Local host response following an intramammary challenge with Staphylococcus fleurettii and different strains of Staphylococcus chromogenes in dairy heifers

Coagulase-negative staphylococci (CNS) are a common cause of subclinical mastitis in dairy cattle. The CNS inhabit various ecological habitats, ranging between the environment and the host. In order to obtain a better insight into the host response, an experimental infection was carried out in eight healthy heifers in mid-lactation with three different CNS strains: a Staphylococcus fleurettii strain originating from sawdust bedding, an intramammary Staphylococcus chromogenes strain originating from a persistent intramammary infection (S. chromogenes IM) and a S. chromogenes strain isolated from a heifer's teat apex (S. chromogenes TA). Each heifer was inoculated in the mammary gland with 1.0 x 10(6) colony forming units of each bacterial strain (one strain per udder quarter), whereas the remaining quarter was infused with phosphate-buffered saline. Overall, the CNS evoked a mild local host response. The somatic cell count increased in all S. fleurettii-inoculated quarters, although the strain was eliminated within 12 h. The two S. chromogenes strains were shed in larger numbers for a longer period. Bacterial and somatic cell counts, as well as neutrophil responses, were higher after inoculation with S. chromogenes IM than with S. chromogenes TA. In conclusion, these results suggest that S. chromogenes might be better adapted to the mammary gland than S. fleurettii. Furthermore, not all S. chromogenes strains induce the same local host response

Mycolicibacterium smegmatis, basonym Mycobacterium smegmatis, causing pyogranulomatous mastitis and its cross-reactivity in bovine (para)tuberculosis testing

Different mycobacterial species are encountered in bovine medicine. The fastidiously growing mycobacteria (Mycobacterium bovis as the cause of bovine tuberculosis, and Mycobacterium avium ssp. paratuberculosis, MAP, as the cause of paratuberculosis) are well known and targeted in eradication/control or monitoring programs in different countries, whereas the rapidly growing species is only rarely identified from bovine disease. The latter have occasionally been reported as the cause of bovine clinical mastitis, but recent reports are scarce. In this study, Mycolicibacterium smegmatis (basonym Mycobacterium smegmatis) was identified as cause of granulomatous, relapsing clinical mastitis in 2 cows from one Belgian dairy herd. Milk, blood, and fecal samples were collected, as well as tissue samples after the cows were culled. Serological analysis conducted on milk and serum samples resulted in positive reactions for MAP, but negative for Mycobacterium bovis. Production of IFN-gamma showed sensitization with mycobacteria or similar organisms, other than M. bovis, in one cow. Detection of MAP by bacteriological culture and IS900-based quantitative PCR on milk and feces remained negative. In conclusion, this paper describes M. smegmatis as a cause of bovine clinical mastitis in Belgium and suggests cross-reactivity of the intramammary M. smegmatis infection with routinely used serological tests for MAP

Coagulase-negative Staphylococcus species in bulk milk : prevalence, distribution, and associated subgroup- and species-specific risk factors

Coagulase-negative staphylococci (CNS) have become the main pathogens causing bovine mastitis in recent years. A huge variation in species distribution among herds has been observed in several studies, emphasizing the need to identify subgroup- and species specific herd-level factors to improve our understanding of the differences in ecological and epidemiological nature between species. The use of bulk milk samples enables the inclusion of a large(r) number of herds needed to identify herd-level risk factors and increases the likelihood of recovering enough isolates per species needed for conducting subgroup- and, eventually, species-specific analyses at the same time. This study aimed to describe the prevalence and distribution of CNS species in bulk milk samples and to identify associated subgroup- and species-specific herd-level factors. Ninety percent of all bulk milk samples yielded CNS. Staphylococcus equorum was the predominant species, followed by Staphylococcus haemolyticus and Staphylococcus epidermidis. A seasonal effect was observed for several CNS species. Bulk milk samples from herds with a loose-pack or a tiestall housing system were more likely to yield CNS species compared with herds with a freestall barn, except for S. epidermidis, Staphylococcus simulans, and Staphylococcus cohnii. In September, herds in which udders were clipped had lower odds of yielding Staphylococcus chromogenes, S. simulans, and Staphylococcus xylosus, the CNS species assumed to be most relevant for udder health, in their bulk milk than herds in which udder clipping was not practiced. Bulk milk of herds participating in a monthly veterinary udder health-monitoring program was more likely to yield these 3 CNS species. Herds always receiving their milk quality premium or predisinfecting teats before attachment of the milking cluster had lower odds of having S. equorum in their bulk milk. Herds not using a single dry cotton or paper towel for each cow during premilking udder preparation were more likely to have S. cohnii-positive bulk milk. Herds in which flushing with hot water or steam of the milking cluster after having milked a cow with a (sub)clinical mastitis was applied, were less likely to yield S. simulans, S. haemolyticus, and S. cohnii in their bulk milk. Always wearing gloves during milking decreased the odds of having Staphylococcus devriesei-positive bulk milk. Tap water from the public drinking system used as drinking water increased the odds of yielding S. simulans in the bulk milk. In conclusion, CNS are highly prevalent in bulk milk and might originate from the environment for some species (we hypothesize this is true for S. equorum or S. cohnii), or from within the udder (e.g., for S. simulans). Studies collecting bulk milk and quarter milk samples at the same time along with environmental samples are needed to determine the exact origin of the different (subgroups of) CNS species present in bulk milk using strain-typing techniques

