Major Portal Of Entry Research Articles

The pulmonary effects of nickel-containing nanoparticles: Cytotoxicity, genotoxicity, carcinogenicity, and their underlying mechanisms.

With the exponential growth of the nanotechnology field, the global nanotechnology market is on an upward track with fast-growing jobs. Nickel (Ni)-containing nanoparticles (NPs), an important class of transition metal nanoparticles, have been extensively used in industrial and biomedical fields due to their unique nanostructural, physical, and chemical properties. Millions of people have been/are going to be exposed to Ni-containing NPs in occupational and non-occupational settings. Therefore, there are increasing concerns over the hazardous effects of Ni-containing NPs on health and the environment. The respiratory tract is a major portal of entry for Ni-containing NPs; thus, the adverse effects of Ni-containing NPs on the respiratory system, especially the lungs, have been a focus of scientific study. This review summarized previous studies, published before December 1, 2023, on cytotoxic, genotoxic, and carcinogenic effects of Ni-containing NPs on humans, lung cells in vitro, and rodent lungs in vivo, and the potential underlying mechanisms were also included. In addition, whether these adverse effects were induced by NPs themselves or Ni ions released from the NPs was also discussed. The extra-pulmonary effects of Ni-containing NPs were briefly mentioned. This review will provide us with a comprehensive view of the pulmonary effects of Ni-containing NPs and their underlying mechanisms, which will shed light on our future studies, including the urgency and necessity to produce engineering Ni-containing NPs with controlled and reduced toxicity, and also provide the scientific basis for developing nanoparticle exposure limits and policies.

In Vivo Imaging Sheds Light on the Susceptibility and Permissivity of Carassius auratus to Cyprinid Herpesvirus 2 According to Developmental Stage.

Cyprinid herpesvirus 2 (CyHV-2) is a virus that causes mass mortality in economically important Carassius spp. However, there have been no comprehensive studies into host susceptibility or permissivity with respect to developmental stage, and the major portal of viral entry into the host is still unclear. To help bridge these knowledge gaps, we developed the first ever recombinant strain of CyHV-2 expressing bioluminescent and fluorescent reporter genes. Infection of Carassius auratus hosts with this recombinant by immersion facilitated the exploitation of various in vivo imaging techniques to establish the spatiotemporal aspects of CyHV-2 replication at larval, juvenile, and adult developmental stages. While less susceptible than later developmental stages, larvae were most permissive to CyHV-2 replication, leading to rapid systemic infection and high mortality. Permissivity to CyHV-2 decreased with advancing development, with adults being the least permissive and, thus, also exhibiting the least mortality. Across all developmental stages, the skin was the most susceptible and permissive organ to infection at the earliest sampling points post-infection, indicating that it represents the major portal of entry into these hosts. Collectively these findings provide important fundamental insights into CyHV-2 pathogenesis and epidemiology in Carassius auratus with high relevance to other related economically important virus-host models.

Th17-driven immunity to oral candidiasis is dependent on the microbiome and can be triggered by mono-colonization with segmented filamentous bacteria.

Abstract The opportunistic infection oropharyngeal candidiasis (OPC; oral thrush) is a common oral infection among the immunocompromised. The oral cavity is a major entry portal to the body and oral health impacts several aspects of host immunity. While much is now known about microbiome dependent Th17 induction and protection against intestinal candidiasis and other Th17 driven responses, our understanding of the role of the microbiome in oral Th17 induction and oral antifungal immunity is rather rudimentary. We demonstrate a direct role for the microbiome in triggering protection against OPC. While wildtype (WT) mice raised under specific pathogen free (SPF) conditions are resistant to OPC, WT germ free (GF) mice succumbed to infection. Reconstitution of WT-GF mice with segmented filamentous bacteria (SFB) was sufficient to restore protection against OPC with associated increase in Th17 cells, neutrophils, and antimicrobial responses in the oral mucosa. Thus, SFB mono-colonization in WT-GF mice triggers oral mucosal Th17 induction to protect against oral thrush and reveals a role of the microbiome in promoting oral mucosal immunity. Supported by R37-DE022550

Remote effects and biodistribution of pulmonary instilled silver nanoparticles in mice

