Review
Lactobacillus iners Friend or
Foe
Mariya I Petrova12 Gregor Reid3 Mario Vaneechoutte4 and
Sarah Lebeer12*
The vaginal microbial community is typically characterized by abundant lacto
bacilli Lactobacillus iners a fairly recently detected species is frequently
present in the vaginal niche However the role of this species in vaginal health
is unclear since it can be detected in normal conditions as well as during vaginal
dysbiosis such as bacterial vaginosis a condition characterized by an abnor
mal increase in bacterial diversity and lack of typical lactobacilli Compared to
other Lactobacillus species L iners has more complex nutritional requirements
and a Gramvariable morphology L iners has an unusually small genome (ca 1
Mbp) indicative of a symbiotic or parasitic lifestyle in contrast to other lacto
bacilli that show niche flexibility and genomes of up to 3–4 Mbp The presence of
specific L iners genes such as those encoding iron–sulfur proteins and unique
sfactors reflects a high degree of niche specification The genome of L iners
strains also encodes inerolysin a poreforming toxin related to vaginolysin of
Gardnerella vaginalis Possibly this organism may have clonal variants that in
some cases promote a healthy vagina and in other cases are associated with
dysbiosis and disease Future research should examine this friend or foe
relationship with the host
Members of the Vaginal Microbiota
Microbes are recognized as playing an important role in the balance between health and disease in
the human host While the recent advances in DNA sequencing techniques have revealed that an
increased diversity in the gut microbiota is generally linked to less susceptibility to disease the
opposite seems to be true for the vaginal microbiota [12] These studies confirm previous culture
and microscopydependent findings that – under healthy conditions – the vaginal econiche is
dominated by a limited number of Lactobacillus species in most women When diversity increases
so does the risk of bacterial vaginosis (BV) (see Box 1) or bacterial vaginal dysbiosis (see Glossary)
A characteristic of BV is the formation of dense multispecies biofilms [3] The onset of BV and
depletion of lactobacilli can have severe outcomes such as preterm birth [4] reduced birth weight
and increased risk for and transmission of genital infections [56] In contrast various studies have
reported that Lactobacillus strains can promote a healthy vaginal ecosystem by actively interfering
with bacterial fungal and viral pathogens (as reviewed recently by [7])
The most frequently isolated species from the healthy vagina are Lactobacillus crispatus
Lactobacillus gasseri Lactobacillus jensenii and Lactobacillus iners (Figure 1) Efforts have
been made to group or grade vaginal microbiota patterns according to the dominant (Lactoba
cillus) species present as well as to link these patterns or community types to vaginal health
status For example Verhelst et al [8] suggested the existence of different grades based upon
Gram staining sequencing of 16S rRNA genes and culture In their grades Ia and Iab L
crispatus is the dominant Lactobacillus species followed by L jensenii while in grade Ib L iners
Trends
Recent microbiome data substantiate
previous culturedependent and cul
tureindependent data that the vaginal
econiche is generally dominated by
Lactobacillus species in women with
out vaginal dysbiosis
Lactobacillusdominated vaginal micro
biota profiles can be further divided in
community types dominated by either
Lactobacillus crispatus [2_TDDIFF]Lactobacillus
iners [2_TDDIFF]Lactobacillus gasserior
[2_TDDIFF]Lactobacillus jenseni
An L inersdominated vaginal commu
nity type seems to be less stable or more
in transition than the other community
types and more associated with vaginal
dysbiosis
Genome sequencing revealed that L
iners has an unusually small genome
indicative of a parasitic or symbiotic
lifestyle in the human vagina
Biochemical and functional assays
suggest that L iners bacteria contain
features of probiotic lactobacilli as well
as of vaginal pathogens
1KU Leuven Centre of Microbial and
Plant Genetics Kasteelpark Arenberg
20 box 2460 B 3001 Leuven Belgium
2University of Antwerp Department of
Bioscience Engineering
Groenenborgerlaan 171 B2020
Antwerp Belgium
3Western University Departments of
Microbiology & Immunology and
Surgery and The Lawson Health
