TEAT DIPPING AGENT

TECHNICAL AREA

[0001] The present invention relates to the area teat dipping agents for lactating animals. More in particular it relates to a teat dipping agent based on components which are new within this technical area. This new teat dipping agent possesses several advantages in relation to known teat dipping agents.

BACKGROUND OF THE INVENTION

[0002] Mastitis is an inflammatory reaction of udder tissue and is the most common and most costly disease among lactating cows over the world. The inflammation is a reaction of the lactating tissues on the presence of infectious microorganisms. A large number of different bacteria have been identified as mastitis pathogens. They have been divided into four different groups, contagious, environmental, opportunistic and other bacteria. The majority of the mastitis infections are caused by S. aureus. Another contagious mastitis pathogen is Streptococcus agalactiae. Among the environmental bacteria there are other streptococci and the coliform bacteria, such as Escherichia coli and Klebsielle pneumoniae.

[0003] A large number of different disinfectants (most frequently chlorohexidine or iodophors) are used for dipping the teats immediately after milking in order to prevent bacteria from penetrating into the teat canal and further to lactating tissues. These disinfectants have a killing effect in direct contact between disinfectant and bacterium. In spite of routine use of these agents a number of bacteria escaped the killing effect, i.e. the known agents are not sufficiently effective, which can be due to insufficient amount of active components and the fact that the agents do not reach sufficient contact with the infected sites. It is also known that the effect of these agents fades out very quickly and that renewed contamination of the teats takes place shortly after the treatment. Small wounds and skin tissues on the teats can act as reservoirs for certain bacteria, and live stock with infected teat wounds often show higher mastitis frequencies than other live stock. Furthermore, iodine and chlorohexidin can result in taste changes of the milk and relatively small quantities of iodine and chlorohexidin in milk can cause problems in the manufacture of dairy products.

DESCRIPTION OF THE INVENTION

[0004] The present invention thus relates to a new teat dipping composition for lactating animals, which composition represents a substantial contribution to the technology within the area since it has been found to show improvement in several respects compared to existing teat dipping compositions.

[0005] More in particular, it has, in accordance with the invention, surprisingly been found that a combination of chitosan and a group of polysaccharides functions quite well as active components in such teat dipping composition. Several different effects and advantages have been observed. Quite generally, it means that the composition according to the invention shows higher activity than. known teat dipping agents of the type chlorohexidin and iodophors. An effect in this context is that the new teat dipping composition forms a long time barrier at the teat canal opening and prevents bacteria from penetrating into the teat canal. Furthermore, the composition has a growth inhibiting effect on bacteria and a healing effect on skin wounds further strengthening the protection. This means that the composition can be used for a pure bacteria barrier or bacteriostatic effect, by which follows that it can be used as a prophylactic agent against mastitis. In addition, the composition assists in the healing of wounds and skin fissures. Furthermore, the composition can be given such consistency that it is easy to apply to the teats, for example by dipping or spraying, at the same time as it imparts a softening effect on the teats.

[0006] It can be added in this context with regard to prior art that a composition similar in composition is know per se from for example WO96/02260. However, this known composition has not been referred to or remotely indicated as a teat dipping composition, much less is there in the prior art any indication about the fact that it in such context even shows higher activity than known teat dipping agents of the type chlorohexidin and iodophors.

[0007] The expression teat dipping in connection with the present invention shall be interpreted broadly and in accordance with the terminology used within this technical area. Thus, the composition is not only intended for a dipping of the teats in same but it can, of course, also be applied in another way, such as by spraying, but still fall within the recognised term teat dipping composition or agent.

[0008] A first object of the invention is, with other words, to provide a teat dipping composition intended to be used for its bacteria barrier effect or bacteriostatic effect.

[0009] Another object of the invention is to provide a composition which, when applied, results in a wound healing effect.

[0010] Yet an object of the invention is to provide a composition which, when applied, results in a prophylactic action against mastitis.

[0011] Another object of the invention is to provide a composition, which has such consistency that it is extremely well suited for dipping or spray application on the teats of lactating animals, at the same time as it thereby imparts a softening effect.

[0012] A further object of the invention is to provide a composition which assists in a faster and qualitatively improved healing of wounds on the teats.

[0013] Other objects and advantages of the invention will be clear from the more detailed description thereof below.

DETAILED DESCRIPTION OF THE INVENTION

[0014] More in particular, the invention relates to use of a composition comprising chitosan in combination with a polysaccharide selected from heparin, heparan sulphate and dextran sulphate, as an active component in a solvent, for the manufacture of a teat dipping solution for lactating animals, particularly cows.

