Future perspectives on infections associated with gastrointestinal tract diseases.

There are a vast number of infectious agents that are associated with gastrointestinal (GI) tract diseases. The epidemiology of GI diseases is changing, with a greater number of conditions increasing in incidence. Challenges exist with establishing cause-and-effect relationships because of the ubiquitous nature of these organisms and the milieu in which they exist. Advances in technology should provide novel methods for identifying and diagnosing these organisms and the relationship they have with a specific digestive disease. Crown

Changing burden of gastrointestinal disease

In 2004, in the United States, there were 72 million presentations with a primary diagnosis of a digestive disease and 104 million presentations with combined gastrointestinal (GI) tract diseases and other diseases (Table 1 ). It was also found that those who are older tend to have more GI problems, there was no difference in the rates of digestive disease between the African Americans and the whites, and women were 20% more likely to present than men with digestive diseases. Thus, more than one-third (35%) of all presentations are for digestive diseases. In 2009, in the United States, the cancer statistics revealed 275,720 new cases of GI cancer, with colorectal and pancreatic cancer in the top 10 for both men and women. There were 135,830 deaths due to GI cancer, with colorectal, pancreatic, hepatic, and esophageal cancers in the top 10 for both men and women, except for esophageal cancer, which was only listed for men (Fig. 1 ). Furthermore, 2 of these 3 cancers that have an increasing mortality were GI tract cancers for both genders, with esophageal and hepatic cancers among men and pancreatic and hepatic cancers among women. Worldwide the rates of digestive diseases are staggering.

Table 1

Burden of selected digestive diseases in the United States, 2004

Digestive Disease	Deaths, Underlying Causea	Years of Potential Life Lost to Age 75 Yearsa	Ambulatory Care Visits, All-Listed Diagnosesb	Hospital Discharges, All-Listed Diagnosesc
All Digestive Diseases	236,164	2,007,500	104,790,000	13,533,000
All Digestive Cancers	135,107	945,200	4,198,000	726,000
Colorectal Cancer	53,226	333,000	2,589,000	255,000
Pancreatic Cancer	31,800	206,800	415,000	68,000
Esophageal Cancer	13,667	113,800	372,000	44,000
Gastric Cancer	11,253	84,200	141,000	31,000
Primary Liver Cancer	6323	72,400	63,000	33,000
Bile Duct Cancer	4954	32,900	—	17,000
Gall Bladder Cancer	1939	10,900	—	6000
Cancer of the Small Intestine	1115	9300	—	9000
Liver Disease	36,090	559,100	2,398,000	759,000
All Viral Hepatitis	5393	101,800	3,510,000	475,000
Hepatitis C	4595	87,500	2,747,000	419,000
Hepatitis B	645	11,800	729,000	69,000
Hepatitis A	58	800	—	10,000
GI Infections	4396	12,800	2,365,000	450,000
Peptic Ulcer Disease	3692	19,700	1,473,000	489,000
Pancreatitis	3480	42,800	881,000	454,000
Diverticular Disease	3372	8600	3,269,000	815,000
Abdominal Wall Hernia	1172	6900	4,787,000	372,000
Gastroesophageal Reflux Disease	1150	6000	18,342,000	3,189,000
Gallstones	1092	4400	1,836,000	622,000
All Inflammatory Bowel Disease	933	9100	1,892,000	221,000
Crohn Disease	622	7000	1,176,000	141,000
Ulcerative Colitis	311	2000	716,000	82,000
Appendicitis	453	5000	782,000	325,000
All Functional Intestinal Disorders	423	2500	11,648,000	1,241,000
Chronic Constipation	137	900	6,306,000	700,000
Irritable Bowel Syndrome	20	0	3,054,000	212,000
Hemorrhoids	14	200	3,275,000	306,000

Vital statistics of the United States.

The National Ambulatory Medical Care Survey and the National Hospital Ambulatory Medical Care Survey.

The Healthcare Cost and Use Project Nationwide Inpatient Sample.

Fig. 1

The 10 leading cancer types among the estimated new cancer cases and deaths, by sex, in the United States in 2009.

