Molecular Darwinian evolution of virulence in Yersinia pestis

25 Pages·2009·0.66 MB·English

IAI Accepts, published online ahead of print on 16 March 2009
Infect. Immun. doi:10.1128/IAI.01477-08
Copyright © 2009, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.
1
2
3
4
5
6
7 Molecular Darwinian evolution of virulence in Yersinia pestis
8
9
D
10 o
w
11 Dongsheng Zhou, and Ruifu Yang nlo
a
12 d
e
13 State Key Laboratory of Pathogen and Biosecurity, d
f
r
o
14 Beijing Institute of Microbiology and Epidemiology, Beijing 100071, China m

15 h
t
t
p
16 :
/
/
ia
17 i.
a
18 Correspondence: sm
.
19 Dongsheng Zhou, Associate Professor, Ph.D. o
r
g
20 E-mail: [email protected] /
o
n
21 Tel: [phone]
J
a
22 n
u
a
23 Ruifu Yang, Professor, Ph.D. r
y

2
24 E-mail: [email protected] 2
,

25 Tel: [phone] 2
0
1
9

b
y

g
u
e
s
t
1
26 The genus Yersinia consists of fifteen species (www.bacterio.cict.fr/xz/yersinia.html),
27 and only three of them, Y. pestis, Y. pseudotuberculosis, and Y. enterocolitica, are
28 pathogenic to mammals including humans. The Y. pseudotuberculosis-Y. pestis
29 evolutionary linkage diverged from Y. enterocolitica between 41 and 186 million years
30 ago, while Y. pestis from Y. pseudotuberculosis within the last 1,500 to 20,000 years (1,
31 65). In accordance with this evolutionary cascade, the widely genetic diversity exists D
o
w
32 between Y. pseudotuberculosis, and Y. enterocolitica, while a very closely genetic n
lo
a
33 similarity is found between Y. pseudotuberculosis and Y. pestis. Y. pseudotuberculosis d
e
d

34 only causes non-fatal gastrointestinal disease in mammalian hosts including humans, and fr
o
m
35 the disease is transmitted by the food-borne route. Y. pestis causes plague that is one of h
t
t
p
36 the most deadly diseases (47). Three pandemics of plague have been recorded in the ://
ia
i.
37 human history, which have claimed hundreds of thousands of lives (47). Plague is a a
s
m
38 typical enzootic disease [an infection of the animal population (s) in one or more .o
r
g
/
39 confined natural foci without the need for external inputs], and the epidemics of rodent o
n

J
40 plague are restricted in various enzootic plague foci especially in Asia, America and a
n
u
a
41 Africa (80). Compared to its progenitor Y. pseudotuberculosis, Y. pestis utilizes a r
y

2
42 radically different mechanism of transmission in rodent reservoirs that rely primarily 2
,

2
0
43 upon bite of flea vectors. This review deals with how genetic changes (gene inactivation, 1
9

b
44 loss and acquisition) and remodeling of gene regulation promote Y. pestis to switch from y

g
u
45 an enteric lifestyle to a mammalian blood-borne lifestyle that relies on vector-borne e
s
t
46 transmission.
47
48 Progression of plague infection
49 Rodents and humans acquire Y. pestis by the bite of infected flea, contact with infected
50 tissues, or inhalation of respiratory droplets or aerosols, with manifestations of bubonic,
2
51 septicemic and pneumonic plague (47). After the flea biting, there is an initial
52 subcutaneous and intradermal colonization, and then the bacteria migrate into the regional
53 lymph nodes, and inflammation, cellulitis and occasionally large carbuncles develop
54 around the bubo (bubonic plague) (60). Without the timely effective treatment, the
55 bacteria will rapidly escape from containment in the lymph node, and spread systemically
56 through the blood to various organs, causing fatal sepsis (septicemic plague) (82). An D
o
w
57 intracellular growth of Y. pestis in macrophages at early stages of infection is thought to n
lo
a
58 be a shelter for this pathogen to proliferate and to synthesize virulence determinants, d
e
d