Research Institute London 268
Grosvenor Street London ON Canada
N6A 4V2
4Ghent University Laboratory of
Bacteriology Research Faculty of
Medicine & Health Sciences De
Pintelaan 185 Medical Research
Building 2 B9000 Gent Belgium
*Correspondence
sarahlebeer@uantwerpenbe (S Lebeer)
182 Trends in Microbiology March 2017 Vol 25 No 3 httpdxdoiorg101016jtim201611007
© 2016 Elsevier Ltd All rights reservedGlossary
Biofilms are communities of
adhering bacteria forming robust
layers on abiotic and biotic surfaces
The adherent cells are surrounded by
a selfproduced matrix of extracellular
polymeric substances Biofilms are
major clinical issues since the
pathogens in the biofilms are difficult
to eradicate
Clustered regularly interspaced
short palindromic repeats
(CRISPR) are segments of
prokaryotic DNA containing short
repetitions of base sequences
CRISPR proteins are considered to
be important antibacteriophage
defense systems in bacteria
Dysbiosis in this review the term is
used to refer to an imbalance in the
composition of the vaginal microbiota
such as during bacterial vaginosis
(BV) Besides BV another common
but not wellstudied vaginal
dysbiosis condition is aerobic
vaginitis (AV) This is an
immunological disorder mainly
characterized by culture by a
dominance of aerobic bacteria such
as Streptococcus spp
Staphylococcus spp and
Escherichia coli
European Food and Safety
Authority (EFSA) is a European
agency funded by the European
Union providing independent
scientific opinions and advice that
form the basis for European policies
and legislation associated with the
food chain
Nugent score is used to diagnose
BV and is based on the Gram
staining of vaginal smears It includes
the microscopic quantitation of
bacterial cell morphology types
yielding a score between 0 and 10 A
score of 0–3 is considered normal
4–6 is intermediate and 7–10 is
considered BV
Probiotics live microorganisms
which when administrated to the
host in adequate amounts confer
health benefits Most of the time
Lactobacillus species or
Bifidobacterium species
Qualified presumption of safety
(QPS) is a generic assessment
system within EFSA that can be
applied to all requests received for
the safety assessments of
microorganisms introduced into the
food chain
and L gasseri are predominant Grade II represents an intermediate status between grade I and
grade III with the presence of L iners L gasseri L crispatus Atopobium vaginae Gardnerella
vaginalis Actinomyces neuii and Peptoniphilus spp Grade III is characterized by BVassociated
species (G vaginalis Aerococcus christensenii A vaginae Bacteroides ureolyticus Dialister
sp Mobiluncus curtisii Prevotella bivia and Varibaculum cambriense) and Lactobacillus
species mainly L iners Using deepsequencing approaches Hummelen et al [9] pointed
out different bacterial abundance profiles associated with health and BV The findings of these
groups have been confirmed using deepsequencing approaches by Ravel et al [1] who
proposed five major community state types in healthy asymptomatic women in their dataset
that is community state types I (262) II (63) III (341) and V (53) dominated by L
crispatus L gasseri L iners and L jensenii respectively (Figure 1) The remaining 27 of the
women forming community state type IV were part of a heterogeneous group characterized by
strictly anaerobic bacteria including Prevotella Dialister Atopobium Gardnerella Mega
sphaera and Peptoniphilus as well as L crispatus and L iners [1] This last group most
probably corresponds to BV typically characterized by an increased diversity of species with
the most commonly isolated species being G vaginalis P bivia A vaginae Bacteroides spp
Dialister spp Mobiluncus spp Eggerthella spp and Megasphaera spp as well as Bacterial
VaginosisAssociated Bacteria 1 2 and 3 (BVAB 1 BVAB 2 and BVAB 3) [1011] (Figure 1) The
detection of L iners species in both BV patients and healthy subjects as well as its very different
genomic and phenotypic characteristics compared to other Lactobacillus species [12] raises
the question of not only how this species can adapt so effectively to the vagina but also whether
this species can actually play an active role in dysbiotic onset as well as recovery from dysbiosis
This review aims to present an overview of different arguments that point towards a dual role for
this species in the vaginal niche
History and Discovery of L iners