From 1979 to 1989, in the United States, a decrease was observed in the ambulatory care visits and hospital discharges for digestive diseases. These rates remained constant between 1990 and 1999, until 2000 when the rates climbed dramatically and was still increasing in 2004 (Fig. 2 ). During this period, substantial increases in the prevalence were observed for certain GI tract diseases, including gastroesophageal reflux disease (GERD) with an increase of 376 per 100,000 population, hepatitis C with 79 per 100,000 population, chronic constipation with 62 per 100,000 population, intestinal infections with 41 per 100,000 population, and pancreatitis with 23 per 100,000 population.

Fig. 2

For all digestive diseases, age-adjusted rates of ambulatory care visits and hospital discharges with all-listed diagnoses in the United States from 1979 to 2004.

The prevalence of digestive diseases around the world is enormous and varies from country to country (Table 2 ). Worldwide there has been a dynamic shift in the epidemiology of GI tract diseases, with some diseases such as peptic ulcer decreasing dramatically since the discovery of Helicobacter pylori infection and a larger number of conditions increasing, such as GERD, nonalcoholic fatty liver disease, diverticular disease, Barrett esophagus, cholelithiasis, alcoholic liver disease, hepatitis C, chronic pancreatitis, esophageal cancer and colorectal cancer.3, 4, 5, 6, 7, 8 In conjunction with this increasing incidence of digestive diseases are the re-emergence of certain infectious agents (Box 1 ) (eg, cholera) and the identification of new agents (eg, H pylori, Laribacter, Campylobacter concisus), which are associated with GI tract diseases. Since the discovery of H pylori there has been an enormous interest in the relationship between microorganisms and GI tract diseases, including cancers.

