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L Crispatus And L Iners Probiotics

In Vitro Functional Evaluation

Probiotics: Mayo Clinic Radio

Evaluation of pH-Reducing Capacity

In order to study the capacity of lactobacilli strains to reduce pH levels, standardized cultures of the strains were grown in de Man, Rogosa and Sharpe medium anaerobically for 17 h at 37°C. Final pH was measured with the aid of a pHmeter. Assays were performed in triplicate. After performing experiments separately, the combination of the most efficient pH-reducing strains was tested. To perform this assay, both strains were co-cultivated in MRS medium for 17 h at 37°C, and final pH was measured with the aid of a pHmeter.

Identification of SCFAs and Lactate Production

To obtain the organic acid profile of selected bacteria, each strain was cultured anaerobically at 37°C in 1L fermentors , in the MRS. For each strain, two conditions were assayed: with and without pH control. In both cases, pH levels were monitored during the assay. Samples were collected throughout the assay.

For organic acid quantification, 0.8 mL of cell-free supernatant was mixed by vortexing with 0.2 mL of a mixture containing 5% meta-phosphoric acid, copper sulfate and 50 mM 4-methyl valeric acid as an internal standard. Samples were then filtered by 0.45 m pore size and diluted 1/2 and 1/10 in MilliQ water.

Alterations Of Microbiota At The Genital And Rectal Sites In Hiv

The mucosal immune system of the female reproductive tract is one of the first lines of defense against HIV-infection and other pathogens . Lying superficial to the epithelial cells of the vaginal tract is the microbiota, which exists in a symbiotic relationship with the female host. This vaginal microbiota living on the mucus layer helps to inactivate the HIV by secreting H2O2 or by decreasing the pH of the environment, among other mechanisms . In fact, lactic acid and short-chain fatty acids produced by the vaginal microbiota have reportedly antimicrobial and immune modulatory activities, indicating their potential as biomarkers of disease and/or disease susceptibility . Microbiota present in female genital tract is also associated with vaginal antiretroviral drug concentrations in HIV-infected women on antiretroviral therapy .

The current concept of a healthy vaginal microbiota includes a low-diversity, Lactobacillus rich environment. Four species of Lactobacilli are known to be the most common dominant species in the vaginal microbiome of Caucasian, Asian, Black, and Hispanic women . However, alterations of diet, inflammation, menstrual cycle, usage of hormonal contraceptives, and infection by other viruses such as HPV or HSV will affect the composition and activity of the vaginal microbiota, which may enhance the chance of HIV infection .

Sandra Borges, … Paula Teixeira, in, 2016

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Conflict Of Interest Statement

EC, SG, and DR are employed by Biopolis IM, MG-M, and CS are employed by Igenomix CS-D is employed by Ferring Pharmaceuticals. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Reduction Of Intracellular Ca2+ Concentration In Vitro

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As previously reported, i can regulate the chemotactic behavior of sperm and is pivotal to its function . Thus, we further studied the effect of adhered bacteria on i. Our results indicated that the adhered bacteria caused a reduction of human sperm i within 1 min, and L group had the lowest sperm i compared with the other groups at 2 min . As shown in Figures 2C,D, the mean sperm numbers of penetrating 1% methylcellulose medium at 1 and 2 cm away from base of the tube for the controls were 489 and 166, respectively. VNP20009 DNase I and E. coli O157:H7 reduced 50 and 80% of the sperm numbers compared with the control group at 2 cm, while L. crispatus reduced the sperm numbers by approximately 90%.

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Assessment Of Sperm Motility

A computer-assisted sperm analysis system was used to detect and analyze sperm motility after incubation with the tested bacteria for 2, 4, and 6 h. The sperm parameters related to total motility, progressive motility, linear velocity, and immotile sperm were measured. At least 200 sperms were examined in each trial. Each sperm motility assay was performed three times.

