The biofilm is one life form of microorganisms (MOs). On mucous membranes of women without and with endogenous infections, they are part of the normal microbiota and cause pathologies. We have demonstrated previously the participation of biofilms in chronic forms of vulvovaginal candidiasis (VVC), the influence of other microorganisms in its formation and evolution, in bacterial vaginosis (BV) and aerobic vaginitis (AV).
To analyze the endocervical biofilms in women with or without vaginal infections (VI) comparing them with vaginal biofilms.
We studied 22 women, 9 non-pregnant (NP) and 13 pregnant (P). Each patient was gynecologically evaluated, and a vaginal sample (VS) was taken with an aspersorium and an endocervical sample was taken with cytobrush (CB). We performed a fresh examination, pH determination and amine test. Both samples were inoculated in suitable culture medium. After each one, Gram staining and optical microscopy with crystal violet were performed for the study of BF. These were put into Sabouraud broth. All samples were incubated at 35°C for 20-24 hours.
We have discovered 9 women without pathology and with normal microbiota (NM) and 13 with vaginal infections (VI): bacterial vaginosis (BV) - 6 (4P); vulvovaginal candidiasis (VVC) - 4 (3P); vaginitis and intermediate microbiota (IMB) - 3 (1E). The notable differences were: inflammatory response in the cytobrush compared to the one found in the vaginal samples of women with vaginal infections (10/13), including women with bacterial vaginosis who did not have inflammatory response in the vaginal sample. In the cytobrush of women with normal microbiota, this response occurred only in 1 case (1/9). It was also observed the formation of microfilms of Gram-positive cocci (mostly spp) in the cytobrush of 84.6% (11/13) of the women with vaginal infections and in 66.6% (6/9) of the women with normal microbiota. Among the latter, mixed biofilms were observed in 3 cases with the presence of Gram-positive Bacilli ( (anaerobic) or ).
Something that called our attention was that the formation of biofilms of and other species of and in the cytobrush of women with vaginal infections in whose vaginal samples these microorganisms were not observed nor recovered significantly. This is a risk since they can initiate an upper genital tract infection (UGTI). In the 4 P with BV, this risk is added to the risk associated with the BV. The question is whether the complications arising from this in pregnancy are not a result of such behavior. In the women with normal microbiota, the biofilms that have Gram-positive cocci can also represent a notable risk in the moment of performing instrumental procedures.
Las biopelículas constituyen una de las formas de vida de los microorganismos (MOs). En las mucosas de mujeres sin y con infecciones endógenas, integran la microbiota normal y desarrollan patologías. Previamente hemos demostrado la participación de las mismas en las formas crónicas de las candidiasis vulvovaginales (CVV), la influencia de otros microorganismos en su conformación y evolución, en la vaginosis bacteriana (VB) y en las vaginitis aeróbicas (VA).
Analizar las biopelículas endocervicales en mujeres sin y con infecciones vaginales (IV), comparándolas con las BP vaginales.
Estudiamos 22 mujeres, 9 no embarazadas (NE) y 13 embarazadas (E). Cada paciente fue estudiada ginecológicamente y se tomó muestra vaginal (MV) con hisopo y muestra endocervical con citobrush (EC). Se realizó examen en fresco, determinación del pH y prueba de aminas. Ambas muestras fueron inoculadas en medios de cultivo adecuados. A cada muestra se efectuó la coloración de Gram y se realizó la capa celular sobre el dispositivo de vidrio (DV) para el estudio de las BP. Los DV se colocaron en caldo Sabouraud. Todas las muestras se incubaron a 35°C durante 20-24 horas.
Encontramos 9 mujeres sin patología y con microbiota normal (MN) y 13 con infecciones vaginales (IV): vaginosis bacteriana (VB) - 6 (4E); candidiasis vulvovaginal (CVV) - 4 (3E); vaginitis y microbiota intermedia (VMI) - 3 (1E). Las diferencias notables fueron: hallazgo de respuesta inflamatoria en el EC comparada con la encontrada en la MV en las mujeres con IV (10/13), incluyendo a las mujeres con VB que no presentan respuesta inflamatoria en la MV. En el EC de las mujeres con MN dicha respuesta ocurrió sólo en 1 caso (1/9); y formación de BPs de cocos Gram-positivos (la mayoría spp) en el EC del 84,6% (11/13) de las mujeres con IV y en el 66,6% (6/9) de las mujeres con MN. En estas últimas se observaron BPs mixtas en 3 casos con la presencia de bacilos Gram-positivos, ( (anaerobio) o ).
Llama la atención la formación de BP de y otras especies de y en el EC de mujeres con IV en cuyas MV no se observan ni se recuperan significativamente estos microorganismos. Esto constituye un riesgo ya que las mismas pueden iniciar una infección del tracto genital superior (ITGS). En las 4E con VB, este riesgo se suma al de la VB y cabe preguntarse si las complicaciones derivadas de la misma en la gestación no son el producto de dicho comportamiento. En las mujeres con MN las BP de cocos Gram-positivos podrían representar un riesgo notable para el desarrollo de ITGS en el momento de efectuar maniobras instrumentales.
