The objective of this pilot study was to evaluate the influence of sampling technique and exposure to different bedding types on the milk microbiome of healthy primiparous cows. Primiparous Holstein cows (n = 20) with no history of clinical mastitis or monthly somatic cell counts >150,000 cells/mL were selected for this study. From each enrolled cow, a composite milk sample was aseptically collected from all 4 mammary quarters (individual quarter somatic cell counts <100,000 cells/mL), 1 individual quarter milk sample was collected using conventional aseptic technique, and 2 individual quarter milk samples were collected directly from the gland cistern using a needle and vacuum tube. All milk samples were cultured using standard milk microbiological techniques and DNA was extracted. Extracted DNA was subjected to PCR and next-generation sequencing for microbiota determination. All samples yielded relatively little total DNA. Amplification of PCR was successful in 45, 40, and 83% of composite, conventional, and cisternal samples, respectively. Bacteria were successfully cultured from 35% of composite milk samples but from none of the quarter milk samples collected using conventional or cisternal sampling techniques. Bacterial DNA sequences were assigned to operational taxonomic units (OTU) based on 97% sequence similarity, and bacterial richness and diversity were determined. Most samples were dominated by low-prevalence OTU and of the 4,051 identified OTU, only 14 were prevalent at more than 1% each. These included bacteria typically recovered from environmental sources. Chao richness was greatest in composite samples and was 636, 347, and 356 for composite, conventional quarter, and cisternal milk samples, respectively. Shannon diversity was similar among sample types and ranged from 3.88 (quarter) to 4.17 (composite). Richness and diversity did not differ by bedding type among cisternal samples, but the power of this pilot study was limited due to small sample size. Despite the small sample size, for milk samples collected from the gland cistern, overall bacterial community composition differed among bedding types. These results demonstrate that sampling technique and bedding type may be associated with the microbiota detected in bovine milk, and we suggest that these variables should be considered in designing and reporting studies about the milk microbiota.
Steroid-induced glaucoma is an iatrogenic condition resulting from the use of glucocorticoids. Glucocorticoids such as dexamethasone (DEX) 1 raise intraocular pressure (IOP) in ϳ40% of patients in the general population, and ϳ6% of these patients will go on to develop glaucoma (1, 2). This condition is similar to primary open angle glaucoma (1-3), and is caused by a restriction in fluid outflow through the trabecular meshwork (TM), resulting in an imbalance between the amount of aqueous humor produced and the amount drained. This imbalance results in a higher IOP.It is thought that an alteration in the cytoskeletal structure or contractile properties of TM cells may result in the disruption of normal fluid flow. In support of this idea, cross-linked actin networks, referred to as CLANs, have been observed with increased frequency in the TM of glaucomatous patients and in glucocorticoid treated anterior segments as well as in TM cells in culture. CLANs are thought to alter the contractility of the TM by holding the cells in a rigid conformation, making the cells unresponsive to the change in pressure and blocking the aqueous humor outflow pathway (1,4,5). Thus, agents such as H7 and the latrunculins A and B, which disrupt the organization of the cytoskeleton, decrease IOP in porcine and monkey cultured anterior segments (6 -9).Control of the actin cytoskeleton is mediated by the Rho family of small GTPases. The Rho effector ROCK has been shown to play a part in TM contractility and modulation of IOP. Inhibition of ROCK using a dominant negative mutant or the inhibitor Y-27632 causes TM cells to "relax" by decreasing actin stress fiber formation and phosphorylation of myosin light chain (MLC) (10, 11). ROCK inhibition also decreases IOP in cultured human and porcine anterior segments (10, 11). In contrast, constitutively active RhoA (RhoA V14) increases From the ‡Departments
The objective of this longitudinal cohort study was to describe the milk microbiota of dairy cow mammary glands based on inflammation status before and after the dry period. Individual mammary quarters were assigned to cohorts based on culture results and somatic cell count (SCC) at dryoff and twice in the first 2 weeks post-calving. Mammary glands that were microbiologically negative and had low SCC (< 100,000 cells/mL) at all 3 sampling periods were classified as Healthy (n = 80). Microbiologically negative mammary glands that had SCC ≥150,000 cells/mL at dryoff and the first post-calving sample were classified as Chronic Culture-Negative Inflammation (CHRON; n = 17). Quarters that did not have both culture-negative milk and SCC ≥ 150,000 cells/mL at dryoff but were culture-negative with SCC ≥ 150,000 at both post-calving sampling periods were classified as Culture-Negative New Inflammation (NEWINF; n = 6). Mammary glands with bacterial growth and SCC ≥ 150,000 cells/mL at all 3 periods were classified as Positive (POS; n = 3). Milk samples were collected from all enrolled quarters until 150 days in milk and subjected to microbiota analysis. Milk samples underwent total DNA extraction, a 40-cycle PCR to amplify the V4 region of the bacterial 16S rRNA gene, and next-generation sequencing. Healthy quarters had the lowest rate of PCR and sequencing success (53, 67, 83, and 67% for Healthy, CHRON, NEWINF, and POS, respectively). Chao richness was greatest in milk collected from Healthy quarters and Shannon diversity was greater in milk from Healthy and CHRON quarters than in milk collected from glands in the NEWINF or POS cohorts. Regardless of cohort, season was associated with both richness and diversity, but stage of lactation was not. The most prevalent OTUs included typical gut- and skin-associated bacteria such as those in the phylum Bacteroidetes and the genera Enhydrobacter and Corynebacterium. The increased sequencing success in quarters with worse health outcomes, combined with the lack of bacterial growth in most samples and the high PCR cycle number required for amplification of bacterial DNA, suggests that the milk microbiota of culture-negative, healthy mammary glands is less abundant than that of culture-negative glands with a history of inflammation.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.