Silver nanoparticles (AgNPs) are the most commonly used nanoparticles (NPs) owing to their anti-microbial properties, and the pulmonary system provides a major portal of entry for these NPs used in aerosolized products. AgNPs have the potential to cause pulmonary toxicity, cross the alveolar-capillary barrier, and distribute to remote organs following pulmonary exposure. The mechanism underlying the effects of AgNPs, secondary to lung exposure, on the major organs including liver, spleen, kidney and brain, however, is still not completely understood. The aim of this study was to analyze the organ toxicity and distribution of pulmonary exposure to single dose of 5 mg/kg AgNPs (10 nm) with varying coatings (polyvinylpyrrolidone and citrate), at different time points (1 and 7 days), in Balb/C mice. Silver ions (Ag+) were used as ionic control. Histological evidence of inflammation was observed in lungs for both types of AgNPs. Markers of inflammation including tumor necrosis factor α (TNF-α) and interleukin 6 (IL-6) were significantly increased in lung, brain and liver in AgNPs exposed animals. Ag+ ions caused significant increase of TNF-α and IL-6 in the spleen and kidney. Significant increase of reduced glutathione, nitric oxide, and 8-isoprostane was observed in most of the organs investigated. Furthermore, AgNPs induced DNA damage and apoptosis in the lung, liver and brain. The biodistribution showed that, AgNPs were distributed mainly in the spleen, liver, lung and little in kidney and brain. Comparatively, reduced amount of Ag was detected in most organs 7 days after exposure, except for AgAc in the kidney and brain. In conclusion, pulmonary exposure to AgNPs caused oxidative stress markers, inflammation, DNA damage and biodistribution in remote organs. These findings provide a novel mechanistic insight into the pathophysiological effects and tissue distribution of lung exposure to AgNPs.

From Initiators to Effectors: Roadmap Through the Intestine During Encounter of Toxoplasma gondii With the Mucosal Immune System.

The gastrointestinal tract is a major portal of entry for many pathogens, including the protozoan parasite Toxoplasma gondii. Billions of people worldwide have acquired T. gondii at some point in their life, and for the vast majority this has led to latent infection in the central nervous system. The first line of host defense against Toxoplasma is located within the intestinal mucosa. Appropriate coordination of responses by the intestinal epithelium, intraepithelial lymphocytes, and lamina propria cells results in an inflammatory response that controls acute infection. Under some conditions, infection elicits bacterial dysbiosis and immune-mediated tissue damage in the intestine. Here, we discuss the complex interactions between the microbiota, the epithelium, as well as innate and adaptive immune cells in the intestinal mucosa that induce protective immunity, and that sometimes switch to inflammatory pathology as T. gondii encounters tissues of the gut.

HOVERING THREAT OF CORONAVIRUS (COVID-19) AROUND PAKISTAN

January 2020 observed the onset of a new epidemic in Wuhan city of China where hundreds of people were noticed to be suffering from pneumonia like Severe acute respiratory syndrome (SARS) which later on termed as New Corona Virus Disease 2019 (COVID-19) by World Health Organization (WHO).1 The outbreak in China shook the World as many countries planned on restricting international travel and closing businesses momentarily.2 The outbreak of Covid-19, nevertheless, infected every country of the world around 20 million cases worldwide and about 0.7 million confirmed deaths with approximately 19% mortality rate and WHO declared it a pandemic on March 11, 2020.3 Pakistan is the world's fifth-most-populous country with 207,774,520 human population. The rising whirl of Covid-19 hit Pakistan following the end of February 2020. Experts predicted the trade operations between Pakistan and China, the students and businesspersons (who frequently cross China-Pakistan border for academic and commercial purposes) to be the vulnerable portal of Covid-19 in Pakistan. Nevertheless, the Shia pilgrims visiting Iran, where immense religious congregations led to untold Covid-19 surge, were the major entry portal of Covid-19 in Pakistan and officials never reported any Covid-19 case of Chinese transmission in Pakistan. National Command and Control Centre reported around more than 280  thousand Covid-19 cases with 6,139 deaths on August 13, 2020 overall.4 The Covid-19 is on daily exponential rise in Pakistan and the limited resources, lack of equipment and human resource, unavailability of diagnostic kits and medicines have deteriorated the situation. Pakistan does not have an effective infection disease surveillance system to meet the millennium developmental goals in healthcare. Pakistan is struggling to control diseases such as hepatitis, diarrhea, respiratory diseases, typhoid, measles and many mosquito-borne diseases thereof.5 The present outbreak of Covid-19 has affected the whole world although its incidence is low in Pakistan which is continuously struggling to deal with multiple socio-economic factors including poverty, overpopulation, low literacy about common health and health care system issues natural disasters and lack of decisive precautionary approaches. Currently, there is no cure or vaccine available for the virus and therefore “prevention is better than cure”. The preventive measures including lockdown strategies will further add economic crisis to the government and people. The dual frontiers of pandemic and economy could not be combated simultaneously; therefore, the government itself is trying to make a balanced strategy, which could minimally affect the lives of people on both issues. The international funding agencies should take serious notice of this pandemic and its rise in poor country like Pakistan with 220 million populations and come forward to assist Pakistan in fighting out Covid-19 as they are contributing in the eradication of other disease such as tuberculosis, human immunodeficiency virus infection and malaria. The smart lockdown proved to be the best strategies to combat this pandemic and have produced significant results in not only reducing the spread of disease but also running the wheel of economy. The government of Pakistan is further planning to narrow the margin of lockdown to micro smart lockdown to boost up economic activities and eradicate the load of Covid-19.