Table 2

Estimates of digestive disease burden around the world

Country/Region	Extrapolated Prevalence	Population Estimated Used
Digestive Diseases in North America (Extrapolated Statistics)
United States of America	64,776,924	293,655,405a
Canada	7,170,854	32,507,874b
Digestive Diseases in Europe (Extrapolated Statistics)
Austria	1,803,256	8,174,7622
Belgium	2,282,707	10,348,276b
Britain (United Kingdom)	13,295,008	60,270,708 for UKb
Czech Republic	274,892	10,246,178b
Denmark	1,194,130	5,413,392b
Finland	1,150,259	5,214,512b
France	13,328,869	60,424,213b
Greece	2,348,719	10,647,529b
Germany	18,181,898	82,424,609b
Iceland	64,845	293,966b
Hungary	2,213,023	10,032,375b
Liechtenstein	7375	33,436b
Ireland	875,637	3,969,558b
Italy	12,806,795	58,057,477b
Luxembourg	102,063	462,690b
Monaco	7118	32,270b
Netherlands (Holland)	3,599,602	16,318,199b
Poland	8,520,517	38,626,349b
Portugal	2,321,502	10,524,145b
Spain	8,885,465	40,280,780b
Sweden	1,982,294	8,986,400b
Switzerland	1,643,573	7,450,867b
United Kingdom	13,295,008	60,270,708b
Wales	643,676	2,918,000b
Digestive Diseases in the Balkans (Extrapolated Statistics)
Albania	781,942	3,544,808b
Bosnia and Herzegovina	89,913	407,608b
Croatia	991,956	4,496,869b
Macedonia	450,018	2,040,085b
Serbia and Montenegro	2,388,066	10,825,900b
Digestive Diseases in Asia (Extrapolated Statistics)
Bangladesh	31,178,044	141,340,476b
Bhutan	482,110	2,185,569b
China	286,510,493	1,298,847,624b
East Timor	224,834	1,019,252b
Hong Kong SAR	1,512,159	6,855,125b
India	234,942,036	1,065,070,607b
Indonesia	52,599,913	238,452,952b
Japan	28,088,161	127,333,002b
Laos	1,338,555	6,068,117b
Macau SAR	98,224	445,286b
Malaysia	5,188,782	23,522,482b
Mongolia	606,907	2,751,314b
Philippines	19,023,902	86,241,697b
Papua New Guinea	1,195,649	5,420,280b
Vietnam	18,234,440	82,662,800b
Singapore	960,417	4,353,893b
Pakistan	35,116,837	159,196,336b
North Korea	5,006,812	22,697,553b
South Korea	10,639,799	48,233,760b
Sri Lanka	4,390,845	19,905,165b
Taiwan	5,018,346	22,749,838b
Thailand	14,308,570	64,865,523b
Digestive Diseases in Eastern Europe (Extrapolated Statistics)
Azerbaijan	1,735,673	7,868,385b
Belarus	2,274,379	10,310,520b
Bulgaria	1,658,376	7,517,973b
Estonia	295,955	1,341,664b
Georgia	1,035,417	4,693,892b
Kazakhstan	3,340,522	15,143,704b
Latvia	508,743	2,306,306b
Lithuania	795,860	3,607,899b
Romania	4,931,371	22,355,551b
Russia	31,758,982	143,974,059b
Slovakia	1,196,375	5,423,567b
Slovenia	443,707	2,011,473b
Tajikistan	1,546,666	7,011,556b
Ukraine	10,529,134	47,732,079b
Uzbekistan	5,825,826	26,410,416b
Digestive Diseases in Australasia and Southern Pacific (Extrapolated Statistics)
Australia	4,392,605	19,913,144b
New Zealand	880,989	3,993,817b
Digestive Diseases in the Middle East (Extrapolated Statistics)
Afghanistan	6,289,781	28,513,677b
Egypt	16,790,606	76,117,421b
Gaza Strip	292,277	1,324,991b
Iran	14,890,412	67,503,205b
Iraq	5,597,358	25,374,691b
Israel	1,367,428	6,199,008b
Jordan	1,237,765	5,611,202b
Kuwait	497,988	2,257,549b
Lebanon	833,209	3,777,218b
Libya	1,242,261	5,631,585b
Saudi Arabia	5,690,280	25,795,938b
Syria	3,974,310	18,016,874b
Turkey	15,197,187	68,893,918b
United Arab Emirates	556,745	2,523,915b
West Bank	509,824	2,311,204b
Yemen	4,417,249	20,024,867b
Digestive Diseases in South America (Extrapolated Statistics)
Belize	60,208	272,945b
Brazil	40,610,537	184,101,109b
Chile	3,490,578	15,823,957b
Colombia	9,333,258	42,310,775b
Guatemala	3,150,131	14,280,596b
Mexico	23,152,850	104,959,594b
Nicaragua	1,182,299	5,359,759b
Paraguay	1,365,742	6,191,368b
Peru	6,075,949	27,544,305b
Puerto Rico	859,844	3,897,960b
Venezuela	5,518,541	25,017,387b
Digestive Diseases in Africa (Extrapolated Statistics)
Angola	2,421,739	10,978,552b
Botswana	361,595	1,639,231b
Central African Republic	825,547	3,742,482b
Chad	2,104,090	9,538,544b
Congo Brazzaville	661,332	2,998,040b
Congo Kinshasa	12,864,050	58,317,030b
Ethiopia	15,736,007	71,336,571b
Ghana	4,578,756	20,757,032b
Kenya	7,275,464	32,982,109b
Liberia	747,934	3,390,635b
Niger	2,506,000	11,360,538b
Nigeria	3,915,519	125,750,356b
Rwanda	1,817,354	8,238,673b
Senegal	2,393,855	10,852,147b
Sierra Leone	1,297,916	5,883,889b
Somalia	1,831,897	8,304,601b
Sudan	8,635,623	39,148,162b
South Africa	9,804,809	44,448,470b
Swaziland	257,920	1,169,241b
Tanzania	7,956,793	36,070,799b
Uganda	5,821,380	26,390,258b
Zambia	2,432,137	11,025,690b
Zimbabwe	809,969	12,671,860b

Abbreviation: SAR, special administrative region.

US Census Bureau, population estimates, 2004.

US Census Bureau, international database, 2004.

