MASTITIS: A DEVASTATING DISEASE OF DAIRY ANIMAL

 

S.K. Rajak^1*, A.K. Singh² and P.K. Bharti³

1-Subject Matter Specialist (Veterinary Science), DrRPCAU, Pusa, Samastipur

2 – Assistant Professor (Animal Nutrition), FVAS, RGSC BHU, Varanasi

3- Scientist SS (LPM), ICAR-Mahatma Gandhi Integrated Farming Research Institute, Motihari.

*Corrosponding author email Id- shailendra06rajak@gmail.com

 

Mastitis is one of the most economically distressing diseases of dairy cattle. It is defined as the inflammation of the udder which may result due to microbial, thermal or physical causes. The udder becomes inflamed and bacteria invade the teat canal and mammary glands. These bacteria multiply and produce toxins that cause injury to the milk secreting tissue which cause increase in the number of leukocytes or somatic cells in the milk, reducing its quantity and adversely affecting the quality of milk and milk byproducts. Affected quarters show 30% less productivity and cow drops about 15% production. Prevention and control of mastitis are warranted in the post WTO scenario, to export good quality dairy products. Mastitis prevention and control is one of the top challenges for dairy farmers in India. Several attempts and procedures are being used all over the world not only in India to get mastitis free clean milk production. Various research and development agencies, government, private sector agencies, co-operatives and farmers are increasingly paying attention to mastitis management for clean milk production.

MASTITIS

Mastitis (Mast: breast, itis: inflammation) is an inflammatory condition of the udder, which may be due to microbial, thermal or physical causes. It is the most common and most expensive disease affecting dairy cattle throughout the world. Mastitis is caused by various bacteria that can invade the udder, multiply there, and produce harmful substances that result in inflammation. Mastitis reduces milk yield and alter milk composition (Ibrahim, 2017).

Causative agents of mastitis

Mastitis is one of the major health concern in dairy production sytem across the world. The predominant causal organisms are cell-walled pathogens, although mycoplasma, yeast and algae have also been reported to cause mastitis (Barkema et al., 1998). Interestingly, about 137 species and subspecies of potential pathogens can be found linked for the occurrence of infection of the udder. Further, mastitis in dairy herds is generally of two types: contagious mastitis and environmental mastitis (Bengtssm et al., 2009 and Persson et al., 2011).

Contagious Mastitis: Contagious mastitis is caused primarily by Staphylococcus aureus and Streptococcus agalactiae, Mycoplasma bovis and other Mycoplasma species. Characteristic feature of contagious mastitis are (1) a high prevalence of intra-mammary infection (IMI) during lactation, (2) a high Somatic Cell Count (3) infections of long duration, (4) low proportion of infections resulting in clinical mastitis (infections mostly subclinical) and (5) a low prevalence of infection during the dry period.

Contagious mastitis can be divided into three types:

Clinical mastitis:  Clinical mastitis is characterized by sudden onset, swelling, pain, redness of the udder, reduced and altered milk secretion from the affected quarters. The milk may have clots, flakes or of watery inconsistency and accompanied by fever, depression and anorexia.

Clinical mastitis can be divided into three types:

1. Per-acute mastitis: It is characterized by gross inflammation, reduction in milk yield and changes in milk composition. The sign and symptoms of this condition is characterized by systemic signs like fever, depression, shivering and loss of appetite and loss of weight.
2. Acute mastitis: Similar to per-cute mastitis, but with lesser systemic signs like fever and mild depression.

    iii. Sub-acute mastitis: In this, the mammary gland inflammation signs are minimal and no visible systemic signs.

Subclinical mastitis: The subclinical mastitis is characterized by having no visible signs either in the udder or in the milk, but the milk production decreases and the SCC increases. It  has greater impact on older milch animals than the animal in first lactation. The incidence of  subclinical mastitis is 15 to 40 times more than the clinical form. It adversely affects milk quality and production because of longer duration, difficult to detect and constitutes a reservoir of microorganisms that lead to chronic mastitis condition and infection of other animals within the herd.

Chronic mastitis: An inflammatory process that exists for months and may continue from lactation to lactation. It exists as subclinical but may exhibit periodical flare-ups sub-acute or acute form, which last for a short period of time.

