Control Tools

• Diagnostics availability

• Commercial diagnostic kits available worldwide

  A number of diagnostics tools/kits are commercially available. Availability may differ between countries and over time so an exhaustive list will not be provided here but rather, a broad classification by host, pathogen or immunological targets. Host level, detection of inflammation: indicators of inflammation (e.g. AMBIC vision mastitis detectors which, when installed in milk tubes, detect clots associated with clinical mastitis). California Mastitis Test (CMT) can be used to identify high SCC quarters. Sensors within AMS can notify of potential cases of mastitis. Pathogen detection methods: Microbiological methods: Bacteriological culture in dedicated diagnostic laboratories or in veterinary clinics. Selective and chromogenic media for in-clinic or on-farm culture and detection of mastitis pathogens, some of which can distinguish S. aureus and Streptococci, or streptococcal species, while others only indicate growth of gram-positive organisms. Nucleic acid amplification tests: Detection of nucleic acid of mastitis pathogens including S. aureus and Streptococci in milk samples, such as PCR and LAMP. Does not confirm the presence of live organisms. Multiplex PCR is available via professional diagnostic laboratories. Pathogen identification: Following pathogen detection, pathogen identification is routinely done by MALDI-ToF MS in professional veterinary diagnostic laboratories in some countries. Immunodiagnostics: detection of host antibodies or use of antibodies to detect the pathogen or its proteins, such as enterotoxins (e.g. the detection of S. aureus enterotoxins in milk e.g. Dairy Staphylococcus aureus Enterotoxin Typing ELISA Test Kit). Some tests may be ill-suited to small ruminants.

  GAPS

  Regulation for diagnostic tests. Rapid in-parlour tests. MALDI-ToF MS directly on milk samples.

• Diagnostic kits validated by International, European or National Standards

  There are diagnostic tests that conform to European standards or have USDA approval, but there are no international standards such as WOAH for diagnostic testing.

• Diagnostic method(s) described by International, European or National standards

  Standard bacteriological analysis of milk samples (plate count; focuses on milk quality rather than mastitis diagnostics). Somatic Cell Count determination.

• Commercial potential for diagnostic kits worldwide

  Several kits available; uptake variable.

  GAPS

  Insight into benefits (e.g., antimicrobial stewardship) and limitations (e.g., reduced surveillance, reduced veterinary involvement) of on-farm diagnostics

• DIVA tests required and/or available

  Not applicable at present.

• Vaccines availability

• Commercial vaccines availability (globally)

  For S. aureus two vaccines are available. In Europe StartVac® (HIPRA) contains inactivated Escherichia coli and S. aureus. In the USA, LysiginTM (Boehringer Ingelheim) contains inactivated S. aureus. For S. uberis the UBAC® (HIPRA) vaccine contains lipoteichoic acid from biofilm adhesion component of S. uberis, strain 5616. There may be vaccines available in some countries that we are not aware of.

  GAPS

  There is a need for effective and affordable vaccines, with independent scientific evaluation of efficacy in field trials. Clearly defined criteria for testing/validating new vaccines as they come to market. Vaccines that enhance clearance of established infection (therapeutic effect), possibly in conjunction with other treatment modalities.

• Commercial vaccines authorised in Europe

  STARTVAC® (Hipra), based on killed bacteria.

• Marker vaccines available worldwide

  None.

• Effectiveness of vaccines / Main shortcomings of current vaccines

  Current S. aureus vaccines primarily stimulate humoral immune responses. Trials on the efficacy of the vaccines against IMI have been equivocal. In some studies, a reduction in the severity or duration of infection has been reported. However, in others no benefit of the vaccine was discernible. Startvac was reported to increase milk yield and decrease the severity of clinical mastitis in one study; however, this was in herds where clinical IMI was predominantly due to E. coli. For S. uberis, vaccination was reported to be associated with a moderate reduction in clinical signs of disease, reduced bacterial shedding and increased milk yield after experimental challenge with a heterologous strain. The beneficial effect of vaccination was only evident from day 4 post-challenge. No large field trials of vaccine efficacy have been conducted. Commercial vaccines for S. aureus and S. uberis provide short-term protection only with vaccination each lactation required. None provide sterile immunity.

  GAPS

  Lack of independent assessment of vaccine efficacy in field studies. Lack of multivalent vaccines, i.e. targeting multiple major gram-positive mastitis pathogens.

• Commercial potential for vaccines

  There is a need for efficacious vaccines because of the high prevalence and economic cost of S. aureus and streptococcal mastitis worldwide. Poor treatment success for some mastitis pathogens and societal, regulatory or legal requirement to reduce reliance on antimicrobial agents in veterinary medicine add to the need for effective vaccines.

  GAPS

  Vaccine administration or efficacy requirements may vary based on regional differences in pathogen prevalence or disease manifestation, e.g., subclinical versus clinical; chronic versus acute IMI; heifer versus cow. For small ruminants, to be cost-effective, the vaccine should be very cheap.

• Regulatory and/or policy challenges to approval

  Pathways for regulatory approval exist, but return on investment may not be sufficient to cover the costs of vaccine development and regulatory approval.

• Opportunity for barrier protection

  May improve if a more thorough understanding of the host-pathogen interaction and immunity to S. aureus and streptococci can be elucidated. Vaccines are often seen as a means to reduce antimicrobial use. The use of vaccines to potentiate antimicrobial therapy has not been fully evaluated in vaccine studies. As treatment efficacy can be as high as 80-90% in early stages of infection (weeks), the use of a vaccine to extend this window would extend the opportunity for detection and treatment, contributing to prevention of chronicity and prevention of transmission, thereby potentially reducing overall antimicrobial use.

  GAPS

  Previous unsuccessful attempts suggest that novel approaches are needed in addition to improved understanding of host-pathogen interactions, the relationship between mucosal and mammary immune responses, and what constitutes a protective immune response. Vaccines that protect against multiple gram-positive pathogen species e.g. trained immunity (innate immunity immunological memory) or multivalent vaccines. Vaccines that only require only a single or limited number of administrations for ease of use and to manage costs of vaccination. Vaccines that enhance clearance of established infection (therapeutic effect), possibly in conjunction with other treatment modalities. Vaccines that do not require injection (e.g., nasal vaccine to stimulate mucosal immunity).

• Pharmaceutical availability

• Current therapy (curative and preventive)

  Numerous treatment products are commercially available for cattle. Sheep and goats are minor species and most antimicrobial use is off-label. In addition to antimicrobial treatment (curative and preventive), non-steroidal anti-inflammatory products are used, both for pain management and to limit negative impacts of mastitis on fertility. Both lactating and dry cow therapies are widely available. In dry cows, internal teat sealants are used as prevention tools Prevention is described in section “Main means of prevention, detection and control/Sanitary measures”. For contagious mastitis, antimicrobial treatment or culling of infected animals contributes to prevention. In northern Europe and in other parts of the world, excellent control of contagious mastitis has been achieved without ongoing reliance on routine use of antimicrobials. Indeed, in northern Europe, blanket antimicrobial dry cow treatment has never been practiced. Instead, routine use of diagnostics, prevention of transmission, and genetic selection have been key components of mastitis control programs. This requires investment in recruitment, retention, and training of farm staff, veterinarians, and other technical advisors or diagnostic experts.

  GAPS

  As detailed in previous sections, including Diagnostic tests for gram-positive mastitis that are ideally high throughput, highly sensitive and highly specific (not affected by sample contamination) to allow for rapid detection and intervention. Studies on the motivation of farmers and veterinarians. Strain-specific markers of transmissibility and probability of cure that can be used routinely at low cost

• Future therapy

  Improved therapies that utilize flexible treatments that are pathogen dependent will be needed in the future. The use of improved diagnostics and vaccines will be needed to maximize treatment efficacy. The use of peptide antimicrobials may offer the option of no withdrawal times, blanket fresh cow therapy and heifer treatments. Novel antimicrobial compounds that act intracellularly are under development. Bacteriophages, and more recently endolysins, have been proposed as alternative treatments for bovine infections, particularly during the dry period. Bacteriophages are self-proliferating entities that typically have narrow bacteriocidal specificity at species or strain specificity level. Bacteriophages do not penetrate eukaryotic cells (i.e., they cannot act intracellularly) but disrupt cellular processes in bacterial cells. Time from administration to bacterial eradication may be long. Bacterial resistance to phages can arise through the natural co-evolution between the virus and its host. Phages may be less effective as antibiofilm agents if they lack certain enzymes, such as depolymerases, which are crucial for penetrating biofilms. Additionally, phages may interact with the immune system, making them susceptible to clearance by antibodies, further limiting their effectiveness. There is growing evidence of efficacy of phage therapy of S. aureus in human patients, but treatment is still highly individualised and experimental and not suitable for routine use. Endolysins are phage products (hydrolytic enzymes that cleave the prokaryotic cell wall at the end of the lytic cycle). They have relatively broad lytic activity, do not proliferate, and target bonds in the peptidoglycan. Bactericidal activity is rapid (within seconds of contact). Modification of endolysins can enhance intracellular efficacy. Resistance has not been reported. Endolysins are relatively effective antibiofilm agents with higher destruction of biofilms. They are immunogenic, but with a lower degree of antibody neutralization than phages. Because they are not self-replicating, they can be administered at a defined concentration at a site of infection and in blood circulation.

  GAP

  Most animal health companies rely heavily on their parent firms to provide new antimicrobial agents. Since most companies have exited antimicrobial discovery, there are few compounds available to develop and lack of trained personnel in animal health companies with experience in antimicrobial discovery. If new antimicrobials were to be discovered, their use would likely be restricted, e.g. reserved for human use, to preserve their efficacy. This mean that there is no financial incentive to develop new compounds as there will be limited return on investment. Further development and engineering is needed to enable use of endolysins targeting gram-positive bacteria causing mastitis in ruminants. Production costs associated with endolysins may pose a significant limitation. Further investigation is warranted into the potential synergistic use of endolysins, bacteriophages, and other antimicrobial compounds, whether for lactation or in conjunction with internal teat sealants.

