Diagnosis of bovine mastitis: from laboratory to farm.

Accurate diagnosis of disease is the major step between the cause and cure of disease. An economical, reliable, and rapid diagnostic tool is fundamental for the management of udder health. The earlier the disease is identified, the less will be the damage; keeping this in mind, many efforts are being made to develop reliable diagnostic tools for use on farm. However, traditional gold standard methods including somatic cell count and microbial culturing are still in use. They are partially being replaced with polymerase chain reaction and sequencing-based tests. Nanotechnology and protein-based tests have also gained lot of attention and some of them are potential candidate of future diagnostic tests for bovine mastitis. Research laboratories are struggling to develop simple, economical, and user-friendly biosensor-based methods that can be performed on farm for rapid diagnosis. The combination of both genomic and proteomic approaches, together with further involvement of nanotheranostic technologies and other sensors, will assist in the quest of better mastitis diagnostic tools.

A novel multiplex PCR assay for simultaneous detection of nine clinically significant bacterial pathogens associated with bovine mastitis.

For rapid and simultaneous detection of nine bovine mastitic pathogens, a sensitive and specific multiplex PCR assay was developed. The assay was standardized using reference strains and validated on mastitic milk cultures which were identified to species level based on 16S rRNA sequencing. Multiplex PCR assay also efficiently detected the target bacterial strains directly from milk. The detection limit of the assay was up to 50 pg for DNA isolated from pure cultures and 104 CFU/ml for spiked milk samples. As estimated by latent class analysis, the assay was sensitive up to 88% and specific up to 98% for targeted mastitic pathogens, compared with the bacterial culture method and the 16S rRNA sequence analysis. This novel molecular assay could be useful for monitoring and maintaining the bovine udder health, ensuring the bacteriological safety of milk, and conducting epidemiological studies.

Causes, types, etiological agents, prevalence, diagnosis, treatment, prevention, effects on human health and future aspects of bovine mastitis.

Mastitis is among the most common and challenging diseases of dairy animals. It is an inflammation of udder tissues due to physical damage, chemical irritation, or infection caused by certain pathogens. Bovine mastitis has been known for ages, but its complex etiology and multi-factorial nature make it difficult to control. Mastitis may have a negative impact on human health by inducing antibiotic-resistant pathogens that may spread, which is threatening. Researchers are continuously struggling to devise suitable methods for mastitis control. Management strategies are mainly focused on disease prevention by farm management which includes proper hygiene, trained staff to monitor minor changes in the udder or milk, and better diagnostic and treatment methods. New technologies which have the potential to unravel this complicated disease include improved diagnostic tools, based on advanced genomics or proteomics, prevention, based on vaccines and immune modulators, and metabolic products of probiotics such as bacteriocins and gene therapy.

Development and validation of a loop-mediated isothermal amplification assay for the detection of Mycoplasma bovis in mastitic milk.

Mycoplasma mastitis is often difficult to control due to a lack of rapid and accurate diagnostic tools. The aim of the current study was to develop a loop-mediated isothermal amplification (LAMP) assay for the detection of Mycoplasma bovis (M. bovis) in mastitic milk. The assay was developed using primers designed for three different target genes: uvrC, 16S rRNA, and gyrB, and validated using mastitic milk samples previously found positive for the target pathogen. Specificity of the developed assay was determined by testing cross-reactivity of LAMP primers against closely related bovine mastitis bacterial pathogens. The sensitivity was found to be higher compared to conventional polymerase chain reaction (PCR). The LAMP assay was also capable of detecting M. bovis in PCR-negative milk samples of cows with clinical mastitis. The uvrC primers were found to be more sensitive, while gyrB primers were more specific; however, 16S rRNA primers were less specific and sensitive compared to either uvrC or gyrB primers. Cohen's kappa values for uvrC, gyrB, and 16S rRNA primers used in the LAMP assays were 0.940, 0.970, and 0.807, respectively. There was a high level of agreement between the test results and the true-disease status as indicated by the receiver operating characteristic (ROC) curve. Our findings suggest that the newly developed LAMP assays targeting the uvrC and gyrB genes could be a useful tool for rapid and accurate diagnosis of mastitis caused by M. bovis.