L Iners And Biofilm Formation

Biofilms are bacterial structures tightly attached to a surface, and they are known to be more resistant to the host immune response and antibiotic therapy than planktonic cells . It has been shown that biofilm formation on vaginal epithelial cells is strongly associated with vaginal infections . There is sufficient evidence that BV associates with the presence of a dense polymicrobial biofilm, in which G. vaginalis is the dominant bacterial strain on the vaginal epithelium . It has been hypothesized that Gardnerella spp. initiate biofilm formation, which supports the attachment of other BV-associated bacteria to the vaginal epithelium, further enhancing the biofilm thickness . Moreover, Gardnerella biofilms serve as barriers to antibiotics and function to protect other BVAB by preventing the penetration of antibiotics . It is generally believed that the high rate of BV recurrence is due to the formation of biofilms that protect the bacteria from antibiotic treatment, and even serve as a reservoir for pathogen regrowth .

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Urinary Tract Disorders: Vaginitis

Vaginal mucosa is rich in nutrients such as glucose or amino acids, which allows for the colonization by bacteria that forms the vaginal microbiota. Lactobacilli are the predominant microorganisms with the main following strains: Lactobacillus iners, Lactobacillus jensenii, Lactobacillus crispatus, L. gasseri, Lactobacillus vaginalis, and L. salivarius. In addition, occasionally other species coming from the digestive tract are also found, such as L. rhamnosus, L. casei, and Lactobacillus plantarum .

The presence of lactobacilli in the vaginal mucosa limits its colonization by potentially pathogen bacteria, since lactobacilli compete with those bacteria for nutrients, blocking their adhesion, and they also produce antimicrobial compounds and acids that decrease the pH, inhibiting the growth of pathogens .

All these protective mechanisms have made that lactobacilli are proposed for the maintenance of the vaginal mucosa integrity and prevention of disorders related to alteration of vaginal microbiota, such as vaginitis and vaginosis.

It has been suggested that the use of probiotics after antibiotic treatment allows a recolonization of the vaginal mucosa and causes a pH decrease, limiting the recovery of pathogens. For that reason probiotics could be useful in the prevention of recurrent urogenital infections.

David H. Martin, Michael J. Ferris, in, 2015

Can Probiotics Clear Human Papillomavirus

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Imagine a virus that most people harbor at some point in their lives, usually unaware of it because our fine-tuned immune systems clear them, yet for an unlucky numberusually womenit can be a death sentence.

The pathogen is called human papillomavirus , the main cause of cervical cancer.

A German virologist named Harald zur Hausen won the Nobel prize for linking the two, just a few decades ago. Cervical cancer numbers have dropped since then thanks to PAP screenings and a vaccine . Still, 10% of HPV infections persist bringing a dire cancer diagnosis to many, often in developing countries.

Exciting new evidence reveals that cervicovaginal microbiota plays a substantial role in the persistence or regression of the virus. In a 2016 Microbiome article, Anita Mitra and colleagues summarized the current literature and discussed potential mechanisms for the involvement of vaginal microbiota in the evolution of CIN and cervical cancer.

Microbiota and HPV

Unlike most sites in the body where diversity of microbes is a good sign, persistence of HPV is more commonly associated with low microbial diversity and dominance by only one or a few species of Lactobacillus.

Impacts on Vaginal Microbiota

Scientists are beginning to untangle possible mechanisms

First, vaginal Lactobacillus prevents colonization of bacterial vaginosis-associated bacterial species via:

  • Acidity or low pH inhibits growth of pathogens.
  • Bacteriocin production alters adhesion and viral infiltration.

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Probiotics For The Treatment Of Bacterial Vaginosis

Probiotics for the Treatment of Bacterial Vaginosis

, Naturopathic Physician and Director of Natural Product Research Consultants, Seattle, WA. Dr. Brown is a consultant to the supplement industry. He reports no relevant financial relationships to any of the probiotic strains discussed in this article.