Acanthamebiasis

Australian bat Lyssavirus

Babesia, atypical

Bartonella henselae

Ehrlichiosis

Encephalitozoon cuniculi

Encephalitozoon hellem

Enterocytozoon bieneusi

H pylori

Hendra or equine morbillivirus

Hepatitis C

Hepatitis E

Human herpesvirus 8

Human herpesvirus 6

Lyme borreliosis

Parvovirus B19

Enterovirus 71

Clostridium difficile

Mumps virus

Streptococcus, group A

Staphylococcus aureus

National Institute of Allergy and Infectious Diseases (NIAID): category A

Bacillus anthracis (anthrax)

Clostridium botulinum toxin (botulism)

Yersinia pestis (plague)

Variola major (smallpox) and other related poxviruses

Francisella tularensis (tularemia)

Viral hemorrhagic fevers

Arenaviruses: lymphocytic choriomeningitis virus, Junin virus, Machupo virus, Guanarito virus, Lassa fever

Bunyaviruses: Hantaviruses, Rift Valley fever, Flaviviruses, dengue virus

Filoviruses: Ebola, Marburg

NIAID: category B

Burkholderia pseudomallei

Coxiella burnetii (Q fever)

Brucella species (brucellosis)

Burkholderia mallei (glanders)

Chlamydia psittaci (psittacosis)

Ricin toxin (from Ricinus communis)

Epsilon toxin of Clostridium perfringens

Staphylococcus enterotoxin B

Typhus fever (Rickettsia prowazekii)

Food- and waterborne pathogens

Diarrheagenic Escherichia coli

Pathogenic vibrios

Shigella species

Salmonella

Listeria monocytogenes

Campylobacter jejuni

Yersinia enterocolitica

Viruses (Caliciviruses, Hepatitis A)

Protozoa: Cryptosporidium parvum, Cyclospora cayetanensis, Giardia lamblia, Entamoeba histolytica, Toxoplasma

Fungi

Microsporidia

Additional viral encephalitides: West Nile virus, La Crosse virus, California encephalitis virus, Venezuelan equine encephalitis virus, Eastern equine encephalitis virus, Western equine encephalitis, Japanese encephalitis virus, Kyasanur forest virus

NIAID: category C

Emerging infectious disease threats such as Nipah virus and additional hantaviruses

NIAID priority areas

Tick-borne hemorrhagic fever viruses: Crimean-Congo hemorrhagic fever virus

Tick-borne encephalitis viruses

Yellow fever

Multidrug-resistant tuberculosis

Influenza

Other rickettsias

Rabies

Prions

Chikungunya virus

Severe acute respiratory syndrome–associated coronavirus

Antimicrobial resistance, excluding research on sexually transmitted organisms

Research on mechanisms of antimicrobial resistance

Studies of the emergence and/or spread of antimicrobial resistance genes within pathogen populations

Studies of the emergence and/or spread of antimicrobial-resistant pathogens in human populations

Research on therapeutic approaches that target resistance mechanisms

Modification of existing antimicrobials to overcome emergent resistance

Antimicrobial research, as related to engineered threats and naturally occurring drug-resistant pathogens, focused on the development of broad-spectrum antimicrobials

Innate immunity, defined as the study of nonadaptive immune mechanisms that recognize, and respond to, microorganisms, microbial products, and antigens

Coccidioides immitis

Coccidioides posadasii

Cause-and-effect issues

One of the main issues associated with infections and disease is determining the relationship of the cause and effect. The landmark article by Sir Austin Bradford Hill in 1965 titled The environment and disease: association or causation? became widely known as the Bradford Hill’s criteria. There were 8 criteria that were required to be met to determine a cause-and-effect relationship (Box 2 ). It is usually difficult to meet all these criteria, particularly when trying to find the cause-and-effect relationships between organisms in the small intestine or colon because of the large number of organisms living in these environments. Even for H pylori infection and the relationship with gastric cancer, although it is currently the only bacterium classified as a class I carcinogen, the evidence supporting this relationship is not complete in terms of Bradford Hill’s criteria.

Consistency: The association is consistent when results are replicated in studies in different settings using different methods.

Strength: This is defined by the size of the risk as measured by appropriate statistical tests.

Specificity: This is established when a single putative cause produces a specific effect.

Dose-response relationship: An increasing level of exposure (in amount and/or time) increases the risk.

Temporal relationship: Exposure always precedes the outcome.

Biologic plausibility: The association agrees with currently accepted understanding of pathobiologic processes. This criterion should be applied with caution.

