

Mycoplasma. Library

can be customized.

Rack of tubes with suspended cells is

inserted into an automated Sample

Preparation Module to be processed

in parallel. The Module isolates and

purifies DNA, specifically digests it,

tags DNA with fluorescent probes, and

elutes the samples for consecutive

measurement with the Detector

Module. The fragments of genomic

DNA are stretched and fluorescent

signatures generated by the probes

are measured one-by-one. Each

signature is compared with internal

database to identify an isolate or

elucidate the composition of microbial

mixture.

rap.ID

Bio Particle

Explorer BPE

Viable Staining and

Imaging LED

Raman

Spectroscopy

Identification and

enumeration

3-10 min

> 150

samples per

8 hr

Cells from

colony, liquid

medium,

product/raw

material,

surfaces

1 cell is

identified and

quantified

> 30 bacteria; can add

new entries

The sample material is collected on

metal foil. Viability staining and

automated image analysis using dark

field illumination detects viable particle

quantity, shape, and size for particles

ranging from 0.5 m and larger.

Raman spectroscopy is then

performed on each viable particle, and

a spectral signature is provided. The

spectral signatures are statistically

correlated to a library of known

microorganisms. Non-destructive for

further analysis.

Thermo Scientific

Nicolet

Fourier Transform

Infrared (FT-IR)

Spectrometry

Identification


colonyNot applicable Bacteria

Cells from isolated colonies are

incubated in liquid medium for 18 hr,

followed by washing, centrifugation

and resuspension in distilled water. 50

uL of the suspension is transferred

onto a special IR-transparent,

reusable sample plate and dried at


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40-45C under vacuum to create a

biofilm. The biofilm is then analyzed

and biomolecules such as proteins,

lipids, carbohydrates and DNA/RNA

are identified and quantified. The

FT-IR spectrum is then compared with

an internal library.

RMMS FOR QUALITATIVE ANALYSIS

Company

Product

Method

App lic ations

Time to

ResultThroughput

Sample Size

or TypeSensitivity

Organisms

DetectedWorkflow

BioLumix

BioLumix

System

Growth-based; CO2

detection and

selective growth

Detection of specific

organisms;

detection of

microbial growth

8-48 hr

32 tests at a

single

temperature

per

instrument

(32

instruments

can be used

with one

computer)

0.1-1.0 mL1 cell after

enrichment

Total aerobic count,

yeast & mold, coliforms,

E. coli, lactic acid

bacteria,

Enterobacteriaceae,

Salmonella,

Pseudomonas,

Staphylococcus

The test sample is added to unique

vials that contain a selective medium

and a dye. Changes in color or

fluorescence, expressed as light

intensity units, are detected by the

optical sensor and represent growing

microorganisms. The total aerobic

count and total yeast/mold vials detect

microbial growth by monitoring the

generation of CO2, and can be used

to screen for an estimation of

organisms in a test sample that are

above or below a certain quantitative

specification. Liquid and diluted solid

samples can be assayed.

Applied

Biosystems

MycoSEQ

PCR

Mycoplasma

detection

< 5 hr

100 l to 10 ml

of cell culture

sample

< 10 CFU or

copy

equivalent/ml

Detection of > 90

Mycoplasma species

The sample is added to a centrifuge

tube and Mycoplasma is concentrated

in a pellet. The Mycoplasma is

enzymatically lysed and purified using

magnetic beads and washing steps.

Real time PCR is performed and the

Mycoplasma target sequence is

detected via use of amplification plots

and melt curve analysis. If a positive

result fir Mycoplasma is obtained, the

purified DNA can be used in the

MicroSEQ for subsequent

identification.

Bactest

Speedy Breedy

Respirometry,

pressure sensing

Detection of

microbial growth;sterility testing;

presence of specific

organisms

4-20 hr 2-4 per day Up to 50 mL1 CFU after

enrichment

Aerobes, facultative

anaerobes, anaerobes,

and microaerophilicbacteria; yeast

The sample is transferred into a

disposable vessel containing a

general or selective medium. The

vessel is then placed into the portable

respirometer and incubated to

encourage microbial growth. Detection

of metabolic activity is determined by

pressure transients relating to

gaseous exchanges within the closed

culture vessel as a result of microbial

respiration. Continuous data collected

is analyzed in real time, and detection

algorithms process the pressure

transients to alert when significant

changes have taken place. The

instrument measures both positive

and negative pressure meaning that

monitoring can be performed on a

range of microbial processes reacting

to differing conditions within the

culture chamber. The system is

applicable for pharmaceuticals, raw

materials, food and beverage,

veterinary, household and hygiene

products.

