Hyeun Bum Kim

Longitudinal investigation of the age-related bacterial diversity in the feces of commercial pigs.

The importance of bacteria in the gastrointestinal tracts of animals is widely acknowledged as important. However, very little is known about composition and distribution of the microbial population in lower intestinal tracts of animals. Because most bacterial species in pig intestines have not been cultured, it has been difficult to analyze bacterial diversity by conventional culture methods. Even with the development of culture independent 16S rRNA gene sequencing, the previous methods were slow and labor intensive. Therefore, high throughput pyrosequencing of 16S rDNA libraries was used in this study in order to explore the bacterial diversity of the pig feces. In our two trials, fecal samples from individual pigs were collected five times at 3-week intervals, and the 16S rRNA genes in the community DNAs from fecal samples were sequenced and analyzed. This longitudinal study design identified that microbial populations in the feces of the each pig continued to change as pigs aged. The variations of bacterial diversity of the animals were affected by less abundant bacterial components of the feces. These results help us to understand the age-related bacterial diversity in the commercial pig feces.

Modulations of the Chicken Cecal Microbiome and Metagenome in Response to Anticoccidial and Growth Promoter Treatment

With increasing pressures to reduce or eliminate the use of antimicrobials for growth promotion purposes in production animals, there is a growing need to better understand the effects elicited by these agents in order to identify alternative approaches that might be used to maintain animal health. Antibiotic usage at subtherapeutic levels is postulated to confer a number of modulations in the microbes within the gut that ultimately result in growth promotion and reduced occurrence of disease. This study examined the effects of the coccidiostat monensin and the growth promoters virginiamycin and tylosin on the broiler chicken cecal microbiome and metagenome. Using a longitudinal design, cecal contents of commercial chickens were extracted and examined using 16S rRNA and total DNA shotgun metagenomic pyrosequencing. A number of genus-level enrichments and depletions were observed in response to monensin alone, or monensin in combination with virginiamycin or tylosin. Of note, monensin effects included depletions of Roseburia, Lactobacillus and Enterococcus, and enrichments in Coprococcus and Anaerofilum. The most notable effect observed in the monensin/virginiamycin and monensin/tylosin treatments, but not in the monensin-alone treatments, was enrichments in Escherichia coli. Analysis of the metagenomic dataset identified enrichments in transport system genes, type I fimbrial genes, and type IV conjugative secretion system genes. No significant differences were observed with regard to antimicrobial resistance gene counts. Overall, this study provides a more comprehensive glimpse of the chicken cecum microbial community, the modulations of this community in response to growth promoters, and targets for future efforts to mimic these effects using alternative approaches.

Microbial shifts in the swine distal gut in response to the treatment with antimicrobial growth promoter, tylosin

Antimicrobials have been used extensively as growth promoters (AGPs) in agricultural animal production. However, the specific mechanism of action for AGPs has not yet been determined. The work presented here was to determine and characterize the microbiome of pigs receiving one AGP, tylosin, compared with untreated pigs. We hypothesized that AGPs exerted their growth promoting effect by altering gut microbial population composition. We determined the fecal microbiome of pigs receiving tylosin compared with untreated pigs using pyrosequencing of 16S rRNA gene libraries. The data showed microbial population shifts representing both microbial succession and changes in response to the use of tylosin. Quantitative and qualitative analyses of sequences showed that tylosin caused microbial population shifts in both abundant and less abundant species. Our results established a baseline upon which mechanisms of AGPs in regulation of health and growth of animals can be investigated. Furthermore, the data will aid in the identification of alternative strategies to improve animal health and consequently production.

The intestinal microbiome of the pig

Abstract The intestinal microbiome has been the subject of study for many decades because of its importance in the health and well being of animals. The bacterial components of the intestinal microbiome have closely evolved as animals have and in so doing contribute to the overall development and metabolic needs of the animal. The microbiome of the pig has been the subject of many investigations using culture-dependent methods and more recently using culture-independent techniques. A review of the literature is consistent with many of the ecologic principles put forth by Rene Dubos. Animals develop an intestinal microbiome over time and space. During the growth and development of the pig, the microbiome changes in composition in a process known as the microbial succession. There are clear and distinct differences in the composition of the pig intestinal microbiome moving from the proximal end of the intestinal tract to the distal end. The majority (>90%) of the bacteria in the pig intestinal microbiome are from two Phyla: Firmicutes and Bacteroidetes. However, the ileum has a high percentage of bacteria in the phylum Proteobacterium (up to 40%). Perturbations to the microbiome occur in response to many factors including stresses, treatment with antibiotics, and diet.

The pig gut microbial diversity: Understanding the pig gut microbial ecology through the next generation high throughput sequencing

The importance of the gut microbiota of animals is widely acknowledged because of its pivotal roles in the health and well being of animals. The genetic diversity of the gut microbiota contributes to the overall development and metabolic needs of the animal, and provides the host with many beneficial functions including production of volatile fatty acids, re-cycling of bile salts, production of vitamin K, cellulose digestion, and development of immune system. Thus the intestinal microbiota of animals has been the subject of study for many decades. Although most of the older studies have used culture dependent methods, the recent advent of high throughput sequencing of 16S rRNA genes has facilitated in depth studies exploring microbial populations and their dynamics in the animal gut. These culture independent DNA based studies generate large amounts of data and as a result contribute to a more detailed understanding of the microbiota dynamics in the gut and the ecology of the microbial populations. Of equal importance, is being able to identify and quantify microbes that are difficult to grow or that have not been grown in the laboratory. Interpreting the data obtained from this type of study requires using basic principles of microbial diversity to understand importance of the composition of microbial populations. In this review, we summarize the literature on culture independent studies of the pig gut microbiota with an emphasis on its succession and alterations caused by diverse factors.