For many years culture and microscopydependent approaches aimed at the detection of
lactobacilli in the human vagina failed to identify L iners It was not until 1999 that the species
was named L iners (from iners inert lazy) following Falsen and coworkers [13] who isolated
seven strains from healthy vaginal and urinary tract samples The organism was described as
Grampositive with rodshaped cells occurring mostly as single cells in pairs or in short chains
The growth of the strains occurred after 24 h of incubation under anaerobic conditions on blood
agar and the colonies were small smooth and not pigmented According to one report the
Box 1 Bacterial Vaginosis
One of the most common vaginal disorders is bacterial vaginosis (BV) resulting in millions of health care visits annually
around the world BV is a complex polymicrobial condition characterized by disruption of the vaginal econiche resulting
in a reduction in lactobacilli and an overgrowth of strict or facultatively anaerobic bacteria such as Gardnerella spp
Atopobium spp Prevotella spp and Mobiluncus spp as well as other taxa such as Clostridium Megasphera spp and
Leptotrichia spp [5758] In addition cultureindependent techniques have identified three potentially BVassociated
bacteria (BVAB) BVAB1 BVAB2 and BVAB3 The number of sexual partners a new partner hormonal changes
personal hygiene smoking and antibiotic treatment have been reported to be factors important for the development of
BV but the exact cause of BV is still unknown [5960] A hallmark of BV is the presence of a dense polymicrobial biofilm on
the vaginal epithelial surface assumed to be initiated by G vaginalis strains which then become a scaffold for other
species to adhere to [6162] It is still uncertain whether these species are actually involved in the development of BV or if
they are opportunistic organisms which are well adapted to the changes in the vaginal environment When symptomatic
BV is characterized by vaginal discomfort and homogeneous malodorous vaginal discharge However in many cases BV
is asymptomatic and most of the time noninflammatory When asymptomatic it is still debated whether this type of
microbiota is reflecting an asymptomatic state of BV or rather a healthy state Even though this type of microbiota is
nonlactobacilli dominated the abundant species might be able to maintain the protective function of the vaginal niche
that is a low vaginal pH by production of lactic acid [36] but this remains to be confirmed However women harboring
this diverse microbiota type can be of high risk of various vaginal and urinary tract infections increased risk of sexually
transmitted infections such as Neisseria gonorrhoeae Chlamydia trachomatis HIV and HSV2 and increased rates of
early pregnancy loss and preterm delivery [763] Therefore close and regular monitoring of women harboring a diverse
type of microbiota is required
Trends in Microbiology March 2017 Vol 25 No 3 183species is not always clearly Grampositive and at least some isolates seem to have a
coccobacillary rather than bacillary morphology possibly one of the reasons why the species
has been overlooked by Gram staining of vaginal smears of women with vaginal dysbiosis [14]
The cell morphology and surface properties of many Lactobacillus species and strains can vary
for example under in vivo versus in vitro conditions [15] This could also be hypothesized for L
iners strains because genome analyses suggest large strain differences [16] This ambiguity in
cell morphology and Gramstaining properties of L iners is important to consider since many
diagnostic determinations of vaginal health depend on these characteristics In fact the
commonly used Nugent score is based on Gram staining of the vaginal fluid and differentiation
of lactobacilli as presumptive Grampositive rods versus other cell morphology types [17]
In addition to its peculiar cell morphology the genome size of L iners is also unique among
Lactobacillus species L iners strain AB1 has by far the smallest Lactobacillus genome
determined so far consisting of a 13 Mbp single chromosome [12] This genome appears
to have undergone one or more rapid evolutionary events resulting in massive gene loss but it
also acquired genes for optimal survival in the vagina such as genes encoding iron–sulfur
Healthy state Healthy or transional
Vaginal microbiota
Dysbiosis–BV state