[0015] The composition according to the invention is useful for all kinds of lactating animals but, of course, in the first place cows. It is further presented in the form of a solution of the active component in a solvent, said solution with regard to consistency or viscosity generally being adapted to the method of application in a conventional manner.

[0016] From the above it is clear that the invention primarily is based on the specific combination of chitosan and the indicated polysaccharide and that primarily it is not restricted to any specific ratio between said components. The ratio between the components and the total quantity of same in the solution are therefore, in other words, to be determined by the expert in each individual case depending on the effect desired.

[0017] However, in accordance with the invention it is primarily a question of using the stated components in such quantities that the solution when applied results in a bacteria barrier effect or bacteriostatic effect.

[0018] Yet another embodiment means that the composition comprises chitosan and the stated polysaccharide in such quantities that the solution when applied results in prophylactic effect against mastitis.

[0019] Although the invention is not restricted to any specific total concentration of the active components it is generally suitable that the total concentration of chitosan plus polysaccharide in the solution is within the range 0.2-10, particularly 0.2-5, for example 0.25-3 percent by weight, based on the total weight of the solution, where the upper limit is mainly dictated by economic factors.

[0020] According to a preferred embodiment of the invention the weight ratio chitosan:polysaccharide in the composition according to the invention lies within the range 100:1-10:1, particularly 60:1-15:1.

[0021] With regard to the component chitosan it is constituted by a polysaccharide consisting of 1.4-β-bound D-glucose amine units. Said polysaccharide is linear and the properties can vary according to the degree of N-acetylation. In the nature all amino groups are acetylated and the polysaccharide is then called chitin corresponding to the shell of inter alia insects and shellfish. Chitosan is manufactured by N-deacetylation of chitin. Commercially, chitosan is recovered from crab and shrimpshells which are waste products from the fishing industry. When treating chitin with alkali, usually sodium hydroxide, N-deacetylation takes place, i.e. acetamido groups are converted into amino groups. By controlling the conditions for this alkali treatment chitosans of varying degree of N-acetylation can be manufactured.

[0022] In connection with the invention said chitosan, according to a preferred embodiment, has an N-acetylation degree of at most about 90%. In particular, said N-acetylation degree is at most about 50%, in some cases at most about 25%.

[0023] Accordingly, the polysaccharide used in the teat dipping composition according to the invention is selected from heparin, heparansulphate and dextransulphate, and these are commercially available on the market from several manufacturers. Within the framework of the invention are, furthermore, partially hydrolysed forms of these polysaccharides as long as they essentially maintain the activity desired in the context. The expression polysaccharide shall, in other words, be interpreted broadly in connection with the invention. The particularly preferred polysaccharide is, however, dextransulphate since it shows very good activity at the same time as it is considerably cheaper than the other polysaccharides.

[0024] As a carrier or matrix the composition according to the invention contains a conventional one, usually an aqueous carrier, for example pure water.

[0025] The solution is imparted suitable consistency or viscosity suitable for the application method used and this can be obtained according to conventional technology. As exemples of viscosity controlling agents for use in the composition according to the invention, there may be mentioned hemicellulose types, for example arabinoxylanes and glucomannanes; plant gum materials, for example guar gum and johannistree gums; cellulose types and derivatives thereof, for-example methyl cellulose, ethyl cellulose, hydroxyethyl cellulose or carboxymethyl cellulose; starch and starch derivatives, for example hydroxyethyl starch or crosslinked starch; microbial polysaccharides, for example xantan rubber, curdlan, pullulan or dextran; and glycerol. Particularly preferred viscosity controlling agents are, however, different types of cellulose and derivatives thereof, particularly methyl cellulose, and glycerol.

[0026] Since the composition described above can be used as a barrier-forming teat dipping agent for lactating animals without any therapeutic effect another aspect of the invention relates to the use of the composition as defined above as a barrier-forming or bacteriostatic teat dipping agent for lactating animals, particularly cows.

[0027] Since the composition, furthermore, can be utilised to counteract mastitis according to another aspect of the invention it represents the use of said composition for the manufacture of a teat dipping agent having prophylactic action against mastitis of lactating animals, particularly cows.

[0028] A further aspect of the invention is represented by a method for prophylactic treatment of lactating animals, particularly cows, against mastitis, where on the teats of said animals a composition according to the above definition in a quantity effective for preventing the generation of mastitis is applied.

[0029] The composition according to the invention can be prepared by dissolving the active components and, optionally, additives, such as for example viscosity controlling agents, in the selected solvent in the desired concentration. An advantageous method in connection with the composition according to the invention resides in dissolving chitosan and polysaccharide separately in a suitable quantity of solvent, for example about half each of the quantity solvent totally used. The additives, for example the viscosity controlling agents, are thereby dissolved together with the polysaccharide before both solutions of chitosan and polysaccharide are admixed to a common solution which is carefully admixed, for example in a mixer, to a final teat dipping solution.

