Добавил:
Sekretar
kiopkiopkiop18@yandex.ru
t.me/Prokururor I Вовсе не секретарь, но почту проверяю
Опубликованный материал нарушает ваши авторские права? Сообщите нам.
Вуз:
Предмет:
Файл:Ординатура / Хирургия / Библиотека им академика М.И. Перельмана / Книга_998_Библиотеки_им_академика_М_И_Перельмана
.pdf
Anti-Biolm Activity ofViruses,
https://t.me/medicina_free
Bacteria, Fungi, andLichens:
Mechanisms andImpact
onClinical Practice
OanaSăndulescu, AncaStreinu-Cercel,
MihaiSăndulescu, andAdrianStreinu-Cercel
11
11.1 Introduction
Biolms are pluricellular structures displaying
sophisticated regulatory mechanisms that allow
the survival of bacteria or fungi in hostile environments such as those found in human hosts during
clinical infection. When adopting a sessile lifestyle, bacteria gain the adaptive ability to tolerate
a wide range of antimicrobials, becoming increasingly resilient. In such cases, antimicrobial treatment may fail not necessarily due to resistance but
rather through tolerance and target evasion [1, 2].
Biolms have different characteristics in Grampositive [3] and Gram-negative germs [4, 5] and,
consequently, different mechanism may be
required to ght biolm-driven infections.
In the clinic, there is an acute need to nd new
options for the treatment of biolm-driven infections, and research on biolm-active agents is
well underway. Theoretically, if the three-
O. Săndulescu (*) · A. Streinu-Cercel
A. Streinu-Cercel
Department of Infectious Diseases I, National
Institute for Infectious Diseases “Prof. Dr. Matei
Balș”, Carol Davila University of Medicine and
Pharmacy, Bucharest, Romania
e-mail: oana.sandulescu@umfcd.ro;
anca_sc@germs.ro; astreinucercel@germs.ro
M. Săndulescu
Department of Implant Prosthetic Therapy, Carol
Davila University of Medicine and Pharmacy,
Bucharest, Romania
e-mail: mihai.sandulescu@umfcd.ro
dimensional biolm structure is specically targeted, the remaining planktonic cells can be
easily reached by common antimicrobials, and
the infectious process can thus be stopped.
However, despite the abundant research on this
topic, the transition from bench to bedside is not
always as straightforward. Through this chapter,
we aim to characterize the existing body of
knowledge on the topic of natural anti-biolm
agents, by reviewing the specic literature, in
order to identify the main types of agents, their
mechanisms and their potential clinical role and
impact on medical practice.
11.2 Viruses
A well-described category of natural anti-biolm
agents is that of bacteriophages, which are viruses
infecting bacterial cells and either destroying
these bacterial cells or circumventing their ability
to form biolms.
As bacteriophages display target specicity,
different bacteriophages target different bacteria.
Most of the literature on this topic specically
discusses bacterial lysis, but a lot of recent work
has also focused on their specic ability to inhibit
biolm formation or to contribute to the disruption of mature biolm. As bacteriophages play a
wide range of roles, they can also be involved in
biolm-building activities, by contributing to
polymer assembly, and increasing the amount of
© Springer Nature Switzerland AG 2019
D. Duscher, M. A. Shiffman (eds.), Regenerative Medicine and Plastic Surgery,
https://doi.org/10.1007/978-3-030-19958-6_11
113
114
https://t.me/medicina_free
O. Săndulescu et al.
extracellular DNA as is the case with
Pseudomonas aeruginosa’s internal phage Pf [6].
This subchapter will however focus on the antibiolm properties of bacteriophages.
One of the best-characterized actions of bacteriophages is bacterial lysis, which leads to a
decrease in bacterial load, similar to the mechanisms of other antimicrobial agents. However,
particularly in infections with Gram-negative
germs, lysis can lead to release of endotoxin [7],
specically its lipid A component [8], potentially
associating exaggerated proinammatory cytokine responses; Escherichia coli, for example,
can display on its surface up to 106 lipid A residues [9] and induce a strong host response.
Therefore, the use of lysis-decient bacteriophages has been proposed as option for decreasing endotoxin release [8], and promising results
have been shown in a murine peritonitis model
[10]. Nevertheless recent data suggest that in E.
coli clinical isolates the release of endotoxin with
the use of therapeutic virulent phages (LM33_P1
and 536_P1) may be comparable to that associated with amikacin use, and two- to fourfold
lower compared to carbapenem (specically, imipenem) use [11].
Bacteriophage mixtures have been studied in
clinical trials and are already marketed in countries such as Georgia, or used as experimental
adjunctive local treatment in patients who fail
conventional antimicrobial therapy in a few other
countries, including Romania [12–15] and
Poland [7], where most of the invitro and clinical
experience is available for Gram-positives, but an
extending body of work also addresses Gramnegatives [7].
Apart from bacteriophage mixtures, or cocktails, specic bacteriophage-encoded enzymes
have been studied for the anti-biolm activity.
Among these, depolymerases have been, until
recently, by far the best studied, and are known
for their activity against carbohydrates such as
those found in capsular polysaccharides and
extracellular polymeric substances (EPS). A
thorough review by Pires et al. [16] classied
depolymerases into three main categories:
O-glycosyl hydrolases (divided into six groups,
among which three are more frequently encoun-
tered: sialidases or neuraminidases, levanases,
and peptidases, and three are less common: xylosidases, dextranases, and rhamnosidases), polysaccharide lyases (divided into three groups:
hyaluronate lyases, alginate lyases, and pectin/
pectate lyases), and other types of enzymes, such
as lipases. A large number of the described
depolymerases are encoded by bacteriophages
from the Caudovirales order, and are constituents
of the tail structure. For example, multiple types
of tail ber and tailspike proteins have been
reported to have depolymerase, or, specically,
endoglycosidase activity [17]. The roles that
depolymerases play in biolm control are twofold. First, they can degrade the EPS and decrease
the viscosity of the biolm matrix, leading to better diffusion of both bacteriophages and other
antimicrobials in the bacterial biomass. Second,
they can degrade capsular polysaccharides and
facilitate bacteriophage adsorption and entry into
bacterial cells [17], where bacteriophage-induced
bacterial lysis can now occur. Therefore, depolymerases can be further studied for prospective
application in clinical practice either as part of
bacteriophage therapy or, potentially, as puried
enzyme extracts or recombinant depolymerases.
