Institute of Bacteriology and Mycology University of Leipzig Microbiological and immunological results of dairy cows fed one year with Effective Microorganisms (EM)

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Institute of Bacteriology and MycologyUniversity
of Leipzig Microbiological and immunological
results of dairy cows fed one year with Effective
Microorganisms (EM)

• Monika Krueger, Karen Thaens, W. Schroedl

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Microbiological and immunological results of
dairy cows fed one year with Effective
Microorganisms (EM)
Introduction Microorganisms of the
gastrointestinal tract (GIT) influence health and
performance of animals and human beings. They
live in a close relationship to their
hosts. Quality and quantity of the
microorganisms influence digestion, metabolism
and immuno-logical reactivity of the host.
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Microbiological and immunological results of
dairy cows fed one year with Effective
Microorganisms (EM)

• The GIT is the focus of interaction of the
  outside to the inside. Food modulates
  microorganisms and their products metabolites,
  hormones, toxins, etc. supporting or disturbing
  numerous metabolic, immune and nervous functions
  of the GIT.

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Microbiological and immunological results of
dairy cows fed one year with Effective
Microorganisms (EM)

• High-yielding dairy cows are influenced by diets
  rich of fat, proteins and carbohydrates. These
  diets promote lipolytic, proteolytic and
  saccharolytic microorganisms.
• They overgrow and damage the gastrointestinal
  barrier.
• Microorganisms and their toxins translocate. They
  damage the immune system and they are the cause
  of milk losses and diseases.

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Microbiological and immunological results of
dairy cows fed one year with Effective
Microorganisms (EM)

• Objective I
• In a German dairy farm, cows and their calves
  were infected by Salmonella Zerro, a relatively
  unknown serotype, over a period of several
  months.
• At the beginning of the infection some calves
  and cows diseased severely. At present,
  salmonellosis is eradicated.

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Microbiological and immunological results of
dairy cows fed one year with Effective
Microorganisms (EM)

• Objective II
• Measures
• Effective Microorganisms (EM) were given in
  feed from
• 27. 03.03 - 25.02.04,
• November 03 Lactulose application in
• feed over 6 weeks,
• November 03 vaccination against sal-
• monellosis by farm specific vaccine.

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Microbiological and immunological results of
dairy cows fed one year with Effective
Microorganisms (EM)

• Material and Methods
• A dairy farm of about 1400 cows and their calves
  in Saxony was involved in these investigations.
• Faeces and blood plasmas of 20 randomly selected
  cows of the highest yielding group, 15 cows of
  the medium yielding group, 15 heifers, 15 dried
  off cows and 15 cows immediately after birth were
  investigated.

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Microbiological and immunological results of
dairy cows fed one year with Effective
Microorganisms (EM)
Material and Methods
Schedule of investigation - before EM
application - 6 weeks after permanent EM
application - 6 months after permanent EM
application - 12 months after permanent EM
application
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Material and methodsNumber of investigated
animals
Microbiological and immunological results of
dairy cows fed one year with Effective
Microorganisms (EM)
27.03. 14.05. 24.09. 25.02.
number of all animals 80 72 56 47
group I 20 20 19 18
group II 15 14 10 6
group III 15 13 9 9
group IV 15 12 8 6
group V 15 13 10 8
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Microbiological and immunological results of
dairy cows fed one year with Effective
Microorganisms (EM)
Material and Methods

• The following parameters were used to
• evaluate the effect of EM

Faeces Parameter Total aerobic bacterial
count Gram-negative bacterial count Total
anaerobic bacterial count Clostridium perfringens
Method Nutritional-agar (SIFIN) McConkey-agar
(SIFIN) Columbia-agar (OXOID) Blood-agar with
Neomycine and Polymyxine B
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Microbiological and immunological results of
dairy cows fed one year with Effective
Microorganisms (EM)
Material and methods
Method
Parameter

• MRS-agar (OXOID)
• MRS-agar (OXOID)
• Sabouraud-agar (SIFIN)
• Rappoport enrichment medium, Kauffmann-agar
  (SIFIN)
• Two component-agar acc. to Stolp 1963
• Staphylococcus-Medium No. 110 (OXOID
• Neomycine, Desoxycholat Nutritional-agar

• Lactobacillus spp.
• Bifidobacterium spp.
• Yeasts
• Salmonella serotypes
• Bdellovibrio spp.
• Staphylococci
• Bacteriodes spp.

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ies
Microbiological and immunological results of
dairy cows fed one year with Effective
Microorganisms (EM)
Material and methods

• Blood
• Parameters
• CRP
• Haptoglobin
• LPS binding Protein
• IgG and IgM anti-LPS antibodies E. coli J5
• IgG and IgM anti-
• Salmonella Cerro antibodies