EXAMPLES

[0030] The invention will now be further illustrated by the following non-limiting examples.

Example 1

[0031] General procedure

[0032] Vertical agar plates are covered with chitosan-dextransulphate solution before they are dipped in concentrated bacteria solutions and after said dipping. Chitosan-dextransulphate is compared with commercial iodine solution used as a teat dipping agent, and the agar plates are treated in exactly the same manner with iodine solution. The agar plates are incubated in a heating cabinet over night and the number of bacteria colonies are calculated and recorded as cfu/mL of the bacteria solution. Controls are untreated agar plates.

[0033] Materials and reagents

[0034]

E
-coli, isolated from mastitis infection (Mastitlaboratoriet, SVA) S.aureus, isolated from mastitis infection (Mastitlaboratoriet, SVA) Strept. agalactiae, Strept. dysgalactiae and Strept. uberis, isolated from mastitis infection (Mastitlaboratoriet, SVA) Soy broth, Tryptic soy broth, Difco 0370-01-1 Verical agar plates, Uricult Orion UF009

[0035] Sterile bench, Kojair, LAF 7022 Photometer, Shimadzu UV-1201 Shake incubator, Orbit Environ-Shaker, Lab-Line In struments Inc. Heat cabinet, Memmert

[0036] NaCl, Riedel-de Haën 31434 Chitosan, Seacure 343, Pronova Biopolymer Acetic acid, Riedel-de Haën NaOH Methyl cellulose, Sigma M-0387 Glycerol, Johnson Matthey Electronics 13797 Dextran sulphate, Pharmacia iodine solution, Dipal, Alfa Laval 997515-80

[0037] Procedure

[0038]

E.coli
and S.aureus and three different streptococci (Strept. agalactiae, Strept. dysgalactiae and Strept. uberis) are sterile transferred (sterile bench) with loop from storage agar to separate sterilized trypton soy broth and incubated over night at 37° C. in a shake incubator. The following morning the bacteria are washed three times with a physiological salt solution. The optical density at 650 nm was measured with a photometer, and by means of standard curves the bacteria density was adjusted.

[0039] Chitosan-dextran sulphate solution

[0040] 1 g chitosan was dissolved in 50 mL sterile water, and the pH was adjusted to 5.8. 0.5 g glycerol and 0.1 g methylcellulose and 0.02 g dextransulphate were dissolved in 50 mL sterile water. Equal amounts of chitosan solution and glycerol +methyl cellulose +detransulphate solution were mixed in a mixer.

[0041] Iodine solution

[0042] The base solution was diluted according to the instruction 6 times with water for giving a solution for use.

[0043] Bacteria tests

[0044] (i) Bacterial growth under chitosan-dextran sulphate film.

[0045] Each bacterial type was treated separately according to the following:

[0046] Three agar plates were lowered into the bacteria solution, and two of the agar plates were then dipped three times in each of the above solutions. One agar plate was left untreated.

[0047] (ii) Bacteria growth on top of chitosan-dextran sulphate film.

[0048] Each type of bacterium was treated separately according to the following:

[0049] Two agar plates were dipped three times in each of the above solutions. One agar plate was left untreated. All three agar plates were then immediately quenched in the bacteria solution.

[0050] The agar plates were incubated over night in a heat cabinet at 37° C. Then the plates and the number of colonies were inspected and registered.

[0051] Results

1TABLE 1Bacterial growth under chitosan-dextran sulphate filmCFU/mLChitosan-dextranPatogeneControlsulphateIodineE. coli10 000 00000S. aureus 1 000 00000Streptococci10 000 00000

[0052]

2

TABLE 2

Bacterial growth on top of chitosan-dextran sulphate film

CFU/mL

Chitosan

dextran

Patogen
Controll
sulphate
Iodine

E. coli

10 000 000
0
10 000 000

S. aureus

1 000 000
0
1 000 000

Streptococci
10 0000 000
0
10 000 000

[0053] The results are presented in FIGS. 1 and 2.

Example 2

[0054] Materials and methods

[0055] 6 cows of an age of 3-5 years were involved in the study. Five of the cows were of SRB-rase (Svensk Rödbrokig Boskap) and the sixth was a crossing between the SRB and SLB (Svensk L{dot over (a)}glands Boskap). The animals were acquired from one and the same stock and were bought 12 days before initiating the experiments. The cows were stabled and a maintenance ration of fodder was composed similar to the maintenance ration of fodder that the cows had had in their original stock. The maintenance ration of fodder consisted of hay, grass ensilage, rich food and concentrate. The cows were collared in long stalls with rubber mat and cutter shavings as a litter bed.

