Subclinical Mastitis In Sheep Research Articles

204 Detecting subclinical mastitis in meat-breed ewes using infrared thermography

Abstract Mastitis, the inflammation of the mammary gland most commonly due to bacterial infection, is an important welfare concern in meat-breed sheep. The objective of our study was to evaluate udder surface temperature obtained from infrared thermography as a diagnostic tool for subclinical mastitis. We predicted that udder halves with subclinical mastitis would have greater surface temperatures than healthy udder halves. We enrolled 39 meat-breed ewes (10 Hampshire, 24 Polypay, 5 Targhee; 2 to 9 yr of age) at the University of Wisconsin-Madison sheep unit. All procedures were approved by the UW-Madison Institutional Animal Care and Use Committee. Milk samples from each udder half were collected in duplicate weekly beginning 2 to 4 d postpartum until 8 wk postpartum. Immediately before milk sampling, the udder was photographed from the rear using a thermal camera. A sample was considered a true infection if at least 100 CFU/mL of a bacterial species were isolated. From each photo, we obtained values for minimum, maximum, and mean skin surface temperature for the right and left udders. Three linear mixed models were fit using minimum, maximum, and mean temperatures as the response variables. Fixed effects were infection status (infected or not), weeks postpartum (wk 1-8), and ambient temperature, with a random effect of udder half nested within animal ID. Contrary to our predictions, infection status did not predict udder skin surface temperature (P > 0.12). Healthy udder halves had a mean temperature of 35.5 ± 0.1°C (mean ± SE), minimum temperature of 29.9 ± 0.3°C, and maximum temperature of 37.8 ± 0.1°C. Infected udder halves had a mean temperature of 35.4 ± 0.1°C, minimum temperature of 30.5 ± 0.3°C, and maximum temperature of 37.7 ± 0.1°C. Our results contradict findings in dairy cows, and suggest more research is needed to evaluate the relationship between udder surface temperature and subclinical mastitis in sheep.

Alternative therapeutic approach of ovine subclinical mastitis using the ethanolic roots extract of Capparis spinosa.

Over the past decades, Capparis spinosa has been considered a traditional therapy for relieving different illnesses. Mastitis causes a decrease in milk production and is usually treated with injectable and intra-mammary antibiotics. Investigating the therapeutic effects of C. spinosa root extract on subclinically mastitic ewes. Totally, 164 lactating ewes were selected randomly from the flocks that existed in some areas belonging to Al-Najaf City (Najaf, Iraq) from September to December (2022). Each study animal was subjected to direct sampling of milk before and once each week for 6 weeks (42 days) post treatment to be tested directly by the California mastitis test (CMT). Concerning phytochemical testing of ethanolic root extract, the findings revealed a significant increase in the concentration of alkaloids, flavonoids, polyphenols, and tannins when compared to other components such as coumarins, saponin, glycosides, amino acids, and steroids. In this study, there were 44.51% infected ewes with subclinical mastitis, involving 25.61%, 13.41%, and 5.49% for scores 1, 2, and 3, respectively. In comparison with pre-treatment week, insignificant alteration was seen in the values of all scores in therapeutic week 1. However, significant differences were initiated in values of score 0 in week 2; score 0 and score 2 in week 3; score 0, score 1, and score 2 in week 4; and values of all scores in weeks 5 and 6. This represents the first Iraqi study aimed at the treatment of subclinical mastitis in sheep using the root extract of C. spinosa. Phytochemical testing of ethanolic extract revealed the presence of variable amounts of chemical compounds that reflect their effects on treated animals by decreasing the number of infected ewes with the disease. Moreover, studies are greatly important to estimate the therapeutic effects of other parts of C. spinosa such as leaves and seeds, on the disease and other animal diseases.