Further data is needed to ascertain the degree to
which they retain their biological activity under
in vivo conditions, but a number of studies do
point towards a preserved activity of recombinant
enzymes in decreasing virulence in E. coli K1
[18, 19]. Specic examples of potential clinical
applications include alginate lyase in the
reduction of exopolysaccharides produced by P.
aeruginosa mucoid strains from patients with
cystic brosis [20, 21], or CHAPK murein peptidase (cysteine, histidine-dependent amido hydrolase/peptidase) derived from anti-staphylococcal
bacteriophage K, which inhibits biolm
formation and disrupts mature methicillin-resistant S. aureus (MRSA) biolm [22]. The K
bacteriophage- derived modied murein hydrolase domain has also been combined invitro with
a cell wall-binding domain derived from lysostaphin, to generate the chimeric protein P128,
which was able to induce a 95.5% reduction in
mature 48-h biolm by S. aureus isolates from
chronic rhinosinusitis [23].
11 Anti-Biolm Activity ofViruses, Bacteria, Fungi, andLichens: Mechanisms andImpact onClinical…
https://t.me/medicina_free
115
Lysozymes such as Cpl-1 and Cpl-7 have
shown anti-biolm activity on Streptococcus
pneumoniae, S. pseudopneumoniae, and S. oralis
14–16-h biolms [24].
Bacteriophage-encoded endolysins have
long been described as potential antimicrobial
agents, through their lytic activity resulting
from the hydrolysis of peptidoglycan layers.
Recent data also point towards their potential
role as anti- biolm agents, again, either as part
of bacteriophage therapy or through their
administration as puried extracts, and a notable example is that of endolysin MR-10, which
has been shown to decrease bacterial biomass
in mature 7-day-old MRSA biolm, and has
been proposed for its potential use in sequential
treatment, following initial administration of an
antimicrobial (specically, minocycline) [25].
Another study also demonstrated the activity of
nine other endolysins on S. aureus mature 24-h
biolm; among these recombinant peptidoglycan hydrolases containing the SH3b domains,
four appeared highly active (LysK, lysostaphin,
Twort, phiSH2), while others demonstrated a
concentration- dependent activity (80ɑ, phi11,
P68, 2638A, and WMY) [26]. Further data on
endolysins show that LysH5 is also active on
mature 24-h S. aureus and S. epidermidis biolms, albeit at a lower extent when compared to
lysostaphin; furthermore, LysH5 also targets
persister cells, and does not lead to biolm
induction when administered at sub-inhibitory
concentrations [27]. By comparison, SAP-2 is
as efcient as lysostaphin on 2-day mature S.
aureus biolm [28]. PlyGRCS also induces a
rapid decrease in S. aureus mature 24-h biolm,
with a reduction of the biomass to half in as
little as 1h [29].
As described above, peptidoglycan hydrolases
have been intensely studied for their potential
role in the management of Gram-positive biolmrelated infections. The structural characteristics
of Gram-negative germs make them less susceptible to endolysins, as their outer membrane efciently covers the peptidoglycan layer. Different
strategies for facilitating the action of endolysins
on Gram-negative bacteria have been assessed;
an example is pretreatment with outer membrane
permeabilizers, including chelators, such as ethylene diamine tetraacetic acid disodium salt
dihydrate (EDTA), or polycationic agents, such
as polymyxins, aminoglycosides, or lysine polymers [30], but recombinant proteins such as
LysPA26 may also display stand-alone anti-biolm activity on P. aeruginosa [31].
Apart from the already well-described
bacteriophage- derived enzymes, other types of
bacteriophage proteins, specically tail tubular
proteins such as TTPAgp31 from Klebsiella
pneumoniae bacteriophage KP32 and TTPAgp44
from K. pneumoniae bacteriophage KP34 have
been shown to display dual function, with structural and enzymatic activity alike [7]. In a recent
study, Brzozowska etal. (2017) [7] have shown
that TTPAgp31 degrades multiple types of K.
pneumoniae polysaccharides, including capsular,
cell-free (slime), and lipo-polysaccharides
through an α-1,4-glucosidase activity, and displays activity on 20-h mature biolm, decreasing
the biomass by 80% for K. pneumoniae, 50% for
S. aureus, and 60% for Enterococcus faecalis,
while TTPAgp44 hydrolyzes E. faecium capsular
polysaccharides through a glucohydrolase-like
activity and also displays activity on mature 20-h
biolm, reducing the bacterial biomass by 80%
for E. faecium, 40% for P. aeruginosa, and 40%
for Bacillus subtilis.
11.3 Bacteria
In natural environments, bacteria often come into
contact with each other, and they can display a
complex range of interactions, from collaborating within microbial consortia, to competing
with each other for scavenging resources, or even
directly attacking each other by synthetizing specic molecules, bioactive peptides, or by secondary metabolites.
LytA, an N-acetylmuramoyl--alanine amidase, is a pneumococcal autolysin which,
when purified and administered under invitro
conditions, decreases pneumococcal biofilm
biomass by 80%, and also displays synergy
with the Cpl-1 bacteriophage-derived lysozyme [24].
116
https://t.me/medicina_free
O. Săndulescu et al.
Bacillus aneurinolyticus, or Bacillus brevis,
produce a wide array of secondary metabolites.
Among these, tyrocidines TrcA, TrcB, and gramicidin S signicantly inhibit biolm formation by
C. albicans, while gramicidin S is also able to
fully eradicate 24-h mature biolm, although all
studied tyrocidines (TrcA, TrcB, TrcC, TpcC,
and PhcA) display some bactericidal effect on
mature biolm, in the range of 28–74% reduction
[32]. When looking specically at TrcA, TrcB,
and TrcC, they eradicate 55–74% of mature C.
albicans biolms, and they display synergy with
caspofungin and amphotericin B [32].
Bacillus safensis, a soil-dwelling germ, can
also inhibit biolm formation and impair yeastto- hypha transition in C. albicans; it also inhibits
biolm formation and capsule formation by
Cryptococcus neoformans, potentially by impacting the accumulation of glucuronoxylomannan
and its organization into a matrix [33].
P. aeruginosa displays an anti-biolm effect
on Aspergillus fumigatus through the produc-
tion of pyoverdine, which acts as a siderophore,
decreasing the iron concentrations and inducing
iron starvation in A. fumigatus [34]. Extracellular
products such as polysaccharides from P. aeru-
ginosa PAO1 can disperse mature 24-h S. epi-
dermidis biolm [35]. Furthermore, anti-biolm
effects of either planktonic or biolm-associated polysaccharide extracts or Gram-negative
lipopolysaccharides have also been demonstrated for a wider range of bacteria-bacteria
interactions, whereby one germ’s products or
components inhibit the other’s biolm mode of
growth [36].
