Molecular detection of Enteropathogens from diarrheic stool of HIV positive patients in Gondar, Ethiopia.

BACKGROUND: Infectious diarrhea is a common problem in the developing world, especially among people living with HIV/AIDS. Traditional diagnostic methods such as stool culture and microscopic examination are limited by resources and poor sensitivity. The use of molecular diagnostics for enteropathogen detection in this region of sub-Saharan Africa has not been fully explored. We sought to identify risk factors and characterize enteropathogens from diarrheic stools of HIV-positive patients in Gondar, Ethiopia using multiplex molecular panels targeting key infectious agents. METHODS: A cross-sectional study of 100 stool samples was performed. Samples were collected consecutively from HIV- positive patients presenting with diarrhea at University of Gondar Hospital clinic, a major center in NW Ethiopia. Genomic DNA was extracted from stool and processed using a multiplex molecular panel Allplex™ [Seegene, Canada]. Correlations between patient characteristics, symptoms, public health risk factors, and enteropathogen type (s) were studied. Eighty-six samples were successfully analyzed by molecular methods. RESULTS: The mean age was 35 with 43% male. Eighty percent lived in an urban area, 18% had access to well water only, and 81% practiced proper hand hygiene. The majority of patients (72%) were receiving HAART with a median CD4 cell count of 362/µL. Multiple pathogens were detected in 94% of specimens, with an average of 5 enteropathogens per sample. Common bacteria, viruses, and parasites detected were Shigella spp./enteroinvasive E. coli (80%), enterotoxigenic E. coli (73%), Norovirus (16%) and B. hominis (62%). CD4 cell count < 500/ µL was associated with the presence of viruses (p = 0.004) and the absence of STEC (p = 0.010). The use of HAART or CD4 levels was not associated with the number of enteropathogens detected. CONCLUSIONS: Diarrheic stool from HIV-positive outpatients in Gondar, Ethiopia had on average 5 enteropathogens present in their stool. Shigellaspp./enteroinvasive E. coli and enterotoxigenic E. coli are the major pathogens, not dissimilar to immunocompetent individuals in low income countries.

Synbiotic therapy decreases microbial translocation and inflammation and improves immunological status in HIV-infected patients: a double-blind randomized controlled pilot trial.

BACKGROUND: HIV-infection results in damage and dysfunction of the gastrointestinal system. HIV enteropathy includes pronounced CD4+ T-cell loss, increased intestinal permeability, and microbial translocation that promotes systemic immune activation, which is implicated in disease progression. A synbiotic is the combination of probiotics and prebiotics that could improve gut barrier function. Our study goal was to determine whether the use of a synbiotic, probiotics or a prebiotic can recover immunological parameters in HIV-infected subjects through of a reduction of microbial translocation and pro-inflammatory cytokine production. METHODS: A randomized, double-blind controlled study was performed; twenty Antiretroviral treatment-naïve HIV-infected subjects were subgrouped and assigned to receive a synbiotic, probiotics, a prebiotic, or a placebo throughout 16 weeks. RESULTS: We had no reports of serious adverse-events. From baseline to week 16, the synbiotic group showed a reduction in bacterial DNA concentrations in plasma (p = 0.048). Moreover, the probiotic and synbiotic groups demonstrated a decrease in total bacterial load in feces (p = 0.05). The probiotic group exhibited a significant increment of beneficial bacteria load (such as Bifidobacterium; p = 0.05) and a decrease in harmful bacteria load (such as Clostridium; p = 0.063). In the synbiotic group, the CD4+ T-cells count increased (median: +102 cells/µL; p = 0.05) and the level of Interleukin 6 cytokine decreased significantly (p = 0.016). CONCLUSIONS: Our study showed a significant increase in CD4+ T lymphocyte levels in the synbiotic group, which could delay the initiation of antiretroviral therapy and decrease costs in countries with limited resources.

Altered host-microbe interaction in HIV: a target for intervention with pro- and prebiotics.