Antilisterial activity of raw sheep milk from two native Epirus breeds: Culture-dependent identification, bacteriocin gene detection and primary safety evaluation of the antagonistic LAB biota

Raw milk from native small ruminant breeds in Epirus, Greece, is a valuable natural source of autochthonous lactic acid bacteria (LAB) strains with superior biotechnological properties. In this study, two bulk milks (RM1, RM2) from two local sheep yards, intended for traditional Kefalotyri cheese production, were preselected for bacteriocin-like antilisterial activity by in vitro tests. Their antagonistic LAB biota was quantified followed by polyphasic (16S rRNA gene sequencing; IGS for Enterococcus; a multiplex-PCR for Leuconostoc) identification of 42 LAB (RM1/18; RM2/24) isolates further evaluated for bacteriocin encoding genes and primary safety traits. Representative isolates of the numerically dominant mesophilic LAB were Leuconostoc mesenteroides (10) in both RMs, Streptococcus parauberis (7) in RM2, and Lactococcus lactis (1) in RM1; the subdominant thermophilic LAB isolates were Enterococcus durans (8), E. faecium (6), E. faecalis (3), E. hirae (1), E. hermanniensis (1), Streptococcus lutetiensis (2), S. equinus (1) and S. gallolyticus (1). Based on their rpoB, araA, dsr and sorA profiles, six Ln. mesenteroides strains (8 isolates) were atypical lying between the subspecies mesenteroides and dextranicum, whereas two strains profiled with Ln. mesenteroides subsp. jonggajibkimchii that is first-time reported in Greek dairy food. Two RM1 E. faecium strain biotypes (3 isolates) showed strong, enterocin-mediated antilisterial activity due to entA/entB/entP possession. One E. durans from RM1 possessed entA and entP, while additional nine RM2 isolates of the E. faecium/durans group processed entA or entP singly. All showed direct (cell-associated) antilisterial activity only, as also both S. lutetiensis strains from RM2 did strongly. Desirably, no LAB isolate was β-hemolyrtic, or cytolysin-positive, or possessed vanA, vanB for vancomycin resistance, or agg, espA, hyl, and IS16 virulence genes. However, all three E. faecalis from RM2 possessed gelE and/or ace virulence genes. In conclusion, all Ln. mesenteroides strains, the two safe, enterocin A-B-P-producing E. faecium strains, and the two antilisterial S. lutetiensis strains should be validated further as potential costarter or adjunct cultures in Kefalotyri cheese. The prevalence of α-hemolytic pyogenic streptococci in raw milk, mainly S. parauberis in RM2, requires consideration in respect to subclinical mastitis in sheep and the farm hygiene overall.

Longitudinal Study of Antibiotic Resistance of Staphylococci from Cases of Subclinical Mastitis in Sheep in Greece: Incidence and Risk Factors.

The present paper extends a previous publication on a field study of subclinical mastitis in sheep and focuses on the following laboratory characteristics of the staphylococcal isolates: antibiotic resistance and association with biofilm formation. The specific objectives of the present study were (a) to describe the incidence of isolation of antibiotic-resistant staphylococci from cases of mastitis throughout the milking period in dairy sheep flocks and (b) to identify relevant risk factors, which would contribute to the sustainable control of the infection. Staphylococcal isolates from subclinical mastitis were evaluated for antibiotic resistance to 18 antibiotics. Antibiotic resistance was detected in 57 of the 179 staphylococcal isolates from subclinical mastitis (31.8%). Resistance was recorded against 11 antibiotics, most often against ampicillin (63.2% of resistant isolates), penicillin (63.2%) and tetracycline (47.4%). Isolates resistant to ampicillin and penicillin were recovered in all 12 farms. Twenty-one multidrug-resistant isolates (11.7%) were also recovered. The incidence risk of isolation of staphylococci resistant to at least one (any) antibiotic throughout the study period was 23.8%. The incidence risk of isolation of staphylococci resistant to oxacillin was 5.0%; that of isolation of multidrug-resistant staphylococci was 8.8%. With regard to increased incidence risk of isolation of staphylococci resistant to at least one (any) antibiotic and increased incidence risk of isolation of staphylococci resistant to oxacillin, the omission of anti-staphylococcal mastitis vaccination of ewes emerged as a risk factor. With regard to increased incidence risk of isolation of multidrug-resistant staphylococci, the following variables emerged as risk factors: (a) higher number of antibiotics used on the farm for the treatment of mastitis and (b) younger age of lambs taken away from their dam. Most biofilm-forming antibiotic-resistant staphylococci were recovered from farms where anti-staphylococcal mastitis vaccination was not applied (55.9% versus 44.1% from farms where anti-staphylococcal mastitis vaccination was applied).

