Haptoglobin and lactate dehydrogenase measurements in milk for the identification of subclinically diseased udder quarters

Diagnosis of subclinical mastitis is of increasing importance and appropriate detection methods are needed. Both haptoglobin (Hp), an acute phase protein in cattle, as well as lactate dehydrogenase (LDH), an ubiquitous enzyme, can be successfully used to detect clinical mastitis. The present paper describes quantification of Hp and LDH in milk samples from healthy and subclinically diseased udder quarters. Hp was analysed in the laboratory using an ELISA. The activity of LDH was measured in raw milk directly in the milking parlor. Both parameters were suitable to distinguish between sterile samples and bacteriologically positive samples. The ability to differentiate between minor and major pathogens was better for Hp in skim milk than for LDH in raw milk. Hp and somatic cell count (SCC) as well as LDH and SCC were positively correlated (r = 0.8 and r = 0.76, respectively). Subclinical mastitis was defined as follows: SCC > 100 × 103 cells/ml and bacteriological positive findings in two out of three weekly samples. Sensitivity and specificity were above 85% for Hp and above 81% for LDH. Using a less rigid classification to define mastitis, i.e. SCC < 200 × 103 cells/ml and two out of three weekly samples bacteriologically positive, sensitivity for Hp improved (89%) and remained unchanged for LDH. Both parameters are useful parameters for the diagnosis of subclinical mastitis. LDH activity in raw milk was less sensitive and specific than Hp but the method described herein offers the opportunity to measure LDH activity directly in the milking parlor and might therefore be suitable for on-line system developments.

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Using dairy herd improvement records and clinical mastitis history to identify subclinical mastitis infections at dry-off

Journal of Dairy Research ◽

10.1017/s0022029908003257 ◽

2008 ◽

Vol 75 (2) ◽

pp. 240-247 ◽

Cited By ~ 21

Author(s):

Audrey H Torres ◽

Päivi J Rajala-Schultz ◽

Fred J DeGraves ◽

Kent H Hoblet

Keyword(s):

Dairy Herd ◽

Cost Effective ◽

Subclinical Mastitis ◽

Clinical Mastitis ◽

Somatic Cell Counts ◽

Dry Cow Therapy ◽

Cell Counts ◽

On Farm ◽

First 90 Days ◽

Major Pathogens

Interest in selective dry cow therapy (SDCT) has been increasing owing to concerns over development of antimicrobial resistance. Implementation of SDCT, however, requires a quick and cost-effective on-farm method for identifying cows for treatment and cows that can be left without treatment. The objective of the present study was to evaluate the use of clinical mastitis (CM) history and somatic cell counts (SCC) from monthly Dairy Herd Improvement (DHI) records in identification of infected and uninfected cows at dry-off. A total of 647 Holstein cows were classified as uninfected or infected at dry-off based on CM history and varying number of monthly SCC records (with three different SCC cut-offs). Cows were considered uninfected based on the following criteria: (1) SCC <100 000 cells/ml and no CM during the lactation; (2) SCC <200 000 cells/ml and no CM during the lactation; (3) as criterion two, but additionally a cow was also considered uninfected if it experienced a case of CM during the first 3 months of the lactation and the SCC was <100 000 cells/ml for the rest of the lactation; (4) SCC <300 000 cells/ml and no CM during the lactation; otherwise they were considered infected. Infected and uninfected cows at dry-off were most efficiently identified using three months' SCC records with a threshold of 200 000 cells/ml for cows without CM during the lactation and a threshold of 100 000 cells/ml during the rest of lactation for cows with CM during the first 90 days in milk. Moreover, this criterion also most efficiently identified cows infected with major pathogens only at dry-off. The success of the criteria used for identifying infected and uninfected cows will, however, depend on herd characteristics, such as prevalence of infection and type of pathogens present in the herd.

