Establishment of Health Utility Indices for Post-Infectious Functional Gastrointestinal Disorders in Active Duty US Military.

INTRODUCTION: Knowledge of disease burden attributable to functional gastrointestinal disorders (FGD) in travelers is lacking, despite the high incidence of travelers' diarrhea (TD) associated with increased FGD risk. One tool for assessing the impact of disease on health-related quality of life is the health utility index (HUI), which values health states based on preferential health outcomes. Health utilities can be used as preference weights in the estimation of quality-adjusted life-years (QALYs). METHODS: Six months following travel to Egypt or Turkey, 120 US military personnel completed a survey on TD during deployment, health-related quality of life (SF-36), and the onset of functional bowel disorders (Rome II). Elements from the SF-36 were used to develop SF-6D values, which were combined with health state valuations to enable calculation of HUI scores for each subject. Mean index scores were compared across functional outcomes, specific symptoms, and demographic profiles. RESULTS: The presence of FGD significantly reduced index scores, with irritable bowel syndrome (IBS) and dyspepsia showing the greatest impact (-0.17 and -0.19, respectively) compared with those with no FGD (p < 0.05). Importantly, however, several individuals met multiple FGD outcome definitions. Additionally, a number of symptoms associated with abnormal bowel habits and abdominal pain were associated with reduced index scores regardless of outcome. CONCLUSION: FGD are associated with significant morbidity as assessed by HUIs. Given the strong link between TD and FGD as well as the large number of travelers from the developed to the developing world, additional study is needed to further understand this association and efforts aimed at primary disease prevention are warranted.

Seroepidemiologic survey for Coxiella burnetii among US military personnel deployed to Southwest and Central Asia in 2005.

We used a seroepidemiologic study to estimate Q fever (Coxiella burnetii) seroprevalence, seroincidence, and risk factors for seroconversion in two deployed military populations in 2005. The first study group resided in an area with a known Q fever outbreak history (Al Asad, Iraq). Of this population, 7.2% seroconverted for an incidence rate of 10.6 seroconversions per 1,000 person-months. The second population included personnel transiting through Qatar on mid-deployment leave from southwest/central Asia. In this group, we found 2.1% prevalence with 0.92 seroconversions per 1,000 person-months. However, no significant risk factors for Q fever seroconversion were found in either population.

Genotypic characterization of Egypt enterotoxigenic Escherichia coli isolates expressing coli surface antigen 6.

INTRODUCTION: One approach to control enterotoxigenic Escherichia coli (ETEC) infections has been to develop vaccines focused on inducing protective immunity against surface expressed antigenic factors. One such factor is coli surface antigen 6 (CS6); ETEC isolates expressing CS6 may also simultaneously co-express surface antigens CS4 or CS5. However, there is little information regarding the inter-relationships of isolates expressing the CS6 antigen alone or in combination with CS4 or CS5. METHODOLOGY: A total of 62 CS6-associated ETEC isolates were evaluated for their antimicrobial susceptibility, mechanisms of resistance, toxin genes, colonization factor expression, and XbaI-pulsed-field gel electrophoretic profiles. RESULTS: We observed 46 XbaI profiles; 31 were exclusive to ETEC expressing CS6 alone and 15 among the ETEC co-expressing CS4 or CS5. Nearly half (47%) of these isolates were resistant to ampicillin, a third (37%) of the isolates were resistant to trimethoprim-sulfamethoxazole, and 24% of the isolates were tetracycline-resistant. A blaTEM gene was detected in 24 (83%) ampicillin-resistant isolates. Trimethoprim-sulfamethoxazole-resistant isolates (n = 23) carried either sulI (n = 1, 4%), sulII (n = 8, 35%) or both genes (n = 10, 43%); 4 had no detectable sul gene. CONCLUSIONS: Our results show a lack of clonality among Egypt CS6 E. coli isolates and supports the use and the further research on vaccines targeting this cell surface antigen.

The impact of post-infectious functional gastrointestinal disorders and symptoms on the health-related quality of life of US military personnel returning from deployment to the Middle East.