Longitudinal Study of Subclinical Mastitis in Sheep in Greece: An Investigation into Incidence Risk, Associations with Milk Quality and Risk Factors of the Infection.

The objectives of this work were (a) to describe the incidence risk of subclinical mastitis in dairy flocks throughout the milking period, (b) to present potential associations of subclinical mastitis with the quality of milk and (c) to identify risk factors for high-incidence risk of the infection. A longitudinal study was performed in 12 dairy sheep flocks in Greece. Mammary secretion samples from 240 ewes and bulk-tank milk samples were collected in four repeated visits over a period of six months for bacteriological, chemical and cytological examinations. Overall, the incidence risk of subclinical mastitis throughout the study period was 51.7%, and it varied among farms from 25.0% to 75.0%. The respective figure for staphylococcal subclinical mastitis was 48.8%. The incidence risk of recurrence of subclinical mastitis among ewes in the flocks was 35.4%. The most frequently identified bacteria from cases of subclinical mastitis were S. aureus and S. simulans; of the mastitis-causing staphylococcal isolates, 65.4% were biofilm-forming. Somatic cell counts in bulk-tank milk progressively increased as the lactation period advanced, with significant increases seen on the third and fourth visits to the farms. Somatic cell counts in bulk-tank milk correlated well with prevalence of subclinical mastitis in flocks. A significant inverse correlation and a significant positive correlation were seen for prevalence of subclinical mastitis versus total protein content or added water in bulk-tank milk. During multivariable analysis, younger age of newborns when taken away from the dam and consequently delayed start of milking of ewes, omission of anti-mastitis vaccination of ewes and lack of employed staff on the farms emerged to be significantly associated (positively) with increased incidence risk of subclinical mastitis.

O-099 Prevalence and aetiology of subclinical mastitis in sheep, as found in an extensive field investigation

O-099 Prevalence and aetiology of subclinical mastitis in sheep, as found in an extensive field investigation

Coagulase Negative Staphylococci Causing Subclinical Mastitis in Sheep: Prevalence, Phenotypic and Genotypic Characterization

Coagulase Negative Staphylococci Causing Subclinical Mastitis in Sheep: Prevalence, Phenotypic and Genotypic Characterization

Use of photodynamic therapy and photobiomodulation as alternatives for microbial control on clinical and subclinical mastitis in sheep.

Evaluate the effects of antimicrobial photodynamic therapy (aPDT) and photobiomodulation (PBM) as alternatives in the treatment of mastitis in sheep. A total of 100 sheep were evaluated, and four teats with clinical mastitis and 16 teats with subclinical mastitis were selected. Milk was collected for isolation and identification of microorganisms. They were grown on TSA, EMB, and MacConkey agar for 24h, and the microorganisms were identified by Gram stain and biochemical tests. The ceilings were subdivided into four groups: G1, treatment with photosensitizer; G2, treatment with PBM (diode laser λ = 660nm); G3, aPDT with methylene blue, and G4, control group. Milk samples were collected before, 24 and 48h after treatments. Cases of subclinical mastitis presented coagulase-negative Staphylococcus and Streptococcus spp, and clinical mastitis had Escherichia coli grow from the samples. The treatments decrease the total bacterial count of negative coagulase Staphylococcus, Streptococcus spp, and Escherichia coli. Comparing the treatments, aPDT stood out, as it was able to photoinactivate all bacteria. Treatment with methylene blue photosensitizer, PBM, and aPDT induced the initial microbial reduction, but aPDT was more effective 48h after treatment.

Molecular and phylogenetic analysis of methicillin resistant Staphylococcus aureus isolated from subclinical mastitis in lactating ewes