The intestinal immune system is severely affected by HIV and circulating microbial products from the intestinal tract that provide an ongoing source of systemic inflammation and concomitant viral replication. In addition, HIV-infected individuals can have a deregulated immune response that may hamper the anti-viral capacity of the host. Various probiotic organisms and prebiotic agents have been shown to enhance intestinal epithelial barrier functions, reduce inflammation, and support effective Th-1 responses. As these characteristics may benefit HIV patients, this review aims to provide a theoretical framework for the development of probiotic and prebiotic interventions specifically for this population.

Chronic diarrhea and AIDS: insights into studies with non-human primates.

Diarrhea is the pathophysiological reaction of host's gastrointestinal tract to a variety of external stimuli. Classified as a clinical syndrome, diarrhea is the leading cause of mortality and morbidity worldwide. Clinical manifestations can occur in two major forms: A) acute, which usually resolves in less than three weeks and B) chronic, which can last for months. Because of its impact on the host immune system, acquired immune deficiency syndrome (AIDS) is currently the major cause of chronic diarrhea in many parts of the world. It is estimated that up to 90% of HIV-infected individuals with symptoms of AIDS exhibit clinical diarrhea [9, 74, 55]. In SIV-infected rhesus macaques, intense infiltration of intestinal lamina propria with virus-containing lymphocytes and macrophages can be found within days after experimental virus inoculation [25, 57]. In addition to acute enteropathy syndrome, viral infection ultimately leads to other alterations of the gastrointestinal tract including persistent and/or chronic diarrhea, a condition similar to untreated AIDS of human patients. In this short review, the chronic diarrhea is presented from the perspective of the non-human primate or simian model of AIDS (SAIDS), and its most common opportunistic and pathogenic co-infections.

Isolation of potential bacterial pathogens from the stool of HIV-infected and HIV-non-infected patients and their antmicrobial susceptibility patterns in Jimma Hospital, south west Ethiopia.

Diarrhea is a major clinical problem in HIV-infected patients. There is a need to monitor antimicrobial susceptibility patterns of enteric bacterial pathogens in order to ensure appropriate treatment and control of infections. The objectives of this study was to identify and determine the magnitude of potential enteric pathogens including Salmonella, Shigella, Campylobacter and other species in HIV-infected and HIV-non-infected patients with diarrhea, to evaluate the current antimicrobial susceptibility pattern of the clinical isolates and the association of enteric bacterial pathogens in HIV infected patients with diarrhea. A cross-sectional study was conducted from Feb-July 2001 on 372 consecutive HIV seropositive and seronegative patients presenting at Jimma hospital for different illnesses. Patients were selected based on their serological tests for HIV. Sample of faeces specimens were collected and inoculated onto standard culture media as well as onto Skirrow's medium for isolation of Campylobacter species. Salmonella and Shigella species were tested for antimicrobial susceptibility using disc agar diffusion technique recommended by Kirby-Bauer. Stool specimens were also smeared and stained by Zehl-Neelson staining technique for the identification of Mycobacterium species. Among the 99 HIV-infected patients with diarrhea, 25 (25.0%) of them had enteric bacteria among which 8(8.1%) were Salmonella, 4(4.0%) Shigella and 13(13.1%) Campylobacter species. Mycobacterium species were identified in 3(3.0%) of stool specimens obtained from HIV-infected patients with diarrhea and another 3 species were detected in HIV-infected patient without diarrhea. Salmonella species were isolated with higher prevalence in HIV-infected than in HIV non-infected patients. These Salmonella isolates were 100% susceptible to Amikacin, Gentamicin, Nalidixic acid and Kanamycin while Shigella isolates were 100% susceptible for Gentamycin and Kanamycin only. Unlike Salmonella, Shigella and Campylobacter species showed higher prevalence rates in HIV non-infected patients. Enteric bacterial pathogens account for about one fourth diarrhea in HIV infected patients in Jimma hospital. The finding of this investigation also confirmed earlier observations of wide spread resistance to the commonly used drugs in this region.

[Chronic diarrhea: value of microbiology in diagnosis]. / Chronische Diarrhoe: Stellenwert der Mikrobiologie in der Diagnostik.