Pathogenesis and Possible Drug Targets for Covid-19

Coronavirus (CoVs) is a large family of enveloped, single-stranded, positive-sense RNA viruses that infect a wide range of vertebrates. They are extensively found in bats and also in many other birds and mammals including humans. SARS-CoV-2 is a global pandemic and originated from Wuhan States of China. The SARS-CoV-2 is more genetically similar to zoonotic SARS-CoV and less similar to MERS-CoV. The viral surface spike protein of SARS-CoV-2 binds to the human angiotensin-converting enzyme-2 (ACE-2) receptor of Type II alveolar cells of the lungs and it appears to be the major portal of entry by this virus. The subsequent activation of the spike protein by transmembrane protease-2 and in addition to lung, ACE-2 is highly expressed in heart followed by kidney and intestinal epithelium. SARS-CoV-2 infects more men than women due to ACE-2 receptor on the cells increased with age and generally it was higher in men than in women. The incubation period for this virus varies from place to place and asystematic symptoms are also commonly seen in infected patients. There are a number of pharmaceuticals already being tried and are in different phase levels of testing, but a better understanding of the underlying pathobiology is required. In this circumstance, this article will briefly review the underlying principle for ACE-2 receptor as a specific target. Despite ACE-2 serving as the portal for infection, the role of ACE inhibitors or angiotensin receptor blockers requires further investigation.

The Contribution of Pulmonary Dendritic Cells to the Induction and Regulation of T Cell Responses During Respiratory Infection

Abstract The respiratory tract represents a major portal of entry for pathogens, including Group A Streptococci (GAS). GAS represents a significant burden in cases of primary infection as well as cases of secondary bacterial pneumonia following influenza virus infection. Control of infection and eventual resolution is dependent on the local adaptive immune response. This is facilitated by the uptake and subsequent presentation of antigen by dendritic cells (DC) to T cells. Furthermore, after GAS infection, DC drive T cell development towards a Th17 immune response that is associated with pathogen control. Our hypothesis is that pulmonary DC, both resident and recruited, contribute locally to the maintenance and regulation of an effector CD4 T cell response in the context of a GAS infection. Toward this end, our studies are defining the contribution of local DC: T cell interactions in shaping the phenotype and magnitude of Th17 and Tfh responses within the lungs during GAS infection. An improved understanding of the contribution of these DC to the development and regulation of these CD4 T cells responses during GAS infection should elaborate strategies to the development of better therapies.

Staphylococcus aureus Bloodstream Infection in Patients with Atopic Dermatitis, or: Think Twice Before Placing a Venous Catheter into Lesional Atopic Skin

summary Due to the high rate of colonization, patients with atopic dermatitis are at risk of S. aureus bloodstream infection, the most severe manifestation of S. aureus infections. Intravascular devices and the skin are the major portals of entry for S. aureus in AD. With prompt and adequate diagnostic and therapeutic management, mortality is lower than in Non-AD patients with SAB and severe sequelae can be averted. Important preventive strategies include AD treatment to reduce S. aureus colonization and meticulous care of intravascular catheters, which should not be placed in/on lesional atopic skin.