Bacterial vaginosis is a common vaginal infection causing significant gynecologic and obstetric morbidity. Though no single pathogen has been identified as the causative agent of BV, Gardnerella vaginalis and Atopobium vaginae are commonly associated with the condition.1 Typical symptoms are said to include vaginal malodor, itching, dysuria, and thin discharge, but a substantial number of women with BV are asymptomatic.2 BV has been associated with pelvic inflammatory disease, infections following gynecologic surgery, and preterm birth.3 Estimates suggest that 40% of cases of spontaneous preterm labor and preterm birth may be associated with BV.4 Some evidence also suggests the presence of BV may increase risk of HIV-infection.5

Treatment for the condition consists of metronidazole or clindamycin, administered orally or intravaginally. However, these antimicrobials are sometimes ineffective and many women have high recurrence rates following treatment .6 A study of women with BV found that an adherent biofilm of primarily G. vaginalis and A. vaginae persisted 3 weeks after treatment with metronidazole.7

L Iners And Vaginal Dysbiosis

Compared with intestinal microflora, a typical feature of the vaginal microbial environment in healthy individuals is its extremely low bacterial diversity . There are five major community-state types in healthy premenopausal women, namely, L. crispatus-dominated CST I, L. gasseri-dominated CST II, L. iners-dominated CST III, and L. jensenii-dominated CST V, whereas CST IV is characterized by the absence of Lactobacillus species . Vaginal dysbiosis, which is defined by a high bacterial diversity and a mixture of anaerobic bacteria, is frequently associated with a variety of gynaecological diseases .

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L Iners And Infertility

Previous studies have reported that up to 40% of patients who failed assisted reproduction by in vitro fertilization had an abnormal reproductive tract microbiome . Vaginal dysbiosis, including an elevated pH value, increased flora diversity, BV, vulvovaginal candidiasis, and trichomonal vaginitis, are recognized as risk factors for infertility . Campisciano et al. reported that the abundance of L. iners was associated with an increased infertility rate . Chen et al. also recently reported that the L. iners-dominated vaginal microbiome was associated with tubal infertility and C. trachomatis infection . As a transitional species, L. iners may facilitate the transition between an abnormal and a normal vaginal microbiome under treatment or artificially high estrogen levels that are needed for IVF . It is believed that the L. iners-dominated vaginal microbiome is an unfavorable factor for

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Follow all directions on the product package. Refer to the label directions for your specific product to see if the dose should be swallowed whole, chewed, sprinkled onto food or mixed with liquid. If you are using the liquid form of this medication, carefully measure the dose using a special measuring device/spoon. Do not use a household spoon because you may not get the correct dose. If you have any questions, ask your doctor or pharmacist.

Some probiotic products may contain live bacteria . Antibiotics may prevent these products from working well. Take any product containing live bacteria at least 2 to 3 hours before or after taking antibiotics. Follow the directions for your specific product.

If you are taking this product for diarrhea due to antibiotics, do not use it if you have a high fever or for more than 2 days, unless directed by your doctor. You may have a serious problem that requires medical treatment.

If your condition lasts or gets worse, or if you think you may have a serious medical problem, seek immediate medical attention.

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Evaluation Of Antagonistic Effect Of Strains On Pathogens

Bacterial Strains and Growth Conditions

Lactobacilli strains were grown using MRS medium supplemented with 0.05% of cysteine, and incubated anaerobically at 37°C for 48 h. The antagonistic effect of analyzed strains was tested against Atopobium vaginae DSM 15829T, Gardnerella vaginalis DSM 4944, Propionibacterium acnes CECT 5684 and Streptococcus agalactiae CECT 183.

Pathogens were cultivated at 37°C anaerobically in BHI medium , supplemented with bovine serum , and cysteine in the case of A. vaginae and G. vaginalis. Lactobacilli strains were grown on MRS medium and incubated anaerobically at 37°C for 17 h.

Primary Cultures of Human Endometrial Epithelial Cells

Endometrial tissue was obtained at day 15 of the menstrual cycle from healthy donors aged 1835 years-old. Subjects diagnosed with endometriosis and/or endometritis were excluded. The study was carried out in accordance with the recommendations of local Ethical Committee at IVI Valencia, Spain with written informed consent from all subjects. All subjects gave written informed consent in accordance with the Declaration of Helsinki. The protocol was approved by the local Ethical Committee at IVI Valencia, Spain .