Coherence: The association should be compatible with existing theory and knowledge.

Experiment: The condition can be altered by an appropriate experimental regimen. Experiment is possibly the most important support for a causal relationship.

Organisms associated with GI tract diseases

There are a large number of organisms believed to be responsible for diseases of the digestive system. Some of these organisms are true pathogens, whereas others are merely commensal in nature and are unlikely to ever produce any pathologic condition. Table 3 shows the various types of microbes that are associated with diseases of the GI tract covered in this issue; it is by no means all-inclusive but provides the current magnitude of an ever-increasing field of research. At present, some of these diseases are only associated with a single group of organisms (eg, irritable bowel syndrome), whereas other diseases are affected by all groups of organisms (eg, appendicitis).

Table 3

Organisms associated with GI tract disease in humans

GI Tract Disease	Bacteria	Virus	Parasite	Fungi
Esophageal Cancer	α-hemolytic streptococcus, β-hemolytic streptococcus, Bacteroides fragilis, Bacteroides melaninogenicus, Bacteroides sp, Clostridium sp, coagulase-negative Staphylococcus, Corynebacterium sp, Escherichia coli, Fusobacteria sp, Haemophilus influenzae, Lactobacillus sp, Neisseria catarrhalis, nonhemolytic streptococcus, Peptococcus, Pneumococcus, Proteus mirabilis, Staphylococcus albus, Staphylococcus aureus, Streptococcus pyogenes, Streptococcus viridans, Candida albicans Mycobacterium avium, Mycobacterium tuberculosis	Cytomegalovirus, Epstein-Barr virus, Herpes simplex virus, Varicella-zoster virus	Cryptosporidium	Histoplasma capsulatum,
Gastric Cancer	H pylori	Epstein-Barr virus	—	—
Cholangiocarcinoma	—	Hepatitis C virus, Hepatitis B virus	Clonorchis sinensis, Opistochus vivarini	—
Gall Bladder Disease	E coli, H pylori, Helicobacter sp, Enterobacteriaceae, Leptospira, Salmonella enteritidis, Salmonella typhi, Staphylococcus aureus, Micrococcus sp	Cytomegalovirus, Epstein-Barr virus, Dengue virus	C sinensis, O vivarini, Ascaris lumbricoides, Dolosigranulum pigrum	Actinomyces sp, Candida sp,
Hepatocellular Carcinoma	—	Hepatitis B virus, Hepatitis C virus	—	—
Acute Pancreatitis	Mycoplasma pneumoniae, S typhi, Leptospira, Yersinia enterocolitica, Yersinia pseudotuberculosis, Campylobacter jejuni, M tuberculosis, M avium, Legionella sp, Brucellosis, Actinomyces, Nocardia	Measles virus, Coxsackie B virus, hepatitis B virus, Cytomegalovirus, herpes simplex virus, varicella virus, human immunodeficiency virus, Epstein-Barr virus, vaccinia, rubella, adenovirus	A lumbricoides, Echinococcus granulosus	Aspergillus sp, Cryptococcus neoformans, Coccidioides immitis, Paracoccidioides brasiliensis, Histoplasma capsulatum, Pneumocystis carinii
Small Intestinal Bacterial Overgrowth	Streptococcus sp, E coli, Staphylococcus sp, Micrococcus sp, Klebsiella sp, Methanobrevibacter smithii, Bacteroides sp, Firmicutes sp	—	—	—
Irritable Bowel Syndrome	Salmonella sp, Campylobacter sp, Shigella sp, Enterobacteriaceae, Clostridia	—	—	—
Inflammatory Bowel Disease	E coli, M avium, Streptococcus sp, Clostridia, Actinobacteria, Proteobacteria, Clostridium leptum, Faecalibacterium prausnitzii, Bacteroides, Fusobacteria	—	—	—
Appendicitis	Y enterocolitica, Y pseudotuberculosis, Actinomyces israelii, Mycobacterium, C jejuni, Clostridium difficile, Salmonella sp, B fragilis	Adenovirus, cytomegalovirus, measles virus (rubeola virus)	A lumbricoides, Enterobius vermicularis, Strongyloides stercoralis, Schistosomiasis haematobium, Entamoeba histolytica, Trichuris sp	Mucormycosis, histoplasma capsulatum
Colorectal Cancer	Helicobacter hepaticus, Enterococcus faecalis, Streptococcus bovis, H pylori, Clostridium septicum, E coli, Streptococcus sanguis, Streptococcus salivarius	Human papillomavirus, JC virus, Epstein-Barr virus, cytomegalovirus	—	—