Kolodkin-Gal et al. [37] have shown that
biolm- grown Bacillus subtilis produces
D-amino acids once the biolm reaches a mature
state (5–8days growth). Among these D-amino
acids, a spontaneously occurring mixture of
D-leucine, D-methionine, D-tyrosine, and
D-tryptophan is able to disperse mature biolm
and also, when extracted and puried, it also
inhibits biolm formation. The mixture’s antibiolm activity is explained through the incorporation of these biolm-disassembling D-amino
acids into the cell wall on the third day of growth.
Once incorporated, they subsequently impair the
anchoring into the cell wall of the amyloid bers
formed by the TasA protein, which is the main
component of B. subtilis biolms, along with
exopolysaccharides [38]. The study by KolodkinGal etal. [37] also tested the efcacy of this mixture of D-amino acids in inhibiting biolm by
other bacterial species, and they found that both
D-tyrosine and the D-amino acid mixture were
able to prevent biolm formation by S. aureus
and P. aeruginosa.
Oral microbiota may play a role in preventing
dental caries, by inhibiting biolm formation by
Streptococcus mutans. Multiple species of lactobacilli have been studied for their anti-biolm
properties, either through coculturing, administration of cell-free supernatant, or extraction of
bacterial products such as bacteriocins. For
example, Ahn etal. have shown that Lactobacillus
plantarum inhibits the production of exopolysaccharide from sucrose by S. mutans [39], while
Was et al. have shown that four Lactobacillus
species inhibit biolm formation, namely L. sali-
varius, L. reuteri, L. plantarum subspecies plantarum, and L. casei subspecies casei, while only
three of these are also active on mature overnight
S. mutans biolm: L. salivarius, L. reuteri, and L.
plantarum subspecies plantarum, through the
same mechanism of reducing exopolysaccharide
formation [40].
Salivary isolates of L. paracasei, L. rhamnosus,
and L. fermentum also inhibit C. albicans biolm
formation through their production of exometabolites and organic acids, both when cocultured with
the fungi and when administered as mature 24-h
growth supernatant [41]. Matsubara etal. have also
shown that L. rhamnosus, L casei, and L. acidophi-
lus inhibit biolm formation and are active on
mature C. albicans biolm, while also hindering its
yeast-to-hyphae differentiation, which is an important anti- virulence effect [42], and this was also
demonstrated by Vilela etal. specically for L. aci-
dophilus, both invitro and in an experimental candidiasis model of Galleria mellonella [43]. The
supernatant derived from certain probiotic lactobacilli (Lactobacillus gasseri and Lactobacillus
rhamnosus) can inhibit biolm formation and disrupt mature 24-h biolm of Candida non-albicans
biolms, specically C. tropicalis, C. krusei, and
11 Anti-Biolm Activity ofViruses, Bacteria, Fungi, andLichens: Mechanisms andImpact onClinical…
https://t.me/medicina_free
117
C. parapsilosis, both alone and in a mixed plurispecies biolm model [44].
Mixed biolms can be encountered in clinical
practice, and Krzyściak etal. [45] have studied
the dental caries biolm, showing that the presence of S. mutans increases the number of C.
albicans colonies and increases the overall biolm mass. They also showed that coculturing
with Lactobacillus salivarius signicantly
decreased biolm formation by S. mutans and C.
albicans alone, or in mixed biolms.
L. fermentum was used for purication of a
bacteriocin, namely fermencin SD11, which displays antimicrobial activity on oral bacteria such
as S. mutans, S. sobrinus, periopathogenic bacteria
such as A. actinomycetemcomitans, Fusobacterium
nucleatum, and Porphyromonas gingivalis, as well
as C. albicans, but has not yet been further studied
for a potential anti-biolm activity [46]. However,
other bacteriocins have been shown to display
anti-biolm properties, as is the case with sonorensin, produced by a marine isolate of Bacillus
sonorensis, MT93, which is bactericidal to both
metabolically active and dormant S. aureus and E.
coli strains, and also inhibits biolm formation by
S. aureus [47]. Specic strains of L. fermentum
have also been shown to produce bacteriocins able
to inhibit biolm formation by P. aeruginosa
PAO-1 [48], and L. kunkeei also inhibits biolm
formation by P. aeruginosa and attenuates infec-
tion in a Galleria mellonella model [49]. When
extracted or puried for standalone administration, bacteriocins can be considered as postbiotics,
as they are bacterial products or by-products of
probiotic bacterial metabolism [50].
Okuda et al. [51] have studied two class I
bacteriocins (lantibiotics): nisin A produced by
Lactococcus lactis and nukacin ISK-1 produced
by Staphylococcus warneri ISK-1, and a class II
bacteriocin, lacticin Q, produced by Lactococcus
lactis QU 5. In their study, only nisin A and lacticin Q were bactericidal on mature 24-h
biolm- embedded S. aureus, and their activity
could be explained by their pore-forming potential, which is not present for nukacin ISK-1.
However, none of the tested bacteriocins were
able to completely eradicate mature biolm in
this study.
A bacteriocin produced by L. plantarum
ST8SH displayed potent anti-biolm activity on
Listeria monocytogenes strains, and synergy with
vancomycin [52]. Another L. plantarum isolate
(CIRM653) decreased 24-h mature K. pneu-
moniae biolm by 77.8%, leading to bacterial
dispersal, but also to an increased rate of gastrointestinal colonization by K. pneumoniae in a
murine model [53].
Another class of bacteriocins, sactibiotics, are
small antimicrobial peptides. Such an example is
hyicin 4244, produced by Staphylococcus hyicus
4244, which showed strong inhibition of biolm
formation, and strong activity on 24-h mature
biolm produced by clinical isolates of S. aureus
and S. saprophyticus [54].