BACKGROUND AND AIM: Mental and physical health-related quality of life (HRQOL) are important health impact measures following military deployment. While conditions such as post-traumatic stress disorder (PTSD) are known to adversely affect QOL, little is known about the effect of post-infectious functional gastrointestinal disorders (PI-FGID). Our aim was to evaluate the risk of PI-FGID and its impact on HRQOL among military personnel returning from deployment. METHODS: A cross-sectional cohort of active-duty military deployed to Egypt or Turkey between 2004 and 2005 was asked to complete a questionnaire (Rome II and SF-36 instruments) on travelers' diarrhea (TD) during deployment and FGID symptoms and HRQOL 6 months after returning from deployment. RESULTS: A total of 121 military personnel returning from Egypt (n = 33) and Turkey (n = 88) completed the post-deployment questionnaire. Nearly half (48.3%) met the definition for an FGID at the time of the survey, and 53% of individuals reporting one or more episodes of TD during deployment developed an FGID, compared to 33% of those not reporting TD (odds ratio [OR] 2.2, P = 0.08). Compared to those not meeting the FGID criteria, those with post-deployment FGID had lower mean mental HRQOL scores (-13.4%, P < 0.0001) and lower physical HRQOL scores (-7.2%, P = 0.004). CONCLUSIONS: There was a high prevalence of FGID symptoms in military personnel returning from deployment, and TD was a noted risk factor. FGID and symptoms decreased QOL, with mental HRQOL being affected more than physical HRQOL. These findings require further research in order to assess the long-term impact of these and other post-infectious sequela related to TD during deployments among returning veterans.

A prospective study of acute diarrhea in a cohort of United States military personnel on deployment to the Multinational Force and Observers, Sinai, Egypt.

To better understand the epidemiology of diarrhea in deployed personnel to the Middle East, a prospective cohort study of travelers' diarrhea (TD) was conducted between May 2004 and January 2005 at the Multinational Force and Observers (MFO) camp in the southern Sinai. A baseline entry questionnaire and stool specimen was provided on study entry, and volunteers were followed every 6 weeks. Of 211 volunteers, 145 (68.7%) completed one or more follow-up visits. In total, 416 follow-up surveys were completed, which described an overall incidence of 25.2 episodes per 100 person months (95% confidence interval = 21.2-30.0). Additionally, stools were collected in 72 of 77 diarrhea-associated clinic visits, with bacterial pathogens most commonly isolated (enterotoxigenic Escherichia coli in 30 [42%] samples and Campylobacter jejuni in 7 [10%] samples) Despite modern preventive methods, diarrhea is still a common problem for deployed US military personnel in Egypt, frequently resulting in diminished ability to work.

Detection of gyrA mutation among clinical isolates of Campylobacter jejuni isolated in Egypt by MAMA-PCR.

INTRODUCTION: Campylobacter spp are the major cause of enteritis in humans and more than 90% of reported infections are caused by Campylobacter jejuni. Fluoroquinolones such as ciprofloxacin are the antibiotics of choice for treatment. An increase in the frequency of ciprofloxacin-resistant Campylobacter has been reported globally due to a single base mutation (C-257 to T) in codon 86 of the quinolone resistance determining region (QRDR) of the gyrA gene altering the amino acid sequence from threonine at position 86 to isoleucine (Thr-86 to Ile). METHODOLOGY: Campylobacter spp (n = 118) were selected from a collection of Egyptian isolates spanning 1998 to 2005. The presence of C. jejuni gyrA gene was confirmed in each isolate by a PCR assay amplifying 368 bp portion of the gyrA gene. C to T alteration was detected by the mismatch amplification mutation assay MAMA PCR. The MIC of nalidixic acid (NA) and ciprofloxacin (CIP) was determined by E-test. RESULTS: C. jejuni gyrA gene was detected in 100 of the Campylobacter spp studied; the other 18 isolates were found to be Campylobacter coli by lpxA PCR. The mutation was detected in 89 C. jejuni resistant isolates with MIC values (NA; 8 - >256 µg/ml) and (CIP; 4 - >32 µg/ml). The other 11 sensitive C. jejuni isolates with MIC values (NA; 0.38 - 3 µg/ml) and (CIP; 0.03 - 0.125 µg/ml) were not amplified by the MAMA primers. There was 100% congruence with MAMA PCR, MIC results and gyrA gene sequence analysis. CONCLUSIONS: In Egypt the main mechanism for resistance to fluoroquinolones is an alteration in the gyrA QRDR. MAMA PCR provides an economical and rapid means for screening fluoroquinolone resistance.

Universal detection and identification of avian influenza virus by use of resequencing microarrays.

Zoonotic microbes have historically been, and continue to emerge as, threats to human health. The recent outbreaks of highly pathogenic avian influenza virus in bird populations and the appearance of some human infections have increased the concern of a possible new influenza pandemic, which highlights the need for broad-spectrum detection methods for rapidly identifying the spread or outbreak of all variants of avian influenza virus. In this study, we demonstrate that high-density resequencing pathogen microarrays (RPM) can be such a tool. The results from 37 influenza virus isolates show that the RPM platform is an effective means for detecting and subtyping influenza virus, while simultaneously providing sequence information for strain resolution, pathogenicity, and drug resistance without additional analysis. This study establishes that the RPM platform is a broad-spectrum pathogen detection and surveillance tool for monitoring the circulation of prevalent influenza viruses in the poultry industry and in wild birds or incidental exposures and infections in humans.