In the lower genital tract, the microorganisms (MOs) that compose the normal or usual microbiota can colonize or infect the vaginal mucosa, making biofilms (BF) of single species or mixed ones1. Both the colonizing MOs and those producing vaginal and endocervical infections can determine, in pregnant women during pregnancy and at labor, the contraction of congenital or perinatal infections.
A BF is a very dynamic sessile community of MOs, characterized by cells that are irreversibly joined to a substrate or interface between them, saturated in an extracellular matrix of polymerized substances produced by them and that show a changed phenotype with regard to the growth and gene transcription index2,3,4,5. They can have an important role, both in the infections and for protection. In general, the lower genital tract content is studied taking into consideration the planktonic MOs, but not the BF, since we do not know many physiopathological aspects that happen during the colonization and/or infections.
In the vagina, BFs of lactobacilli are responsible for the wider production of lactic acid that decreases the vaginal pH and prevents, thus, the colonization by pathogenic or potentially pathogenic MOs6.
The usual endocervical microbiota has not been deeply studied and we can assume that it can allow adherence of other MOs that are different from the vagina, due to its more alkaline pH and histological configuration with the cylindrical epithelium tissue. Therefore, in the pathology due to sexually transmitted infections (STIs), like gonorrhea and chlamydia, produced by
Is has not been reliably known if the same happens to endogenous infections, since it was always investigated the vaginal tract and not the endocervical one. Probably, the establishment of BF of other MOs in the endocervix is mediated by similar mechanisms, which according to environmental conditions can be changed by the presence of some adhesins, such as those we have seen in these STI agents.
To analyze the behavior of MOs as BF in the endocervix (BFC) in women with and without endogenous vaginal infections (VI) comparing them with the vaginal BF.
We have studied 22 women, 9 that non-pregnant (NP) and 13 pregnant (P). Each patient was gynecologically studied. It was taken their vaginal sample (VS) with cotton swabs and endocervical sampling with cytobrush (EC). The following exams were performed: wet mount test, pH determination, and amine testing with HOK at 10%. Both samples were inoculated in suitable culture medians: Sabouraud agar, Cystine Lactose Electrolyte Deficient (CLED) agar, trypticase soy agar (TSA). Gram coloration was performed with each sample and the optical microscopy on the glass equipment (GE) for the BF analysis. The GEs were put in Sabouraud broth. All the samples were incubated at 35oC for 20 to 24 hours.
We found 9 women (5 NP and 4 P) without pathology and NM and 13 with VI: bacterial vaginosis (BV) 6 (4P); vulvovaginal candidiasis (VVC) 4 (3P); vaginitis and intermediate microbiota (VIM) 3 (1P) (
The most important differences were:
• Inflammatory response in EC comparable to that found in MV in women with VI (10/13), women with BV that did not present inflammatory response in the MV were also included ( • Formation of BF of Gram-positive cocci (most of them were
• Inflammatory response in EC comparable to that found in MV in women with VI (10/13), women with BV that did not present inflammatory response in the MV were also included (
• Formation of BF of Gram-positive cocci (most of them were
In the cases 12 and 16, one can see the relevant difference on the varied formation of biofilms at vaginal and endocervical levels (
The formation of BF is an important characteristic of NM constitution from the mucosae in our organism. As it is known, the BFs have two kinds of behaviors in our organism, like NM or a pathogenicity and resistance factor in places that are usually sterile or with prosthesis10,11. Their formation participates in the infectious pathology of the lower genital tract together with immunological and allergy factors12.
We studied the in vitro vaginal BFs both in special equipment and in optical microscopy13,14,15,16. We have demonstrated that in mixed BFs of yeasts and
Our findings call the attention to the BF formation of Gram-positive cocci. From the 17 cases of Gram-positive cocci, 11 were of
The inflammatory response in endocervix in BV cases, when absent, could be happening due to the blockage of interleukin (IL)-8 that is in the vagina and not in the endocervix. Although it is increased in the IL-1 β vagina and therefore we expect an inflammatory response, it is not produced through the hydrolytic enzymes of anaerobic bacteria that, together with
Another theme for discussion with regard to BF from the genital tract is the possible influence in the appearance of late sepsis in newborns. Sepsis in such age range is divided into: early sepsis that is manifested in the first 72 hours for 7 days, and the late one, whose incidence peaks are between the second and third weeks21. The Gram-negative bacilli were the most important representations in the 1960s together with emergence of group B
In general, it is confirmed that MOs in neonatal late sepsis come from the environment25 or the use of central venous catheters, mechanical ventilation, parenteral nutrition or other invasive procedures26,27,28,29. Few investigators associate the presence of such MOs with a possible colonization from the labor channel.
Detection of SCN BF in the endocervix of a pregnant woman suggests a risk since the MOs that form them may persist, firstly as colonizers and then as infections, in neonates that normally are treated with anti-microbes with difficulties of approaching the BFs.
In the common studies regarding lower genital tract infections and during pregnancy, it is more common to give more attention to vaginal infection and forget about investigating what is happening in the endocervix, with the exception of NG or CT. However, endocervix is part of the labor channel and eventually MOs present may affect the fetus before or in labor.
The described findings allow proposing that further endocervical investigation be performed, independently from the one that is done to investigate NG and CT, especially in women with risks of premature birth or birth of low weight infants.
Formation of BF of
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