Group I
L crispatusdominated microbiota
pH 4
Group II
L gasseridominated microbiota
pH 5
Group III
L jenseniidominated microbiota
pH 47
Group IVB diverse microbiota
pH 53
Group III
L inersdominated microbiota
Moderate lacc acid producon
Isolated from both healthy and BV state
Dominang spp following BV treatment
Isolated from transional type
vaginal microbiota
Overgrowth facultave or strict anaerobes
Gardnerella spp Atopobium Spp
Prevotella spp Mobiluncus spp
Megasphera spp BVAB123
Group IVA diverse microbiota
pH 45
Figure 1 Composition of Vaginal Microbiota The vaginal microbiota of premenopausal women can be divided into different community types but their exact
number is still under discussion because it depends on the dataset and methods used The most commonly identified dominant species governing these community
types belong to Lactobacillus crispatus [2_TDDIFF]Lactobacillus gasseri [2_TDDIFF]Lactobacillus jensenii and [2_TDDIFF]Lactobacillus iners [18936] In addition nonLactobacillus dominated vaginal
microbiota community types have also been reported For example group IVA is characterized by a modest proportion of either L crispatus L iners or other
Lactobacillus species as well as low numbers of strict anaerobic bacteria Group IVB is dominated by diverse bacteria belonging mainly to the genera Atopobium
Prevotella Parvimonas Sneathia GardnerellaorMobiluncus [36] Different community types can be associated with health and dysbiosis and a possible transition state
between the two Since L iners has often been isolated during menstruation and in microbiota profiles being in transition between healthy and BV state here we focus on
its role in maintaining vaginal health
184 Trends in Microbiology March 2017 Vol 25 No 3proteins and unique sfactors (see below) Subsequent studies have confirmed that the small
genome size of L iners AB1 appears to be a species characteristic since similar sizes (13
Mbp) have been reported for all other L iners genomes in the National Center for Biotechnology
Information (NCBI) The small genome is similar in size to that of human symbionts and parasites
while facultative human symbionts have intermediate genome sizes [18] Therefore L iners
could be a genuine vaginal symbiont but also a potential opportunistic parasitic’ pathogen
Dynamic Presence in the Vagina in Health and Disease
The original identification of L iners occurred in 2002 in four out of 23 Swedish women with a
normal Nugent score using randomly amplified polymorphic DNA (RAPD) analysis with an
identification by means of temporal temperature gradient gel electrophoresis [19] That same
year L iners was detected using denaturing gradient gel electrophoresis in Canadian women
with a normal Nugent score and in three subjects with a BV score [20] Since then L iners has
been found to be one of the most frequently isolated organisms in the vagina [21–26] This is a
universal finding from samples collected in Nigeria [25] Brasil [27] and China [21] and in
American women of different ethnic origin [1] Although Srinivasan et al [28] reported that the
vagina of 93 of American women without BV were dominated by L iners or L crispatus they
noted that women with high levels of L iners could be either BVpositive or BVnegative and
have either low or high pH levels [28] The presence of L iners during BVassociated conditions
[8914202930] is thus well documented Nevertheless this does not mean that it plays a
causative role in dysbiosis (Figure 2 Key Figure) With an estimated 50 women reportedly
harbouring L iners [13132] its continued presence during BV is perhaps not surprising This
clear adaptation to the vagina is reflected in a study showing similar prevalence of L iners in
women with normal intermediate and BV Nugent scores [33] Nevertheless not merely the
presence but also the relative abundance of L iners is probably important More research is thus
required to quantify the relative and absolute numbers of L iners under different conditions
Shipitsyna et al [10] reported on the depletion of L iners during BV arguing against it being a key
etiologic agent in the condition However others have suggested that when the microbiota is
dominated by L iners it is more likely to shift towards dysbiosis – unlike when L crispatus is
dominant [34] A semiquantitative study (using speciesspecific qPCRs) of Belgian (mostly