BD Diagnostic

Systems

BACTEC

Growth-based; CO2

detection

Detection of

microbial growth;

sterility testing

8-48 hr

240-1200 per

incubation

period

1-10 mL or gm1 CFU after

enrichmentBacteria, yeast, mold

Samples are added directly to bottles

of liquid culture media and incubated

in the system. During microbial

growth, CO2in the closed container

accumulates and is detected by a

fluorometric sensor. The system

automatically monitors the sensor

every 10 minutes, and the generation

of CO2indicates the presence of


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growing microorganisms.

bioMrieux

Bactometer

Growth-based;

measurement of

electrical impedance

Detection of

microbial growth

6-48 hr

64-512

samples per

incubation

period

1 mL1 CFU after


Samples are added to media in a

specialized holder that also contain

two electrodes at the bottom of each

sample test well. The holder is placed

in an incubator and continuously

monitored. Growth is detected by

monitoring the movement of ions

between electrodes (conductance), or

the storage of charge at the electrode

surface (capacitance).

bioMrieux

BacT/ALERT 3D

Dual-T

Growth-based; CO2

detection

Detection of

microbial growth;

sterility testing

24-96 hr

480-1440 per

incubation

period

1-10 mL or gm1 CFU after


Samples are added directly to bottles

of liquid culture media and incubated

in the system (one of two

temperatures). During microbial

growth, CO2in the closed container

accumulates and diffuses into a

colorimetric sensor at the base of the

bottle. Hydrogen ions interact with the

sensor resulting in a decrease in pH,

causing the sensor to change to a

yellow color. The system automatically

monitors the sensor every 10 minutes,

and the generation of CO2indicates

the presence of growing

microorganisms.

BIOTECON

Hygiene

Screening

System

PCR

Bacterial detection

90 min 160 per dayCells from

colony

Corynebacterium,

Staphylococcus,

Macrococcus,

Micrococcus, Kocuria,

and Kytococcus

Bacterial colonies growing on agar

plates are suspended in buffer, and

the suspension is placed in a reaction

tube. Primers, fluoresence resonance

energy transfer (FRET) probes and

Taq polymerase are added to the

reaction tube. PCR amplification and

detection is carried out in a Roche

Diagnostics LightCycler 2.0

Carousel-Based System. The

amplification cycles are monitorined

via fluorescence, and melting curves

are used to determine what target

sequences/organisms are present.

CCM MuScan

Innosieve

Diagnostics Test

Kits

Viability staining and

solid phase

cytometry;

fluorescence

microscopy

Bioburden of raw

material andin-process samples,

finished product,

EM, water, sterility

testing

10-65 minutes100 per 8

hours

Filterable

samples; 1 uL

to 1L

1 - 105cells

Bacteria, yeast, fungal

spores

The test sample is

filtered/concentrated through a

0.45um micro sieve. Microorganisms

are retained on the membrane and

subsequently labeled with a viability

stain and/or a species-specific stain.

Available are total viable count, total

live-dead ratio, total species-specific

count and total species-specific viable

count. After staining a scanning period

using MuScan digital fluorescent

microscopy at specific excitation and

emission wavelengths is performed.

The image processing software

analyzes fluorescent objects on size,

shape and fluorescent signals of all

microbes. The assay can be

user-customized with different types of

fluorescent stains, labeled antibodies,

DNA-probes, etc., depending on the

organism(s) to be detected. Examples

of test kits for specific organisms

include Legionella, Salmonella,

Listeria, Chronobacter and E. coli. The

method is non-destructive such that

detected microbes can be

subsequently cultured.

ceeram

ceeramTools

RT-PCR Real

Time Detection

Kits

Real time RT-PCR

Viral detection

5 hrs

Depends on

thermocycler

used

2 gr digestive

tissue, 25 gr

fruit or

vegetable, 1 L

water

5 genome

copies

Norovirus GI, GII;

Hepatitis A, E;

Enterovirus;Adenovirus; Sapovirus;

Aichivirus; Rotavirus;

Circovirus; Brachyspira;

Giardia;

Cryptosporidium

Elution, concentration, extraction,

purification, amplification,

quantification and interpretation. Any

matrix (food, environmental, health)

may be assayed, including shellfish,

vegetable, herbs and spices, water,

sludge and surfaces. The detection

kits can be used with most PCR

instrumentation.


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Celsis

International Ltd.

Celsis Advance

System using

RapiScreen

ATP

bioluminescence

Bioburden of water,

raw materials,

in-process samples,

Microbial Limits


7/14

used to run an assay contain precise

amounts of LAL reagent, chromogenic

substrate and control standard

endotoxin (CSE). Pipette 25 L of a

sample into each of the four sample

reservoirs of a cartridge. The reader

draws and mixes the sample with the

reagents. After mixing, the optical

density of the wells is measured and

analyzed against an internally-

archived standard curve.

