Diagnosis of Strongyloides stercoralis infection.

Strongyloides stercoralis infects 30 million people in 70 countries. Infection usually results in asymptomatic chronic disease of the gut, which can remain undetected for decades. However, in patients receiving long-term corticosteroid therapy, hyperinfection can occur, resulting in high mortality rates (up to 87%). Strongyloidiasis is difficult to diagnose because the parasite load is low and the larval output is irregular. Results of a single stool examination by use of conventional techniques fail to detect larvae in up to 70% of cases. Several immunodiagnostic assays have been found ineffective in detecting disseminated infections and show extensive cross-reactivity with hookworms, filariae, and schistosomes. Although it is important to detect latent S. stercoralis infections before administering chemotherapy or before the onset of immunosuppression in patients at risk, a specific and sensitive diagnostic test is lacking. This review describes the clinical manifestations of strongyloidiasis, as well as various diagnostic tests and treatment strategies.

Isolation of a cDNA encoding an IgG immunoreactive antigen (zinc finger protein) of Strongyloides stercoralis.

A full length cDNA encoding an IgG immunoreactive antigen of Strongyloides stercoralis is described. A clone containing 1,328 bp insert was selected following screening of S. stercoralis cDNA library with an IgG fraction obtained from a pool of 78 S. stercoralis positive human sera samples. The nucleotide sequence of the 1,328 bp insert was found to be 70.5% A/T, reflecting a characteristic A/T codon bias of S. stercoralis. The nucleotide sequence of this insert identified a cDNA coding for a zinc finger protein. The conceptually translated amino acid sequence of the open reading frame for the IgG immunoreactive antigen of S. stercoralis encodes a 211 amino acid residue protein with an apparent molecular weight of 22.8 kDa and a predicted isoelectric point of 8.71. The diagnostic potential of this IgG immunoreactive antigen of S. stercoralis is also discussed.

Cloning and expression of isocitrate lyase from human round worm Strongyloides stercoralis.

A full length cDNA (1463 bp) encoding isocitrate lyase (EC 4.1.3.1) of Strongyloides stercoralis is described. The nucleotide sequence of this insert identified a cDNA coding for the isocitrate lyase. The conceptually translated amino acid sequence of the open reading frame for S. stercoralis isocitrate lyase encodes a 450 amino acid residue protein with an apparent molecular weight of 50 kDa and a predicted pl of 6.39. The sequence is 69% A/T, reflecting a characteristic A/T codon bias of S. stercoralis. The amino acid sequence of S. stercoralis isocitrate lyase is compared with bifunctional glyoxylate cycle protein of Caenorhabditis elegans and isocitrate lyases from Chlamydomonas reinhardtii and Myxococcus xanthus. The full length cDNA of S. stercoralis was expressed in pRSET vector and bacteriophage T7 promoter based expression system. S. stercoralis lyase recombinant protein, purified via immobilized metal affinity chromatography, showed a molecular mass of 50 kDa on polyacrylamide gels. The role of isocitrate lyase in the glyoxylate cycle and energy metabolism of S. stercoralis is also discussed.

A cDNA encoding the highly immunodominant antigen of Strongyloides stercoralis: gamma-subunit of isocitrate dehydrogenase (NAD+).

A full length cDNA encoding the highly immunodominant 41 kDa antigen of Strongyloides stercoralis (P5), recognized by 83% of human patients [Siddiqui et al. (1997) Parasitol Res 83:655-658], is obtained. A clone containing a 1371 bp insert was selected following screening of the S. stercoralis cDNA library with antibodies specific to antigen P5. The nucleotide sequence of this insert identified a cDNA coding for the gamma-subunit of isocitrate dehydrogenase (NAD+), GenBank Accession Number AF176568. The conceptually translated amino acid sequence of the open reading frame for the gamma-subunit of S. stercoralis isocitrate dehydrogenase (NAD+) encodes a 388 amino acid residue protein with an apparent molecular weight of 43 kDa and a predicted pI of 7.15. The sequence is 71% A/T, reflecting the characteristic A/T codon bias of S. stercoralis. The amino acid sequence of the S. stercoralis gamma-subunit of isocitrate dehydrogenase (NAD+) is compared with those of Caenorhabditis elegans, rat and human NAD(+)-ICDH. The diagnostic potential of the S. stercoralis gamma-subunit of isocitrate dehydrogenase (NAD+) is also discussed.

Long-term trends in susceptibility of Moraxella catarrhalis: a population analysis.