Acute gastrointestinal infection, respiratory illness, and noncombat injury among US military personnel during Operation Bright Star 2005, in Northern Egypt.

BACKGROUND: In the fall 2005, approximately 7,500 US military personnel participated in an exercise in the Egyptian desert. The epidemiology of disease and noncombat injury among deployed troops is important in the context of assessing current mitigation strategies and the development of future ones. METHODS: To assess the prevalence and impact of diarrhea and enteropathogen distribution, we conducted a case series study. To assess the relative impact of diarrhea compared to respiratory infection and injury, we conducted a post-deployment survey and compared these data to clinic-based syndromic surveillance data. RESULTS: We enrolled 43 patients with acute diarrhea, 21 (49%) having one or more pathogens isolated. Enterotoxigenic Escherichia coli (n= 16), enteroaggregative E coli (n= 3), and Shigella spp. (n= 3) were the most common pathogens identified. Respiratory illness had the highest incidence (73 episodes/100 person-months) compared to diarrhea (35 episodes/100 person-months) and noncombat injury (17 episodes/100 person-months), though noncombat injury more frequently resulted in lost duty days and health-care utilization. CONCLUSIONS: Noncombat injuries and illnesses have had a significant impact on military missions and continue to result in force health protection challenges today. Future studies are needed to test and evaluate countermeasures to mitigate these illnesses and injuries to increase the health of the individuals and optimize mission readiness.

Azithromycin and loperamide are comparable to levofloxacin and loperamide for the treatment of traveler's diarrhea in United States military personnel in Turkey.

BACKGROUND: The recommended treatment for traveler's diarrhea is the combination of an appropriate antibiotic (usually a fluoroquinolone) and loperamide. Azithromycin compared favorably with fluoroquinolones in trials that did not include the use of loperamide, but combination therapy has not, to our knowledge, been studied to date. METHODS: A randomized, double-blind trial was conducted at Incirlik Air Base, Turkey, fromJ une 2003 through August 2004. Adults from the United States with noninflammatory diarrhea were randomized to receive a single dose of azithromycin (1000 mg; 106 persons) or levofloxacin (500 mg; 101 persons) plus loperamide (4 mg initially and as needed thereafter). Volunteers maintained a symptom diary and were evaluated on days 1, 3, and 7 after treatment. RESULTS: No differences were noted with respect to pretreatment symptoms or pathogen distribution. Enterotoxigenic Escherichia coli was the most common pathogen isolated (from 45% of patients in the azithromycin group and 42% of patients in the levofloxacin group), and Campylobacter species was the second most common pathogen isolated (from 6% of patients in the azithromycin group and 9% of patients in the levofloxacin group). Median time to last diarrheal stool (azithromycin group, 13 h; levofloxacin group, 3 h), median time to resolution of associated symptoms (2 days), and additional loperamide usage (azithromycin group, 39% of patients; levofloxacin group, 34% of patients) were similar between groups. Azithromycin use was associated with more nausea in the 30 min after dosing (azithromycin group, 8% of patients; levofloxacin group, 1% of patients; Pp.004), but no vomiting or other adverse events were noted in either group. CONCLUSIONS: Single-dose treatment with azithromycin (1000 mg) and loperamide is as effective as single-dose treatment with levofloxacin (500 mg) and loperamide for noninflammatory diarrhea. Although nausea after dosing is uncommon, it is more frequently associated with azithromycin than with levofloxacin. Future studies should focus on determining whether lower doses of azithromycin would decrease the frequency of nausea and decrease treatment costs without affecting efficacy.

Incidence, etiology, and impact of diarrhea among deployed US military personnel in support of Operation Iraqi Freedom and Operation Enduring Freedom.

A health assessment survey was collected from US military personnel deployed to the Middle East taking part in the "Rest and Recuperation" program or on temporary assignment to Camp As Sayliyah Doha, Qatar, from January to December 2004. In addition, a concurrent clinic-based observational study was conducted to determine pathogen etiology and potential risk factors. From 28,322 health assessment surveys, overall self-reported incidence of diarrhea was 4.9 cases per 100 person-months. Disease incidence increased with rank and was higher in Iraq compared with Afghanistan. During this period, 109 US military personnel with acute diarrhea and 85 asymptomatic personnel were enrolled in the observational study. Enterotoxigenic E. coli (ETEC) was the predominant pathogen (32%), followed by enteroaggregative E. coli (12%) and Salmonella spp. (6%). These data are consistent with previous reports implicating ETEC as the primary cause of acute diarrhea for military personnel deployed to this region.

