Early impairment of gut function and gut flora supporting a role for alteration of gastrointestinal mucosa in human immunodeficiency virus pathogenesis.
ABSTRACT Our results show that impairment of the gastrointestinal tracts in human immunodeficiency virus (HIV)-positive patients is present in the early phases of HIV disease. This impairment is associated with alterations in gut microbiota and intestinal inflammatory parameters. These findings support the hypothesis that alterations at the gastrointestinal-tract level are a key factor in HIV pathogenesis.
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ABSTRACT: HIV causes rapid CD4+ T cell depletion in the gut mucosa, resulting in immune deficiency and defects in the intestinal epithelial barrier. Breakdown in gut barrier integrity is linked to chronic inflammation and disease progression. However, the early effects of HIV on the gut epithelium, prior to the CD4+ T cell depletion, are not known. Further, the impact of early viral infection on mucosal responses to pathogenic and commensal microbes has not been investigated. We utilized the SIV model of AIDS to assess the earliest host-virus interactions and mechanisms of inflammation and dysfunction in the gut, prior to CD4+ T cell depletion. An intestinal loop model was used to interrogate the effects of SIV infection on gut mucosal immune sensing and response to pathogens and commensal bacteria in vivo. At 2.5 days post-SIV infection, low viral loads were detected in peripheral blood and gut mucosa without CD4+ T cell loss. However, immunohistological analysis revealed the disruption of the gut epithelium manifested by decreased expression and mislocalization of tight junction proteins. Correlating with epithelial disruption was a significant induction of IL-1β expression by Paneth cells, which were in close proximity to SIV-infected cells in the intestinal crypts. The IL-1β response preceded the induction of the antiviral interferon response. Despite the disruption of the gut epithelium, no aberrant responses to pathogenic or commensal bacteria were observed. In fact, inoculation of commensal Lactobacillus plantarum in intestinal loops led to rapid anti-inflammatory response and epithelial tight junction repair in SIV infected macaques. Thus, intestinal Paneth cells are the earliest responders to viral infection and induce gut inflammation through IL-1β signaling. Reversal of the IL-1β induced gut epithelial damage by Lactobacillus plantarum suggests synergistic host-commensal interactions during early viral infection and identify these mechanisms as potential targets for therapeutic intervention.PLoS Pathogens 08/2014; 10(8):e1004311. · 8.14 Impact Factor
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ABSTRACT: Survival rates among HIV patients have significantly improved since the introduction of antiretroviral therapy (ART) in HIV management. However, persistent disease progression and clinical complications in virally suppressed individuals point to additional contributing factors other than HIV replication; microbial translocation is one such factor. The role of underlying commensal microbes and microbial products that traverse the intestinal lumen into systemic circulation in the absence of overt bacteraemia is under current investigation. This review focuses on current knowledge of the complex microbial communities and microbial markers involved in the disruption of mucosal immune T-cells in the promotion of inflammatory processes in HIV infections. Unanswered questions and aims for future studies are addressed. We provide perspective for discussing potential future therapeutic strategies focused on modulating the gut microbiota to abate HIV disease progression.Gastroenterology research and practice. 01/2014; 2014:803185.
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ABSTRACT: Current pathogenetic aspects on HIV infection highlight the importance of a chronic immune activation ultimately leading to T lymphocyte homeostasis disruption and immune deregulation associated with disease manifestations and progression. It is widely accepted that this continuous immune activation in HIV infection is principally driven by the phenomenon of pathological microbial translocation (MT).Infection 07/2014; · 2.44 Impact Factor
JOURNAL OF CLINICAL MICROBIOLOGY, Feb. 2008, p. 757–758
Copyright © 2008, American Society for Microbiology. All Rights Reserved.
Vol. 46, No. 2
Early Impairment of Gut Function and Gut Flora Supporting a Role
for Alteration of Gastrointestinal Mucosa in Human
Immunodeficiency Virus Pathogenesis?