Environmental Mastitis Pathogens: It is primarily caused by environmental streptococci including Streptococcus uberis, Streptococcus dysgalactiae subsp. dysgalactiae, and coliforms including Escherichia coli and Klebsiella species (Hogan and Smith, 1987). These organisms which are absent normally on the skin or in the udder but when enter the teat canal after cow comes in contact with a contaminated environment. The pathogens normally found in dung, bedding materials and feed. Of the total mastistis cases only 10% are occurred due to environmental mastitis in the herd. Infections generally occur between milking and during the milking process. Characteristic features of environmental mastitis are- (1) a low prevalence of Intra Mammary Infection during lactation, (2) a low SCC, (3) infections of short duration, (4) many Intra Mammary Infection resulting in clinical mastitis,  (5) a high prevalence of infection during the dry period.

 

Some other important etiological agent of mastitis is-

Virus:  Vesicular Stomatitis, Infectious bovine Rhinotrachitis, Bovine herpes virus etc.

Fungus:   Tricosporium spp, Aspergillus spp, Candida spp etc.

Mycoplasmas : M. bovis, M. californicum, M. canadense, M. bovigenitalium, M. alkalescens, M. arginini, M. bovihirnis, M. dispar, bovine group 7, and F-38

Algae: Prototheca sps. (Anderson and Walker, 1998)

Yeast: Cryptococcus neoformans and Candida albicans (Yeh et al., 1998)

 

ECONOMIC IMPACT OF MASTITIS

Mastitis is known as one of the most expensive disease affecting dairy industry. It was estimated that mastitis reduces milk by 21% and butter fat by 25% (Blood and Rodostits, 1989 and Dua, 2001). Losses due to mastitis milk in terms of abnormal characteristic and higher cost of veterinary aid is a barrier in development of dairy industry. In spite of economic losses, presence of different bacteria may be harmful to human consumption as well. Thus, in addition to its economic effect, bovine mastitis is important from the view of public health consideration. The economic losses are huge due to unmarketable milk or milk-products contaminated with antibiotic residues originating from treatment in the developing nations as well as from the use of antibiotics as growth promoters in dairy feedlots particularly in the developed world. The prolonged use of antibiotics in the treatment of mastitis has led to the additional problem like emergence of antibiotic resistant strains and entering the food chain (White and Mc Dermont, 2001). Many organisms associated with mastitis also have zoonotic importance and can cause diseases like Brucellosis, Tuberculosis, Leptospirosis, Q-fever etc. (Sharif and Umer, 2009). The losses are not only economic, but other issues such as animal health and welfare, milk quality, antibiotic usage and the image of the dairy sector are important reasons to focus on mastitis control.

FACTORS AFFECTING THE SUSCEPTIBILITY TO MASTITIS

The large number of predisposing factors may contribute to the emergence of mastitis in dairy cattle like physiological, genetic, pathological or environmental as mentioned below:

Age of the cow

It has been reported that occurrence of mastitis in infected quarters increases with advancing age in cows and was observed to be highest at the age of 7 years (Sachukken et al., 1989). This may be due to an increased cellular response to intra-mammary infection or due to permanent udder tissue damage resulting from the primary infection. Efficient innate host defense mechanisms of the younger animals are one possibility that makes them less susceptible to infection (Dullin et al., 1988).

Inherited features of the bovine

Various genetic traits may also have a considerable impact upon the susceptibility of the animal to mastitis. These genetic traits include the natural resistance, teat shape and conformation, positioning of udders, relative distance between teats, milk yield and fat content of milk. High milk yielders with higher fat content are reported to be more susceptible to mastitis (Grohn et al., 1990). The conformation of the udder and shape of the teat are inherited characteristics that may also affect susceptibility to mastitis.

Stage of lactation

The incidence of mastitis is reported to be higher immediately after parturition, early lactation and during the dry period, especially the first 2-3 weeks probably due to increased oxidative stress and reduced antioxidant defense mechanisms during early lactation (Sharma et al., 2011)

Mammary regression

There are significant functional changes in the udder during the early and late lactation and dry period, which increases the cow’s susceptibility to infections. Lactating cows under stress show premature mammary regression which compromises udder’s natural defense mechanisms leading to invasion of the teat canals by potential pathogens (Giesecke et al., 1994). The same condition prevails during the healing process of lesions because the resistance to causal agents remains less effective (Sordillo, 2005).

Milking machine

Extraneous factors such as the milking habits of farmers and faulty milking machines favour the pathogens for entry into mammary gland and proliferate potentially leading to mastitis. In farms where machines are used for milking needs to maintain physiologically optimal pressure otherwise excess pressure may lead to injury to teat (Blood and Rodostits, 1989). Proper installation as well as the correct maintenance of milking machines is important to avoid an inadequate vacuum level, teat and tissue damage and incomplete milking (International Dairy Federation, 1987).