• Commercial potential for pharmaceuticals

  Low due to regulatory hurdles for new antimicrobial agents in veterinary medicine, and new actives may be restricted to humans. EU regulation prohibits prophylactic use. Commercial potential in sheep and goats is extremely low.

  GAP

  Narrow spectrum antibacterials that are not considered critical for human use are needed.

• Regulatory and/or policy challenges to approval

  Current EU and US policies are not conducive to the development of new antimicrobial agents in veterinary medicine.

• Commercial feasibility (e.g manufacturing)

  Yes

• Opportunities for new developments

  New classes of antimicrobial agents that provide high levels of efficacy with minimized zoonotic issues.

  GAP:

  Overall, the AH industry does not have active Antibacterial Discovery programs looking for new mastitis therapeutics.

• New developments for diagnostic tests

• Requirements for diagnostics development

  GAPS

  Rapid, sensitive, specific and cost-effective laboratory, cow-side or in-line testing is needed to help manage infected individuals and herds with S. aureus and streptococcal mastitis. Educated veterinarians capable of interpreting the results of the novel assays in the context of the overall management and mastitis control program.

• Time to develop new or improved diagnostics

  Years

• Cost of developing new or improved diagnostics and their validation

  N/A

• Research requirements for new or improved diagnostics

  Better understanding on how diagnostics inform herd management.

• Technology to determine virus freedom in animals

  Not relevant to staphylococcal or streptococcal mastitis. Note that a PCR test is available to detect avian influenza A virus in bovine milk (in use as of March 2024)

• New developments for vaccines

• Requirements for vaccines development / main characteristics for improved vaccines

  Entry level S. aureus and streptococci vaccines should reduce the severity of clinical mastitis (mostly S. aureus), reduce the incidence of new infections (either by reducing shedding and transmission, or by enhancing host response to new infections). If possible, clear existing IMI (therapeutic vaccine), either alone or in conjunction with antimicrobial or other therapies. Cost-effective, stable at room temperature, single or limited number of doses.

  GAPS

  A whole-genome/proteomic approach to reverse vaccinology has resulted in some success for other organisms and would be a rational way to design an effective subunit vaccine by objectively testing the protective efficacy of surface-presented proteins. Investigation of the potential of immunologically subdominant antigens to circumvent interference from immune imprinting. Trained immunity-based vaccines which confer non-specific protection against IMI. Appropriate small animal disease model (mice are not natural S. aureus hosts).

• Time to develop new or improved vaccines

  Years or decades

  Vaccines against staphylococci and streptococci have been studied for decades and very few of the recently developed vaccines have made it to commercialization either in human or veterinary medicine. Recent studies in humans have proposed vaccine failure may be due to non-protective imprint from prior host-S. aureus interaction which may advance the development of S. aureus vaccines. However, the likelihood of a new efficient vaccine reaching the marketplace in the near future is difficult to estimate.

  GAPS

  Owing to the lack of understanding of protective immunity and of the reasons why previous vaccines fell short of expectation, vaccine development resorts to hit-and-miss approach or contested rationales.

• Cost of developing new or improved vaccines and their validation

  R & D and eventual large scale testing would potentially cost millions of Euros.

• Research requirements for new or improved vaccines

  New knowledge and tools are available to investigate the bovine immune responses. This can give rise to the hope that some success will come despite previous failures.

  GAPS

  There are a number of knowledge gaps impeding the rational design of vaccine strategy, involving both pathogenesis mechanisms and vaccine-induced immune mechanisms protecting the mammary gland. Thorough understanding of the host-pathogen interaction is needed before proceeding with rational vaccine development: Understanding of the diversity of S. aureus and streptococcal strains infecting dairy ruminants and the antigenic variation of surface-presented and secreted proteins Understanding of the molecular basis for pathogenesis of mastitis including genome-scale analysis of proteins involved in host-pathogen interactions Testing of selected antigens individually and in combination for ability to induce protective immune response Examine potential for conserved antigens as vaccine components Better knowledge of the (protective) immune response (cellular and humoral) Investment in basic research on cell-mediated immunity in the ruminant species New adjuvant favouring a protective long-lasting immune response Strategies that target bacterial nutrient use and multiplication, or bacterial adherence or invasion in host cells may need to be re-visited to identify mechanisms that could be exploited for vaccination, e.g. by inducing antibodies that would obstruct essential bacterial processes.

• New developments for pharmaceuticals

• Requirements for pharmaceuticals development

  Depends on avenue taken. Identification of a new antimicrobial agent, if not available from human health programs, may require development of veterinary specific analogous programs. For gram-positive mastitis pathogens, narrow spectrum antimicrobials that do not exceed high importance for human medicine in the WHO classification are the preferred treatment option. Such products are already available. Penicillin resistance is rare in streptococci and penicillin-resistant S. aureus shows limited response to treatment regardless of the compound used, making culling (and prevention) the preferred approach at the time of writing. Use of broad-spectrum products or compounds with critical importance to human medicine or higher importance ranking is difficult to justify as there is no obvious benefit from an animal health perspective, while there is a clear concern from a human health perspective.

  GAP

  Lack of personnel in animal health companies with antibacterial discovery experience. Products for treatment of penicillin-resistant S. aureus.

• Time to develop new or improved pharmaceuticals

  Some novel antimicrobial compounds are currently under development but require several years of in vitro and in vivo testing. May need 10-15 years.

• Cost of developing new or improved pharmaceuticals and their validation

  Potentially millions of Euros (30-50).

• Research requirements for new or improved pharmaceuticals

  Need to determine classes with necessary spectrum of action with reduced conflicts with uses in human medicine.

Disease details

• Description and characteristics

• Pathogen

  Staphylococcus aureus is a gram-positive pathogen that causes a variety of diseases in humans and animals. In dairy cows, sheep, goats, water buffaloes and camels, the most economically important pathology caused by S. aureus is mastitis. S. aureus is catalase positive, generally but not always coagulase positive, and haemolytic. Streptococci are also gram-positive pathogens. The major species associated with mastitis are Streptococcus agalactiae, Streptococcus dysgalactiae subsp. dysgalactiae, and Streptococcus uberis. They are catalase negative, and may be haemolytic (most S. agalactiae), positive for the CAMP reaction (Christie–Atkins–Munch-Peterson reaction; most S. agalactiae., some S. uberis), and/or esculin positive (S. uberis). The term “environmental streptococci” has been used to describe bacteria belonging to several genera, including Enterococcus and Lactococcus but is both scientifically and epidemiologically inaccurate. The term “Streptococcus and Streptococcus-like organisms (SSLO)” is a scientifically accurate descriptor for members of the genera Streptococcus, Enterococcus and Lactococcus. Although there are some publications on Streptococcus canis, Enterococcus faecium, Enterococcus faecalis, Lactococcus lactis and Lactococcus garviae as causes of bovine mastitis, they are beyond the scope of the current gap analysis.

  GAPS

  With the advent of mastitis diagnostics based on matrix-assisted laser desorption ionisation time of flight mass spectrometry (MALDI ToF MS), a growing number of Streptococcus, Enterococcus and Lactococcus species is identified in association with mastitis. For most of those species, there is very limited knowledge of control methods (prevention or treatment).

• Variability of the disease

  S. aureus and S. agalactiae are host generalists that have a wide host spectrum with people as their primary host, and dairy cattle and other animal species as secondary host. Within both bacterial species, host generalist clades exist, which affect people as well as cattle, as well as host specialist clades, which are largely or exclusively limited to a single host species or host group (e.g., small ruminants). The distribution of strains within each pathogen species differs between countries and within and between continents. Occurrence of S. dysgalactiae subsp. dysgalactiae and S. uberis is largely limited to ruminants. The epidemiology of S. uberis mastitis varies considerably between different countries and production systems. Large, standardised databases and typing systems exist for S. aureus and S. agalactiae because they are also human pathogens. These include multilocus sequence typing (MLST) and core genome multilocus sequence typing (cgMLST). PubMLST also includes typing databases for S. uberis and for S. dysgalactiae. The latter is dominated by the multi-host, zoonotic pathogen S. dysgalactiae subsp. equisimlilis, and contains limited information about the mastitis pathogen S. dysgalactiae subsp. dysgalactiae.

  GAPS

  There is a lack of standardised typing systems that can be used in a diagnostic setting for definitive typing of mastitis isolates at subspecies level with the aim of predicting to inform case- or herd management based on transmissibility or prognosis. The only exception at the time of writing is the use of penicillin resistance as prognostic marker in S. aureus, which is associated with poor treatment outcomes.

• Stability of the agent/pathogen in the environment

  For S. aureus and S. agalactiae, the main reservoir in a cattle herd is the infected mammary gland, which forms the basis of prevention strategies that aim to control contagious transmission, whereas additional reservoirs or sources are relatively more important for S. dysgalactiae and S. uberis. Non-mammary reservoirs exist for each of these species. S. aureus can be found on ruminant skin, skin wounds (e.g., hock lesions) and, particularly for small ruminants, in nares; on milking equipment, in bedding, on flies, and in people. Duration of environmental survival is poorly characterised and may differ between strains, possibly in association with biofilm formation, and environmental conditions, such as cleaning and disinfection of teat skin and milking equipment. Strains of human origin, esp. methicillin resistant S. aureus (MRSA) can survive for weeks on fabrics and plastics. The mixed contagious/environmental epidemiology of S. aureus is supported by epidemiological data about sources and the impact of control strategies, as well as strain typing data that demonstrate genetic heterogeneity of S. aureus within herds. Although S. agalactiae is often described as an obligate intramammary pathogen, some strains, notably sequence type (ST)103 are also carried in the gastro-intestinal tract of dairy cattle. S. agalactiae may be found on mucosa, in faeces, on equipment, and in drinking water. Humans may also serve as reservoirs of S. agalactiae for dairy cattle. In dairy camels, nasopharyngeal carriage is common. Environmental survival is poorly characterised, but survival in surface water for several weeks has been described. The reservoirs of S. dysgalactiae are relatively poorly studied but include intact skin, wound, and cubicle bases in freestall herds, as well as infected mammary glands. Reservoirs of S. uberis include infected mammary glands, bovine faeces, and anything contaminated with bovine faeces, including the animals’ legs, paddocks, bedding, flies, laneways etc. The quantity and diversity of S. uberis in the environment correlates with the intensity of cow traffic: more cows means more S. uberis. Documented survival time in the environment is approximately 3 weeks.