Chronic diarrhoea of the adult is defined as diarrhea during 30 days or longer. Frequent causes of chronic diarrhea in the immunocompetent adult without recent travel to developing countries are noninfectious processes, including laxatives misuse, diseases causing chronic maldigestion, osmotically active artificial sweeteners (i.e. sorbitol), hormonal disorders or drugs with intestinal side effects. Infectious agents as the cause of chronic diarrhea are important in two populations, namely in travelers returning from tropical countries bearing a significant risk of intestinal parasitic infections and in immunocompromised patients, especially AIDS patients with CD4 cell counts below 50 per microliter. Intestinal parasites and C. difficile, Y. enterocolitica, Shigellae and Cytomegalovirus are the most important causative agents of chronic diarrhea. Intestinal pathogens were identified in 46% of chronic, but only in 16.5% of acute diarrhea episodes of HIV-infected patients. An extensive medical history including recent travel as well as the detailed characteristics of onset of symptoms and of their time course is essential for the diagnosis. All patients should have a complete differential blood count, ESR, determination of electrolytes, liver enzymes, creatinine, blood glucose, and serum albumin. Tests to exclude hyperthyriodism, or pancreatic insufficiency as well as a d-xylose absorption test can be included, if appropriate. Microbiological-parasitological investigations are obligatory in patients with chronic diarrhea returning from countries with increased risk of traveler diarrhea, in cases of suspected immunodeficiency, if sudden onset of symptoms with fever is reported, after antibiotic treatment, and in children below six years of age. As a rule, stool specimens are appropriate, for the detection of cytomegalovirus colonic biopsies are necessary. In the latter case colonosigmoidoscopy has no diagnostic advantage. One single stool specimen is sufficient for the detection of bacteria or toxins, in contrast to parasitological investigations, where only three consecutive specimens provide sufficient diagnostic sensitivity.

Gastrointestinal manifestations of HIV infection.

Since the beginning of the AIDS pandemic, gastrointestinal (GI) problems have been among the most common features of the disease. Despite the introduction of highly active antiretroviral therapy (HAART) in 1995 and 1996, most HIV-infected patients continue to have GI complications. The clinician must be able to diagnose and treat the opportunistic gastrointestinal infections and neoplasms that occur in the advanced AIDS patient, as well as the treatment-induced symptoms and non-HIV-related GI disorders that predominate in early HIV disease. This review addresses the GI manifestations of HIV, with particular emphasis on new developments in the era of highly effective therapy.

[Endoscopic evaluation of chronic human immunodeficiency virus-related diarrhea]. / La place de l'endoscopie dans le diagnostic etiologique des diarrhées chroniques des sidéens.

Chronic diarrhea is an important clinical problem in patients infected with HIV. Data assessing the diagnostic yield of upper and lower endoscopy are limited. We reported 10 cases of HIV-infected patient referred to our hospital for chronic diarrhea from March 1995 to June 1999. 60% of the pathogens were identified obviously by stool studies. Cryptosporidium and Mycobacterium avium intracellulare (MAI) were the most common organisms. In this study, endoscopy identified 2 additional cases of MAI and one of 5 cryptosporidia detected in stool. Immunologic test identified a CMV infection in one case. Stool tests and endoscopy identified obviously 80% of the pathogens. Most investigators and us agree that stool studies should be the first diagnostic test. In patients with negative stool studies, lower endoscopy is more cost-effective than upper endoscopy and indicated as an initial exam.

A prospective study of endoscopy in HIV-associated diarrhea.