Bidobacteria have also been studied for their
potential anti-biolm activity, albeit to a lesser
extent than lactobacilli. Kim et al. have shown
that Bidobacterium longum cell extracts can
inhibit biolm formation by enterohemorrhagic
E. coli O157:H7 by 36%, and attenuate its virulence in a Caenorhabditis elegans model [55],
Among cyanobacteria, Spirulina platensis can
display antimicrobial [56] and antifungal properties [57], and its methanolic exact has recently
been shown to inhibit P. aeruginosa biolms by
decreasing the amount of EPS [56]. Furthermore,
the aqueous extract of Spirulina platensis has
been used in the biosynthesis of silver nanoparticles, which were then used to coat Foley catheters in combination with amikacin and
nitrofurantoin, and displayed a complete inhibition of colonization or biolm formation by uropathogenic E. coli for 14days, in a murine model
of UTI [58].
Other types of silver nanoparticles have been
biosynthesized from Streptomyces calidiresistens
supernatant, and have been shown to inhibit biolm formation by S. aureus, E. coli, and C. albi-
cans, albeit the degree of biolm inhibition was
signicantly inuenced by the type of Streptomyces
calidiresistens strain used for biosynthesis [59].
Streptomyces hawaiiensis produces acyldep-
sipeptides (ADEPs) [60]; among these, a
semi- synthetic derivative, ADEP4, binds to the
ClpP protease and activates proteolysis, leading
to the destruction of bacterial cells, both
118
https://t.me/medicina_free
O. Săndulescu et al.
metabolically active and inactive, and specically bacterial persisters [61], and its association with rifampin fully eradicated S. aureus
biolm invitro and in a murine thigh infection
model [61].
11.4 Fungi andLichens
In the phylogenetic tree of life, Bacteria and
Eukaryota represent different domains, each
comprising multiple life forms. Fungi are part of
the Eukaryota domain, and can be involved in
clinical infections in humans. However, recent
research has shown that some of their cell wall
components or some of their secondary metabolites may display important roles in limiting
infections or biolms. Here, we will briey
describe anti-biolm compounds isolated from
either clinically relevant fungi or lichenassociated fungi.
Different members of the Penicillium genus
produce different biolm-active compounds,
including the dipeptide cis-cyclo (LeucylTyrosyl), which inhibits biolm formation by S.
epidermidis [62], or norlichexanthone, a non-
reduced tricyclic polyketide isolated from
Penicillium algidum, which inhibits biolm formation and virulence traits such as neutrophil
lysis by MRSA [63]. Other members of this
genus produce shearinines, secondary metabolites that can inhibit yeast-to-hyphae transition
and biolm formation, and disrupt 48-h mature
biolm in C. albicans, while also displaying synergy with amphotericin B [64]. The hyphal transition and biolm formation can also be inhibited
by other alkaloid and polyketide secondary fungal metabolites, waikialoid A and waikialide A,
produced by members of the Aspergillus genus;
however, these metabolites are not active on
mature biolm [65].
Mannoprotein is a surfactant which has been
extracted from Saccharomyces cerevisiae cell
wall. Mannoprotein does not possess antimicrobial activity on S. aureus or S. epidermidis but it
does inhibit biolm formation, and disrupt
mature staphylococcal biolms, potentially by
inuencing cell surface hydrophobicity [66].
Metabolites from Plectosphaerella cucume-
rina such as patulin and emodin can specically
inhibit biolm formation, disrupt mature 24-h
biolm, and inhibit the production of virulence
factors such as protease, elastase, and pyocyanin,
by P. aeruginosa PAO1 without displaying anti-
bacterial activity [67]. Terreic acid, a secondary
metabolite of Aspergillus terreus, can inhibit bio-
lm formation by E. coli [68].
Farnesol, a sesquiterpene from C. albicans or
C. dubliniensis, can inhibit biolm formation by
other Candida isolates [69], but also by
Pneumocystis jirovecii [69, 70], S. epidermidis,
or S. mutans [69]. Other fungi-derived terpenes
have also been reported to display anti-biolm
activity, including guignardone N and guignardic
acid, produced by Guignardia spp., which display synergy with uconazole in the inhibition of
C. albicans biolm [69, 71].
Lichens have also been studied for their capacity to produce anti-biolm compounds, and a
recent study by Millot et al. has identied four
acetone lichen extracts, from Cladonia uncialis,
Evernia prunastri, Ramalina fastigiata, and
Xanthoparmelia conspersa, that showed promis-
ing anti-biolm activity on C. albicans, through a
non-lethal effect. The main metabolites identied
in the acetone extracts were squamatic acid and
usnic acid, evernic acid and usnic acid, evernic
acid and usnic acid, and stictic acid and usnic
acid, respectively [72].
Potentially one of the best characterized secondary metabolites of licheni-associated fungi,
usnic acid has been studied for its anti-biolm
properties. It inhibits biolms by most group A
streptococci [73], S. aureus strains isolated from
patients with cystic brosis [74], C. albicans
[75], C. orthopsilosis [76], but not by C. krusei
[77], while data for C. parapsilosis is contradictory [77]. It has been loaded onto magnetic
nanoparticles [78], carboxylated poly(-lactide)
microparticles used for disrupting 24-h mature S.
epidermidis biolm [79], and used for coating
magnetic polylactic-co-glycolic acid-polyvinyl
alcohol (PLGA-PVA) microsphere thin lms
[80], and used for surface coating of zirconium
dioxide bearing and barium sulfate bearing bone
cement, to prevent biolm formation by MRSA
11 Anti-Biolm Activity ofViruses, Bacteria, Fungi, andLichens: Mechanisms andImpact onClinical…
https://t.me/medicina_free
119
[81]; however, it failed to inhibit biolm formation by S. aureus when used loaded onto polyurethane surfaces [82].
Retigeric acid B, a pentacyclic triterpenoid
isolated from the lichen Lobaria kurokawae, synergistically attenuates yeast-to-hyphae transition
and biolm formation by C. albicans, together
with uconazole [83].
Evernic acid, a secondary metabolite isolated
from lichens from the Evernia genus, inhibits
biolm formation and quorum sensing of
Pseudomonas aeruginosa PAO1 [84], and a simi-
lar but even stronger effect has been demonstrated by the same author group for zeaxanthin,
a tetraterpenoid isolated from the Cladonia
genus, among other lichens [85].
Pyridoxatin, a product isolated from an endolichenic fungus from the Acremonium genus, inhibits biolm formation and growth of C. albicans by
inhibiting ergosterol synthesis [86]. Diorcinol D, a
diphenyl ether derivative isolated from the lichen
endophytic fungus Aspergillus versicolor, displays
synergy with uconazole on C. albicans in disrupting 24-h mature biolm, and in reversing azole
resistance, potentially by inhibiting efux pumps
and ergosterol biosynthesis [87].