[0056] Five cows were in the first lactation and one cow was in its third lactation. The stage of lactation varied between the second to the seventh lactation month. The yield of milk in the start of the experiments varied between 28-48 kgs ECM. The udder health was documented to be good before the start of the experiments, with negative cultivation tests and CMT-values of 1 and 2. However, coagulase negative staphylococci grew in one udder section (forward left) on one cow, together with increased cell contents (CMT 3), in view of which this udder section was excluded from the experiment.

[0057] A teat dipping solution containing (percent by weight) 1% chitosan, 0.02 dextran sulphate, 0.1% methyl cellulose and 0.5% glycerol was prepared.

[0058] Milking was carried out at 0730 and at 1530. Milk samples were taken directly after each milking situation for bacterial growth and analysis of the cell contents.

[0059] The cell contents were determined with a fluorooptoelectronic method (Fossomatic 90, A/S N. Foss Electric Denmark). The bacteriological analysis was carried out by smearing 10 μl of the milk sample on to a bovine blood agar plate with esculine added and was incubated at 3-7° C. for two days. Recordings were made after 24 and 48 hours and any mastitis pathogenic bacteria were isolated and identified. In connection with bacterial growth also routine CMT-analysis was made (California Mastit Test)

[0060] The study was carried out in three experimental rounds. In the first experimental round the animals were divided up so that three cows with together 12 udder sections were teat-dipped diagonally right forward and left rear, and three cows were teat-dipped reversed diagonally, i.e. left forward and right rear (see Table 5). The other 11 udder sections were left untreated as controls.

[0061] To the second experimental round four of the cows having 10 udder sections not infected in the first experiment were used. The udder sections were randomly distributed so that five were treated with the new teat dipping agent and five constituted controls. Two cows contributed with 3 udder sections each and two udder sections each and the two others with two udder sections each, see Table 5.

[0062] For the third experimental round four of the cows were used with seven udder sections which had not become infected during the previous experimental rounds. All 7 udder sections were treated with commercial teat dipping agent containing iodine. Two cows contributed with one udder section each and the two others with 2 and 3 udder sections, respectively, each, see Table 8.

[0063] In order to make the udder sections more sensitive to infection during the experiment keratin was removed from the teat canal on all udder sections. This was done with a darning-needle (John James & Sons, Ltd, Studley, Warwickshire, size 16). Keratin was removed from all teat canals after each morning milking.

[0064] In the first experimental round the keratin removal was terminated day 8. For two cows which up to day 7 had not become infected keratin removal was carried out both morning and night day 7 and 8.

[0065] In the second experimental round the keratin removal was terminated day 6, here double keratin removals did not have to be performed in order to infect the cows.

[0066] In the third experimental round the keratin removal was terminated day 8. Keratin removal was carried out both morning and night day 7 and 8.

[0067] Ten minutes after the teat dipping procedure all teats were dipped for 10 seconds in a bacteria suspension, containing about 1-7×107 CFU/ml of a β-hemolysing Streptococcus agalactiae strain (S-B8).

[0068] The criterium for infection of an udder section was two subsequent milk samples having bacterial growth or increased cell contents. A cow with symptom for acute clinical mastitis was treated with antibiotic and increased milking frequency.

[0069] Experimental schedule Morning milking

[0070] 1. After milking the teats were cleaned with 70% ethanol. The ethanol was allowed to evaporate and milk samples were then taken in test tubes from each udder quarter.

[0071] 2. The teats were then carefully washed with 70% ethanol. A sterile darning-needle was inserted into the teat canal, rotated and then withdrawn. All teats were treated in this manner.

[0072] 3. The pre-determined teats were immediately after point 2 dipped with teat dipping agent. The control teats were left untreated.

[0073] 4. After 10 minutes all teats were dipped for 10 seconds in the bacterium suspension with 1-7×107 CFU/ml of 8-hemolysing Streptococcus agalactiae.

[0074] Night milking

[0075] 1. After milking the teats were cleaned with 70% ethanol. The ethanol was allowed to evaporate and milk samples were then taken in test tubes from each udder quarter.

[0076] 2. The predetermined teats were dipped immediately after point 1 with teat dipping agent. The control teats were left untreated.

[0077] 3. After 10 minutes all teats were dipped in bacterium suspension for 10 seconds. Statistic evaluation has been made by means of x2-analysis according to the following formula.