A retrospective, population analysis of antimicrobial susceptibility patterns was performed on Moraxella catarrhalis isolates recovered from a single medical centre to detect temporal trends and infer potential mechanisms of reduced susceptibility. The duration of this study, June 1984 to July 1994, encompassed the period during which the frequency of beta-lactamase production expanded from 30 to 96% in the population. MICs of penicillin G, cefamandole, ceftriaxone, amoxycillin/clavulanate, imipenem, clarithromycin, tetracycline, ciprofloxacin and trimethoprim/sulphamethoxazole for a representative sample of 375 isolates were determined. Analyses were conducted to test for variation in susceptibility among isolates, correlations of susceptibility levels among different antimicrobial agents, and temporal patterns in susceptibility. All antimicrobials except clarithromycin displayed significant differences among isolates within years, and mean MICs of all antimicrobial agents except tetracycline and clarithromycin varied significantly between years. Temporal trends to a reduction in susceptibility were detected to four of five beta-lactam antimicrobials (all except cefamandole). Significant correlations in MICs were uncovered among all pairs of four beta-lactam antimicrobials in both producers and non-producers of beta-lactamase. In contrast, cefamandole MICs were correlated only with ceftriaxone and penicillin, and these were limited to beta-lactam producing isolates; cefamandole and amoxycillin/clavulanate showed a correlation limited to non-producing isolates. For some antimicrobials, trends toward decreasing susceptibility may have been caused by an increased proportion of beta-lactamase producing isolates in the population, but the observation of significant decreases in susceptibility limited to beta-lactamase-producing isolates suggests that the underlying factors were different forms of beta-lactamase, beta-lactamase-dependent modifiers and/or additional factors.

A cDNA encoding a nuclear hormone receptor of the steroid/thyroid hormone-receptor superfamily from the human parasitic nematode Strongyloides stercoralis.

A cDNA encoding a nuclear hormone receptor of the steroid/thyroid receptor superfamily was obtained from third-stage larvae(L3) of the parasitic roundworm Strongyloides stercoralis. A recombinant clone was isolated via screening of an S. stercoralis cDNA library with a polymerase chain reaction (PCR)-generated probe. The insert of 2,583 bp contained the complete coding sequence of the receptor homologue. The conceptually translated amino acid sequence of this open reading frame encodes a 753-amino-acid-residue protein with an apparent molecular weight of 83.6 kDa and a predicted isoelectric point (pI) of 8.52. The coding sequence is 69% AT and the noncoding sequence is 72% AT, reflecting a characteristic A/T codon bias of S. stercoralis. In this report the amino acid sequence of the S. stercoralis nuclear hormone receptor of the steroid/thyroid receptor superfamily is compared with that of nuclear hormones of Caenorhabditis elegans, human orphan nuclear receptors, and insect ecdysone receptors. The potential role of steroids in the induction of hyperinfection syndrome is also discussed.

Oral use of interferon-alpha stimulates ISG-15 transcription and production by human buccal epithelial cells.

ISG-15 is a 15-kDa protein encoded by an interferon (IFN)-stimulated gene (ISG), which is transcriptionally regulated by IFN-alpha and IFN-beta. Considered as part of the cytokine network, ISG-15 has the potential to amplify the immunomodulatory effects of these IFNs by enhancing IFN-gamma production, natural killer cell proliferation, and lymphokine-alphactivated killer cell cytotoxicity. To understand better the mechanism(s) of action of orally administered IFN-alpha, we have studied the effect of IFN-alpha on ISG-15 gene expression by human buccal epithelial cells (BEC). For in vitro studies, ISG-15 mRNA and protein levels were measured in BEC incubated for 0.5, 2, and 9 h with 100 or 1,000 IU/ml of human lymphoblastoid IFN-alpha. For in vivo studies, ISG-15 mRNA was measured in BEC samples collected at baseline, and 0.5, 2, and 9 h after 5-20 min of oral rinsing with 10 ml of IFN-alpha (1,000 IU/ml). ISG-15 mRNA was measured by reverse transcriptase polymerase chain reaction (RT-PCR), and ISG-15 protein production by Western Blot analysis. IFN-alpha augmented BEC ISG-15 gene expression in a concentration dependent manner both in vivo and in vitro. We conclude that orally administered IFN-alpha exerts its immunomodulatory effects in humans in part by upregulating the production of ISG-15 by BEC, thereby enhancing the immune reactivity of mucosa-associated lymphocytes.

Interferon-alpha upregulates gene expression of aquaporin-5 in human parotid glands.