Plane of Nutrition

The quality and plane of nutrition appears to be an important factor that influences clinical manifestation of mastitis in heifers and cows, although no direct relationship has been reported between the incidence of mastitis and feeding  either high energy or high protein feed in cows.

Vitamin E is one of the important supplements in dairy feed to boost the immune response of cows as it has been reported to enhance the neutrophil function as well as the phagocytic properties of neutrophils after parturition (Spears and Weiss, 2008). Vitamin E in symbiotic relationship with selenium acts as an anti-oxidant by preventing oxidative stress. Various studies has led to the conclusion that selenium when added to feed of cows are more effective at killing mastitis causing microorganisms ans incidences of mastitis (Grasso et al., 1990).

Beta-carotene and Vitamin A have also been found to be effective in preventing the occurrence of mastitis, which may be due to their antioxidant and immune-enhancing properties that contribute to enhance the mucosal surface integrity of the mammary gland (Sordillo et al., 1997). Zinc and copper are also reported to contribute mammary gland health by promoting cellular repair, wound healing and reduction in SCC (Scaletti et al., 2003).Various studies demonstrated that feed supplemented with copper when fed to heifers reduces the severity of subclinical mastitis as well as clinical mastitis induced by Escherichia coli (Upadhayay et al., 2008).

Weather and climate

The incidence of mastitis is greatly influenced by the weather conditions and prevailing climatic conditions. Heat, humidity, cold and draught are the important predisposing factors as stress lowers the immunity in the animals. A higher incidence of mastitis has been reported to occur particularly during summer and rainy months, as heat and humidity increases, the bacterial multiplication as well as the load of pathogens in the environment (Godden et al., 2003). However, Ranjan et al (2011) has reported a higher incidence of coliform mastitis during the cold months when the temperature was less than 21°C.

DIAGNOSIS OF BOVINE MASTITIS

Healthy udders are economically profitable because of better quality product and better cow health. Monitoring of unhealthy udder in dairy cows starts with visual observation. Abnormal milk, even the smallest clot is a signal that something is wrong within that quarter and requires immediate attention and needful action. Monitoring udder health is very difficult without reliable and affordable diagnostic methods. Thus, there is need for improvement for the accuracy, cost price or convenience and availability of these methods.

1. Somatic cell count (SCC):- SCC has been known to be one of the most important indicator of intra mammary infections (Schukken et al., 2003; 2009). An easy, cheap, and quick platform test to estimate SCC on farm is the California Mastitis Test (CMT). This method is based on a semi-quantitative SCC measure by forming a stringy mass from the reagent (3% sodium lauryl sulphate) and DNA out of disrupted cells. Although the CMT test is not very complicated but its execuation accuracy is always debateable.
2. Bacteriological culturing (BC):- Bacteriological culturing is mostly used as a diagnostic tool to rule out the mastitis problems. It can be executed at herd, as well as cow and quarter level, each with its own specific goal. Further, BC results give herd-based information that can be helpful in optimizing the treatment of future mastitis cases. For effective use of bacteriological culturing as a diagnostic tool, milk samples have to be collected from the affected cows and quarters at the correct point of time.
3. Electrical Conductivity (EC) Test

EC has been used for detection of bovine mastitis on the phenotypic level. This test measures the increase in conductance in milk which may be caused by the elevation in levels of ions such as sodium, potassium, calcium, magnesium and chloride during inflammation. It can be used ‘on-site’. Non-mastitis-related variations in EC possess problems in diagnosis.

4. Biomarkers

This is rapid laboratory based assays to detect mastitis in dairy animals. Biomarkers are used to detect enzymes, such as Serum Amyloid A (SAA), Haptoglobin Lactate dehydrogenase, N-acetyl- β-d-glucosaminidase and Alkaline phosphatase. Each of these parameters is valuable in order to detect changes in the milk indicating mastitis. They are good in the accuracy but the cost of testing challenges its rapid use.

5. Proteomic Techniques

Advances in relevant proteomics techniques, such as two-dimensional gel electrophoresis (2D-GE) and mass spectroscopy (MS) have led to the identification of several new proteins involved in mastitis. They could possibly be used as markers for its detection of mastitis. The results showed that there is an up regulation of k-casein and a down regulation of cytochrome C oxidase and annexin V in animal tissues that are mastitis-infected (Yang et al., 2009).