The environment is a first order issue for lung health

In this issue of the Journal we publish the first of a series of State of the Art articles on environmental health. This is a topic that is front and centre for those concerned about lung health.Not only are the airways and lungs the major portal of entry for airborne pollutants, they are also a major site of disease caused by air pollutants, including asthma-like syndromes, chronic obstructive pulmo-nary disease, pneumonitis, pneumoconiosis and lungmalignancy. Of all the environmental causes of ill-health, air pollutants are, by far, the major contributor to death and burden of disease. Furthermore, the burden of this exposure falls particularly on those living in low- and middle-income countries, where the capacity to reduce exposure and mitigate the effects is weakest. This series is thus a timely reminder of the importance of the environment to lung health

Silica nanoparticles disrupt OPT-2/PEP-2-dependent trafficking of nutrient peptides in the intestinal epithelium

Despite of the increasing application of silica nanoparticles and identification of oral exposure as a major entry portal, we lack understanding of nanosilica effects in the gut. Thus, we investigated biointeractions of nanosilica with single intestinal cells. The invertebrate nematode Caenorhabditis elegans was chosen as model organism with a tractable intestine and realistic target organism of nanomaterials in the environment. We found that nanosilica impairs the intestinal uptake of oligopeptides. Downstream to absorption by the apical OPT-2/PEP-2 transporter dipeptides were trapped in aberrant vesicles that grow over time and reach diameters of ≥6 μm. The peptide vesicles do not correspond to known organelles such as gut granules and form independently of related gene products GLO-1 or GLO-3. Formation of aberrant peptide vesicles also occurred independently of insulin/IGF-I receptor (DAF-2) signaling and daf-2 loss of function mutants showed specific vesicle patterns including distinct localization along the apical membrane of single intestinal cells. As malnutrition of exposed C. elegans manifested in reduced growth and a petite phenotype similar to OPT-2/PEP-2 transporter deficient mutants, we conclude that nanosilica-induced peptide vesicles represent a new compartment of di- and tripeptide trapping which disrupts hydrolysis of nutrient peptides and metabolism.

A survey of fungal microbiota in airways of healthy volunteer subjects from Puglia (Apulia), Italy

BackgroundThe human respiratory tract represents the major portal of entry for numerous microorganisms, primarily those occurring as airborne particles such as viral and bacterial entities, or fungal spores. Microorganism characteristics coupled with the local host immune response will determine whether they will be cleared or adhere and colonize the airways leading to acute or chronic pulmonary disease. Like bacteria, fungi can cause severe lung diseases, but their infection rates are much lower. The lung microbiota is commonly sampled using relatively invasive bronchoscopic procedures. Exhaled breath condensate (EBC) collection offers a potentially less invasive alternative for lung microbiota sampling. This study tries to determine the composition of fungal communities in a cohort of healthy adult volunteer subjects from Puglia (Apulia), Italy.MethodsFungi diversity in 27 EBC samples collected from Italian adult volunteers was investigated using conventional microbiological culturing and DNA sequencing approach.ResultsTen tested subjects (37,03%) turned out to present fungi in the EBC. We observed complex fungal communities, in which more than 10% of the isolated species are represented by Aspergillus sydowii (14,8%) and Cladosporium spp (11,11%). Three subjects that showed fungal presence in EBC have been diagnosed with a respiratory disease.ConclusionsWe present a survey of an important scientific field in its early stages that is fungal contamination of airways of healthy subjects in a small geographic area. Furthermore, we interpreted our results to highlight the potential role of fungi in the context of respiratory diseases.

Development, Homeostasis, and Functions of Intestinal Intraepithelial Lymphocytes.

The intestine is continuously exposed to commensal microorganisms, food, and environmental agents and also serves as a major portal of entry for many pathogens. A critical defense mechanism against microbial invasion in the intestine is the single layer of epithelial cells that separates the gut lumen from the underlying tissues. The barrier function of the intestinal epithelium is supported by cells and soluble factors of the intestinal immune system. Chief among them are intestinal intraepithelial lymphocytes (iIELs), which are embedded in the intestinal epithelium and represent one of the single largest populations of lymphocytes in the body. Compared with lymphocytes in other parts of the body, iIELs exhibit unique phenotypic, developmental, and functional properties that reflect their key roles in maintaining the intestinal epithelial barrier. In this article, we review the biology of iIELs in supporting normal health and how their dysregulation can contribute to disease.