Enteric pathogens associated with diarrhea in children in Fayoum, Egypt.

In a cross-sectional study of children <60 months old from Fayoum, Egypt, presenting with diarrhea, 46% (162/356) had detectable enteric pathogens. Bacterial pathogens were identified in 25% (89/356), whereas rotavirus and Cryptosporidium were detected in 21% (54/253) and 15% (39/253), respectively. Cryptosporidium is an important pathogen in this region.

Enterotoxigenic Escherichia coli colonization factor types collected from 1997 to 2001 in US military personnel during operation Bright Star in northern Egypt.

Operation Bright Star (OBS) is a biennial, multinational exercise in Egypt involving 15000 US troops. Consistent with past observations in deployed troops, diarrhea is the most significant cause of morbidity. Focused efforts are ongoing to develop vaccines against the most common pathogens affecting our troops. As part of these efforts, diarrhea surveillance was conducted during OBS to monitor pathogens associated with illness and to identify new vaccine targets. A retrospective review was conducted of prior studies with similar methods. Soldiers with diarrhea presenting to the OBS clinic provided a stool sample that was inoculated into Carey-Blair transport media. Within 3 days, the Cary-Blair tubes were transported to the Naval Medical Research Unit no. 3 in Cairo where bacterial culture was performed. As part of the evaluation, 5 Escherichia coli-like colonies were collected and tested for toxin production using the GM1-ELISA. Toxin-positive isolates were further tested for colonization factors (CF) by a dot-blot assay using a standardized panel of monoclonal antibodies against CFA/I, CS1-CS7, CS17, CS8 (CFA/III), CS12 (PCFO159), and CS14 (PCFO166). Enterotoxigenic E. coli (ETEC) was the most frequently isolated pathogen during each OBS from which data were collected. The rate of ETEC-associated diarrhea ranged from 22% to 58%. Over time, there were dramatic shifts in the frequency and distribution of CFs. Over the 5 years of study, an increasing number of ETEC isolates had no known CF identified, and in 2001, only 40% of ETEC was associated with known CFs. The most commonly identified CF was CS6. Diarrheal disease, particularly ETEC, continues to be a common malady among US military personnel deployed to Egypt. We have identified ETEC CF types, especially CS6, which should be considered potential vaccine candidates. However, despite intensive testing, CFs could not be identified in most of the ETEC isolated, highlighting the need for further studies to identify novel CFs and alternative vaccine targets.

Impact of illness and non-combat injury during Operations Iraqi Freedom and Enduring Freedom (Afghanistan).

Historically, non-combat injuries and illnesses have had a significant impact on military missions. We conducted an anonymous cross-sectional survey to assess the prevalence and impact of common ailments among U.S. military personnel deployed to Iraq or Afghanistan during 2003-2004. Among 15,459 persons surveyed, diarrhea (76.8% in Iraq and 54.4% in Afghanistan), respiratory illness (69.1%), non-combat injuries (34.7%), and leishmaniasis (2.1%) were commonly reported. For all causes, 25.2% reported that they required intravenous fluids, 10.4% required hospitalization, and 5.2% required medical evacuation. Among ground units, 12.7% reported that they missed a patrol because of illness, and among air units, 11.7% were grounded because of illness. The incidence of diarrhea and respiratory infections doubled from the pre-combat to combat phases, and the perceived adverse impact of these illnesses on the unit increased significantly during the combat phase. Despite technologic advances in warfare and preventive medicine, illness and non-combat injuries have been common during operations in Iraq and Afghanistan, resulting in frequent transient decreases in operational efficiency.

Identification of a fibronectin-binding domain within the Campylobacter jejuni CadF protein.

