Listeria monocytogenes

L.monocytogenes morphology

Introduction: history • Murray in 1926: – Bacterium monocytogenes – blood monocytes in rabbits – human and animal pathogen

• Pirie in 1940: – Listeria monocytogenes – Joseph Lister

• Since 1980 – foodborn pathogen

Introduction: general • Cause of listeriosis • Found in environment – water, soil, sudge, …

Found in animal feaces • Taxonomy: – Kingdom: Division: Class: Order: Family: Genus: Species:

Bacteria Firmicutes Bacilli Bacillales Listeriaceae Listeria L. monocytogenes

Listeria monocytogenes • L.monocytogenes is the only important human pathogen among the six species: Listeria monocytogenes, Listeria innocua, Listeria ivanovii, Listeria seeligeri, Listeria welshimeri and Listeria grayi. Only two species of the genus are generally considered to be pathogenic, L. monocytogenes in humans and L. ivanovii in other mammals. L.monocytogenes is a Grampositive , facultatively anaerobic, catalase-positive, oxidasenegative, non-sporeformer. • L.monocytogenes elaborates a 58 kDa β-haemolysin, listeriolysin O, which acts synergistically with the haemolysin produced by Staphylococcus aureus

Characteristics Basic features • • • • • • • • • • • •

Gram positive Facultatively anaerobe Catalase positive Oxidase negative Non-spore former Coccoid to rod shaped 0,4 – 0,5 µm x 0,5 – 2,0 µm Cultured at 20 – 25°C Petritious flagella and tumbling motility T-range: grow 0 – 42 °C, Opt: 30 – 35°C Min pH: 4,4 – 5,6 pH Salt tolerated

Characteristics Biochemical facts enter the the • Foodborn: The pathogen can be present in the original foodstuf or enter foodchain during processing.  Lactating cows can shed L. monocytogenes in milk as a consequence of mastitis for for aa – rather long time.  Soft cheese such as Camembert can be contaminated on the surface during ripening; –

Food items which may contain L. monocytogenes

Food items which in general are free of Listeriae

Sausages (salami, pate¤)

All kinds of food immediately after heating

Raw meat, in particular turkey and chicken

Pasteurized milk, yoghurt (industrial products!)

Sandwiches

Hard cheese

Lettuce, raw mushrooms

Chocolate, marmalade, cookies

Raw milk and products made from this material

Raw carrots

Soft cheese (Munster, Roquefort, Camembert, Brie)

Raw apples

Fresh cheese (ricotta, feta)

Raw tomatoes

Seafood (mussels, salmon) All kind of meals which are conserved after heating

Characteristics Pathogenesis and clinical features The manifestations of listeriosis include: – Septicemia (blood poisoning) – Meningitis – Encephalitis (an acute inflammation ot the brain) – Intrauterine or cervical infections in pregnant women  spontaneous abortion (2nd/3rd trimester) or stillbirth. Symptoms: • Influenza-like symptoms: persistent fever. • Gastrointestinal symptoms: nausea, vomiting, and diarrhea

Characteristics Pathogenesis and clinical features Incubation Period: • Is variable and ranges from 3 to 70 days, with the median incubation period being three weeks. • In infections during pregnancy, the mother usually survives. Treatment: – Parenteral penicillin or ampicillin with aminogycosides. – Trimethoprim-sulfamethoxazole for patients allergic to penicillin.

Prevention: • Similar to prevent from other foodborne illnesses, such as salmonellosis.

L.monocytogenes • Organism grows over a wide range of temperature from 0 – 42oC with an optimum between 30 and 35oC . • Organism is ubiquitous in the environment. Isolation from fresh and salt water, soil, sewage sludge, decaying vegetation, silage. • Oportunistic pathogen – incubation periods for disease from 1 day to 90 days. Symptoms vary from a mild, flu-like illness to mengitis and meningoencephalitis. • Attack : pregnant women, children (0 – 2 years) or elderly (more 65 years) and the immunocompromised population.

