Search

Gnotobiotics or gnotobiology is a research field exploring organisms with a known microbiological state. In animal research, the development of gnotobiotics started in the late 19th century with the rederivation of germ-free guinea pigs. Cutting-edge achievements were accomplished by scientists in the Laboratories of Bacteriology at the University of Notre Dame (LOBUND). The primary goals of gnotobiotics were not only the development of the equipment required for long-term husbandry but also phenotypic characterization of germ-free animals. The first isolators were designed by Reynolds and Gustafsson as rigid-wall stainless steel autoclave-like chambers, which were subsequently replaced by Trexler's flexible-film polyvinyl plastic isolators. Flexible-film or semi-rigid isolators are commonly used today. The long-term maintenance of gnotobiotic rodents is performed in positive-pressure isolators. However, to facilitate gnotobiotic experimental procedures, short-term husbandry systems have been developed. Gnotobiotic animal husbandry is laborious and requires experienced staff. Germ-free animals can be rederived from existing rodent colonies by hysterectomy or embryo transfer. The physiology and anatomy of germ-free rodents are different from those of specified pathogen-free (SPF) rodents. Furthermore, to guarantee gnotobiotic status, the colonies need to be regularly microbiologically monitored. Today, gnotobiotics provides a powerful tool to analyse functional effects of host-microbe interactions, especially in complex disease models. Gnotobiotic models combined with 'omics' approaches will be indispensable for future advances in microbiome research. Furthermore, these approaches will contribute to the development of novel therapeutic targets. In addition, regional or national gnotobiotic core facilities should be established in the future to support further applications of gnotobiotic models. [ABSTRACT FROM AUTHOR]

The assembly of horizontally transmitted endophytic fungi within plant tissues may be affected by "biotic filtering". In other words, only particular endophytic fungal taxa from the available inoculum pool may be able to colonize a given plant species. We tested that hypothesis in Bromus tectorum, an important invasive species in the arid, western United States. We collected seed from Bromus tectorum and sources of inoculum for endophytic fungi including soil and various kinds of plant litter at a field site in central Utah. We characterized, using Illumina sequencing, the endophytic fungal communities in the various inoculum sources, inoculated Bromus tectorum seedlings under gnotobiotic conditions with the various sources, and then characterized the communities of endophytic fungi that assembled in their roots and leaves. Different inoculum sources containing significantly different endophytic fungal communities produced complex communities of endophytic fungi in leaves and roots of Bromus tectorum. In leaves, the communities assembling from the various inoculum sources were not significantly different from each other and, in roots, they were only slightly different from each other, mainly due to variation in a single fungal OTU, Coprinopsis brunneofibrillosa. Consequently, there was significantly more variation in the structure of the communities of endophytic fungi among the inoculum sources than in the resultant endophytic fungal communities in the leaves and roots of Bromus tectorum. These results are consistent with biotic filtering playing a significant role in endophytic fungal community assembly. [ABSTRACT FROM AUTHOR]

Gnotobiotics is the scientific study of animals or other organisms that are raised in germ free environments or ones that contain only specifically known germs. The gnotobiotic laboratory animal is potentially a very valuable tool for investigating any suspected interaction between the host and its associated microflora or between different components of that flora. However, like many other good ideas, the production of gnotobiotes is simple in concept but complicated in execution. In the early stages the greatest obstacles to the general use of germ free animals were the expense and the restricted amount of space that could be maintained free from contaminants. Nowadays, with modern isolators and facilities it is easier to produce gnotobiotic animals at relatively modest price. [ABSTRACT FROM AUTHOR]