OBJECTIVE: Diarrhea commonly occurs in persons with human immunodeficiency virus (HIV) infection. The optimal use of endoscopic procedures remains poorly studied for patients with HIV-related diarrhea. The purpose of this study is to compare the diagnostic yield of a complete endoscopic work-up including an esophagogastroduodenoscopy and colonoscopy to a more limited approach of biopsies obtainable by flexible sigmoidoscopy. METHODS: A prospective study of 79 patients with HIV-related diarrhea. Upper endoscopy and colonoscopy were performed with tissue biopsies labelled according to location within the colon or small intestine. RESULTS: A new infection was diagnosed in 22 of 79 patients (28%). Biopsy of the left colon yielded an enteric pathogen in 17 of 22 patients (sensitivity: 77%) and in 15 of 15 patients with cytomegalovirus colitis (sensitivity: 100%). Combined left and right colonic biopsies had a sensitivity of 82%. Combined colonic and terminal ileum biopsies missed no pathogens. Duodenal biopsies yielded no additional pathogens beyond those identified by colonoscopy and terminal ileal biopsy. Patients with a new pathogen diagnosed had significantly lower CD4 lymphocyte counts as compared to patients without a new pathogen (p = 0.001). CONCLUSIONS: For patients with CD4 counts < 100/mm3 and unexplained AIDS-related diarrhea, flexible sigmoidoscopy with biopsy is a sufficiently thorough endoscopic evaluation.

Prevalence of intestinal pathogens in HIV patients with diarrhea: implications for treatment.

Patients infected with the human immunodeficiency virus (HIV) commonly experience diarrhea at some time during their illness. A variety of enteric pathogens are identified in 50-80% of these patients, depending on the intensity of the diagnostic work-up that is done. In addition to the common enteric pathogens, several unusual enteric pathogens are recognized to cause diarrhea especially in HIV patients. These include protozoan parasites such as Cryptosporidia, Isospora belli, Cyclospora cayatenensis and Microsporidium species bacteria such as enteropathogenic Escherichia coli and Mycobacterium avium-intracellulare, fungi including Candida albicans and Histoplasma capsulatum, and viruses such as astroviruses and caliciviruses. Diagnosis of these infections sometimes involves special procedures not readily available every where, and empiric therapy based on knowledge of the likely pathogens has been advocated for developing countries. This article reviews the currently available data on geographic variation of enteric pathogens in HIV patients with diarrhea and outlines a rational strategy for empiric therapy of these patients.

HIV enteropathy: comparative morphometry of the jejunal mucosa of HIV infected patients resident in the United Kingdom and Uganda.

AIMS: To compare jejunal mucosal morphometry in HIV infected patients resident in London and Uganda. PATIENTS: Twenty HIV positive patients from London and 16 from Uganda were studied, and compared with HIV negative control subjects from both sites. METHODS: Stools and biopsy specimens were examined for enteropathogens. Surface area to volume (S:V) ratio was estimated morphometrically, mean crypt length of jejunal biopsy specimens was measured, and HIV infected cells detected immunohistochemically were quantified. RESULTS: Enteric pathogens were detected in none of the London patients, and in three Ugandan patients. S:V ratio was lower, and mean crypt length higher, in the specimens of London patients than in normal subjects, but there was no difference in S:V ratio or mean crypt length between Ugandan patients and controls. A negative correlation was present between S:V ratio and mean crypt length in all biopsy specimens analysed. HIV infected cells were detected only in lamina propria. CONCLUSION: Infection of cells in the lamina propria of the jejunum with HIV stimulates crypt cell proliferation, and a fall in villous surface area. The mucosal response to HIV is masked by other pathogens in the African environment.

Managing and treating. AIDS-related diarrhea.

Diarrhea is a significant problem for most people with AIDS. Its causes are multifactorial, ranging from infectious organisms to malignancies and functional problems. A thorough evaluation will usually determine the etiology and guide treatment. With HAART, dramatic improvements in immune status have caused a decline in opportunistic infections, including those that cause diarrhea. It is a sign of hope for all patients with HIV.

Small intestine pathogens in AIDS: conventional and opportunistic.

The small intestine, coming in direct contact with ingested potential pathogens, depends on active mucosal immunity to withstand invasion and damage. In patients with AIDS and severe impairment of immunoregulatory lymphocytes, proliferation of protozoal, viral, bacterial, and fungal pathogens produces diarrhea and malabsorption. When noninvasive tests of stool and blood fail to identify responsible organisms, endoscopy can reveal mucosal lesions which are suggestive if not diagnostic. Cryptosporidium, cf2E. intestinalis, cf1CMV, MAC, and other infections can be identified by intestinal biopsy quicker and often at lower overall cost than they can be by culture.