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Virulence Determinants and Plasmid-Mediated Colistin Resistance mcr Genes in Gram-Negative Bacteria Isolated From Bovine Milk

Frontiers in Cellular and Infection Microbiology ◽

10.3389/fcimb.2021.761417 ◽

2021 ◽

Vol 11 ◽

Author(s):

Yasmine H. Tartor ◽

Rasha M. A. Gharieb ◽

Norhan K. Abd El-Aziz ◽

Hend M. El Damaty ◽

Shymaa Enany ◽

...

Keyword(s):

Virulence Genes ◽

Raw Milk ◽

Subclinical Mastitis ◽

Clinical Mastitis ◽

Gram Negative Bacteria ◽

Colistin Resistance ◽

Genetic Characteristics ◽

Gram Negative ◽

E Coli ◽

Resistant Strains

A major increase of bacterial resistance to colistin, a last-resort treatment for severe infections, was observed globally. Using colistin in livestock rearing is believed to be the ground of mobilized colistin resistance (mcr) gene circulation and is of crucial concern to public health. This study aimed to determine the frequency and virulence characteristics of colistin-resistant Gram-negative bacteria from the milk of mastitic cows and raw unpasteurized milk in Egypt. One hundred and seventeen strains belonging to Enterobacteriaceae (n = 90), Pseudomonas aeruginosa (n = 10), and Aeromonas hydrophila (n = 17) were screened for colistin resistance by antimicrobial susceptibility testing. The genetic characteristics of colistin-resistant strains were investigated for mcr-1–9 genes, phylogenetic groups, and virulence genes. Moreover, we evaluated four commonly used biocides in dairy farms for teat disinfection toward colistin-resistant strains. Multidrug-resistant (MDR) and extensive drug-resistant (XDR) phenotypes were detected in 82.91% (97/117) and 3.42% (4/117) of the isolates, respectively. Of the 117 tested isolates, 61 (52.14%) were colistin resistant (MIC >2 mg/L), distributed as 24/70 (34.29%) from clinical mastitis, 10/11 (90.91%) from subclinical mastitis, and 27/36 (75%) from raw milk. Of these 61 colistin-resistant isolates, 47 (19 from clinical mastitis, 8 from subclinical mastitis, and 20 from raw milk) harbored plasmid-borne mcr genes. The mcr-1 gene was identified in 31.91%, mcr-2 in 29.79%, mcr-3 in 34.04%, and each of mcr-4 and mcr-7 in 2.13% of the colistin-resistant isolates. Among these isolates, 42.55% (20/47) were E. coli, 21.28% (10/47) A. hydrophila, 19.12% (9/47) K. pneumoniae, and 17.02% (8/47) P. aeruginosa. This is the first report of mcr-3 and mcr-7 in P. aeruginosa. Conjugation experiments using the broth-mating technique showed successful transfer of colistin resistance to E. coli J53-recipient strain. Different combinations of virulence genes were observed among colistin-resistant isolates with almost all isolates harboring genes. Hydrogen peroxide has the best efficiency against all bacterial isolates even at a low concentration (10%). In conclusion, the dissemination of mobile colistin resistance mcr gene and its variants between MDR- and XDR-virulent Gram-negative isolates from dairy cattle confirms the spread of mcr genes at all levels; animals, humans, and environmental, and heralds the penetration of the last-resort antimicrobial against MDR bacteria. Consequently, a decision to ban colistin in food animals is urgently required to fight XDR and MDR bacteria.