Introduction: properties • Gram positive, facultative anaerobe • Non-sporeformer • Rod shaped with flagella – tumbling mobility

• Growth temperature: 0 – 42°C – optimum: 30 – 35°C

• Min pH for growth: 4.4 – 5.6

• D60: few minutes D70: few seconds • Other properties: – catalase +, oxidase –, hemolysis

Foods containing L. monocytogenes • Raw vegetables – cabbage, tomatoes, lettuce, …

• Meat – delicatesses: salami, ham, paté, … – pork sausages, chicken nuggets, …

• Dairy products – pasteurised milk – soft cheeses: brie, camenbert, … – even butter

Listeriosis • Flu-like illness, meningitis, meningoencephalitis, sepsis, (endocarditis) • Patients – pregnant women – newborn and old people – immunocompromised

• Incubation: 1 – 90 days • Infective dose: not known

Listeriosis • Symptoms: – fever, headache, sometimes stomacache – pregnant women: abortion, stillbirth, premature labour – newborn: pneumonia, sepsis, abscesses, meningitis, infection of nervous system

• Treatment with antibiotics – ampicillin, vancomycin, …

• Mortality 20-25% – meningitis 70% – sepsis 50% – neonatal infections 80%

!!! Early diagnosis is rather rare !!!

Listeriosis: lifecycle

Methodology Traditional Identification • Microscopic examination of plates illuminated from below at 45°  blue-grey to blue-green

Conformation • Sugar-fermentation test ( distinguish from other Listeria species)

Methodology Modern Isolation: Two of the most widely-used culture reference methods for detection of Listeria in all foods are – FDA bacteriological and analytical method (BAM) – the International Organization of Standards (ISO) 11290 method

Identification: – CAMP test ( figure) – Chromogenic substrates – Antibody-based tests • •

Enzyme-linked immunosorbent assay (ELISA ) Immuno-capture

– Molecular tests • •

DNA hybridization Polymerase chain reaction (PCR)

Methodology Modern Future directions for the detection and identification of L. monocytogenes •

Tests targeting RNA – Reverse transcription (RT)-PCR – Real-time PCR – Nucleic acid sequence-based amplification (NASBA)

•

DNA microarrays – PCR-based microarrays – Oligonucleotide-based microarrays

Detection: standard • Isolation / enrichment (48h) – buffered Listeria enrichment broth base (BLEB) – slow: enrichment at 4°C – faster: antibiotics at optimal growth temperature

• Incubation 24-48h at 30°C – selective agents: lithium chloride, polyethanol, antibiotics, … – elective agents: aesculin, ferric ammonium citrate – agar: PALCAM, MOX, …

Detection: standard • Identification – blue-green colonies (Henry illumination) – black halo (aesculetin + iron) – other Listeria: yellow (mannitol fermentation)

• Confirmation – catalase, oxidase, nitrate reduction, CAMP tests, … – fast methods

!!! Takes 5-7 days !!!

Detection: new methods • DNA probes – hybridisation

• Recombinant DNA – plasmid

• New, faster media – Rapid'L.Mono Medium

• Commercial kits – AccuProbeTM, GeneTrak

Conclusion 

L. monocytogenes is a wide spread organism, able to grow in many foods



Listeriosis 

dangerous  difficult to recognise  pregnant women, newborn and eldery people, immunocompromised 

Standard detection methods take a long time

Listeria monocytogenes Scanning EM showing Flagella

L.monocytogenes • Association with food : documented were: coleslaw salad, raw vegetable (celery, tomatoes, lettuce), dairy products as raw milk, soft cheeses, smoked salmon, pork tongue in aspic. • Ability to multiply at refrigerator temperatures • Morrtality: 20 – 40%

Detection Listeria monocytogenes according to ISO 11290-1:1996

Characteristics of L. monocytogenes  Gram-positive facultative anaerobic bacterium  resistant to environmental conditions  0,5 – 45 °C  pH from 4,3 to 9,6  high concentration of NaCl (10 – 15 %)  Found in the environments (studied, not known exactly) from there it can infect food

Characteristics of L. monocytogenes  Intracellular parasite - causative agent of alimentary disease listeriosis  High mortality (~ 30 %) in the threaten groups  elder people, babies, persons with weakened or insufficiently developped immune system  abortions at pregnant women

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Food safety criteria COMMISSION REGULATION (EC) No 2073/2005 of 15 November 2005 on microbiological criteria for foodstuffs (Text with EEA relevance)

ANNEX I Microbiological criteria for foodstuffs Chapter 1. Food safety criteria Chapter 2. Process hygiene criteria Chapter 3. Rules for sampling and preparation of test samples COMMISSION REGULATION (EC) No 1441/2007 of 5 December 2007 amending Regulation (EC) No 2073/2005 on microbiological criteria for foodstuffs (Text with EEA relevance) COMMISSION REGULATION (EU) No 365/2010 of 28 April 2010 amending Regulation (EC) No 2073/2005 on microbiological criteria for foodstuffs as regards Enterobacteriaceae in pasteurised milk and other pasteurised liquid dairy products and Listeria monocytogenes in food grade salt (Text with EEA relevance)

