Comparison and Utilization of Repetitive-Element PCR Techniques for Typing Lactobacillus Isolates from the Chicken Gastrointestinal Tract

ABSTRACT Three repetitive-element PCR techniques were evaluated for the ability to type strains of Lactobacillus species commonly identified in the chicken gastrointestinal tract. Enterobacterial repetitive intergenic consensus PCR (ERIC-PCR) produced species- and strain-specific profiles for Lactobacillus crispatus, Lactobacillus gallinarum, Lactobacillus johnsonii, and Lactobacillus reuteri isolates. The technique typed strains within these species equally as well as pulsed-field gel electrophoresis. DNA concentration and quality did not affect the ERIC-PCR profiles, indicating that this method, unlike other high-resolution methods, can be adapted to high-throughput analysis of isolates. Subsequently, ERIC-PCR was used to type Lactobacillus species diversity of a large collection of isolates derived from chickens grown under commercial and necrotic enteritis disease induction conditions. This study has illustrated, for the first time, that there is great strain diversity within each Lactobacillus species present and has revealed that chickens raised under commercial conditions harbor greater species and strain diversity than chickens raised under necrotic enteritis disease induction conditions.

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Quantitative Detection of Clostridium perfringens in the Broiler Fowl Gastrointestinal Tract by Real-Time PCR

Applied and Environmental Microbiology ◽

10.1128/aem.71.7.3911-3916.2005 ◽

2005 ◽

Vol 71 (7) ◽

pp. 3911-3916 ◽

Cited By ~ 71

Author(s):

Mark G. Wise ◽

Gregory R. Siragusa

Keyword(s):

Gastrointestinal Tract ◽

Real Time ◽

Clostridium Perfringens ◽

Real Time Pcr ◽

Limit Of Detection ◽

Quantitative Detection ◽

Necrotic Enteritis ◽

Analytical Sensitivity ◽

Quantitative Real Time Pcr ◽

Pcr Inhibitor

ABSTRACT Strains of Clostridium perfringens are a frequent cause of food-borne disease and gas gangrene and are also associated with necrotic enteritis in chickens. To detect and quantify the levels of C. perfringens in the chicken gastrointestinal tract, a quantitative real-time PCR assay utilizing a fluorogenic, hydrolysis-type probe was developed and utilized to assay material retrieved from the broiler chicken cecum and ileum. Primers and probe were selected following an alignment of 16S rDNA sequences from members of cluster I of the genus Clostridium, and proved to be specific for C. perfringens. The assay could detect approximately 50 fg of C. perfringens genomic DNA and approximately 20 cells in pure culture. Measurements of the analytical sensitivity determined with spiked intestinal contents indicated that the consistent limit of detection with ileal samples was approximately 102 CFU/g of ileal material, but only about 104 CFU/g of cecal samples. The decreased sensitivity with the cecal samples was due to the presence of an unidentified chemical PCR inhibitor(s) in the cecal DNA purifications. The assay was utilized to rapidly detect and quantify C. perfringens levels in the gut tract of broiler chickens reared without supplementary growth-promoting antibiotics that manifested symptoms of necrotic enteritis. The results illustrated that quantitative real-time PCR correlates well with quantification via standard plate counts in samples taken from the ileal region of the gastrointestinal tract.

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Characterization of a Novel Fructosyltransferase from Lactobacillus reuteri That Synthesizes High-Molecular-Weight Inulin and Inulin Oligosaccharides

Applied and Environmental Microbiology ◽

10.1128/aem.68.9.4390-4398.2002 ◽

2002 ◽

Vol 68 (9) ◽

pp. 4390-4398 ◽

Cited By ~ 104

Author(s):

S. A. F. T. van Hijum ◽

G. H. van Geel-Schutten ◽

H. Rahaoui ◽

M. J. E. C. van der Maarel ◽

L. Dijkhuizen

Keyword(s):