The intestinal microbiota and its functions are intricately interwoven with host physiology. Colonizing rodents with donor microbiota provides insights into host-microbiota interactions characterization and the understanding of disease physiopathology. However, a better assessment of inoculation methods and recipient mouse models is needed. Here, we compare the engraftment at short and long term of genetically obese mice microbiota in germ-free (GF) mice and juvenile and adult specific pathogen free (SPF) mice. We also tested the effects of initial microbiota depletion before microbiota transfer. In the present work, donor microbiota engraftment was better in juvenile SPF mice than in adult SPF mice. In juvenile mice, initial microbiota depletion using laxatives or antibiotics improved donor microbiota engraftment 9 weeks but not 3 weeks after microbiota transfer. Microbiota-depleted juvenile mice performed better than GF mice 3 weeks after the microbiota transfer. However, 9 weeks after transfer, colonized GF mice microbiota had the lowest Unifrac distance to the donor microbiota. Colonized GF mice were also characterized by a chronic alteration in intestinal absorptive function. With these collective results, we show that the use of juvenile mice subjected to initial microbiota depletion constitutes a valid alternative to GF mice in microbiota transfer studies. [ABSTRACT FROM AUTHOR]

This brief review is dedicated to the legacy of Prof. Jaroslav Šterzl and his colleagues, who laid the foundation for gnotobiology in the former Czechoslovakia 55 years. Prof. Sterzl became one of the founders of modern Czechoslovak immunology, which was characterized by work on a wide range of problems needing to be solved. While examining the mechanisms of innate immunity, he focused his studies on the induction of antibody production by immunocompetent cells involved in adaptive immune transmission while using the model of pig fetuses and germ-free piglets and characterizing immunoglobulins in the sera of these piglets. Although not fully appreciated to this day, his experimental proof of the hypothesis focused on the common precursor of cell-forming antibodies of different isotypes was later confirmed in experiments at the gene level. Prof. Sterzl's work represented a true milestone in the development of not solely Czechoslovak but also European and global immunology. He collaborated closely with the World Health Organization for many years, serving there as leader of the Reference Laboratory for Factors of Innate Immunity. [ABSTRACT FROM AUTHOR]

Differences in the responses of conventional and germfree male Sprague-Dawley rats to acute injury induced by alpha-naphthylisothiocyanate (ANIT), a well-characterized biliary epithelial toxicant, were evaluated. Conventional and germfree rats were dosed once orally with 50 mg/kg of ANIT or corn oil alone and serially sacrificed daily for the next 3 days. Germfree rats treated with ANIT tended to have greater increases in virtually all liver and biliary-related analytes compared with conventional rats treated with ANIT; however, significant differences were found only in a few of these analytes including increased bile acids on day 3, total bilirubin on day 4, glutamate dehydrogenase (GLDH) on day 3, and reduced paraoxonase 1 (PON1) on days 2 and 3. Histologic differences between the conventional and germfree rats were modest, but most pronounced on day 2 (24-hr post dosing). Based on subjective scoring, biliary necrosis, neutrophilic cholangitis, and portal tract edema were more severe in germfree rats at 24 hr post dosing compared with conventional rats. Biliary epithelial replication did not differ between treated groups, however. Overall, germfree rats had a modestly greater level of biliary tract injury based on subjective histologic scoring and clinical chemistry measurements following an acute exposure to the well-characterized biliary toxin, ANIT; however, the difference between the ANIT-treated germfree and conventional groups was modest and most evident only within the first day following exposure. These findings suggest that the microbiome did not significantly affect ANIT-induced acute biliary tract injury in the conditions of this study. [ABSTRACT FROM AUTHOR]