Effect of Antifungal Treatment in a Diet-Based Murine Model of Disseminated Candidiasis Acquired via the Gastrointestinal Tract.

Candida albicans, normally found as a commensal in the gut, is a major human fungal pathogen responsible for both mucosal and systemic infections in a wide variety of immunocompromised individuals, including cancer patients and organ transplant recipients. The gastrointestinal tract represents a major portal of entry for the establishment of disseminated candidiasis in many of these individuals. Here we report the development of a diet-based mouse model for disseminated candidiasis acquired via the gastrointestinal tract. Using this model, as well as an appropriate immunosuppression regimen, we demonstrate that dissemination of C. albicans from the gastrointestinal tract can result in mortality within 30 days postinfection. We also show a significant increase in fungal burden in systemic organs, but not gastrointestinal tract organs, upon immunosuppression. Importantly, we demonstrate that the administration of two widely used antifungals, fluconazole and caspofungin, either pre- or postimmunosuppression, significantly reduces fungal burdens. This model should prove to be of significant value for testing the ability of both established and experimental therapeutics to inhibit C. albicans dissemination from the gastrointestinal tract in an immunocompromised host as well as the subsequent mortality that can result from disseminated candidiasis.

Ultra-deep and quantitative saliva proteome reveals dynamics of the oral microbiome.

BackgroundThe oral cavity is home to one of the most diverse microbial communities of the human body and a major entry portal for pathogens. Its homeostasis is maintained by saliva, which fulfills key functions including lubrication of food, pre-digestion, and bacterial defense. Consequently, disruptions in saliva secretion and changes in the oral microbiome contribute to conditions such as tooth decay and respiratory tract infections. Here we set out to quantitatively map the saliva proteome in great depth with a rapid and in-depth mass spectrometry-based proteomics workflow.MethodsWe used recent improvements in mass spectrometry (MS)-based proteomics to develop a rapid workflow for mapping the saliva proteome quantitatively and at great depth. Standard clinical cotton swabs were used to collect saliva form eight healthy individuals at two different time points, allowing us to study inter-individual differences and interday changes of the saliva proteome. To accurately identify microbial proteins, we developed a method called “split by taxonomy id” that prevents peptides shared by humans and bacteria or between different bacterial phyla to contribute to protein identification.ResultsMicrogram protein amounts retrieved from cotton swabs resulted in more than 3700 quantified human proteins in 100-min gradients or 5500 proteins after simple fractionation. Remarkably, our measurements also quantified more than 2000 microbial proteins from 50 bacterial genera. Co-analysis of the proteomics results with next-generation sequencing data from the Human Microbiome Project as well as a comparison to MALDI-TOF mass spectrometry on microbial cultures revealed strong agreement. The oral microbiome differs between individuals and changes drastically upon eating and tooth brushing.ConclusionRapid shotgun and robust technology can now simultaneously characterize the human and microbiome contributions to the proteome of a body fluid and is therefore a valuable complement to genomic studies. This opens new frontiers for the study of host–pathogen interactions and clinical saliva diagnostics.Electronic supplementary materialThe online version of this article (doi:10.1186/s13073-016-0293-0) contains supplementary material, which is available to authorized users.

Glycosaminoglycans and infection.

Glycosaminoglycans (GAGs) are complex linear polysaccharides expressed in intracellular compartments, at the cell surface, and in the extracellular environment where they interact with various molecules to regulate many cellular processes implicated in health and disease. Subversion of GAGs is a pathogenic strategy shared by a wide variety of microbial pathogens, including viruses, bacteria, parasites, and fungi. Pathogens use GAGs at virtually every major portals of entry to promote their attachment and invasion of host cells, movement from one cell to another, and to protect themselves from immune attack. Pathogens co-opt fundamental activities of GAGs to accomplish these tasks. This ingenious strategy to subvert essential activities of GAGs likely prevented host organisms from deleting or inactivating these mechanisms during their evolution. The goal of this review is to provide a mechanistic overview of our current understanding of how microbes subvert GAGs at major steps of pathogenesis, using select GAG-pathogen interactions as representative examples.

An innate antiviral pathway acting before interferons at epithelial surfaces.