Andrea Gori,1,2* Camilla Tincati,2Giuliano Rizzardini,3Carlo Torti,4Tiziana Quirino,5
Monique Haarman,6Kaouther Ben Amor,6Jacqueline van Schaik,6Aldwin Vriesema,6
Jan Knol,6Giulia Marchetti,7Gjalt Welling,8and Mario Clerici9
Division of Infectious Diseases, Department of Internal Medicine, San Gerardo Hospital, University of Milano-Bicocca, Monza,
Italy1; Department of Internal Medicine, Clinic of Infectious Diseases, San Paolo Hospital, University of Milan, Milan, Italy2;
1st Division of Infectious Diseases, Luigi Sacco Hospital, Milan, Italy3; Clinic of Infectious Diseases, University of Brescia,
Brescia, Italy4; Infectious Diseases Unit, Busto Arsizio Hospital, Busto Arsizio, Italy5; Numico Research BV, Wageningen,
The Netherlands6; Department of Clinical Science, Luigi Sacco Hospital, University of Milan, Milan, Italy7; Department of
Medical Microbiology, University Medical Center Groningen, University of Groningen, Groningen, The Netherlands8;
and Department of Science Technology Biomedicine, University of Milan, Milan, Italy9
Received 30 August 2007/Returned for modification 8 October 2007/Accepted 29 November 2007
Our results show that impairment of the gastrointestinal tracts in human immunodeficiency virus (HIV)-
positive patients is present in the early phases of HIV disease. This impairment is associated with alterations
in gut microbiota and intestinal inflammatory parameters. These findings support the hypothesis that alter-
ations at the gastrointestinal-tract level are a key factor in HIV pathogenesis.
The hallmark of human immunodeficiency virus (HIV) in-
fection is the depletion of CD4?T cells, which occurs through-
out the entire disease. Recent data have shown that primary
HIV infection is associated with a preferential depletion of
CD4?T cells in the gastrointestinal (GI) tract, where more
than 60% of T lymphocytes reside, suggesting a pivotal role for
the early impairment of the gut-associated lymphoid tissue in
HIV disease (4). However, the precise pathogenic mechanisms
underlying the profound CD4?T-cell loss observed in the
various stages of disease are only partially understood.
A characteristic feature of HIV disease is chronic immune
activation, which might play a crucial role in the CD4?T-cell
depletion. It has been recently hypothesized that a breakdown
of the GI mucosal barrier may contribute to chronic immune
activation (4). Thus, the exposure of peripheral immune cells
to microbial products after gut injury may result in the abnor-
mal activation of such cells.
The healthy GI tract is colonized by a large variety of com-
mensal microbes that impact on the development of the
humoral and cellular mucosal immune system (11, 12). This
microbiota is shielded from the immune system via a strong
mucosal barrier. Infections and antibiotics are known to alter
both the normal GI-tract barrier and the microbiota, which
may result in possible immune abnormalities. If HIV infection
itself impaired the GI barrier and the composition of the gut
microbiota, the breakdown of the GI mucosa would result in
the sudden and chronic exposure of peripheral lymphocytes to
an abnormal intestinal microbiota (4).
To verify this hypothesis, we studied the effect of HIV in-
fection on the commensal intestinal microbiota and on intes-
tinal inflammation by analyzing the composition of GI tract
microbiota and by measuring the levels of fecal calprotectin in
57 healthy, asymptomatic HIV-positive, antiretrovirus-naive
individuals (average CD4?T-lymphocyte count and HIV RNA
levels of 520 cells/?l and 28,393 copies/ml, respectively) en-
rolled in a trial in Italy (COPA study). The results were com-
pared to historical data from the general population not suf-
fering from HIV infection. Informed consent was obtained
from all of the patients.
The fecal microbiota of the HIV patients was analyzed by
either fluorescence in situ hybridization or quantitative real-
time PCR (Q-PCR). Preparation of fixed fecal cells and DNA
extraction were performed as described previously (6). For
fluorescence in situ hybridization, a set of specific rRNA-tar-
geted oligonucleotides probes were used to quantify Can-
dida albicans (Caal [5?-GCCAAGGCTTATACTCGCT-3?]),
bifidobacteria (Bif164 [5?-CATCCGGCATTACCA CCC]),
lactobacilli (Lab158 [5?-GGTATTAGCAYCTGTTTCCA]),
and total bacteria (Eub338 (5?-GCTGCCTCC CGTAGGA
GT)) (1, 7, 8, 10). The relative abundance of bacterial groups
was determined as the proportion of cells hybridizing with the
specific probe to the total cells positive for the Eub338 probe.
For the quantification of Pseudomonas aeruginosa, a Q-PCR-
based method was used as described by Pirnay et al. (14).
Calprotectin was measured by using a commercial enzyme-
linked immunosorbent assay kit (PhiCal, Eurospital, Italy).