This study was conducted for estimation of subclinical mastitis prevalence caused by Staphylococcus aureus (S.aureus) in apparent normal lactating ewes in the areas around Baghdad (Al-Rasheed, Al-Yousifya, Al-Latifya). 400 milk samples from 200 ewes were collected and examined by California mastitis test (CMT), Bacteriological examination was done by culturing on Blood agar, Mannitol salt agar, Staph-110 agar and Chrome agar then bacteria examined by Gram stain and biochemical test. Antibiotic sensitivity test and molecular assay by PCR were done to detect methicillin- resistant strains with the phylogenetic study. The results showed that 49/400 samples gave positive to CMT in the field and positive to S. aureus isolation. The percentage of subclinical mastitis in ewes was 12.25% in this study. All isolated bacteria were resistant to methicillin 100%. The result of PCR revealed that 16.32 % (8/49) of the MRSA were possess mecA gene, while no isolate carried the mecC gene, PCR-product for 8 positive samples of mecA target gene were sequenced, analyzed, and reported in Genbank-NCBI and obtained accession number and became a reference to Iraq and the Middle East and the world. Ongoing work will add to this set more type strains are published and it is available for download at ID: MZ359756.1, M Z359757.1, MZ359758.1, MZ359759.1, MZ359760.1, MZ359761.1, MZ359762.1, MZ359763.1 (mecA gene). Seven samples that match the global results by 100%, and one sample contains a mutation at the site 1539, Nucleotide. Our conclusion revealed that detection of subclinical mastitis in sheep very important caused by S. aureus and all S. aureus was resistant to methicillin and some strains possess mecA gene. However, absence of mec A in other MRSA isolates requires detecting the alternative genetic possibilities related to the resistance profile and fragment of gene used.

Эффективность препарата Мастинол-форте при субклиническом мастите у овец

Эффективность препарата Мастинол-форте при субклиническом мастите у овец

Antimicrobial activity and therapeutic efficacy of thiacycline in subclinical mastitisin sheep

Relevance. The work is devoted to the results of studying of the antimicrobial and therapeutic efficacy of the drug thiacycline in subclinical mastitis in sheep. The aim of our research еsеwas to study the antimicrobial activity and therapeutic efficacy of thiacycline in subclinical mastitis in lactating ewes.Methods. The study of antimicrobial activity was carried out by the method of serial dilutions in a liquid nutrient medium, on the basis of the Caspian zonal NIVI in the laboratory on the study of diseases of non-infectious etiology of farm animals. The therapeutic efficacy of thiacycline was studied in the SEC "Bukhti" of the Gunib district of Dagestan Republic on ewes of the Dagestan rock, aged from 3 to 5 years, with subclinical mastitis, in the amount of 51 heads. Animals on the basis of analogues were divided into two groups: experience (n = 26), control (n = 25). The animals of the experimental group were treated with thiacycline, the drug was administered intramuscularly, once a day, in dose 0.1 ml/kg. The test combination drug thiacycline developed by VIC. The control group was treated with bicillin-3, in dose 600,000 units per head, with an interval 72 hours.Results. The complex antibacterial drug thiacycline has a wide spectrum and high antimicrobial effect against museum strains and potential causative agents of sheep mastitis. The therapeutic efficacy of thiacycline in the treatment of subclinical mastitis in sheep – 97.0%, which is on 13.0% higher compared to bicillin-3. On the 7th day after the last administratin of preparation all the physicochemical parameters of milk corresponded to the physiological norm.

Effect of subclinical mastitis on the yield and cheese-making properties of ewe’s milk

Abstract This review covers an update of scientific knowledge about productive and technological consequences of subclinical mastitis in sheep milk. The literature reports individual milk yield losses of 2.6–43.1 %, being modulated by several factors as infection severity, production level, causal agents, and unilateral or bilateral IMI (1 or 2 infected glands, respectively). A compensatory increase of milk production from the uninfected gland when only one half was infected has been quantified in 6.6 %, compared with healthy halves of control sheep. This compensatory adaptation highlights the risk of underestimating subclinical mastitis in sheep. The mammary gland response is quick and milk yield losses in absolute terms remained constant within the following weeks, both when infection appear during lactation and when it is present from lambing. With respect to the changes on main components in milk due to subclinical mastitis it has been clearly established a decrease in the concentration of lactose and an increase of that of whey proteins. The role played by lactose as an osmotic regulator results in a more accentuated decrease of its concentration in milk. This is why lactose is considered at present as a reliable potential indicator of subclinical mastitis. Whey proteins increase as a result of the increase of the blood-milk barrier permeability and the increased proteolysis of caseins. However, the content in milk of fat and casein are modified depending on the magnitude of milk yield reduction, being affected by a concentration or dilution effect. In any case, the ratio casein to protein (parameter independent of the milk volume) decreases as a result of infection. The impairment of physical and chemical characteristics due to decreased udder health status is the responsible of the negative effect of increased SCC on the coagulation properties of milk, the curd yield and the quality of cheese. Low ratio of casein to protein in high bulk tank SCC milk enhances the extension of the rennet coagulation time (RCT) and curd firming time (k20) because there are more serum proteins and the stability of casein micelles are reduced as a result of hydrolysis. Those changes in turn led to poor syneresis, lower cheese yield, increased moisture content and lower fat and protein content in cheese. Finally, there is a favourable relationship between lactose and milk technological properties because the decrease of this component in case of mastitis is associated with an increase of milk pH. Thus, the three parameters, SCC, pH, and lactose affect, contemporarily and independently, milk quality and coagulation properties, and this is why have been highlighted as potential indicators traits for improving cheese-making ability of sheep milk.