Charles River

Laboratories

EndoSafe PTS

Gram ID

LAL assay

Rapid Gram staining

3-7 min4 every 3-7

min25 L Gram + and - bacteria

Using a loop, place an isolated colony

into 2-3 mLs of saline or LRW

(concentration should be 0.5

McFarland equivalence turbidity

standard units). Add 25 l each of 4

samples/organisms to 4 channels in

the disposable cartridge. Results for

Gram +, Gram , or yeast/mold are

displayed on the screen of a portable,

handheld spectrophotometer.

Charles River

Laboratories

EndoSafe PTS

Glucan Assay

LAL assay

Detection of glucan

30 min 2 per hr 25 L 10-1,000 pg/mLGlucans from yeast and

mold

Portable, handheld spectrophotometer

that utilizes disposable cartridges.

Rapid, in-process test designed for

investigational purposes to detect

(1,3)--D glucans from the cell wallsof most yeasts and molds.

Dupont Qualicon

BAX System Q7

PCR

Detection of

microorganisms

1.5-2.5 hr; 48

hr for

yeast/mold

96 per

1.5-2.5 hr10-50 L

Salmonella, Listeria

monocytogenes,

Camplyobacter

jejuni/coli, E. coli

O157:H7, Enterobacter

sakazakii,


yeast and mold, Vibrio

Samples are enriched in media to

provide enough DNA for analysis and

to eliminate false positives from dead

cells. The enriched samples are

heated in a lysis solution to release

DNA. PCR tablets, which contain all

the reagents necessary for PCR plus

fluorescent dye, are hydrated with

lysed sample and processed in the

cycler/detector. PCR amplifies a DNA

fragment that is specific to a target

organism. The amplified DNA

generates a fluorescent signal, and

results are displayed as positive ornegative results. The system uses

SYBR Green, Taqman and Scorpion

probes, facilitating the detection of

multiple species in a single sample.

Greiner Bio-One

CytoInspect

PCR and microarray

analysis

Mycoplasma

detection and

identification

5 hr 100 per day

Detection of > 90

species of Mycoplasma

and identification of 40

Mycoplasma species

A microarray based test kit for the

detection and identification of

mycoplasma species in cell cultures

and other biological materials. DNA is

extracted and PCR performed using

primers specific for conserved and

species-specific regions of the

16S-23S rRNA intergenic transcribed

spacer (ITS) of Mycoplasma DNA.

The fluorescently labeled fragments

are then hybridized to the microarraychip. The chip contains probes for

both species-specific targets and a

universal probe for all Mycoplasma.

Hitachi

BioMAYTECTOR

ATP

bioluminescence

Detection of

microorganisms

90 min5-6 samples

per 8 hr

Volumetric air

sample (e.g., 1

cubic meter)

1 cell (1

attomole ATP)

Bacteria, bacterial

spores

The system utilizes a highly-sensitive

ATP measuring instrument (100 to

200 times higher sensitivity than

conventional ATP methods) for

detecting airborne bacteria. A unique

spore germination induction

processing method is used to detect

spores. A 1 cubic meter of air is

collected within 10 minutes and

particles are deposited onto a

collection carrier. ATP from dead

bacteria are eliminated while ATPfrom viable bacteria are extracted. An

optical detection system measures

light intensity within 90 minutes.

Hyglos

ELISA

3.5 hrs96 samples

every 3 hrs100 l 0.05-500 EU/mL Endotoxin

Uses a microplate that is pre-coated

with a phage-derived receptor protein


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EndoLISADetection of

endotoxin

which has a high affinity and

specificity for the conserved core

region of LPS (endotoxin). When the

sample matrix is added to the

microplate, the LPS is bound to the

phage protein. Any sample matrix with

potentially interfering components is

then removed by a washing step. The

subsequent detection by recombinant

Factor C and a fluorescence substrate

is left unaffected by inhibitors,

facilitating a reliable quantification ofendotoxin in the sample.

Innosieve

Diagnostics

Real-Time Q-PCR

Detection Kits

Q-PCR

Bacterial detection

3-5 hrs (15-25

cycles)

Depends on

thermocycler

used

25 g

< 1 cell / 25 g

(after

pre-enrichment)

< 10 cells / g or

10

non-degraded

genomes (no

enrichment)

Campylobacter jejuni,

Clostridium perfringens,

Cronobacter sakazakii,

E. coli, Listeria innocua,

Legionella

pneumophila,

Legionella spp., Listeria

monocytogenes,

Salmonella sp.,


Vibrio alginolyticus,

Vibrio cholerae, V.

cholerae tox, Vibrio

parahaemolyticus,Vibrio vulnificus, MRSA

Taqman-based Q-PCR detection kits.