Viability Assessment of hEECs

Colonization Assays

Quantification of Organic Acids

Quantification of Selected Populations by Real Time PCR

Probiotics As Functional Foods

safety issues .

Tripathi and Giri suggested that the intended health benefits of probiotics can only be obtained when the food contains the required minimum viable microorganism count at the time of consumption. To deliver the health benefits, probiotic foods need to contain an adequate amount of live bacteria, at least 106107 cfu/g, although there are recent convincing data on beneficial immunological effects derived from dead cells . Also, US FDA has recommended that the minimum probiotic count in a probiotic food should be at least 106 cfu/mL but depending on the amount ingested and taking into account the effect of storage on probiotic viability, a daily intake of 108109 cfu/mL probiotic microorganisms is essential to achieve probiotic action in the human organism . However, it has been stated that probiotic products should be consumed regularly with an approximate amount of 100 g/day in order to deliver about 109 cfu/mL viable cells into the intestine .

P. Pérez-Matute, … J.A. Oteo, in, 2019

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Culture Characteristics And Gram

L. iners was first described by Falsen et al. in 1999 in vaginal and urinary tract specimens . This species had escaped our attention for a long time due to its inability to grow on MRS agar under the same culture conditions as other Lactobacillus species . Nevertheless, L. iners is characterized by small, smooth, circular, translucent, and non-pigmented colonies after 24 h of anaerobic incubation on blood agar . Indeed, most L. iners isolates can grow on MRS agar upon the addition of 15% sheep and human blood . In addition, Yoshimura et al. demonstrated that L. iners can grow on MRS agar under anaerobic conditions for a period of at least 7 days, which is evidently longer than that of other Lactobacillus species. In MRS broth with 0.5% cysteine as the reducing agent, which created the anaerobic conditions, L. iners grows slowly to its highest concentration of only 107 CFU/ml and then growth decreases after 12 h .

Isolation And Identification Of Strains

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Strain Isolation

In total, 14 samples were obtained from vaginal samples. In order to obtain isolates, vaginal samples were grown on MRS medium supplemented with 0.05% of cysteine and incubated anaerobically at 37°C for 72 h. Fourteen strains were obtained from the 100 isolates recovered. Once recovered, strains were long-term stored in glycerol for further analysis. The isolates obtained were classified on the basis of Gram staining and cell morphology. Fourteen Gram-positive, non-sporulated rods were classified as presumptive lactobacilli and thus selected for further identification.

Identification and Taxonomic Characterization of Isolates

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Genome Size And Function

L. iners has the smallest genome of 1.3 Mbp on a single chromosome among the Lactobacillus species identified so far, with its pangenome count of 2300 genes and average GC content of ~33.3% . This low genome size is comparable to those of human symbionts and parasites, and is strongly indicative of a more parasitic, host-dependent lifestyle . The genome of L. iners seems to have undergone rapid evolution events that resulted in large-scale gene loss and genome reduction, as well as the acquisition of genes, such as iron-sulfur genes, for specific adaption to the vaginal econiche .

Effect Of Tested Bacteria On The Composition Of The Vaginal Microbiota

To determine the effect of bacterial transplantation on vaginal microbial composition, we performed 16S rRNA gene sequencing of DNA isolated from vaginal secretion samples from the C, V, VD, E, and L groups. As shown in Figures 5A,B, the alpha-diversity of microorganism was higher in the C and L groups compared with the V, VD, and E groups. The NMDS analysis indicated that the samples in the C and L groups were clustered together, while the samples in the V, VD, and E groups were diverged from the C group . Besides this, the Venn diagram showed that 761 common OTUs were identified in all groups, among which 48 OTUs were identified in the C and L groups, 17 OTUs were identified in the C and V groups, 28 OTUs were identified in the C and VD groups, and 13 OTUs were identified in the C and E groups . At different taxonomic levels, we found that the number of microbial groups contained in the V, VD, and E groups were reduced compared with the C and L groups .

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