Effect of a pre-milking teat foam and a liner disinfectant on the presence of mesophilic and (proteolytic) psychrotrophic bacteria prior to milking

Contamination of raw milk by psychrotrophs can lead to the production of heat-resistant proteases and subsequent spoilage of UHT milk. Therefore, this research communication evaluated the effect of a pre-milking teat disinfectant (active components: L-(+)-lactic acid and salicylic acid) and a liner disinfectant (active components: peracetic acid and hydrogen peroxide) on the number of mesophilic and (proteolytic) psychrotrophic bacteria prior to milking. The teat orifices of 10 cows were sampled using a swabbing procedure before and after treatment with a pre-milking teat disinfectant on six subsequent days. On the teat orifices, there was a small but statistically significant decrease in the psychrotrophic bacterial counts between pre and post dipping. No differences were observed for the mesophilic bacterial counts and proteolytic active counts. Liners were also sampled using swabs pre and post disinfection. No statistically significant decrease in the bacterial counts was observed post liner disinfection, although there was a numerical decrease. Sixty-two percent of the proteolytic psychrotrophs were pseudomonads: 16.5% of which were P. fragi, 14.3% P. lundensis, 10.0% P. fluorescens and 2.9% P. putida. Trinitrobenzenesulfonic acid (TNBS) analysis revealed a wide variety in proteolytic activity (from 0 to 55 mu mol glycine/ml milk) and the presence of high producers. It can be concluded that there was only a minor effect of teat and liner disinfection on the psychrotrophic bacterial counts indicating that the measures presented did not result in a reduction of the targeted bacteria on teat orifices and liners

Reduction of Mycobacterium avium ssp. paratuberculosis in colostrum : development and validation of 2 methods, one based on curdling and one based on centrifugation

The aim of this study was to develop and validate 2 protocols (for use on-farm and at a central location) for the reduction of Mycobacterium. avium ssp. paratuberculosis (MAP) in colostrum while preserving beneficial immunoglobulins (IgG). The on-farm protocol was based on curdling of the colostrum, where the IgG remain in the whey and the MAP bacteria are trapped in the curd. First, the colostrum was diluted with water (2 volumes colostrum to 1 volume water) and 2% rennet was added. After incubation (1 11 at 32 degrees C), the curd was cut and incubated again, after which whey and curd were separated using a cheesecloth. The curd was removed and milk powder was added to the whey. Approximately 1 log reduction in MAP counts was achieved. A reduction in total proteins and IgG was observed due to initial dilution of the colostrum. After curd formation, more than 95% of the immunoglobulins remained in the whey fraction. The semi-industrial protocol was based on centrifugation, which causes MAP to precipitate, while the IgG remain in the supernatant. This protocol was first developed in the laboratory. The colostrum was diluted with skimmed colostrum (2 volumes colostrum to 1 volume skimmed colostrum), then skimmed and centrifuged (at 15,600 x g for 30 min at room temperature). We observed on average 1.5 log reduction in the MAP counts and a limited reduction in proteins and IgG in the supernatant. To obtain a semi-industrial protocol, dairy pilot appliances were evaluated and the following changes were applied to the protocol: after 2:1 dilution as above, the colostrum was skimmed and subsequently clarified, after which the cream was heat treated and added to the supernatant. To investigate the effect of the colostrum treatment on the nutritional value and palatability of the colostrum and the IgG transfer, an animal experiment was conducted with 24 calves. Six received the dam's colostrum, 6 were given untreated purchased colostrum (control), and 2 groups of 6 calves received colostrum treated according to both of the above-mentioned methods. No significant differences were found between the test groups and the dam's colostrum group in terms of animal health, IgG uptake in the blood serum, milk; or forage uptake. Two protocols to reduce MAP in colostrum (for use on-farm or at a central location) were developed. Both methods preserve the vital IgG

Application of LC-HRMS identified marker peptides in an LC-MS/MS method for detection and quantification of heat-resistant proteolytic activity in raw milk

For the dairy industry, it would be useful to be able to determine the heat-resistant proteolytic activity of the raw milk upon arrival at the plant, preferably before processing (within 24 h of arrival). This would allow producers to choose the most optimal processing of the raw milk and hence its fate, together with a more accurate prediction of shelf-life. A liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the detection and quantification of Pseudomonas-derived heat-resistant proteolytic activity in raw milk based on three new marker peptides, identified using liquid chromatography coupled to high-resolution mass spectrometry (LC-HRMS), was developed. This method involved a sample preparation where existing peptides are removed from the milk sample, after which the heat-resistant protease-containing fraction is mixed with a casein substrate. After incubation, the increase of the peptide markers is determined using an LC-MS/MS method. The limit of detection (LOD) and limit of quantification (LOQ) of this method were determined using synthetically produced peptides (with an average LOD between 0.1 and 5 ng/mL and an average LOQ between 0.1 and 10 ng/mL)