When necessary, use pre-enrichment

of the sample in an appropriate

(selective) medium (1:10 w/v: 25 g

sample + 225 ml medium) for 20- 24

hours at appropriate temperature (the

actual method is provided for each

test kit). This is followed by filtration of

100 ml with an appropriate membrane

system (e.g., 0.45 m, cellulose

nitrate filter). The detection kits can be

used with most PCR instrumentation.

JMAR

BioSentry

Light scattering

Detection of water

pathogens

2 minutesContinuous

monitoringProcess water 600 CFU/mL

Cryptosporidium,

Giardia, E. coli,

Salmonella, Shigella,

Pseudomonas,

Legionella, other

rod-shaped bacteria

The fully automated instrument

functions as a real-time, continuous

flow water-monitoring system. As the

slip stream passes through the flow

cell, it also passes through a laser

beam. When particles 0.4 microns to

10 microns in size are present, a

specific multi-angle light-scatter

pattern will be captured by the units

photodetector. This pattern is then

analyzed and compared to the

Bio-Optical Signatures database using

proprietary algorithms. From this

analysis, relative concentration is

calculated and detected particles are

classified as a bacteria, spore,

protozoan or unknown. The system

does not provide viability data as it

cannot differentiate between live and

dead microorganisms.

Lonza

MycoAlert

ATP

bioluminescence

Mycoplasma

detection

20 min 24 per 8 hr

100 L of

culturesupernatant

< 50 CFU/mL

> 90 species of

Mycoplasma

Viable Mycoplasma are lysed and the

organism's enzymes react with the

MycoAlert Substrate catalyzing the

conversion of ADP to ATP. By

measuring the level of ATP in a

sample both before and after the

addition of the MycoAlert Substrate

a ratio can be obtained which is

indicative of the presence or absence

of Mycoplasma. If these enzymes are

not present, the second reading

shows no increase over the first, while

reaction of mycoplasmal enzymes

with their specific substrates in the

MycoAlert Substrate, leads to

elevated ATP levels.

Micro

Identification

Technologies

Light scattering

Detection ofmicroorganisms

10 min 48 per 8 hr Cells from

colony

10-50 cells

E. coli,

Cryptosporidium,

Giardia

Cells from an isolated colony are

suspended in filtered water in a

sample vial and placed into the

instrument. 35 photo detectors in five

concentric arcs that surround the

sample vial collect Mie scattering light

intensities that are generated when a

cell intersects a red laser beam. The

shape, size and internal/external

structures of microorganisms will

provide a unique scattering signature,

which are compared with an internal

database.


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Merck Millipore

MilliPROBE

PCR

Mycoplasma

detection

4 hr 24 per day

Up to 20 mL of

filterable

sample

< 1-10 CFU/mL

10,000 copies

of Mycoplasma

rRNA

> 90 species of

Mycoplasma

Introduce sample into the

centrifugation assay device to

separate eukaryotic cells from

Mycoplasma. Lysis reagent and

primers for the target ribosomal RNA

sequence is added. Magnetic particles

capture and purify the RNA target.

The purified RNA is added to a

96-well plate, in addition to

amplification mix and enzyme reagent.

Real-Time Transcription Mediated

Amplification (TMA) is performed, andthe fluorescence signal from a

molecular torch probe increases as

amplification occurs. If the

Mycoplasma rRNA target is amplified,

a positive result is reported.

Pall Corporation

Pallchek Rapid

Microbiology

System

ATP

bioluminescence

Sterility testing,

bioburden (product

and surface),

non-sterile product

release, biological

indicator,

preservative

effectiveness,

sanitization

monitoring

1 min

following

24-48 hr

enrichment

and

membrane

filtration steps

100 per day

1-300 mL.

Protocol

adapted for

solids and

non-filterable

samples.

1 CFU after an

enrichment of

24 to 48 hr

Bacteria (aerobic and

anaerobic), yeast and

mold

Samples (filterable or non-filterable)

are enriched in growth media.

Incubated media is then filtered.

Microorganisms collected on the

membrane are lysed with extractant.

Luciferin and luciferase enzyme is

added and light is detected with the

Pallchek Luminometer. A

photomultiplier tube amplifies the

photons and results are reported as

Relative Light Units (RLU).Additional

identification is possible using the

residual broth left in the

Microfunnel.