\sum \frac{{(O - E)}^{2}}{E}

[0078] and 95% confidence interval was calculated by means of the formula

(p1−p2)±1,96×se(p1−p2)

[0079] wherein se(p1−p2) has been calculated as

\sqrt{\frac{p_{1} (1 - p_{1})}{n_{1}} + \frac{p_{2} (1 - p_{2})}{n_{2}}}

[0080] se standard error

[0081] n1=number of infected non-treated udder sections

[0082] n2=number of infected treated udder sections

[0083] p1=number of non-treated udder sections/n1

[0084] p2=number of treated udder sections/n2

[0085] Results

[0086] In total 14 of the 33 udder sections were infected during the two experimental rounds, of these were 12 not teat dipped and two were teat dipped. All cows in experimental rounds 1 and 2 became infected. In experimental round 3 two of the cows became infected.

[0087] The number of days between start of experiments and up to infection of udder sections varied between 2 and 9 days.

3TABLE 3The number of infected udder sections after treat-ment with teat dipping agent compared with no teat dip-ping (control). Experimental rounds 1 and 2 are assembledto one table.TreatedControlTotalInfected21214Not infected14519Total161733

[0088]

4

TABLE 4

The number of infected udder sections after treat-

ment with commercial teat dipping agent. Experimental

round 3.

Treated

Infected
2

Not infec
5

Total
7

[0089] Experimental round 1

[0090] Of the control udder sections 7/12 were infected and of teat dipped udder sections 1/11 were infected.

[0091] Five cows were infected with Streptococcus agalactiae in one udder section, the udder sections infected were controls.

[0092] One cow (No. 335) was infected in 3 udder sections, 2 control udder sections and 1 teat dipped udder section. The udder section of cow 335 which was strongly infected and reacted with the highest cell contents in the milk was one control section, see tables 5 and 6.

[0093] Experimental round 2

[0094] Out of the control udder sections 5/5 were infected and out of teat dipped udder sections 1/5 were infected, see table 7.

[0095] Cow 334 was infected in all 3 udder sections part-taking in the experiment, 2 udder sections were controls and 1 udder section was treated. It was the treated udder section that became most infected and responded with a strong inflammatory reaction in the form of swelling, soreness and increased cell contents.

[0096] The other three cows in the experiment were infected in one control udder section each, see table 7.

[0097] Experimental round 3

[0098] 10 Out of the 7 udder sections treated with commercial iodine teat dipping agent, 2 udder sections were infected, see Table 8.

[0099] The result of the %2-analysis for the assembled experimental results from rounds 1 and 2 gave an x2-value of 11.39, which corresponds to a probability value of <0.1% at one degree of liberty. I.e., the probability to obtain such a deviating value as 11.39, if ho (the zero hypothesis) starts from the assumption that there is no difference between treated and untreated group, is less than 0.1%.

[0100] When calculating 95% confidence interval for the difference in proportion (p1×p2) between the two experimental groups the result was 0.581±1.96×0.138, i.e. between 0.311 and 0.851.

[0101] Only 7 udder sections were part of experimental round 3. These were constituted by udder sections resisting infection for the longest period of time and were therefore the result of a selection. In view of this no statistic evaluation of this experimental round was made.

[0102] The infection frequency of the different experimental rounds is summarized in FIG. 3.

[0103] Discussion

[0104] All cows originated from the same live stock and had thereby the same immunological background. This was an advantage since one could expect that the cows would react in essentially the same manner during the readjustment period to the new environment including new maintenance ration of fodder. This resulted, together with the maintenance ration of fodder that was similar to that the cows were used to, in a situation whereby the time for buying the cows to the start of the experiments could be kept short (by 10 days).

[0105]

Streptococcus agalactiae
was selected in order that the animals with high probability would not have been subjected to this bacterium earlier, it is easy to find in cultivation (β-hemolysing), it is strictly udder specific and it is documented sensitive to penicillin.

[0106] The cows which were infected in 1 udder section in experimental rounds 1 and 2 became so in the untreated udder section, whereas the cows receiving infection in teat dipped udder sections became simultaneously infected in untreated udder sections.

[0107] In order to increase the infection risk of two cows which in the first experimental round were not infected up to day 7, the removal of keratin was extended to two times a day. The rate of generation of teat canal keratin is high (12-36h) and probably varies between individuals in view of which they have different sensibility for similar challenges in this context. Also the quantity of keratin, the majority of the keratin and composition thereof can vary between individuals.

[0108] It can be noted that animals infected late in the first experiment were also infected late in the second experiment.