Caucasian) women sampled throughout the menstrual cycle found a clearly inversed preva
lence between L iners and L crispatus with the latter largely absent during episodes of BV
during the menses and after sexual intercourse whereby in most of those instances L iners
became the predominant species (together with G vaginalis) [229] L iners appears to be better
adapted than L crispatus to changing vaginal conditions such as during BV
The correlation between vaginal microbiota dominated by L iners and sexually transmitted
infections (STIs) has also been investigated For example Borgdorff et al [35] have recently
reported that L crispatusdominated vaginal microbiota and to a lesser extent L iners
dominated microbiota are associated with a lower prevalence of HIV HSV2 HPV and bacterial
STIs (Neisseria gonorrhoeae Chlamydia trachomatis and Trichomonas vaginalis combined)
However viral and bacterial STIs were significantly increased in dysbiotic clusters A similar trend
was found for HIV1 RNA shedding in the genital tract of HIVpositive women with microbiota
dominated by L crispatus and L iners showing a lower likelihood of genital HIV1 RNA shedding
[35]
Functional analyses are required to better understand how L iners responds to vaginal con
ditions An important study by Macklaim et al [16] reported that during BV genes for the uptake
of mannose and maltose as well as genes for the breakdown of glycogen were strongly
upregulated in L iners compared to healthy conditions To explore whether L iners also plays an
active role in driving the changing conditions during BV in addition to merely responding to it
Trends in Microbiology March 2017 Vol 25 No 3 185Key Figure
Lactobacillus iners Friend or Foe
Normal
microbiota
Beneficial
lactobacillus
BV
microbiota
Transional
microbiota
Opportunisc
pathogen
Hormonal changes
Personal hygiene
Other unknown factors
Anbioc treatment
Other unknown factors
Lactobacillus iners
Genes for opmal survival in the vaginal niche
1 Ironsulfur cluster assembly system
2 Unique σfactors
3 Alkalineshock proteins
heatshock proteins
and chaperonins
4 Fibronecn binding protein
5 Cholesteroldependent cytolysin (inerolisin)
6 Mucin glycogen and glycerol transports
and metabolic enzymes
7 Mannose type PTS
8 Genes belonging to CRISPR system
1 Cholesteroldependent cytolysin (inerolysin)
2 Mucin and glycerol transport and metabolic enzymes
3 Genes belonging to CRISPR system
4 Mannosetype PTS
5 Genes for uptake and metabolism of glycogen
Genes upregulated during BV condions
Hormonal changes
Sexual intercourse
Smoking
Personal hygiene
Ethnicity
Other unknown factors
Anbioc treatment
Figure 2 [3_TDDIFF]L iners is reported to be a dominant species in the vaginal microbiota of healthy women However L iners is also often isolated from the vaginal microbiota of
women with BV characterized by a diverse set of bacteria Of interest L iners is also reported to be a dominant species in the transitional type of the vaginal microbiota
[284064] or during menses [293637] suggesting that this species is very flexible and can easily adapt to the fluctuating vaginal niche The genome analysis supports
both a beneficial and an ambiguous role for L iners in the vaginal niche For example the genome of L iners encodes proteins predicted to be involved in the optimal
adaptation to the vaginal niche such as iron–sulfur proteins and unique sfactors [4_TDDIFF]By [5_TDDIFF]contrast the small genome size could indicate that L iners is a symbiotic species
optimally adapted to the vaginal niche [1216] L iners also encodes some genes suggesting that it could be an opportunistic pathogen of which inerolysin a potential
cholesteroldependent cytolysin is the best documented [1251] Taken together the presence of L iners in the vaginal niche is not a good biomarker for vaginal health in
contrast to the presence of other frequently identified Lactobacillus species Abbreviation PTS phosphoenolpyruvatedependent transport system
186 Trends in Microbiology March 2017 Vol 25 No 3sequential samples would need to be collected Alterations in mucus pH menstrual blood and
epithelial sloughing occur in the vagina and external factors such as seminal fluid all can change
the environment and perhaps elicit metabolic adaptation by L iners which could then encour
age proliferation of BV pathogens Indeed L iners numbers and gene expression can be