Pall Corporation

GeneDisc Rapid

Microbiology

System

qPCR

Screening of

pathogens (e.g.

Specified

Microorganisms,

USP )

3 to 8 hours

including

sample

filtration,

nucleic acid

prep, and

PCR

96 per PCR

run

1-300 mL.Protocol

adapted for

solids and

non-filterable

samples.

1 CFU afterenrichment for 6

to 24 hours.

>100 CFU

without

enrichment.

E. coli, Salmonella,

Pseudomonas

aeruginosa,


Candida albicans,

Aspergillus brasiliensis,

STEC and non-STEC,

E. coli 0157, Listeria,

Legionella,

Enterococcus,

Cyanobacteria

Samples are filtered and

microorganisms collected on the

membrane are lysed by a combination

of reagents, sonication and heating.

The DNA and Master Mix (polymerase

and deoxynucleotides) are added to

the GeneDisc plate, pre-loaded with

the primers and probes. The plate is

transferred into the GeneDisc Cycler,

and it rotates through four

temperature zones during the PCR

amplification process. When the target

DNA sequence (from the

microorganism of interest) is

amplified, a fluorescent signal from

the probe will increase and is

measured in real time. Current plate

comes pre-loaded with 6 probes for

compendial specified microorganisms.

The GeneDisc Cycler can measure

fluorescent signal from each probe

simultaneously in real time resulting in

detection and positive identification of

microorganisms in a single PCR run of

less than 1 hour.

Roche

MycoTool

PCR

Mycoplasma

detection

< 5 hr

1 mL of cell

culture

containing 5

106cells/mL

< 1 CFU/mL> 90 species of

Mycoplasma

A 1 mL of sample of cell culture with a

concentration of 5 106cells/mL is

treated with lysis buffer. Mycoplasma

DNA is then purified and the target

DNA is amplified via PCR. The

resulting amplicons are evaluated

using gel electrophoresis.

Soleris

Neogen

Growth-based,

media based

detection via CO2or

pH

Quality, spoilage

and sterility

microbial testing

3 to 48 hours

Up to 128 per

unit; up to 4

units per

computer

0.1-5.0 mL1 CFU after

enrichment

Total aerobic count,

sterility, yeast & mold,

coliforms, E. coli, lactic

acid bacteria,

Enterobacteriaceae,

Pseudomonas,

Staphylococcus,

aciduric organisms

Samples are inoculated directly into a

vial which contains ready to use

media and a detection system. The

optical assay measures microbial

growth by monitoring pH or CO2that

generate a color change as

microorganisms proliferate.

Vivione

Biosciences, LLC

RAPID-B

Flow cytometry

Incoming inspection,

sterility confirmation,

bioburden, drug

20 min for

quantitative

results

Up to 8 hr for

96-160 per 8

hrs75-225 uL

1 CFU after

enrichment

TB, Chlamydia and

Staphylococcus for

clinical samples.

Salmonella, E. Coli

O157, non-O157

Samples and reagent are added to a

vial and mixed (5-10 min). The

RAPID-B system analyzes the sample

using flow cytometry, with results

available within 3-5 minutes. Light


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development,

process control

qualitative

results

STECs,

Staphylococcus, and

Vibrio for food samples

scatter resolution of around 130nm

shows separation of bacteria

populations via size and refractive

index alone. The system utilizes a

multi-parametric detection approach: a

phenotypic library to identify the

bacteria, an immunoprobe to

specifically tag the target organism

and a DNA dye to ascertain the live or

dead cells of the target organism. The

workflow is also based on the limit of

detection (LOD) required and thetarget of interest (may include

enrichment, filtration, centrifugation

and dilution).

RMMS FOR QUANTITATIVE ENUMERATION

Company

Product

Method

App lic ations

Time to

ResultThroughput

Sample Size

or TypeSensitivity

Organisms

DetectedWorkflow

Advencis

Lynx Technology

Growth based &

staining free;

High-Magnification

Imaging

Bioburden of raw

material, in-process

samples & finished

product, water

analysis

24-48 hr 240 per 48 hr

Filtrable

samples, solid

medium

1 CFU after

growth

Bacteria, yeast, mold,

spores

Samples are prepared according to

the compendial method (membrane

filtration or direct inoculation on solid

medium). The system incubates theplates (from 20 to 55C) and

automatically acquires high magnified

images (X50) of micro-colonies. The

image processing software analyzes

information on size and shape and

provides an enumeration for each

plate every 30 minutes. The system is

non-destructive, allowing for follow up

analysis.