Development of the infant small intestine is influenced by bacterial colonization. To promote establishment of optimal microbial communities in preterm infants, knowledge of the beneficial functions of the early gut microbiota on intestinal development is needed. The purpose of this study was to investigate the impact of early preterm infant microbiota on host gut development using a gnotobiotic mouse model. Histological assessment of intestinal development was performed. The differentiation of four epithelial cell lineages (enterocytes, goblet cells, Paneth cells, enteroendocrine cells) and tight junction (TJ) formation was examined. Using weight gain as a surrogate marker for health, we found that early microbiota from a preterm infant with normal weight gain (MPI-H) induced increased villus height and crypt depth, increased cell proliferation, increased numbers of goblet cells and Paneth cells, and enhanced TJs compared with the changes induced by early microbiota from a poor weight gain preterm infant (MPI-L). Laser capture microdissection (LCM) plus qRT-PCR further revealed, in MPI-H mice, a higher expression of stem cell marker Lgr5 and Paneth cell markers Lyz1 and Cryptdin5 in crypt populations, along with higher expression of the goblet cell and mature enterocyte marker Muc3 in villus populations. In contrast, MPI-L microbiota failed to induce the aforementioned changes and presented intestinal characteristics comparable to a germfree host. Our data demonstrate that microbial communities have differential effects on intestinal development. Future studies to identify pioneer settlers in neonatal microbial communities necessary to induce maturation may provide new insights for preterm infant microbial ecosystem therapeutics. [ABSTRACT FROM AUTHOR]

This article examines a specific technology, the germ-free "isolator," tracing its development across three sites: (1) the laboratory for the production of standard laboratory animals, (2) agriculture for the efficient production of farm animals, and (3) the hospital for the control and prevention of cross-infection and the protection of individuals from infection. Germ-free technology traveled across the laboratory sciences, clinical and veterinary medicine, and industry, yet failed to become institutionalized outside the laboratory. That germ-free technology worked was not at issue. Working, however, was not enough. Examining the history of a technology that failed to find widespread application reveals the labor involved in aligning cultural, societal, and material factors necessary for successful medical innovation. [ABSTRACT FROM AUTHOR]

The article presents a discussion of the concept of standardization through mechanization, examining its application to the biological sciences, wherein biological engineering technologies are used to develop standard forms of germ-free animals to be used in experimental science. Particular focus is given to the role played by the scientists James Reyniers and Charles Trexler in developing germ-free animals. Other topics include a description of Reyniers' germ-free steel-isolator systems for animals, his role in developing the Laboratories of Bacteriology at the University of Notre Dame (LOBUND), and innovations made by Trexler in the field of gnotobiotics.

Previous studies have suggested that intestinal microbiota modulates colonic epithelium renewal. The objective of our work was to study the effects of microbiota on colonic epithelium structure and cell cycle-related proteins by using gnotobiotic rats. Colonic crypts and amount of cell cycle-related proteins were compared between germ-free (GF), conventional (CV), and conventionalized rats by histochemistry and Western blot. Ki67 and proliferating cell nuclear antigen (PCNA) were used as surrogates for proliferative cells; p21cip1 and p27kip1 were markers of cell cycle arrest; anti- and proapoptotic proteins, Bcl2 and Bax, respectively, were also studied. We observed 40% increase of the crypt proliferative area 2 days after inoculation of GF rats with a complex microbiota. This recruitment of proliferative cells may account for the 30% increase of crypt depth observed between CV and GF rats. The hyperproliferative boost induced by microbiota was compensated by a fourfold increase of p21cip1 and p27kip1 involved in cell cycle arrest and a 30% drop of antiapoptotic Bcl2 protein while Bax was unchanged. Inductions of p21cip1, p27kip1, and PCNA protein were not paralleled by an increase of the corresponding mRNA. We also showed that p21cip1 induction by microbiota was partially restored by Bacteroides thetaiotaomicron, Ruminococcus gnavus, and Clostridium paraputrificum. Colonization of the colon by a complex microbiota increases the crypt depth of colon epithelium. This event takes place in conjunction with a multistep process: a hyperproliferative boost accompanied by compensatory events as induction of p21cip1 and p27kip1 and decrease of Bcl2. [ABSTRACT FROM AUTHOR]