59 enabling the releasing bacteria to acquire the ability to annihilate the host immune fr
o
m
60 response (39). In addition, secondary pneumonic plague could result from haematogenous h
t
t
p
61 spread from the bubo to lung, presenting in patients as severe bronchopneumonia, ://
ia
i.
62 cavitation, or consolidation with production of bloody or purulent sputum (82). Primary a
s
m
63 pneumonic plague could directly caused be the inhalation of infectious droplets or .o
r
g
/
64 aerosols, with symptoms including acute pneumonia, intraalveolar hemorrhage and edema, o
n

J
65 profound lobular exudation, fibrin deposition and bacillary aggregation (33). Both a
n
u
a
66 primary and secondary pneumonic plagues are highly contagious by the airborne r
y

2
67 transmission for close contacts. 2
,

2
0
68 1
9

b
69 Ecological and epidemiological differences between Y. pestis and Y. pseudotuberculosis y

g
u
70 Animals, food and the abiotic environment are Y. pseudotuberculosis reservoirs from e
s
t
71 which epizootic and human infection may arise, and the disease is mild and transmitted by
72 the food-borne route. In humans, typical symptoms include fever and right-sided
73 abdominal pain. In rare cases the disease may cause skin complaints (erythema nodosum),
74 joint stiffness and pain (reactive arthritis), or spread of bacteria to the blood (bacteremia).
75 Due to acute and systemic infection, the mortality rate of plague reaches to 70-100%
3
76 without treatment depending on routes of infection. Y. pestis has a limited ability to live in
77 the environment although there was evidence that Y. pestis could live in soil for up to 30
78 weeks (3). Maintenance of plague in enzootic plague foci is almost absolutely dependent
79 upon cyclic transmission between fleas and mammals (80). Blocked fleas are important
80 for transmission of plague (24). Blockage of fleas (heavy proliferation of bacteria in the
81 adhesive biofilms in the proventriculus) makes them feel hungry and repeatedly attempt to D
o
w
82 feed, and the plague bacilli will be pumped into host body during these futile feeding n
lo
a
83 attempts (25). The development of a heavy bacteremia (a bacterial concentration reaches d
e
d

84 at least 106 cfu/ml) in hosts is necessary to reliably infect the fleas (38). Such high level of fr
o
m
85 bacteremia raises the risk of hosts’ rapid death. Nevertheless, once some fleas achive h
t
t
p
86 feeding prior to the host’s death, they will seek alternative hosts, thereby increasing the ://
ia
i.
87 likelihood of transmission to other individuals of the hosts (15). a
s
m
88 Generally, it takes about two weeks for blockage to develop, which is not sufficient to .o
r
g
/
89 explain the rapid rate of spread that typifies plague epidemics and epidemics. Various o
n

J
90 species of rodent fleas are immediately infectious after biting a septicaemic host and a
n
u
a
91 transmits efficiently for at least 4d post-infection, and accordingly the mode of r
y

2
92 ‘early-phase transmission’ by unblocked fleas is proposed (14). Early-phase transmission 2
,

2
0
93 helps explain not only the rapid spread that typifies plague epidemics but also previous 1
9

b
94 inconsistencies between the rate of pathogen spread expected by the blocked fleas. It was y

g
u
95 suggested that the mechanical transmission by unblocked fleas is significant during e
s
t
96 epidemics that represents the periods when Y. pestis can spread rapidly across landscapes,
97 but that transmission by blocked fleas is important primarily during inter-epizootic
98 transmission (15). In addition, a combination of early-phase transmission and blocking
99 probably helps to understand the observed high mortality rates of susceptible host
100 populations, including humans during the Black Death (74).
4
101 The rodent reservoirs, the flea vectors and Y. pestis constitute a well-balanced
102 biocommunity in the plague foci. Y. pestis possesses its potential to attack humans, and
103 the human infection usually occurs with the transmission of the pathogen from rodents.
104 Although cases of human plague can be well controlled by timely antibiotic
105 administration, plague still remains a significant concern of public health because it can
106 be transmitted from person to person through respiratory droplets and used for D
o
w
107 bioterrorism and biological warfare (68). n
lo
a
108 d
e
d