  GAP

  The relative contribution of extra-mammary colonization and/or environmental contamination as a reservoir for intramammary infection (IMI) is not well-defined at herd- or strain level. This is particularly important for S. aureus and S. uberis. There is a need for methods that can be used routinely in diagnostic microbiology to differentiate between contagious and environmental modes of transmission. The environmental survival time of mastitis pathogens under different conditions (climatic, environmental, management) is poorly known. A lack of systematic environmental sampling contributes to these knowledge gaps.

• Species involved

• Animal infected/carrier/disease

  Mastitis is the main disease caused by S. aureus in ruminants, including cows, sheep, goats, camels and water buffaloes. Mastitis can be subclinical or clinical and may be very acute and severe, particularly in heifers. Many other animal species can be affected by S. aureus, including horses, pigs, dogs, cats, rabbits and poultry (for humans: see 8.2). The organism may cause a wide range of conditions, as well as asymptomatic carriage. Pigs and poultry are major carriers of ST398 (including methicillin susceptible and methicillin resistant strains), which may spill over into dairy cattle. S. agalactiae is a major pathogen of dairy cattle and water buffalo (subclinical mastitis), camels (subclinical mastitis, skin and soft tissue infections), and frogs and fishes (encephalitis). S. agalactiae spill-over has been described into horses, dogs, cats, rats, mice, monkeys, aquatic mammals, and crocodiles, generally with humans as reservoir. Infection in pigs and porcupines has also been documented for a limited number of clades (clonal complex (CC) 103). S. dysgalactiae subsp. dysgalactiae is largely limited to dairy cattle and sheep. In cattle, mastitis (subclinical or clinical) is the main manifestation. In sheep, polyarthritis in lambs (“joint ill”) is the main disease manifestation, rather than mastitis. S. uberis is almost exclusively associated with carriage or mastitis (subclinical and clinical) in cattle, buffalo and sheep.

  GAPS

  Although host-jumps (evolutionary adaptation) and interspecies transmission (current events) have been described for S. aureus and S. agalactiae, both in the scientific literature and anecdotally, their frequency and relative importance as contributors to the epidemiology of mastitis are poorly quantified. This is especially relevant for mastitis caused by MRSA, and in herds or dairy production systems where S. agalactiae re-emerges.

• Human infected/disease

  In humans, S. aureus is both a commensal and a major pathogen responsible for a wide range of clinical diseases, such as skin and soft-tissue infections, sepsis, meningitis, pneumonia, endocarditis, and septic arthritis. S. aureus is capable of producing a variety of toxins which results in severe food-poisoning (incl. diarrhoea, vomiting) and, TSS (toxic shock syndrome). Dairy foods may be a source of staphylococcal food poisoning due to presence of S. aureus of bovine origin or contamination with S. aureus of human origin during harvesting or processing. Staphylococcal enterotoxins are heat stable and remain intact during pasteurisation. Humans can be a source of MRSA (e.g., ST5, ST8) for dairy cattle. S. agalactiae is better known in human medicine as Group B Streptococcus or GBS. It is carried asymptomatically by approximately 20-40% of the male and female population, with differences between age groups, countries and ethnicities. The gastrointestinal tract is the most common carriage site, followed by the urogenital tract, and oropharynx. Clinical manifestations of GBS disease include (1) maternal, foetal and neonatal disease; (2) opportunistic disease in non-pregnant adults, including skin and soft tissue infections, urinary tract infections, and sepsis in immunocompromised individuals; and (3) invasive foodborne GBS disease derived from raw seafood (ST283). GBS is a leading cause of neonatal and infant sepsis and meningitis globally. There is no direct evidence for zoonotic GBS infection, other than via the foodborne route for ST283, which had not been documented in ruminants at the time of writing. There is debate about evolutionary relationships between human and bovine strains of GBS. S. dysgalactiae subsp. dysgalactiae is not zoonotic (shared between humans and animals), in contrast to S. dysgalactiae subps. equisimilis. S. uberis is not zoonotic. Some strains are used as probiotics in commercially available human mouth wash products.

  GAPS

  The dynamics of transfer of S. aureus or S. agalactiae strains from humans to dairy ruminants and vice versa deserves to be investigated on a regular basis using modern genotyping methods, such as whole genome sequencing (WGS).

• Vector cyclical/non-cyclical

  Flies have been shown to be colonized and act as possible vectors for the transmission of S. aureus in cases of bovine mastitis. Fly control can contribute to mastitis prevention. Flies also contribute to the summer mastitis complex, which mostly affects non-lactating cattle (dry cows, heifers) with S. dysgalactiae subsp. dysgalactiae, Trueperella pyogenes, and Peptostreptococcus indolicus as potential contributing etiological agents. Transmission of S. dysgalactiae by wasps has been documented in a mastitis outbreak that primarily affected heifers.

• Reservoir (animal, environment)

  See Section “Stability of the agent/pathogen in the environment”

• Description of infection & disease in natural hosts

• Transmissibility

  Transmission of S. aureus and streptococci happens primarily during the milking process. The bacteria are spread from infected quarters to uninfected quarters via teat cup liners, milkers' hands, and wash cloths. Several existing control strategies are aimed at reducing this likelihood including 1. Identification of infected quarters or animals so that they can be removed from the remainder of the herd through treatment, drying off, temporary segregation or culling 2. Appropriate settings and proper use of the milking equipment to reduce the risk of pressure gradients that may contribute to dispersal of bacteria-laden milk droplets 3. Postmilking teat-disinfection to kill bacteria that may be deposited on teat skin during the milking process. Additional measures, such as backflushing of units between cows, may also contribute to a reduction in transmission. Further detail on use of sanitary measures to prevent transmission is provided in Section “Main means of Prevention, detection and control/Sanitary measures”. Transmissibility is a function of pathogen species and herd management, with highest transmissibility for S. agalactiae, followed by S. aureus, then S. dysgalactiae, and finally S. uberis. Most S. agalactiae problems are due to contagious transmission, although rare examples of environmental or anthroponotic (reverse zoonotic) transmission have been described. S. aureus is generally considered contagious but ca. 20% of S. aureus problems are attributed to environmental origins in some countries. Moreover, in many herds, multiple strains of S. aureus co-exist, often with one or two predominant strains and multiple strains limited to one or a few animals. This implies that there are differences in transmissibility at strain level. S. uberis transmissibility is variable, with over 50% of clinical mastitis due to S. uberis attributed to contagious transmission in England, compared to no contagious transmission in New Zealand. Transmissibility of S. dysgalactiae is thought to be intermediate between S. aureus and S. uberis. Transmission via bedding or vectors cannot be differentiated from transmission via the milking machine based on molecular typing of mastitis isolates alone but requires additional epidemiological and bacteriological investigations, such as isolation and typing of bacteria from milking unit liners, bedding material, or insects.

  GAPS

  There is no standardised typing method that can be used to predict the transmissibility of staphylococcal or streptococcal strains without comparative analysis of multiple isolates per herd. Molecular indicators for transmissibility would be useful to inform herd management.

• Pathogenic life cycle stages

  Neither S. aureus nor streptococci form spores. There is debate as to whether biofilm plays a role in the life cycle of these organisms. Biofilm formation has been demonstrated in vitro, generally on plastic surfaces and often under static conditions. It may occur on-farm in milking equipment or water troughs. Biofilm formation may affect antimicrobial efficacy in vitro. Whether biofilm formation plays a role during intramammary infection or in response to antimicrobial treatment is unknown. For S. aureus, small colony variants (SCV) have been described. Their appearance on agar is fried-egg like, similar to Mycoplasma, and they go undetected in standard culture-based diagnostic assays.

  GAPS

  Knowledge of the biological or in vivo relevance of biofilm formation as observed in vitro. The epidemiological relevance of SCV of S. aureus in treatment response and transmission is unknown. Knowledge of the biological or in vivo relevance of biofilm formation as observed in vitro. The epidemiological relevance of SCV of S. aureus in treatment response and transmission is unknown.

• Signs/Morbidity

  All four major gram-positive pathogens can cause subclinical mastitis (inflammatory response as evidenced by elevated somatic cell counts) or clinical mastitis, which can be mild (milk affected), moderate (milk and mammary gland affected), or severe (milk and mammary gland affected plus systemic effects). S. agalactiae primarily causes subclinical mastitis, although human-adapted strains that infect cows may cause clinical mastitis. S. dysgalactiae causes subclinical and mild clinical mastitis. S. uberis causes subclinical to moderate clinical mastitis. S. aureus may cause mastitis of any severity, including gangrenous mastitis. Different strains of S. aureus appear to be associated with different levels of severity within individual herds. Gangrenous mastitis, which is often fatal, generally occurs around calving, suggesting that environmental sources may play a role as opposed to strains that are transmitted during milking. Severe forms (gangrenous signs) are much more frequent in goats and particularly sheep than in cows. In cows, S. aureus mastitis may become chronic with formation of purulent lesions or abscesses, which may be palpable, and which are refractory to antimicrobial treatment. All four major gram-positive pathogens can cause subclinical mastitis (inflammatory response as evidenced by elevated somatic cell counts) or clinical mastitis, which can be mild (milk affected), moderate (milk and mammary gland affected), or severe (milk and mammary gland affected plus systemic effects). S. agalactiae primarily causes subclinical mastitis, although human-adapted strains that infect cows may cause clinical mastitis. S. dysgalactiae causes subclinical and mild clinical mastitis. S. uberis causes subclinical to moderate clinical mastitis. S. aureus may cause mastitis of any severity, including gangrenous mastitis. Different strains of S. aureus appear to be associated with different levels of severity within individual herds. Gangrenous mastitis, which is often fatal, generally occurs around calving, suggesting that environmental sources may play a role as opposed to strains that are transmitted during milking. Severe forms (gangrenous signs) are much more frequent in goats and particularly sheep than in cows. In cows, S. aureus mastitis may become chronic with formation of purulent lesions or abscesses, which may be palpable, and which are refractory to antimicrobial treatment.