Mucosal surfaces are exposed to environmental substances and represent a major portal of entry for microorganisms. The innate immune system is responsible for early defense against infections and it is believed that the interferons (IFNs) constitute the first line of defense against viruses. Here we identify an innate antiviral pathway that works at epithelial surfaces before the IFNs. The pathway is activated independently of known innate sensors of viral infections through a mechanism dependent on viral O-linked glycans, which induce CXCR3 chemokines and stimulate antiviral activity in a manner dependent on neutrophils. This study therefore identifies a previously unknown layer of antiviral defense that exerts its action on epithelial surfaces before the classical IFN response is operative.

Directing vaccine immune responses to mucosa by nanosized particulate carriers encapsulating NOD ligands

Mucosal surfaces are a major portal of entry for many pathogens that are the cause of infectious diseases. Therefore, effective vaccines that induce a protective immune response at these sites are much needed. However, despite early success with the live attenuated oral polio vaccine over 50 years ago, only a few new mucosal vaccines have been subsequently licensed. Development of new adjuvants, comprising antigen delivery platforms and immunostimulatory molecules, are critical for the successful development of new mucosal vaccines. Among them, biodegradable nanoparticle delivery systems are promising and NOD-like receptors are considered as potential new targets for immunostimulatory molecules.In this work, different NOD1 and NOD2 ligands were encapsulated in polylactic acid (PLA) nanoparticles, coated with HIV-1 gag p24 antigen. We showed that these new formulations are able to induce proliferation of HIV-specific T cells from HIV+ individuals as well as autophagy. In vivo, these formulations highly enhanced p24-specific systemic and mucosal immune responses in mice not only after mucosal administration but also after immunization via the parenteral route.Our results provide a rational approach for combining nanosized particulate carriers and encapsulated NOD receptor ligands as potent synergistic tools for induction of specific mucosal immunity.

Distinct Murine Mucosal Langerhans Cell Subsets Develop from Pre-dendritic Cells and Monocytes

Langerhans cells (LCs) populate the mucosal epithelium, a major entry portal for pathogens, yet their ontogeny remains unclear. We found that, in contrast to skin LCs originating from self-renewing radioresistant embryonic precursors, oral mucosal LCs derive from circulating radiosensitive precursors. Mucosal LCs can be segregated into CD103(+)CD11b(lo) (CD103(+)) and CD11b(+)CD103(-) (CD11b(+)) subsets. We further demonstrated that similar to non-lymphoid dendritic cells (DCs), CD103(+) LCs originate from pre-DCs, whereas CD11b(+) LCs differentiate from both pre-DCs and monocytic precursors. Despite this ontogenetic discrepancy between skin and mucosal LCs, the transcriptomic signature and immunological function of oral LCs highly resemble those of skin LCs but not DCs. These findings, along with the epithelial position, morphology, and expression of the LC-associated phenotype strongly suggest that oral mucosal LCs are genuine LCs. Collectively, in a tissue-dependent manner, murine LCs differentiate from at least three distinct precursors (embryonic, pre-DC, and monocytic) in steady state.

Influence of Nevirapine on Gastrointestinal Function

In recent times there has been a growing research interest in Nevirapine an antiretroviral medication that prevents human immunodeficiency virus cells from multiplying in the blood. Nevirapine comes as tablet and a suspension (liquid) often taken by mouth, it is taken with or without once a day for 2 weeks and twice a day after the first 2 weeks. It is best to swallow nevirapine with liquids such as water, milk or soda. Feeding experiments in various animal species and humans have highlighted the beneficial role of nevirapine on health. These benefits include significant reduction in acquired immune deficiency syndrome-related morbidity and mortality. However, nevirapine hypersensitivity reaction can lead to morbidity through treatment interruption, inconvenience and loss of productivity. Additionally, nevirapine has an excellent bioavailability and a long half-life. This increases the risk of non-nucleoside reverse transcriptase inhibitor resistance when patients discontinue all the other antiretroviral drugs in the regimen at the same time. Thus, this can lead to decreased therapeutic options if nevirapine resistance develops. The gastrointestinal tract is a major portal of entry of nutrients into the body and nevirapine first make contact with the gastrointestinal tract. The health of an individual is dependent on the nutrient absorbed and hence functional integrity of the gastrointestinal tract. Studies have revealed that the interaction with drug have led to disruption of gastrointestinal function.