Microbiota analysis of environmental slurry and its potential role as a reservoir of bovine digital dermatitis pathogens

ABSTRACT At present, very little information exists regarding what role the environmental slurry may play as an infection reservoir and/or route of transmission for bovine digital dermatitis (DD), a disease which is a global problem in dairy herds. To investigate whether DD-related bacteria belong to the indigenous microbiota of the dairy herd environment, we used deep amplicon sequencing of the 16S rRNA gene in 135 slurry samples collected from different sites in 22 dairy farms, with and without DD-infected cows. Both the general bacterial populations and digital dermatitis-associated Treponema were targeted in this study. The results revealed significant differences in the bacterial communities between the herds, with only 12 bacterial taxa shared across at least 80% of all the individual samples. These differences in the herd microbiota appeared to reflect mainly between-herd variation. Not surprisingly, the slurry was dominated by ubiquitous gastrointestinal bacteria, such as Ruminococcaceae and Lachnospiraceae . Despite the low relative abundance of spirochetes, which ranged from 0 to 0.6%, we were able to detect small amounts of bacterial DNA from DD-associated treponemes in the slurry. However, the DD-associated Treponema spp. were detected only in samples from herds with reported DD problems. These data indicate that treponemes involved in the pathogenesis of DD are not part of the normal environmental microflora in dairy herds without clinical DD and, consequently, that slurry is not a primary reservoir of infection. IMPORTANCE Bovine digital dermatitis (DD), a dermal disease which causes lameness in dairy cattle, is a serious problem worldwide. To control this disease, the infection reservoirs and transmission routes of DD pathogens need to be clarified. The dairy herd slurry may be a pathogen reservoir of DD-associated bacteria. The rationale for the present study was, therefore, to examine whether DD-associated bacteria are always present in slurry or if they are found only in DD-afflicted herds. The results strongly indicated that DD Treponema spp. are not part of the indigenous slurry and, therefore, do not comprise an infection reservoir in healthy herds. This study applied next-generation sequencing technology to decipher the microbial compositions of environmental slurry of dairy herds with and without digital dermatitis. </jats:p

In situ prebiotics for weaning piglets: In vitro production and fermentation of potato galactorhamnogalacturonan

Postweaning diarrhea (PWD) in pigs is a leading cause of economic loss in pork production worldwide. The current practice of using antibiotics and zinc to treat PWD is unsustainable due to the potential of antibiotic resistance and ecological disturbance, and novel methods are required. In this study, an in vitro model was used to test the possibility of producing prebiotic fiber in situ in the gastrointestinal (GI) tract of the piglet and the prebiotic activity of the resulting fiber in the terminal ileum. Soluble fiber was successfully produced from potato pulp, an industrial waste product, with the minimal enzyme dose in a simulated upper GI tract model extracting 26.9% of the initial dry matter. The fiber was rich in galactose and galacturonic acid and was fermented at 2.5, 5, or 10 g/liter in a glucose-free medium inoculated with the gut contents of piglet terminal ileum. Fermentations of 5 g/liter inulin or 5 g/liter of a purified potato fiber were used as controls. The fibers showed high fermentability, evident by a dose-dependent drop in pH and an increase in the organic acid content, with lactate in particular being increased. Deep sequencing showed a significant increase in the numbers of Lactobacillus and Veillonella organisms and an insignificant increase in the numbers of Clostridium organisms as well as a decrease in the numbers of Streptococcus organisms. Multivariate analysis showed clustering of the treatment groups, with the group treated with purified potato fiber being clearly separated from the other groups, as the microbiota composition was 60% Lactobacillus and almost free of Clostridium. For animal studies, a dosage corresponding to the 5-g/liter treatment is suggested

A detailed investigation of the porcine skin and nose microbiome using universal and <i>Staphylococcus</i> specific primers

Abstract MRSA is an increasing problem in humans as well as livestock. The bacterial co-colonization of the skin in MRSA carriers has been poorly investigated and moreover, there have been no methods for high resolution investigations of the Staphylococcus genus apart from tediously culturing or doing multiple PCRs. On 120 samples from pig ear, skin and nose, we generated amplicons from the V1-V2 region of the 16S rRNA gene to gather an overview of the genus-level microbiome, along with using MRSA specific plates to count MRSA. In parallel with this, amplicons of the tuf gene were generated, targeting only a region of the tuf gene found only in the Staphylococcus genus. Using these methods, we determined a core microbiota across the healthy pig and determined the Staphylococcus genus to be dominated by S. equorum. Moreover, we found Streptococcus to be inversely associated with Staphylococcus and MRSA, suggesting a role for this genus in combating MRSA. In this work, we have thoroughly investigated the skin and nose microbiome of the pig and developed a high throughput method for profiling the Staphylococcus genus which we believe will be useful for further investigations

A detailed investigation of the porcine skin and nose microbiome using universal and <i>Staphylococcus</i> specific primers

Abstract MRSA is an increasing problem in humans as well as livestock. The bacterial co-colonization of the skin in MRSA carriers has been poorly investigated and moreover, there have been no methods for high resolution investigations of the Staphylococcus genus apart from tediously culturing or doing multiple PCRs. On 120 samples from pig ear, skin and nose, we generated amplicons from the V1-V2 region of the 16S rRNA gene to gather an overview of the genus-level microbiome, along with using MRSA specific plates to count MRSA. In parallel with this, amplicons of the tuf gene were generated, targeting only a region of the tuf gene found only in the Staphylococcus genus. Using these methods, we determined a core microbiota across the healthy pig and determined the Staphylococcus genus to be dominated by S. equorum. Moreover, we found Streptococcus to be inversely associated with Staphylococcus and MRSA, suggesting a role for this genus in combating MRSA. In this work, we have thoroughly investigated the skin and nose microbiome of the pig and developed a high throughput method for profiling the Staphylococcus genus which we believe will be useful for further investigations

In situ prebiotics: enzymatic release of galacto-rhamnogalacturonan from potato pulp in vivo in the gastrointestinal tract of the weaning piglet

Prebiotics may be efficient for prevention of intestinal infections in humans and animals by increasing the levels of beneficial bacteria and thereby improving gut health. Using purified prebiotics may however not be cost-effective in the livestock production industry. Instead, prebiotic fibres may be released directly in the gastro-intestinal tract by feeding enzymes with a suitable substrate and allowing the prebiotics to be produced in situ. Using low doses, 0.03 % enzyme-to-substrate ratio, of the enzymes pectin lyase and polygalacturonase in combination with potato pulp, a low-value industrial by-product, we show that high molecular weight galacto-rhamnogalacturonan can be solubilized in the stomach of weaning piglets. The release of this fiber is in the order of 22–38 % of the theoretical amount, achieved within 20 min. The catalysis takes place mainly in the stomach of the animal and is then followed by distribution through the small intestines. To our knowledge, this is the first paper describing targeted production of prebiotics in an animal model