5TABLE 5Analysis results of cell contents, CMT and bacterialgrowth in cows Nos. 306, 331 and 333 from the first experimentround. Cows Nos. 306 and 331 were teat dipped on right rearand left forward teat, respectively, cow No. 333 was teatdipped on right forward and left rear teat, respectively.Day23242526272829Time ofCow #exp.ampmampmampmampmampmampmampm306CellHf973041731931071356515010713880172102261cont.•Hb158331208221237303128179193245124143108220Vb20721615822012516592887220590160231459599441021135277958•Vf10992133764299620082679105228121970714980134711342238396175CMTHf23222211222213•Hb22221211222223Vb12122455555545•Vf33344435555544Bact.Hfgrowth•HbVbSrSrSrSrE SrEE•VfSSSSrSrSrSrSr331CellHf721265310160936212884111639931102cont.•Hb41604968466654825770668934108Vb304226454556661141941591261941259922497•Vf46574274486744126478250913884CMTHf12121211211212•Hb11121111111112Vb11111211221255•Vf12232211111113Bact.Hfgrowth•HbVbSrSr ESr N•Vf333Cell•Hf9238717319812013113116981108861524596contHb95386194181155175110158531045410548114•Vb1696542042511952831201791291498014379130Vf11373925526715625213829780142851315992CMT•Hf23332211121211Hb13333211121211•Vb23333211221223Vf1333221221111Bact.•HfgrowthHb•VbVfDay3012Time ofCow #exp.ampmampmampm306CellHf1192058819236100cont.•Hb149217116173153214Vb262540801379294116081692•Vf27242793140219579351410CMTHf221211•Hb222111Vb343222•Vf343112Bact.Hfgrowth•HbVb•VfSr331CellHf102118851185077cont.•Hb1362461391665864Vb238283255641565254502189813532•Vf961671381876762CMTHf111111•Hb211111Vb555555•Vf221111Bact.Hfgrowth•HbVbSr NSrN SrN SrSrE•Vf333Cell•Hf7896487869125cont.Hb11213313250993987830290•Vb255160122242165305Vf12811558144127175CMT•Hf111111Hb111555•Vb222211Vf211111Bact.•HfgrowthHbSr NSr NSr NSr•VbVfam = morning pm = afternoon Hf = right forward teat Hb = right rear teat Vb = left rear teat Vf = left forward teat Cell count = counted in 1000; CMT = California Mastitis Test; Bact. growth, Sr = Streptococcus agalactiae, S = Staphylococcus spp.; E = Ethacilin; • = teat-dipped; N = Novocillin

[0109]

6

TABLE 6

Analysis results of cell contents, CMT and bacterial

growth in cow Nos. 334, 335 and 338 from the first experiment

round. Cow Nos. 306 and 335 were teat dipped on right forward

and left rear teat, respectively, cow No. 338 was teat dipped

on right rear and left forward teat, respectively.

Day

23

24

25

26

27

28

29

Time of

Cow #
exp.