109 Y. pestis virulence determinants shared by Y. pseudotuberculosis fr
o
m
110 Y. pestis has developed specialized strategies for virulence in hosts and transmission h
t
t
p
111 by flea (Table 1), and many of these determinants were harbored in the genome of Y. ://
ia
i.
112 pseudotuberculosis. a
s
m
113 Colonization and dissemination .o
r
g
/
114 The major adhesin and invasin, YadA and Inv, specific for gastrointestinal infection o
n

J
115 are inactivated in Y. pestis (see below), but this pathogen still have additional proteins a
n
u
a
116 [Ail (31), YadBC (20) and YapE (35)] that account for bacterial colonization and r
y

2
117 dissemination during infection. 2
,

2
0
118 Intracellular growth 1
9

b
119 The ability to replicate in macrophages is conserved in Y. pestis and Y. y

g
u
120 pseudotuberculosis (53). RipABC (54), MgtCB (22), Ugd (22), Yfe (48), Feo (48) have e
s
t
121 been shown to be required for the replication of Y. pestis in macrophages. Both MgtCB
122 and Ugd are positively regulated be the PhoP/PhoQ two-component system (37) that is
123 important for survival under conditions of macrophage-induced stress and virulence in Y.
124 pestis (44).
125 Annihilation of host immune response
5
126 The plasmid pCD1-borne type III secretion system (T3SS) is composed of a secretion
127 machinery, a set of translocation proteins, a control system, and six Yop effector proteins
128 (56). Through the T3SS, pathogenic yersiniae inject effectors into the cytosol of
129 eukaryotic cells when docking at the surface of host cells, and the injected YOPs mediate
130 suppression of phagocytosis and inflammatory reaction (56). Y. pestis utilizes T3SS to
131 selectively destroy innate immune cells that represent the first line of host defense, D
o
w
132 thereby preventing adaptive responses and precipitating the fatal outcome of plague (40). n
lo
a
133 Y. pesitis still employs pH6 antigen fimbriae to function as antiphagocytic factor d
e
d

134 independent of YOPs (27). fr
o
m
135 The Tc genes were first identified in the insect pathogen and encode a protein h
t
t
p
136 complex toxic to insects. Tc proteins in Y. pseudotuberculosis and Y. pestis are not ://
ia
i.
137 insecticidal toxins but have evolved the toxicity to mammalian cells (23). a
s
m
138 Heavy proliferation of Y. pestis in the bloodstream is essential for its transmission by .o
r
g
/
139 fleas. Resistance to complement-mediated lysis (serum resistance) is required for o
n

J
140 bacterial survival in mammalian blood. The Ail protein (4) and the lipopolysaccharide a
n
u
a
141 (LPS) (50) promotes serum resistance, which appears to be a conserved mechanism in r
y

2
142 pathogenic yersiniae. 2
,

2
0
143 Iron uptake 1
9

144 In mammals, iron is bound to Fe3+-binding proteins and hemopoteins, and thus free by

g
u
145 iron is too rare to sustain bacterial growth. Iron acquisition is critical for the survival of e
s
t
146 pathogenic bacteria during infection. A large array of iron acquisition systems have been
147 characterized or annotated in Y. pestis (21), and at least two (Ybt and Yfe) of them were
148 proven to be required for full virulence (5). Ybt, also known as high pathogenicity island
149 (HPI) (59), is essential to iron acquisition at the site of the flea bite and in the lymphatic
150 system, while Yfe is likely used in the later stages of the disease, i.e., blood-borne
6
151 systemic dissemination (5).
152
153 Lateral acquisition of novel virulence determinants by Y. pestis
154 Lateral gene transfer directly introduce foreign DNA elements into the host genome,
155 which will effectively alter pathogenic characters of bacterial species (42). Y. pestis has
156 acquired two unique virulence plasmids, pPCP1 and pMT1, through lateral gene transfer. D
o
w
157 pPCP1 encodes plasminogen activator (Pla), while pMT1 encodes murine toxin (Ymt) n
lo
a
158 and F1 capsule (Table 1). d
e
d