6. Immunoassays

Numerous immunoassays have been developed for the detection of pathogens in milk for monitoring its quality (Duarte et al., 2015). More than one hundred known organisms can be responsible for causing mastitis but ELISAs have only been developed for some of the most prevalent pathogens, such as S. aureus, E. coli and L. monocytogenes. Multiplex PCR and ‘real-time’ PCR assays that can simultaneously detect different mastitis-causing organisms in milk samples have limited and the most recently developed assay is capable of detecting 11 of the major mastitis-associated pathogens, including E. coli, S. aureus, Streptococcus agalactiae and Streptococcus uberis (Koskinen et al., 2009).

7. Infra-Red thermography

Infra-red thermography (IRT) is a non-invasive method for measuring radiated heat emitted by the skin that reflects subcutaneous circulation and metabolism. Radiated heat emitted by the udder during clinical mastitis can be detected with IRT (Zaninelli et al., 2018). IRT technique did not require milk or milch animal, which provides a possibility for detecting mastitis during the dry period and before the first calving. Preliminary studies on the detection of clinical mastitis using IRT have shown promising results but IRT was not appropiate for detection of subclinical mastitis (Scott et al., 2000 and Polat et al., 2010).

8. Future developments in diagnosis

New developments such as the use of fuzzy logic and Bayesian Networks have been used to detect mastitis but to improve the efficacy of results information from other sources are necessary to distinguish healthy cows, cows with subclinical and cows with clinical mastitis before using this technique (De Mol and Woldt, 2001). The automation of the detection of mastitis with fuzzy logic models using traits with regard to performance (milk yield, water and dry matter intake) or behaviour (feeding) did not fit the available methods in practice and needs some more validation and modification to be fit for the dairy farmers and industry (Steeneveld et al., 2008).

CONTROL OF MASTITIS

The fundamental principle of mastitis control is, either decreasing the exposure of the potential pathogens or by increasing resistance of dairy cows teat/udder to infection. Controlling mastitis is involves many number of steps better referred to as a control programme. Success of such programme depends upon the economic, practicality and effectiveness under most management conditions and capable of reducing new infections. The programme should also be able to shorten the duration of pre-existing infections, reduce the incidence of clinical mastitis and be subject to easy modification. In attempting to control different types of infections, it is important to consider the source and means of transmission of the disease. Organisms that cause mastitis live in different environments (manure, bedding, skin, etc.). General cleanliness of cows and their housing as well as good management procedures especially at milking are effective ways of controlling the spread of mastitis. A key factor in preventing mastitis is to reduce the number of bacteria present around the teat end. This is particularly important in the transition period that will be about two weeks before calving to two weeks after.

The following points should be practiced to control the mastitis-

1. Preparation of animal

Clipping the hair on the udders, the flanks and inside the hind-legs should be done. Animal should be clean properly before milking.

2. Milk cows with clean, dry teats and teat ends-

The procedures of milking are very important for the prevention of mastitis and insuring complete milk removal from the udder.  Firstly the teats and teat ends should be washed with clean and fresh water and dried completely before the milk is taken by hand or machine. Emphasis should be placed on the teat ends. An additional positive step may be pre-dipping with a sanitizing solution similar to those used for post-milking teat dipping. Pre-dipping with teat dip has become popular.

 

3. Prevent transfer of pathogens from cow to cow during milking

During the milking process, preventing spread mastitis from one infected cow to non-infected cows by milking the non-infected cow first then infected cow. We should use towels, paper, cloth, wipes etc. in preparation of the udder and teats.

4. Prevent injury to the teats and udder during milking

Any injuries to the teats or teat ends and udder will eventually end up with a new case of mastitis. It is important to follow the proper milking techniques (attachment, alignment and removal of machines), proper milking machine design/function, routine, timely changing of inflations and continuous maintenance of the milking equipment (cleaning pulsators etc.). It is advisable to assess the teat end condition regualarly. Environmental sources of mastitis like bedding, housing, free stall design and maintenance should also be controlled.

5. Housing

Providing adequate space, ventilation, bedding, and lighting to ensure cleanliness and comfort at all times. Overcrowding should be avoided.