HEp-2 cell-adherent Escherichia coli and intestinal secretory immune response to human immunodeficiency virus (HIV) in outpatients with HIV-associated diarrhea.

HEp-2 cell-adherent Escherichia coli and the human immunodeficiency virus (HIV) itself have recently been incriminated as causes of chronic HIV-associated diarrhea. This study sought to determine the prevalence of these two agents among HIV-infected patients with diarrhea in an outpatient setting in the United States and to compare their prevalence to that of other commonly recognized enteropathogens known to be present in this population. HEp-2 cell-adherent E. coli was found in 20 of 83 (24.1%) patients with diarrhea. A diffuse pattern of adherence was the most common, found in 14 of 20 (70%) patients, followed by a localized adherence pattern (6 of 20; 30%). An intestinal secretory immune response against the p24 antigen of HIV was found in 9 of 34 (27.5%) patients with HIV-associated diarrhea. The following pathogens or products were also detected in lower frequencies: Cryptosporidium spp. (10.8%), Clostridium difficile toxin (8.8%), microsporidia (6%), Isospora belli (3.6%), Blastocystis hominis (2.4%), Giardia spp. (1.2%), Salmonella spp. (1.2%), and Mycobacterium spp. (1.2%). The role of HEp-2 cell-adherent E. coli and HIV enteric infections in patients with HIV-associated diarrhea deserves further study.

Morphological injury of the intestinal mucosa and infection in patients with AIDS. The role of combined tissue and stool examination.

AIMS: To define the relationship between morphological injury of the intestinal mucosa and infections in AIDS patients. METHODS: Forty-nine AIDS patients were examined by upper gastrointestinal (GI) endoscopy and 8 of them also by lower GI endoscopy. Biopsy specimens, taken from the lower duodenum, esophagus and rectum, were studied by light (L.M.) and transmission electron microscopy (T.E.M.). Stool examination for microorganisms was routinely performed in all patients. RESULTS: Microorganisms were detected in 37 of the 49 patients (75.5%) by combined tissue and stool examination. The histological study revealed villous atrophy, inter- and intra-enterocyte oedema and epithelial degenerative changes in most of the patients whether or not they had detectable microorganisms. CONCLUSIONS: Combined methods (endoscopy, L.M. and T.E.M., studies of tissue samples, microbiological study of stool samples) may be used to improve the documentation of infections and morphological injury of the intestinal mucosa in AIDS patients.

Results of culture form colonoscopically obtained specimens for bacteria and fungi in HIV-infected patients with diarrhea.

BACKGROUND: The aim of our study was to determine the diagnostic yield of culture for bacteria and fungi from colonic biopsy specimens in 290 consecutive HIV-infected patients with diarrhea. METHODS: During each colonoscopy, three biopsy specimens were homogenized and cultured on media for Salmonella and Shigella and for Campylobacter and Yersinia, on Loewenstein medium and on Sabouraud medium. RESULTS: Cultures were found positive for one (n = 32) or two (n = 5) infectious agents in 37 cases, i.e., in 12.8% of the patients. Bacteria were isolated in 24 cases, and identified as Campylobacter jejunl-coli (n = 14), Salmonella (n = 2), Shigella (n = 1), or Pseudomonas aeruginosa (n = 7). Among the 14 patients with C. jejuni-coli intestinal infection, 11 had normal-appearing mucosa at colonoscopy, and 3 had a concomitant stool culture negative for Campylobacter. Mycobacterial cultures were positive for Mycobacterium avium intracellulare in 6 patients, who were already known as having a disseminated M. avium intracellulare infection from positive blood cultures. Fungal cultures were positive for Candida in 10 cases, without clear clinical significance. CONCLUSIONS: The overall yield of culture for bacterial pathogens from colonic tissue in HIV-infected patients with diarrhea is low, but some individual cases of C. jejuni-coli infections may be detected from colonic tissue culture and not diagnosed by concomitant stool culture.