  GAPS

  It is unknown to what extent manifestation (subclinical, mild, moderate, severe) and duration of infection are driven by host, pathogen, or management characteristics. There are no standardized diagnostic markers for severity based on typing of bacterial isolates. What makes small ruminants more prone to severe S. aureus mastitis (or less susceptible to subclinical S. aureus mastitis) has not been identified.

• Incubation period

  Incubation period (i.e., time until manifestation of clinical signs) may depend on pathogen species, pathogen strain, host genetics (at least for S. aureus), host immune status, and physiology. Infection may remain subclinical, in which case there is no incubation time in a clinical sense. In experimentally induced infections, incubation time ranges from 12 to 72 h for clinical mastitis based on studies with S. aureus and S. uberis, if clinical signs occur. A low inoculum (< 100 colony forming units) may be sufficient to achieve this. Bacterial amplification may be detectable prior to changes in cytokine levels or clinical signs (clotting or discolouration of milk, swelling of gland, systemic signs in cow).

  GAPS

  Events taking place during the lag phase between intrusion of staphylococci or streptococci into the mammary gland and the onset of the host inflammatory response or clinical signs are not well understood. Insight into expression of growth factors, adhesins, invasins, immune evasins, or other virulence factors during the early stages of infection might help to inform novel control strategies, whether based on breeding for resistance, vaccination or treatment.

• Mortality

  Direct mortality in bovine dairy herds due to gram-positive mastitis is low, with the exception of gangrenous mastitis due to S. aureus. Most mastitis-associated mortality is due to culling of animals with treatment non-responsive chronic intramammary infection. This is done to prevent within-herd transmission, or negative impact on bulk milk quality. The outcome of treatment differs between pathogens and production systems, with better prognosis for S. agalactiae (90% or more), than S. dysgalactiae (ca. 75%) and S. uberis (ca. 50% in systems with contagious S. uberis). For S. aureus, treatment prognosis and hence the likelihood of culling are heavily dependent on pathogen factors (penicillin resistance of the bacteria reduces the probability of cure, making culling the preferred option), and host factors (duration and location of the intramammary infection, parity). Those factors are well-studied, but poorly utilised in on-farm decision making. Direct mortality can be high in heifers, ewes and goats due to peracute (gangrenous) mastitis.

  GAPS

  Tools for early detection of subclinical and clinical mastitis are available. In addition, the knowledge to predict the usefulness of antimicrobial treatment is largely available. There is a gap in implementation of such tools and knowledge, in part because it is difficult to monetise. As pressure to reduce antimicrobial use grows, social sciences insight may be needed to enhance uptake of existing tools and knowledge.

• Shedding kinetic patterns

  Most infections are detected when they are chronic with varying degrees of bacterial shedding (concentrations) in milk. Shedding is largely persistent in infections caused by S. agalactiae and S. dysgalactiae, very frequent in infections caused by S. uberis (>85% of milk samples from infected mammary glands test positive for bacterial growth), and can be variable in infections caused by S. aureus. Most S. aureus infections, however, are accompanied by persistent shedding. Variable shedding may contribute to false-negative bacterial detection results based on culture or PCR in milk from infected quarters. PCR may give positive results if bacteria are present but not culturable. Spontaneous cure with cessation of shedding is possible, although in a small percentage (< 20%) of infections confirmed by several consecutive samplings. A lengthy dry period in ewes may favour spontaneous cure.

  GAPS

  Drivers of variable shedding levels in S. aureus or S. uberis infections are unknown. The impact of shedding patterns on mastitis diagnostics is poorly quantified.

• Mechanism of pathogenicity

  S. aureus mastitis is an inflammatory disease of ducts and alveoli, due to multiplication of staphylococci in milk and possibly on the mammary epithelium, with the possibility of intracellular bacterial survival and replication, particularly in elongated (myoepithelial) cells. Inflammation, which is triggered by non-specific response to staphylococcal MAMPs (Microbe Associated Molecular Patterns) and exoproteins (proteases and toxins), can be complicated by acquired immunity (hypersensitivity). Micro-abscesses may develop in the secretory tissue during chronic infections. The detrimental effects on mammary secretory tissue of an excessive inflammatory reaction are difficult to evaluate. The ability of S. aureus to invade and survive in mammary epithelial cells, demonstrated by in vitro experiments, is likely to have a major impact on chronic infection and resistance to antimicrobials. The pathophysiology of streptococcal infections is disputed, with some experts describing streptococci as “replicating irritants” in the alveoli, and others emphasising their ability to adhere to mammary epithelial cells (cuboidal or alveolar cells), possibly followed by cellular invasion, and intracellular survival and replication. Different schools of thought regarding pathogenic mechanisms have informed different approaches to vaccine development, e.g. targeting nutritional requirements of bacteria, or adherence and invasion of host epithelial cells (as observed in vitro). Intracellular occurrence of bacteria may also affect efficacy of antimicrobial treatment. In streptococci, nutrient utilisation appears to be a key virulence factor, with all bovine streptococci (but not S. agalactiae from humans or fish) carrying the lactose operon. This operon is shared between streptococcal species infecting the mammary gland through lateral gene transfer. Vaccine strategies aimed at depriving streptococci of essential nutrients have been explored and are distinct from those aiming to induce opsonising antibodies or a cellular response.

  GAP

  Pathogen- and host factors that contribute to pathogenicity are poorly understood and need to be known for rational vaccine design.

• Zoonotic potential

• Reported incidence in humans

  Not quantified but thought to be rare. See Section “Human infection/disease” for human infection and the possibility of interspecies transmission.

• Risk of occurence in humans, populations at risk, specific risk factors

  Staphylococcal food poisoning due to enterotoxins produced by S. aureus is possible, and the EU has specified legal limits for S. aureus in dairy products (COMMISSION REGULATION (EC) No 2073/2005). Populations at risk of interspecies transmission of S. aureus or S. agalactiae may, at least in theory, include those in direct contact with animals, those consuming raw milk, and those with immunosuppression or comorbidities. No data is available on any of those risk factors in relation to mastitis as incidence data in humans are not recorded. S. dysgalactiae subsp. dysgalactiae and S. uberis are not pathogenic to humans.

• Symptoms described in humans

  See Section “Human infection/disease”

• Likelihood of spread in humans

  In the evolutionary biology and epidemiology literature, there are more indications of human-to-animal transmission than of animal-to-human transmission of S. aureus and S. agalactiae. Some researchers have stated that “animal S. aureus always evolve from human strains”. Others have found that the major contemporary endemic clones of S. aureus causing bovine mastitis around the world can be traced back to 4 independent host-jump events from humans that occurred up to 2,500 y ago. In addition to human-to-animal spill over, there is the possibility of adaptation and amplification in the ruminant host, followed by-spill back. Examples include emergence of S. aureus ST97 in humans in Italy (possibly with pigs as intermediary host between cattle and humans), and emergence of the methicillin-resistant and mecC-carrying S. aureus clades such as CC130 and ST425.

  GAPS

  The dynamics of transfer of S. aureus or S. agalactiae strains from humans to dairy ruminants and vice versa deserves to be investigated on a regular basis using modern genotyping methods, such as whole genome sequencing (WGS). In addition, the risk of onward transmission, and the relative importance of inter-species transmission compared to intra-species transmission need to be considered to prioritise control strategies.

• Impact on animal welfare and biodiversity

• Both disease and prevention/control measures related

  Impact of animal welfare depends on the form of the disease, with increasing pain associated with/indicative of increasing severity. In experimental studies, administration of lipoteichoic acid from S. aureus does not elicit a strong pain response, in contrast to administration of lipopolysaccharide associated with gram-negative mastitis pathogens. As a result, most clinical studies of pain and pain mitigation in association with mastitis focus on gram-negative pathogens. Mild clinical mastitis and subclinical mastitis can be associated with changes in lying behaviour and in nociceptive thermal threshold in dairy cows, indicating that subclinical or mild gram-positive mastitis may affect animal welfare. Drying off of individual quarters to prevent transmission of pathogens can be associated with signs of pain. Culling affects longevity. In humans, longevity is increasingly seen as a welfare indictor. In animals, longevity as welfare indicator is probably a proxy for the welfare issues associated with the condition that leads to culling, here: mastitis.

  GAPS

  The impact of gram-positive mastitis on cow welfare and pain is poorly known. The potential benefit of non-steroidal anti-inflammatory drugs on temperature, rumen function, SCC, milk production, reproduction, behaviour, and pain sensitivity in animals during gram-positive mastitis is largely unknown.

• Endangered wild species affected or not (estimation for Europe / worldwide)

  S. aureus, including MRSA, has been identified in a wide range of wildlife species, but their conservation status and the contribution of dairy ruminants to transmission is unknown. S. agalactiae has been identified in wild fishes and sea mammals (grey seal, sperm whale, bottlenose dolphin) but is rarely definitively identified as the cause of death. Humans and farmed fishes rather than dairy cattle are the most likely source. S. agalactiae has recently been detected in crested porcupines in Italy. They are classed as species of lower risk by the International Union for Conservation of Nature (IUCN). The source of infection in porcupines is unknown and could be human or bovine.

• Slaughter necessity according to EU rules or other regions

  Chronic or treatment-refractory mastitis is a cause of premature culling for reasons of milk production, milk quality, antimicrobial use, and risk of pathogen transmission. Recourse to euthanasia remains infrequent but is justified when sepsis or gangrenous mastitis develops.

• Geographical distribution and spread

• Current occurence/distribution

  Worldwide. Prevalence varies between countries, partly in relation to differences in production and animal health systems and diagnostic infrastructure.