6. Early detection of new infections (clinical and subclinical) and treatment

Quick detection of mastitis will prohibit severe mastitis occurrences. Milking may begin with a checking of all quarters for mastitis. Any cows that show clinical mastitis should be examined and appropriate action should taken thereafter. Visual checking for inflamed quarters is done by milkers and herd health people regularly. Early detection  of mastitis may may be done by fore milking, observation of the udder and teats, California mastitis test, various forms of electronic somatic cell counting or electrical conductivity. Milker should be trained to use these techniques. Just after confirmation of mastitis, treatment with appropriate therapy should be started as early as possible.

7. Milking

Cows should be milk completely to prevent the occurrence of new cases. Standard milking techniques should be used like, apply milking units properly and make adjustments to prevent the admittance of air into the teat cup liners and prevent liner slip.

8. Provide adequate nutrition to preclude increased susceptibility to mastitis

The research finding indicated that nutrition is linked to mastitis in the dairy cow and it was found that negative energy balance and elevated serum beta-hydroxybutyrate have been concerned in reducing leukocyte functionality; thus increasing the risk of diseases like mastitis. Similarly, many trace minerals and vitamins have a direct effect on the immune system function and hence could have a direct effect on the ability of a cow to fight against  mastitis.

In addition, beta-carotene have anti-oxiant property and  helps in reducing superoxide formation within the phagocyte (Sordillo et al., 1997). Copper is another mineral associated in neutrophil production and affects phagocyte killing ability and is required for antibody development and lymphocyte replication.

The research evidence indicated that nutrition and feeding management  have a direct relationship with mastitis and we should more vigilant in formulating  the ration for heifer and older cows separately due to differences in nutrient requirements and metabolism. Feeding balanced diet to the heifer is very much  important for optimal immunity at or near calving. However, there is need of hour to study further to define the role of nutrition in animal health and specifically in mastitis.

Suggested levels of supplementation (amount/cow/day):

Selenium 6 mg (Lactating cows) , 3 mg (Dry cows) , Copper 200-250 mg ,Zinc 900-1200 mg Vitamin A 100,000 – 150,000 IU ,Vitamin E 500 IU (Lactating cows) ,1000 IU (Dry cows).

9. Fly control

Some flies spread the infection from infected cow to healthy one, particularly summer mastitis and other pathogen including S. aureus from one source to the teat ends of heifers or cows. The basic principle of fly control involves prevention of breeding sites through routine removal of manure and decaying feeds.

10. Vaccination

Various vaccines have been tried to prevent mastitis but it is clear now that a single vaccine will not prevent mastitis caused by the different pathogens and their different mechanisms of pathogenesis (Heath, 2011). Several efforts have been made to develop a vaccine against mastitis, but none have claimed satisfactory outcomes neither in the field nor on backyard farms (Lee et al., 1998). So vaccination is an important aspect in disease prevention but in mastitis vaccination is not much successful due to its multifactorial complex etiology. However, vaccine against major mastitis pathogens like Staphylococcus aureus, Streptococcus uberis, E. coli have been developed, but they are costly.

11. Dry cow therapy (DCT)

Dry cow therapy with antibiotics has been suggested as one of the options to control intra-mammary infections (IMI) and prevent development of mastitis. During the dry period, the cow is at the greatest risk of acquiring new intra-mammary infections with both gram-positive and gram-negative environmental or contagious pathogens. It has been reported that about 61% of new infections are acquired during this period (Todhunter et al.,1991). Treatment during dry periods is advantageous because it allows treatment of infections with antibiotics without the need to discard milk from treated quarters. Antibiotics are administered towards the end of lactation and may remain in the udder in concentrations high enough to kill pathogenic bacteria for 20-70 days (Todhunter et al., 1991). Dry cow therapy eliminated almost 100% of mastitis caused by S. agalactiae (Oliver et al., 1988). However, DCT is comparatively less successful to prevent S aureus mastitis than Streptococcal mastitis.

12. Lactation therapy

                 Treatment of mastitis during lactation with antibiotics is referred as “lactation therapy”. This therapy has proven useful in reducing the SCC in milk and thereby maintains the quality of milk (Sol et al., 1997). However, lactation therapy for subclinical mastitis is not suggested due to poor efficacy and non economic. Various factors viz.  Somatic cell count in milk during treatment, stage of lactation, immune status of the animal, age of the cow and type of pathogen also play an important role in the success or failure of lactation therapy.