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ЕТІОЛОГІЧНІ ЧИННИКИ МАСТИТІВ КОРІВ УКРАЇНСЬКОЇ ЧОРНО–РЯБОЇ МОЛОЧНОЇ ПОРОДИ

Scientific Messenger of LNU of Veterinary Medicine and Biotechnology ◽

10.15421/nvlvet7046 ◽

2016 ◽

Vol 18 (3(70)) ◽

pp. 191-196

Author(s):

V. Panevnyk ◽

T. Suprovych

Keyword(s):

Staphylococcus Aureus ◽

Somatic Cells ◽

Raw Milk ◽

Subclinical Mastitis ◽

Index Number ◽

Clinical Form ◽

Leading Role ◽

Skin Contamination ◽

Major Pathogens ◽

And Staphylococcus Aureus

The article shows data on the microbial landscape and quantity of somatic cells milk in different forms of mastitis in cows. Index number of somatic cells (SCC) in the raw milk of cows in the country is only used to establish the quality milk. They are key safety indicators that are directly related to udder cow disease, especially subclinical mastitis. Research has established that the number of SCC in healthy animals ranges from 84000 cells/ml to 436000 cells/ml. Over the course of subclinical turned from 508000 cells/ml to 756000 cells/ml. Animals with clinical form of mastitis were from 876000 cells/ml to 69260000 cells/ml. The 42 cows of the first lactation average SCC was 143000 cells/ml, and 47 of fifth lactation cows – 213000 cells/ml.The microflora in the breast can get in different ways: galactogenous – through teat channel hematogenous ahd lymphogenous ways. The leading role galactogenous way in which the pathogens penetrate from the environment through teat channel. This contributes to the udder skin contamination by microorganisms. Activators of subclinical mastitis were Staphylococcus aureus 31.8% and Streptococcus agalactiae 40.9%. In the clinical course of mastitis major pathogens were Escherichia coli – 34.8% and Staphylococcus aureus – 41.3%. Selected cultures of microorganisms were sensitive to cephalexin, gentamicin, oxacillin, rifampicin, enrofloksacin.

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L-lactate dehydrogenase and N-acetyl-β-D-glucosaminidase activities in bovine milk as indicators of non-specific mastitis

Journal of Dairy Research ◽

10.1017/s0022029906001956 ◽

2006 ◽

Vol 73 (4) ◽

pp. 431-440 ◽

Cited By ~ 56

Author(s):

Mizeck GG Chagunda ◽

Torben Larsen ◽

Martin Bjerring ◽

Klaus L Ingvartsen

Keyword(s):

Lactate Dehydrogenase ◽

Post Partum ◽

Bovine Milk ◽

Clinical Signs ◽

Threshold Value ◽

Clinical Mastitis ◽

Factors Affecting ◽

Udder Health ◽

Ldh Activity ◽

High Level

Systematic factors affecting the activities of L-lactate dehydrogenase (LDH) and N-acetyl-β-D-glucosaminidase (NAGase) and somatic cell count (SCC), the association between the activities of LDH and NAGase and SCC with respect to udder health status, and the ability of LDH and NAGase to classify cows in udder health categories for early detection of mastitis were studied. A dataset of records from 74 Danish Holstein, 76 Danish Red and 47 Jersey cows on one research farm was used. Cows were grouped into healthy and clinically mastitic. A healthy cow was defined as having no veterinary treatment and SCC<100000 cells/ml. A clinically infected cow was one receiving veterinary treatment after showing clinical signs of mastitis and SCC >800000 cells/ml. Breed, month of production, and days in milk significantly influenced (P<0·001) LDH activity, NAGase activity and SCC in both healthy and clinically mastitic cows. In healthy cows, LDH activity, NAGase activity and SCC started at a high level immediately after calving and decreased to low levels approximately 30–40 d post partum. All the three parameters increased due to clinical mastitis. NAGase activity had numerically higher variation in healthy cows than in clinically mastitic cows (CV=56·2% v. CV=53·5%). The relationship between LDH activity and SCC was stronger in milk from clinically mastitic than from healthy cows (r=0·76 v. r=0·48 and r=0·67 v. r=0·44 for correlation of observed values and residuals, respectively). LDH activity had higher sensitivity than NAGase activity (73–95% v. 35–77%) while specificities were in a similar range (92–99%). Further, sensitivities for LDH activity were more robust to changes in the threshold value than those for NAGase activity. Opportunities for automated, in-line real-time mastitis detection are discussed.