am
pm
am
pm
am
pm
am
pm
am
pm
am
pm
am
pm

334
Cell
•Hf
191
144
156
187
126
72
80
106
66
47
105
105
103
60

cont.
Hb
83
87
97
98
63
65
56
78
80
73
67
106
81
108

•Vb
108
114
109
107
111
84
65
78
91
79
134
74
83
126

Vf
117
164
130
143
16959
43705
44471
40411
35281
22651
23493
18960
16792
17835

CMT
•Hf
1
2
2
2
1
1
1
1
1
1
2
2
2
1

Hb
1
3
2
2
1
1
1
1
1
1
1
1
1
1

•Vb
1
3
2
2
1
1
1
1
2
1
1
1
1
1

Vf
1
2
1
1
5
5
5
5
5
5
5
5
5
5

Bact.
•Hf

growth
Hb

Sr

•Vb

Vf

E
Sr
Sr E
Sr
Sr E
Sr

335
Cell
•Hf
87
86
64
83
318
1029
4187
4733
3369
2980
2289
2139
1032
1398

cont
Hb
100
101
58
96
14956
35956
45703
37195
32044
17631
9923
10081
6821
7330

•Vb
68
94
42
51
60
159
76
111
97
99
56
80
50
68

Vf
80
75
38
67
688
1037
2204
2314
2061
2321
1353
2123
804
962

CMT
•Hf
1
2
1
1
3
4
5
5
4
5
4
3
3
3

Hb
1
1
1
1
5
5
5
5
5
5
5
5
5
4

•Vb
1
1
1
1
1
1
1
1
2
2
1
1
1
2

Vf
1
1
1
1
3
4
4
4
3
5
3
3
3
3

Bact.
•Hf

Sr
Sr

growth
Hb

Sr E
Sr
Sr E
Sr
Sr
E

•Vb

Vf

Sr
Sr

338
Cell
Hf
33
83
82
74
33
58
26
69
31
40
29
48
64
132

cont.
•Hb
193
130
77
118
49
65
21
74
97
73
43
80
68
99

Vb
146
90
69
146
112
150
62
130
49
105
44
98
133
121

•Vf
53
87
66
82
424
1685
2935
2005
1445
1348
650
994
395
392

CMT
Hf
1
1
1
1
1
1
1
1
1
1
1
1
1
3

•Hb
1
1
1
1
1
1
1
1
1
1
1
1
1
3

Vb
1
1
1
1
2
1
1
1
1
1
1
1
2
1

•Vf
1
1
1
1
3
4
4
4
3
3
2
3
2
2

Bact.
Hf

growth
•Hb

Vb

•Vf
S

S
S

Day

30

1

2

Time of

Cow #
exp.

am
pm
am
pm
am
pm

334
Cell
•Hf
106
110
124
71
82
151

cont.
Hb
85
97
78
49
63
88

•Vb
106
73
81
105
65
75

Vf
8841
13602
4894
6013
1821
6099

CMT
•Hf
1
1
1
1
1
1

Hb
1
1
1
1
1
1

•Vb
1
1
1
1
1
1

Vf
5
5
4
3
3
3

Bact.
•Hf

growth
Hb

•Vb
S

Vf

335
Cell
•Hf
749
1325
675
964
483
10731

cont
Hb
4418
6865
3245
6150
2186
2855

•Vb
60
77
73
82
66
70

Vf
530
921
869
22227
19869
16789

CMT
•Hf
3
3
3
3
3
5

Hb
4
4
4
4
3
3

•Vb
1
1
1
2
1
1

Vf
3
3
3
5
5
5

Bact.
•Hf

Sr
Sr (2)

growth
Hb

E

•Vb

Vf

Sr
Sr
Sr
Sr

338
Cell
Hf
38
75
88
8862
30894
17377

cont.
•Hb
128
153
198
204
136
209

Vb
94
136
91
138
352
299

•Vf
257
501
387
692
566
621

CMT
Hf
1
1
1
5
5
5

•Hb
1
1
1
1
1
1

Vb
2
2
1
1
1
1

•Vf
3
2
3
2
2
3

Bact.
Hf

Sr
Sr
Sr E
Sr

growth
•Hb

Vb

S

•Vf
S

S

am = morning

pm = afternoon

Hf = right forward teat

Hb = right rear teat

Vb = left rear teat

Vf = left forward teat

Cell cont = counted in 1000;

CMT = California Mastitis Test;

Bact. growth, Sr = Streptococcus agalactiae, S = Staphylococcus spp.;

E = Ethacilin;

• = teat-dipped;

N = Novocillin

[0110]

7

TABLE 7

Analysis results of cell contents, CMT and bacterial

growth in cow Nos. 306, 331, 334 and 338 from the second ex-

periment round. Cow No. 306 was teat dipped on right forward

teat, 331 was teat dipped on right forward and right rear

teat, respectively, 334 and 338 were teat dipped on right rear

teat.

Day

13

14

15

16

17

18

19

Time of

Cow #
exp.