Preliminary study of the effect of sow washing, as performed on the farm, on livestock-associated methicillin-resistant Staphylococcus aureus skin status and strain diversity

Washing sows (n = 12 per herd) on four Belgian pig farms positive for methicillin-resistant Staphylococcus aureus (MRSA) had no significant effect on MRSA status of the sow's skin (P = .32) or nares (P = 1.00). In 64% of cases, the same strain was detected before and after washing

Methicillin-resistant Staphylococcus aureus (MRSA) ST398 in pig farms and multispecies farms

During the last few years, methicillin-resistant Staphylococcus aureus (MRSA) ST398 has been isolated frequently from livestock, especially from pigs and to a lesser extent from cattle and poultry. To gain insight into the distribution of this bacterium in pig farms versus multispecies farms, 30 Belgian farms (10 pig, 10 pig/poultry and 10 pig/cattle farms) were screened for the presence of MRSA. On each farm, 10 nasal swabs were taken from pigs. When present, cattle (n = 10) were sampled in the nares and poultry (n = 10) in the nares, earlobes and cloaca. A selection of the obtained isolates were further characterized using multilocus sequence typing (MLST), spa typing, SCCmec typing, pulsed field gel electrophoresis (PFGE), multiple-locus variable-number tandem repeat analysis (MLVA) and antimicrobial susceptibility testing. On 26 of 30 farms, MRSA was isolated from pigs. Furthermore, MRSA was also isolated from poultry and cattle on one pig/poultry and five pig/cattle farms, respectively. All tested MRSA isolates belonged to ST398. Eight spa types (t011, t034, t567, t571, t1451, t2974, t3423 and t5943) were detected, among which t011 predominated. SCCmec cassettes type IVa and V were present in 20% and 72% of the isolates, respectively. When combining the results of the two remaining typing methods, PFGE and MLVA, eighteen genotypes were obtained of which one genotype predominated (56% of the positive farms). All MRSA isolates were resistant to tetracycline. Resistance to trimethoprim, aminoglycosides, macrolides, lincosamides, fluoroquinolones and chloramphenicol was also observed. In conclusion, there was no effect of the farm type on the MRSA status of the pigs. A statistically significant difference was observed when comparing the pig/poultry or the pig/cattle MRSA status on the multispecies farms. Additionally, a wide variety of MRSA ST398 strains was found within certain farms when combining different typing methods

Prevalence and genetic diversity of livestock-associated methicillin-resistant Staphylococcus aureus on Belgian pork

Since the first description of livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA), a high prevalence was observed in pigs. At present, questions remain about the transmission of LA-MRSA to the general human population through pork. The objectives of the present study were to determine the prevalence of LA-MRSA in Belgian pork and to determine the role of the pork production chain and butcheries in transmission of LA-MRSA to the human population. Pig meat samples (chops, bacon, minced pork, ribs, forelimbs, and ears; n=137) originating from four butcheries (A through D) were spread plated on ChromID MRSA plates both before and after overnight enrichment culture. Suspect colonies were confirmed using a MRSA-specific triplex PCR assay and a CC398-specific PCR assay. The isolates (n = 147) were further characterized by SCCmec typing, multiple-locus variable number tandem repeat analysis, and antimicrobial susceptibility testing, a selection of isolates were subjected to pulsed-field gel electrophoresis and spa typing. Direct plating revealed a MRSA prevalence of 8%. After enrichment, MRSA was isolated from 98 (72%) of 137 samples of which the majority were from rib, ear, and forelimb. The majority (97%) of obtained isolates belonged to CC398, the main LA-MRSA type. A high level of genetic diversity was noted among the isolates from one butchery. Thirty antimicrobial susceptibility profiles were found; 13 and 9% of the isolates had Cip-Tet-Tri and Gen-Kan-Tet-Tob-Tri profiles, respectively. These results indicate the importance of enrichment for MRSA detection of pork. The observed genetic diversity of the isolates indicated that the pork production chain can be considered a source of multiple MRSA types that could be transmitted to the human population through cross-contaminated meat

Cohort study for the presence of livestock-associated MRSA in piglets: effect of sow status at farrowing and determination of the piglet colonization age

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