AES Chemunex

D-Count and

BactiFlow


flow cytometry

Bioburden of rawmaterial and

in-process samples,

finished product,

EM, water

30 min

150 per day

(BactiFlow)300 per day

(D-Count)

Liquids,

usually less

than 1 mL

10-50 cel ls Bacteria, yeast, mold

Viable cells in a liquid sample are

labeled with a non-fluorescent

substrate. Within the cytoplasm of

metabolically active cells, the

substrate is enzymatically cleaved (by

esterase) to release a fluorochrome.Cells with intact membranes will retain

the fluorescent label. The labeled

organisms pass through a argon ion

laser in the flow cell. Two fluorescence

detectors provide an enumeration in

cells per mL.

AES Chemunex

ScanRDI


solid phase

cytometry

Bioburden of raw

material and

in-process samples,

finished product,


testing

1.5-3 hr 30 per dayFilterable

samples1 cell


spores

The test sample is filtered through a

polyester membrane. Microorganisms

retained on the filter are labeled with a

non-fluorescent substrate. Within the

cytoplasm of metabolically active

cells, the substrate is enzymatically

cleaved (by esterase) to release a

fluorochrome. Cells with intact

membranes will retain the fluorescentlabel. An argon laser scans the

surface of the membrane within 3

minutes, and viable cells are detected.

Auto-fluorescent particles, membrane

fluorescence and background noise

are rejected and a total viable count is

reported. Viable cells may be

subsequently observed using a

phase-contrast microscope and an

automated stage.

Battelle

REBS

Raman

spectroscopy

Identification and

enumeration

3 min160 every 8

hr

Cells from

colony, liquid

medium,

product/raw

material,

surfaces

1 cell isidentified and

quantified

Alcaligenes,

Pseudomonas,

Brevundimonas,Candida, E. coli,

Bacillus, Ralstonia,

vegetative and spore

forms.

Sample material is retained on a

supported film. The area is examined

for microscopic particulates using

Raman spectroscopy and a spectral

signature is provided for eachparticulate. The spectral signatures

are statistically correlated to a library

of known microorganisms. No need

for Gram staining. Mixed cultures and

be identified and enumerated.


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BD Diagnostics

FACSMicroCount


flow cytometry

Fermentation

growth monitoring,

bioburden screening

of raw materials,

in-process samples,

finished product,

water analysis,

probiotics and stock

culture enumeration

4 min

quantitative;

24 hr for

enrichment

based

qualitative test

(presence /

absence)

12-15 per hr

for

quantitative

test;

20 samples

per hr for

qualitative

test

Up to 3 mL

30 to 106

quantitative;

1 cell qualitative

with enrichment

Bacteria, yeast, mold

For a quantitative test, the sample is

first diluted in buffer. For a qualitative

test (presence/absence testing), the

sample is diluted and inoculated into a

proprietary enrichment media, which

is then incubated for 24-48 hours.

Three mL of the diluted sample is then

loaded in the instrument for either test.

The automated system tags nucleic

acid of all microorganisms with

fluorescent stain. Sample is then

loaded into flow cytometer chamber

where a 639 nm red laser hits tagged

microorganisms to produce

fluorescence and side scatter signals.

Signals are captured for each

microorganism to reflect one count on

intensity plot. Total microbial count is

available in both tabular and graphical

formats.

BioVigilant

IMD-A

Light scattering

Active air monitoring

Instantaneous

Continuous

or episodic

monitoring

IMD-A 350

(28.3 L/min)

IMD-A 300

(1.15 L/min)

1 cellBacteria, yeast, mold,

spores

Air is drawn into the instrument.

Particles that pass through a 405 nm

diode laser are sized (0.5 to 10

microns) and enumerated using a Mie

scattering particle counter. At thesame time, particles that contain

biological targets, such as NADH,

riboflavin, and dipicolinic acid, will

auto-fluoresce as they pass through

the laser, and a separate fluorescence

detector will record these as viable

microorganisms. The system;

therefore, provides simultaneous

viable and total particulate data per

cubic volume of air. Results are

obtained in real-time, and there are no

consumables, reagents or media.

CCM MuScan

Innosieve

Diagnostics Test

Kits


solid phase

cytometry;

fluorescence

microscopy

Bioburden of raw

material and

in-process samples,

finished product,

EM, water, sterilitytesting

10-65 minutes100 per 8

hours

Filterable

samples; 1 uL

to 1L

1 - 105cells

Bacteria, yeast, fungal

spores

The test sample is

filtered/concentrated through a

0.45um micro sieve. Microorganismsare retained on the membrane and

subsequently labeled with a viability

stain and/or a species-specific stain.

