Role of Indole Production on Virulence of Vibrio cholerae Using Galleria mellonella Larvae Model.

Cell to cell communication facilitated by chemical signals plays crucial roles in regulating various cellular functions in bacteria. Indole, one such signaling molecule has been demonstrated to control various bacterial phenotypes such as biofilm formation and virulence in diverse bacteria including Vibrio cholerae. The present study explores some key factors involved in indole production and the subsequent pathogenesis of V. cholerae. Indole production was higher at 37 °C than at 30 °C, although the growth at 37 °C was slightly higher. A positive correlation was observed between indole production and biofilm formation in V. cholerae. Maximum indole production was detected at pH 7. There was no significant difference in indole production between clinical and environmental V. cholerae isolates, although indole production in one environmental isolate was significantly different. Both growth and indole production showed relevant changes with differences in salinity. An indole negative mutant strain was constructed using transposon mutagenesis and the direct effect of indole on the virulence of V. cholerae was evaluated using Galleria mellonella larvae model. Comparison to the wild type strain, the mutant significantly reduced the mortality of G. mellonella larvae which regained its virulence after complementation with exogenous indole. A gene involved in indole production and the virulence of V. cholerae was identified.

Identification and methionine analog tolerance of environmental bacterial isolates selected on methionine analog containing medium.

Methionine is the first limiting amino acid in poultry feed. Currently, methionine supplement is synthesized from an expensive chemical process requiring hazardous chemicals. Therefore, the objectives of this study were isolation of methionine producing bacteria from environmental samples and quantification of methionine production in these isolated bacteria. MCGC medium was selected as the isolation medium for methionine-producing bacteria by using Corynebacterium glutamicum ATCC13032 and Escherichia coli ATCC23798 as the positive and negative controls, respectively. Thirty-nine bacterial strains were obtained from environmental samples. Only strains A121, A122, A151 and A181 were able to tolerate up to 0.1% (w/v) of ethionine or norleucine. These isolated strains were identified by sequencing small subunit rRNA genes. The results revealed that bacterial strains A121, A122, A151and A181 were Klebsiella species, Acinetobacter baumannii, A. baumannii and Pseudomonas aeruginosa, respectively. When methionine production in strains A121 and A181 was quantitated, strains A121 and A181 generated methionine up to 31.1 and 124.6 µg/ml, respectively.

Ecology and characteristics of methanogenic archaea in animals and humans.

In this review, the molecular techniques used in animal-based-methanogen studies will be discussed along with how methanogens interact not only with other microorganisms but with their animal hosts as well. These methods not only indicate the diversity and levels of methanogens, but also provide insight on their ecological functions. Most molecular techniques have been based on either 16S rRNA genes or methyl-coenzyme M reductase, a ubiquitous enzyme in methanogens. The most predominant methanogens in animals belong to the genus Methanobrevibacter. Besides methanogens contributing to overall H2 balance, methanogens also have mutual interactions with other bacteria. In addition to shared metabolic synergism, the host animal retrieves additional energy from the diet when methanogens are co-colonized with other normal flora. By comparing genes in methanogens with other bacteria, possible gene transfer between methanogens and other bacteria in the same environments appears to occur. Finally, diets in conjunction with the genetics of methanogens and hosts may represent the biological framework that dictate the extent of methanogen prevalence in these ecosystems. In addition, host evolution including the immune system could serve as an additional selective pressure for methanogen colonization.

Characterization of isolated yeast growth response to methionine analogs.

Methionine is one of the first limiting amino acids in poultry nutrition. The use of methionine-rich natural feed ingredients, such as soybean meal or rapeseed meal may lead to negative environmental consequences. Amino acid supplementation leads to reduced use of protein-rich ingredients. The objectives of this study were isolation of potentially high content methionine-containing yeasts, quantification of methionine content in yeasts and their respective growth response to methionine analogs. Minimal medium was used as the selection medium and the isolation medium of methionine-producing yeasts from yeast collection and environmental samples, respectively. Two yeasts previously collected along with six additional strains isolated from Caucasian kefir grains, air-trapped, cantaloupe, and three soil samples could grow on minimal medium. Only two of the newly isolated strains, K1 and C1, grew in minimal medium supplied with either methionine analogs ethionine or norleucine at 0.5% (w/v). Based on large subunit rRNA sequences, these isolated strains were identified as Pichia udriavzevii/Issatchenkia orientalis. P. kudriavzevii/I. orentalis is a generally recognized as a safe organism. In addition, methionine produced by K1 and C1 yeast hydrolysate yielded 1.3 ± 0.01 and 1.1 ± 0.01 mg g(-1) dry cell. Yeast strain K1 may be suitable as a potential source of methionine for dietary supplements in organic poultry feed but may require growth conditions to further increase their methionine content.

Identification and quantification of methanogenic Archaea in adult chicken ceca.

By using molecular methods for the identification and quantification of methanogenic archaea in adult chicken ceca, 16S rRNA genes of 11 different phylotypes, 10 of which were 99% similar to Methanobrevibacter woesei, were found. Methanogen populations, as assessed by cultivation, and the 16S rRNA copy number were between 6.38 and 8.23 cells/g (wet weight) and 5.50 and 7.19 log(10)/g (wet weight), respectively.

Effects of nitro compounds and feedstuffs on in vitro methane production in chicken cecal contents and rumen fluid.

Short-chain volatile fatty acids (VFA) and methane are the products from a wide variety of microorganisms living in the gastrointestinal tract. The objective of this study was to examine effects of feedstuff and select nitro compounds on VFA and methane production during in vitro incubation of laying hen cecal contents and rumen fluid from cattle and sheep. In the first experiment, one of the three nitro compound was added to incubations containing cecal contents from laying hens supplemented with either alfalfa (AF) or layer feed (LF). Both feed material influenced VFA production and acetic acid was the primary component. Incubations with nitro ethanol and 2-nitropropanol (NP) had significantly (P<0.05) higher propionate concentrations than incubations with added nitroethane (NE). The results further indicated that incubations containing LF produced significantly (P<0.05) more butyrate than incubations with added AF. Addition of NP and LF to incubations of avian cecal flora may promote Gram-positive, saccharolytic, VFA-producing bacteria, especially Clostridium spp. which is the predominant group in ceca. Similar to VFA production, both feed materials fostered methane production in the incubations although methane was lower (P<0.05) in incubations with added nitro compound, particularly NE. In experiments 3-8, NE was examined in incubations of bovine or ovine rumen fluid or cecal contents containing either AF or LF. Unlike cecal contents, LF significantly (P<0.05) supported in vitro methane production in incubations of both rumen fluids. The results show that NE impedes methane production, especially in incubations of chicken cecal contents.