Comparison of microscopic and immunoassay examination in the diagnosis of intestinal protozoa of humans in Mansoura, Egypt.

Protozoal diseases are prevalent globally and especially in developing countries that have relatively lower socioeconomic populations such as Egypt. Direct microscopic examination (DME) is used for the detection and identification of protozoa but lacks sufficient reliability, and thus may be detrimental in obtaining accurate diagnostic or epidemiological data. In this study, we determine the prevalence of infections by Giardia intestinalis, Cryptosporidium sp., and Entamoeba histolytica in humans in Egypt. Furthermore, we determine the reliability of DME in determining infections caused by these protozoa and compare the results to enzyme linked Immunosorbent assays (ELISA). Our results indicate that the prevalence of giardiasis, cryptosporidiosis, and entamoebiasis is 38, 22, and 16 %, respectively. The sensitivity and specificity of DME for detection of G. intestinalis is 45 and 99 %, for Cryptosporidium 66 and 99 %, and for Entamoeba 45 and 100 %, respectively. Our findings demonstrate that ELISA is more reliable for diagnostic and epidemiologic study purposes.

The role of lipooligosaccharide phosphorylcholine in colonization and pathogenesis of Histophilus somni in cattle.

Histophilus somni is a Gram-negative bacterium and member of the Pasteurellaceae that is responsible for respiratory disease and other systemic infections in cattle. One of the bacterium's virulence factors is antigenic phase variation of its lipooligosaccharide (LOS). LOS antigenic variation may occur through variation in composition or structure of glycoses or their substitutions, such as phosphorylcholine (ChoP). However, the role of ChoP in the pathogenesis of H. somni disease has not been established. In Haemophilus influenzae ChoP on the LOS binds to platelet activating factor on epithelial cells, promoting bacterial colonization of the host upper respiratory tract. However, ChoP is not expressed in the blood as it also binds C-reactive protein, resulting in complement activation and killing of the bacteria. In order to simulate the susceptibility of calves with suppressed immunity due to stress or previous infection, calves were challenged with bovine herpes virus-1 or dexamethazone 3 days prior to challenge with H. somni. Following challenge, expression of ChoP on the LOS of 2 different H. somni strains was associated with colonization of the upper respiratory tract. In contrast, lack of ChoP expression was associated with bacteria recovered from systemic sites. Histopathology of cardiac tissue from myocarditis revealed lesions containing bacterial clusters that appeared similar to a biofilm. Furthermore, some respiratory cultures contained substantial numbers of Pasteurella multocida, which were not present on preculture screens. Subsequent biofilm experiments have shown that H. somni and P. multocida grow equally well together in a biofilm, suggesting a commensal relationship may exist between the two species. Our results also showed that ChoP contributed to, but was not required for, adhesion to respiratory epithelial cells. In conclusion, expression of ChoP on H. somni LOS contributed to colonization of the bacteria to the host upper respiratory tract, but phase variable loss of ChoP expression may help the bacteria survive systemically.

Chronic microsporidial enteritis in a missionary from Mozambique.

Microsporidiosis often occurs in immunocompromised persons but may also occur in those who are immunocompetent. Infection by Microsporidia involves a variety of organs and systems, most notably, intestine, lung, kidney, brain, sinuses, muscle, and eyes. Enterocytozoon bieneusi and Encephalitozoon intestinalis are associated with gastroenteritis, and Enterocytozoon hellem and Encephalitozoon cuniculi are associated with keratoconjunctivitis. We report a case of chronic microsporidiosis in a 28-year-old woman missionary from Mozambique who came to our diagnostic laboratory with nausea, lower abdominal pain, and frequent bowel movements. Over two years, the patient was clinically assessed and treated for malaria and giardiasis without laboratory diagnosis while in Mozambique. Identification of the causative agent of her condition was not attempted during the course of her illness in Mozambique. Furthermore, adverse effects of malaria and giardiasis medications may have exacerbated the chronic illness in this patient and mimicked chronic microsporidiosis.

Antioxidant nanoparticles for control of infectious disease.