Available are total viable count, total

live-dead ratio, total species-specific

count and total species-specific viable

count. After staining a scanning period

using MuScan digital fluorescent

microscopy at specific excitation and

emission wavelengths is performed.

The image processing software

analyzes fluorescent objects on size,

shape and fluorescent signals of all

microbes. The assay can be

user-customized with different types of

fluorescent stains, labeled antibodies,

DNA-probes, etc., depending on the

organism(s) to be detected. Examples

of test kits for specific organisms

include Legionella, Salmonella,

Listeria, Chronobacter and E. coli. The

method is non-destructive such that

detected microbes can be

subsequently cultured.

Lonza

MicroCompass II

Note that this

RMM is currently

being redesigned

and is currently

unavailable.

RT-PCR

Estimation of cell

count

4-5 hr 72 per 8 hr ~ 1 mL 100 cells Bacteria, yeast, mold

Microorganisms are captured from

filterable samples via spin filtration,

and are then chemically lysed to

release nucleic acid. Alternate

procedures are used for non-filterable

samples. Nucleic acid extraction and

purification is fully automated using

magnetic beads and washing steps.

Purified DNA is amplified using PCR

and ribosomal RNA is amplified using

reverse transcriptase (RT) PCR. PCR

is monitored in real-time (via a MGB


11 of 14 12.02.2014 13:47


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Eclipse probe), and the number of

cycles where the fluorescence rate

increases above a threshold value is

used to estimate the number of viable

cells in the original sample.

Appropriate studies are required to

correlate the number of PCR cycles

with cell concentration.

Millipore

Milliflex Rapid

Growth-based; ATP

bioluminescence

Bioburden of raw

material and

in-process samples,

finished product,


24-28 hr 60 per dayFilterable

samples

1 CFU after

growthBacteria, yeast, mold

Utilizes a membrane filter to capture

individual cells. Filter the sample and

place the membrane on an

appropriate agar medium to allow the

growth of micro-colonies. Micro-

colonies are then treated with an

ATP-releasing reagent, followed by

the addition of luciferin and luciferase.

Photons of light from each micro-

colony are detected by a luminometer,

and a cell count is reported. May be

able to continue incubation to form

larger colonies for subsequent

microbial identification.

Millipore

Milliflex Quantum

Growth-based;

viability staining and

cellular fluorescence

Bioburden of raw

material and

in-process samples,

finished product,


24-28 hr 60 per dayFilterable

samples

1 CFU after

growthBacteria, yeast, mold

Filter the sample, and incubate the

membrane an an agar cassette to

form micro-colonies. Add

non-fluorescent substrate andincubate an additional 30 min. Within

the cell, the substrate is enzymatically

cleaved, releasing a free fluorochrome

into the microorganism cytoplasm. As

fluorochrome accumulates inside the

cells, the signal is naturally amplified.

The membrane is placed in a reader

and exposed to the excitation

wavelength of the fluorochrome.

Fluorescent micro-colonies are then

automatically enumerated.

Non-destructive; can continue to

incubate media to obtain colonies for

microbial identification.

Pall Corporation

GeneDisc Rapid

MicrobiologySystem

qPCR

Estimation of cell

count

3 to 8 hours

including

sample

filtration,

nucleic acidprep, and

PCR

96 per PCR

run

1-300 mL.

Protocol

adapted for

solids and

non-filterable

samples.

1 CFU after

enrichment for 6

to 24 hours.

>100 CFU

without

enrichment.

E. coli, Salmonella,

Pseudomonas

aeruginosa,


Candida albicans,

Aspergillus brasiliensis,

STEC, E. coli 0157,

Listeria, Legionella,

Enterococcus,

Cyanobacteria

Samples are filtered andmicroorganisms collected on the

membrane are lysed by a combination

of reagents, sonication and heating.

DNA is purified and concentrated by

further filtration. The DNA and Master

Mix (polymerase and

deoxynucleotides) are added to the

GeneDisc plate, preloaded with the

primers and probes. The plate is

transferred into the GeneDisc Cycler,

and it rotates through four

temperature zones during the qPCR

amplification process. With each

amplification cycle a fluorescent signal

is generated from the probe. The point

at which the signal reaches above the

background is called Cycle Threshold

(Ct) value. The higher the DNA copy

(e.g., higher microorganisms), the less

amplification cycles will be required to

reach the Ct value (i.e., faster

detection). Built-in software calculates

the number of genomic copies (e.g.,

amount of DNA, which is directly

related to the number of microbial

cells/CFUs) present in the initial

sample. Identification is possible by

DNA sequencing of the remaining

DNA sample.