11.5 Conclusions
A multitude of natural compounds have been
studied for their potential use as anti-biolm
agents, and promising data show effect on nascent
biolm for most substances, but also on mature
biolm for some of the studied products.
However, further research is still needed for most
of these compounds, as the available body of
knowledge is mostly based on invitro studies, or
in vivo euarthropode, nematode, or murine
models.
References
1. Săndulescu O.Managing sticky situations—anti-bio-
lm agents. Germs. 2016;6(2):49.
2. Nunez-Nunez M, Navarro MD, Palomo V, Rajendran
NB, Del Toro MD, Voss A, Sharland M, Sifakis F,
Tacconelli E, Rodriguez-Bano J.The methodology of
surveillance for antimicrobial resistance and healthcare-associated infections in Europe (SUSPIRE): a
systematic review of publicly available information.
Clin Microbiol Infect. 2018;24(2):105–9.
3. Săndulescu O, Streinu-Cercel A, Săndulescu M,
Neguț A, Calistru P, Berciu I, Preoțescu L, StreinuCercel A. Quorum sensing and biolm formation
in Staphylococcus species. Therap Pharmacol Clin
Toxicol. 2015;19(2):45–51.
4. Bleotu C, Chiriuc M, Mircioaga D, Săndulescu O,
Aldea I, Banu O, Ion D, Diaconu C, Lazar V. The
inuence of nutrient culture media on Escherichia
coli adhesion and biolm formation ability. Rom
Biotechnol Lett. 2017;22(2):12483–91.
5. Săndulescu O. Global distribution of antimicro-
bial resistance in E. coli. J Contemp Clin Pract.
2016;2(2):69–74.
6. Secor PR, Sweere JM, Michaels LA, Malkovskiy
AV, Lazzareschi D, Katznelson E, Rajadas J,
Birnbaum ME, Arrigoni A, Braun KR, Evanko SP,
Stevens DA, Kaminsky W, Singh PK, Parks WC,
Bollyky PL. Filamentous bacteriophage promote
biolm assembly and function. Cell Host Microbe.
2015;18(5):549–59.
7. Brzozowska E, Pyra A, Pawlik K, Janik M, Gorska S,
Urbanska N, Drulis-Kawa Z, Gamian A. Hydrolytic
activity determination of Tail Tubular Protein A of
Klebsiella pneumoniae bacteriophages towards saccharide substrates. Sci Rep. 2017;7(1):18048.
8. Colavecchio A, Goodridge LD. Phage therapy
approaches to reducing pathogen persistence and
transmission in animal production environments:
opportunities and challenges. Microbiol Spectr.
2017;5:3.
PFS-0017-2017.
9. Ramachandran G. Gram-positive and gram-negative
bacterial toxins in sepsis: a brief review. Virulence.
2014;5(1):213–8.
10. Matsuda T, Freeman TA, Hilbert DW, Duff M, Fuortes
M, Stapleton PP, Daly JM. Lysis-decient bacteriophage therapy decreases endotoxin and inammatory
mediator release and improves survival in a murine
peritonitis model. Surgery. 2005;137(6):639–46.
11. Dufour N, Delattre R, Ricard JD, Debarbieux L.The
lysis of pathogenic Escherichia coli by bacteriophages releases less endotoxin yhan by beta-lactams.
Clin Infect Dis. 2017;64(11):1582–8.
12. Neguț A, Streinu-Cercel A, Săndulescu O, Moţoi M,
Berciu I, Popa M, Streinu-Cercel A.Bacteriophages–
novel biotechnology tools available in clinical practice in Romania. Rom Biotechnol Lett.
2017;22(2):12492–5.
13. Negut AC, Chiriuc MC, Sandulescu O, StreinuCercel A, Oprea M, Dragulescu EC, Gheorghe I,
Berciu I, Coralia B, Popa M, Otelea D, Talapan D,
Dorobat O, Codita I, Popa MI.Bacteriophage-driven
inhibition of biolm formation in Staphylococcus
strains from patients attending a Romanian reference
center for infectious diseases. FEMS Microbiol Lett.
2016;363:18. pii: fnw193
https://doi.org/10.1128/microbiolspec.
120
https://t.me/medicina_free
O. Săndulescu et al.
14. Negut AC, Sandulescu O, Popa M, Streinu-Cercel A,
Alavidze Z, Berciu I, Bleotu C, Popa MI, Chiriuc
MC. Experimental approach for bacteriophage susceptibility testing of planktonic and sessile bacterial
populations—study protocol. Germs. 2014;4(4):92–6.
15. Săndulescu O, Bleotu C, Matei L, Streinu-Cercel A,
Oprea M, Drăgulescu EC, Chiriuc MC, Rala A,
Pirici D, Tălăpan D, Dorobăț OM, Neguț AC, Oțelea
D, Berciu I, Ion DA, Codiță I, Calistru PI.Comparative
evaluation of aggressiveness traits in staphylococcal
strains from severe infections versus nasopharyngeal
carriage. Microb Pathog. 2016;102:45–53.
16. Pires DP, Oliveira H, Melo LD, Sillankorva S,
Azeredo J. Bacteriophage-encoded depolymerases:
their diversity and biotechnological applications.
Appl Microbiol Biotechnol. 2016;100(5):2141–51.
17. Yan J, Mao J, Xie J. Bacteriophage polysaccharide
depolymerases and biomedical applications. Bio
Drugs. 2014;28(3):265–74.
18. Mushtaq N, Redpath MB, Luzio JP, Taylor
PW. Prevention and cure of systemic Escherichia
coli K1 infection by modication of the bacterial phenotype. Antimicrob Agents Chemother.
2004;48(5):1503–8.
19. Zelmer A, Martin MJ, Gundogdu O, Birchenough
G, Lever R, Wren BW, Luzio JP, Taylor
PW. Administration of capsule-selective endosialidase E minimizes upregulation of organ gene expression induced by experimental systemic infection with
Escherichia coli K1. Microbiology. 2010;156(Pt
7):2205–15.
20. Glonti T, Chanishvili N, Taylor PW. Bacteriophagederived enzyme that depolymerizes the alginic
acid capsule associated with cystic brosis isolates
of Pseudomonas aeruginosa. J Appl Microbiol.
2010;108(2):695–702.
