Molecular assessment of bacterial vaginosis by Lactobacillus abundance and species diversity.
BACKGROUND
To date, women are most often diagnosed with bacterial vaginosis (BV) using microscopy based Nugent scoring or Amsel criteria. However, the accuracy is less than optimal. The aim of the present study was to confirm the identity of known BV-associated composition profiles and evaluate indicators for BV using three molecular methods.


METHODS
Evaluation of indicators for BV was carried out by 16S rRNA amplicon sequencing of the V5-V7 region, a tailor-made 16S rRNA oligonucleotide-based microarray, and a PCR-based profiling technique termed IS-profiling, which is based on fragment variability of the 16S-23S rRNA intergenic spacer region. An inventory of vaginal bacterial species was obtained from 40 females attending a Dutch sexually transmitted infection outpatient clinic, of which 20 diagnosed with BV (Nugent score 7-10), and 20 BV negative (Nugent score 0-3).


RESULTS
Analysis of the bacterial communities by 16S rRNA amplicon sequencing revealed two clusters in the BV negative women, dominated by either Lactobacillus iners or Lactobacillus crispatus and three distinct clusters in the BV positive women. In the former, there was a virtually complete, negative correlation between L. crispatus and L. iners. BV positive subjects showed cluster profiles that were relatively high in bacterial species diversity and dominated by anaerobic species, including Gardnerella vaginalis, and those belonging to the Families of Lachnospiraceae and Leptotrichiaceae. Accordingly, the Gini-Simpson index of species diversity, and the relative abundance Lactobacillus species appeared consistent indicators for BV. Under the conditions used, only the 16S rRNA amplicon sequencing method was suitable to assess species diversity, while all three molecular composition profiling methods were able to indicate Lactobacillus abundance in the vaginal microbiota.


CONCLUSIONS
An affordable and simple molecular test showing a depletion of the genus Lactobacillus in combination with an increased species diversity of vaginal microbiota could serve as an alternative and practical diagnostic method for the assessment of BV.
Molecular Cell Physiology, Faculty of Earth and Life Sciences, VU University, Amsterdam, The Netherlands.|Department of Public Health, Erasmus MC, University Medical Center Rotterdam, Rotterdam, The Netherlands.
Joke A M
JA
Dols
Molecular Cell Physiology, Faculty of Earth and Life Sciences, VU University, Amsterdam, The Netherlands.
Douwe
D
Molenaar
STI Outpatient Clinic, Public Health Service of Amsterdam (GGD Amsterdam), Amsterdam, The Netherlands.
Jannie J
JJ
van der Helm
Netherlands Organisation for Applied Scientific Research (TNO), Microbiology and Systems Biology, Utrechtseweg 48, 3704HE, Zeist, The Netherlands.
Martien P M
MP
Caspers
Netherlands Organisation for Applied Scientific Research (TNO), Microbiology and Systems Biology, Utrechtseweg 48, 3704HE, Zeist, The Netherlands.
Alie
A
de Kat Angelino-Bart
Netherlands Organisation for Applied Scientific Research (TNO), Microbiology and Systems Biology, Utrechtseweg 48, 3704HE, Zeist, The Netherlands.
Frank H J
FH
Schuren
STI Outpatient Clinic, Public Health Service of Amsterdam (GGD Amsterdam), Amsterdam, The Netherlands.|Naturalis Biodiversity Center, Darwinweg 2, Leiden, The Netherlands.
Adrianus G C L
AG
Speksnijder
Molecular Cell Physiology, Faculty of Earth and Life Sciences, VU University, Amsterdam, The Netherlands.|Synthetic Systems Biology, Swammerdam Institute for Life Sciences, University of Amsterdam, Amsterdam, The Netherlands.|Manchester Centre for Integrative Systems Biology, University of Manchester, Manchester, UK.
Hans V
HV
Westerhoff
Department of Public Health, Erasmus MC, University Medical Center Rotterdam, Rotterdam, The Netherlands.
Jan Hendrik
JH
Richardus
Leiden Cytology and Pathology Laboratory, Leiden, The Netherlands.|, Present address: Achter de Hor 2, 9304 TN, Lieveren, The Netherlands.
Mathilde E
ME
Boon
Canadian Center for Human Microbiome and Probiotic Research, Lawson Health Research Institute, London, Ontorio, Canada; Department of Microbiology and Immunology, Division of Urology, Department of Surgery, Western University, London, Ontario, Canada.
Gregor
G
Reid
STI Outpatient Clinic, Public Health Service of Amsterdam (GGD Amsterdam), Amsterdam, The Netherlands.|Amsterdam Centre for Infection and Immunity (CINIMA), Academic Medical Center (AMC), University of Amsterdam, Amsterdam, The Netherlands.|Department of Dermatology, Academic Medical Center (AMC), University of Amsterdam, Amsterdam, The Netherlands.
Henry J C
HJ
de Vries
Molecular Cell Physiology, Faculty of Earth and Life Sciences, VU University, Amsterdam, The Netherlands. remco.kort@tno.nl.|Netherlands Organisation for Applied Scientific Research (TNO), Microbiology and Systems Biology, Utrechtseweg 48, 3704HE, Zeist, The Netherlands. remco.kort@tno.nl.|Micropia, Natura Artis Magistra, Plantage Kerklaan 38-40, 1018 CZ, Amsterdam, The Netherlands. remco.kort@tno.nl.
Remco
R
Kort
D000293:Adolescent; D000328:Adult; D001419:Bacteria / Q000145:classification / Q000235:genetics* / Q000302:isolation & purification; D016000:Cluster Analysis; D004269:DNA, Bacterial / Q000302:isolation & purification / Q000378:metabolism; D005260:Female; D006194:Gardnerella vaginalis / Q000235:genetics / Q000302:isolation & purification; D006801:Humans; D007778:Lactobacillus / Q000235:genetics / Q000302:isolation & purification; D064307:Microbiota; D008875:Middle Aged; D020411:Oligonucleotide Array Sequence Analysis; D016133:Polymerase Chain Reaction; D012336:RNA, Ribosomal, 16S / Q000032:analysis / Q000235:genetics / Q000378:metabolism; D017422:Sequence Analysis, DNA; D013045:Species Specificity; D016585:Vaginosis, Bacterial / Q000175:diagnosis* / Q000382:microbiology; D000070658:Vigna / Q000382:microbiology; D055815:Young Adult
D004269:DNA, Bacterial; D012336:RNA, Ribosomal, 16S
16S rRNA amplicon sequencing; Bacterial vaginosis; Gardnerella vaginalis; Gini-Simpson index; IS-profiling; Lactobacillus crispatus; Lactobacillus iners; Nucleotide-based microarrays; Vaginal microbiota