Future challenges

There are a variety of methodological and technical issues related to infectious agents and their role in digestive diseases. For diseases of the colon, the major limitation remains the inability to completely identify these organisms. Identification of bacteria was mainly conducted using culture-based methods. Now, the focus in identification of bacteria is increasingly based on using molecular techniques. Many of these techniques allow the detection and identification of viable but nonculturable cells that are metabolically active but not reproducing. Gene sequencing using single-stranded RNA has been a key method in being able to elucidate multitudes of organisms that remain unknown. At present, there are approximately 9000 bacterial species, and this number is estimated as just the tip of the iceberg. The development of molecular methods offers great promise not only in research and development but also in the diagnostic setting (eg, stool samples) (Table 4 ).11, 12 Clearly, metagenomics, in which genetic material is directly retrieved from environmental sources, will play a critical role in the future development of determining infectious agents of the GI tract. The use of high-throughput technology has already produced important findings in relation to the GI tract microflora, including the differences between adults and children, with numerous uncultured organisms being the crux of the normal human adult gut flora which remain stable but other organisms change depending on environmental and genetic factors, whereas in infants there appear to be a constant transformation of organisms over time (Figs. 3 and 4 ). There have been several new detection methods developed, with some of these using nanoscale electrochemical detectors and others using DNA sensors (extrachromosomal DNA). The use of stable-isotope probing is also being investigated, but even this technique has limitations. Although these technologies are increasing the understanding of the gut microflora, there remains large gaps of knowledge regarding the metabolic functions of these organisms and the relationship they have with human GI disease. These will be extremely fruitful areas of research and development in the coming years.

Table 4

Automatic nucleic acid extraction methods from bacteria

Instrument	Method	Number of Samples	Time Required
ABI PRISM 6100 nucleic acid PrepStation (Applied Biosystems)	Silica membrane bind/elute protocols with vacuum processing (RNA and DNA)	Up to 96	30 min
ABI PRISM 6700 automated nucleic acid workstation (Applied Biosystems)	Silica membrane bind/elute protocols with vacuum processing (RNA and DNA)	Up to 96	90 min
BioRobot EZ1 workstation (QIAGEN)	Silica membrane bind/elute protocols using magnetic-particle handling (RNA and DNA)	1–6	15–20 min
iPrep Purification Instrument (Invitrogen)	Based on a unique, ionizable nucleic acid-binding ligand whose charge can be switched based on the pH of the surrounding medium (DNA)	Up to 12	18 min
KingFisher ML/96 (Thermo Scientific)	Silica membrane bind/elute protocols using magnetic-particle handling (RNA and DNA)	1–96	20–30 min
MagNA pure compact/LC (Roche Applied Science)	Silica membrane bind/elute protocols using magnetic-particle handling (RNA and DNA)	1–32	15–40 min
Maxwell 16 Instrument (Promega)	Silica membrane bind/elute protocols using magnetic-particle handling (RNA and DNA)	Up to 16	30 min
NucliSens miniMAG (BioMérieux)	Silica membrane bind/elute protocols using magnetic-particle handling (RNA and DNA)	Up to 12	45 min
QIAcube (QIAGEN)	Silica membrane bind/elute protocols with built in centrifuge (RNA and DNA)	Up to 12	60 min, user-developed protocols
X-Tractor Gene RNA/DNA Extraction System (Corbett Life Science)	Silica membrane bind/elute protocols with vacuum processing (RNA and DNA)	8–96	1 h

Fig. 3

High-throughput analysis of human GI tract microbiota via brute force sequencing and phylogenetic microarray analysis. SSU rRNA, small subunit ribosomal RNA.

Fig. 4

Metagenomics and other community-based “omics” approaches. SSU rRNA, small subunit ribosomal RNA.