Food safety criteria • • • •

Listeria monocytogenes All ready-to-eat foods - Target: Reduction of human listeriosis Although the disease is relatively rare, listeriosis is a significant public health concern Clinical severity and high mortality (20-30%)

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Food safety criteria 2073/2005 Food category

1.1 Ready-to-eat foods intended for infants and ready-to-eat foods for special medical purposes4

Microorgani sm/their toxins, metabolites

Sampling plan 1

Limits 2

n

c

m

L. monocytoge nes

10

0

Absence in 25 g

M

Analytical reference method 3

Stage where the criterion applies

EN/ISO 11290-1

Products placed on the market during their shelf-life

(1) n = number of units comprising the sample; c = number of sample units giving values over m or between m and M. (2) For 1.1-1.24 m=M. (3) The most recent edition of the standard shall be used. (4) Regular testing against the criterion is not useful in normal circumstances for the following ready-eat food - those which have received heat treatment or other processing effective to eliminate L. monocytogenes, when contamination is not possible after this treatment (e.g. Products heat treated in their final package) - fresh, uncut and unprocessed vegetables and fruits, excluding sprouted seeds - bread, biscuits and similar products - bottled or packed waters, soft drinks, beer, cider, wine, spirits, and similar products, - live bivalve molluscs COMMISSION REGULATION (EU) No 365/2010 of 28 April 2010 In footnote 4 of Chapter 1 the following indent is added: ‘— food grade salt’

Food safety criteria 2073/2005 Food category

1.2 Ready-to-eat foods able to support the growth of L. monocytogenes, other than those intended for infants and for special medical purposes

Microorga nism/their toxins, metabolite s

Sampling plan 1

Limits 2

n

c

m

L. monocytog enes

5

0

5

0

Analytical reference method 3

Stage where the criterion applies

100 cfu/g 5

EN/ISO 11290-2 6

Products placed on the market during their shelf-life

Absence in 25 g 7

EN/ISO 11290-1

Before the food has left the immediate control of the food business operator, who has produced it

M

(5) This criterion applies if the manufacturer is able to demonstrate to the satisfaction of the competent authority, that the product will not exceed the limit 100 cfu/g throughout the shelf-life. The operator may fix intermediate limits during the process that should be low enough to guarantee that the limit of 100 cfu/g is not exceeded at the end of the shelf-life. (6) 1 ml of inoculum is plated on a Petri dish of 140 mm diameter or on three Petri dishes of 90 mm diameter. (7) This criterion applies to products before they have left the immediate control of the producing food business operator, when he is not able to demonstrate, to the satisfaction of the competent authority, that the product will not exceed the limit of 100 cfu/g throughout the shelf-life.

Food safety criteria 2073/2005 Food category

1.3Ready-to-eat foods unable to support the growth of L. monocytogenes, other than those intended for infants and for special medical purposes 4, 8

Microorganis m/their toxins, metabolites

Sampling plan 1

Limits 2

n

c

m

5

0

100 cfu/g

M

Analytical reference method 3

Stage where the criterion applies

EN/ISO 11290-2 6

Products placed on the market during their shelf-life

(4) Regular testing against the criterion is not useful in normal circumstances for the following ready-eat food - those which have received heat treatment or other processing effective to eliminate L. monocytogenes, when contamination is not possible after this treatment (e.g. products heat treated in their final package) - fresh, uncut and unprocessed vegetables and fruits, excluding sprouted seeds - bread, biscuits and similar products - bottled or packed waters, soft drinks, beer, cider, wine, spirits, and similar products, - live bivalve molluscs. (6) 1 ml of inoculum is plated on a Petri dish of 140 mm diameter or on three Petri dishes of 90 mm diameter. (8) Products wih pH≤4,4 or aw ≤0,92, products with pH≤5,0 and aw≤0,94 products with a shelf-less than five days are automatically considered to belong to this category. Other categories of products can also belong to this category, subject to scientific justification.