affected by the stages of the menstruation cycle [23637] Alternatively the physiological
changes could promote BV and thereafter L iners simply adapts to it Resolving the order
of these reactions will be necessary to determine the extent to which L iners is deleterious to the
host if at all
By definition a pathogen should oppose injure or impede the host with the net result being a
negative outcome Suffering from BVinduced preterm labour would be deemed such a negative
outcome but the mere domination of the vaginal microbiota by L iners in women with a higher
risk of preterm labour still would not count as sufficient cause and effect [38] Such a correlation
would be strengthened if L iners had been shown to cause endometritis or other reproductive
tract infections but this has not been the case so far
[6_TDDIFF]By [5_TDDIFF]contrast the argument that L iners is a beneficial species is supported by studies showing
that the organism is associated with homeostasis Ferris et al [39] analyzed the vaginal
microbiota of patients treated with metronidazole gel and found at 30 days posttreatment
that L iners was the only dominant species This could indicate that L iners is the only
metronidazoleresistant Lactobacillus species Jakobsson et al [40] proposed that L iners
may become or is a dominant species when the microbiota is in a transitional stage between
abnormal and normal (Figure 2) The idea is that by surviving under BV conditions [41] L iners
helps recovery to a lactobacillidominated microbial community [37] The fact that postBV
treatment the microbiota remains L inersdominated and not L crispatusdominated might
support the argument in favor of L iners as a friend’ However it raises the question of why does
L crispatus not behave in this way survive BV and help the recovery process It could also be
that L crispatus is more susceptible than L iners to the metronidazole therapy during BV and the
other stressful conditions during BV A detailed comparative genomic analysis might answer this
question
Moreover the dynamics of L iners versus L crispatus during menses needs to be further
explored In one study L crispatus appears to decline during menses while G vaginalis
increases along with L iners and subsequently the concentration of both species decreases
after menses [37] In other studies L crispatus has also been reported to decline 100fold during
menses while the numbers of L iners strongly increase [22936] However Chaban et al [42]
did not observe significant changes in the vaginal microbiota during specific menstrual phases
for 18 out of 26 women In their study four women (15) that had clear changes in the
microbiota composition transitioned from an L inersdominated to an L crispatusdominated
status This might indicate that although temporal changes can be observed withinsubject
microbiota variation over time is still consistently lower than betweensubject variation both in
microbial composition and metabolic function [43]
Gajer et al [36] observed that vaginal communities dominated by L crispatus can transform to a
community state dominated by L iners or to a community state characterized by a modest
proportion of either L crispatus or L iners Another indication of the lower protective value of
colonization with L iners is that a community group dominated by L iners appears to shift more
often to one dominated by diverse bacteria belonging to the genera Atopobium Prevotella
Parvimonas Sneathia GardnerellaorMobiluncus Analysis of the metabolite profiles showed
that high levels of lactate were maintained even during menses when communities dominated by
L crispatus transformed to L iners However a decrease in lactate and an increase in succinate
and acetate were noted during major changes in the composition of the vaginal microbiota that
Trends in Microbiology March 2017 Vol 25 No 3 187lasted for several weeks characterized by a shift of the microbiota dominated by L iners to one
dominated by Atopobium sp [36] Such microbial dynamics have not been reported at any other
body site to our knowledge and they require further exploration
L iners has been more often detected in Black African and AfroAmerican women compared to
Caucasian or Asian women For example a few studies show that black women are less likely to
carry L crispatus L jensenii L gasseri andor L vaginalis and more likely to carry L iners
[1284445] However the reasons for such a differences between ethnic groups is still not