13. Teat sealer

 The development of internal and external teat sealants for use during the dry period is a promising progress towards control of mastitis (Izak et al., 2012). However, lack of persistence is the main drawback of external teat sealers. Bismuth subnitrate as an internal teat sealer used in field conditions was reported to reduce new infections up to ten fold. Internal teat sealer used with long acting antibiotics during the dry period showed a 30% and 33% reduction in new intra-mammary infections and incidence of clinical mastitis, respectively.  Bismuth subnitrate combined with cloxacillin as dry cow therapy demonstrated reduction in both clinical and subclinical cases of mastitis (Runciman et al., 2010).

14. Genetic selection of cattle for resistance to clinical mastitis

Generally, breeding of farm animals has been confined to increase the milk production (Adams et al., 1998). But at the same time, the incidence of many infectious diseases increases, including bovine mastitis. To overcome this problem, a strategy based on enhancing the overall immune response- including antibody-mediated immune response (AMIR) and cell-mediated immune response (CMIR) has been proposed. However, a negative genetic correlation between AMIR and CMIR has been recorded, making a balanced genetic selection more complex and requiring further investigations (Adams et al.,1998).

TREATMENTS OF MASTITIS

Antibiotics

Antibiotics ranging from narrow to broad spectrum have been used extensively over the past 40 years in the control of bovine mastitis (Barkeman, 2006). However, because of the emerging antibiotic resistance due to their overuse and the induction of prolonged persistent antibiotic resistance in bio-films by many mastitis-causing pathogens, as demonstrated recently for S aureus isolated from cases of bovine mastitis effectiveness of antibiotic therapy has been compromised (Tiwari et al., 2013). The most commonly used antibiotics on conventional dairies were Penicillin (86%), Cephalosporin (78%) and Tetracyclines (41%). Conventional dairy herds used 98% of the intra-mammary dry cow antibiotic treatment while only 6.3% of the organic herds used intra-mammary dry cow therapy. The organic herds used non-antibiotics products for dry cow therapy (Gruet et al., 2001). Antibiotics can be used at two ihibit the infection in animals at two point of cycle: (a) To treat outbreaks of mastitis in milking cows as soon as they occur, and (b) To reduce subclinical infection during the dry period and hence increase the cow’s productive life (Barlow, 2011).

Bacteriophage therapy for mastitis associated infections

Specific bacteriophages were used in the treatment of mastitis caused by bacterial infection. Few studies have been carried out using bacteriophages to treat mastitis caused by S aureus infection have yielded variable results. Kwiatek et al. (2012) isolated and characterized a bacteriophage from the milk of cows suffering from mastitis with broad-spectrum activity against methicillin- resistant S aureus (MRSA). It requires additional research to explore the therapeutic potential of bacteriophages to treat clinical and subclinical mastitis.

 

Conclusions

There have been continuous shift in the predominance of etiology for mastitis. Better understanding of the host responses to intra-mammary infections and treatment regimens leading to adoption of various control and prevention measures. To control the mastitis effectively require knowledge from a wide range of fields, like infectious pressure, milking procedures, resistance, detection, diagnosis and treatment. However, present knowledge alone is not enough to ensure effective mastitis control. Further, research needs to be inspired by and implemented in dairy farming practice both at the industry and institutional level for the betterment and profit of the dairy sector. Thus, effective control of mastitis, needs cooperation and communication among scientists, veterinarians, extension specialists and dairy farmers worldwide.

References:

Adams, L.G. and Templeton, J.W (1998). Genetic resistance to bacterial diseases of animals. Rev Sci Tech 17: 200-219.

Anderson, K. L. and Walker, R.L (1988). Sources of Prototheca spp in a dairy herd environment. J. Am. Vet. Med. Assoc. 193:553

Barkema, H.W., Schukken,Y.H., Lam,T.J., Beiboer, M.L and  Wilmink, H (1998). Incidence of clinical mastitis in dairy herds grouped in three categories by bulk milk somatic cell counts, J Dairy Sci. 81: 411-41

Barkeman, H.W., Schukken,Y.H., Zadoks, R.N (2006). Invited Review: The role of cow, pathogen, and treatment regimen in the therapeutic success of bovine Staphylococcus aureus mastitis. J Dairy Sci. 89: 1877-1895.

Barlow, J (2011). Mastitis therapy and antimicrobial susceptibility: a multi species review with a focus on antibiotic treatment of mastitis in dairy cattle. Journal of mammary gland biology and neoplasia, 16 (4), 383-407.