Aims: This study was conducted to test the virulence of luminescent (L) and non-luminescent (NL) isogenic strains of Vibrio campbellii LMG21363, Vibrio harveyi BB120 (wild type) and quorum-sensing mutant strains derived from the wild type such as Vibrio harveyi BB152, BB170, MM30 and BB886. Methods and Results: The NL strains could be obtained by culturing rifampicin-resistant luminescent strains in the dark under static condition. The virulence of the L and NL strains was tested in gnotobiotic Artemia franciscana larvae challenged with 104 CFU ml−1 of bacteria. All luminescent isogenic tested strains showed higher virulence compared to the NL strains. The virulence of L and NL V. campbellii and V. harveyi BB120 was also tested in specific pathogen-free juvenile shrimp upon intramuscular injection with 106 CFU of bacteria. In contrast with Artemia, there was no significant difference in mortality between the groups challenged with L and NL strains ( P > 0·05). The non-luminescent strains were not able to revert back to the luminescent state and quorum sensing did not influence this phenotypic shift. Conclusions: Luminescent Vibrio strains can switch to a non-luminescent state by culturing them in static conditions. The NL strains become less virulent as verified in Artemia. Significance and Impact of the Study: The luminescent state of Vibrio cells in a culture needs to be verified in order to assure maintenance of virulence. [ABSTRACT FROM AUTHOR]

Antibiotics have traditionally been used for growth promotion in the pork industry; however, their use in animal feed has recently been limited because of human health concerns. The intestinal microbiota plays an important role in mediating many physiological functions such as digestion and animal growth. It was hypothesized that use of antibiotics as growth promotants and subsequent variations in intestinal microbiota induce significant changes in the intestinal glycoconjugate composition, which ultimately affects animal growth and disease susceptibility. The aim of this study was to characterize the lectin binding profiles of the ileum of weanling pigs in response to the absence of intestinal microbiota and to the use of the antibiotic chlortetracycline as growth promotant. Eighteen half-sib piglets obtained by cesarean section were divided into 3 treatment groups (n = 6) and maintained as control, antibiotic-fed, and gnotobiotic piglets until 5 wk of age. The glycoconjugate composition of the ileal tissues was examined by lectin histochemistry. Lycopersicon esculentum lectin, Jacalin, Pisum sativum agglutinin, Lens culinaris agglutinin (LCA), and Sambucus nigra lectin showed significant differences (P < 0.05) in binding intensities on the dome and villous epithelium between the treatment groups. Griffonia simplicifolia lectin I, Glycine maxi agglutinin, and Arachis hypogea agglutinin exhibited differences (P < 0.05) between treatment groups in lectin binding on goblet cells. Triticum vulgaris agglutinin, Pisurn sativum agglutinin, and LCA showed significant differences (P < 0.05) in binding intensities on dome, corona, and follicular regions of the ileum among treatment groups of animals. Overall, ileal tissues from gnotobiotic piglets expressed significantly weaker (P < 0.05) lectin binding for many lectins compared with control and antibiotic groups. This suggests that the intestinal microbiota plays an important role in the expression of sugar moieties in the intestine. Lectins LCA, Phaseolus vulgaris Leucoagglutinin, and Maackia amurensis lectin II showed significant differences (P < 0.05) in lectin bindings between control and antibiotic-fed piglets. This indicates that chlortetracycline as a growth promotant induces biologically relevant changes in the lectin binding profile of the ileum. These findings will help in further understanding the role of the gut microbiota and the mechanisms of action of antibiotics as growth promotants in pigs. [ABSTRACT FROM AUTHOR]

The culture of aquatic organisms is still hampered by the occurrence of unpredictable diseases in their early life stages, which are responsible for massive mortalities and considerable economic losses. A better understanding of the host–microbe interactions is certainly essential to develop effective solutions of disease control for the aquaculture industry. As demonstrated in terrestrial animals, the use of gnotobiotic systems (animals cultured in axenic conditions or with a known microflora) can be an excellent tool to extent the understanding of the mechanisms involved in host–microbe interactions and to evaluate new treatments of disease control. Several aquatic animals were cultured so far in germ-free conditions, such as fish, molluscs, crustaceans, rotifers, echinoderms, cnidarians, turbellarians, ascidians and echiurans. The aim of the present review is to recapitulate the findings obtained with gnotobiotic aquatic animals over the last decades, with special emphasis to the host–microbe interactions, as well as the perspectives for future research in this field. In addition, the procedures utilized to culture axenic aquatic animals and to verify contaminations are summarized, and the standardization of these procedures is proposed. [ABSTRACT FROM AUTHOR]