21. Latka A, Maciejewska B, Majkowska-Skrobek G,
Briers Y, Drulis-Kawa Z. Bacteriophage-encoded
virion-associated enzymes to overcome the carbohydrate barriers during the infection process. Appl
Microbiol Biotechnol. 2017;101(8):3103–19.
22. Keary R, Sanz-Gaitero M, van Raaij MJ, O’Mahony
J, Fenton M, McAuliffe O, Hill C, Ross RP, Coffey
A. Characterization of a bacteriophage-derived
murein peptidase for elimination of antibiotic-resistant Staphylococcus aureus. Curr Protein Pept Sci.
2016;17(2):183–90.
23. Drilling AJ, Cooksley C, Chan C, Wormald PJ,
Vreugde S. Fighting sinus-derived Staphylococcus
aureus biolms in vitro with a bacteriophagederived muralytic enzyme. Int Forum Allergy Rhinol.
2016;6(4):349–55.
24. Domenech M, Garcia E, Moscoso M.In vitro destruction of Streptococcus pneumoniae biolms with bacterial and phage peptidoglycan hydrolases. Antimicrob
Agents Chemother. 2011;55(9):4144–8.
25. Chopra S, Harjai K, Chhibber S.Potential of sequential treatment with minocycline and S. aureus
specic phage lysin in eradication of MRSA biolms: an in vitro study. Appl Microbiol Biotechnol.
2015;99(7):3201–10.
26. Schmelcher M, Shen Y, Nelson DC, Eugster MR,
Eichenseher F, Hanke DC, Loessner MJ, Dong S,
Pritchard DG, Lee JC, Becker SC, Foster-Frey J,
Donovan DM. Evolutionarily distinct bacteriophage endolysins featuring conserved peptidoglycan
cleavage sites protect mice from MRSA infection. J
Antimicrob Chemother. 2015;70(5):1453–65.
27. Gutierrez D, Ruas-Madiedo P, Martinez B, Rodriguez
A, Garcia P. Effective removal of staphylococcal biolms by the endolysin LysH5. PLoS One.
2014;9(9):e107307.
28. Son JS, Lee SJ, Jun SY, Yoon SJ, Kang SH, Paik HR,
Kang JO, Choi YJ.Antibacterial and biolm removal
activity of a podoviridae Staphylococcus aureus bacteriophage SAP-2 and a derived recombinant cellwall-degrading enzyme. Appl Microbiol Biotechnol.
2010;86(5):1439–49.
29. Linden SB, Zhang H, Heselpoth RD, Shen
Y, Schmelcher M, Eichenseher F, Nelson
DC.Biochemical and biophysical characterization of
PlyGRCS, a bacteriophage endolysin active against
methicillin-resistant Staphylococcus aureus. Appl
Microbiol Biotechnol. 2015;99(2):741–52.
30. Briers Y, Walmagh M, Lavigne R. Use of bacteriophage endolysin EL188 and outer membrane permeabilizers against Pseudomonas aeruginosa. J Appl
Microbiol. 2011;110(3):778–85.
31. Guo M, Feng C, Ren J, Zhuang X, Zhang Y, Zhu Y,
Dong K, He P, Guo X, Qin J.A novel antimicrobial
endolysin, LysPA26, against Pseudomonas aerugi-
nosa. Front Microbiol. 2017;8:293.
32. Troskie AM, Rautenbach M, Delattin N, Vosloo JA,
Dathe M, Cammue BP, Thevissen K. Synergistic
activity of the tyrocidines, antimicrobial cyclodecapeptides from Bacillus aneurinolyticus, with
amphotericin B and caspofungin against Candida
albicans biolms. Antimicrob Agents Chemother.
2014;58(7):3697–707.
33. Mayer FL, Kronstad JW. Disarming fungal pathogens: Bacillus safensis inhibits virulence factor
production and biolm formation by Cryptococcus
neoformans and Candida albicans. MBio. 2017;8(5):
e01537–17.
34. Sass G, Nazik H, Penner J, Shah H, Ansari SR,
Clemons KV, Groleau MC, Dietl AM, Visca P, Haas
H, Deziel E, Stevens DA. Studies of Pseudomonas
aeruginosa mutants indicate pyoverdine as the central
factor in inhibition of Aspergillus fumigatus biolm. J
Bacteriol. 2018;200(1):e00345–17.
35. Qin Z, Yang L, Qu D, Molin S, Tolker-Nielsen T.
Pseudomonas aeruginosa extracellular products
inhibit staphylococcal growth, and disrupt established
biolms produced by Staphylococcus epidermidis.
Microbiology. 2009;155(Pt 7):2148–56.
36. Rendueles O, Kaplan JB, Ghigo JM.Antibiolm polysaccharides. Environ Microbiol. 2013;15(2):334–46.
37. Kolodkin-Gal I, Romero D, Cao S, Clardy J, Kolter R,
Losick R.D-amino acids trigger biolm disassembly.
Science. 2010;328(5978):627–9.
38. Romero D, Aguilar C, Losick R, Kolter R.Amyloid
bers provide structural integrity to Bacillus
11 Anti-Biolm Activity ofViruses, Bacteria, Fungi, andLichens: Mechanisms andImpact onClinical…
https://t.me/medicina_free
121
subtilis biolms. Proc Natl Acad Sci U S A.
2010;107(5):2230–4.
39. Ahn KB, Baik JE, Park OJ, Yun CH, Han SH.
Lactobacillus plantarum lipoteichoic acid inhibits
biolm formation of Streptococcus mutans. PLoS
One. 2018;13(2):e0192694.
40. Was R, Abd El-Rahman OA, Zafer MM, Ashour
HM. Probiotic Lactobacillus sp. inhibit growth,
biolm formation and gene expression of cariesinducing Streptococcus mutans. J Cell Mol Med.
2018;22(3):1972–83.
41. Rossoni RD, de Barros PP, de Alvarenga JA, Ribeiro
FC, Velloso MDS, Fuchs BB, Mylonakis E, Jorge
AOC, Junqueira JC. Antifungal activity of clinical Lactobacillus strains against Candida albicans
biolms: identication of potential probiotic candidates to prevent oral candidiasis. Biofouling.
2018;34(2):212–25.
42. Matsubara VH, Wang Y, Bandara HM, Mayer MP,
Samaranayake LP. Probiotic lactobacilli inhibit
early stages of Candida albicans biolm development by reducing their growth, cell adhesion, and
lamentation. Appl Microbiol Biotechnol. 2016;
100(14):6415–26.