Molecular Weight ◽

High Molecular Weight ◽

Lactobacillus Reuteri ◽

Bacterial Strains ◽

Potential Health ◽

Isolation And Characterization ◽

A Cell ◽

Encoding Gene ◽

First Time

ABSTRACT Fructosyltransferase (FTF) enzymes produce fructose polymers (fructans) from sucrose. Here, we report the isolation and characterization of an FTF-encoding gene from Lactobacillus reuteri strain 121. A C-terminally truncated version of the ftf gene was successfully expressed in Escherichia coli. When incubated with sucrose, the purified recombinant FTF enzyme produced large amounts of fructo-oligosaccharides (FOS) with β-(2→1)-linked fructosyl units, plus a high-molecular-weight fructan polymer (>107) with β-(2→1) linkages (an inulin). FOS, but not inulin, was found in supernatants of L. reuteri strain 121 cultures grown on medium containing sucrose. Bacterial inulin production has been reported for only Streptococcus mutans strains. FOS production has been reported for a few bacterial strains. This paper reports the first-time isolation and molecular characterization of (i) a Lactobacillus ftf gene, (ii) an inulosucrase associated with a generally regarded as safe bacterium, (iii) an FTF enzyme synthesizing both a high molecular weight inulin and FOS, and (iv) an FTF protein containing a cell wall-anchoring LPXTG motif. The biological relevance and potential health benefits of an inulosucrase associated with an L. reuteri strain remain to be established.

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Differential Metabolism of Exopolysaccharides from Probiotic Lactobacilli by the Human Gut Symbiont Bacteroides thetaiotaomicron

Applied and Environmental Microbiology ◽

10.1128/aem.00149-15 ◽

2015 ◽

Vol 81 (12) ◽

pp. 3973-3983 ◽

Cited By ~ 30

Author(s):

Alicia Lammerts van Bueren ◽

Aakanksha Saraf ◽

Eric C. Martens ◽

Lubbert Dijkhuizen

Keyword(s):

Gastrointestinal Tract ◽

Carbohydrate Metabolism ◽

Lactobacillus Reuteri ◽

Bacterial Species ◽

Transport Systems ◽

Bacteroides Thetaiotaomicron ◽

Human Gut ◽

Positive Health ◽

Content Type ◽

Commensal Species

ABSTRACTProbiotic microorganisms are ingested as food or supplements and impart positive health benefits to consumers. Previous studies have indicated that probiotics transiently reside in the gastrointestinal tract and, in addition to modulating commensal species diversity, increase the expression of genes for carbohydrate metabolism in resident commensal bacterial species. In this study, it is demonstrated that the human gut commensal speciesBacteroides thetaiotaomicronefficiently metabolizes fructan exopolysaccharide (EPS) synthesized by probioticLactobacillus reuteristrain 121 while only partially degrading reuteran and isomalto/malto-polysaccharide (IMMP) α-glucan EPS polymers.B. thetaiotaomicronmetabolized these EPS molecules via the activation of enzymes and transport systems encoded by dedicated polysaccharide utilization loci specific for β-fructans and α-glucans. Reduced metabolism of reuteran and IMMP α-glucan EPS molecules may be due to reduced substrate binding by components of the starch utilization system (sus). This study reveals that microbial EPS substrates activate genes for carbohydrate metabolism inB. thetaiotaomicronand suggests that microbially derived carbohydrates provide a carbohydrate-rich reservoir forB. thetaiotaomicronnutrient acquisition in the gastrointestinal tract.

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Study on a Fermented Whole Wheat: Phenolic Content, Activity on PTP1B Enzyme and In Vitro Prebiotic Properties

Molecules ◽

10.3390/molecules24061120 ◽

2019 ◽

Vol 24 (6) ◽

pp. 1120 ◽

Cited By ~ 3

Author(s):

Diletta Balli ◽

Maria Bellumori ◽

Paolo Paoli ◽

Giuseppe Pieraccini ◽

Monica Di Paola ◽

...

Keyword(s):