Aquaporins are a family of homologous membrane proteins that function as highly selective water channels. Aquaporin-5 (AQP5) is uniquely present in lacrimal and salivary glands, where it accounts for normal tear and saliva production. We tested the hypothesis that orally administered human interferon-alpha (HuIFN-alpha) benefits persons with xerostomia by augmenting the production of AQP5 protein by parotid gland epithelium. Cells from three human parotid glands were cultured with and without human lymphoblastoid IFN-alpha, and assayed for AQP5 mRNA levels by reverse transcriptase polymerase chain reaction (RT-PCR), and AQP5 protein levels by Western blot. Intracellular localization of AQP5 protein was done using confocal microscopy. The functional integrity of the glandular tissue was confirmed by RT-PCR analysis of alpha-amylase 1 and basic proline-rich protein transcripts. AQP5 was constitutively expressed in human parotid gland tissue, with AQP5 protein restricted to the plasma membranes and cytoplasmic vesicles of acinar cells. IFN-alpha augmented AQP5 transcription and protein production in a concentration-dependent manner, and increased the size of intensity of staining of AQP5-containing cytoplasmic vesicles in acinar cells. We conclude that IFN-alpha upregulates AQP5 gene expression in human parotid acinar cells in vitro. To our knowledge, this is the first demonstration that IFN-alpha regulates the gene expression of an aquaporin.

Clinical manifestations of IgE hypogammaglobulinemia.

BACKGROUND: Although IgE has been shown to play a role in the expulsion of intestinal parasites in experimental animals, its overall contribution to host defense in humans remains a subject of controversy. In order to clarify the potential role of IgE in host defense, we have studied the clinical characteristics of patient with serum IgE levels of < 2.5 IU/mL, using patients with normal or elevated IgE levels as controls. OBJECTIVE: To determine the clinical characteristics of IgE deficiency. METHODS: Serum IgE levels were measured in 420 adult patients seen in our Allergy-Immunology Clinic over a period extending from January, 1990 to March, 1996. All subjects were examined by one of the authors (JKS or GHK) using a standardized history and physical examination form. Patients with IgE levels of < 2.5 IU/mL also had measurements of serum IgG, IgG subclasses, IgA and IgM. All IgE-deficient patients and 73% of those with normal to elevated IgE levels underwent RAST and/or skin testing for Type I hypersensitivity, and, where clinically indicated, had serum drawn for autoimmune serologic profiles. Infectious complications were documented by culture. The American Rheumatology Association criteria were used to establish a diagnosis of autoimmune disease. RESULTS: Forty-four patients were found to have IgE levels of < 2.5 IU/mL; 57% of these had depressed serum levels of other immunoglobulins, and 43% had isolated IgE deficiencies. Respiratory symptoms were equally common in IgE-deficient patients and in patients with normal to elevated IgE levels. IgE-deficient patients, however, were more likely to complain of arthralgias (P < .0001), chronic fatigue (P < .0001), and symptoms suggestive of airway infection (P = .0119). Compared with controls, patients with IgE deficiency had a higher prevalence of autoimmune disease (46% versus 15%) (P < .0001) and nonallergic reactive airway disease (73% versus 20%) (P < .0001). There was no difference in the prevalence of these disease in patients with selective IgE deficiency as compared with those with IgE deficiency complicated by deficits in other immunoglobulin classes. IgE-deficient patients with multiple immunoglobulin deficiencies, however, were more likely to have serious infection involving both the upper and lower respiratory tract than those with isolated IgE deficiency. CONCLUSIONS: IgE-deficient patients have an increased prevalence of multiple immunoglobulin deficits, autoimmune disease, and nonallergic reactive airway disease when compared with a clinic population of patients with normal to elevated IgE levels.

Strongyloides stercoralis: identification of antigens in natural human infections from endemic areas of the United States.

Using Western-blot analysis, we identified eight immunodominant antigens (apparent molecular weights 96, 86, 75, 56, 41, 32, 28, and 26 kDa) of Strongyloides stercoralis in natural human infections. For this study, 78 individual serum samples were obtained from S. stercoralis-infected patients residing in endemic areas of the United States. Poly A+ RNA was translated in vitro in the rabbit-reticulocyte lysate system. The newly synthesized translation products were immuno-precipitated with S. stercoralis human infection sera. All eight of the identified antigens were detected in the immunoprecipitates. The potential of these antigens as targets for immunodiagnosis is also discussed.

Antibiotic susceptibility patterns of community-acquired respiratory isolates of Moraxella catarrhalis in western Europe and in the USA. The Alexander Project Collaborative Group.