Initially, we focused our attention on two opportunistic
pathogens: P. aeruginosa and C. albicans. The presence of
these species was found to be very high in this HIV-positive
patient population compared to levels reported for healthy
individuals. P. aeruginosa was identified in 92% of all fecal HIV
samples analyzed. In healthy individuals, P. aeruginosa was
* Corresponding author. Mailing address: Division of Infectious
Diseases, Department of Internal Medicine, San Gerardo Hospital,
University of Milano-Bicocca, Via Solferino 16-20052, Milan, Italy.
Phone: 39 039 2333794. Fax: 39 039 2333898. E-mail: andrea.gori
?Published ahead of print on 19 December 2007.
found in only 20% of samples (unpublished data) using the
same Q-PCR method. Moreover, P. aeruginosa accounted for
0.7% ? 0.13% of the total microbiota in our HIV positive
group and represented a 10-fold increase compared to the
levels in the healthy individuals. Similarly, C. albicans was
detected in 100% of the feces samples of our HIV-positive
group, whereas Bernhardt and Knoke (3) reported that in the
general population only 40% are positive carriers of Candida.
More importantly, the average count of C. albicans was almost
10,000-fold higher in the HIV-positive group (ranging from
[9.1 ? 1.0] ? 107to [8.9 ? 1.0] ? 107cells/g [wet weight] of
feces) than reported for the HIV-negative general population
(?104cells/g [wet weight] of feces) (3).
Further support of the hypothesis of an impairment of the
GI microbiota in HIV-infected patients was revealed by data
showing lower levels in other microbial species, e.g., bifidobac-
teria and lactobacilli, compared to the levels reported for the
general population. Both species have been shown to have a
positive influence on mucosal immune function and gut health
(2, 5, 15, 17). The relative amount of bifidobacteria in the
HIV-positive population investigated during our trial was 2.5%
(95% confidence interval ? 1.4 to 4.2%) of the total fecal
bacteria, while the lactobacilli were nearly undetectable at
0.02% (95% confidence interval ? 0.009 to 0.05%). It is well
documented that in the general population bifidobacteria can
vary between 5 and 10% of the total bacterial community,
while lactobacilli represent 1 to 2% (9, 13). Our data thus
suggest that in HIV-infected individuals the composition of the
fecal microbiota is atypical at an early stage of infection.
Levels of fecal calprotectin, a protein secreted by recruited
neutrophils in the intestinal lining indicative of intestinal in-
flammation and used as a marker of mucosal inflammatory
activity in patients with inflammatory bowel disease (4, 11, 16,
17), were also analyzed. The cutoff for this marker is at 50 ?g/g
(wet weight) of feces, with a median value in healthy individ-
uals of 26 ?g/g (16). Our results revealed that in our HIV
patient group half of the subjects (27 of 53) had increased fecal
calprotectin levels (?50 ?g/g). Even 34% (18 of 53) of the HIV
patients had levels over 100 ?g/g (wet weight) of feces, a
finding that is clearly indicative of a significant GI inflamma-
tion. Since intestinal inflammation is known to reduce the
intestinal barrier function, these data confirm the breakdown
of the intestinal barrier.
Abnormal immune activation represents one of the main
factors associated with HIV disease progression. A better un-
derstanding of the mechanisms driving chronic activation of
the immune system could shed light onto the mechanism(s)
associated with immune cell depletion. The hypothesis that
chronic activation stems from a damaged mucosal barrier and
stimulation of immune cells by microbial products is therefore
The results presented here clearly show that impairment of
the GI tract in HIV-positive patients is present already in the
early phases of HIV disease. This impairment is associated
with elevated levels of intestinal inflammatory parameters and
clear alterations in the gut commensal microbiota, confirming
a possible correlation between intestinal microbial alteration,
GI mucosal damage, and immune activation status. These find-
ings strongly support the recent hypothesis that alterations at
the GI-tract level are a key factor in the pathogenesis of
chronic HIV infection.
We thank Mauro Moroni for helpful discussion and suggestions. We
are grateful to Alan Michael Rosen for critical reading of the manu-
script and valuable grammatical advice. We are also grateful to Daria
Trabattoni for her continuous help and to Tiziana Formenti for excel-
1. Amann, R., B. M. Fuchs, and S. Behrens. 2001. The identification of micro-
organisms by fluorescence in situ hybridisation. Curr. Opin. Biotechnol.