ISO methods for detection L. monocytogenes • ISO 11290-1:1996 • Microbiology of food and animal feeding stuffs -- Horizontal method for the detection and enumeration of Listeria monocytogenes -- Part 1: Detection method • ISO 11290-1:1996/Amd 1:2004 • Modification of the isolation media and the haemolysis test, and inclusion of precision data • ISO 11290-2:1998 • Microbiology of food and animal feeding stuffs -- Horizontal method for the detection and enumeration of Listeria monocytogenes -- Part 2: Enumeration method • ISO 11290-2:1998/Amd 1:2004 • Modification of the enumeration medium

ISO 11290-1:1996 Test portion and initial suspension Primary enrichment (resuscitation) Selective primary enrichment medium: Fraser broth with reduced concentration of selective agents (half Fraser broth) Secondary enrichment Selective secondary enrichment medium with full concentration of selective agents (Fraser broth) Plating out and identification First medium: Oxford agar Second medium: Palcam agar Confirmation of Listeria spp. Solid culture medium: Tryptone soya yeast extract agar (TSYEA) Catalase reaction Gram staining Motility test (if necessary)-tryptone soya yeast extract broth (TSYEAB) or optional motility agar Confirmation of Listeria monocytogenes Haemolysis test Carbohydrate utilisation CAMP test

ISO 11290-1:1996/Amd 1:2004 Plating out and identification First medium: Agar Listeria according to Ottaviani and Agosti (ALOA) Second medium: any other solid selective medium at the choice of the laboratory complementary to ALOA (such as Oxford or PALCAM)

Fraser broth FRASER Listeria Selective Enrichment Broth (base) (g/litre) the high nutrient content and the large buffer capacity = optimum growth conditions for Listeria Proteose peptone 5.0; peptone from casein 5.0; yeast extract 5.0; meat extract 5.0; Sodium chloride 20.0; disodium hydrogen phosphate 9.6; potassium dihydrogen phosphate 1.35; esculin 1.0; lithium chloride 3.0. pH: 7.2 ± 0.2 at 25°C. The prepared broth is clear to almost clear and yellowish-brown. FRASER Listeria Supplements (per vial/l for half-concentrated Fraser broth) Ammonium iron(III) citrate Supplement: Ammonium iron(III) citrate 500 mg – dissolved in 1 ml sterile distilled water Selective Supplement: Acriflavine 12.5 mg; nalidixic acid 10 mg - dissolved in 1 ml sterile distilled water

Fraser broth Lithium chloride, nalidixic acid and acriflavine hydrochloride = large inhibition of accompanying bacteria Esculin and amonium iron(III) citrate = β-D-glucosidase activity of Listeria sp. glucose esculin is cleaved into esculetin and glucose esculetin then forms an olive-green to black complex with the iron(III) ions blackening of the broth is observed

Fraser broth

Oxford agar Oxford Listeria Selective Agar, Base (g/litre) Peptone 23.0; starch 1.0; sodium chloride 5.0; agar-agar 13.0 (= Columbia agar); esculin 1.0; ammonium iron(III) ; citrate 0.5; lithium chloride 15.0. Oxford-Listeria-Selective-Supplement Composition (per vial for 500 ml base) Cycloheximide 200.0 mg, Colistin sulfate 10.0 mg, Acriflavine 2.5 mg, Cefotetan 1.0 mg, Fosfomycin 5.0 mg. (disolve in 5 ml mixture 1:1 of ethanol+sterile distilled water) Lithium chloride, acriflavine, colistin sulfate, cefotetan, cycloheximide and fosfomycin Supression of the growth of the common bacteria (e.g. Gram negative bacteria and a greater part of Gram positive bacteria) Hydrolysing esculin to esculetin – a black complex with iron(III)ions= production of brown-green coloured colonies with a black halo (esculin splitting) of Listeria monocytogenes

Oxford agar

Listeria innocua ATCC 33090

Listeria monocytogenes ATCC 19118

Plating out

PALCAM agar PALCAM Listeria Selective Agar Base acc. to VAN NETTEN et al. Peptone 23.0; yeast extract 3.0; starch 1.0; sodium chloride 5.0; agar-agar 13.0 (= Columbia Agar); D(-)mannitol 10.0; ammonium iron(III) citrate 0.5; esculin 0.8; glucose 0.5; lithium chloride 15.0; phenol red 0.08 (g/l) PALCAM Listeria Selective Supplement acc. to VAN NETTEN et al. (per vial/0,5 l) Polymixin-B-sulfate 5.0 mg; ceftazidime 10.0 mg; acriflavine 2.5xmg – dissolve in 1 ml of sterile distilled water The prepared plates (incl. supplement) are clear and dark-red. Polymyxin, acriflavin, ceftazidime and lithium chloride – inhibition of the Gram-negative and most of the Gram-positive accompanying bacteria. L. monocytogenes - breaking esculin to glucose and esculetin - esculetin forms an olive-green to black complex with iron(III) Mannitol-positive accompanying bacteria (e.g. as staphylococci) grow as yellow colonies, if they are not inhibited.