Particle

Measuring

Systems

BioLaz

Real-Time

Microbial Monitor

Light scattering


InstantaneousContinuous

monitoring3.6 L/min 1 cell


spores

Air is drawn into the instrument

sensing area via a stainless steel

sample probe. As the air passes

through the system it is illuminated by

a laser. Biological particles that

contain NADH or riboflavin will

auto-fluoresce as they pass through


12 of 14 12.02.2014 13:47


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the laser. Those fluorescing biological

particles are then counted in one of

two size channels. The system can be

integrated into an existing

Environmental Monitoring data

management platform or can be used

with a local PC with available interface

software.

rap.ID Particle

Systems

rap.ID

Viable Staining and

Imaging LED

Raman

Spectroscopy

Identification and

enumeration

3-10 min300-600 ID's

per hour

Cells from

colony, liquid

medium,

product/raw

material,

surfaces

1 cell is

identified and

quantified

> 150 bacterial and

spore entries;

customizable

The sample material is collected on

metal foil using impaction or filtration

methods. Viability staining and

automated image analysis using dark

field illumination detects viable particle

quantity, shape, and size ranging from

0.5 m and larger. Raman

spectroscopy is then performed on

each viable particle and a spectral

signature is provided. The spectral

signatures are statistically correlated

to a library of known microorganisms.


Rapid Micro

Biosystems

The Growth

Direct System

Growth-based;

automated detection

of cellular

auto-florescence

Water analysis,

Bioburden, Sterility,

Environmental

Monitoring

Final results in

one-half the

time of the

compendial

method

Positive

results within

hours

400 samples

for 3-day EM

Test

280 samples

for 5-day

water or

bioburden

tests

20-40

samples per

day for

Sterility Test

Filterable

samples

Standard

Environmental

Monitoring

samples

1 CFU


all organisms that grow

on agar media

Samples are prepared as per the

compendial method and loaded into

the system which manages the

incubation and colony enumeration for

the sample. Proprietary digital imagingtechnology automatically enumerates

micro-colonies in one-half the time

than traditional visual plate counting

methods. The sample is collected onto

a filter placed onto an agar medium

cassette with an optically clear lid.

Illumination with blue light excites

micro-colonies to auto-fluoresce

(without the addition of any reagents),

which are enumerated by a CCD

imaging system. The system

automatically incubates and analyzes

each cassette for the formation of

micro-colonies over time. Particles

that do not grow in size over time are

ignored. Non-destructive; can

continue to incubate media to obtain

colonies for microbial identification.

TSI Inc.

BioTrak

Real-Time Viable

Particle Counter

Light scattering


Instantaneous

Continuous

or episodic

monitoring

28.3 L/min 1 cellBacteria, yeast, mold,

spores

Air is drawn into the instrument and

both viable and total particulates are

simultaneously detected, sized and

enumerated via a 685 nm laser diode

(for particle sizing) and a 405 nm laser

diode (for viability detection). The size

range for detection is from 0.5 to 25

microns. The counting efficiency is

50% at 0.5 microns and 100% for

particles > 0.75 microns (per ISO

21501-4 and JIS B9921). The system

can be calibrated using NIST

traceable standards. Also included is

an integrated particle collection filter

(gelatin filter) to capture

microorganisms for subsequent

growth and identification.

Vivione

Biosciences, LLC

RAPID-B

Flow cytometry

Incoming inspection,

sterility confirmation,

bioburden, drug

development,

process control

20 min for

quantitative

results

Up to 8 hr for

qualitative

results

96-160 per 8

hrs75-225 uL

1 CFU after

enrichment

TB, Chlamydia and

Staphylococcus for

clinical samples.

Salmonella, E. Coli

O157, non-O157

STECs,

Staphylococcus, and

Vibrio for food samples

Samples and reagent are added to a

vial and mixed (5-10 min). The

RAPID-B system analyzes the sample

using flow cytometry, with results

available within 3-5 minutes. Light

scatter resolution of around 130nm

shows separation of bacteria

populations via size and refractive

index alone. The system utilizes a

multi-parametric detection approach: a

phenotypic library to identify the

bacteria, an immunoprobe to

specifically tag the target organism

and a DNA dye to ascertain the live or

dead cells of the target organism. The

workflow is also based on the limit of


13 of 14 12.02.2014 13:47


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k

detection (LOD) required and the

target of interest (may include

enrichment, filtration, centrifugation

and dilution).


14 of 14 12 02 2014 13:47

rmm product matrix - rapid microbiology and rapid microbiological methods

Documents

cells originatingfrom

resulting sequence

pcr products

microbial suspension

fungiusing pcr

dna sample

gene sequence of thetarget

needfor gram staining