Subclinical mastitis in sheep: etiology and association with milk somatic cell count and ewe productivity in three research flocks in the Western United States.

Subclinical mastitis in sheep: etiology and association with milk somatic cell count and ewe productivity in three research flocks in the Western United States.

Metataxonomic and immunological analysis of milk from ewes with or without a history of mastitis

Mastitis is a highly prevalent condition that has a great impact on milk production and animal welfare, and often requires substantial management efforts. For this reason, it is generally considered an important threat to the dairy industry. Many microbial, host, and environmental factors can protect against, predispose to, or influence the development of mastitis. The objective of this work was to characterize the milk microbiota of Manchega ewes, and to compare samples from animals with and without a history of mastitis. We analyzed milk samples from 36 ewes belonging to 2 different farms (18 ewes from each farm) using culture-dependent and culture-independent techniques. We also analyzed several immune compounds to investigate associations of mastitis with 3 main variables: farm; history of mastitis or no mastitis; and parity number. Both culture-dependent and culture-independent techniques showed that ewe milk harbored a site-specific complex microbiota and microbiome. Staphylococcus epidermidis was the main species driving the difference between farm A (where it was the dominant species) and B (where it was not). In contrast, samples from farm B were characterized by the presence of a wide spectrum of other coagulase-negative staphylococci. Some of these species have already been associated with subclinical intramammary infections in ruminants. Of the 10 immune compounds assayed in this study, 3 were related to a history of mastitis [IL-8, IFN-γ, and IFN-gamma-induced protein 10 (IP-10)]. Increases in IL-8 concentrations in milk seemed to be a feature of subclinical mastitis in sheep, and in this study, this immune factor was detected only in samples from ewes with some episodes of mastitis and from the group with the highest somatic cell count. We also observed a positive correlation between the samples with the highest somatic cell count and IFN-γ and IP-10 levels. Our results suggest that these 3 compounds could be used as biomarkers for the negative selection of mastitis-prone animals, particularly when somatic cell count is very high.

Slime-producing staphylococci as causal agents of subclinical mastitis in sheep

Hitherto, research work in slime production from staphylococcal strains of mastitis origin has focused in laboratory properties of these organisms. Objective of present work was to study subclinical mastitis in sheep, caused specifically by slime-producing staphylococci: to investigate its frequency and to identify potential factors playing a role therein. Slime production was evaluated in 708 staphylococcal isolates recovered from cases of subclinical mastitis in a field study in 2198 ewes performed in an extensive countrywide field investigation across Greece. Isolates were studied by means of microbiological and molecular methods. Of these strains, 262 were characterised as slime-producing, 227 as weak slime-producing and 219 as non slime-producing. Most frequently detected genes were eno and icaB; Staphylococcus aureus possessed more genes than coagulase-negative strains; greater number of genes was detected in slime-producing than in weak slime-producing or non-slime-producing strains. Subclinical mastitis caused specifically by slime-producing staphylococci was detected in 337 ewes: prevalence in population sampled was 0.153. A multivariable mixed-effects model revealed that milking mode (highest prevalence in hand-milked flocks) and flock management system (highest prevalence in semi-intensive flocks) were the two factors associated with increased prevalence of mastitis in flocks. The results confirmed the significance of slime producing staphylococcal strains of importance in the aetiology of subclinical mastitis of sheep. Hand-milking was identified as the most important factor predisposing to that infection.