The establishment of β-glucuronidase activity was investigated in intestinal contents from gnotobiotic mice monoassociated either with Lactobacillus rhamnosus ATCC 53103 (LGG) or with Escherichia coli X7 for different periods of time. Additionally, the enzyme activity was investigated after the animals had been conventionalized. Neither the monoassociated animals nor the conventionalized animals had reached normal levels of β-glucuronidase activity after the colonization period (3 and 7 days, or 7 and 14 days, respectively). A great inter-individual variation of the enzyme activity was observed in the group that was conventionalized for longer (14 days), mainly in the caecum. These results indicate a very slow establishment of the β-glucuronidase activity in the intestinal tract of mice after exposure to either individual bacterial strains or exclusively to the environment. [ABSTRACT FROM AUTHOR]

We studied the synergistic effects of Japanese green tea extract (JGTE) and levofloxacin (LVFX) against enterohemorrhagic Escherichia coli (EHEC) infection in a gnotobiotic mouse model. Mice fed on JGTE conferred a significant degree of protection against an oral challenge with EHEC. Complete elimination of the bacteria from the mice, was however, difficult. The combination of JGTE and LVFX increased the survival rate and reduced damage to target organs. Thus, dietary supplementation with JGTE improved the therapeutic effects of antibiotic treatment. [ABSTRACT FROM AUTHOR]

Pea proteins have been considered for the introduction into the human diet only recently. This protein source was tested on nutritional and digestive parameters in heteroxenic male Fischer rats inoculated with a human faecal microflora from a methane producer. Compared to soybean proteins, pea proteins have similar effects on the rat’s endogenous and bacterial digestive patterns. Compared to the pea proteins, a diet containing a standard meat meal enhanced the pH and the production of ammonia, while a lyophilized beef meat enhanced that of urea. The diet containing the standard meat decreases short-chain fatty acids and modifies the ratio of caecal short-chain fatty acids. Both animal diets decreased the specific activities of pancreatic proteases such as chymotrypsin (EC 3.4.21.1), trypsin (EC 3.4.21.4), and carboxypeptidase A (EC 3.4.17.1) when compared to the diet containing the pea isolate. In conclusion, the whole composition of the diet, more than the origin of the dietary protein, influences the rat’s digestive pattern. [ABSTRACT FROM AUTHOR]

Eight chlorococcalean algae and 5 rhodophycean algae have been grown in axenic cultures. These organisms have been "fingerprinted" using a pyrolysis-gas-liquid chromatographic analysis. Each alga has a distinctive pyrogram which characterizes it both quantitatively and qualitatively. The programs are given and the significance for possible future uses of this technique in developmental, evolutionary and systematic studies with algae is discussed. [ABSTRACT FROM AUTHOR]

The article provides information on about the IV International Symposium on Germfree Research entitled "Biological of Gnotobiotic Environments," held in New Orleans, Louisiana on April 16-20, 1972 sponsored by the Louisiana State Medical Center.

The development of the art of gnotobiotics has facilitated the production of germfree animals and has encouraged their use as research tools. The eventual elimination of the anomalies of germfree life by the reintroduction of the essential microflora will produce a controllable experimental animal. [ABSTRACT FROM AUTHOR]

The article discusses gnotobiotics, the study of organisms that involves careful control of germs present in gnotobiotic animals including mice, guinea pigs and chicks. The procedure involves removal of a young animal from its mother's womb through careful surgical approach to avoid exposing to mother's vagina and skin. It mentions that gnotobiotic animals give researchers the ability to compare germ-free individuals and introduce microorganisms at a time.