Bengtsson, B., Unnerstad, H. E., Ekman, T., Artursson, K., Nilsson-Öst, M., & Waller, K. P. (2009). Antimicrobial susceptibility of udder pathogens from cases of acute clinical mastitis in dairy cows. Veterinary microbiology, 136 (1-2) :142-149.

Blood,D.C., Radostits, O.M (1989). Veterinary medicine. In: textbook of the diseases of cattle, sheep, pigs, goats and horses, (7^th edn), 501

De,Mol, R.M. and Woldt, W.E (2001). Application of fuzzy logic in automated cow status monitoring, J.of Dairy Sci.84, 400-410

Dua, K (2001). Incidence, etiology and estimated loss due to mastitis in India- An update-Indian Dairyman 53: 41-48

Duarte, C. M., Freitas, P. P., and Bexiga, R (2015). Technological advances in bovine mastitis diagnosis: an overview. Journal of veterinary diagnostic investigation, 27(6) :665-672.

Dulin,A.M., Paape.M.J., Nickerson, S.C (1988). Comparison of phagocytosis and chemiluminescence by blood and mammary gland neutrophils from multiparous and nulliparous cows,  American J. Vet. Res. 49: 172-177

Giesecke, W.H., Du Preez, J.H. and Petzer, I.M (1994). Practical Mastitis Control in Dairy Herds. Diagnosis of udder health problems, Butterworth Publishers: Durban, South Africa

Godden, S., Rapnicki, P., Stewart, S., Fetrow, J. and Johnson, A (2003). Effectiveness of an internal teat seal in the prevention of new intra mammary infections during the dry and early-lactation periods in dairy cows when used with a dry cow intra mammary antibiotic, J. Dairy Sci. 86: 3899-3911

Grasso, P.J., Scholz. R.W., Erskine,R. J. and Eberhart, R. J (1990). Phagocytosis, bactericidal activity, and oxidative metabolism of milk neutrophils from dairy cows fed selenium-supplemented and selenium-deficient diets, American J. Vet. Res. 51: 269-274

Grohn,Y.T., Erb, H.N., Mc.Culloch,C .E. and Saloniemei, H.S (1990). Epidemiology of mammary gland disorders in multiparous Finnish Ayrshire cows. Prev. Vet. Med. 8: 241-252

Gruet, P., Maincent, P., Berthelot, X., Kaltsatos, V (2001) Bovine mastitis and intra-mammary drug delivery: review and perspectives. Adv Drug Deliv Rev 50: 245-259.

Heath, P.T (2011) An update on vaccination against group B streptococcus. Expert Rev Vaccines 10: 685-694.

Hogan, J.S., Smith, K.L (1987). A practical look at environmental mastitis. Compendium on Continuing Education for the Practicing Veterinarian 9: F342.

Ibrahim, N (2017). Review on mastitis and its economic effect. Can. J. Sci. Res, 6 ;13-22.

International Dairy Federation (1987). Bovine mastitis: Definition and guidelines for diagnosis. Bulletin of the International Dairy Federation, Document 211

Izak,E., Castello, E., Veneranda, G (2012). Mastitis in Dairy: efficacy and economic benefit of an internal teat sealant during drought and rainy weather dry period on incidence of clinical mastitis.

Koskinen, M.T., Holopainen,J., Pyorala, S., Bredbacka, P. and Pitkala, A (2009). Analytical specificity and sensitivity of a real-time polymerase chain reaction assay for identification of bovine mastitis pathogens. J Dairy Sci. 92: 952-959

Kwiatek,M., Parasion, S., Mizak,L.,Gryko,R.,Bartoszcze, M (2012) Characterization of a bacteriophage, isolated from a cow with mastitis, that is lytic against Staphylococcus aureus strains. Arch. Virology 157: 225-234.

Lee,J.C., Perez,N.E.,Hopkins,C.A.,Pier,G.B (1988) Purified capsular polysaccharide-induced immunity to Staphylococcus aureus infection, J. Infect Dis. 157: 723-730.

Oliver, S.P., Lewis, M.J., Gillespie, B.E., Dowlen,H.H, Jaenicke, E.C (2003) Prepartum antibiotic treatment of heifers: milk production, milk quality and economic benefit. J Dairy Sci. 86: 1187-1193.

Persson, Y., Nyman, A. K. J., & Grönlund-Andersson, U. (2011). Etiology and antimicrobial susceptibility of udder pathogens from cases of subclinical mastitis in dairy cows in Sweden. Acta Veterinaria Scandinavica, 53(1): 36.