clear but could be based on differences in innate and adaptive immune system epithelial cell
surface quantity of vaginal secretion and other genetically determined differences between
hosts
Peculiar Genetic and Biochemical Features
The genome of L iners AB1 contains several genes putatively involved in optimal survival in the
fluctuating vaginal environment (Figure 2) For example L iners AB1 encodes an iron–sulfur
protein cluster (Fe–S) mainly detected in vaginal Lactobacillus isolates since the complete
system is rarely detected in lactobacilli inhabiting other niches [12]Fe–S cluster proteins are
suggested to be involved in resistance to oxidative stress which could be elevated in the vaginal
niche where high levels of H2O2 are produced by other lactobacilli [12] In addition L iners AB1
contains two alkaline shock proteins various heatshock proteins coldshock proteins and
universal stress proteins The presence of these stressresistance proteins is not uncommon in
lactobacilli [15] but the specific combination and regulated expression might promote enhanced
tolerance to vaginal environmental fluctuations such as in pH mucus concentration hormones
and infection [12] Several alternative sfactors (RpoE RpoH and RpoN) typically involved in
stress resistance were also detected in the genome of L iners AB1 Each sfactor is used by
bacterial cells to initiate transcription of certain genes depending on the gene itself and on the
environmental signals needed to initiate the transcription RpoE is mainly detected in Gram
negative bacteria where it is involved in maintenance of cell envelope integrity in stress
conditions [4647] RpoH has also been reported to play a role in the survival of mainly
Gramnegative bacteria during elevated temperature and concentrations of H2O2 [48] Genes
responsible for the uptake and metabolism of DNA were found to be affected by sigHLsa (RpoH)
in Lactobacillus sakei [49] RpoN acts as a major regulator of alternative carbohydrate uptake in
L plantarum WCFS1 such as for the mannose phosphoenolpyruvatedependent transport
system (PTS) [50] Thus in addition to stress resistance the better survival or adaptation of L
iners during BV could also be explained by superior metabolic adaptation to the changing
carbohydrate sources in this environment
During BV andor menses it can be speculated that the overgrowth of other bacteria and their
utilization of substrates has a major impact on L iners metabolic activity For example the BV
associated pathogen G vaginalis cannot grow in ironlimiting conditions but can use iron
sources such as hemoglobin for growth Srinivasan et al [37] suggest that one mechanism for
acquiring iron by G vaginalis is to lyse host cells such as erythrocytes by secreting vaginolysin (a
member of the cholesteroldependent cytolysin toxins) This is consistent with the observation
that G vaginalis increases with menses (and therefore vaginal blood) Since L iners grows best
on nutrientrich media such as blood agar and appears to require iron for its growth it is
tempting to speculate that growth of L iners is improved by the released iron from the
degradation of erythrocytes by G vaginalis which would explain the cooccurrence of these
two species during menses and possibly during BV L iners strains have also been shown to
secrete inerolysin which is a toxin with comparable characteristics to that of vaginolysin [51]
Inerolysin is a poreforming toxin that is activated by reducing agents and inhibited by an excess
of cholesterol It can lyse murine ovine and human erythrocytes in a dosedependent manner
in contrast to vaginolysin which is human specific Inerolysin is active across a pH range of 45 to
188 Trends in Microbiology March 2017 Vol 25 No 360 and therefore has optimal activity at the normal vaginal pH between 4 and 5 Inerolysin
seems to be encoded by one of the specific L iners genes that are strongly upregulated during
BV [16] which might suggest that L iners may have an unappreciated role in BV and might
contribute to the pathogenesis of the condition
The ability of pathogenic bacteria to adhere to surfaces is well documented as a virulence factor
and L iners AB1 has been shown to strongly adhere to human fibronectin while other vaginal
strains of L cripatus ATCC 3800 L reuteri RC14 and L rhamnosus GR1 did not [52] L iners