Lactobacillus Reuteri ◽

Tyrosine Phosphatase ◽

Bacterial Species ◽

Food Ingredient ◽

Microbial Composition ◽

Acidic Hydrolysis ◽

Inhibitory Power ◽

Methyl Ferulate ◽

First Time

Fermented cereals, staple foods in Asia and Africa, are recently receiving a growing interest in Western countries. The object of this work is the characterization of a fermented wheat used as a food ingredient and dietary supplement. To this aim, the phenolic composition, the activity on protein tyrosine phosphatase 1B (PTP1B), an enzyme overexpressed in type-II diabetes, the in vitro prebiotic properties on Lactobacillus reuteri and the microbial composition were investigated. Basic and acidic hydrolysis were tested for an exhaustive recovery of bound phenols: the acidic hydrolysis gave best yields. Methyl ferulate and neocarlinoside were identified for the first time in wheat. The inhibitory power of the extracts of several batches were investigated on PTP1B enzyme. The product was not able to inhibit the enzyme, otherwise, for the first time, a complete inhibition was observed for schaftoside, a major C-flavonoid of wheat. The microbial composition was assessed identifying Lactobacillus, Enterococcus, and Pediococcus as the main bacterial species. The fermented wheat was a suitable substrate for the grown of L. reuteri, recognized for its health properties in the human gut. The proposed method for phenols is easier compared to those based on strong basic hydrolysis; our results assessed the bound phenols as the major fraction, differently from that suggested by the literature for fermented cereals.

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Molecular Epidemiology of Rotavirus A Strains Pre- and Post-Vaccine (Rotarix®) Introduction in Mozambique, 2012–2019: Emergence of Genotypes G3P[4] and G3P[8]

Pathogens ◽

10.3390/pathogens9090671 ◽

2020 ◽

Vol 9 (9) ◽

pp. 671 ◽

Cited By ~ 2

Author(s):

Eva D. João ◽

Benilde Munlela ◽

Assucênio Chissaque ◽

Jorfélia Chilaúle ◽

Jerónimo Langa ◽

...

Keyword(s):

Acute Diarrhea ◽

Sentinel Site ◽

Predominant Genotype ◽

Post Vaccination ◽

Rotavirus A ◽

Strain Diversity ◽

Monovalent Vaccine ◽

Group A ◽

Expanded Program On Immunization ◽

First Time

Group A rotavirus (RVA) remains the most important etiological agent associated with severe acute diarrhea in children. Rotarix® monovalent vaccine was introduced into Mozambique’s Expanded Program on Immunization in September 2015. In the present study, we report the diversity and prevalence of rotavirus genotypes, pre- (2012–2015) and post-vaccine (2016–2019) introduction in Mozambique, among diarrheic children less than five years of age. Genotyping data were analyzed for five sentinel sites for the periods indicated. The primary sentinel site, Mavalane General Hospital (HGM), was analyzed for the period 2012–2019, and for all five sites (country-wide analyses), 2015–2019. During the pre-vaccine period, G9P[8] was the most predominant genotype for both HGM (28.5%) and the country-wide analysis (46.0%). However, in the post-vaccine period, G9P[8] was significantly reduced. Instead, G3P[8] was the most common genotype at HGM, while G1P[8] predominated country-wide. Genotypes G9P[4] and G9P[6] were detected for the first time, and the emergence of G3P[8] and G3P[4] genotypes were observed during the post-vaccine period. The distribution and prevalence of rotavirus genotypes were distinct in pre- and post-vaccination periods, while uncommon genotypes were also detected in the post-vaccine period. These observations support the need for continued country-wide surveillance to monitor changes in strain diversity, due to possible vaccine pressure, and consequently, the effect on vaccine effectiveness.

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Candida albicans forms biofilms on the vaginal mucosa

Microbiology ◽

10.1099/mic.0.039354-0 ◽

2010 ◽

Vol 156 (12) ◽

pp. 3635-3644 ◽

Cited By ~ 168

Author(s):

M. M. Harriott ◽

E. A. Lilly ◽

T. E. Rodriguez ◽

P. L. Fidel ◽

M. C. Noverr

Keyword(s):

Biofilm Formation ◽

Ex Vivo ◽

Microscopic Analysis ◽

Vaginal Mucosa ◽

First Time ◽

Established Role ◽

Type Strains

Current understanding of resistance and susceptibility to vulvovaginal candidiasis challenges existing paradigms of host defence against fungal infection. While abiotic biofilm formation has a clearly established role during systemic Candida infections, it is not known whether C. albicans forms biofilms on the vaginal mucosa and the possible role of biofilms in disease. In vivo and ex vivo murine vaginitis models were employed to examine biofilm formation by scanning electron and confocal microscopy. C. albicans strains included 3153A (lab strain), DAY185 (parental control strain), and mutants defective in morphogenesis and/or biofilm formation in vitro (efg1/efg1 and bcr1/bcr1). Both 3153A and DAY815 formed biofilms on the vaginal mucosa in vivo and ex vivo as indicated by high fungal burden and microscopic analysis demonstrating typical biofilm architecture and presence of extracellular matrix (ECM) co-localized with the presence of fungi. In contrast, efg1/efg1 and bcr1/bcr1 mutant strains exhibited weak or no biofilm formation/ECM production in both models compared to wild-type strains and complemented mutants despite comparable colonization levels. These data show for the first time that C. albicans forms biofilms in vivo on vaginal epithelium, and that in vivo biotic biofilm formation requires regulators of biofilm formation (BCR1) and morphogenesis (EFG1).