The binding of Campylobacter jejuni to fibronectin (Fn), a component of the extracellular matrix, is mediated by a 37 kDa outer membrane protein termed CadF for Campylobacter adhesion to Fn. Previous studies have indicated that C. jejuni binds to Fn on the basolateral surface of T84 human colonic cells. To further characterize the interaction of the CadF protein with Fn, enzyme-linked immunosorbent assays were performed to identify the Fn-binding domain (Fn-BD). Using overlapping 30-mer and 16-mer peptides derived from translated cadF nucleotide sequence, maximal Fn-binding activity was localized to four amino acids (AA 134-137) consisting of the residues phenylalanine-arginine-leucine-serine (FRLS). A mouse alpha-CadF peptide polyclonal antibody (M alpha-CadF peptide pAb) was generated using FRLS containing peptides and found to react with viable C. jejuni as judged by indirect fluorescent microscopy, suggesting that the FRLS residues are surface-exposed. Binding of CadF to purified Fn and INT 407 human epithelial cells was significantly inhibited with peptides containing the Fn-BD. Moreover, a CadF recombinant variant protein, in which the Phe-Arg-Leu residues (CadF AA 134-136) were altered to Ala-Ala-Gly, exhibited a 91% decrease in Fn-binding activity as compared with the wild-type CadF protein. Collectively, these data indicate that the FRLS residues (CadF AA 134-137) of the C. jejuni CadF protein possess Fn-binding activity.

Secretion of virulence proteins from Campylobacter jejuni is dependent on a functional flagellar export apparatus.

Campylobacter jejuni, a gram-negative motile bacterium, secretes a set of proteins termed the Campylobacter invasion antigens (Cia proteins). The purpose of this study was to determine whether the flagellar apparatus serves as the export apparatus for the Cia proteins. Mutations were generated in five genes encoding three structural components of the flagella, the flagellar basal body (flgB and flgC), hook (flgE2), and filament (flaA and flaB) genes, as well as in genes whose products are essential for flagellar protein export (flhB and fliI). While mutations that affected filament assembly were found to be nonmotile (Mot-) and did not secrete Cia proteins (S-), a flaA (flaB+) filament mutant was found to be nonmotile but Cia protein secretion competent (Mot-, S+). Complementation of a flaA flaB double mutant with a shuttle plasmid harboring either the flaA or flaB gene restored Cia protein secretion, suggesting that Cia export requires at least one of the two filament proteins. Infection of INT 407 human intestinal cells with the C. jejuni mutants revealed that maximal invasion of the epithelial cells required motile bacteria that are secretion competent. Collectively, these data suggest that the C. jejuni Cia proteins are secreted from the flagellar export apparatus.

Maximal adherence and invasion of INT 407 cells by Campylobacter jejuni requires the CadF outer-membrane protein and microfilament reorganization.

The binding of Campylobacter jejuni to fibronectin (Fn), a component of the extracellular matrix, is mediated by a 37 kDa outer-membrane protein termed CadF for Campylobacter adhesion to fibronectin. The specificity of C. jejuni binding to Fn, via CadF, was demonstrated using antibodies reactive against Fn and CadF. More specifically, the anti-CadF antibody reduced the binding of two C. jejuni clinical isolates to immobilized Fn by greater than 50 %. Furthermore, a C. jejuni wild-type isolate, in contrast to the isogenic CadF mutant, was found to compete with another C. jejuni wild-type isolate for host cell receptors. Given the relationship between the pericellular Fn matrix and the cytoskeleton, the involvement of host cell cytoskeletal components in C. jejuni internalization was also examined. Cytochalasin D and mycalolide B microfilament depolymerizing agents resulted in a significant reduction in C. jejuni invasion. Studies targeting paxillin, a focal adhesion signalling molecule, identified an increased level of tyrosine phosphorylation upon C. jejuni infection of INT 407 cells. Collectively, these data suggest CadF promotes the binding of C. jejuni to Fn, which in turn stimulates a signal transduction pathway involving paxillin.

Fibronectin-facilitated invasion of T84 eukaryotic cells by Campylobacter jejuni occurs preferentially at the basolateral cell surface.

Previous studies have indicated that the ability to bind to fibronectin is a key feature in successful cell invasion by Campylobacter jejuni. Given the spatial distribution of fibronectin and the architecture of the epithelium, this suggests the possibility that C. jejuni cell invasion might preferentially occur at the basolateral cell surface. To test this hypothesis, we examined the interaction of C. jejuni with T84 human colonic cells. When grown under the appropriate conditions, T84 cells form a polarized cell monolayer. C. jejuni translocation of a T84 cell monolayer appeared to occur via a paracellular (extracellular) route as opposed to a transcellular (intracellular) route based on the finding that a C. jejuni noninvasive mutant translocated as efficiently as its isogenic parent. Additional studies revealed that two distinct C. jejuni wild-type isolates could compete with one another for host cell receptors, whereas a C. jejuni fibronectin-binding-deficient mutant could not compete with a wild-type isolate for host cell receptors. Further, C. jejuni adherence and internalization were significantly inhibited by antifibronectin antibodies but only when cells were first treated with EGTA to expose basolateral cell surfaces. Together, these results support the theory that C. jejuni invasion occurs preferentially at the basolateral surface of eukaryotic cells.