43. Vilela SF, Barbosa JO, Rossoni RD, Santos JD,
Prata MC, Anbinder AL, Jorge AO, Junqueira JC.
Lactobacillus acidophilus ATCC 4356 inhibits biolm formation by C. albicans and attenuates the
experimental candidiasis in Galleria mellonella.
Virulence. 2015;6(1):29–39.
44. Tan Y, Leonhard M, Moser D, Ma S, Schneider-Stickler
B.Inhibitory effect of probiotic lactobacilli supernatants on single and mixed non-albicans Candida species biolm. Arch Oral Biol. 2018;85:40–5.
45. Krzyściak W, Kościelniak D, Papież M, Vyhouskaya
P, Zagórska-Świeży K, Kołodziej I, Bystrowska B,
Jurczak A.Effect of a Lactobacillus salivarius probiotic on a double-species Streptococcus mutans
and Candida albicans caries biolm. Nutrients.
2017;9(11):E1242.
46. Wannun P, Piwat S, Teanpaisan R. Purication,
characterization, and optimum conditions of fermencin SD11, a bacteriocin produced by human
orally Lactobacillus fermentum SD11. Appl Biochem
Biotechnol. 2016;179(4):572–82.
47. Chopra L, Singh G, Kumar Jena K, Sahoo
DK.Sonorensin: A new bacteriocin with potential of
an anti-biolm agent and a food biopreservative. Sci
Rep. 2015;5:13412.
48. Sharma V, Harjai K, Shukla G.Effect of bacteriocin
and exopolysaccharides isolated from probiotic on P.
aeruginosa PAO1 biolm. Folia Microbiol (Praha).
2018;63(2):181–90.
49. Berrios P, Fuentes JA, Salas D, Carreno A, Aldea P,
Fernandez F, Trombert AN.Inhibitory effect of biolm-forming Lactobacillus kunkeei strains against
virulent Pseudomonas aeruginosa invitro and in honeycomb moth (Galleria mellonella) infection model.
Benef Microbes. 2017;9(2):257–68.
50. Patel RM, Denning PW. Therapeutic use of prebiotics, probiotics, and postbiotics to prevent necrotiz-
ing enterocolitis. what is the current evidence? Clin
Perinatol. 2013;40(1):11–25.
51. Okuda K, Zendo T, Sugimoto S, Iwase T, Tajima A,
Yamada S, Sonomoto K, Mizunoe Y. Effects of bacteriocins on methicillin-resistant Staphylococcus
aureus biolm. Antimicrob Agents Chemother.
2013;57(11):5572–9.
52. Todorov SD, de Paula OAL, Camargo AC, Lopes
DA, Nero LA.Combined effect of bacteriocin produced by Lactobacillus plantarum ST8SH and vancomycin, propolis or EDTA for controlling biolm
development by Listeria monocytogenes. Rev Argent
Microbiol. 2018;50:48–55.
53. Lagrafeuille R, Miquel S, Balestrino D, VareilleDelarbre M, Chain F, Langella P, Forestier
C. Opposing effect of Lactobacillus on in vitro
Klebsiella pneumoniae in biolm and in an in vivo
intestinal colonisation model. Benef Microbes.
2018;9(1):87–100.
54. Duarte AFS, Ceotto-Vigoder H, Barrias ES, SoutoPadron T, Nes IF, Bastos M. Hyicin 4244, the rst
sactibiotic described in staphylococci, exhibits an
anti-staphylococcal biolm activity. Int J Antimicrob
Agents. 2018;51:349–56.
55. Kim Y, Lee JW, Kang SG, Oh S, Grifths MW.
Bidobacterium spp. inuences the production of
autoinducer-2 and biolm formation by Escherichia
coli O157:H7. Anaerobe. 2012;18(5):539–45.
56. Lewis Oscar F, Nithya C, Alharbi SA, Alharbi NS,
Thajuddin N.In vitro and in silico attenuation of quorum sensing mediated pathogenicity in Pseudomonas
aeruginosa using Spirulina platensis. Microb Pathog.
2018;116:246–56.
57. Marangoni A, Foschi C, Micucci M, Nahui Palomino
RA, Gallina Toschi T, Vitali B, Camarda L, Mandrioli
M, De Giorgio M, Aldini R, Corazza I, Chiarini A,
Cevenini R, Budriesi R.In vitro activity of Spirulina
platensis water extract against different Candida species isolated from vulvo-vaginal candidiasis cases.
PLoS One. 2017;12(11):e0188567.
58. Mala R, Annie Aglin A, Ruby Celsia AS, Geerthika S,
Kiruthika N, VazagaPriya C, Srinivasa KK.Foley catheters functionalised with a synergistic combination of
antibiotics and silver nanoparticles resist biolm formation. IET Nanobiotechnol. 2017;11(5):612–20.
59. Wypij M, Swiecimska M, Czarnecka J, Dahm H, Rai
M, Golinska P. Antimicrobial and cytotoxic activity of silver nanoparticles synthesized from two
haloalkaliphilic actinobacterial strains alone and
in combination with antibiotics. J Appl Microbiol.
2018;124:1411–24.
60. drugs KCTA. Persisters come under re. Nat Rev
Drug Discov. 2014;13(1):18–9.
61. Conlon BP, Nakayasu ES, Fleck LE, LaFleur MD,
Isabella VM, Coleman K, Leonard SN, Smith RD,
Adkins JN, Lewis K.Activated ClpP kills persisters
and eradicates a chronic biolm infection. Nature.
2013;503(7476):365–70.
62. Scopel M, Abraham WR, Henriques AT, Macedo
AJ.Dipeptide cis-cyclo(Leucyl-Tyrosyl) produced by
sponge associated Penicillium sp. F37 inhibits biolm
122
https://t.me/medicina_free
O. Săndulescu et al.
formation of the pathogenic Staphylococcus epidermidis. Bioorg Med Chem Lett. 2013;23(3):624–6.
63. Baldry M, Nielsen A, Bojer MS, Zhao Y, Friberg C,
Ifrah D, Glasser Heede N, Larsen TO, Frokiaer H,
Frees D, Zhang L, Dai H, Ingmer H.Norlichexanthone
reduces virulence gene expression and biolm
formation in Staphylococcus aureus. PLoS One.
2016;11(12):e0168305.