159 Pla is essential for bubonic and primary pneumonic plague (but not primary and fr
o
m
160 secondary septicemic forms), since it specifically promotes Y. pestis to disseminate from h
t
t
p
161 peripheral infection routes (34, 61). At 37°C but not 26°C, Y. pestis expresses a ://
ia
i.
162 capsule-like antigen, called F1 antigen. F1 provides Y. pestis the ability to block the a
s
m
163 phagocytosis by a mechanism different from those of T3SS and pH6 antigen (13). Ymt .o
r
g
/
164 does not play a role in the mouse infection (57), but shows phospholipase D (PLD) o
n

J
165 activity and is required for survival of Y. pestis in the fleas (26). It was thought that a
n
u
a
166 intracellular PLD activity appeared to protect Y. pestis from a cytotoxic digesting product r
y

2
167 of plasma in the flea gut (26). 2
,

2
0
168 Unexpectedly, only two chromosomal regions seem to be specific to Y. pestis (76) 1
9

b
169 (Table 1). They are located in two different genomic islands probably acquired through y

g
u
170 lateral gene transfer (45). These two Y. pestis-specific chromosomal regions deserve e
s
t
171 more attentions to investigate their roles in virulence and/or transmission by flea.
172 However, it has been argued that analysis of more bacterial strains could further reduce
173 the number of Y. pestis-specific chromosomal genes, perhaps to zero (7, 45).
174
175
7
176 Decay of redundant or deleterious functions in Y. pestis
177 About 13% of Y. pseudotuberculosis genes no longer functions (inactivated or absent)
178 in Y. pestis CO92 (45). Genome decay (gene loss and inactivation) appears to be closely
179 linked to flea-borne transmission and increased virulence of Y. pestis (Table 1).
180 Gene inactivation
181 yadA and inv encode major adhesin/invasin in Y. pseudotuberculosis, and enable this D
o
w
182 enteropathogen to specifically adhere to surfaces of host intestines and invade lining n
lo
a
183 epithelial cells. Both of them are inactivated in Y. pestis [30]. Urease plays its role in d
e
d

184 using urea as a source of nitrogen. Production of urease by the ure operon is necessary fr
o
m
185 for oral transmission of Y. pseudotuberculosis, but it is inactivated in Y. pestis due to the h
t
t
p
186 mutation causing a premature stop codon in ureD [33]. Since Y. pestis spends its life ://
ia
i.
187 most exclusively in a flea-host-flea cycle, the organism can lose with impunity the a
s
m
188 function of urease needed for survival in natural environments. .o
r
g
/
189 Y. pestis expresses rough LPS lacking the O-antigen, due to the inactivation of several o
n

J
190 genes in the O-antigen gene cluster (52). Y. pseudotuberculosis produces a rough LPS at a
n
u
a
191 37ºC but not at 26 ºC and a variable number of LPS genes are defective when comparing r
y

2
192 various biovars of Y. pestis (64). Expression of rough LPS is essential for Pla activity and 2
,

2
0
193 virulence in Y. pestis (32). A pathogenic advantage of rough LPS in Y. pestis is to enable 1
9

b
194 efficient Pla-mediated bacterial dissemination to cause systemic disease. y

g
u
195 For the blocked fleas, Y. pestis synthesizes an attached biofilm in the flea e
s
t
196 proventriculus and in its midgut posteriorly (25). Three distinct operons, hmsHFRS,
197 hmsT and hmsP are involved in the synthesis of bacterial extracellular matrix that is the
198 primary component of Yesinia biofilm (28). Y. pseudotuberculosis contains all of these
199 hms genes, which are 99% identical to the Y. pestis homologues (9). In addition to Hms,
200 several other proteins (GmhA, SpeAC and YrbH) involved in the biofilm formation by Y.
8
201 pestis are harbored in Y. pseudotuberculosis (Table 1).
202 Only a small portion of Y. pseudotuberculosis strains is able to form biofilm on
203 Caenorhabditis elegans, none of them has the ability to form adhesive biofilms in the
204 fleas (17). The transcriptional regulator RcsA (69) and the glycosyl hydrolase NghA (16)
205 have been shown to inhibit Yersisnia biofilm formation, but both of them are functional
206 in Y. pseudotuberculosis but inactivated in Y. pestis (16, 69). Expression of functional D
o
w
207 RcsA or NghA in Y. pestis strongly represses biofilm formation and abolishes flea n
lo
a
208 blockage (16). Therefore, Y. pestis has evolved the changes in regulatory functions on d
e
d