• Epizootic/endemic- if epidemic frequency of outbreaks

  The herd is the epidemiological unit. Endemic in herds. Outbreaks can develop at the herd level, notably in early lactation in ovine flocks. Elimination of S. agalactiae from individual herds is generally feasible, with extremely low prevalence attained in several European countries. Denmark is the only country with a national S. agalactiae monitoring program, conducted through annual bulk milk surveillance of all active dairy cattle herds. Dairy cattle herds can have low incidence of S. aureus intramammary infections for long periods. This is likely to depend on the contagiousness and ecology of the strains. Environmental strains are more likely to cause transient infection with limited transmission than host-adapted, contagious strains. Outbreaks of S. agalactiae, S. aureus or S. uberis mastitis may occur when measures to control contagious transmission (notably post-milking teat disinfection) are discontinued or ineffective and have been described in herds with milking parlours as well as automated milking systems. Bovine, ovine and caprine flocks are seldom completely free of S. aureus, whether as carriage strain or cause of subclinical mastitis, and clinical mastitis cases appear from time to time. The incidence and prevalence of S. dysgalactiae infections appears to be higher in northern Europe and Canada than in parts of the European and North American continent further south. Increased incidence has been described in association with insect vectors (flies, wasps) and with damage to udder tissue, e.g., resulting from poor claw health.

  GAPS

  There is no standardised typing method that can be used to predict the transmissibility of staphylococcal or streptococcal strains without comparative analysis of multiple isolates per herd. Molecular indicators for transmissibility would be useful to inform herd management. Social science insights may be needed to enhance uptake of existing tools and knowledge to improve control, including control of contagious transmission of S. dysgalactiae and S. uberis.

• Seasonality

  Climate does not seem to be a risk factor.

  When observed, seasonality is in fact related to other factors: calving periods, variation in risk factor exposure (flies for instance), variation of efficacy in prevention measures (seasonal implementation of teat dipping for instance), …

  GAP:

  Influence of heat stress on host susceptibility.

• Speed of spatial spread during an outbreak

  Spread between herds occurs mainly by introduction of infected animals. This may lead to co-circulation of multiple strains of a pathogen, as demonstrated for S. aureus. Co-mingling or use of shared equipment may also result in spread, e.g., during agricultural shows or community pasturing in the summer. Speed of spread within herds depends on herd management and strain transmissibility.

  GAPS

  In some regions, there has been a move from closed herds to the use of contracted heifer farms that supply heifers to dairy farms. The movement of cattle between farms is much higher than 20-30 years ago and needs to be evaluated as a practice. Point-of-care or point-of-need diagnostic could help to reduce biosecurity risks through identification of infected animals prior to movement.

• Transboundary potential of the disease

  Possible but of minor concern because endemic globally.

• Route of Transmission

• Usual mode of transmission (introduction, means of spread)

  At herd-level, introduction is through live animals. Thus, transmission between herds is generally through movement of animals, e.g., purchased animals, or animals that have been at shared facilities (alpine pastures, agricultural shows). Humans may introduce S. agalactiae and MRSA into dairy herds. At animal-level, the port of entry is the teat canal. Contagious transmission results from spread from quarter to quarter and from cow to cow, usually during the milking process. Environmental transmission refers to infection from extra-mammary sources, including animals’ skin, non-ruminant species, bedding, faeces and other environmental sources. See also Section “Transmissibility”.

  GAPS

  Point-of-care tests for detection of infected cows or quarters. Routine availability of diagnostic strain typing to differentiate between contagious and environmental transmission.

• Occasional mode of transmission

  Insect vectors (fly, wasp), human-to-animal, animal-to-animal (e.g., introduction of Streptococcus canis via dogs or cats).

• Conditions that favour spread

  Failure to follow basic hygienic milking procedures such as use of adequately functioning milking machines, post-milking teat antisepsis, milking order, proper use of antimicrobials and culling of refractory infected animals allow the organism to propagate in dairy herds. High prevalence (increases exposure), and poor control of risk factors (teat lesions and “over-milking” especially) favour spread within herds. Animal movements favour spread between herds. Infrequent monitoring of udder health (clinical signs, somatic cell count) contributes to delayed detection of infection and prolonged opportunities for pathogen transmission as well as for infections to become chronic and treatment refractory. Poor environmental hygiene and insect control favours environmental and vector borne transmission, respectively. Nutritional deficiencies, esp. selenium deficiency, may increase susceptibility of animals. Staff shortages may contribute to failure to follow basic hygiene milking procedures. Use of automatic milking systems (“robots”), where a single unit is used to milk a larger number of animals than in a milking parlour, may be associated with increased or decreased risk of transmission, depending on machine settings and maintenance, and pathogen species.

  GAP

  Social and economic insight into the drivers of and the barriers to implementation of mastitis prevention and control measures based on existing knowledge.

• Detection and Immune response to infection

• Mechanism of host response

  Once in the lumen of a healthy mammary gland following penetration through the teat canal, staphylococci multiply in milk. Microbe-associated molecular patterns (MAMPs) activate the innate immune system, which triggers inflammation. The initial immune response is characterized by an influx of leukocytes, mainly neutrophils, in mammary tissue and milk. Phagocytosis and killing of bacteria by neutrophils is the main defence mechanism and is required to control bacterial growth. Usually the initial neutrophilic inflammation is not able to clear S. aureus infection, which evolves into chronic mastitis. With time, foci of tissue fibrosis and/or abscess formation occur, and cell-mediated hypersensitivity may develop in S. aureus mastitis. This outcome appears to be host and strain dependent. Self-cure occurs in a small proportion of cases. Infection of the mammary gland does not induce protection against subsequent S. aureus infections. Yet, antibodies are induced against several staphylococcal constituents. In particular, IgG opsonizing and toxin-neutralizing antibodies are induced, which increase the titres of antibodies pre-existing intramammary infection. Antibody and cell-mediated immune responses of the correct type and proportions are important in the context of chronic S. aureus mastitis. However, the precise adaptive immunological mechanisms that best control mastitis remain elusive. There is significant variation among S. uberis strains in their ability to establish infection and persist in the mammary gland. The role of neutrophil phagocytosis in the control of this pathogen is uncertain with some strains highly resistant to phagocytic killing by neutrophils. Macrophages have been suggested as the primary phagocytic cell although the bactericidal efficacy of mammary macrophages is limited. Induction of the innate immune response to S. uberis is proposed to be via macrophages rather than mammary epithelial cells. Anti-S. uberis antibodies increase post-infection. Recovery from S. uberis IMI has been reported to protect against subsequent clinical mastitis with the homologous strain in an experimental challenge model. However, protection may not extend to heterologous strains, and quarters that had recovered from S. uberis infection were at increased risk of re-infection in field studies. Phagocytosis by neutrophils is considered the main defence mechanism against S. agalactiae. The bovine immune response to S. dysgalactiae remains poorly researched. There is significant variation in the immune response during lactation and the dry period, which is closely related to periods of high susceptibility (periparturient period and during active involution) and relatively high self-cure rates (steady involution). The genetic basis for resistance to mastitis is an active research field. Genetic selection for mastitis resistance is implemented in many countries. Selection is generally based on somatic cell score (SCS) (+/- Genomic Selection) and hence is pathogen non-specific. In some cases, clinical mastitis (CM) is also a selection trait. Both SCS and CM are polygenic traits with many genes of small effect associated with resistance and little overlap across species, breeds and even studies. In dairy sheep a putative causal mutation controlling SCC has been identified in the SOCS2 gene but other causative mutations remain to be identified.

  GAPS

  It seems worth: Defining the role of cell-mediated immunity in protection and immuno-pathology. There is increasing evidence of the critical role of T cells, especially γδ T cells and their subpopulations, in protective immunity against S. aureus infections in humans and mice; however, our knowledge regarding ruminant mastitis is quite limited. Identifying the immune response(s) leading to clearing of mammary gland infection and appropriate correlates of protection. Designing specific antigen/adjuvant/route of administration combinations to elicit the appropriate immune response. Increasing our understanding of mammary gland immunity during the dry period, especially during the involution period. Deciphering the role of the teat apex and teat canal in mastitis pathogenesis and bovine mammary gland protective immunity. Better understanding how the bovine teat apex, teat canal, and teat cistern serve as primary defence mechanisms against mastitis pathogens, and how we can manipulate these mechanisms to strengthen host defence. The use of a multiomics approach, including single-cell RNA sequencing and epigenetics to enhance our understanding of ruminant mastitis pathogenesis and mammary gland protective immunity. There is a lack of bovine immunological reagents and development of such would facilitate investigation of the bovine immune response to S. aureus/Streptococci. Metagenomic approaches have revealed genetic signatures from a diverse array of bacterial genera in milk. Whether these are commensals or opportunists and if they constitute a true ‘mammary microbiome’ is the subject of debate. Their role, if any, in modulating mammary immune responses and inhibiting pathogens (i.e. competitive inhibition) remains largely undetermined. The genetic basis for resistance to mastitis remains to be elucidated with a view to identifying favourable alleles for selection or even generation of resistant stock via gene editing. Genetic evaluation for pathogen-specific mastitis resistance would be beneficial. Host-driven immunotherapy should be investigated, including the potential use of immune checkpoints. The potential use of stem cell-based therapies for immune modulation and tissue regeneration in bovine mastitis remains an unexplored research gap.

• Immunological basis of diagnosis

  Anti-staphylococcal or streptococcal antibodies titres increase as mastitis develops. Pre-existing antibodies are present in serum of all cows against many S. aureus antigens. In milk of a healthy gland, antibody titres correlate with blood titres, due to transudation of plasmatic antibodies and to selective transport of IgG1. In infected glands, milk titres depend more on exudation of plasma than on local synthesis. As a result, any inflammation of the mammary gland provokes an increase in milk antibody titres to S. aureus, a phenomenon which complicates the use of antibodies for immunological diagnosis. Recruitment of leucocytes in milk by the innate immune system, a very sensitive marker of inflammation of the mammary gland, is a reliable and commonly utilised means of detecting mastitis, although not specific to the causal agent.