Some coagulase-negative Staphylococcus species affect udder health more than others

A longitudinal study in 3 dairy herds was conducted to profile the distribution of coagulase-negative Staphylococcus (CNS) species causing bovine intramammary infection (IMI) using molecular identification and to gain more insight in the pathogenic potential of CNS as a group and of the most prevalent species causing IMI. Monthly milk samples from 25 cows in each herd as well as samples from clinical mastitis were collected over a 13-mo period. Coagulase-negative staphylococci were identified to the species level using transfer-RNA intergenic spacer PCR. The distribution of CNS causing IMI was highly herd-dependent, but overall, Staphylococcus chromogenes, Staphylococcus xylosus, Staphylococcus cohnii, and Staphylococcus simulans were the most prevalent. No CNS species were found to cause clinical mastitis. The effect of the most prevalent species on the quarter milk somatic cell count (SCC) was analyzed using a linear mixed model, showing that Staph. chromogenes, Staph. simulans, and Staph. xylosus induced an increase in the SCC that is comparable with that of Staphylococcus aureus. Almost all CNS species were able to cause persistent IMI, with Staph. chromogenes causing the most persistent infections. In conclusion, accurate species identification cannot be ignored when studying the effect of CNS on udder health, as the effect on SCC differs between species and species distribution is herd-specific. Staphylococcus chromogenes, Staph. simulans, and Staph. xylosus seem to be the more important species and deserve special attention in further studies. Reasons for herd dependency and possible cow- and quarter-level risk factors should be examined in detail for the different species, eventually leading to cost-benefit analyses for management changes and, if needed, treatment recommendations

Short communication: Species group-specific predictors at the cow and quarter level for intramammary infection with coagulase-negative staphylococci in dairy cattle throughout lactation

Coagulase-negative staphylococci (CNS) are frequently isolated from quarters with subclinical mastitis, teat apices, and the cows' environment. Virulence, ecology, epidemiological behavior, and effect on udder health vary between different CNS species. Staphylococcus chromogenes, Staph. simulans, and Staph. xylosus are frequently present in milk and have a more substantial effect on quarter milk somatic cell count than other species. Therefore, these species are considered the "more relevant" CNS. As species-specific factors associated with CNS intramammary infection (IMI) have not yet been identified and susceptibility for IMI differs between cows and quarters, this study aimed to identify predictors for CNS IMI at the cow and quarter level (some of them changing over time) with a specific focus on the aforementioned more relevant CNS. Precise data were available from a longitudinal study (3,052 observations from 344 quarters from 86 dairy cows belonging to 3 commercial dairy herds). All CNS were molecularly identified to the species level, and multivariable, multilevel logistic regression models taking into account the longitudinal nature of the data, were fit to study the likelihood of infection. Staphylococcus chromogenes, Staph. xylosus, and Staph. cohnii were the most frequently isolated species from CNS IMI in older cows, whereas Staph. chromogenes, Staph. xylosus, and Staph. simulans were the main species found in IMI in heifers. Quarters from heifers (as opposed to multiparous cows), from heifers and multiparous cows in third or fourth month in lactation (as opposed to early lactation, <60 d in milk), and with an increasing quarter milk SCC were more likely to be infected with the more relevant CNS species. Quarter milk SCC was identified as the sole statistically significant predictor for IMI with other CNS species, although the size of the effect was lower [odds ratio of 1.6 (1.4-1.9) vs. 2.1 (1.8-2.5)] than the effect for IMI with the more relevant CNS. As a strong herd effect was present, studying herd-level predictors is warranted

Performance of API Staph ID 32 and Staph-Zym for identification of coagulase-negative staphylococci isolated from bovine milk samples

In this study, the accuracy of two phenotypic tests, API Staph ID 32 and Staph-Zym, was determined for identification of coagulase-negative staphylococci (CNS) from bovine milk samples in comparison with identification based on DNA-sequencing. A total of 172 CNS isolated from bovine milk were classified into 17 species. The most frequently isolated species based on rpoB sequencing were Staphylococcus chromogenes and Staphylococcus epidermidis, followed by Staphylococcus xylosus, Staphylococcus warneri and Staphylococcus equorum (37, 13, 9, 8 and 6% of isolates, respectively). The API Staph ID 32 correctly identified 41% of the CNS isolates. Best agreement with rpoB sequence based species identification was found for S. epidermidis, Staphylococcus hyicus and S. xylosus (100, 89 and 87%, respectively). The positive predictive value was 89, 100 and 52%, respectively. Poor sensitivity was observed for 3 of the 5 most frequently found species, S. chromogenes (37%), Staphylococcus warneri (15%) and S. equorum (0%) albeit with specificity of 100%. The Staph-Zym needed additional tests for 66% of the isolates and identified 31% of the CNS isolates correctly. Good sensitivity was found for S. epidermidis, S. simulans and S. xyloxus (100, 78 and 73%, respectively). The positive predictive value was 89, 78 and 98%, respectively. Poor sensitivity was observed for S. chromogenes, S. warneri and S. equorum (0, 54 and 0%, respectively) but with a specificity of 100, 99 and 100%, respectively. Both phenotypic tests misidentified a large proportion of CNS isolates and were thus unsuitable for identification of CNS species from bovine milk samples