• Method
• ELISA
• ELISA
• ELISA
• ELISA
• ELISA

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Microbiological and immunological results of
dairy cows fed one year with Effective
Microorganisms (EM)
Quantitative results of microbiological investigat
ions of faeces from cows treated with EMa
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Effects of EMa on total aerobic bacterial counts
in faeces of cows
H1 H2 H3 H4
median 4,79 x 105 4,17 x 106 1,07 x 106 2,45 x 106


lg/g
p lt 0,05 to H1
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Effects of EMa on anaerobic total bacterial
counts in faeces of cows
H1 H2 H3 H4
median 2,00 x 106 1,51 x 107 9,12 x 106 1,82 x 107


lg/g
p lt 0,05 to H1
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Effects of EMa on counts of lactobacilli in
faeces of cows
H1 H2 H3 H4
median 1,32 x 105 1,05 x 106 3,89 x 105 1,07 x 105


lg/g
p lt 0,05 to H1
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Effects of EMa on bifidobacteria countsin faeces
of cows
H1 H2 H3 H4
median 4,57 x 105 1,41 x 106 7,94 x 106 1,78 x 107



lg/g
p lt 0,05 to H1
H3, H4 p lt 0,05 to H2
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Effects of EMa on aerobic Gram-negative bacterial
counts in faeces of cows
H1 H2 H3 H4
median 1,78 x 104 3,09 x 105 3,89 x 104 3,98 x 104

lg/g
p lt 0,001 to H1, H3 und H4
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Effects of EMa on yeast counts in faeces of cows
H1 H2 H3 H4
median 3,02 x 103 5,89 x 103 4,27 x 103 3,98 x 103

lg/g
p lt 0,05 to H1, H3 und H4
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Effects of EMa on Bacteroides counts in faeces of
cows
H1 H2 H3 H4
median 1,17 x 104 1,66x 104 5,37 x 103 2,75 x 103

lg/g

p lt 0,05 to H2
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Effects of EMa on staphylococcal counts in
faeces of cows
H1 H2 H3 H4
median 9,12 x 104 1,07 x 105 1,15 x 104 1,17 x 105
lg/g
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Effects of EMa on Clostridium perfringens
countsin faeces of cows
H1 H2 H3 H4
median 1,66 x 104 5,50 x 103 7,08 x 103 6,03 x 103
lg/g
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Effects of EMa on alkalic phosphatase in faeces
of cows
U/g
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Results EMa in feed of cows significantly
increased the total aerobic and anaerobic
bacterial counts in faeces of cows at 6 weeks and
12 months of application. Significant increases
of lactobacilli and bifidobacteria were seen in
faeces 6 weeks and 12 months ( only
bifidobacteria) of application.
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Results Gram-negative aerobic bacteria and
yeasts were significantly increased at 6 weeks of
application. Bacteroides, staphylococci and
Clostridium perfringens were significantly
reduced at 12 months of application only in case
of bacteroides but the isolation number of
C.perfringens (35 to 22) and staphylococci (20
to 12) were reduced from the beginning to the
end of our investigations.
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Results of bacteriological investigations of
group I and V
Salmonella Bdellovibrio Protozoa
H1 0 0 0
H2 0 0 0
H3 7 0 15
H4 0 0 11
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Microbiological and immunological results of
dairy cows fed one year with Effective
Microorganisms (EM)
Quantitative results of volatile fatty acids of
faeces from cows treated with EMa
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Effects of EMa on the amount of volatile fatty
acids in faeces of cows (µmol/g)










p lt 0,05 to H1
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Effects of EMa on the amount of volatile fatty
acids in faeces of cows (mol)





p lt 0,05 to H1
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Results EMa were effective on the amount of
volatile fatty acids in faeces of our cows.The
amounts of acetic acid, propionic acid and
butyric acid increased during the whole time of
investigation. The relationship of acetic to
butyric acid was reduced. This means more butyric
acid was provided for the colonic enterocytes.
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Microbiological and immunological results of
dairy cows fed one year with Effective
Microorganisms (EM)
Quantitative results of immunological parameters
of blood plasma of cows treated with EMa
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C-reactive Protein


?
? Beginning of the vaccination
p lt 0,05 to H2
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Haptoglobin



?
?
p lt 0,05 to H2
Beginning of the vaccination
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LPS-binding Protein

?
Beginning of the vaccination
?
p lt 0,05 to H2
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IgG-anti-LPS E.coli,J5

?
Beginning of the vaccination
?
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IgG-anti-S.Cerro


?
H4 p lt 0,05 to H2 und H3
?
Beginning of vaccination
p lt 0,05 to H1
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IgM-anti-S.Cerro



?

Beginning of vaccination
?
p lt 0,05 to H1
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Results The acute phase proteins C-reactive
protein and haptoglobin increased at 6 and 12
months of EMa application in comparison to 6
weeks. LPS binding protein decreased at 12 months
of EMa application. This means that towards the
end of our investigations the cows were more able
to stabilize their innate unspecific immune
system.
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Results In comparison to this, the specific
immunological parameters - IgG-anti-LPS-antibodie
s and IgM-anti-S. Cerro-antibodies were reduced
and IgG-anti-S. Cerro-antibodies were increased.
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Conclusions I The application of EMa by feed
to high yielding cows for about one year was
suitable to increase healthful bacteria as
lacto-bacilli and bifidobacteria. The increase
of healthful bacteria was accompanied by
increasing amounts of volatile fatty acids.
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Conclusions II EMa were able to induce more
propionic and butyric acids in relationship to
acetic acid. During application of EMa the acute
phase proteins C-reactive protein and haptoglobin
increased and LBP decreased, the specific
antibodies against Salmonella Cerro increase
(IgM, 6 weeks and 6 months) or decrease (IgG, 12
months) because of growing defense capacity of
the animals.
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Conclusions III Altogether the combined
measurements of EMa, Lactulose and vaccination
against Salmonella Cerro were able to defense
salmonellosis and to stabilize the health status
of the investigated animals.