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Determination of lactate dehydrogenase (LDH) activity in milk by a fluorometric assay

Journal of Dairy Research ◽

10.1017/s0022029905000865 ◽

2005 ◽

Vol 72 (2) ◽

pp. 209-216 ◽

Cited By ~ 37

Author(s):

Torben Larsen

Keyword(s):

Lactate Dehydrogenase ◽

Large Scale ◽

Raw Milk ◽

Milk Samples ◽

Ldh Activity ◽

Straight Line ◽

Methodological Problems ◽

Assay Precision ◽

Detailed Kinetics

Indigenous L-lactate dehydrogenase (LDH) in milk originates mainly from somatic cells, leucocytes and invading microorganisms. Its activity may be used for detection of mastitis. However, existing methods to measure LDH activity in milk both need pretreatment of the samples and still suffer from methodological problems. The present paper describes a fast, reliable method for determination of LDH activity, suitable for milk samples. The method is based on fluorometric determination of enzyme kinetics when L-lactate is converted to pyruvate. The assay uses raw milk without pretreatment and the method is easily adjustable to large-scale analyses on micro assay plates. Detection is based on (straight line) linear response within 4–7 min of initiation of the reaction. A substrate concentration of 35 mM in the reaction mixture was considered to be optimal for the assay. Intra plate assay precision was approx. 6% (CV) and the inter plate precision approx. 10%. Known inhibitors of LDH activity (oxidative direction), i.e., oxalic acid, oxamate, and pyruvate, were tested in different concentrations in order to verify the specificity of the response. The detailed kinetics of samples analysed indicated that the isoenzyme composition may have differed between milk samples, and that this composition may have been altered in high activity samples.

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Changes in the inline lactate dehydrogenase according to the cow’s production and reproduction status

Acta Veterinaria Brno ◽

10.2754/avb201988040369 ◽

2019 ◽

Vol 88 (4) ◽

pp. 369-375

Author(s):

Dovilė Malašauskienė ◽

Vida Juozaitienė ◽

Mindaugas Televičius ◽

Arūnas Rutkauskas ◽

Mingaudas Urbutis ◽

...

Keyword(s):

Lactate Dehydrogenase ◽

Dairy Cows ◽

Milk Yield ◽

High Sensitivity ◽

Cost Effective ◽

Subclinical Mastitis ◽

Milk Productivity ◽

Somatic Cell Counts ◽

Cell Counts ◽

Ldh Activity

The aim of the present study was to investigate inline lactate dehydrogenase (LDH) changes in clinically healthy dairy cows according to the number and stage of lactations, milk yield, and the reproduction status The LDH activity (μmol/min per litre) was measured using the dry-stick technology. A total of 378 cows were selected and classified according to their reproductive status into the following groups: fresh (1–44 days after calving); open (45–65 days after calving); inseminated (1–35 days after insemination); pregnant (35–60 days after insemination). According to their milk productivity, the cows were classified into the following groups: <15 kg/d, 15–25 kg/d, >25–35 kg/d, and >35 kg/d. They were milked with a DeLaval milking robot in combination with a Herd Navigator analyser. The results showed that the inline LDH concentration had a tendency to increase along with the increase in the number of lactation periods (P < 0.05). The highest level of LDH was observed in fresh cows 5–10 days in milk (DIM), and the highest LDH concentration was found in the milk of fresh cows. A positive statistically reliable relationship was found between the milk yield and LDH concentration (P < 0.05); LDH and milk somatic cell counts (SCC) were positively related in all groups of cows, although LDH concentration and SCC were the highest correlated variables in inseminated cows (P < 0.05). The present study shows that measuring LDH activity in milk is both easy and cost effective with high sensitivity and specificity, having a great potential as a diagnostic tool for detection of subclinical mastitis.