Longitudinal Analysis of the Lung Microbiome in Lung Transplantation

Lung transplant recipients experience poor long-term survival, largely due to chronic rejection. The pathogenesis of chronic rejection is incompletely understood, but bacterial colonization of the lung is associated with chronic rejection, while antibiotic use slows its progression. The lung harbors a bacterial community, termed the microbiome, which is present both in health and disease. We hypothesize that the lung microbiome will change following transplantation, and these changes may correspond to the development of rejection. Twelve bronchoalveolar lavage fluid (BALF) samples were obtained from four patients at three time points after transplantation, and two BALF samples were obtained from healthy, nontransplant controls. The microbiome of each sample was determined by pyrosequencing the 16S rRNA gene hypervariable 3 region. The data were analyzed using mothur, Ribosomal Database Project Classifier, Fast UniFrac, and Metastats. Transplanted lungs contained more bacterial sequences and demonstrated more microbial diversity than did control lungs. Bacteria in the phyla Proteobacteria (class Betaproteobacteria) predominated in the transplant samples. In contrast, the microbiome of the healthy lung consisted of the phyla Proteobacteria (class Gammaproteobacteria) and Firmicutes. The microbiome of the transplanted lung is vastly different from that of healthy lungs, mainly due to the presence of the family Burkholderiaceae in transplant samples.

Pyrosequencing-based analysis of fecal microbial communities in three purebred pig lines

This study examined the fecal bacterial diversity of 15-weekold pigs from three purebred lines: Duroc, Landrace, and Yorkshire. Taxon-dependent and -independent analyses were performed to evaluate differences in the fecal bacterial communities and to identify bacterial genera that can be used to discriminate breeds, following high-throughput pyrosequencing of 16S rRNA genes. Among the breeds evaluated, Landrace had the most diverse bacterial community composition. Prevotella, Blautia, Oscillibacter, and Clostridium were detected in all samples regardless of breed. On the other hand, Catenibacterium, Blautia, Dialister, and Sphaerochaeta were differentially detected among breeds, as demonstrated by the canonical loading plot. The discriminant analysis of principal components plot also showed clear separation of the three purebred pig lines, with a certain degree of similarity between Landrace and Yorkshire pigs and a distinct separation between Duroc pigs and the other two breeds. Other factors not related to breed, such as season or time of sampling and pen effects, may contribute to shaping the gut microbiota of pigs.

Comparative efficacy of three commercial PCV2 vaccines in conventionally reared pigs.

The aim of this study was to compare the efficacy of three commercial porcine circovirus type 2 (PCV2) vaccines. Twenty 3-week old pigs were injected with one of three different vaccines, A, B, C, or were unvaccinated controls. Pigs in group A were given a second dose at 6 weeks of age according to the manufacturers protocol. Antibody titers in all pigs declined gradually, but increased sharply by 22 weeks of age. Viremia in the three vaccinated groups at 22 weeks of age was significantly less than in the control group. The average daily gains of pigs in groups A-C, and in controls were 0.81 ± 0.03, 0.80 ± 0.05, 0.78 ± 0.04, and 0.74 ± 0.05 kg, respectively. It was concluded that PCV2 vaccine reduced viremia and improved weight gain, but differences in weight gain among vaccinated groups were not statistically significant.

Changes in the Porcine Intestinal Microbiome in Response to Infection with Salmonella Enterica and Lawsonia Intracellularis

Salmonella enterica is a leading cause of food borne illness. Recent studies have shown that S. enterica is a pathogen capable of causing alterations to the composition of the intestinal microbiome. A recent prospective study of French pork production farms found a statistically significant association between Lawsonia intracellularis and carriage of S. enterica. In the current study the composition of the gut microbiome was determined in pigs challenged with S. enterica serovar Typhimurium and or L. intracellularis and compared to non-challenged control pigs. Principal coordinate analysis demonstrated that there was a disruption in the composition of the gut microbiome in the colon and cecum of pigs challenged with either pathogen. The compositions of the microbiomes of challenged pigs were similar to each other but differed from the non-challenged controls. There also were statistically significant increases in Anaerobacter, Barnesiella, Pediococcus, Sporacetigenium, Turicibacter, Catenibacterium, Prevotella, Pseudobutyrivibrio, and Xylanibacter in the challenged pigs. To determine if these changes were specific to experimentally challenged pigs, we determined the compositions of the fecal microbiomes of naturally infected pigs that were carriers of S. enterica. Pigs that were frequent shedders of S. enterica were shown to have similar fecal microbiomes compared to non-shedders or pigs that shed S. enterica infrequently. In a comparison of the differentially abundant bacteria in the naturally infected pigs compared to experimentally challenged pigs, 9 genera were differentially abundant and each exhibited the same increase or decrease in abundance between the two groups. Thus, there were similar changes in the GI microbiome associated with carriage of S. enterica regardless of whether the pigs were experimentally challenged with S. enterica or acquired it naturally.

Update on Antimicrobial Resistance in Clostridium difficile: Resistance Mechanisms and Antimicrobial Susceptibility Testing

ABSTRACT Oral antibiotics such as metronidazole, vancomycin and fidaxomicin are therapies of choice for Clostridium difficile infection. Several important mechanisms for C. difficile antibiotic resistance have been described, including the acquisition of antibiotic resistance genes via the transfer of mobile genetic elements, selective pressure in vivo resulting in gene mutations, altered expression of redox-active proteins, iron metabolism, and DNA repair, as well as via biofilm formation. This update summarizes new information published since 2010 on phenotypic and genotypic resistance mechanisms in C. difficile and addresses susceptibility test methods and other strategies to counter antibiotic resistance of C. difficile.