am
pm
am
pm
am
pm
am
pm
am
pm
am
pm
am
pm

306
Cell
•Hf
89
160
141
235
91
131
129
143
108
95

cont.
Hb
78
131
127
123
31822
38666
39981
16350
19969
12399

CMT
•Hf
2
3
2
3
2
2
2
2
2
2

Hb
2
3
1
2
3
5
5
5
5
5

Bact.
•Hf

growth
Hb

E
Sr
Sr E
Sr

E

E

E

331
Cell
•Hf
58
112
76
72
69
97
46
88
56
95
62
96
148
6

cont
•Hb
75
109
92
74
73
75
69
1011
68
133
133
130
211
174

Vf
50
67
65
55
51
39
33
48
41
50
375
26511
44130
35022

CMT
•Hf
1
3
1
1
2
2
1
1
2
2
2
1
2
1

•Hb
2
2
1
1
3
2
1
1
2
2
1
2
2
3

Vf
1
1
1
2
1
1
1
1
1
1
4
5
5
5

Bact.
•Hf

growth
•Hb

Vf

Sr E
Sr
Sr E
Sr

334
Cell
Hf
116
149
113
159
108
143
144
174
112

cont
•Hb
47
62
56
71
6870
26446
43123
36795
27753

Vb
47
62
86
333
2686
2550
1459
1561
1816

CMT
Hf
1
1
1
2
2
3
1
1
1

•Hb
1
1
1
1
4
5
5
5
5

Vb
1
1
1
3
3
3
3
3
3

Bact.
Hf

Sr
Sr

growth
•Hb

E
Sr
Sr E
Sr
Sr
E

E

E

Vb

S
Sr

338
Cell
•Hb
50
135
29
64
44
56
28
34
33
47
39
56
110
145

cont.
Vb
32
88
26
35
26
57
18
43
21
47
35
9548
29784
25099

CMT
•Hb
1
1
1
1
1
1
1
1
1
1
1
1
3
3

Vb
1
1
1
1
1
1
1
1
1
1
1
5
5
5

Bact.
•Hb

growth
Vb

Sr E
Sr
Sr E
Sr

am = morning

pm = afternoon

Hf = right forward teat

Hb = right rear teat

Vb = left rear teat

Vf = left forward teat

Cell cont = counted in 1000;

CMT = California Mastitis Test;

Bact. growth, Sr = Streptococcus agalactiae;

S = Staphylococcus spp.;

E = Ethacilin;

• = teat-dipped;

N = Novocillin

[0111]

8

TABLE 8

Analysis results of cell contents, CMT and bacterial

growth in cow Nos. 306, 331, 333 and 335 from the third ex-

periment round with commercial teat dipping composition. Cow

No. 306 was teat dipped on right forward teat, cow No. 331 on

right forward and right rear teat, respectively, cow No. 333

on right forward, left rear teat and left forward teat, re-

spectively, and cow No. 335 on left rear teat..

Day

28

29

30

31

1

2

3

4

Time of

Cow #
exp.

am
pm
am
pm
am
pm
am
pm
am
pm
pm
am
pm
am
pm
am

306
Cell
Hf
96
144
107
155
57
132
114
114
125
152
62
66
129
89
115
104

cont.

CMT
Hf
1
1
1
1
1
1
1
1
1
1
1
1
2
1
2
2

Bact.
Hf

Sr

growth

331
Cell
Hf
42
64
82
153
59
125
58
86
53
158
72
88
248
129
224
121

cont
Hb
51
67
86
109
86
113
74
91
66
107
100
61
114
101
143
107

CMT
Hf
2
1
1
1
1
1
1
2
1
1
1
2
2
2
2
2

Hb
2
1
1
1
1
2
1
2
1
1
1
2
2
2
3
2

Bact.
Hf

growth
Hb

333
Cell
Hf
49
95
34
104
44
139
103
159
107
199
250
93
217
134
207
192

cont
Vb
73
150
85
131
73
115
91
114
44
83
83
72
135
141
208
164

Vf
81
133
82
195
68
195
119
156
106
184
167
146
263
172
278
198

CMT
Hf
1
2
1
1
1
3
2
3
1
2
2
2
2
3
3
2

Vb
2
2
3
1
3
2
1
3
1
2
2
2
2
3
3
2

Vf
3
2
1
1
3
3
2
3
2
3
2
2
3
3
1
2

Bact.
Hf

growth
Vb

Vf

335
Cell
Vb
115
133
118
144
106
144
225
333
265
456
316
434
605
251
280
240

cont.

CMT
Vb
3
2
3
1
3
3
2
3
2
3
2
2
3
3
1
2

Bact.
Vb

Sr

growth

Day

5

6

Time of

Cow #
exp.

pm
am
pm

306
Cell
Hf
12055
38455

cont.

CMT
Hf
5
5

Bact.
Hf
Sr
Sr

growth

331
Cell
Hf
208
148
271
143

cont
Hb
187
114
136
118

CMT
Hf
1
2
3

Hb
2
2
2

Bact.
Hf

growth
Hb

333
Cell
Hf
258
179
326
195

cont
Vb
288
118
147
81

Vf
278
137
237
94

CMT
Hf
3
2
3

Vb
2
2
2

Vf
2
2
2

Bact.
Hf

growth
Vb

Vf

335
Cell
Vb
7997
23298

cont.

CMT
Vb
5
5

Bact.
Vb
Sr
Sr

growth

am = morning

pm = afternoon

Hf = right forward teat

Hb = right rear teat

Vb = left rear teat

Vf = left forward teat

Cell cont = counted in 1000;

CMT = California Mastitis Test;

Bact. growth, Sr = Streptococcus agalactiae, S = Staphylococcus spp.;

E = Ethacilin;

TEAT DIPPING AGENT

Information

Publication Number

Date Filed

Date Published

Inventors

CPC

US Classifications

International Classifications

Abstract

Description

Claims

Priority Claims (1)

PCT Information