2. Bernet, M., D. Brassart, J. Neeser, and A. Servin. 1993. Adhesion of human
bifidobacterial strains to cultured human intestinal epithelial cells and inhi-
bition of enteropathogen-cell interactions. Appl. Environ. Microbiol. 59:
3. Bernhardt, H., and M. Knoke. 1997. Mycological aspects of gastrointestinal
microflora. Scand. J. Gastroenterol. Suppl. 222:102–106.
4. Brenchley, J. M., D. A. Price, T. W. Schacker, T. E. Asher, G. Silvestri, S.
Rao, Z. Kazzaz, E. Bornstein, O. Lambotte, D. Altmann, B. R. Blazar, B.
Rodriguez, L. Teixeira-Johnson, A. Landay, J. N. Martin, F. M. Hecht, L. J.
Picker, M. M. Lederman, S. G. Deeks, and D. C. Douek. 2006. Microbial
translocation is a cause of systemic immune activation in chronic HIV in-
fection. Nat. Med. 12:1365–1371.
5. De Vrese, M., and P. R. Marteau. 2007. Probiotics and prebiotics: effects on
diarrhea. J. Nutr. 137:803S–811S.
6. Haarman, M., and J. Knol. 2005. Quantitative real-time PCR assays to
identify and quantify fecal Bifidobacterium species in infants receiving a
prebiotic infant formula. Appl. Environ. Microbiol. 71:2318–2324.
7. Harmsen, J. H. M., P. Elfferich, F. Schut, and G. W. Welling. 1999. A 16S
rRNA-targeted probe for detection of lactobacilli and enterococci in faecal
samples by fluorescent in situ hybridization. Microbiol. Ecol. Health Dis.
8. Hogardt, M., K. Trebesius, A. M. Geiger, M. Hornef, J. Rosenecker, and
J. Heesemann. 2000. Specific and rapid detection by fluorescent in situ
hybridization of bacteria in clinical samples obtained from cystic fibrosis
patients. J. Clin. Microbiol. 38:818–825.
9. Kolida, S., D. Meyer, and G. R. Gibson. 2007. A double-blind placebo-
controlled study to establish the bifidogenic dose of inulin in healthy humans.
Eur. J. Clin. Nutr. 61:1189–1195.
10. Langendijk, P., F. Schut, G. Jansen, G. Raangs, G. Kamphuis, M. Wilkin-
son, and G. Welling. 1995. Quantitative fluorescence in situ hybridization of
Bifidobacterium spp. with genus-specific 16S rRNA-targeted probes and its
application in fecal samples. Appl. Environ. Microbiol. 61:3069–3075.
11. Macpherson, A. J., and N. L. Harris. 2004. Interactions between commensal
intestinal bacteria and the immune system. Nat. Rev. Immunol. 4:478–485.
12. Mueller, C., and A. J. Macpherson. 2006. Layers of mutualism with com-
mensal bacteria protect us from intestinal inflammation. Gut 55:276–284.
13. Mueller, S., K. Saunier, C. Hanisch, E. Norin, L. Alm, T. Midtvedt, A.
Cresci, S. Silvi, C. Orpianesi, M. C. Verdenelli, T. Clavel, C. Koebnick,
H.-J. F. Zunft, J. Dore, and M. Blaut. 2006. Differences in fecal microbiota
in different european study populations in relation to age, gender, and
country: a cross-sectional study. Appl. Environ. Microbiol. 72:1027–1033.
14. Pirnay, J.-P., D. de Vos, L. Duinslaeger, P. Reper, C. Vandenvelde, P.
Cornelis, and A. Vanderkelen. 2000. Quantitation of Pseudomonas aerugi-
nosa in wound biopsy samples: from bacterial culture to rapid ‘real-time’
polymerase chain reaction. Crit. Care 4:255–261.
15. Saavedra, J. M., N. A. Bauman, J. A. Perman, R. H. Yolken, J. M. Saavedra,
N. A. Bauman, and I. Oung. 1994. Feeding of Bifidobacterium bifidum and
Streptococcus thermophilus to infants in hospital for prevention of diarrhoea
and shedding of rotavirus. Lancet 344:1046–1049.
16. Ton, H., Ø. Brandsnes, S. Dale, J. Holtlund, E Skuibina, H Schjonsby, and
B. Johne. 2000. Improved assay for fecal calprotectin. Clin. Chim. Acta
17. Vaughan, E. E., M. C. de Vries, E. G. Zoetendal, K. Ben-Amor, A. D. L.
Akkermans, and W. M. de Vos. 2002. The intestinal LABs. Antonie van
758NOTES J. CLIN. MICROBIOL.