Detailed data from experimentally-induced mastitis in ewes, with the aim to evaluate cathelicidin-1 in milk

Bacteriological, cytological and proteomics data have been obtained from ewes in two experiments, after intramammary challenge with Mannheimia haemolytica or Staphylococcus chromogenes. Animals were sampled before and sequentially after challenge. Conventional techniques were employed for bacterial isolation and somatic cell counting in milk samples; milk whey samples were subjected to proteomics evaluation by using two-dimensional gel electrophoresis and MALDI-TOF mass spectrometry. There was a correlation between leucocyte content and cathelicidin-1 spot densities in milk samples, although the protein was detected in milk earlier than the increase in leucocyte content. There was also a significant association between presence of mastitis in a mammary gland and detection of cathelicidin-1 in the respective milk sample; the degree of association was greater during the first 24 h post-inoculation. The data are further discussed in the research article “Detection of cathelicidin-1 in the milk as an early indicator of mastitis in ewes” [1].


a b s t r a c t
Bacteriological, cytological and proteomics data have been obtained from ewes in two experiments, after intramammary challenge with Mannheimia haemolytica or Staphylococcus chromogenes. Animals were sampled before and sequentially after challenge. Conventional techniques were employed for bacterial isolation and somatic cell counting in milk samples; milk whey samples were subjected to proteomics evaluation by using twodimensional gel electrophoresis and MALDI-TOF mass spectrometry. There was a correlation between leucocyte content and cathelicidin-1 spot densities in milk samples, although the protein was detected in milk earlier than the increase in leucocyte content. There was also a significant association between presence of mastitis in a mammary gland and detection of cathelicidin-1 in the respective milk sample; the degree of association was greater during the first 24 h post-inoculation. The data are further discussed in the research article "Detection of cathelicidin-1 in the milk as an early indicator of mastitis in ewes" [1]. © 2020 The Author(s). Published by Elsevier Inc. This is an open access article under the CC BY license (http://creativecommons. org/licenses/by/4.0/).

Data
In two experiments, we performed intramammary challenge of ewes with Mannheimia haemolytica or Staphylococcus chromogenes; subsequently, mastitis was induced [1], as confirmed by clinical, microbiological and cytological findings (Tables 1e4). Presence of cathelicidin-1 in milk was also evaluated (Tables 5 and 6, Figs. 1e4).
The data have provided evidence of a significant association between cell content and detection of cathelicidin-1 in respective milk sample (Figs. 5 and 6), as well as between presence of mastitis in a mammary gland and detection of cathelicidin-1 in the respective milk sample (Table 7). The association was stronger in samples collected during the first 24 h post-inoculation than in samples collected thereafter (Table 8). There was a slight increase in cathelicidin-1 levels, when a higher challenge dose of M. haemolytica was used (Experiment I).
Data indicated a correlation between CMT scores and cathelicidin-1 spot densities in milk samples: the correlation coefficient for both experiments was r ¼ 0.398 (P < 0.001); the respective values for Specifications  Value of the Data This is the only dataset available from experimentally induced mastitis, detailing cathelicidin-1 presence in milk, with early start of monitoring post-challenge, to fully evaluate the course of cathelicidin-1 presence in milk. The data can be used by researchers working in the development of diagnostic techniques for mastitis, based on detection of cathelicidin-1 in milk.
i. s.: inoculated side of the udder, c. s.: uninoculated side of the udder. þ: isolation of the challenge pathogen, -: no bacterial isolation. experiments 1 and 2 were r ¼ 0.272 (P ¼ 0.023) and r ¼ 0.540 (P < 0.001). In experiment 2, the correlation coefficients when data from ewes inoculated with M. haemolytica or S. chromogenes were taken separately were r ¼ 0.604 and 0.704 (P < 0.001), respectively. There was also evidence of correlation between somatic cell counts and cathelicidin-1 spot densities in milk samples in experiment 2. The correlation coefficient was r ¼ 0.565 (P < 0.001).  Mastitis definition: mastitis was defined in ewes with (i) clinically evident abnormalities in mammary gland or mammary secretion or (ii) with no clinical abnormalities, but in which a bacteriologically positive milk sample with concurrently increased cell content (CMT score ! 'l' or cell counts ! 0.5 Â 106 cells mL-1) plus increased neutrophil and lymphocyte proportion (!65% of all leucocytes) in Giemsa-stained milk films was detected.

Experimental design, materials and methods
In experiment I, M. haemolytica (1000e1250 c.f.u.) was deposited into the teat duct of ewes (n ¼ 5) on Day 0 (D0). In experiment II, M. haemolytica (50e80 c.f.u.) or S. chromogenes (1 Â 10 6 e2 Â 10 6 c.f.u) was inoculated into in the gland cistern of ewes (n ¼ 3 for each pathogen) also on D0. In all cases, mastitis was induced, as confirmed by microbiological and cytological examination of milk samples, which were collected on D0 þ 12 h, D1, D2, D3 and D4 (experiment 1) or on D0 þ 3 h, D0 þ 6 h, D0 þ 9 h, D0 þ 12 h and D1 (experiment 2). The uninoculated mammary gland (contralateral) was used as uninfected control. Increased cell content and recovery of the challenge pathogens were simultaneously recorded. Milk whey prepared from the samples was processed for proteomics examination.
Proteomics analysis for detection of cathelicidin-1 was performed as detailed by . Two-dimensional gel electrophoresis was used initially.
In experiment 1, image analysis was performed as detailed by  and included all the surface of each gel; spots corresponding to cathelicidin-1 were identified. In experiment 2, image analysis was limited in the region of each gel, where cathelicidin-1 had been located during experiment 1. Spot optical densities obtained from PD Quest v.8.0 for each spot of interest on each gel on D0 or sequentially after challenge, were recorded. In case of multiple spots indicative of the same protein, densities of all spots were taken into account. The spot volume was used as the analysis parametre to quantify protein expression.