209 biofilm development to ensure stable biofilm formation in the flea proventriculus and fr
o
m
210 resulting in efficient arthropod-borne transmission. h
t
t
p
211 Gene loss ://
ia
i.
212 Hexa-acylated LPS observed in many Gram-negative pathogens is able to activate the a
s
m
213 TLR4 signaling and to further stimulate the host innate immune response (55). The .o
r
g
/
214 acyltransferase LpxL is required for addition of the secondary acyl chains to the o
n

J
215 tetra-acylated precursor (55). The lpxL gene is absent from Y. pestis, and makes this a
n
u
a
216 pathogen to produce tetra-acylated LPS that inhibits the TLR4 activation, which allows r
y

2
217 this pathogen to evade protective inflammatory response and establish fatal infection 2
,

2
0
218 (41). 1
9

b
219 Five additional Y. pseudotuberculosis-specific chromosomal loci (R1, R3, ORF2, y

g
u
220 ORF3 and ORF4) required for its survival, optimal growth, or virulence are absent from e
s
t
221 Y. pestis (51) (Table 1). ORF3 and ORF4 with unknown function are essential for the
222 viability of Y. pseudotuberculosis, while ORF2 (a putative pseudouridylate synthase
223 involved in RNA stability) and R3 (a genomic region composed mostly of genes of
224 unknown functions) are necessary for its optimal growth in a chemically defined medium
225 (51). Deletion of R1 (a genomic region responsible for the methionine salvage pathway)
9
226 alters the mutant’s virulence, suggesting that the availability of free methionine is
227 severely restricted in vivo (51).
228
229 Remodeling of gene regulation
230 Virulence determinants are tightly and coordinately regulated during infection.
231 Virulence-related regulators can sense the host signals, e.g. changes in temperature, and D
o
w
232 then differentially regulate not only virulence genes but other wide sets of genes required n
lo
a
233 for adaptation to host niche (84). A number of virulence-required regulators have been d
e
d

234 characterized in Y. pestis and Y. pseudotuberculosis (Table 1), indicating that remodeling fr
o
m
235 of gene regulation contributes to the denoted differences between these two pathogens h
t
t
p
236 (Figure 1). ://
ia
i.
237 Integration of laterally acquired virulence genes a
s
m
238 The global transcriptional regulators CRP (49, 79) and RovA (8, 73) are conserved .o
r
g
/
239 and required for virulence in the three pathogenic Yersinia. In addition, the Y. pestis CRP o
n

J
240 is 98.6% identical to the E. coli one with the same length, and CRP from these two a
n
u
a
241 bacteria share an identical consensus box sequence (TGTGA-N6-TCACA) that represents r
y

2
242 the conserved signals for CRP recognition of promoter DNA (79). Through 2
,

2
0
243 regulator-promoter DNA interaction in Y. pestis, CRP activates two laterally acquired 1
9

b
244 plasmid genes pla and pst (30, 79), while RovA up-regulates a genomic region y

g
u
245 YPO2272-2281 (8). The prophage YPO2272-2281 is acquired by the Y. pestis ancestor, e
s
t
246 and its genome forms an unstable episome in Antiqua and Medievalis whereas is stably
247 integrated in Orientalis (12, 36). The acquisition of this prophage does not correlate to
248 flea transmission, but contributes to virulence in mice (12). These ‘newly’ acquired
249 virulence genes have evolved to integrate themselves into the ‘ancestral’ Yersinia
250 regulatory cascade. The plague pathogen integrates laterally acquired genes to coordinate
10