  GAPS

  To find a S. aureus and/or S. uberis antigen inducing antibodies during infection, but not recognized by pre-infection serum (giving rise to sero-conversion). Identify S. aureus and streptococci antigens associated with a favourable prognosis, particularly those linked to a high odds ratio for cure. Penside device (e.g. lateral flow assay) for direct immunodetection of specific pathogens in milk.

• Main means of prevention, detection and control

• Sanitary measures

  Most recommended sanitary measures were introduced decades ago. 1. Prepare teats properly prior to milking. Udders should be dry, and teats should be cleaned and dried prior to machine attachment. In some countries, pre-milking teat-disinfection is allowed and recommended. 2. Use adequately sized, properly functioning milking equipment. Use milking machines in a proper manner on properly prepared cows. Wear clean gloves during the milking process. The proper maintenance and use of milking equipment are crucial to preventing new intramammary infections. Avoid unnecessary air admission into the teat cups during unit attachment, machine stripping and unit take-off that can cause irregular vacuum fluctuations. 3. Disinfect teats. Use an effective product after every milking. Post-milking teat disinfection is the single most effective practice to reduce the rate of new intramammary infections by contagious pathogens. A good product should also include skin care components, as healthy skin is more resilient to bacterial colonisation than cracked or chapped teat skin. 4. Assess clinical cases for treatment decisions. Work together with the herd veterinarian to design a management protocol for the treatment of mild, moderate, and severe cases of clinical mastitis. Selective treatment of clinical mastitis is highly recommended to reduce the use of antibiotics. Cows affected by severe cases of clinical mastitis should be treated. Moderate and mild cases of clinical mastitis can be treated in udder quarters with a higher chance of cure. A poor prognosis is associated with older cows (parity >2), chronic infections (multiple quarters affected, multiple previous cases of clinical mastitis, very high SCC, long duration of SCC elevation) and, in the case of S. aureus, with penicillin resistance. Early detection and treatment improve the prognosis. Longer duration of treatment will enhance the probability of cure, esp. for S. aureus and S. uberis, but the cost-benefit balance (milk withhold, selection pressure for antimicrobial resistance) may not justify this. 5. Use dry cow therapy. For prudent use of antibiotics, selective dry cow treatment should be considered. In Europe, blanket use of antimicrobial prophylaxis, e.g., for dry cow treatment, is not allowed (Regulation EU 2019/6). 6. Consider culling chronically infected cows. Cows infected with low or non-responsive mastitis pathogens, such as treatment-nonresponsive streptococci, chronic S. aureus and Mycoplasma spp., should be considered for culling due to the risk they pose to non-infected cows/udder quarters in the herd. 7. Maintain a closed herd. If new animals are purchased, quarantine and test them before adding them to the herd. 8. Maintain a clean, dry and comfortable environment. Ensure appropriate bedding management (if housing is used), clear and dry traffic areas, and proper stall size and design, providing adequate space for all cows. 9. Establish an active milk quality program. The herd veterinarian, together with the farm team, should establish achievable goals for udder health and maintain good data records. Communication is a crucial step in achieving successful control of mastitis. 10. Milk infected cows last, in a separate group, or with a separate cluster, or disinfect the cluster after milking an infected cow. 11. Blitz therapy should be considered for controlling S. agalactiae mastitis, especially in low and middle-income countries. Blitz therapy is the treatment of all affected cows at once. This requires good diagnostics, good hygiene during treatment, and good follow-up on the outcome of treatments. It also requires arrangements for disposal of milk from treated cows.

  GAPS

  The effectiveness of the control measures may depend on the properties (contagiousness, ability to colonize sites other than the mammary gland) of the strains prevailing in herds. Therefore, it would be useful to be able to assess the ecological behaviour of isolates (such as infection dynamics at the herd level), i.e. through genotyping. The adoption of automatic milking systems (AMS) is increasing worldwide. Further studies are needed to assess their impact on mastitis control and detection, and how to improve it, as they have been associated with an increased milk SCC in some countries. There is a need for better understanding of strategies to monitor bedding hygiene under different climate conditions, and for evidence-based benchmarks to interpret bedding culture reports, particularly concerning streptococci and streptococci-like organisms. There is a gap in understanding farmers' behaviours, identifying barriers to change, and determining motivators for implementing effective mastitis control strategies.

• Mechanical and biological control

  The most effective control is to prevent new infections by minimizing or eliminating conditions that contribute to the exposure of teat ends to bacteria so that they can colonise the teat end or penetrate the teat canal. This applies to bacteria that are spread by contagious transmission and to environmental organisms. Internal teat sealants, as increasingly used in dry cows to prevent infection, are the prime example of biological control. In some countries, they are used in combination with antimicrobials to minimize the risk of iatrogenic intramammary infection, but in Europe this is not allowed. Antimicrobials can be used to treat existing infections, mechanical teat sealants (mostly internal sealants based on bishmuth subnitrate; external teat sealant exist too) can be used to prevent new infections. Certain nutritional components enhance the animal's resistance to mastitis. Supplementation of the diet with vitamin E and selenium, vitamin A and beta-carotene, and balancing dietary copper and zinc content to meet requirements have reduced mastitis. Vector control may contribute to prevention of mastitis caused by S. aureus and S. dysgalactiae. The role of vaccination and biologicals as treatment options is discussed in subsequent section.

  GAPS

  Influence of nutrition on immunological status. The impact of genetic and epigenetic modulation of the immune response on the outcome of infection after exposure. Value of inclusion of biologicals (e.g., phages, bacteriocins, bacteriolysins) in mechanical control tools (internal teat sealants).

• Diagnostic tools

  Diagnosis of inflammation is routinely performed by measurement of indicators of inflammation, such as milk somatic cell counts (SCCs). However, this is not pathogen- specific. Etiological diagnosis is currently by bacteriological analysis or PCR. Bacteriological analysis requires collection of aseptically taken milk samples from individual mammary quarters, which may be combined into a composite or cow-level sample. The risk of contamination during sampling (cow-side conditions), phases of low shedding and time to get results are limitations of laboratory-based bacteriological diagnosis. Bacterial identification at the genus and species level can be greatly facilitated by using the Matrix-Assisted Laser Desorption/Ionization Time of Flight Mass Spectrometry (MALDI-ToF MS), provided the data banks are updated for bacteria of animal origin. Identification of streptococci other than S. agalactiae, S. dysgalactiae and S. uberis may not always be reliable. Availability of bacterial isolates allows for conduct of antimicrobial resistance tests and strain typing. Such additional assays can help to inform on prognosis, case management and herd management as detailed in other sections. Polymerase chain reaction (PCR) is routinely used for mastitis diagnostics in northern Europe, using both aseptically collected samples and non-aseptically collected samples, e.g., those used for herd testing (SCC measurement). Carry-over between samples that are not aseptically collected may result in false-positive results and results should be confirmed at animal level to inform culling decisions. Some PCR-based diagnostics can routinely identify penicillin resistance in S. aureus. PCR can detect DNA from non-viable bacteria, or bacteria that have entered a viable, non-culturable state. The use of on-farm culture methods is growing in several countries, particularly for establishing clinical mastitis treatment decisions and reducing the use of antimicrobials. At the time of writing, all on-farm methods were based on growth of organisms. A variety of systems is available, incl. petrifilm, agar and “dipstick” based platforms, among others. Incubators can be miniaturised and hold one sample at time, or they can hold multiple stacks of agar plates. Media may be selective (suppress or allow growth of specific species) and/or chromogenic (colour indicator for genus or species) to help with identification of outcomes of interest. Outcomes range from gram-positive (yes/no) to detailed genus and species identification (e.g., multiple species of streptococcus and staphylococcus). Read-outs can be based on visual inspection of line on a strip or colours on a plate, to automated cloud-based analysis of growth patterns.

  GAPS

  The use of inflammation indicators in the diagnosis of mastitis in small ruminants is less precise than in cows; therefore, novel diagnostic tools should be regarded as a priority. There is a need for novel, affordable, sensitive, specific, user-friendly, rapid, equipment-free, and end-user deliverable (ASSURED criteria) diagnostic tools for identifying IMIs at dry-off to strengthen the implementation of selective dry cow therapy, and for targeted treatment of clinical mastitis during lactation. A rapid, cow-side or in-line, diagnostic test would allow timely implementation of targeted treatment. The level of detail (e.g., gram-positive/gram-negative, genus, species, strain) required from the test depends on the decision making that is linked to the test. On-farm diagnostics need to be both sensitive and specific to avoid false-negative and, more importantly, false positive diagnoses. Ideally, false-positive results due to contamination would be identified. Diagnostics need to be very cheap to allow routine use (esp. when used in-line), and quick, ideally allowing decision making during or immediately after milking. Nucleic acid amplification tests (NAAT), including PCR and loop-mediated isothermal amplification (LAMP) deserve research and development. Issues inherent to the technique (high sensitivity implying ‘sensitivity’ to sample contamination; inability to distinguish dead from live bacteria) will deserve due consideration. On-farm biosafety guidelines and measures are needed for handling and disposal of on-farm culture of hazard group 2 organisms, which include S. aureus and S. agalactiae. Culture-free on-farm diagnostics, e.g. based on metabolite, peptide and protein profiles, that are specific to the pathogen (at the level that is relevant to decision making) rather than severity of the host response. Exploring how machine learning and big data can provide a more accurate and earlier diagnosis of mastitis, particularly for different udder pathogens. Socio-economic and legal aspects of on-farm testing, including, but not limited to, culture of hazard group 2 organisms in non-professional laboratories, antimicrobial stewardship, drivers for uptake, and implications for the business model of veterinarians as advisors and as actors that generate income from antimicrobial sales.