Detection of quantitative trait loci and putative causal variants affecting somatic cell score in dairy sheep by using a 50K SNP chip and whole-genome sequencing

This study presents a scan of the ovine genome to identify quantitative trait loci (QTL) influencing the somatic cell score (SCS), a classical indicator of subclinical mastitis in sheep, and a subsequent high-resolution analysis of one of the identified QTL regions based on the analysis of whole-genome sequence data sets. A half-sib commercial population of Churra sheep genotyped with a 50K SNP chip was analyzed using linkage analysis (LA) and combined linkage and linkage disequilibrium analysis (LDLA). By LA, 2 5% chromosome-wide significant QTL on OAR5 and OAR25 and one 5% genome-wide significant QTL on ovine chromosome 20 (OAR20) were detected, whereas 22 significant associations were identified by LDLA. Two of the associations detected by LDLA replicated LA-detected effects (OAR20, OAR25). We compared the detected associations with previously reported QTL in sheep and cattle, and functional candidate genes were identified within the estimated confidence intervals. We then performed a high-resolution analysis of the OAR20 QTL region, the most significant QTL region identified by LA that replicated a QTL previously described in Churra sheep for SCS using microsatellite markers. For that, 2 segregating trios of 2 segregating families for the OAR20 QTL (each including the Qq sire and 2 daughters, QQ and qq) were selected for whole-genome sequencing. The bioinformatic analysis of the 6 sequenced samples performed across the genomic interval considered (14.2-41.7 Mb) identified a total of 227,030 variants commonly identified by 2 independent software packages. For the 3 different concordance tests considered, due to discrepancies regarding the QTL peak in the segregating families, the list of mutations concordant with the QTL segregating pattern was processed to identify the variants identified in immune-related genes that show a moderate/high impact on the encoded protein function. Among a list of 85 missense variants concordant with the QTL segregation pattern that were within candidate immune-related genes, 13 variants distributed across 7 genes [PKHD1, NOTCH4, AGER, ENSOARG00000009395 (HLA-C, Homo sapiens), ENSOARG00000015002 (HLA-B, H. sapiens), MOG, and ENSOARG00000018075 (BoLA, Bos taurus, orthologous to human HLA-A] were predicted to cause deleterious effects on protein function. Future studies should assess the possible associations of the candidate variants identified herein in commercial populations with indicator traits of udder inflammation (SCS, clinical mastitis).

Somatic cell count in the diagnosis of subclinical mastitis in sheep of different breeds

The aims of this study were to determine the occurrence of subclinical mastitis in sheep of different breeds and the values for somatic cell count (SCC) in milk for the diagnosis of the disease at lactation and weaning, a fundamental prerequisite for identifying animals in need of control measures. Milk samples were obtained from 1,457 mammary halves of Santa Inês, Texel, Ile de France, and Dorper sheep at two different periods, during the second week of lactation and at weaning. After teats antisepsis, the samples were collected, and identification of the infectious etiology of mastitis and determination of SCC were performed. Microorganisms were identified in 117/762 (15.3%) mammary halves in the second week of lactation and in 86/694 (12.4%) at weaning. Coagulase-negative staphylococci (CoNS) were the etiological agents with the highest incidence alone and in association with other microorganisms, with percentages of 58.1% and 60.6%, respectively. The Santa Inês presented a higher incidence of subclinical mastitis when compared to the other breeds. The cut-off values of SCC for subclinical mastitis were determined at both sampling periods and varied according to stage of lactation, as well breed. These results illustrate the lack of a universal value that can be used for the diagnosis of mastitis and suggests the need for permanent follow-up in herds in order to control the disease.