enhanced the adhesion of G vaginalis to HeLa cells in contrast to L crispatus but only when G
vaginalis was added after the lactobacilli were allowed to adhere first [53] Moreover G vaginalis
displaced L crispatus but not L iners from HeLa cells [53] Rather than being a virulence factor
this adhesion could simply be one of the mechanisms utilized by L iners to retain its presence in
the vagina Of note fibronectin has been reported to increase 30fold in the vaginal epithelium
during menses [52] This may explain the increased levels of L iners during menses and its
increased prevalence over L crispatus
Another important aspect that needs to be taken into account is that BV is often associated with
a high phage load [54] Although the role of bacteriophages in BV pathogenesis needs to be
further documented Macklaim et al [16] reported that CRISPR (clustered regularly inter
spaced short palindromic repeat) genes generally responsible for antibacteriophage
defense are highly expressed in L iners under BV conditions but not in health suggesting
a very efficient ability to respond to changes in the phage load during BV Protein members from
the CRISPR defense system are present in only [7_TDDIFF]four out of 12 L iners genome sequences (as
determined by mapping the RNAseq to 12 available L iners genomes at the time of their study
available in NCBI) indicating that this phage resistance is not a universal phenotype It would be
interesting to explore whether this antiviral defense is more associated with strains that prolifer
ate during vaginal dysbiosis
Taken together various genetic and functional studies point towards a remarkable environ
mental adaptation of L iners to the vaginal niche It remains to be elucidated to what extent this
represents pathogenicity or strong colonization capacity especially since various – including yet
unknown – factors during BV might be toxic or inhibitory for the other lactobacilli but not for L
iners
Concluding Remarks
The genus Lactobacillus has a remarkable track record of safety due to its widespread use in the
preparation of fermented foods and its application as probiotics Thirtythree Lactobacillus
species have a qualified presumption of safety’ (QPS) status granted by the European
Food and Safety Authority (EFSA) but to date L iners is not one of these [55] Indeed at
present the available literature is insufficient to classify L iners as a microbiota and Lactobacillus
friend’ Given its relative difficulty in propagation and its ambiguous properties such as the
presence of inerolysin a cytotoxin it is unlikely that L iners will be straightforwardly developed as
a probiotic Nevertheless the available literature is also insufficient to classify l iners as a foe’
Further clarification of its role in health and disease and identification of clones that are
associated with one or the other is thus warranted in the future (see Outstanding Questions)
Of note at present deepsequencing approaches are limited to identification at species level (at
best) The recently developed DADA2 pipeline [56] – which can detect unique ribosomal
sequence variants and which was applied on vaginal microbiota samples as first case study
– is an interesting step forward However even more detailed approaches that can discriminate
different clones or strains of L iners are warranted for future vaginal microbiome studies While
there is some circumstantial evidence of an association between the presence of L iners and
vaginal dysbiosis there is insufficient data to indicate causeandeffect pathogenesis Further
Outstanding Questions
Which factors underlie the instability of
vaginal microbiota community types
dominated by L iners Are bacterio
phages involved
Do different strains of L iners exist
namely beneficial and pathogenic
How to recognize these different
strains and how to select the beneficial
ones
How does L iners organize its gene
expression during environmental
changes such as during bacterial
vaginosis
What is the host response against L
iners – does the host consider the spe
cies as a friend or as a foe
Why does L iners appears more com
monly in Black African and AfroAmeri
can women
Trends in Microbiology March 2017 Vol 25 No 3 189elucidation is also necessary given that other microbially targeted therapies such vaginal transfer
or vaginal seeding are increasingly explored
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