• Vaccines

  Vaccines based on killed bacterins are currently approved in several countries. Their efficacy is either limited and fickle or in need of large-scale field evaluation. Numerous attempts to induce protection with subunit vaccines have been done or are ongoing, so far without convincing results. Since the mammary gland is the predominant reservoir for contagious transmission, a vaccine that prevents intramammary infection or clears it very shortly after infection is needed. None of the vaccines studied to date have achieved this goal. Most vaccines rely on humoral immunity, however there is increasing evidence of the critical role of cell-mediated immunity on protective immunity.

  GAPS

  There is an urgent need for effective vaccines against S. aureus, S. uberis and S. dysgalactiae. Although less of a priority in Europe, many other dairy industries would benefit greatly from a S. agalactiae vaccine. Investigating how new vaccine technologies, such as subunit vaccines, viral vectors, and replicant mRNA, could provide advantages over traditional bacterins for mastitis vaccines is a crucial area of research. A careful evaluation of why previous vaccine attempts have largely failed would be useful. Recent progress in immunology, in particular cell-mediated immunity, should be taken into account to devise new approaches for development of effective vaccines. The choice of relevant antigens remains a major and challenging step for the developing an effective vaccine. Mapping anti-S. aureus and anti-streptococci T and B cell epitopes and antigens using rational, non-empirical serum immunoproteomics and immunopeptidomics approaches, with a focus on protective (likely subdominant) epitopes. Identifying antigens associated with hosts that mounted successful immune responses (i.e., phenotypically and genotypically resistant dairy cows) to infection could be an intriguing target for effective vaccine development. Investigate whether an immunological imprint (i.e., colonisation or previous infection) further dictates the efficacy of ruminant mastitis vaccines. Understanding the impact of different adjuvants and routes of administration on mastitis vaccine efficacy, particularly through prime-and-pull strategies that target mucosal immunity, is a critical area for further research.

• Therapeutics

  Numerous antimicrobial compounds and formulations are approved for the treatment of mastitis in both lactating and dry cows. Availability, regulation and attitudes towards use of antimicrobials differ between countries. Surveillance of mastitis pathogens and antimicrobial resistance profiles shows different patterns in different countries. In vitro antimicrobial resistance among mastitis-causing pathogens is not the primary reason for treatment failure. Access of antimicrobials to infection foci (sequestration of alveoli by duct clogging, scar tissue barrier) and growth phase (e.g., adherent, intracellular or in biofilm, if biofilm occurs in vivo) may contribute to treatment failure. Late diagnosis of infection, late onset of treatment, and short duration of therapy contribute to poor treatment outcomes. Standard commercial treatment is limited to 2-3 days whereas several scientific studies have shown that cure rates are higher with 5-8 day treatment or a combination of local and systemic antibiotics. The economic justification for such approaches is weak, and they may promote selection for antimicrobial resistance. The concerns regarding antimicrobial resistance (AMR) and antibiotic residues in milk samples are increasing in relation to public health, despite limited evidence of a contribution of bovine mastitis to AMR in human pathogens. Due to these changing attitudes, antimicrobial use quota and a ban on prophylactic use of blanket dry cow treatment have come into place, and there is growing interest in on-farm diagnostics, vaccines, and biologicals for mastitis treatment.

  GAPS

  The distribution and the dynamics of antibiotics in the mammary gland is poorly understood. There is an increased need for surveillance of the spread of antimicrobial resistance in bovine mastitis pathogens, preferably integrated into a One Health framework.

• Biosecurity measures effective as a preventive measure

  Milking time hygiene and monitoring of introduction of dairy animals in herds – see Section “Transmissibility” Within-herd heterogeneity of S. agalactiae (documented in, e.g., South America and China) and S. aureus strains may be associated with biosecurity practices within the herds, such as the purchase of animals for expansion or replacement. For S. aureus, a distinction may need to be made between environmental S. aureus (multiple strains with only one or two animals per strain) and co-circulation of contagious strains (two or more strains with three or more cows per strains).

• Border/trade/movement control sufficient for control

  Not applicable.

• Prevention tools

  Milking time hygiene, environmental hygiene, teat disinfection, culling, segregation and separate milking, intramammary therapy. The main point is that control programs must reduce both the rate of new infections and the duration of existing infections. Antimicrobial treatment of infected udder quarters can reduce pathogen spread within the herd, unless the risk of transmission is very high (exceeds indirect benefit of treatment) or very low (no indirect benefit of treatment). Antimicrobial treatment of subclinical mastitis is not supported in all countries. In dry cows, internal teat sealants can be used to prevent intramammary infection (Mechanical and biological control)

• Surveillance

  Possible at national, bulk milk, cow and quarter level using measures of inflammation (somatic cell count) or infection (culture, PCR). Denmark is the only country known to have a nation-wide surveillance system for S. agalactiae. Surveillance of antimicrobial resistance is possible at herd level, national level, and supranational level (European reporting systems).

  GAP

  Longitudinal, large-scale surveillance programs designed to determine the overall prevalence of mastitis pathogens and bacterial straits, including S. aureus and streptococci-like bacteria, using precise bacterial identification such as MALDI-ToF MS, alongside whole-genome sequencing for phygenetic, resistome, mobilome and virulome analysis

• Past experiences on success (and failures) of prevention, control, eradication in regions outside Europe

  Eradication programs tend to be herd-based and in some herds have been successful, particularly for S. agalactiae (e.g., using blitz-therapy). In other herds, maintaining a low prevalence of disease is the best that can be achieved, especially for S. aureus, S. uberis and S. dysgalactiae. Country-level eradication does not seem to be attainable, although several European countries and Canada have come close to eradication of S. agalactiae from their dairy herds. Because S. agalactiae and S. aureus are human commensals, complete eradication from an inhabited country is not possible. Changes in regulatory limits on bulk milk SCC are reflected in changes in actual BMSCC in developed countries. Uptake of existing control measures is improved when a regulatory incentive is in place, especially in developed countries.

• Costs of above measures

  Dependent on country, herd, disease prevalence and on the approach used, as well as the availability and costs of, e.g., labour and diagnostics, and the price of milk and cull cows. Because many of the control measures detailed above will have benefits beyond the control of staphylococcal and streptococcal mastitis, actual costs can be difficult to discern on a pathogen basis.

• Disease information from the WOAH

• Disease notifiable to the WOAH

  No.

• WOAH disease card available

  N/A

• WOAH Terrestrial Animal Health Code

  N/A

• WOAH Terrestrial Manual

  N/A

• Socio-economic impact

• Zoonosis: impact on affected individuals and/or aggregated DALY figures

  No data

• Zoonosis: cost of treatment and control of the disease in humans

  No data

• Direct impact (a) on production

  Costs associated with mastitis include current and future milk production losses, pharmaceuticals, discarded milk, veterinary services, labour, milk quality deficits, loss of premium price for good milk quality, penalty for poor milk quality, investment in mastitis management materials and infrastructure, diagnostic testing, and cattle replacement. Estimates of the costs associated with cases of clinical and subclinical mastitis vary widely, are limited in the literature, and are somewhat herd and pathogen specific. According to a recent review on the economic effects of bovine mastitis (Sandeberg et al., 2023) the cost per clinical and/or sub-clinical mastitis case in high income countries is between €63-€485. The average cost per case of clinical mastitis due to gram positive pathogens has been estimated as $134USD (Cha et al., 2011). These data underline the important economic impact of mastitis on the dairy industry, but when considering the costs of treating cases of mastitis, it must also be considered that treatment is not uniformly efficacious.

  GAPS

  Cost-benefit data to accompany efficacy trial data for vaccines, treatments, nutritional or management interventions. Lack of data on the cost of mastitis caused by different pathogens.

• Direct impact (b) cost of private and public control measures

  There are some national mastitis control programmes (e.g. Countdown Downunder (Australia), CellCheck (Ireland), Norwegian Mastitis control programme. Denmark is the only country with a specific S. agalactiae control program.

• Indirect impact

  Increasing consumer concern for dairy cow health, antimicrobial usage and welfare. Possible “image” crisis related to multidrug resistant pathogens in case of mention of cow-origin or presence in cows. Milk and milk products can be a source of S. aureus food poisoning. Although most cases are related to contamination during preparation and processing of milk products, milk from infected glands (subclinical mastitis) can be the source of contamination, and European guidelines limit the S. aureus content of milk and raw milk cheeses.

  GAPS

  Enterotoxin-producing S. aureus strains most often contaminate food products during preparation and processing. Consequently, it is important to trace the origin of the contamination to its source, using strain-typing approaches or preferably using WGS.

• Trade implications

• Impact on international trade/exports from the EU

  Varying milk somatic cell count limits worldwide potentially hamper international trade of milk and milk products from countries with higher legal limits to countries with lower legal limits. Examples of legal limits include Europe (Council Directive 92/46/EEC) - 400,000 cells/mL; USA - 750,000 cells/mL; and Brazil - 500,000 cells/mL. Possible impact on the trade of cheese made with raw milk. - Staphylococcal foodborne intoxication (milk products were involved in 1–9 % (mean 4.8 %) of all the incriminated foods) Source: https://food.ec.europa.eu/ As exporter, the EU is not at risk because no trade partner has higher standards for SCC, staphylococcal toxins, or antimicrobial use.

• Impact on EU intra-community trade

  None

• Impact on national trade

  None

• Links to climate

  Seasonal cycle linked to climate

  For contagious transmission, incidence is largely independent of the lactation cycle or nutritional status of animals, with the possible exception of selenium deficiency in heifers, and vector borne transmission. In some countries, e.g., The Netherlands, S. dysgalactiae is more common during the winter housing season and S. uberis is more common during the summer pasture season. It is not known whether this is due to seasonal fluctuation in exposure or host susceptibility, both of which may be affected by diet. Seasonality will differ between management systems and climates.

  GAPS

  The role of diet in shedding of – and susceptibility to S. dysgalactiae and S. uberis is unknown. Influence of heat stress on host susceptibility.

• Distribution of disease or vector linked to climate

  There are regional differences in prevalence of streptococcal and staphylococcal species but there is insufficient data to differentiate the impact of national wealth, technical and economic development of the dairy industry, regulation, cultural, and other factors that may affect distribution and control strategies from direct climate effects.