Polat, B., Colak, A., Cengiz, M., Yanmaz, L. E., Oral, H., Bastan, A. and Hayirli, A (2010). Sensitivity and specificity of infrared thermography in detection of subclinical mastitis in dairy cows. J Dairy Sci. 93(8): 3525-3532.

Ranjan,R., Gupta, M.K., Singh, K.K (2011) Study of bovine mastitis in different climatic conditions in Jharkhand, India. Vet. World 4: 205-208.

Runciman, D.J., Malmo, J., Deighton, M (2010). The use of an internal teat sealant in combination with cloxacillin dry cow therapy for the prevention of clinical and subclinical mastitis in seasonal calving dairy cows. J Dairy Sci., 93: 4582-4591.

Scaletti, R.W., Trammell, D.S., Smith, B.A., Harmon, R.J (2003) Role of dietary copper in enhancing resistance to Escherichia coli mastitis. J Dairy Sci., 86: 1240-1249

Schukken, Y.H., Grommers, F.J., Van de Geer, D., Brand, A (1989) Incidence of clinical mastitis on farms with low somatic cell counts in bulk milk, Vet. Record 125: 60-63

Schukken, Y.H., Hertl, J., Bar, D., Bennett, G.J., González, R.N (2009) Effects of repeated gram-positive and gram-negative clinical mastitis episodes on milk yield loss in Holstein dairy cows. J Dairy Sci., 92: 3091-3105.

Schukken, Y.H., Wilson, D. J., Welcome, F (2003). Monitoring udder health and milking quality using somatic cell counts. Veterinary Research 34: 579-596

Scott, S.L., Schaefer, A.L., Tong, A.K.W and Lacasse P (2000) Use of infrared thermography for early detection of mastitis in dairy cows. Can J Anim Sci. 80: 764-765

Sharif, A and Umer, M, Md. (2009) Mastitis control in dairy production, J Agril Soc Sci 5: 102-105

Sharma, N., Singh, N.K., Singh, O.P., Pandey,V. and Verma, P.K (2011). Oxidative stress and antioxidant status during transition period in dairy cows. Asian-Aust J Anim Sci. 24: 479-484

Sol, J., Sampimon, O.C., Snoep, J.J., Schukken, Y.H (1997). Factors associated with bacteriological cure during lactation after therapy for subclinical mastitis caused by Staphylococcus aureus. J Dairy Sci., 80: 2803-2808.

Sordillo, L.M (2005). Factors affecting mammary gland immunity and mastitis susceptibility. Liv. Prod. Sci. 98: 89-99

Sordillo, L.M, Shafer-Weaver, K. and De Rosa, D (1997). Immunobiology of the mammary gland. J Dairy Sci.,  80: 1851-1865

Spears, J.W., Weiss, W.P (2008) Role of antioxidants and trace elements in health and immunity of transition dairy cows. Veterinary Journal, 176: 70-76

Steeneveld, W., van der Gaag, L.C, Barkema, H.W (2008). Using cow-specific risks to support the detection of clinical mastitis on farms with an automatic milking system. Proceedings Mastitis Control 2008, The Hague, the Netherlands 370

Tiwari, J.G., Babra, C., Tiwari, H.K, Williams, V., Wet, S.D (2013). Trends In Therapeutic and Prevention Strategies for Management of Bovine Mastitis: An Overview. J Vaccines Vaccin 4: 176. doi:10.4172/2157-7560.1000176

Todhunter, D.A., Smith, K.L.,Hogan,J.S.,Schoenberger, P.S (1991). Gram-negative bacterial infections of the mammary gland in cows. Am J Vet Res 52: 184-188.

Upadhayay, A.K., Gangwar, P., Kumar, M (2008). Supplementation to prevent subclinical mastitis. Vet World 1: 40-41

White, D.G. and Mc Dermott, P.F (2001). Emergence and transfer of antibiotic resistance. Journal of dairy science, 84:E151-E5

Yang, Y. X., Zhao, X.X. and Zhang, Y (2009). Proteomic analysis of mammary tissues from healthy cows and clinical mastitic cows for identification of disease related proteins. Vet Res Commun 33: 295-303

Zaninelli, M., Redaelli, V., Luzi, F., Bronzo, V., Mitchell, M., Dell’Orto, V. and Savoini, G (2018). First evaluation of infrared thermography as a tool for the monitoring of udder health status in farms of dairy cows. Sensors, 18(3):862.