Plating out

Listeria innocua ATCC 33090

Listeria monocytogenes ATCC 19118

Plating out

ALOA Chromocult® Listeria Selective Agar corresponds to Agar Listeria acc. to Ottaviani and Agosti Peptone from meat 18.0; peptone from casein 6.0; yeast extract 10.0; sodium pyruvate 2.0; glucose 2.0; magnesium glycerophosphate 1.0; magnesium sulphate 0.5; sodium chloride 5.0; lithium chloride 10.0; disodium hydrogen phosphate 2.5; 5-bromo-4-chloro-3-indolyl-β-D glucopyranoside 0.05; agar-agar 13.0 (g/l). Do not autoclave . Do not overheat. Chromogenic medium. The antibiotic solution sensitive to heat. Poor plates directly after adding the antibiotic and enrichment solution

Chromocult® Listeria Agar Selective Supplement Composition (per vial for 500 ml) Amphotericin B 5 mg; Ceftazidime 10 mg; Nalidixic acid sodium salt 10 mg; Polymyxin B sulphate 38350 U (dissolved in 4 ml sterile demineralised water/ethanol mixture (1:1) for 500 ml) Chromocult® Listeria Agar Enrichment Supplement (vial for 20 ml) L-α-phosphatidylinositol 1 g (per 500 ml)

ALOA Basis - good growth for a broad range of bacteria 5-bromo-4-chloro-3-indolyl-β-D-glucopyranoside – substrate for β-Dglucosidase - β-Dglucosidase -positive bacteria as Listeriae grow on the medium in the form of blue-green colonies. L. monocytogenes and L. innocua show virtually uninhibited growth, whilst the growth of other listeriae may be retarded (L. ivanovii) or inhibited (L. seeligeri). Selective supplement - antibiotics – inhibition effect Amphotericin (yeasts and moulds), Polymyxin B (gram-negative bacteria), .nalidixic acid (gram-negative bacteria, some gram- positive bacteria) , ceftazidime (gram-negative and gram-positive bacteria). Enrichment supplement L. monocytogenes and L. ivanovii - enzyme phosphatidylinositol phospholipase C (PI-PLC) described as a virulence factor. This phospholipase activity results in the formation of opaque haloes around colonies of L. monocytogenes (and retardelly of L. ivanovii).

ALOA All colonies which appear blue-green with an opaque halo on the medium are counted as suspect L. monocytogenes colonies (typical colonies). The suspect colonies must be confirmed, as L. ivanovii and some other bacteria, e.g. bacilli, may show the typical colony pattern of L. monocytogenes.

Listeria innocua ATCC 33090

Listeria monocytogenes ATCC 13932

Plating out

ISO 11290-1:1996 Test portion x g or x ml + Primary enrichment medium (half Fraser broth) Incubation at 30 °C for 24 h ±3 h 0,1 ml of culture in 10 ml of secondary enrichment medium (Fraser broth) (subculture)

Plating out on

Incubation at 35 °C or 37 °C for 48 h ±3 h

– Listeria according to Ottaviani - and Agosti (ALOA) –and second selective medium (Oxford, Palcam)

Plating out on – Listeria according to Ottaviani - and Agosti (ALOA) –and second selective medium (Oxford, Palcam)

Incubation of (ALOA) for 24 h ± 3 h and, if necessary, additional 24 h ± 3 h at 37 °C Oxford agar - 35°C up to 48 h aerobically Palcam agar - 35°C for up to 48 hours preferably under microaerophilic conditions

Incubation of (ALOA) for 24 h ± 3 h and, if necessary, additional 24 h ± 3 h at 37 °C Oxford agar - 35°C up to 48 h aerobically Palcam agar - 35°C for up to 48 hours preferably under microaerophilic conditions

Confirmation

Listeria spp. genus confirmation Selection for confirmation: take from each plate of each selective medium five colonies presumed to be Listeria spp. Inoculation on TSYEA (tryptone soya yest extract agar)- a general purpose medium which supports the growth of a wide variety of microorganisms Tryptone 17.0, Dipotassium Phosphate 2.5, Yeast Extract 6.0 Glucose Monohydrate 2.5, Sodium Chloride 5.0, Bacteriological Agar 15.0, Soy Peptone 3.0 (g/l) Incubation at 35 °C or 37 °C for 18-24 hours or until growth is satisfactory. Typical colonies are in 1 mm to 2 mm in diameter, convex, colourless and opaque with an entire edge.