The Prevalence and Etiology of Subclinical Mastitis in Sheep and Goats

From September 2016 to August 2017, four hundred and twenty four milk samples were collected from mammary halves of 212 apparently healthy lactating animals (108 ewes and 104 does) from small private flocks in different localities at Sharkia Governorate. All samples were investigated by California mastitis test (CMT) and bacteriological examination to determine prevalence and etiology of subclinical mastitis (SCM) in sheep and goats and to estimate the mean somatic cell count (SCC) of infected milk samples with different pathogens. The prevalence rates of SCM using CMT were 44 (40.7%) in ewes [59 milk samples, (27.3%)] and 50 (48.1%) in does [62 milk samples, (29.8%)]. Furthermore, out of ewes’ and does’ milk samples examined by bacteriological culture 54 (25.0%) and 56 (26.9%) samples were bacteriologically positive, respectively. The isolated bacterial causing SCM in ewes’ milk samples were E. coli (44.4%), S. aureus (38.9%), Streptococcus spp. (27.8%), coagulase negative Staphylococci (26.0%), Citrobacter spp. (3.7%) and Enterobacter spp. (1.9%). The identified isolates from does’ milk samples were S. aureus (46.4%), E. coli (26.8%), Streptococcus spp. (25.0%), coagulase negative Staphylococci (19.6%), Citrobacter spp. (5.4%) and Enterobacter spp. (3.6%). The geometric mean of SCC of milk samples harbored major pathogens was significantly (P < 0.05) higher than those harbored minor pathogens. Polymerase Chain Reaction (PCR) was applied on a total of 26 isolates from the most recovered species associated with mastitic milk samples of ewes and does. Suspected S. aureus, E. coli, Strep. agalactiae, Strep. dysagalactiae, and Strep. uberis isolates showed characteristic bands at 270 bp, 366 bp, 487 bp, 279 bp, and 723 bp which were specific for the used genes, respectively. In conclusion: a great attention should be directed to the early diagnosis of SCM in ewes and does.

Extensive countrywide field investigation of subclinical mastitis in sheep in Greece

The objectives of this work were (1) to investigate prevalence of subclinical mastitis, (2) to identify etiological agents involved, and (3) to study factors potentially predisposing ewes to subclinical mastitis. Milk samples were collected from 2,198 ewes in 111 farms with a total population of 35,925 ewes, in all 13 administrative regions of Greece, for bacteriological and cytological examination. Prevalence of subclinical mastitis was 0.260. Main etiological agents were staphylococci (Staphylococcus aureus and coagulase-negative species), which accounted for 0.699 of all isolates recovered; prevalence of staphylococcal mastitis was 0.191. In a multivariable mixed-effects analysis, the primary factor found to be associated with increased prevalence of subclinical mastitis was the management system practiced in flocks (flocks under a semi-intensive system had the highest prevalence). Other factors that were included in the multivariable model were the stage of lactation period (ewes in the 2nd month postpartum showed the highest prevalence) and application of postmilking teat dipping. In contrast, measures taken at the end of a lactation period (e.g., intramammary administration of antimicrobial agents) were not found to have an effect on prevalence of subclinical mastitis. The results confirmed the significance of subclinical mastitis as a frequent problem of ewes, with staphylococci as the primary etiological agent. The findings confirm the multifactorial nature of subclinical mastitis and indicate that its control should rely on many approaches.

Relationship between virulence factor genes in coagulase-negative Staphylococcus spp. and failure of antimicrobial treatment of subclinical mastitis in sheep

ABSTRACT: Coagulase-negative Staphylococcus spp. (CNS) are the main microorganisms involved in ovine mastitis. Treatment at the end of lactation can contribute towards cure and prevention of subclinical cases during the subsequent lactation. However, virulence factors and resistance mechanisms presented by CNS can decrease cure rates. The aims of the study were to identify the species of CNS in milk of mastitic ewes with and without antimicrobial treatment, and to investigate the presence of genes relating to resistance of β-lactam antimicrobials, formation of biofilms, production of enterotoxins and production of the toxic shock syndrome toxin. Cases of failure in the treatment were related with the presence/absence of the respective genes. Sixty sheep were divided into three groups: G1, without treatment; G2, animals treated via the intramammary route with 100mg of cloxacillin during drying off; and G3, sheep treated via the intramammary route with 50 mg of nanoparticulate cloxacillin. Milk samples were gathered during drying off and 15 and 30 days after the parturition of the subsequent lactation. The analyses to identify the species of CNS were carried out by means of the internal transcribe spacer technique and the investigation of the genes responsible for the virulence factors and resistance to oxacillin was performed using the polymerase chain reaction (PCR) technique. No sample was positive for the mecA gene. The only gene relating to production of enterotoxins was sec. Among the genes relating to production of biofilm, icaD was the only one identified in the three experimental groups. Staphylococcus warneri was the main species of CNS isolated during the pre and post-partum periods of the sheep. The species carrying genes relating to production of enterotoxins and biofilms were present in uncured sheep.