Eight hundred and eighteen Moraxella catarrhalis strains, isolated in 1992 and 1993 at 15 centres in Western Europe and the USA were tested for beta-lactamase production and resistance to 15 antibiotics. The proportion of beta-lactamase producing strains in Europe rose significantly from 70% in 1992 to 82% in 1993, whilst in the USA the increase (85-92%) was not significant. Penicillin and amoxycillin resistance was more prevalent in the USA than in Europe. All penicillin-resistant strains isolated in the USA exhibited beta-lactamase activity, whilst 8% of beta-lactamase-negative strains isolated in Europe were also penicillin resistant. All but three isolates were sensitive to cefaclor, cefuroxime, cefixime and ceftriaxone, all but one were sensitive to ofloxacin and all were sensitive to ciprofloxacin and amoxycillin cluvulanate. Resistance to erythromycin was not encountered, although 19 strains had MICs > or = 0.5 mg/L. Of these, 15 came from European centres. Almost all strains were highly susceptible to clarithromycin, azithromycin, doxycycline and co-trimoxazole.

Effect of interferon alpha on HLA-DR expression by human buccal epithelial cells.

/We have studied the effect of interferon alpha (IFN-alpha) on MHC class II expression by human buccal epithelial cells (BEC), and mRNA expression by BEC and mucosal-associated mononuclear cells (MAMC). In 6 experiments, freshly collected BEC were suspended at a concentration of 1.0 x 10(5)/ml in RPMI 1640 and incubated in the presence of 0-10,000 IU/ml of human lymphoblastoid IFN-alpha (HuIFN-alpha). Zero and six hour samples were analyzed by single color flow cytometry using FITC-labeled murine IgG1 monoclonal antibody to HLA-DR. Preparations were also analyzed for expression of cytokine transcripts (IL-2, IL-4, IL-5, IL-6, IL-8, IFN-gamma, GM-CSF) by reverse transcriptase polymerase chain reaction (RT-PCR). Increasing concentrations of IFN-alpha resulted in proportionate increases in the percentage of HLA-DR + BEC (r = 0.7897, p = 0.0627) and in the percentage of HLA-DR+ staining at higher intensities (10(1) to 10(2) log fluorescence intensity) (LFI) (r = 0.4010, p = 0.0424). The percentage of HLA-DR + BEC rose from a mean of 1.5% with no IFN-alpha to 7% with 10,000 IU/ml IFN-alpha (p < 0.05). The percentage of HLA-DR + BEC staining at 10(1) to 10(2) LFI rose from a mean of 8.3% with no added IFN-alpha to 19.2% with 10,000 IU/ml IFN-alpha (p < 0.05). Unstimulated BEC constitutively expressed IL-8 and GM-CSF. IFN-alpha stimulated preparations also expressed IFN-gamma, possibly due to the presence of MAMC, which comprised 2-9% of the total cell population. These data indicated that HuIFN-alpha upregulates MHC class II expression by human BEC, possibly by enhancing IFN-gamma production by MAMC present in the culture preparations.

Value of the agar plate method for the diagnosis of intestinal strongyloidiasis.

An agar plate method for the diagnosis of intestinal strongyloidiasis was compared to the standard formalin-ethyl acetate concentration method. A total of 13 of 225 patients with eosinophilia had positive stools for strongyloides larva by agar plate compared to six of 225 by the formalin-ethyl acetate method (P = .0455). Nine positive stool specimens by the agar plate method were tested by the Baermann technique, and five were positive. The agar plate method is a sensitive and efficient technique for the diagnosis of strongyloidiasis.

Infections in the nursing home.

Infections are a common cause of morbidity and mortality in nursing home patients. A variety of factors predispose these patients to infections. Infections commonly encountered among these patients include pneumonia, urinary tract infections, tuberculosis, and gastrointestinal and skin infections. Preventive measures and infection control techniques offer protection to both patients and employees.

Percutaneous endoscopic gastrostomy as an unrecognized source of methicillin-resistant Staphylococcus aureus colonization.

Methicillin-resistant Staphylococcus aureus (MRSA) caused colonization or infection around the gastrostomy site of seven hospitalized patients, five of whom were in the long-term care unit. All cultures of gastrostomy sites were retrospectively reviewed, and 28% had MRSA. The gastrostomy site was responsible for 6.3% of all MRSA cultures, and 12.5% of all MRSA-positive patients with gastrostomy site cultures had involvement at that site. The implications of MRSA and gastrostomy tubes are discussed.