Listeria spp. genus confirmation Catalase reaction Resuspend one colony in a drop of H2O2 solution Listeria spp. is catalase positive – gas bubbles (oxygen) If catalase present, oxygen and water is produced by its activity Gram staining Gram positive slim, short rods Motility test (if necessary) Colony suspended in a tube containing TSYEB (Casein Peptone 17.00 Soy Peptone 3.00 Yeast Extract 6.00 Dipotassium Phosphate 2.50 Sodium Chloride 5.00 D-Glucose 2.50 g/l) and incubate at 25 °C for 8 to 24 h until a cloudy medium is observed. Deposit a drop of this culture onto a clean glass microscope slide, place a coverslip and examine in the microscope – Listeria spp. as slim, short rods with tumbling motility. Motility agar: stab the motility agar and incubate for 48 h at 25 °C. Listeria spp. are motile, giving a typical umbrella-like growth pattern. If growth is not sufficient, incubate for up to an additional 5 days and observe the stab again.

Listeria monocytogenes confirmation Haemolysis Species

Production of acid

CAMP test

Rhamnose

Xylose

S. aureus

R. equi

L. monocytogenes

+

+

-

+

-

L. innocua

-

V

-

-

-

L. ivanovii

+

-

+

-

+

L. seeligeri

+

-

+

(+)

-

L. welshimerii

(+)

V

+

-

-

L. grayi subsp.grayi

-

-

-

-

-

L. grayi subsp.marrayi

-

V

-

-

-

V: variable reaction, (+): weak reaction, +: > 90 % of positive reactions, -: no reactions NOTE: There exist rare strains of L. monocytogenes which do not show β-haemolysis or a positive reaction to the CAMP test under the conditions described in this part of ISO 11290.

Confirmation of L. monocytogenes Haemolysis test Stab one space for each culture on blood sheep agar together with positive and negative control. After incubation at 35 or 37 °C for 24±2 h on blood sheep agar L. monocytogenes show narrow, clear, light zones of β-haemolysis- positive control L. innocua show no clear zone – negative control L. seeligeri show a weak zone of β-haemolysis L. ivanovii usually show wide, clearly delineated zones of β-haemolysis Carbohydrate utilisation inoculate each of the carbohydrate utilization broths with L-rhamnose or D-xylose Incubate at 35 °C or 37 °C up tu 5 days. Positive reaction are indicated by the evolution of the coulour from purple to yellow, mostly within 24 h to 48 h.

Validation of Microbiological analytical methods for SFDA, Riyadh, 9-21 September 2011

CAMP test L. monocytogenes

Rhodococus equi

L. ivanovii L. inocua sheep blood agar

1 cm

tested isolates (L. inocua) Staphylococcus aureus

1-2 mm

The Christie-Atkins-Munch-Peterson = (CAMP) test . β-hemolytic Staphylococcus aureus and non-hemolytic Rhodococcus equi Incubation at 35° C for 24-48 h L. monocytogenes (listeriolysin O - erythrocytes hemolysis) and L. seeligeri hemolytic reactions are slightly enhanced in the zone influenced by the S. aureus streak , while for L. ivanovii hemolytic reaction is strongly enhanced in the zone influenced by R. equi streak. The other species remain non-hemolytic (or weak hemolysis without synergism in L. welshimerii).

Listeria monocytogenes confirmation API Listeria – 24-hour identification of all Listeria species Identification of all species of Listeria in 24 hours. Biomérieux - patented biochemical test (DIM) Reliable results: specific tests distinguish the genus Listeria from other genera of similar morphology; the standardized inoculum with a low bacterial concentration (0.5 MF) avoids mixed cultures and subcultures SinglePath L.Mono Example of lateral flow assay - the specific detection of Listeria monocytogenes after enrichment in Fraser or BLEB Broth for 48 hours