• Outbreaks linked to extreme weather

  Likely when teat chapping is induced by very low temperatures, or dry conditions. Heat stress is associated with immunosuppression, which could increase susceptibility to new intramammary infections.

  GAP

  Impact of heat stress on susceptibility to gram-positive mastitis.

• Sensitivity of disease or vectors to the effects of global climate change (climate/environment/land use)

  Not identified

• Main perceived obstacles for effective prevention and control

  Sanitary and hygienic measures are labour-intensive and need to be implemented consistently in a market with economic and political pressures and labour shortages. Scientific evidence for efficacy of existing S. aureus and S. uberis vaccines is limited. Pathogen-based diagnostics have limitations, notably limits to sensitivity, limits to specificity (esp. problematic in contaminated samples), and the occurrence of culture-negative/pathogen negative clinical mastitis in a considerable (20-50%) of cases. Lack of breeding programs for a higher resistance to mastitis in some countries. Selection is usually based on SCC (heritability of 10-15%) although in some countries such as Norway selection is also based on clinical mastitis (heritability of 3-5%).

  GAPS

  Lack of socio-economic incentive for uptake of control measures. Devise prevention practices adapted to robotic milking. Lack of cheap, safe, user-friendly and rapid on-farm tests. Difficulty of early detection of new subclinical infections. Implementing effective communication and training programs to change farmers' behaviour and attitudes is crucial for mastitis control. Addressing cultural, social and economic determinants is a significant gap in current control measures, as these factors profoundly influence the adoption of best practices. The challenge lies in overcoming entrenched behaviours and attitudes to achieve long-term improvements in udder health. Significant work in this area has been conducted in some countries (e.g., The Netherlands, UK) but is not universally integrated in development of disease control tools.

• Main perceived facilitators for effective prevention and control

  The application of newer technologies to provide improved diagnostics as well as a better understanding of the immune response are needed to allow the development of improved therapies and vaccines. Importance of regular control of the milk quality by means of somatic cell count or other detection of inflammation. Cultural or socio-economic incentives

  GAPS

  Rapid diagnostic cow-side tests. Effective vaccine. Economic, regulatory or social incentive for uptake of existing control measures. Molecular markers of prognosis and transmissibility.

Risk

• Given the complexities of the host-pathogen interaction and the general lack of efficacy of current treatment and vaccination strategies, there is always the risk of newly developed technologies failing to meet the needs of the dairy industry.

  Regulatory and political environment that is hostile to the development of new antimicrobial agents in veterinary medicine.

Main critical gaps

• • Improved understanding of the immunology of the mammary gland is needed. Current lack of understanding of how strain-to-strain variations (bacterial genetics and virulence factors) and host-pathogen interactions lead to different clinical outcomes. Recent progress in immunology should be taken into account to devise new approaches for development of effective vaccines.
  • Global analysis of the capacity for transmission between different host species including humans and global analysis of the evolution and geographic spread of strains and the identification of new emerging strains, including LA-MRSA.
  • Improved diagnostics: cheap, fast, specific, sensitive, cow-side/in-line
  • More information is needed on genetic basis for resistance or susceptibility to mastitis, with a view to identifying candidate markers or SNPs for selection.
  • Investing in antibacterial discovery programs to discover and develop new, more effective antibacterial agents for the treatment of S. aureus mastitis.

• main critical gaps

  Diagnostics – affordable, sensitive, specific, user-friendly, quick or routine tools to inform early detection and decision making with regards to management of individual animals and herds based on likelihood of transmission and/or cure. This may include
  • improved use of sensor technology, big data and machine learning
  • nucleic acid amplification testing
  • metabolic and/or proteomic fingerprint-based assays that preclude the need for bacterial growth
  • knowledge of molecular markers of virulence, transmissibility or probability of cures.
  Treatment – currently available offerings, especially narrow-spectrum compounds that are not of critical importance to human medicine, are largely fit for purpose, with room for additional treatment options for penicillin-resistant S. aureus mastitis, and for future replacement of antimicrobials with alternatives, e.g.
  • engineered lysogens
  • phage therapy
  Vaccines – require improved understanding of host-pathogen interaction and immune response, including
  • host genetic determinants of pathogen-specific susceptibility to IMI
  • identification of immune responses that lead to clearing of IMI, and associated correlates of protection
  • characterisation of the role of cell-mediated immunity in immunopathology, prevention and cure of IMI
  • mapping of anti-staphylococcal and anti-streptococcal B and T-cell epitopes and antigens using rational, non-empirical approaches and reverse vaccinology with a focus on identifying protective (and likely subdominant) epitopes
  • understanding of the role of immunological imprinting and its impact on the efficacy of mastitis vaccines
  • investigation of the impact of adjuvants and route of administration on vaccine efficacy

Conclusion

• Conclusion summary (s)

  Mastitis in dairy cattle remains a complex and globally significant disease, involving several pathogens such as S. aureus and Streptococcus species (S. agalactiae, S. uberis, S. dysgalactiae). Staphylococcus aureus is a multifaceted pathogen capable of expressing a wide array of virulence factors that allow it to evade both immune responses and antimicrobial treatments. Streptococcus species also play a critical role, especially S. agalactiae, which is known for its contagious nature and involvement in subclinical mastitis and S. uberis which, like S. aureus and S. dysgalactiae, can have contagious as well as environmental modes of transmission. The major gram-positive pathogens share common challenges in terms of diagnostic precision, control measures, and treatment options. Advances in genomic tools such as whole-genome sequencing (WGS) have enabled a better understanding of the genetic diversity of these pathogens, aiding in the development of more targeted epidemiological interventions. Genotyping methods are being employed to distinguish between isolates and define clusters linked to contagiousness, pathogenicity, and treatment success. Despite these advances, the links between genotype and clinical outcomes, such as virulence or contagiousness, remain poorly understood for both S. aureus and streptococci. The combination of genomic insights with improved diagnostic tools and herd management practices could significantly advance mastitis control. Current preventive measures, particularly those aimed at reducing the transmission of S. aureus and S. agalactiae during milking, have shown significant success in countries with a highly developed dairy industry and associated monitoring and animal health systems. However, a more comprehensive understanding of the ecology of both S. aureus and Streptococcus strains, including those residing outside the mammary gland, is needed. The environmental reservoirs for S. aureus, S. uberis and S. dysgalactiae also present challenges, requiring strategies tailored to farm environments. In addition to hygiene management and prevention of transmission, the development of vaccines and antimicrobial treatments remains a priority. The limited efficacy of existing vaccines and the societal requirement to reduce the use of antimicrobials underscore the need for multifaceted approaches, which integrate genomics, immunology, and herd management. Therefore, there is need for development of novel vaccines and insight in the underpinning host-pathogen interactions and protective immunity. Understanding these interactions is crucial for creating vaccines that provide strong and lasting protection, as well as novel host-directed immunotherapy that modify host cellular processes essential for pathogen survival or replication, or that target the host immune response through immunotherapies, either to enhance immunity or reduce immunopathology. Further investment in research is essential to achieve these goals, which includes studying genetic diversity of pathogens, immune mechanisms involved in host-pathogen interactions, refining diagnostic techniques, and developing more effective and long-lasting vaccines. This comprehensive approach will be key to effectively reducing the global burden of mastitis and overcoming the challenges posed by antimicrobial resistance or reduced access to antimicrobials for veterinary use.

  Diagnostics – affordable, sensitive, specific, user-friendly, quick or routine tools to inform early detection and decision making with regards to management of individual animals and herds based on likelihood of transmission and/or cure. This may include

  • improved use of sensor technology, big data and machine learning
  • nucleic acid amplification testing
  • metabolic and/or proteomic fingerprint-based assays that preclude the need for bacterial growth
  • knowledge of molecular markers of virulence, transmissibility or probability of cures
  Diagnostics may not need to be genus-, species- or strain-specific as long as they provide relevant information for on-farm decision making. Treatment – currently available offerings, especially narrow-spectrum compounds that are not of critical importance to human medicine, are largely fit for purpose, with room for additional treatment options for penicillin-resistant S. aureus mastitis, and for future replacement of antimicrobials with alternatives, e.g.
  • engineered lysogens
  • phage therapy
  Vaccines – require improved understanding of host-pathogen interaction and immune response, including
  • host genetic determinants of pathogen-specific susceptibility to IMI
  • identification of immune responses that lead to clearing of IMI, and associated correlates of protection
  • characterisation of the role of cell-mediated immunity in immunopathology, prevention and cure of IMI
  • mapping of anti-staphylococcal and anti-streptococcal B and T-cell epitopes and antigens using rational, non-empirical approaches and reverse vaccinology with a focus on identifying protective (and likely subdominant) epitopes
  • understanding of the role of immunological imprinting and its impact on the efficacy of mastitis vaccines
  • investigation of the impact of adjuvants and route of administration on vaccine efficacy

Sources of information

• Expert group composition

  Expert group members are included where permission has been given:

  1. Fernando Nogueira de Souza, University of São Paulo, Brazil – [Leader]
  2. Ruth Zadoks, The University of Sydney, Australia – [co-leader]
  3. Sarne de Vliegher, Ghent University, Belgium
  4. Orla Keane, Teagasc, Ireland
  5. Vinícius da Silva Duarte, Norwegian University of Life Sciences, Norway
  6. Rui Manuel Santana Zepeda, Hipra
  7. Abhijit Gurjar, Zoetis
  8. Leydson Martins, Vaxxinova

• Reviewed by

  Project Management Board

• Date of submission by expert group

  April 2025

• References

  Recommended citation:

  “Nogueira de Souza F, Zadoks R., De Vliegher S., Keane O., da Silva Duarte V., Santana Zepeda R.M.,Gurjar A., Martins L., 2025. DISCONTOOLS chapter on Staphylococcus aureus and Streptococcus mastitis https://www.discontools.eu/database/69-staphylococcus-aureus-mastitis.html.”