64. You J, Du L, King JB, Hall BE, Cichewicz RH.Smallmolecule suppressors of Candida albicans biolm
formation synergistically enhance the antifungal
activity of amphotericin B against clinical Candida
isolates. ACS Chem Biol. 2013;8(4):840–8.
65. Wang X, You J, King JB, Powell DR, Cichewicz
RH. Waikialoid A suppresses hyphal morphogenesis and inhibits biolm development in pathogenic
Candida albicans. J Nat Prod. 2012;75(4):707–15.
66. Walencka E, Wieckowska-Szakiel M, Rozalska S,
Sadowska B, Rozalska B.A surface-active agent from
Saccharomyces cerevisiae inuences staphylococcal
adhesion and biolm development. Z Naturforsch C.
2007;62(5–6):433–8.
67. Zhou J, Bi S, Chen H, Chen T, Yang R, Li M, Fu
Y, Jia AQ. Anti-biolm and antivirulence activities
of metabolites from Plectosphaerella cucumerina
against Pseudomonas aeruginosa. Front Microbiol.
2017;8:769.
68. Sharma R, Lambu MR, Jamwal U, Rani C, Chib
R, Wazir P, Mukherjee D, Chaubey A, Khan IA.
Escherichia coli N-acetylglucosamine-1-phosphateuridyltransferase/glucosamine-1-phosphate-acetyltransferase (GlmU) inhibitory activity of terreic acid
isolated from Aspergillus terreus. J Biomol Screen.
2016;21(4):342–53.
69. Bergamo Estrela A, Abraham W. Fungal metabolites for the control of biolm infections. Agri.
2016;37(6):37.
70. Cushion MT, Collins MS, Linke MJ. Biolm formation by Pneumocystis spp. Eukaryot Cell.
2009;8(2):197–206.
71. Li TX, Yang MH, Wang XB, Wang Y, Kong
LY. Synergistic antifungal meroterpenes and dioxolanone derivatives from the endophytic fungus
Guignardia sp. J Nat Prod. 2015;78(11):2511–20.
72. Millot M, Girardot M, Dutreix L, Mambu L, Imbert
C. Antifungal and anti-biolm activities of acetone
lichen extracts against Candida albicans. Molecules.
2017;22(4):E651.
73. Nithyanand P, Beema Shafreen RM, Muthamil S,
Karutha Pandian S. Usnic acid, a lichen secondary
metabolite inhibits Group A Streptococcus biolms.
Antonie Van Leeuwenhoek. 2015;107(1):263–72.
74. Pompilio A, Pomponio S, Di Vincenzo V, Crocetta
V, Nicoletti M, Piovano M, Garbarino JA, Di
Bonaventura G. Antimicrobial and antibiolm
activity of secondary metabolites of lichens against
methicillin-resistant Staphylococcus aureus strains
from cystic brosis patients. Future Microbiol.
2013;8(2):281–92.
75. Nithyanand P, Beema Shafreen RM, Muthamil S,
Karutha Pandian S. Usnic acid inhibits biolm for-
mation and virulent morphological traits of Candida
albicans. Microbiol Res. 2015;179:20–8.
76. Pires RH, Lucarini R, Mendes-Giannini MJ.Effect
of usnic acid on Candida orthopsilosis and C.
parapsilosis. Antimicrob Agents Chemother.
2012;56(1):595–7.
77. Kvasnickova E, Matatkova O, Cejkova A, Masak
J. Evaluation of baicalein, chitosan and usnic acid
effect on Candida parapsilosis and Candida kru-
sei biolm using a Cellavista device. J Microbiol
Methods. 2015;118:106–12.
78. Taresco V, Francolini I, Padella F, Bellusci M, Boni
A, Innocenti C, Martinelli A, D’Ilario L, Piozzi
A. Design and characterization of antimicrobial
usnic acid loaded-core/shell magnetic nanoparticles.
Korean J Couns Psychother. 2015;52:72–81.
79. Martinelli A, Bakry A, D’Ilario L, Francolini I,
Piozzi A, Taresco V.Release behavior and antibiolm
activity of usnic acid-loaded carboxylated poly(Llactide) microparticles. Eur J Pharm Biopharm.
2014;88(2):415–23.
80. Grumezescu V, Holban AM, Grumezescu AM, Socol
G, Ficai A, Vasile BS, Trusca R, Bleotu C, Lazar V,
Chiriuc CM, Mogosanu GD. Usnic acid-loaded
biocompatible magnetic PLGA-PVA microsphere
thin lms fabricated by MAPLE with increased resistance to staphylococcal colonization. Biofabrication.
2014;6(3):035002.
81. Kim S, Greenleaf R, Miller MC, Satish L, Kathju
S, Ehrlich G, Post JC, Sotereanos NG, Stoodley
P. Mechanical effects, antimicrobial efcacy
and cytotoxicity of usnic acid as a biolm prophylaxis in PMMA. J Mater Sci Mater Med.
2011;22(12):2773–80.
82. Francolini I, Norris P, Piozzi A, Donelli G, Stoodley
P. Usnic acid, a natural antimicrobial agent able
to inhibit bacterial biolm formation on polymer surfaces. Antimicrob Agents Chemother.
2004;48(11):4360–5.
83. Chang W, Li Y, Zhang L, Cheng A, Liu Y, Lou
H. Retigeric acid B enhances the efcacy of
azoles combating the virulence and biolm formation of Candida albicans. Biol Pharm Bull.
2012;35(10):1794–801.
84. Gokalsin B, Sesal NC.Lichen secondary metabolite
evernic acid as potential quorum sensing inhibitor
against Pseudomonas aeruginosa. World J Microbiol
Biotechnol. 2016;32(9):150.
85. Gokalsin B, Aksoydan B, Erman B, Sesal NC.Reducing
virulence and biolm of Pseudomonas aeruginosa by
potential quorum sensing inhibitor carotenoid: zeaxanthin. Microb Ecol. 2017;74(2):466–73.
86. Chang W, Zhang M, Li Y, Li X, Gao Y, Xie Z,
Lou H. Lichen endophyte derived pyridoxatin
inactivates Candida growth by interfering with
ergosterol biosynthesis. Biochim Biophys Acta.
2015;1850(9):1762–71.
87. Li Y, Chang W, Zhang M, Li X, Jiao Y, Lou
H.Synergistic and drug-resistant reversing effects of
diorcinol D combined with uconazole against Candida
albicans. FEMS Yeast Res. 2015;15(2):fov001.
Соседние файлы в папке Библиотека им академика М.И. Перельмана