The new ground being broken by the field of nanotechnology provides us with numerous prospects for treatment and prevention of infectious diseases. Recent reports have demonstrated that several types of nanoparticles act as potent free radical scavengers and antioxidants. Specific nanoconstructs are also reported to have anti-inflammatory activities. Given these properties, the potential application of antioxidant nanoparticles for controlling infectious diseases are discussed in this review. Numerous pathogenic agents establish their virulence and pathogenicity by virtue of their ability to produce free radicals and damage the cells of the immune system. For example, Pseudomonas aeruginosa is a bacterium that produces the toxin pyocyanin, which induces cell damage and compromises the immune system through production of reactive oxygen species (ROS). Nanoparticle antioxidants may provide unique opportunities to counteract the pathogenicity of these types of microorganisms and their formation of biofilms, which are also related to oxygen levels and ROS production. The use of nanoparticles may also play a role in controlling conditions such as ventilation associated pneumonia, where high levels of oxygen induces oxidative stress and inhibits respiratory tract immunity. In contrast, nanoparticle antioxidants, by virtue of their anti-inflammatory activity, may blunt a host's normal immune defenses to certain microorganisms. This review will address this emerging double-edged sword for nanomedicine and its potential role in controlling infectious disease and will address future directions for research in this emerging frontier.

Molecular characterization of phosphorylcholine expression on the lipooligosaccharide of Histophilus somni.

Histophilus somni (Haemophilus somnus) is an important pathogen of cattle that is responsible for respiratory disease, septicemia, and systemic diseases such as thrombotic meningoencephalitis, myocarditis, and abortion. A variety of virulence factors have been identified in H. somni, including compositional and antigenic variation of the lipooligosaccharide (LOS). Phosphorylcholine (ChoP) has been identified as one of the components of H. somni LOS that undergoes antigenic variation. In this study, five genes (lic1ABCD(Hs) and glpQ) with homology to genes responsible for ChoP expression in Haemophilus influenzae LOS were identified in the H. somni genome. An H. somni open reading frame (ORF) with homology to H. influenzae lic1A (lic1A(Hi)) contained a variable number of tandem repeats (VNTR). However, whereas the tetranucleotide repeat 5'-CAAT-3' is present in lic1A(Hi), the VNTR in H. somni lic1A (lic1A(Hs)) consisted of 5'-AACC-3'. Due to the propensity of VNTR to vary during replication and cause the ORF to shift in and out of frame with the upstream start codon, the VNTR were deleted from lic1A(Hs) to maintain the gene constitutively on. This construct was cloned into Escherichia coli, and functional enzyme assays confirmed that lic1A(Hs) encoded a choline kinase, and that the VNTR were not required for expression of a functional gene product. Variation in the number of VNTR in lic1A(Hs) correlated with antigenic variation of ChoP expression in H. somni strain 124P. However, antigenic variation of ChoP expression in strain 738 predominately occurred through variable extension/truncation of the LOS outer core. These results indicated that the lic1(Hs) genes controlled expression of ChoP on the LOS, but that in H. somni there are two potential mechanisms that account for antigenic variation of ChoP.

Construction of a high-efficiency shuttle vector for Histophilus somni.

The genetic manipulation of Histophilus somni is limited due to its high-fidelity restriction-modification system. The broad host-range shuttle plasmid pLS88 is capable of transforming some strains of H. somni, but is an inefficient vector. We have constructed an improved version of pLS88, pNS3K, that transforms H. somni strain 2336 100-fold more efficiently than its predecessor. The transformation efficiency was further increased when pNS3K was isolated from H. somni and retransformed into the same strain. As proof of principle, the lipooligosaccharide biosynthesis gene lob-2A was cloned into pNS3K and expressed in H. somni strain 129Pt, which lacks this gene. Thus, pNS3K is a useful shuttle vector for H. somni and a potential vector for genetic manipulation of this bacterium.

Expression of phosphorylcholine by Histophilus somni induces bovine platelet aggregation.

Histophilus somni-induced platelet aggregation was inhibited by antagonists of the platelet-activating factor (PAF) receptor but not inhibitors of PAF synthesis. In addition, H. somni cells expressing phosphorylcholine (ChoP) induced aggregation, while ChoP(-) H. somni cells did not. This suggests that H. somni ChoP may induce platelet aggregation via interactions with the PAF receptor.