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Organization of the micronuclear genome of oxytricha nova.

Genetics ◽

10.1093/genetics/120.1.123 ◽

1988 ◽

Vol 120 (1) ◽

pp. 123-134

Author(s):

C L Jahn ◽

K E Prescott ◽

M W Waggener

Keyword(s):

Repetitive Dna ◽

Repetitive Sequence ◽

Repetitive Element ◽

Repetitive Sequences ◽

Unique Sequence ◽

Sequence Family ◽

Consensus Map ◽

Ciliated Protozoan ◽

Oxytricha Nova ◽

First Time

Abstract In the hypotrichous ciliated protozoan Oxytricha nova, approximately 95% of the micronuclear genome, including all of the repetitive DNA and most of the unique sequence DNA, is eliminated during the formation of the macronuclear genome. We have examined the interspersion patterns of repetitive and unique and eliminated and retained sequences in the micronuclear genome by characterizing randomly selected clones of micronuclear DNA. Three major classes of clones have been defined: (1) those containing primarily unique, retained sequences; (2) those containing only unique, eliminated sequences; and (3) those containing only repetitive, eliminated sequences. Clones of type one and three document two aspects of organization observed previously: clustering of macronuclear destined sequences and the presence of a prevalent repetitive element. Clones of the second type demonstrate for the first time that eliminated unique sequence DNA occurs in long stretches uninterrupted by repetitive sequences. To further examine repetitive sequence interspersion, we characterized the repetitive sequence family that is present in 50% of the clones (class three above). A consensus map of this element was obtained by mapping approximately 80 phage clones and by hybridization to digests of micronuclear DNA. The repeat element is extremely large (approximately 24 kb) and is interspersed with both macronuclear destined sequences and eliminated unique sequences.

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Survival and persistence of Lactobacillus plantarum 4.1 and Lactobacillus reuteri 3S7 in the gastrointestinal tract of pigs

Veterinary Microbiology ◽

10.1016/j.vetmic.2007.02.022 ◽

2007 ◽

Vol 123 (1-3) ◽

pp. 133-144 ◽

Cited By ~ 23

Author(s):

Maria De Angelis ◽

Sonya Siragusa ◽

Leonardo Caputo ◽

Adriano Ragni ◽

Roberto Burzigotti ◽

...

Keyword(s):

Gastrointestinal Tract ◽

Lactobacillus Plantarum ◽

Lactobacillus Reuteri

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Microscopic, cultural and molecular evidence of disseminated invasive aspergillosis involving the lungs and the gastrointestinal tract

Journal of Medical Microbiology ◽

10.1099/jmm.0.46394-0 ◽

2006 ◽

Vol 55 (7) ◽

pp. 961-964 ◽

Cited By ~ 17

Author(s):

R. S. Jayshree ◽

M. Shafiulla ◽

J. George ◽

J. K. David ◽

P. P. Bapsy ◽

...

Keyword(s):

Gastrointestinal Tract ◽

Invasive Aspergillosis ◽

Aspergillus Flavus ◽

Molecular Evidence ◽

Dna Analyses ◽

Random Amplification ◽

Fungal Hyphae ◽

Gastrointestinal Aspergillosis ◽

Disseminated Disease ◽

First Time

A patient with acute promyelocytic leukaemia developed invasive aspergillosis post chemotherapy during a pancytopenic episode, clinically involving the lungs and the gastrointestinal tract. Dichotomously branched septate fungal hyphae were demonstrated microscopically in stools and sputa. Cultures of the samples yielded Aspergillus flavus, which were identical by RFLP and random amplification of polymorphic DNA analyses and antifungal MICs, proving disseminated disease. To the best of the author's knowledge, this is the first time that boluses of fungal hyphae have been demonstrated microscopically in the stools of a patient with gastrointestinal aspergillosis.

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