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Evaluation of the on-line electrical conductivity of milk in mastitic dairy cows

Acta Veterinaria Hungarica ◽

10.1556/avet.2012.012 ◽

2012 ◽

Vol 60 (1) ◽

pp. 145-155 ◽

Cited By ~ 5

Author(s):

András Gáspárdy ◽

Gil Ismach ◽

Árpád Bajcsy ◽

Gyula Veress ◽

Szilárd Márkus ◽

...

Keyword(s):

Electrical Conductivity ◽

Dairy Cows ◽

Normal Level ◽

Inflammatory Reaction ◽

Somatic Cell Count ◽

Large Scale ◽

Dairy Farm ◽

Subclinical Mastitis ◽

Clinical Mastitis ◽

On Line

Mastitis is a persistent, inflammatory reaction of the udder tissue, which entails a decline in potassium, and is also responsible for a higher somatic cell count (SCC) and electrical conductivity (EC) of milk. The measurement of EC is an indirect, rapid method to detect subclinical mastitis from milk. The aim of this study was to analyse the EC of milk throughout the lactation, around the day of mastitis detection, and also to estimate its heritability based on data from a large-scale dairy farm. Shortly after calving the EC value generally decreases; however, it was discovered that from the thirteenth week onwards, substantial differences arise between the mastitic and healthy groups of cows. The authors observed a significant (P < 0.001) increase in EC before the detection of clinical mastitis. This higher value (around 11 mS) persisted for 4 days, then it gradually returned to the normal level. The EC of milk from daughters sired by different bulls responded differently in case of mastitis. The heritability of EC seems to be high (h2 = 0.56). Therefore, the EC trait can be a beneficial indicator in detecting mastitis and should be considered in sire selection.

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Characterization of Escherichia coli Strains Derived from Cow Milk of Subclinical and Clinical Cases of Mastitis

Applied Sciences ◽

10.3390/app11020541 ◽

2021 ◽

Vol 11 (2) ◽

pp. 541

Author(s):

Katarzyna Grudlewska-Buda ◽

Krzysztof Skowron ◽

Ewa Wałecka-Zacharska ◽

Natalia Wiktorczyk-Kapischke ◽

Jarosław Bystroń ◽

...

Keyword(s):

Escherichia Coli ◽

Bacterial Species ◽

Subclinical Mastitis ◽

Clinical Mastitis ◽

Economic Problem ◽

Cow Milk ◽

P Value ◽

Dairy Herds ◽

E Coli ◽

Bonferroni Test

Mastitis is a major economic problem in dairy herds, as it might decrease fertility, and negatively affect milk quality and milk yield. Out of over 150 bacterial species responsible for the udder inflammation, Escherichia coli is one of the most notable. This study aimed to assess antimicrobial susceptibility, resistance to dipping agents and biofilm formation of 150 E. coli strains isolated from milk of cows with subclinical and clinical mastitis. The strains came from three dairy herds located in Northern and Central Poland. The statistical analyses were performed with post-hoc Bonferroni test and chi-square test (including Yates correction). The data with a p value of <0.05 were considered significant. We found that the tested strains were mostly sensitive to antimicrobials and dipping agents. It was shown that 37.33% and 4.67% of strains were resistant and moderately resistant to at least one antimicrobial agent, respectively. No extended-spectrum beta-lactamases (ESBL)-producing E. coli were detected. The majority of strains did not possess the ability to form biofilm or formed a weak biofilm. The strong biofilm formers were found only among strains derived from cows with subclinical mastitis. The lowest bacteria number was noted for subclinical mastitis cows’ strains, after stabilization with iodine (3.77 log CFU × cm−2) and chlorhexidine (3.96 log CFU × cm−2) treatment. In the present study, no statistically significant differences in susceptibility to antibiotics and the ability to form biofilm were found among the strains isolated from cows with subclinical and clinical mastitis. Despite this, infections in dairy herds should be monitored. Limiting the spread of bacteria and characterizing the most common etiological factors would allow proper treatment.

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